CD45RO enriches for activated, highly mutated human germinal center B cells

PubMed Central

Jackson, Stephen M.; Harp, Natessa; Patel, Darshna; Zhang, Jeffrey; Willson, Savannah; Kim, Yoon J.; Clanton, Christian

2007-01-01

To date, there is no consensus regarding the influence of different CD45 isoforms during peripheral B-cell development. Examining correlations between surface CD45RO expression and various physiologic processes ongoing during the germinal center (GC) reaction, we hypothesized that GC B cells, like T cells, that up-regulate surface RO should progressively acquire phenotypes commonly associated with activated, differentiating lymphocytes. GC B cells (IgD−CD38+) were subdivided into 3 surface CD45RO fractions: RO−, RO+/−, and RO+. We show here that the average number of mutations per IgVH transcript increased in direct correlation with surface RO levels. Conjunctional use of RO and CD69 further delineated low/moderately and highly mutated fractions. Activation-induced cytidine deaminase (AID) mRNA was slightly reduced among RO+ GC B cells, suggesting that higher mutation averages are unlikely due to elevated somatic mutation activity. Instead, RO+ GC B cells were negative for Annexin V, comprised mostly (93%) of CD77− centrocytes, and were enriched for CD69+ cells. Collectively, RO+ GC B cells occupy what seems to be a specialized niche comprised mostly of centrocytes that may be in transition between activation states. These findings are among the first to sort GC B cells into populations enriched for live mutated cells solely using a single extracellular marker. PMID:17644737

Exposure to inorganic mercury in vivo attenuates extrinsic apoptotic signaling in Staphylococcal aureus enterotoxin B stimulated T-cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Laiosa, Michael D.; Eckles, Kevin G.; Langdon, Margaret

2007-12-15

The heavy metal mercury (Hg) is known to have immunomodulatory properties affecting lymphocyte signal transduction, death receptor signaling and autoimmunity. In this study we tested the hypothesis that Hg exposure would attenuate T-cell activation and caspase 8 and 3 activity in response to antigenic stimuli. To test this hypothesis, BALB/cJ mice were exposed to 10 mg/l mercuric chloride (HgCl{sub 2}) in their drinking water for 2 weeks followed by injection with 20 {mu}g of the Staphylococcal aureus enterotoxin B (SEB) superantigen. Eighteen hours after SEB challenge, there was a statistically significant reduction in caspase 8 and caspase 3 enzyme activitymore » in the SEB reactive V{beta}8+ T-cells. The attenuated caspase activity in Hg-exposed mice persisted for 48 h after exposure. Moreover, activation of caspase 8 and caspase 3 was reduced by more than 60% in CD95 deficient MRL/MpJ-Fas{sup lpr} mice demonstrating that caspase 8 and 3 activation in response to SEB is CD95 dependent. In addition to the effects of Hg on caspase activity, expression of the T-cell activation marker CD69 was also attenuated in SEB reactive V{beta}8 T-cells in Hg-exposed mice. Moreover, CD69 expression in MRL/MpJ-Fas{sup lpr} mice was also reduced. Taken together the caspase and CD69 data support a role for CD95 in promoting a proapoptotic and activated state in SEB responsive T-lymphocytes and this state is attenuated by the autoimmune potentiating environmental agent mercury.« less

Comparative analysis of activation induced marker (AIM) assays for sensitive identification of antigen-specific CD4 T cells

PubMed Central

Cirelli, Kimberly M.; Dan, Jennifer M.; Morou, Antigoni; Daigneault, Audrey; Brassard, Nathalie; Silvestri, Guido; Routy, Jean-Pierre; Havenar-Daughton, Colin; Crotty, Shane

2017-01-01

The identification and study of antigen-specific CD4 T cells, both in peripheral blood and in tissues, is key for a broad range of immunological research, including vaccine responses and infectious diseases. Detection of these cells is hampered by both their rarity and their heterogeneity, in particular with regards to cytokine secretion profiles. These factors prevent the identification of the total pool of antigen-specific CD4 T cells by classical methods. We have developed assays for the highly sensitive detection of such cells by measuring the upregulation of surface activation induced markers (AIM). Here, we compare two such assays based on concurrent expression of CD69 plus CD40L (CD154) or expression of OX40 plus CD25, and we develop additional AIM assays based on OX40 plus PD-L1 or 4-1BB. We compare the relative sensitivity of these assays for detection of vaccine and natural infection-induced CD4 T cell responses and show that these assays identify distinct, but overlapping populations of antigen-specific CD4 T cells, a subpopulation of which can also be detected on the basis of cytokine synthesis. Bystander activation had minimal effect on AIM markers. However, some T regulatory cells upregulate CD25 upon antigen stimulation. We therefore validated AIM assays designed to exclude most T regulatory cells, for both human and non-human primate (NHP, Macaca mulatta) studies. Overall, through head-to-head comparisons and methodological improvements, we show that AIM assays represent a sensitive and valuable method for the detection of antigen-specific CD4 T cells. PMID:29065175

Comparative analysis of activation induced marker (AIM) assays for sensitive identification of antigen-specific CD4 T cells.

PubMed

Reiss, Samantha; Baxter, Amy E; Cirelli, Kimberly M; Dan, Jennifer M; Morou, Antigoni; Daigneault, Audrey; Brassard, Nathalie; Silvestri, Guido; Routy, Jean-Pierre; Havenar-Daughton, Colin; Crotty, Shane; Kaufmann, Daniel E

2017-01-01

The identification and study of antigen-specific CD4 T cells, both in peripheral blood and in tissues, is key for a broad range of immunological research, including vaccine responses and infectious diseases. Detection of these cells is hampered by both their rarity and their heterogeneity, in particular with regards to cytokine secretion profiles. These factors prevent the identification of the total pool of antigen-specific CD4 T cells by classical methods. We have developed assays for the highly sensitive detection of such cells by measuring the upregulation of surface activation induced markers (AIM). Here, we compare two such assays based on concurrent expression of CD69 plus CD40L (CD154) or expression of OX40 plus CD25, and we develop additional AIM assays based on OX40 plus PD-L1 or 4-1BB. We compare the relative sensitivity of these assays for detection of vaccine and natural infection-induced CD4 T cell responses and show that these assays identify distinct, but overlapping populations of antigen-specific CD4 T cells, a subpopulation of which can also be detected on the basis of cytokine synthesis. Bystander activation had minimal effect on AIM markers. However, some T regulatory cells upregulate CD25 upon antigen stimulation. We therefore validated AIM assays designed to exclude most T regulatory cells, for both human and non-human primate (NHP, Macaca mulatta) studies. Overall, through head-to-head comparisons and methodological improvements, we show that AIM assays represent a sensitive and valuable method for the detection of antigen-specific CD4 T cells.

Effects of cytarabine on activation of human T cells - cytarabine has concentration-dependent effects that are modulated both by valproic acid and all-trans retinoic acid.

PubMed

Ersvaer, Elisabeth; Brenner, Annette K; Vetås, Kristin; Reikvam, Håkon; Bruserud, Øystein

2015-05-02

Cytarabine is used in the treatment of acute myeloid leukemia (AML). Low-dose cytarabine can be combined with valproic acid and all-trans retinoic acid (ATRA) as AML-stabilizing treatment. We have investigated the possible risk of immunotoxicity by this combination. We examined the effects of cytarabine combined with valproic acid and ATRA on in vitro activated human T cells, and we tested cytarabine at concentrations reached during in vivo treatment with high doses, conventional doses and low doses. T cells derived from blood donors were activated in vitro in cell culture medium alone or supplemented with ATRA (1 μM), valproic acid (500 or 1000 μM) or cytarabine (0.01-44 μM). Cell characteristics were assessed by flow cytometry. Supernatants were analyzed for cytokines by ELISA or Luminex. Effects on primary human AML cell viability and proliferation of low-dose cytarabine (0.01-0.5 μM) were also assessed. Statistical tests include ANOVA and Cluster analyses. Only cytarabine 44 μM had both antiproliferative and proapoptotic effects. Additionally, this concentration increased the CD4:CD8 T cell ratio, prolonged the expression of the CD69 activation marker, inhibited CD95L and heat shock protein (HSP) 90 release, and decreased the release of several cytokines. In contrast, the lowest concentrations (0.35 and 0.01 μM) did not have or showed minor antiproliferative or cytotoxic effects, did not alter activation marker expression (CD38, CD69) or the release of CD95L and HSP90, but inhibited the release of certain T cell cytokines. Even when these lower cytarabine concentrations were combined with ATRA and/or valproic acid there was still no or minor effects on T cell viability. However, these combinations had strong antiproliferative effects, the expression of both CD38 and CD69 was altered and there was a stronger inhibition of the release of FasL, HSP90 as well as several cytokines. Cytarabine (0.01-0.05 μM) showed a dose-dependent antiproliferative effect on AML cells, and in contrast to the T cells this effect reached statistical significance even at 0.01 μM. Even low levels of cytarabine, and especially when combined with ATRA and valproic acid, can decrease T cell viability, alter activation-induced membrane-molecule expression and decrease the cytokine release.

Markers of nonselective and specific NK cell activation.

PubMed

Fogel, Leslie A; Sun, Michel M; Geurs, Theresa L; Carayannopoulos, Leonidas N; French, Anthony R

2013-06-15

NK cell activation is controlled by the integration of signals from cytokine receptors and germline-encoded activation and inhibitory receptors. NK cells undergo two distinct phases of activation during murine CMV (MCMV) infection: a nonselective phase mediated by proinflammatory cytokines and a specific phase driven by signaling through Ly49H, an NK cell activation receptor that recognizes infected cells. We sought to delineate cell surface markers that could distinguish NK cells that had been activated nonselectively from those that had been specifically activated through NK cell receptors. We demonstrated that stem cell Ag 1 (Sca-1) is highly upregulated during viral infections (to an even greater extent than CD69) and serves as a novel marker of early, nonselective NK cell activation. Indeed, a greater proportion of Sca-1(+) NK cells produced IFN-γ compared with Sca-1(-) NK cells during MCMV infection. In contrast to the universal upregulation of Sca-1 (as well as KLRG1) on NK cells early during MCMV infection, differential expression of Sca-1, as well as CD27 and KLRG1, was observed on Ly49H(+) and Ly49H(-) NK cells late during MCMV infection. Persistently elevated levels of KLRG1 in the context of downregulation of Sca-1 and CD27 were observed on NK cells that expressed Ly49H. Furthermore, the differential expression patterns of these cell surface markers were dependent on Ly49H recognition of its ligand and did not occur solely as a result of cellular proliferation. These findings demonstrate that a combination of Sca-1, CD27, and KLRG1 can distinguish NK cells nonselectively activated by cytokines from those specifically stimulated through activation receptors.

Induction of CD69 expression by cagPAI-positive Helicobacter pylori infection

PubMed Central

Mori, Naoki; Ishikawa, Chie; Senba, Masachika

2011-01-01

AIM: To investigate and elucidate the molecular mechanism that regulates inducible expression of CD69 by Helicobacter pylori (H. pylori) infection. METHODS: The expression levels of CD69 in a T-cell line, Jurkat, primary human peripheral blood mononuclear cells (PBMCs), and CD4+ T cells, were assessed by immunohistochemistry, reverse transcription polymerase chain reaction, and flow cytometry. Activation of CD69 promoter was detected by reporter gene. Nuclear factor (NF)-κB activation in Jurkat cells infected with H. pylori was evaluated by electrophoretic mobility shift assay. The role of NF-κB signaling in H. pylori-induced CD69 expression was analyzed using inhibitors of NF-κB and dominant-negative mutants. The isogenic mutants with disrupted cag pathogenicity island (cagPAI) and virD4 were used to elucidate the role of cagPAI-encoding type IV secretion system and CagA in CD69 expression. RESULTS: CD69 staining was detected in mucosal lymphocytes and macrophages in specimens of patients with H. pylori-positive gastritis. Although cagPAI-positive H. pylori and an isogenic mutant of virD4 induced CD69 expression, an isogenic mutant of cagPAI failed to induce this in Jurkat cells. H. pylori also induced CD69 expression in PBMCs and CD4+ T cells. The activation of the CD69 promoter by H. pylori was mediated through NF-κB. Transfection of dominant-negative mutants of IκBs, IκB kinases, and NF-κB-inducing kinase inhibited H. pylori-induced CD69 activation. Inhibitors of NF-κB suppressed H. pylori-induced CD69 mRNA expression. CONCLUSION: The results suggest that H. pylori induces CD69 expression through the activation of NF-κB. cagPAI might be relevant in the induction of CD69 expression in T cells. CD69 in T cells may play a role in H. pylori-induced gastritis. PMID:21990950

In vitro Reactivity to Implant Metals Demonstrates a Person Dependent Association with both T-Cell and B-Cell Activation

PubMed Central

Hallab, Nadim James; Caicedo, Marco; Epstein, Rachael; McAllister, Kyron; Jacobs, Joshua J

2009-01-01

Hypersensitivity to metallic implants remains relatively unpredictable and poorly understood. We initially hypothesized that metal-induced lymphocyte proliferation responses to soluble metal challenge (ions) are mediated exclusively by early T-cell activation (not B-cells), typical of a Delayed-Type-Hypersensitivity response. We tested this by comparing proliferation (6-days) of primary lymphocytes with early T-cell and B-cell activation (48-hours) in three groups of subjects likely to demonstrate elevated metal-reactivity: Group 1(n=12) history of metal-sensitivity with no implant; Group 2a(n=6) well performing metal-on-metal THRs, and Group 2b(n=20) subjects with poorly performing metal-on-polymer total joint arthroplasties (TJA). Group 1 showed 100%(12/12) metal reactivity (Stimulation Index>2) to Ni. Group 2a&2b were 83%(5/6) and 75%(15/22) metal reactive (to Co, Cr or Ni) respectively. Of the n=32 metal reactive subjects to Co, Cr or Ni (SI>2), n=22/32 demonstrated >2-fold elevations in % of T-cell or B-cell activation (CD25+,CD69+) to metal challenge compared to untreated control. 18/22 metal-activated subjects demonstrated an exclusively T-cell or B-cell activation response to metal challenge, where 6/18 demonstrated exclusively B-cell activation and 12/18 demonstrated a T-cell only response, as measured by surface activation markers CD25+ and CD69+. However, there was no direct correlation (R2<0.1) between lymphocyte proliferation and % T-cell or B-cell activation (CD25+:CD69+). Proliferation assays (LTT) showed greater ability to detect metal reactivity than did subject-dependent results of flow-cytometry analysis of T-cell or B-cell activation. The high incidence of lymphocyte reactivity and activation, indicate that more complex than initially hypothesized immune responses may contribute to the etiology of debris induced osteolysis in metal-sensitive individuals. PMID:19235773

Invariant NKT cells from HIV-1 or Mycobacterium tuberculosis-infected patients express an activated phenotype.

PubMed

Montoya, Carlos J; Cataño, Juan C; Ramirez, Zoraida; Rugeles, Maria T; Wilson, S Brian; Landay, Alan L

2008-04-01

The frequency, subsets and activation status of peripheral blood invariant NKT (iNKT) cells were evaluated in pulmonary tuberculosis (TB) patients and in chronically HIV-1-infected subjects. The absolute numbers of iNKT cells were significantly decreased in TB patients and in HIV-1+ individuals who were antiretroviral therapy naive or had detectable viremia despite receiving HAART. iNKT cell subset analysis demonstrated a decreased percentage of CD4(+) iNKT cells in HIV-1+ subjects, and a decreased percentage of double negative iNKT cells in TB patients. Peripheral blood iNKT cells from HIV-1+ and TB patients had significantly increased expression of CD69, CD38, HLA-DR, CD16, CD56, and CD62L. The expression of CD25 was significantly increased only on iNKT cells from TB patients. These findings indicate that peripheral blood iNKT cells in these two chronic infections show an up-regulated expression of activation markers, suggesting their role in the immune response to infection.

Trichloroethylene and Its Oxidative Metabolites Enhance the Activated State and Th1 Cytokine Gene Expression in Jurkat Cells.

PubMed

Pan, Yao; Wei, Xuetao; Hao, Weidong

2015-08-28

Trichloroethylene (TCE) is an occupational and ubiquitous environmental contaminant, and TCE exposure will increase the risk of autoimmune diseases and allergic diseases. T cells play an important role in the pathogenesis of TCE-related immune disorders, but the effect of TCE and its oxidative metabolites, trichloroacetic acid (TCA) and dichloroacetic acid (DCA), on the activation of human T cells is still unknown. In this study, Jurkat cells were pre-treated with TCE, TCA and DCA overnight and then stimulated with phorbol 12-myristate 13-acetate and ionomycin for another 4, 8 and 24 hours. IL-2 secretion was detected by ELISA; the expressions of CD25 and CD69 were tested by flow cytometry; and IFN-γ and IL-2 mRNA expression levels were investigated by real-time PCR. The results showed that TCE and its oxidative metabolites, TCA and DCA, significantly enhanced IL-2 releasing and the expression of T cell activation markers, CD25 and CD69. Consistent with this result, these compounds markedly up-regulated the expression levels of IFN-γ and IL-2 mRNA. Collectively, these findings suggest that TCE and its metabolites, TCA and DCA, might enhance the activation of T cells and disrupt various activities of peripheral T cells.

Trichloroethylene and Its Oxidative Metabolites Enhance the Activated State and Th1 Cytokine Gene Expression inJurkat Cells

PubMed Central

Pan, Yao; Wei, Xuetao; Hao, Weidong

2015-01-01

Trichloroethylene (TCE) is an occupational and ubiquitous environmental contaminant, and TCE exposure will increase the risk of autoimmune diseases and allergic diseases. T cells play an important role in the pathogenesis of TCE-related immune disorders, but the effect of TCE and its oxidative metabolites, trichloroacetic acid (TCA) and dichloroacetic acid (DCA), on the activation of human T cells is still unknown. In this study, Jurkat cells were pre-treated with TCE, TCA and DCA overnight and then stimulated with phorbol 12-myristate 13-acetate and ionomycin for another 4, 8 and 24 hours. IL-2 secretion was detected by ELISA; the expressions of CD25 and CD69 were tested by flow cytometry; and IFN-γ and IL-2 mRNA expression levels were investigated by real-time PCR. The results showed that TCE and its oxidative metabolites, TCA and DCA, significantly enhanced IL-2 releasing and the expression of T cell activation markers, CD25 and CD69. Consistent with this result, these compounds markedly up-regulated the expression levels of IFN-γ and IL-2 mRNA. Collectively, these findings suggest that TCE and its metabolites, TCA and DCA, might enhance the activation of T cells and disrupt various activities of peripheral T cells. PMID:26343699

Weak vaccinia virus-induced NK cell regulation of CD4 T cells is associated with reduced NK cell differentiation and cytolytic activity.

PubMed

Hatfield, Steven D; Daniels, Keith A; O'Donnell, Carey L; Waggoner, Stephen N; Welsh, Raymond M

2018-06-01

Natural killer (NK) cells control antiviral adaptive immune responses in mice during some virus infections, but the universality of this phenomenon remains unknown. Lymphocytic choriomeningitis virus (LCMV) infection of mice triggered potent cytotoxic activity of NK cells (NK LCMV ) against activated CD4 T cells, tumor cells, and allogeneic lymphocytes. In contrast, NK cells activated by vaccinia virus (VACV) infection (NK VACV ) exhibited weaker cytolytic activity against each of these target cells. Relative to NK LCMV cells, NK VACV cells exhibited a more immature (CD11b - CD27 + ) phenotype, and lower expression levels of the activation marker CD69, cytotoxic effector molecules (perforin, granzyme B), and the transcription factor IRF4. NK VACV cells expressed higher levels of the inhibitory molecule NKG2A than NK LCMV cells. Consistent with this apparent lethargy, NK VACV cells only weakly constrained VACV-specific CD4 T-cell responses. This suggests that NK cell regulation of adaptive immunity, while universal, may be limited with viruses that poorly activate NK cells. Published by Elsevier Inc.

T cells in chronic lymphocytic leukemia display dysregulated expression of immune checkpoints and activation markers.

PubMed

Palma, Marzia; Gentilcore, Giusy; Heimersson, Kia; Mozaffari, Fariba; Näsman-Glaser, Barbro; Young, Emma; Rosenquist, Richard; Hansson, Lotta; Österborg, Anders; Mellstedt, Håkan

2017-03-01

Chronic lymphocytic leukemia is characterized by impaired immune functions largely due to profound T-cell defects. T-cell functions also depend on co-signaling receptors, inhibitory or stimulatory, known as immune checkpoints, including cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) and programmed death-1 (PD-1). Here we analyzed the T-cell phenotype focusing on immune checkpoints and activation markers in chronic lymphocytic leukemia patients (n=80) with different clinical characteristics and compared them to healthy controls. In general, patients had higher absolute numbers of CD3 + cells and the CD8 + subset was particularly expanded in previously treated patients. Progressive patients had higher numbers of CD4 + and CD8 + cells expressing PD-1 compared to healthy controls, which was more pronounced in previously treated patients ( P =0.0003 and P =0.001, respectively). A significant increase in antigen-experienced T cells was observed in patients within both the CD4 + and CD8 + subsets, with a significantly higher PD-1 expression. Higher numbers of CD4 + and CD8 + cells with intracellular CTLA-4 were observed in patients, as well as high numbers of proliferating (Ki67 + ) and activated (CD69 + ) CD4 + and CD8 + cells, more pronounced in patients with active disease. The numbers of Th1, Th2, Th17 and regulatory T cells were substantially increased in patients compared to controls ( P <0.05), albeit decreasing to low levels in pre-treated patients. In conclusion, chronic lymphocytic leukemia T cells display increased expression of immune checkpoints, abnormal subset distribution, and a higher proportion of proliferating cells compared to healthy T cells. Disease activity and previous treatment shape the T-cell profile of chronic lymphocytic leukemia patients in different ways. Copyright© Ferrata Storti Foundation.

Proinflammatory T Cell Status Associated with Early Life Adversity.

PubMed

Elwenspoek, Martha M C; Hengesch, Xenia; Leenen, Fleur A D; Schritz, Anna; Sias, Krystel; Schaan, Violetta K; Mériaux, Sophie B; Schmitz, Stephanie; Bonnemberger, Fanny; Schächinger, Hartmut; Vögele, Claus; Turner, Jonathan D; Muller, Claude P

2017-12-15

Early life adversity (ELA) has been associated with an increased risk for diseases in which the immune system plays a critical role. The ELA immune phenotype is characterized by inflammation, impaired cellular immunity, and immunosenescence. However, data on cell-specific immune effects are largely absent. Additionally, stress systems and health behaviors are altered in ELA, which may contribute to the generation of the ELA immune phenotype. The present investigation tested cell-specific immune differences in relationship to the ELA immune phenotype, altered stress parameters, and health behaviors in individuals with ELA ( n = 42) and those without a history of ELA (control, n = 73). Relative number and activation status (CD25, CD69, HLA-DR, CD11a, CD11b) of monocytes, NK cells, B cells, T cells, and their main subsets were assessed by flow cytometry. ELA was associated with significantly reduced numbers of CD69 + CD8 + T cells ( p = 0.022), increased numbers of HLA-DR + CD4 and HLA-DR + CD8 T cells ( p < 0.001), as well as increased numbers of CD25 + CD8 + T cells ( p = 0.036). ELA also showed a trend toward higher numbers of CCR4 + CXCR3 - CCR6 + CD4 T cells. Taken together, our data suggest an elevated state of immune activation in ELA, in which particularly T cells are affected. Although several aspects of the ELA immune phenotype were related to increased activation markers, neither stress nor health-risk behaviors explained the observed group differences. Thus, the state of immune activation in ELA does not seem to be secondary to alterations in the stress system or health-risk behaviors, but rather a primary effect of early life programming on immune cells. Copyright © 2017 by The American Association of Immunologists, Inc.

Human embryonic stem cell-derived neural crest cells capable of expressing markers of osteochondral or meningeal-choroid plexus differentiation.

PubMed

Sternberg, Hal; Jiang, Jianjie; Sim, Pamela; Kidd, Jennifer; Janus, Jeffrey; Rinon, Ariel; Edgar, Ron; Shitrit, Alina; Larocca, David; Chapman, Karen B; Binette, Francois; West, Michael D

2014-01-01

The transcriptome and fate potential of three diverse human embryonic stem cell-derived clonal embryonic progenitor cell lines with markers of cephalic neural crest are compared when differentiated in the presence of combinations of TGFβ3, BMP4, SCF and HyStem-C matrices. The cell lines E69 and T42 were compared with MEL2, using gene expression microarrays, immunocytochemistry and ELISA. In the undifferentiated progenitor state, each line displayed unique markers of cranial neural crest including TFAP2A and CD24; however, none expressed distal HOX genes including HOXA2 or HOXB2, or the mesenchymal stem cell marker CD74. The lines also showed diverse responses when differentiated in the presence of exogenous BMP4, BMP4 and TGFβ3, SCF, and SCF and TGFβ3. The clones E69 and T42 showed a profound capacity for expression of endochondral ossification markers when differentiated in the presence of BMP4 and TGFβ3, choroid plexus markers in the presence of BMP4 alone, and leptomeningeal markers when differentiated in SCF without TGFβ3. The clones E69 and T42 may represent a scalable source of primitive cranial neural crest cells useful in the study of cranial embryology, and potentially cell-based therapy.

CD45RA, a specific marker for leukaemia stem cell sub-populations in acute myeloid leukaemia.

PubMed

Kersten, Bas; Valkering, Matthijs; Wouters, Rolf; van Amerongen, Rosa; Hanekamp, Diana; Kwidama, Zinia; Valk, Peter; Ossenkoppele, Gert; Zeijlemaker, Wendelien; Kaspers, Gertjan; Cloos, Jacqueline; Schuurhuis, Gerrit J

2016-04-01

Chemotherapy resistant leukaemic stem cells (LSC) are thought to be responsible for relapses after therapy in acute myeloid leukaemia (AML). Flow cytometry can discriminate CD34(+) CD38(-) LSC and normal haematopoietic stem cells (HSC) by using aberrant expression of markers and scatter properties. However, not all LSC can be identified using currently available markers, so new markers are needed. CD45RA is expressed on leukaemic cells in the majority of AML patients. We investigated the potency of CD45RA to specifically identify LSC and HSC and improve LSC quantification. Compared to our best other markers (CLL-1, also termed CLEC12A, CD33 and CD123), CD45RA was the most reliable marker. Patients with high percentages (>90%) of CD45RA on CD34(+) CD38(-) LSC have 1·69-fold higher scatter values compared to HSC (P < 0·001), indicating a more mature CD34(+) CD38(-) phenotype. Patients with low (<10%) or intermediate (10-90%) CD45RA expression on LSC showed no significant differences to HSC (1·12- and 1·15-fold higher, P = 0·31 and P = 0·44, respectively). CD45RA-positive LSC tended to represent more favourable cytogenetic/molecular markers. In conclusion, CD45RA contributes to more accurate LSC detection and is recommended for inclusion in stem cell tracking panels. CD45RA may contribute to define new LSC-specific therapies and to monitor effects of anti-LSC treatment. © 2016 John Wiley & Sons Ltd.

Regulatory and activated effector T cells in chronic hepatitis C virus: Relation to autoimmunity

PubMed Central

Fouad, Hanan; El Raziky, Maissa; Hassan, Eman Medhat; Aziz, Ghada Mahmoud Abdel; Darweesh, Samar K; Sayed, Ahmed Reda

2016-01-01

AIM To investigate how Tregs are regulated in chronic hepatitis C virus (HCV) patients via assessment of Tregs markers (granzyme 2, CD69 and FoxP3), Teffs markers [TNFRSF4 (OX40), INFG] and CD4, CD25 genes. METHODS A prospective study was conducted on 120 subjects divided into 4 groups: Group I (n = 30) treatment naïve chronic HCV patients; Group II (n = 30) chronic HCV treated with Peg/Riba; Group III (n = 30) chronic HCV associated with non-organ specific autoantibody and Group IV (n = 30) healthy persons as a control group. Tregs and Teffs markers were assessed in peripheral blood mononuclear cells by quantitative real time reverse transcriptase-polymerase chain reaction. RESULTS Chronic HCV patients exhibited significant higher levels of both Teffs and Tregs in comparison to healthy control group. Tregs markers were significantly decreased in Peg/Riba treated HCV patients in comparison to treatment naïve HCV group. In HCV patients with antinuclear antibody (ANA) +ve, Tregs markers were significantly decreased in comparison to all other studied groups. Teffs markers were significantly elevated in all HCV groups in comparison to control and in HCV group with ANA +ve in comparison to treatment naïve HCV group. CONCLUSION Elevated Tregs cells in chronic HCV patients dampen both CD4+ and CD8+ autologous T cell immune response. Interferon-α and ribavirin therapy suppress proliferation of Tregs. More significant suppression of Tregs was observed in HCV patients with autoantibodies favoring pathological autoimmune response. PMID:27843539

Minocycline attenuates HIV-1 infection and suppresses chronic immune activation in humanized NOD/LtsZ-scidIL-2Rγnull mice

PubMed Central

Singh, Maneesh; Singh, Pratibha; Vaira, Dolores; Amand, Mathieu; Rahmouni, Souad; Moutschen, Michel

2014-01-01

More than a quarter of a century of research has established chronic immune activation and dysfunctional T cells as central features of chronic HIV infection and subsequent immunodeficiency. Consequently, the search for a new immunomodulatory therapy that could reduce immune activation and improve T-cell function has been increased. However, the lack of small animal models for in vivo HIV study has hampered progress. In the current study, we have investigated a model of cord blood haematopoietic progenitor cells (CB-HPCs) -transplanted humanized NOD/LtsZ-scidIL-2Rγnull mice in which progression of HIV infection is associated with widespread chronic immune activation and inflammation. Indeed, HIV infection in humanized NSG mice caused up-regulation of several T-cell immune activation markers such as CD38, HLA-DR, CD69 and co-receptor CCR5. T-cell exhaustion markers PD-1 and CTLA-4 were found to be significantly up-regulated on T cells. Moreover, increased plasmatic levels of lipopolysaccharide, sCD14 and interleukin-10 were also observed in infected mice. Treatment with minocycline resulted in a significant decrease of expression of cellular and plasma immune activation markers, inhibition of HIV replication and improved T-cell counts in HIV-infected humanized NSG mice. The study demonstrates that minocycline could be an effective, low-cost adjunctive treatment to regulate chronic immune activation and replication of HIV. PMID:24409837

Dietary Fructo-Oligosaccharides Attenuate Early Activation of CD4+ T Cells Which Produce both Th1 and Th2 Cytokines in the Intestinal Lymphoid Tissues of a Murine Food Allergy Model.

PubMed

Tsuda, Masato; Arakawa, Haruka; Ishii, Narumi; Ubukata, Chihiro; Michimori, Mana; Noda, Masanari; Takahashi, Kyoko; Kaminogawa, Shuichi; Hosono, Akira

2017-01-01

Fructo-oligosaccharides (FOS) are prebiotic agents with immunomodulatory effects involving improvement of the intestinal microbiota and metabolome. In this study, we investigated the cellular mechanisms through which FOS modulate intestinal antigen-specific CD4+ T cell responses in food allergy, using OVA23-3 mice. OVA23-3 mice were fed an experimental diet containing either ovalbumin (OVA) or OVA and FOS for 1 week. Body weight and mucosal mast cell protease 1 in the serum were measured as the indicator of intestinal inflammation. Single-cell suspensions were prepared from intestinal and systemic lymphoid tissues for cellular analysis. Cytokine production was measured by ELISA. Activation markers and intracellular cytokines in CD4+ T cells were analyzed by flow cytometry. Activated CD4+ T cells were purified to examine cytokine production. Dietary intake of FOS provided moderate protection from the intestinal inflammation induced by the OVA-containing diet. FOS significantly reduced food allergy-induced Th2 cytokine responses in intestinal tissues but not in systemic tissues. FOS decreased OVA diet-induced IFN-γ+IL-4+ double-positive CD4+ T cells and early-activated CD45RBhighCD69+CD4+ T cells in the mesenteric lymph nodes. Furthermore, we confirmed that these CD45RBhighCD69+CD4+ T cells are able to produce high levels of IFN-γ and moderate level of IL-4, IL-10, and IL-13. Dietary intake of FOS during the development of food allergy attenuates the induction of intestinal Th2 cytokine responses by regulating early activation of naïve CD4+ T cells, which produce both Th1 and Th2 cytokines. Our results suggest FOS might be a potential food agent for the prevention of food allergy by modulating oral sensitization to food antigens. © 2017 S. Karger AG, Basel.

Characterization of antibodies against ferret immunoglobulins, cytokines and CD markers.

PubMed

Martel, Cyril Jean-Marie; Aasted, Bent

2009-12-15

Ferret IgG and IgM were purified from normal serum, while ferret IgA was purified from bile. The estimated molecular weights of the immunoglobulin gamma, alpha and mu heavy chains were found to be 54kDa, 69kDa and 83kDa, respectively. For immunological (ELISA) quantification of ferret immunoglobulins, we identified and characterized polyclonal antibodies towards ferret IgG, IgM and IgA. We also identified 22 monoclonal antibodies (mAbs) raised mostly against human CD markers which cross-reacted with ferret leukocytes. These antibodies were originally specific against human CD8, CD9, CD14, CD18, CD25, CD29, CD32, CD44, CD61, CD71, CD79b, CD88, CD104, CD172a and mink CD3. Finally, we identified 4 cross-reacting mAbs with specificities against ferret interferon-gamma, TNF-alpha, interleukin-4 and interleukin-8.

Staphylococcal enterotoxin A-activated regulatory T cells promote allergen-specific TH2 response to intratracheal allergen inoculation.

PubMed

Zeng, Wei-Ping; McFarland, Margaret M; Zhou, Baohua; Holtfreter, Silva; Flesher, Susan; Cheung, Ambrose; Mallick, Avishek

2017-02-01

T H 2 responses are implicated in asthma pathobiology. Epidemiologic studies have found a positive association between asthma and exposure to staphylococcal enterotoxins. We used a mouse model of asthma to determine whether staphylococcal enterotoxins promote T H 2 differentiation of allergen-specific CD4 conventional T (Tcon) cells and asthma by activating allergen-nonspecific regulatory T (Treg) cells to create a T H 2-polarizing cytokine milieu. Ovalbumin (OVA)-specific, staphylococcal enterotoxin A (SEA)-nonreactive naive CD4 Tcon cells were cocultured with SEA-reactive allergen-nonspecific Treg or CD4 Tcon cells in the presence of OVA and SEA. The OVA-specific CD4 T cells were then analyzed for IL-13 and IFN-γ expression. SEA-activated Treg cells were analyzed for the expression of the T H 2-polarizing cytokine IL-4 and the T-cell activation markers CD69 and CD62L. For asthma induction, mice were intratracheally sensitized with OVA or cat dander extract (CDE) alone or together with SEA and then challenged with OVA or CDE. Mice were also subject to transient Treg cell depletion before sensitization with OVA plus SEA. Asthma features and T H 2 differentiation in these mice were analyzed. SEA-activated Treg cells induced IL-13 but suppressed IFN-γ expression in OVA-specific CD4 Tcon cells. SEA-activated Treg cells expressed IL-4, upregulated CD69, and downregulated CD62L. Sensitization with OVA plus SEA but not OVA alone induced asthma, and SEA exacerbated asthma induced by CDE. Depletion of Treg cells abolished these effects of SEA and IL-13 expression in OVA-specific T cells. SEA promoted T H 2 responses of allergen-specific T cells and asthma pathogenesis by activating Treg cells. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Emphysema and soluble CD14 are associated with pulmonary nodules in HIV-infected patients: implications for lung cancer screening.

PubMed

Triplette, Matthew; Sigel, Keith M; Morris, Alison; Shahrir, Shahida; Wisnivesky, Juan P; Kong, Chung Y; Diaz, Phillip T; Petraglia, Alycia; Crothers, Kristina

2017-07-31

Lung cancer screening may benefit HIV-infected (HIV) smokers because of an elevated risk of lung cancer, but may have unique harms because of HIV-specific risk factors for false-positive screens. This study seeks to understand whether inflammatory biomarkers and markers of chronic lung disease are associated with noncalcified nodules at least 4 mm (NCN) in HIV compared with uninfected patients. This is a cohort study of Examinations of HIV-Associated Lung Emphysema (EXHALE), including 158 HIV and 133 HIV-uninfected participants. Participants underwent a laboratory assessment [including measurement of D-dimer, interleukin 6, and soluble CD14 (sCD14)], chest computed tomography (CT), and pulmonary function testing. We created multivariable logistic regression models to determine predictors of NCN in the participants stratified by HIV status, with attention to semiqualitative scoring of radiographic emphysema, markers of pulmonary function, and inflammatory biomarkers. Of the 291 participants, 69 had NCN on chest CT. As previously reported, there was no difference in prevalence of these nodules by HIV status. Emphysema and elevated sCD14 demonstrated an association with NCN in HIV participants independent of smoking status, CD4 cell count, HIV viral load, and pulmonary function. Emphysema and sCD14, a marker of immune activation, was associated with a higher prevalence of NCN on chest CT in HIV participants. Patients with chronic immune activation and emphysema may be at higher risk for both false-positive findings and incident lung cancer, thus screening in this group requires further study to understand the balance of benefits and harms.

Distinct activation phenotype of a highly conserved novel HLA-B57-restricted epitope during dengue virus infection

PubMed Central

Townsley, Elizabeth; Woda, Marcia; Thomas, Stephen J; Kalayanarooj, Siripen; Gibbons, Robert V; Nisalak, Ananda; Srikiatkhachorn, Anon; Green, Sharone; Stephens, Henry AF; Rothman, Alan L; Mathew, Anuja

2014-01-01

Variation in the sequence of T-cell epitopes between dengue virus (DENV) serotypes is believed to alter memory T-cell responses during second heterologous infections. We identified a highly conserved, novel, HLA-B57-restricted epitope on the DENV NS1 protein. We predicted higher frequencies of B57-NS126–34-specific CD8+ T cells in peripheral blood mononuclear cells from individuals undergoing secondary rather than primary DENV infection. However, high tetramer-positive T-cell frequencies during acute infection were seen in only one of nine subjects with secondary infection. B57-NS126–34-specific and other DENV epitope-specific CD8+ T cells, as well as total CD8+ T cells, expressed an activated phenotype (CD69+ and/or CD38+) during acute infection. In contrast, expression of CD71 was largely limited to DENV epitope-specific CD8+ T cells. In vitro stimulation of cell lines indicated that CD71 expression was differentially sensitive to stimulation by homologous and heterologous variant peptides. CD71 may represent a useful marker of antigen-specific T-cell activation. PMID:23941420

Distinct activation phenotype of a highly conserved novel HLA-B57-restricted epitope during dengue virus infection.

PubMed

Townsley, Elizabeth; Woda, Marcia; Thomas, Stephen J; Kalayanarooj, Siripen; Gibbons, Robert V; Nisalak, Ananda; Srikiatkhachorn, Anon; Green, Sharone; Stephens, Henry A F; Rothman, Alan L; Mathew, Anuja

2014-01-01

Variation in the sequence of T-cell epitopes between dengue virus (DENV) serotypes is believed to alter memory T-cell responses during second heterologous infections. We identified a highly conserved, novel, HLA-B57-restricted epitope on the DENV NS1 protein. We predicted higher frequencies of B57-NS1(26-34) -specific CD8(+) T cells in peripheral blood mononuclear cells from individuals undergoing secondary rather than primary DENV infection. However, high tetramer-positive T-cell frequencies during acute infection were seen in only one of nine subjects with secondary infection. B57-NS1(26-34) -specific and other DENV epitope-specific CD8(+) T cells, as well as total CD8(+) T cells, expressed an activated phenotype (CD69(+) and/or CD38(+)) during acute infection. In contrast, expression of CD71 was largely limited to DENV epitope-specific CD8(+) T cells. In vitro stimulation of cell lines indicated that CD71 expression was differentially sensitive to stimulation by homologous and heterologous variant peptides. CD71 may represent a useful marker of antigen-specific T-cell activation. © 2013 John Wiley & Sons Ltd.

Semen CD4+ T Cells and Macrophages Are Productively Infected at All Stages of SIV infection in Macaques

PubMed Central

Bernard-Stoecklin, Sibylle; Gommet, Céline; Corneau, Aurélien B.; Guenounou, Sabrina; Torres, Claire; Dejucq-Rainsford, Nathalie; Cosma, Antonio; Dereuddre-Bosquet, Nathalie; Le Grand, Roger

2013-01-01

The mucosal events of HIV transmission have been extensively studied, but the role of infected cells present in the genital and rectal secretions, and in the semen, in particular, remains a matter of debate. As a prerequisite to a thorough in vivo investigation of the early transmission events through infected cells, we characterized in detail by multi-parameter flow cytometry the changes in macaque seminal leukocytes during SIVmac251 infection, focusing on T cells, macrophages and dendritic cells. Using immunocytofluorescence targeting SIV proteins and real-time quantitative PCR targeting SIV DNA, we investigated the nature of the infected cells on sorted semen leukocytes from macaques at different stages of infection. Finally, we cocultured semen CD4+ T cells and macrophages with a cell line permissive to SIV infection to assess their infectivity in vitro. We found that primary infection induced strong local inflammation, which was associated with an increase in the number of leukocytes in semen, both factors having the potential to favor cell-associated virus transmission. Semen CD4+ T cells and macrophages were productively infected at all stages of infection and were infectious in vitro. Lymphocytes had a mucosal phenotype and expressed activation (CD69 & HLA-DR) and migration (CCR5, CXCR4, LFA-1) markers. CD69 expression was increased in semen T cells by SIV infection, at all stages of infection. Macrophages predominated at all stages and expressed CD4, CCR5, MAC-1 and LFA-1. Altogether, we demonstrated that semen contains the two major SIV-target cells (CD4+ T cells and macrophages). Both cell types can be productively infected at all stages of SIV infection and are endowed with markers that may facilitate transmission of infection during sexual exposure. PMID:24348253

Children with postsurgical capillary leak syndrome can be distinguished by antigen expression on neutrophils and monocytes

NASA Astrophysics Data System (ADS)

Tarnok, Attila; Pipek, Michal; Valet, Guenter; Richter, Jacqueline; Hambsch, Joerg; Schneider, Peter

1999-04-01

Our initial studies indicate that children who develop post- operative capillary leak syndrome (CLS) following cardiac surgery with cardiopulmonary bypass (CPB) can be distinguished based on their pre-operative level of circulating cytokines an adhesion molecules. We tested flow cytometric analysis of surface antigen expression as a potential assay for risk assessment of CLS. 24th preoperative blood samples were stained with monoclonal antibodies for the adhesion molecules ICAM-1, LFA1, MAC1, (beta) -integrin, activation markers CD25, CD54, CD69, HLA- DR, CD14 or CD4. Cells were measured on a dual-laser flow cytometer calibrated with microbeads. Antigen expression was detected as mean fluorescence intensity. The data indicate, that neutrophils of CLS patients express preoperatively higher levels of LFA1 and monocytes higher levels of HLA-DR and activation markers thus are in a state of activation. This could in combination with surgical trauma and CPB lead to their additional stimulation and migration into sites of inflammation and induce postoperative CLS. It is planned to set up a Flow-Classification program for individual risk assessment. By discriminate analysis over 80 percent of the patients were correctly classified. Our preliminary study indicates that flow cytometry with its low samples requirements and rapid access of the results could be a powerful tool to perform risk assessment prior to pediatric open heart surgery.

Immunological parameters in elderly women: correlations with aerobic power, muscle strength and mood state.

PubMed

Raso, Vagner; Natale, Valéria Maria; Duarte, Alberto José da Silva; Greve, Júlia Maria D'Andrea; Shephard, Roy J

2012-05-01

Our objective was to relate immunological data for healthy but sedentary elderly women to aerobic power, strength, and mood state. We measured peak aerobic power and one-repetition maximum strength along with mood (depression and fatigue), quality of life and carbohydrate intake on 42 women aged 60-77 years. Standard immunological techniques determined natural killer cell count and cytotoxic activity (NKCA), proliferative responses to phytohemaglutinin and OKT(3), various lymphocyte subpopulations (CD3(+), CD3(-)CD19(+), CD56(+), CD4(+), CD8(+), CD56(dim) and CD56(bright)), and markers of activation, maturation, down-regulation and susceptibility to apoptosis (CD25(+), CD28(+), CD45RA(+), CD45RO(+), CD69(+), CD95(+), HLA-DR(+)). Correlations of immune parameters with aerobic power and strength were very similar for absolute and relative immunological data. In the group as a whole, the only correlation with aerobic power was -0.35 (relative CD4(+)CD69(+) count), but in subjects with values <22.6 mL kg(-1)min(-1) correlations ranged from -0.57 (relative CD4(+)CD45RO(+)) to 0.92 (absolute CD56(dim)HLA-DR(+)). In terms of muscle strength, univariate correlation coefficients ranged from -0.34 (relative and absolute CD3(+)CD4(+)CD8(+)) to +0.48 (absolute CD3(+)HLA-DR(+)) and +0.50 (absolute CD8(+)CD45RA(+)CD45RO(+)). Neither NKCA nor lymphocyte proliferation were correlated with aerobic power or muscle strength. Although mood state and quality of life can sometimes be influenced by an individual's fitness level, our multivariate analyses suggested that depression, fatigue and quality of life were more important determinants of immune profile than our fitness measures. Psychological changes associated with aging may have a substantial adverse effect upon the immune system, and immunological function may be enhanced more by addressing these issues than by focusing upon aerobic or resistance training. Copyright © 2012 Elsevier Inc. All rights reserved.

Short Communication: Low Expression of Activation and Inhibitory Molecules on NK Cells and CD4(+) T Cells Is Associated with Viral Control.

PubMed

Taborda, Natalia A; Hernández, Juan C; Lajoie, Julie; Juno, Jennifer A; Kimani, Joshua; Rugeles, María T; Fowke, Keith R

2015-06-01

Chronic HIV-1 infection induces severe immune alterations, including hyperactivation, exhaustion, and apoptosis. In fact, viral control has been associated with low frequencies of these processes. Here, we evaluated the expression of activation and inhibitory molecules on natural killer (NK) and CD4(+) T cells and plasma levels of proinflammatory cytokines in individuals exhibiting viral control: a cohort of HIV-1-exposed-seronegative individuals (HESN) and a cohort of HIV controllers. There was lower expression of CD69, LAG-3, PD-1, and TIM-3 in both cohorts when compared to a low-risk population or HIV progressors. In addition, HIV controllers exhibited lower plasma levels of proinflamatory molecules TNF-α and IP-10. These findings suggest that individuals exhibiting viral control have lower basal expression of markers associated with cellular activation and particularly immune exhaustion.

Distinct expression patterns of CD69 in mucosal and systemic lymphoid tissues in primary SIV infection of rhesus macaques.

PubMed

Wang, Xiaolei; Xu, Huanbin; Alvarez, Xavier; Pahar, Bapi; Moroney-Rasmussen, Terri; Lackner, Andrew A; Veazey, Ronald S

2011-01-01

Although the intestinal tract plays a major role in early human immunodeficiency virus (HIV) infection, the role of immune activation and viral replication in intestinal tissues is not completely understood. Further, increasing evidence suggests the early leukocyte activation antigen CD69 may be involved in the development or regulation of important T cell subsets, as well as a major regulatory molecule of immune responses. Using the simian immunodeficiency virus (SIV) rhesus macaque model, we compared expression of CD69 on T cells from the intestine, spleen, lymph nodes, and blood of normal and SIV-infected macaques throughout infection. In uninfected macaques, the majority of intestinal lamina propria CD4+ T cells had a memory (CD95+) phenotype and co-expressed CD69, and essentially all intestinal CCR5+ cells co-expressed CD69. In contrast, systemic lymphoid tissues had far fewer CD69+ T cells, and many had a naïve phenotype. Further, marked, selective depletion of intestinal CD4+CD69+ T cells occurred in early SIV infection, and this depletion persisted throughout infection. Markedly increased levels of CD8+CD69+ T cells were detected after SIV infection in virtually all tissues, including the intestine. Further, confocal microscopy demonstrated selective, productive infection of CD3+CD69+ T cells in the intestine in early infection. Combined, these results indicate CD69+CD4+ T cells are a major early target for viral infection, and their rapid loss by direct infection may have profound effects on intestinal immune regulation in HIV infected patients.

Effect of Priming of Multipotent Mesenchymal Stromal Cells with Interferon γ on Their Immunomodulating Properties.

PubMed

Kapranov, N M; Davydova, Y O; Galtseva, I V; Petinati, N A; Drize, N I; Kuzmina, L A; Parovichnikova, E N; Savchenko, V G

2017-10-01

Multipotent mesenchymal stromal cells (MSCs) are widely used for cell therapy, in particular for prophylaxis and treatment of graft-versus-host disease. Due to their immunomodulatory properties, MSCs affect the composition of lymphocyte subpopulations, which depends on the immunological state of the organism and can change in different diseases and during treatment. Administration of MSCs is not always effective. Treatment of MSCs with different cytokines (in particular IFN-γ) leads to enhancement of their immunomodulatory properties. The aim of this study was to investigate subpopulational alterations and activation markers in lymphocytes (activated and non-activated) after interaction with MSCs and MSCs pretreated with IFN-γ (γMSCs) in vitro. Lymphocytes were co-cultured with MSCs or γMSCs for 4 days. The proportion of CD4+ and CD8 + expressing CD25, CD38, CD69, HLA-DR, and PD-1 and distribution of memory and effector subsets were measured by flow cytometry after co-cultivation of lymphocytes with MSCs or γMSCs. The distribution of lymphocyte subpopulations changes during culturing. In non-activated lymphocytes cultured without MSCs, decrease in the proportion of naïve cells and increase in the number of effector cells was observed. That could be explained as activation of lymphocytes in the presence of serum in culturing medium. Co-culturing of lymphocytes with MSCs and γMSCs leads to retention of their non-activated state. Activation of lymphocytes with phytohemagglutinin increases the number of central memory cells and activates marker expression. Interaction with MSCs and γMSCs prevents activation of lymphocytes and keeps their naïve state. Priming with IFN-γ did not induce MSCs inhibitory effect on activation of lymphocytes.

Role of novel type I interferon epsilon in viral infection and mucosal immunity

PubMed Central

Xi, Yang; Day, Stephanie L; Jackson, Ronald J; Ranasinghe, Charani

2012-01-01

Intranasal infection with vaccinia virus co-expressing interferon epsilon (VV-HIV-IFN-ɛ) was used to evaluate the role of IFN-ɛ in mucosal immunity. VV-HIV- IFN-ɛ infection induced a rapid VV clearance in lung that correlated with (i) an elevated lung VV-specific CD8+CD107a+IFN-γ+ population expressing activation markers CD69/CD103, (ii) enhanced lymphocyte recruitment to lung alveoli with reduced inflammation, and (iii) an heightened functional/cytotoxic CD8+CD4+ T-cell subset (CD3hiCCR7hiCD62Llo) in lung lymph nodes. These responses were different to that observed with intranasal VV-HA-IFN-α4 or VV-HA-IFN-β infections. When IFN-ɛ was used in an intranasal/intramuscular heterologous HIV prime-boost immunization, elevated HIV-specific effector, but not memory CD8+T cells responses, were observed in spleen, genito-rectal nodes, and Peyer's patch. Homing marker α4β7 and CCR9 analysis indicated that unlike other type I IFNs, IFN-ɛ could promote migration of antigen-specific CD8+T cells to the gut. Our results indicate that IFN-ɛ has a unique role in the mucosae and most likely can be used to control local lung and/or gut infections (i.e., microbicide) such as tuberculosis, HIV-1, or sexually transmitted diseases. PMID:22617838

Cyclic adenosine 5'-monophosphate and calcium induce CD152 (CTLA-4) up-regulation in resting CD4+ T lymphocytes.

PubMed

Vendetti, Silvia; Riccomi, Antonella; Sacchi, Alessandra; Gatta, Lucia; Pioli, Claudio; De Magistris, Maria Teresa

2002-12-01

The CTLA-4 (CD152) molecule is up-regulated upon T cell activation and proliferation, and plays a critical role in the inhibition of immune responses. We show in this study that cAMP induces up-regulation of CD152 in human CD4(+) T lymphocytes. This effect occurs in the absence of the up-regulation of CD69 and CD25 activation markers and T cell proliferation. In addition, we found that the Ca(2+) ionophore ionomycin also up-regulates CD152, and that the combination of a cAMP analog or cAMP inducers with ionomycin further enhances the expression of CD152 in resting CD4(+) T lymphocytes. However, cyclosporin A, which inhibits Ca(2+)/calcineurin signaling pathway, fully prevented the ionomycin- but not the cAMP-induced up-regulation of CD152. The effects of cAMP and ionomycin involve increase of both CD152 mRNA transcripts, coding for the membrane and the soluble forms of CD152. Furthermore, we show that CD152 molecules are translocated to the membrane and are functional, as their engagement by specific mAbs prevented NF-kappaB activation by anti-CD3/CD28 stimulation. These findings demonstrate that at least two novel signal pathways regulate CTLA-4 gene expression and CD152 molecule up-regulation in human CD4(+) T lymphocytes, in the absence of full T cell activation.

CD8+ T Cells Primed in the Periphery Provide Time-Bound Immune-Surveillance to the Central Nervous System

PubMed Central

Young, Kevin G.; MacLean, Susanne; Dudani, Renu; Krishnan, Lakshmi; Sad, Subash

2016-01-01

After vaccination, memory CD8+ T cells migrate to different organs to mediate immune surveillance. In most nonlymphoid organs, following an infection, CD8+ T cells differentiate to become long-lived effector-memory cells, thereby providing long-term protection against a secondary infection. In this study, we demonstrated that Ag-specific CD8+ T cells that migrate to the mouse brain following a systemic Listeria infection do not display markers reminiscent of long-term memory cells. In contrast to spleen and other nonlymphoid organs, none of the CD8+ T cells in the brain reverted to a memory phenotype, and all of the cells were gradually eliminated. These nonmemory phenotype CD8+ T cells were found primarily within the choroid plexus, as well as in the cerebrospinal fluid-filled spaces. Entry of these CD8+ T cells into the brain was governed primarily by CD49d/VCAM-1, with the majority of entry occurring in the first week postinfection. When CD8+ T cells were injected directly into the brain parenchyma, cells that remained in the brain retained a highly activated (CD69hi) phenotype and were gradually lost, whereas those that migrated out to the spleen were CD69low and persisted long-term. These results revealed a mechanism of time-bound immune surveillance to the brain by CD8+ T cells that do not reside in the parenchyma. PMID:21715683

Differential Inhibition of T Lymphocyte Proliferation and Cytokine Synthesis by [6]-Gingerol, [8]-Gingerol, and [10]-Gingerol.

PubMed

Bernard, Megan; Furlong, Suzanne J; Power Coombs, Melanie R; Hoskin, David W

2015-11-01

[6]-Gingerol, [8]-gingerol, and [10]-gingerol are pungent components of fresh ginger, extracts of which inhibit various components of the inflammatory response. Because little is known regarding the effect of gingerols with different unbranched alkyl side chain lengths on the activation and effector function of T lymphocytes, we compared the effects of [6]-gingerol, [8]-gingerol, and [10]-gingerol on murine T lymphocyte proliferation, expression of CD25 and CD69 activation markers, cytokine synthesis, and interleukin (IL)-2 receptor signaling. All three gingerols inhibited DNA synthesis by T lymphocytes, as well as interferon-γ synthesis. In contrast, only [8]-gingerol and [10]-gingerol inhibited CD25 and CD69 expression, and IL-2 synthesis. None of the gingerols affected IL-4 synthesis. Exogenous IL-2 enhanced T lymphocyte proliferation in the presence of [6]-gingerol but did not significantly increase T lymphocyte proliferation in the presence of [8]-gingerol or [10]-gingerol. In line with this finding, [8]-gingerol and [10]-gingerol impaired IL-2-induced proliferation of CTLL-2 cells, but constitutive CD25 expression was unaffected, indicating inhibition of IL-2 receptor signaling. In general, [10]-gingerol and [8]-gingerol were more potent inhibitors of T lymphocytes than [6]-gingerol. Suppression of T lymphocyte responses by gingerols suggests that these phytochemicals may be beneficial in chronic inflammatory conditions associated with excessive or inappropriate T lymphocyte activation. Copyright © 2015 John Wiley & Sons, Ltd.

Activating and inhibitory receptors on synovial fluid natural killer cells of arthritis patients: role of CD94/NKG2A in control of cytokine secretion

PubMed Central

de Matos, Cristina Teixeira; Berg, Louise; Michaëlsson, Jakob; Felländer-Tsai, Li; Kärre, Klas; Söderström, Kalle

2007-01-01

Natural killer (NK) cells are activated early during inflammatory events and contribute to the shaping of the ensuing adaptive immune response. To further understand the role for NK cells in inflammation, we investigated the phenotype and function of synovial fluid (SF) NK cells from patients with chronic joint inflammation, as well as from patients with transient inflammation of the knee following trauma. We confirm that synovial NK cells are similar to the well-characterized CD56bright peripheral blood (PB) NK-cell subset present in healthy individuals. However, compared to this PB subset the synovial NK cells express a higher degree of activation markers including CD69 and NKp44, the latter being up-regulated also on CD56bright NK cells in the PB of patients. Activated synovial NK cells produced interferon-γ and tumour necrosis factor, and the production was further up-regulated by antibody masking of CD94/NKG2A, and down-regulated by target cells expressing human leucocyte antigen-E in complex with peptides known to engage CD94/NKG2A. We conclude that synovial NK cells have an activated phenotype and that CD94/NKG2A is a key regulator of synovial NK-cell cytokine synthesis. PMID:17521371

Monocyte-Derived Signals Activate Human Natural Killer Cells in Response to Leishmania Parasites

PubMed Central

Messlinger, Helena; Sebald, Heidi; Heger, Lukas; Dudziak, Diana; Bogdan, Christian; Schleicher, Ulrike

2018-01-01

Activated natural killer (NK) cells release interferon (IFN)-γ, which is crucial for the control of intracellular pathogens such as Leishmania. In contrast to experimental murine leishmaniasis, the human NK cell response to Leishmania is still poorly characterized. Here, we investigated the interaction of human blood NK cells with promastigotes of different Leishmania species (Leishmania major, Leishmania mexicana, Leishmania infantum, and Leishmania donovani). When peripheral blood mononuclear cells or purified NK cells and monocytes (all derived from healthy blood donors from Germany without a history of leishmaniasis) were exposed to promastigotes, NK cells showed increased surface expression of the activation marker CD69. The extent of this effect varied depending on the Leishmania species; differences between dermotropic and viscerotropic L. infantum strains were not observed. Upregulation of CD69 required direct contact between monocytes and Leishmania and was partly inhibitable by anti-interleukin (IL)-18. Unexpectedly, IL-18 was undetectable in most of the supernatants (SNs) of monocyte/parasite cocultures. Confocal fluorescence microscopy of non-permeabilized cells revealed that Leishmania-infected monocytes trans-presented IL-18 to NK cells. Native, but not heat-treated SNs of monocyte/Leishmania cocultures also induced CD69 on NK cells, indicating the involvement of a soluble heat-labile factor other than IL-18. A role for the NK cell-activating cytokines IL-1β, IL-2, IL-12, IL-15, IL-21, and IFN-α/β was excluded. The increase of CD69 was not paralleled by NK cell IFN-γ production or enhanced cytotoxicity. However, prior exposure of NK cells to Leishmania parasites synergistically increased their IFN-γ release in response to IL-12, which was dependent on endogenous IL-18. CD1c+ dendritic cells were identified as possible source of Leishmania-induced IL-12. Finally, we observed that direct contact between Leishmania and NK cells reduced the expression of CD56 mRNA and protein on NK cells. We conclude that Leishmania activate NK cells via trans-presentation of IL-18 by monocytes and by a monocyte-derived soluble factor. IL-12 is needed to elicit the IFN-γ-response of NK cells, which is likely to be an important component of the innate control of the parasite. PMID:29472914

Curcumin Inhibits CD4+ T Cell Activation, but Augments CD69 Expression and TGF-β1-Mediated Generation of Regulatory T Cells at Late Phase

PubMed Central

Kim, Girak; Jang, Mi Seon; Son, Young Min; Seo, Min Ji; Ji, Sang Yun; Han, Seung Hyun; Jung, In Duk; Park, Yeong-Min; Jung, Hyun Jung; Yun, Cheol-Heui

2013-01-01

Background Curcumin is a promising candidate for a natural medicinal agent to treat chronic inflammatory diseases. Although CD4+ T cells have been implicated in the pathogenesis of chronic inflammation, whether curcumin directly regulates CD4+ T cells has not been definitively established. Here, we showed curcumin-mediated regulation of CD2/CD3/CD28-initiated CD4+ T cell activation in vitro. Methodology/Principal Findings Primary human CD4+ T cells were stimulated with anti-CD2/CD3/CD28 antibody-coated beads as an in vitro surrogate system for antigen presenting cell-T cell interaction and treated with curcumin. We found that curcumin suppresses CD2/CD3/CD28-initiated CD4+ T cell activation by inhibiting cell proliferation, differentiation and cytokine production. On the other hand, curcumin attenuated the spontaneous decline of CD69 expression and indirectly increased expression of CCR7, L-selectin and Transforming growth factor-β1 (TGF-β1) at the late phase of CD2/CD3/CD28-initiated T cell activation. Curcumin-mediated up-regulation of CD69 at late phase was associated with ERK1/2 signaling. Furthermore, TGF-β1 was involved in curcumin-mediated regulation of T cell activation and late-phase generation of regulatory T cells. Conclusions/Significance Curcumin not merely blocks, but regulates CD2/CD3/CD28-initiated CD4+ T cell activation by augmenting CD69, CCR7, L-selectin and TGF-β1 expression followed by regulatory T cell generation. These results suggest that curcumin could directly reduce T cell-dependent inflammatory stress by modulating CD4+ T cell activation at multiple levels. PMID:23658623

Attrition of memory CD8 T cells during sepsis requires LFA-1.

PubMed

Serbanescu, Mara A; Ramonell, Kimberly M; Hadley, Annette; Margoles, Lindsay M; Mittal, Rohit; Lyons, John D; Liang, Zhe; Coopersmith, Craig M; Ford, Mandy L; McConnell, Kevin W

2016-11-01

CD8 T cell loss and dysfunction have been implicated in the increased susceptibility to opportunistic infections during the later immunosuppressive phase of sepsis, but CD8 T cell activation and attrition in early sepsis remain incompletely understood. With the use of a CLP model, we assessed CD8 T cell activation at 5 consecutive time points and found that activation after sepsis results in a distinct phenotype (CD69 + CD25 int CD62L HI ) independent of cognate antigen recognition and TCR engagement and likely through bystander-mediated cytokine effects. Additionally, we observed that sepsis concurrently results in the preferential depletion of a subset of memory-phenotype CD8 T cells that remain "unactivated" (i.e., fail to up-regulate activation markers) by apoptosis. Unactivated CD44 HI OT-I cells were spared from sepsis-induced attrition, as were memory-phenotype CD8 T cells of mice treated with anti-LFA-1 mAb, 1 h after CLP. Perhaps most importantly, we demonstrate that attrition of memory phenotype cells may have a pathologic significance, as elevated IL-6 levels were associated with decreased numbers of memory-phenotype CD8 T cells in septic mice, and preservation of this subset after administration of anti-LFA-1 mAb conferred improved survival at 7 d. Taken together, these data identify potentially modifiable responses of memory-phenotype CD8 T cells in early sepsis and may be particularly important in the application of immunomodulatory therapies in sepsis. © Society for Leukocyte Biology.

Attrition of memory CD8 T cells during sepsis requires LFA-1

PubMed Central

Serbanescu, Mara A.; Ramonell, Kimberly M.; Hadley, Annette; Margoles, Lindsay M.; Mittal, Rohit; Lyons, John D.; Liang, Zhe; Coopersmith, Craig M.; Ford, Mandy L.; McConnell, Kevin W.

2016-01-01

CD8 T cell loss and dysfunction have been implicated in the increased susceptibility to opportunistic infections during the later immunosuppressive phase of sepsis, but CD8 T cell activation and attrition in early sepsis remain incompletely understood. With the use of a CLP model, we assessed CD8 T cell activation at 5 consecutive time points and found that activation after sepsis results in a distinct phenotype (CD69+CD25intCD62LHI) independent of cognate antigen recognition and TCR engagement and likely through bystander-mediated cytokine effects. Additionally, we observed that sepsis concurrently results in the preferential depletion of a subset of memory-phenotype CD8 T cells that remain “unactivated” (i.e., fail to up-regulate activation markers) by apoptosis. Unactivated CD44HI OT-I cells were spared from sepsis-induced attrition, as were memory-phenotype CD8 T cells of mice treated with anti-LFA-1 mAb, 1 h after CLP. Perhaps most importantly, we demonstrate that attrition of memory phenotype cells may have a pathologic significance, as elevated IL-6 levels were associated with decreased numbers of memory-phenotype CD8 T cells in septic mice, and preservation of this subset after administration of anti-LFA-1 mAb conferred improved survival at 7 d. Taken together, these data identify potentially modifiable responses of memory-phenotype CD8 T cells in early sepsis and may be particularly important in the application of immunomodulatory therapies in sepsis. PMID:27286793

In vitro Production of IL-6 and IFN-γ is Influenced by Dietary Variables and Predicts Upper Respiratory Tract Infection Incidence and Severity Respectively in Young Adults

PubMed Central

Meng, Huicui; Lee, Yujin; Ba, Zhaoyong; Fleming, Jennifer A.; Furumoto, Emily J.; Roberts, Robert F.; Kris-Etherton, Penny M.; Rogers, Connie J.

2015-01-01

Assessment of immune responses in healthy adults following dietary or lifestyle interventions is challenging due to significant inter-individual variability. Thus, gaining a better understanding of host factors that contribute to the heterogeneity in immunity is necessary. To address this question, healthy adults [n = 36, 18–40 years old, body mass index (BMI) 20–35 kg/m2] were recruited. Dietary intake was obtained via 3-day dietary recall records, physical activity level was evaluated using the International Physical Activity Questionnaire, and peripheral blood mononuclear cells were isolated from peripheral blood. Expression of activation markers on unstimulated immune subsets was assessed by flow cytometry. T-cell proliferation and cytokine secretion was assessed following in vitro stimulation with anti-CD3 or lipopolysaccharide. Furthermore, the incidence and severity of cold or flu symptoms were obtained from self-reported upper respiratory tract infection (URTI) questionnaires. The relationship between activation marker expression on T cells and T-cell effector functions; and in vitro cytokine secretion and URTI was determined by linear or logistic regression. CD69 and CD25 expression on unstimulated T cells was significantly associated with T-cell proliferation and interleukin-2 secretion. Incidence and severity of cold or flu symptoms was significantly associated with in vitro interleukin-6 and interferon-gamma secretion, respectively. Furthermore, host factors (e.g., age, BMI, physical activity, and diet) contributed significantly to the relationship between activation marker expression and T-cell effector function, and cytokine secretion and cold and flu status. In conclusion, these results suggest that lifestyle and dietary factors are important variables that contribute to immune responses and should be included in human clinical trials that assess immune endpoints. PMID:25788896

The level of PPD-specific IFN-gamma-producing CD4+ T cells in the blood predicts the in vivo response to PPD.

PubMed

Martins, Marcia Valéria B S; Lima, Mônica Cristina B S; Duppre, Nadia C; Matos, Haroldo J; Spencer, John S; Brennan, Patrick J; Sarno, Euzenir N; Fonseca, Leila; Pereira, Geraldo M B; Pessolani, Maria Cristina V

2007-05-01

There are no reliable means for detecting subclinical mycobacterial infections. The recent sequencing of several mycobacterial genomes has now afforded new opportunities for the development of pathogen-specific diagnostic tests, critical in the context of leprosy and tuberculosis control. In the present study, we applied a multi-parametric flow cytometric analysis that allowed the investigation of T-cell functions in order to define immunological markers that measure previous exposure to mycobacteria. We compared the in vivo response to PPD, the gold standard skin test reagent for measuring previous exposure to Mycobacterium tuberculosis, with in vitro parameters of leukocyte activation in five PPD positive and five PPD negative healthy volunteers. PPD-stimulated peripheral leukocytes expressing CD4, CD69, cutaneous lymphocyte-associated antigen (CLA) and intracellular IFN-gamma were enumerated in whole blood and compared with the size of in vivo PPD-induced induration and IFN-gamma production levels as measured by ELISA in supernatants of PPD-stimulated peripheral blood mononuclear cells. The reactivity to the tuberculin skin test (TST) was associated with markedly increased frequencies of PPD-responsive activated (CD69+) and IFN-gamma-producing CD4+T cells. Detection of PPD-specific IFN-gamma producing leukocytes was restricted to CD4+T cells and a subset of these cells was shown to express the skin homing molecule CLA. Multiple linear regression modeling of responses to PPD showed the highest association between skin test indurations and frequencies of PPD-responsive IFN-gamma-producing CD4+CD69+ T cells. Our data show that the in vitro enumeration of antigen-specific IFN-gamma-producing CD4+ T cells can provide an alternative to the in vivo tuberculin test for the detection of latent Mycobacterium tuberculosis infection. Moreover, the measurement of these immunological parameters can be useful for the screening of new specific antigens defined by the genome sequence allowing selection of the best candidates for new diagnostics (including new skin tests), and vaccines for leprosy and tuberculosis.

Study of CD69 antigen expression and integrity of leukocyte cellular membrane in stored platelet concentrates following irradiation and treatment with Mirasol® PRT System.

PubMed

Lachert, Elżbieta; Woźniak, Jolanta; Antoniewicz-Papis, Jolanta; Krzywdzińska, Agnieszka; Kubis, Jolanta; Mikołowska, Agata; Letowska, Magdalena

2017-01-01

Leukocytes in transfused blood components, particularly residual lymphocytes, have been shown to contribute to the occurrence of various adverse reactions. One of the most severe is transfusionassociated graft versus host disease (TA-GvHD) following transfusion of blood components contaminated with immunocompetent T lymphocytes. Irradiation is a routine method for protection against TA-GvHD. According to the literature, some pathogen reduction methods have also been proven effective for the inactivation of T lymphocytes, and so they may be considered as an alternative to irradiation. Comparison of CD69 antigen expression and the integrity of the leukocyte cellular membrane in stored platelet concentrates (PCs) following irradiation with the Gammacell 3000 Elan (Nordion Inc., Ottawa, Canada) and treatment with the Mirasol® Pathogen Reduction Technology (PRT) System (Terumo BCT, Lakewood, USA). The study included seven experiments. For each experiment we used 3 PCs, for Mirasol® PRT System treatment (M), for Gammacell 3000 Elan irradiation (R), and for the control (C). 7-amino-actinomycin D (7-AAD, Becton Dickinson, Franklin Lakes, USA) permeability was used to determine lymphocyte viability while CD69 antigen expression was the marker of lymphocyte activation. Analyses of 7-AAD and CD69 antigen expression were performed in a FACS Canto I flow cytometer (Becton Dickinson, USA). During 6 storage days, viable lymphocyte count decreased to 28% (p = 0.001) in the Mirasol® PRT System treated PCs and to 65% (p = 0.004) in the irradiated PCs. A statistically significant increase in CD69 expression in the irradiated PCs was observed; 1.3-fold on day 3 and 1.5-fold on day 6. In the Mirasol ® PRT System treated PCs, no statistically significant increase was observed. The in vitro results suggest that the Mirasol® PRT System is as effective as irradiation due to donor leukocyte inactivation capacity.

Expression of natural killer cell activity with CD107a on ectopic endometrium in woman with endometriosis compared with non-endometriosis

NASA Astrophysics Data System (ADS)

Lubis, H. P.; Aldiansyah, D.; Siregar, H. S.; Rivany, R.; Hariadi, T. S.

2018-03-01

Some factors have an important role in endometriosis pathogenesis; there is an immune cell that plays an important role in endometrial cells that have reflux. Woman with endometriosis experienced the cellular immune disorder. It is suspected that decrease of NK cell in the peritoneal fluid caused by its qualitative defect with CD107a expression as the best marker. The aim of this study was to compare expression of NK Cell activity with CD107a between awoman with endometriosis and non-endometriosis. A case-control study from March until July 2015 in Haji Adam Malik General Hospital. The case group was ectopic endometrial tissue block paraffin and control group was normal endometrial tissue block paraffin. This study included 23 patients in endometriosis group and control group respectively. A majority proportion of CD107a expression in endometriosis group was +1 (16 patients (69.6%)), while the control group was +3 (9 patients (39.1%)). Expression of NK cell activity with CD107a in patients with endometriosis was lower than the control group (p<0.05). It suggested that cellular immune factors may play a role in the pathogenesis of endometriosis.

Thymus-Derived Regulatory T Cell Development Is Regulated by C-Type Lectin-Mediated BIC/MicroRNA 155 Expression

PubMed Central

Sánchez-Díaz, Raquel; Blanco-Dominguez, Rafael; Lasarte, Sandra; Tsilingiri, Katerina; Martín-Gayo, Enrique; Linillos-Pradillo, Beatriz; de la Fuente, Hortensia; Sánchez-Madrid, Francisco; Nakagawa, Rinako; Toribio, María L.

2017-01-01

ABSTRACT Thymus-derived regulatory T (tTreg) cells are key to preventing autoimmune diseases, but the mechanisms involved in their development remain unsolved. Here, we show that the C-type lectin receptor CD69 controls tTreg cell development and peripheral Treg cell homeostasis through the regulation of BIC/microRNA 155 (miR-155) and its target, suppressor of cytokine signaling 1 (SOCS-1). Using Foxp3-mRFP/cd69+/− or Foxp3-mRFP/cd69−/− reporter mice and short hairpin RNA (shRNA)-mediated silencing and miR-155 transfection approaches, we found that CD69 deficiency impaired the signal transducer and activator of transcription 5 (STAT5) pathway in Foxp3+ cells. This results in BIC/miR-155 inhibition, increased SOCS-1 expression, and severely impaired tTreg cell development in embryos, adults, and Rag2−/− γc−/− hematopoietic chimeras reconstituted with cd69−/− stem cells. Accordingly, mirn155−/− mice have an impaired development of CD69+ tTreg cells and overexpression of the miR-155-induced CD69 pathway, suggesting that both molecules might be concomitantly activated in a positive-feedback loop. Moreover, in vitro-inducible CD25+ Treg (iTreg) cell development is inhibited in Il2rγ−/−/cd69−/− mice. Our data highlight the contribution of CD69 as a nonredundant key regulator of BIC/miR-155-dependent Treg cell development and homeostasis. PMID:28167605

Chronic Alcohol Ingestion Delays T Cell Activation and Effector Function in Sepsis.

PubMed

Margoles, Lindsay M; Mittal, Rohit; Klingensmith, Nathan J; Lyons, John D; Liang, Zhe; Serbanescu, Mara A; Wagener, Maylene E; Coopersmith, Craig M; Ford, Mandy L

2016-01-01

Sepsis is the leading cause of death in intensive care units in the US, and it is known that chronic alcohol use is associated with higher incidence of sepsis, longer ICU stays, and higher mortality from sepsis. Both sepsis and chronic alcohol use are associated with immune deficits such as decreased lymphocyte numbers, impaired innate immunity, delayed-type hypersensitivity reactions, and susceptibility to infections; however, understanding of specific pathways of interaction or synergy between these two states of immune dysregulation is lacking. This study therefore sought to elucidate mechanisms underlying the immune dysregulation observed during sepsis in the setting of chronic alcohol exposure. Using a murine model of chronic ethanol ingestion followed by sepsis induction via cecal ligation and puncture, we determined that while CD4+ and CD8+ T cells isolated from alcohol fed mice eventually expressed the same cellular activation markers (CD44, CD69, and CD43) and effector molecules (IFN-γ, TNF) as their water fed counterparts, there was an overall delay in the acquisition of these phenotypes. This early lag in T cell activation was associated with significantly reduced IL-2 production at a later timepoint in both the CD4+ and CD8+ T cell compartments in alcohol sepsis, as well as with a reduced accumulation of CD8dim activated effectors. Taken together, these data suggest that delayed T cell activation may result in qualitative differences in the immune response to sepsis in the setting of chronic alcohol ingestion.

West African Sorghum bicolor Leaf Sheaths Have Anti-Inflammatory and Immune-Modulating Properties In Vitro

PubMed Central

Benson, Kathleen F.; Beaman, Joni L.; Ou, Boxin; Okubena, Ademola; Okubena, Olajuwon

2013-01-01

Abstract The impact of chronic inflammatory conditions on immune function is substantial, and the simultaneous application of anti-inflammatory and immune modulating modalities has potential for reducing inflammation-induced immune suppression. Sorghum-based foods, teas, beers, and extracts are used in traditional medicine, placing an importance on obtaining an increased understanding of the biological effects of sorghum. This study examined selected anti-inflammatory and immune-modulating properties in vitro of Jobelyn™, containing the polyphenol-rich leaf sheaths from a West African variant of Sorghum bicolor (SBLS). Freshly isolated primary human polymorphonuclear (PMN) and mononuclear cell subsets were used to test selected cellular functions in the absence versus presence of aqueous and ethanol extracts of SBLS. Both aqueous and nonaqueous compounds contributed to reduced reactive oxygen species formation by inflammatory PMN cells, and reduced the migration of these cells in response to the inflammatory chemoattractant leukotriene B4. Distinct effects were seen on lymphocyte and monocyte subsets in cultures of peripheral blood mononuclear cells. The aqueous extract of SBLS triggered robust upregulation of the CD69 activation marker on CD3− CD56+ natural killer (NK) cells, whereas the ethanol extract of SBLS triggered similar upregulation of CD69 on CD3+ CD56+ NKT cells, CD3+ T lymphocytes, and monocytes. This was accompanied by many-fold increases in the chemokines RANTES/CCL5, Mip-1α/CCL3, and MIP-1β/CCL4. Both aqueous and nonaqueous compounds contribute to anti-inflammatory effects, combined with multiple effects on immune cell activation status. These observations may help suggest mechanisms of action that contribute to the traditional use of sorghum-based products, beverages, and extracts for immune support. PMID:23289787

Involvement of Mucosal-associated Invariant T cells in Ankylosing Spondylitis.

PubMed

Hayashi, Eri; Chiba, Asako; Tada, Kurisu; Haga, Keiichi; Kitagaichi, Mie; Nakajima, Shihoko; Kusaoi, Makio; Sekiya, Fumio; Ogasawara, Michihiro; Yamaji, Ken; Tamura, Naoto; Takasaki, Yoshinari; Miyake, Sachiko

2016-09-01

Ankylosing spondylitis (AS) is characterized by chronic inflammation of the axial and peripheral joints and ligamentous attachments. Gut immunity is thought to be involved in AS, because a prominent coexistence of gut and joint inflammation has been observed in patients with AS. Mucosal-associated invariant T (MAIT) cells are preferentially located in the gut lamina propria and produce inflammatory cytokines such as interleukin 17 (IL-17) and tumor necrosis factor-α (TNF-α), which are therapeutic targets for AS. This study aimed to investigate the involvement of MAIT cells in AS. The frequency of MAIT cells and their cytokine production were determined in patients with AS and healthy controls (HC). The expression of a MAIT cell activation marker (CD69) was analyzed in patients with AS by using flow cytometry. The frequency of MAIT cells in the peripheral blood was lower in patients with AS compared with HC. The levels of IL-17 produced by MAIT cells after activation were higher in patients with AS than in the HC. CD69 expression on MAIT cells correlated with the Ankylosing Spondylitis Disease Activity Score in patients with AS. These results suggest the involvement of MAIT cells in the pathogenesis of AS.

A controlled clinical trial to evaluate the effect of GanedenBC(30) on immunological markers.

PubMed

Kimmel, M; Keller, D; Farmer, S; Warrino, D E

2010-03-01

GanedenBC(30), a probiotic, has been shown to significantly increase T-cell production of TNF-alpha after ex vivo exposure to a strain of adenovirus (AdenoVI) or influenza A (H3N2 Texas strain [FluTex]). The current controlled study was designed to further evaluate the effect of GanedenBC(30) on immunological marker levels following viral exposure. Ten healthy subjects' baseline immunological marker levels were analyzed. Subjects consumed 1 capsule/day of GanedenBC(30) for 28 days and returned for post-treatment immunological marker evaluation. Subjects' baseline measurements served as their own control. All subjects completed the study with no adverse events; however, one subject was excluded from the final analysis based on a reasonable consideration as an outlier. CD3+CD69+ cells, IL-6, IL-8, interferon-gamma (IFN-gamma) and TNF-alpha levels were increased after exposure to AdenoVI and FluTex. IL-1beta levels also increased after exposure to AdenoVI but were decreased after ex vivo exposure to FluTex. CD3+CD69+ cells increased significantly (P = 0.023) after exposure to both viral strains. Differences in IL-8 levels after FluTex exposure achieved statistical significance (P = 0.039) as did IFN-gamma levels after AdenoVI exposure (P = 0.039). A regimen of one capsule per day containing 500 million CFU of GanedenBC30 may be a safe and effective option for enhancing the immunological response to common viral respiratory tract infections. 2010 Prous Science, S.A.U. or its licensors. All rights reserved.

Immunotherapy expands and maintains the function of high affinity tumor infiltrating CD8 T cells in situ

PubMed Central

Moran, Amy E.; Polesso, Fanny; Weinberg, Andrew D.

2016-01-01

Cancer cells harbor high affinity tumor-associated antigens capable of eliciting potent anti-tumor T cell responses yet detecting these polyclonal T cells is challenging. Therefore, surrogate markers of T cell activation such as CD69, CD44, and PD-1 have been used. We report here that in mice, expression of activation markers including PD-1 is insufficient in the tumor microenvironment to identify tumor-antigen specific T cells. Using the Nur77GFP T cell affinity reporter mouse, we highlight that PD-1 expression can be induced independent of TCR ligation within the tumor. Given this, we characterized the utility of the Nur77GFP model system in elucidating mechanisms of action of immunotherapies independent of PD-1 expression. Co-expression of Nur77GFP and OX40 identifies a polyclonal population of high affinity tumor-associated antigen-specific CD8+ T cells, which produce more IFNγ in situ than OX40 negative and doubles in quantity with anti-OX40 and anti-CTLA4 mAb therapy but not with anti-PD-1 or PD-L1. Moreover, expansion of these high affinity CD8 T cells prolongs survival of tumor bearing animals. Upon chronic stimulation in tumors and after adoptive cell therapy, CD8 TCR signaling and Nur77GFP induction is impaired and tumors progress. However, this can be reversed and overall survival significantly enhanced after adoptive cell therapy with agonist OX40 immunotherapy. Therefore, we propose that OX40 agonist immunotherapy can maintain functional TCR signaling of chronically stimulated tumor resident CD8 T cells thereby increasing the frequency of cytolytic, high affinity, tumor-associated antigen-specific cells. PMID:27503208

CCL21 mediates CD4+ T-cell costimulation via a DOCK2/Rac-dependent pathway.

PubMed

Gollmer, Kathrin; Asperti-Boursin, François; Tanaka, Yoshihiko; Okkenhaug, Klaus; Vanhaesebroeck, Bart; Peterson, Jeffrey R; Fukui, Yoshinori; Donnadieu, Emmanuel; Stein, Jens V

2009-07-16

CD4(+) T cells use the chemokine receptor CCR7 to home to and migrate within lymphoid tissue, where T-cell activation takes place. Using primary T-cell receptor (TCR)-transgenic (tg) CD4(+) T cells, we explored the effect of CCR7 ligands, in particular CCL21, on T-cell activation. We found that the presence of CCL21 during early time points strongly increased in vitro T-cell proliferation after TCR stimulation, correlating with increased expression of early activation markers. CCL21 costimulation resulted in increased Ras- and Rac-GTP formation and enhanced phosphorylation of Akt, MEK, and ERK but not p38 or JNK. Kinase-dead PI3Kdelta(D910A/D910A) or PI3Kgamma-deficient TCR-tg CD4(+) T cells showed similar responsiveness to CCL21 costimulation as control CD4(+) T cells. Conversely, deficiency in the Rac guanine exchange factor DOCK2 significantly impaired CCL21-mediated costimulation in TCR-tg CD4(+) T cells, concomitant with impaired Rac- but not Ras-GTP formation. Using lymph node slices for live monitoring of T-cell behavior and activation, we found that G protein-coupled receptor signaling was required for early CD69 expression but not for Ca(2+) signaling. Our data suggest that the presence of CCL21 during early TCR signaling lowers the activation threshold through Ras- and Rac-dependent pathways leading to increased ERK phosphorylation.

Identification of active and quiescent adipose vascular stromal cells.

PubMed

Lin, Guiting; Xin, Zhongcheng; Zhang, Haiyang; Banie, Lia; Wang, Guifang; Qiu, Xuefeng; Ning, Hongxiu; Lue, Tom F; Lin, Ching-Shwun

2012-02-01

Recent studies have demonstrated the existence of both active and quiescent stem cells in bone marrow, hair follicle and intestine. We attempted to identify active and quiescent vascular stromal cells (VSC) in adipose tissue. For identification of active VSC, adult rats were injected intraperitoneally with thymidine analog 5-ethynyl-2-deoxyuridine (EdU) and their subcutaneous tissue harvested 3 days later. For identification of quiescent VSC, newborn rats were injected intraperitoneally with EdU and their subcutaneous tissue harvested 9 weeks later. The harvested adipose tissues were examined for the co-localization of EdU with VSC marker CD34, smooth muscle marker SMA, endothelial marker RECA and pericyte marker CD140b. In adult rat adipose tissues harvested 3 days after EdU injection, there were 28.80 ± 8.70 (mean ± SD) EdU+ cells/100 × microscopic field, and approximately 6.2% of cell nuclei were labeled with EdU. The percentages of EdU+ cells expressing the following markers were approximately: 84 for CD34, 5.6 for RECA (rat endothelial marker), 3.7 for SMA and 14.8 for CD140b. In the adipose tissues of newborn rats that were harvested 9 weeks after EdU injection, the percentages of EdU+ cells expressing the following markers were approximately: 76 for CD34, 1.8 for RECA, 0 for SMA and 12.9 for CD140b. In both the short-term (active) and long-term (quiescent) EdU-labeled adipose tissues, the EdU label was consistently co-localized with CD34 and in the proximity of CD140b stain or in the adventitia. Both active and quiescent VSC expressed CD34 and localized to capillaries and the adventitia of larger blood vessels.

Evaluation of oral keratinocyte progenitor and T-lymphocite cells response during early healing after augmentation of keratinized gingiva with a 3D collagen matrix - a pilot study.

PubMed

Rusu, Darian; Calenic, Bogdan; Greabu, Maria; Kralev, Alexander; Boariu, Marius; Bojin, Florina; Anghel, Simona; Paunescu, Virgil; Vela, Octavia; Calniceanu, Horia; Stratul, Stefan-Ioan

2016-07-07

The aim of the present study is to analyze the behavior of selected populations of oral keratinocytes and T-lymphocytes, responsible for re-constructing and maintaining the oral epithelial tissue architecture, following augmentation of the keratinized oral mucosa using a 3D-collagen matrix. Different groups of oral keratinocytes were isolated from biopsies harvested from 3 patients before the surgical procedure, as well as 7 and 14 days after the augmentation procedure. T-lymphocytes were isolated from peripheral blood at same timepoints. Keratinocytes were characterized for stem and differentiation markers, such as p63, cytokeratin 10 and 14, and in vitro parameters, such as cell viability, cell size and colony-forming efficiency. T-lymphocytes were analyzed for viability and the expression of various cluster of differentiation markers. The methods included magnetic separation of cell populations, immunofluorescence, flow cytometry, and histology of oral biopsies. Both at 7 and 14 days, the majority of cells that repopulate the matrix were actively proliferating/progenitor oral keratinocytes with the phenotype integrin alfa6beta4 + CD71+. These cells display in vitro characteristics similar to the progenitor cells analyzed before the matrix placement. T-lymphocytes expressed CD8 and CD69 markers, while CD25 was absent. The study shows that two weeks after the collagen membrane placement, the healing process appeared to be histologically complete, with no abnormal immune response induced by the matrix, however, with a higher than usual content of active proliferating cells, the majority of keratinocytes being characterized as transit amplifying cells.

High proportion of PD-1-expressing CD4+ T cells in adipose tissue constitutes an immunomodulatory microenvironment that may support HIV persistence.

PubMed

Damouche, Abderaouf; Pourcher, Guillaume; Pourcher, Valérie; Benoist, Stéphane; Busson, Elodie; Lataillade, Jean-Jacques; Le Van, Mélanie; Lazure, Thierry; Adam, Julien; Favier, Benoit; Vaslin, Bruno; Müller-Trutwin, Michaela; Lambotte, Olivier; Bourgeois, Christine

2017-12-01

We and others have demonstrated that adipose tissue is a reservoir for HIV. Evaluation of the mechanisms responsible for viral persistence may lead to ways of reducing these reservoirs. Here, we evaluated the immune characteristics of adipose tissue in HIV-infected patients receiving antiretroviral therapy (ART) and in non-HIV-infected patients. We notably sought to determine whether adipose tissue's intrinsic properties and/or HIV induced alteration of the tissue environment may favour viral persistence. ART-controlled HIV infection was associated with a difference in the CD4/CD8 T-cell ratio and an elevated proportion of Treg cells in subcutaneous adipose tissue. No changes in Th1, Th2 and Th17 cell proportions or activation markers expression on T cell (Ki-67, HLA-DR) could be detected, and the percentage of CD69-expressing resident memory CD4 + T cells was not affected. Overall, our results indicate that adipose-tissue-resident CD4 + T cells are not extensively activated during HIV infection. PD-1 was expressed by a high proportion of tissue-resident memory CD4 + T cells in both HIV-infected patients and non-HIV-infected patients. Our findings suggest that adipose tissue's intrinsic immunomodulatory properties may limit immune activation and thus may strongly contribute to viral persistence. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

New iridoids from Verbascum nobile and their effect on lectin-induced T cell activation and proliferation.

PubMed

Dimitrova, Petya; Alipieva, Kalina; Grozdanova, Tsvetinka; Simova, Svetlana; Bankova, Vassya; Georgiev, Milen I; Popova, Milena P

2018-01-01

The Verbascum species are widely used traditional herb remedies against respiratory, inflammatory conditions and disorders. In the present study methanol extract of the aerial parts of the endemic Verbascum nobile Velen, was investigated and two novel iridoid glycosides 1 and 2, together with nine known constituents: iridoids, phenylethanoids, and saponins characteristic of Verbascum genus were identified. Further, the biological activity of the extract and selected isolated compounds on concanavalin (Con A)-induced T cell proliferation and activation of human Jurkat T cell line and splenic murine CD3 T cells was evaluated. T cell growth was studied by colorimetric-based WST proliferation assay while DNA content, cell cycling, dynamic of cell proliferation, expression of activation markers, intracellular expression of cytokine IFN-γ, and phosphorylation of ERK were analyzed by flow cytometry. Caspase-mediated apoptosis resulting in a poly (ADP-ribose) polymerase (PARP) cleavage was assessed by colorimetric in-cell kit. It was found that the extract, and all tested compounds (1, 2, 3 and 9) inhibited lectin-induced cell growth of Jurkat T cell line. The novel compounds decreased the frequencies of cells in S phase without causing a significant cell cycle arrest at G1 phase, caspases-mediated apoptosis and/or a profound change in the dynamic of splenic murine CD3 + T cell proliferation. Both compounds showed stronger inhibitory effect on Con A-induced ERK phosphorylation than the known bioactive compounds 3 and 9, and suppressed the expression of early activation marker CD69, the intracellular level of IFN-γ, and the generation of CD3 + IFN-γ + effectors. Our data suggest that the novel iridoid glycosides might have a potential to modulate T cell-related pathologies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Antiretroviral therapy in HIV-1-infected individuals with CD4 count below 100 cells/mm3 results in differential recovery of monocyte activation

PubMed Central

Patro, Sean C.; Azzoni, Livio; Joseph, Jocelin; Fair, Matthew G.; Sierra-Madero, Juan G.; Rassool, Mohammed S.; Sanne, Ian; Montaner, Luis J.

2016-01-01

Reversal of monocyte and macrophage activation and the relationship to viral suppression and T cell activation are unknown in patients with advanced HIV-1 infection, initiating antiretroviral therapy. This study aimed to determine whether reduction in biomarkers of monocyte and macrophage activation would be reduced in conjunction with viral suppression and resolution of T cell activation. Furthermore, we hypothesized that the addition of CCR5 antagonism (by maraviroc) would mediate greater reduction of monocyte/macrophage activation markers than suppressive antiretroviral therapy alone. In the CCR5 antagonism to decrease the incidence of immune reconstitution inflammatory syndrome study, antiretroviral therapy-naïve patients received maraviroc or placebo in addition to standard antiretroviral therapy. PBMCs and plasma from 65 patients were assessed during 24 wk of antiretroviral therapy for biomarkers of monocyte and macrophage activation. Markers of monocyte and macrophage activation were reduced significantly by 24 wk, including CD14++CD16+ intermediate monocytes (P < 0.0001), surface CD163 (P = 0.0004), CD169 (P < 0.0001), tetherin (P = 0.0153), and soluble CD163 (P < 0.0001). A change in CD38+, HLA-DR+ CD8 T cells was associated with changes in CD169 and tetherin expression. Maraviroc did not affect biomarkers of monocyte/macrophage activation but resulted in greater percentages of CCR5-positive monocytes in PBMC. HIV-1 suppression after 24 wk of antiretroviral therapy, with or without maraviroc, demonstrates robust recovery in monocyte subset activation markers, whereas soluble markers of activation demonstrate minimal decrease, qualitatively differentiating markers of monocyte/macrophage activation in advanced disease. PMID:26609048

Activation of Basophils Is a New and Sensitive Marker of Biocompatibility in Hemodialysis

PubMed Central

Aljadi, Zenib; Mansouri, Ladan; Nopp, Anna; Paulsson, Josefin M; Winqvist, Ola; Russom, Aman; Ståhl, Mårten; Hylander, Britta; Jacobson, Stefan H; Lundahl, Joachim

2014-01-01

The hemodialysis procedure involves contact between peripheral blood and the surface of dialyzer membranes, which may lead to alterations in the pathways of innate and adaptive immunity. We aimed to study the effect of blood–membrane interaction on human peripheral basophils and neutrophils in hemodialysis with high- and low-permeability polysulfone dialyzers. The surface expression of CD203c (basophil selection marker) and CD63 (activation marker) after activation by the bacterial peptide formyl-methionyl-leucyl-phenylalanine (fMLP) or anti-Fcε receptor I (FcεRI) antibody and the absolute number of basophils was investigated before and after hemodialysis with each of the dialyzers. Moreover, the expression on neutrophils of CD11b, the CD11b active epitope, and CD88 was analyzed in the same groups of individuals. The expression of CD63 in basophils following activation by fMLP was significantly higher in the patient group compared with that in healthy controls, but no differences were observed after activation by anti-FcεRI. During the hemodialysis procedure, the low-flux membrane induced up-regulation of CD63 expression on basophils, while passage through the high-flux membrane did not significantly alter the responsiveness. In addition, the absolute number of basophils was unchanged after hemodialysis with either of the dialyzers and compared with healthy controls. We found no significant differences in the expression of the neutrophil activation markers (CD11b, the active epitope of CD11b, and CD88) comparing the two different dialyzers before and after dialysis and healthy controls. Together, these findings suggest that alterations in basophil activity may be a useful marker of membrane bioincompatibility in hemodialysis. PMID:24712758

Activation states of blood eosinophils in asthma

PubMed Central

Johansson, Mats W.

2014-01-01

Asthma is characterized by airway inflammation rich in eosinophils. Airway eosinophilia is associated with exacerbations and has been suggested to play a role in airway remodeling. Recruitment of eosinophils from the circulation requires that blood eosinophils become activated, leading to their arrest on the endothelium and extravasation. Circulating eosinophils can be envisioned as potentially being in different activation states, including non-activated, pre-activated or “primed”, or fully activated. In addition, the circulation can potentially be deficient of pre-activated or activated eosinophils, because such cells have marginated on activated endothelium or extravasated into the tissue. A number of eosinophil-surface proteins, including CD69, L-selectin, intercellular adhesion molecule-1 (ICAM-1, CD54), CD44, P-selectin glycoprotein ligand-1 (PSGL-1, CD162), cytokine receptors, Fc receptors, integrins including αM integrin (CD11b), and activated conformations of Fc receptors and integrins have been proposed to report cell activation. Variation in eosinophil activation states may be associated with asthma activity. Eosinophil-surface proteins proposed to be activation markers, with a particular focus on integrins, and evidence for associations between activation states of blood eosinophils and features of asthma are reviewed here. Partial activation of β1 and β2 integrins on blood eosinophils, reported by monoclonal antibodies (mAb) N29 and KIM-127, is associated with impaired pulmonary function and airway eosinophilia, respectively, in non-severe asthma. The association with lung function does not occur in severe asthma, presumably due to greater eosinophil extravasation, specifically of activated or pre-activated cells, in severe disease. PMID:24552191

Normal T-cell activation in elite controllers with preserved CD4+ T-cell counts.

PubMed

Bansal, Anju; Sterrett, Sarah; Erdmann, Nathan; Westfall, Andrew O; Dionne-Odom, Jodie; Overton, Edgar T; Goepfert, Paul A

2015-11-01

HIV elite controllers suppress HIV viremia without antiretroviral therapy (ART), yet previous studies demonstrated that elite controllers maintain an activated T-cell phenotype. Chronic immune activation has detrimental consequences and thus ART has been advocated for all elite controllers. However, elite controllers are not a clinically homogenous group. Since CD4% is among the best predictors of AIDS-related events, in the current study, we assessed whether this marker can be used to stratify elite controllers needing ART. Sixteen elite controllers were divided into two groups based on CD4% (EC > 40% and EC ≤40%), and T-cell subsets were analyzed for markers of memory/differentiation (CD45RA, CCR7, CD28), activation (CD38/HLA-DR), immunosenescence (CD57), costimulation (CD73, CD28) and exhaustion (PD-1, CD160, Tim-3). Monocyte subsets (CD14, CD16) were also analyzed and sCD14 levels were quantified using ELISA. In the EC group, expression of activation, exhaustion, and immunosensescence markers on T cells were significantly reduced compared with the EC group and similar to the seronegative controls. The EC group expressed higher levels of costimulatory molecules CD28 and CD73 and had lower levels of monocyte activation (HLA-DR expression) with a reduced frequency of inflammatory monocyte (CD14 CD16) subset. Furthermore, the EC group maintained a stable CD4% during a median follow-up of 6 years. Elite controllers with preserved CD4T cells (EC) have normal T-cell and monocyte phenotypes and therefore may have limited benefit from ART. CD4% can be an important marker for evaluating future studies aimed at determining the need for ART in this group of individuals.

The feed gas composition determines the degree of physical plasma-induced platelet activation for blood coagulation

NASA Astrophysics Data System (ADS)

Bekeschus, Sander; Brüggemeier, Janik; Hackbarth, Christine; Weltmann, Klaus-Dieter; von Woedtke, Thomas; Partecke, Lars-Ivo; van der Linde, Julia

2018-03-01

Cold atmospheric (physical) plasma has long been suggested to be a useful tool for blood coagulation. However, the clinical applicability of this approach has not been addressed sufficiently. We have previously demonstrated the ability of a clinically accepted atmospheric pressure argon plasma jet (kINPen® MED) to coagulate liver incisions in mice with similar performance compared to the gold standard electrocauterization. We could show that plasma-mediated blood coagulation was dependent on platelet activation. In the present work, we extended on this by investigating kINPen®-mediated platelet activation in anticoagulated human donor blood ex vivo. With focus on establishing high-throughput, multi-parametric platelet activation assays and performing argon feed gas parameter studies we achieved the following results: (i) plasma activated platelets in heparinized but not in EDTA-anticoagulated blood; (ii) plasma decreased total platelet counts but increased numbers of microparticles; (iii) plasma elevated the expression of several surface activation markers on platelets (CD62P, CD63, CD69, and CD41/61); (iv) in platelet activation, wet and dry argon plasma outperformed feed gas admixtures with oxygen and/or nitrogen; (v) plasma-mediated platelet activation was accompanied by platelet aggregation. Platelet aggregation is a necessary requirement for blood clot formation. These findings are important to further elucidate molecular details and clinical feasibility of cold physical plasma-mediated blood coagulation.

Functional and phenotypical analysis of IL-6-secreting CD4+ T cells in human adipose tissue.

PubMed

de Jong, Anja J; Pollastro, Sabrina; Kwekkeboom, Joanneke C; Andersen, Stefan N; Dorjée, Annemarie L; Bakker, Aleida M; Alzaid, Fawaz; Soprani, Antoine; Nelissen, Rob G H H; Mullers, Jan B; Venteclef, Nicolas; de Vries, Niek; Kloppenburg, Margreet; Toes, René E M; Ioan-Facsinay, Andreea

2018-03-01

Emerging evidence indicates that a dynamic interplay between the immune system and adipocytes contributes to the disturbed homeostasis in adipose tissue of obese subjects. Recently, we observed IL-6-secretion by CD4 + T cells from the stromal vascular fraction (SVF) of the infrapatellar fat pad (IFP) of knee osteoarthritis patients directly ex vivo. Here we show that human IL-6 + CD4 + T cells from SVF display a more activated phenotype than the IL-6 - T cells, as evidenced by the expression of the activation marker CD69. Analysis of cytokines secretion, as well as expression of chemokine receptors and transcription factors associated with different Th subsets (Treg, Th1, Th2, Th17 and Tfh) revealed that IL-6-secreting CD4 + T cells cannot be assigned to a conventional Th subset. TCRβ gene analysis revealed that IL-6 + and IL-6 - CD4 + T cells appear clonally unrelated to each other, suggesting a different specificity of these cells. In line with these observations, adipocytes are capable of enhancing IL-6 production by CD4 + T cells. Thus, IL-6 + CD4 + T cells are TCRαβ T cells expressing an activated phenotype potentially resulting from an interplay with adipocytes that could be involved in the inflammatory processes in the OA joint. © 2017 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Interferon-γ production by tubulointerstitial human CD56bright natural killer cells contributes to renal fibrosis and chronic kidney disease progression.

PubMed

Law, Becker M P; Wilkinson, Ray; Wang, Xiangju; Kildey, Katrina; Lindner, Mae; Rist, Melissa J; Beagley, Kenneth; Healy, Helen; Kassianos, Andrew J

2017-07-01

Natural killer (NK) cells are a population of lymphoid cells that play a significant role in mediating innate immune responses. Studies in mice suggest a pathological role for NK cells in models of kidney disease. In this study, we characterized the NK cell subsets present in native kidneys of patients with tubulointerstitial fibrosis, the pathological hallmark of chronic kidney disease. Significantly higher numbers of total NK cells (CD3 - CD56 + ) were detected in renal biopsies with tubulointerstitial fibrosis compared with diseased biopsies without fibrosis and healthy kidney tissue using multi-color flow cytometry. At a subset level, both the CD56 dim NK cell subset and particularly the CD56 bright NK cell subset were elevated in fibrotic kidney tissue. However, only CD56 bright NK cells significantly correlated with the loss of kidney function. Expression of the tissue-retention and -activation molecule CD69 on CD56 bright NK cells was significantly increased in fibrotic biopsy specimens compared with non-fibrotic kidney tissue, indicative of a pathogenic phenotype. Further flow cytometric phenotyping revealed selective co-expression of activating receptor CD335 (NKp46) and differentiation marker CD117 (c-kit) on CD56 bright NK cells. Multi-color immunofluorescent staining of fibrotic kidney tissue localized the accumulation of NK cells within the tubulointerstitium, with CD56 bright NK cells (NKp46 + CD117 + ) identified as the source of pro-inflammatory cytokine interferon-γ within the NK cell compartment. Thus, activated interferon-γ-producing CD56 bright NK cells are positioned to play a key role in the fibrotic process and progression to chronic kidney disease. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

Immunosuppressive Effects of Bryoria sp. (Lichen-Forming Fungus) Extracts via Inhibition of CD8+ T-Cell Proliferation and IL-2 Production in CD4+ T Cells.

PubMed

Hwang, Yun-Ho; Lee, Sung-Ju; Kang, Kyung-Yun; Hur, Jae-Seoun; Yee, Sung-Tae

2017-06-28

Lichen-forming fungi are known to have various biological activities, such as antioxidant, antimicrobial, antitumor, antiviral, anti-inflammation, and anti proliferative effects. However, the immunosuppressive effects of Bryoria sp. extract (BSE) have not previously been investigated. In this study, the inhibitory activity of BSE on the proliferation of CD8 + T cells and the mixed lymphocytes reaction (MLR) was evaluated in vitro. BSE was non-toxic in spleen cells and suppressed the growth of splenocytes induced by anti-CD3. The suppressed cell population in spleen cells consisted of CD8 + T cells and their proliferation was inhibited by the treatment with BSE. This extract significantly suppressed the IL-2 associated with T cell growth and IFN-γ as the CD8 + T cell marker. Furthermore, BSE reduced the expression of the IL-2 receptor alpha chain (IL-2Rα) on CD8 + T cells and CD86 on dendritic cells by acting as antigen-presenting cells. Finally, the MLR produced by the co-culture of C57BL/6 and MMC-treated BALB/c was suppressed by BSE. IL-2, IFN-γ, and CD69 on CD8 + T cells in MLR condition were inhibited by BSE. These results indicate that BSE inhibits the MLR via the suppression of IL-2Rα expression in CD8 + T cells. BSE has the potential to be developed as an anti-immunosuppression agent for organ transplants.

Effect of a high and low dose of caffeine on human lymphocyte activation in response to antigen stimulation.

PubMed

Dulson, Deborah K; Bishop, Nicolette C

2016-02-01

This study investigated the effect of caffeine on antigen-stimulated lymphocyte activation. Six males rested for 3.5 h after ingesting 0 (PLA), 2, or 6 (6CAF) mg·kg(-1) body mass of caffeine. The number of antigen-stimulated NK CD69(+) cells increased in 6CAF at 1 h compared with PLA (P = 0.021). Caffeine did not influence the number of antigen-stimulated CD69(+) T cells or the geometric mean fluorescence intensity expression of CD69 on antigen-stimulated lymphocytes, suggesting caffeine has little effect on antigen-stimulated lymphocyte activation.

Granulysin-Expressing CD4+ T Cells as Candidate Immune Marker for Tuberculosis during Childhood and Adolescence

PubMed Central

Mueller, Henrik; Faé, Kellen C.; Magdorf, Klaus; Ganoza, Christian A.; Wahn, Ulrich; Guhlich, Ute; Feiterna-Sperling, Cornelia; Kaufmann, Stefan H. E.

2011-01-01

Background Granulysin produced by cytolytic T cells directly contributes to immune defense against tuberculosis (TB). We investigated granulysin as a candidate immune marker for childhood and adolescent TB. Methods Peripheral blood mononuclear cells (PBMC) from children and adolescents (1–17 years) with active TB, latent TB infection (LTBI), nontuberculous mycobacteria (NTM) infection and from uninfected controls were isolated and restimulated in a 7-day restimulation assay. Intracellular staining was then performed to analyze antigen-specific induction of activation markers and cytotoxic proteins, notably, granulysin in CD4+ CD45RO+ memory T cells. Results CD4+ CD45RO+ T cells co-expressing granulysin with specificity for Mycobacterium tuberculosis (Mtb) were present in high frequency in TB-experienced children and adolescents. Proliferating memory T cells (CFSElowCD4+CD45RO+) were identified as main source of granulysin and these cells expressed both central and effector memory phenotype. PBMC from study participants after TB drug therapy revealed that granulysin-expressing CD4+ T cells are long-lived, and express several activation and cytotoxicity markers with a proportion of cells being interferon-gamma-positive. In addition, granulysin-expressing T cell lines showed cytolytic activity against Mtb-infected target cells. Conclusions Our data suggest granulysin expression by CD4+ memory T cells as candidate immune marker for TB infection, notably, in childhood and adolescence. PMID:22216262

Inflammation-induced CD69+ Kupffer cell feedback inhibits T cell proliferation via membrane-bound TGF-β1.

PubMed

Zhang, Xiang; Jiang, Zhengping; Gu, Yan; Liu, Yanfang; Cao, Xuetao; Han, Yanmei

2016-12-01

Kupffer cells, tissue-resident macrophage lineage cell, are enriched in vertebrate liver. The mouse F4/80 + Kupffer cells have been subclassified into two subpopulations according to their phenotype and function: CD68 + subpopulation with potent reactive oxygen species (ROS) production and phagocytic capacities, and CD11b + subpopulation with a potent capacity to produce T helper 1 cytokines. In addition, CD11b + Kupffer cells/macrophages may be migrated from the bone marrow or spleen, especially in inflammatory conditions of the liver. For analyzing diverse Kupffer cell subsets, we infected mice with Listeria monocytogenes and analyzed the phenotype variations of hepatic Kupffer cells. During L. monocytogenes infection, hepatic CD69 + Kupffer cells were significantly induced and expanded, and CD69 + Kupffer cells expressed higher level of CD11b, and particularly high level of membrane-bound TGF-β1 (mTGF-β1) but lower level of F4/80. We also found that clodronate liposome administration did not eliminate hepatic CD69 + Kupffer cell subset. We consider the hepatic CD69 + Kupffer cell population corresponds to CD11b + Kupffer cells, the bone marrow-derived population. Hepatic CD69 + Kupffer cells suppressed Ag-nonspecific and OVA-specific CD4 T cell proliferation through mTGF-β1 both in vitro and in vivo, meanwhile, they did not interfere with activation of CD4 T cells. Thus, we have identified a new subset of inflammation-induced CD69 + Kupffer cells which can feedback inhibit CD4 T cell response via cell surface TGF-β1 at the late stage of immune response against infection. CD69 + Kupffer cells may contribute to protect host from pathological injure by preventing overactivation of immune response.

Thiopurine treatment in patients with Crohn's disease leads to a selective reduction of an effector cytotoxic gene expression signature revealed by whole-genome expression profiling.

PubMed

Bouma, G; Baggen, J M; van Bodegraven, A A; Mulder, C J J; Kraal, G; Zwiers, A; Horrevoets, A J; van der Pouw Kraan, C T M

2013-07-01

Crohn's disease (CD) is characterized by chronic inflammation of the gastrointestinal tract, as a result of aberrant activation of the innate immune system through TLR stimulation by bacterial products. The conventional immunosuppressive thiopurine derivatives (azathioprine and mercaptopurine) are used to treat CD. The effects of thiopurines on circulating immune cells and TLR responsiveness are unknown. To obtain a global view of affected gene expression of the immune system in CD patients and the treatment effect of thiopurine derivatives, we performed genome-wide transcriptome analysis on whole blood samples from 20 CD patients in remission, of which 10 patients received thiopurine treatment, compared to 16 healthy controls, before and after TLR4 stimulation with LPS. Several immune abnormalities were observed, including increased baseline interferon activity, while baseline expression of ribosomal genes was reduced. After LPS stimulation, CD patients showed reduced cytokine and chemokine expression. None of these effects were related to treatment. Strikingly, only one highly correlated set of 69 genes was affected by treatment, not influenced by LPS stimulation and consisted of genes reminiscent of effector cytotoxic NK cells. The most reduced cytotoxicity-related gene in CD was the cell surface marker CD160. Concordantly, we could demonstrate an in vivo reduction of circulating CD160(+)CD3(-)CD8(-) cells in CD patients after treatment with thiopurine derivatives in an independent cohort. In conclusion, using genome-wide profiling, we identified a disturbed immune activation status in peripheral blood cells from CD patients and a clear treatment effect of thiopurine derivatives selectively affecting effector cytotoxic CD160-positive cells. Copyright © 2013 Elsevier Ltd. All rights reserved.

CD4 T-helper cell cytokine phenotypes and antibody response following tetanus toxoid booster immunization.

PubMed

Livingston, Kimberly A; Jiang, Xiaowen; Stephensen, Charles B

2013-04-30

Routine methods for enumerating antigen-specific T-helper cells may not identify low-frequency phenotypes such as Th2 cells. We compared methods of evaluating such responses to identify tetanus toxoid- (TT) specific Th1, Th2, Th17 and IL10(+) cells. Eight healthy subjects were given a TT booster vaccination. Blood was drawn before, 3, 7, 14, and 28days after vaccination and peripheral blood mononuclear cells (PBMC) were cultured for 7days with TT, negative control (diluent), and a positive control (Staphylococcus enterotoxin B [SEB]). Activation markers (CD25 and CD69) were measured after 44h (n=8), cytokines in supernatant after 3 and 7days, and intracellular cytokine staining (ICS) of proliferated cells (identified by dye dilution) after 7days (n=6). Vaccination increased TT-specific expression of CD25 and CD69 on CD3(+)CD4(+) lymphocytes, and TT-specific proliferation at 7, 14 and 28days post vaccination. Vaccination induced TT-specific Th1 (IFN-γ, TNF-α, and IL-2) Th2 (IL-13, IL-5, and IL-4), Th17 (IL-17A) and IL-10(+) cells as measured by ICS. TT-specific Th1 cells were the most abundant (12-15% of all TT-specific CD4(+) T-cells) while IL10(+) (1.8%) Th17 (1.1%) and Th2 cells (0.2-0.6%) were less abundant. TT-specific cytokine concentrations in PBMC supernatants followed the same pattern where a TT-specific IL-9 response was also seen. In conclusion, TT booster vaccination induced a broad T-helper cell response. This method of evaluating cytokine phenotypes may be useful in examining the impact of nutrition and environmental conditions on the plasticity of T-helper cell memory responses. Published by Elsevier B.V.

Role of positive selection of thymoma-associated T cells in the pathogenesis of myasthenia gravis.

PubMed

Inada, Keiji; Okumura, Meinoshin; Shiono, Hiroyuki; Inoue, Masayoshi; Kadota, Yoshihisa; Ohta, Mitsunori; Matsuda, Hikaru

2005-06-01

A human thymoma is a thymic epithelial neoplasm and is characterized by its frequent association with myasthenia gravis. The histological characteristic of thymoma is coexistence of a large number of lymphocytes, including CD4(+)CD8(+) double positive T cells, phenotypes of the cortical thymocytes. To elucidate the role of these T lymphocytes in the pathogenesis of thymoma-associated myasthenia gravis, we examined the usage of alphabeta or gammadelta T cell receptor of the T lymphocytes in thymoma in conjunction with the positive selection event. Thymomas were obtained from 28 patients. Nine patients were associated with myasthenia gravis. Lymphocytes were freshly isolated from the tumor tissue and were subjected to four-color flow cytometric analysis. The average proportion of TCRalphabeta(+) cells in thymomas associated with myasthenia gravis was 47.0% and was significantly higher (P = 0.0008) than that without myasthenia gravis (23.4%). Positive selection event was then examined in terms of CD69, a positive selection marker. The mean proportion of TCRalphabeta(+)CD69(+)CD4(+)CD8(-) cells in the myasthenic thymomas (8.22%) was significantly greater (P = 0.015) than the nonmyasthenic thymomas (2.99%). On the other hand, there was not a significant difference in the mean proportion of TCRalphabeta(+)CD69(+)CD4(-)CD8(+) cells between the myasthenic and the nonmyasthenic thymomas. The possible role of development of TCRalphabeta(+) T cells, especially the role of positive selection of TCRalphabeta(+)CD4(+)CD8(-) T cells in thymoma, was suggested in the pathogenesis of thymoma-associated myasthenia gravis.

Enrichment of herpes simplex virus type 2 (HSV-2) reactive mucosal T cells in the human female genital tract.

PubMed

Posavad, C M; Zhao, L; Dong, L; Jin, L; Stevens, C E; Magaret, A S; Johnston, C; Wald, A; Zhu, J; Corey, L; Koelle, D M

2017-09-01

Local mucosal cellular immunity is critical in providing protection from HSV-2. To characterize and quantify HSV-2-reactive mucosal T cells, lymphocytes were isolated from endocervical cytobrush and biopsy specimens from 17 HSV-2-infected women and examined ex vivo for the expression of markers associated with maturation and tissue residency and for functional T-cell responses to HSV-2. Compared with their circulating counterparts, cervix-derived CD4+ and CD8+ T cells were predominantly effector memory T cells (CCR7-/CD45RA-) and the majority expressed CD69, a marker of tissue residency. Co-expression of CD103, another marker of tissue residency, was highest on cervix-derived CD8+ T cells. Functional HSV-2 reactive CD4+ and CD8+ T-cell responses were detected in cervical samples and a median of 17% co-expressed CD103. HSV-2-reactive CD4+ T cells co-expressed IL-2 and were significantly enriched in the cervix compared with blood. This first direct ex vivo documentation of local enrichment of HSV-2-reactive T cells in the human female genital mucosa is consistent with the presence of antigen-specific tissue-resident memory T cells. Ex vivo analysis of these T cells may uncover tissue-specific mechanisms of local control of HSV-2 to assist the development of vaccine strategies that target protective T cells to sites of HSV-2 infection.

Local Inflammatory Cues Regulate Differentiation and Persistence of CD8+ Tissue-Resident Memory T Cells.

PubMed

Bergsbaken, Tessa; Bevan, Michael J; Fink, Pamela J

2017-04-04

Many pathogens initiate infection at mucosal surfaces, and tissue-resident memory T (Trm) cells play an important role in protective immunity, yet the tissue-specific signals that regulate Trm differentiation are poorly defined. During Yersinia infection, CD8 + T cell recruitment to areas of inflammation within the intestine is required for differentiation of the CD103 - CD69 + Trm subset. Intestinal proinflammatory microenvironments have elevated interferon (IFN)-β and interleukin-12 (IL-12), which regulated Trm markers, including CD103. Type I interferon-receptor- or IL-12-receptor-deficient T cells functioned similarly to wild-type (WT) cells during infection; however, the inability of T cells to respond to inflammation resulted in defective differentiation of CD103 - CD69 + Trm cells and reduced Trm persistence. Intestinal macrophages were the main producers of IFN-β and IL-12 during infection, and deletion of CCR2 + IL-12-producing cells reduced the size of the CD103 - Trm population. These data indicate that intestinal inflammation drives phenotypic diversity and abundance of Trm cells for optimal tissue-specific immunity. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Role of immune activation in CD4+ T-cell depletion in HIV-1 infected Indian patients.

PubMed

Vajpayee, M; Kaushik, S; Sreenivas, V; Mojumdar, K; Mendiratta, S; Chauhan, N K

2009-01-01

The correlation of immune activation with CD4(+) depletion and HIV-1 disease progression has been evidenced by several studies involving mainly clade B virus. However, this needs to be investigated in developing countries such as India predominately infected with clade C virus. In a cross-sectional study of 68 antiretroviral treatment naïve, HIV-1 infected Indian patients, we studied the association between CD4(+) T cells, plasma HIV-1 RNA levels, and immune activation markers using unadjusted and adjusted correlative analyses. Significant negative correlations of higher magnitude were observed between the CD4(+) T cell percentages and plasma HIV-1 RNA levels in the study population when adjusted for the effects of immune activation markers. However, the negative association of CD4(+) T cells with immune activation markers remained unaffected when controlled for the effects of plasma HIV-1 RNA levels. Our results support the important role of immune activation in CD4(+) T cell depletion and disease progression during untreated HIV-1 infection.

Intracellular activity of clinical concentrations of phenothiazines including thioridiazine against phagocytosed Staphylococcus aureus.

PubMed

Ordway, Diane; Viveiros, Miguel; Leandro, Clara; Arroz, Maria Jorge; Amaral, Leonard

2002-07-01

The effect of thioridazine (TZ) was studied on the killing activity of human peripheral blood monocyte derived macrophages (HPBMDM) and of human macrophage cell line THP-1 at extracellular concentrations below those achievable clinically. These macrophages have nominal killing activity against bacteria and therefore, would not influence any activity that the compounds may have against intracellular localised Staphylococcus aureus. The results indicated that whereas TZ has an in vitro minimum inhibitory concentration (MIC) against the strains of S. aureus of 18, 0.1 mg/l of TZ in the medium completely inhibits the growth of S. aureus that has been phagocytosed by macrophages. The latter concentration was non-toxic to macrophages, did not cause cellular expression of activation marker CD69 nor induction of CD3+ T cell production of IFN-gamma, but blocked cellular proliferation and down-regulated the production of T cell-derived cytokines (IFN-gamma, IL-5). These results suggest that TZ induces intracellular bactericidal activities independent of the capacity to generate Type 1 responses against S. aureus.

Optimizing of the basophil activation test: Comparison of different basophil identification markers.

PubMed

Eberlein, Bernadette; Hann, Rebekka; Eyerich, Stefanie; Pennino, Davide; Ring, Johannes; Schmidt-Weber, Carsten B; Buters, Jeroen

2015-01-01

Flowcytometric identification of basophils is a prerequisite for measuring activation of basophils with IgE-dependent or IgE-independent stimuli. Aim of this study was to compare different marker combinations in a simultaneous multicolor flowcytometric measurement. Ten patients with a grass pollen allergy and three controls were included in the study. Basophilic cells were gated by using anti-CCR3, anti-IgE, anti-CRTH2, anti-CD203c, and anti-CD3. Cells were activated by a monoclonal anti-FcεRI antibody, N-formyl-methionyl-leucyl-phenylalanine (fMLP), and the allergen extract Phleum pratense. The activation marker anti-CD63 was used. The highest relative number of basophils was found with anti-CCR3+ cells, anti-IgE+ and anti-IgE+ /anti-CD203c+ cells, the lowest with CRTH2+/CD203c+/CD3- cells. A very good and good concordance of CCR3+ cells was seen with CCR3+/CD3- cells and CRTH2+/CD203c+/CD3- cells in all experiments. The contamination of the CCR3+ population with CD3+ cells and the contamination of the IgE+-population with CCR3- cells and CD203- cells were the lowest compared to all other marker combinations. As the highest relative number of basophils was identified by anti-CCR3 followed by the anti-IgE and anti-IgE/antiCD203c positive population in most cases, these markers can generally be recommended for identification of basophils. If a basophil population with very high purity is needed, anti-IgE should be chosen. © 2014 International Clinical Cytometry Society.

Baculovirus directly activates murine NK cells via TLR9.

PubMed

Moriyama, T; Suzuki, T; Chang, M O; Kitajima, M; Takaku, H

2017-04-01

The importance of natural killer (NK) cells in innate immune responses against tumors or viral infections enhances the appeal of NK cell-based immunotherapeutic approaches. We have recently reported that baculovirus (BV)-infected dendritic cells (DCs; BV-DCs) induce antitumor immunity against established tumors in mice. These antitumor effects were CD8 + T-cell and NK cell dependent; however, they were found to be CD4 + T-cell independent. In this study, we investigated the involvement of Toll-like receptor 9 (TLR9) in the process of BV recognition by NK cells. We found that BV directly stimulated NK cells, induced the expression of the activation marker CD69 and promoted interferon-gamma (IFN-γ) production and cytotoxicity. Moreover, TLR9 knockout in mice (tlr9-/- NK cells) inhibited NK cell responses to BV, indicating that TLR9 may have a relevant role in the BV-induced upregulation of NK cell functions. Our data demonstrated for the first time that NK cells directly recognize BV via TLR9, which provides opportunities for the use of this technique as an effective tool for BV-based immunotherapies against malignancies.

T-cell immunity and cytokine production in cosmonauts after long-duration space flights

NASA Astrophysics Data System (ADS)

Morukov, B.; Rykova, M.; Antropova, E.; Berendeeva, T.; Ponomaryov, S.; Larina, I.

2011-04-01

Long-duration spaceflight effects on T-cell immunity and cytokine production were studied in 12 Russian cosmonauts flown onto the International Space Station. Specific assays were performed before launch and after landing and included analysis of peripheral leukocyte distribution, analysis of T-cell phenotype, expression of activation markers, apoptosis, proliferation of T cells in response to a mitogen, concentrations of cytokines in supernatants of cell cultures. Statistically significant increase was observed in leukocytes', lymphocytes', monocytes' and granulocytes' total number, increase in percentage and absolutely number of CD3 +CD4 +-cells, CD4 +CD45RA +-cells and CD4 +CD45RA +/CD4 +CD45RО + ratio, CD4 +CD25 +Bright regulatory cells ( p<0,05) in peripheral blood after landing. T-lymphocytes' capacity to present CD69 and CD25 on its own surfaces was increased for the majority of crewmembers. Analysis of T-cell response to PHA-stimulation in vitro revealed there were some trends toward reduced proliferation of stimulated T-lymphocytes. There was an apparent post flight decrease in secreted IFN-g for the majority of crewmembers and in most instances there was elevation in secreted IL-10. It revealed depression of IFN-g/IL-10 ratio after flight. Correlation analysis according to Spearman's rank correlation test established significant positive correlations ( p<0.05) between cytokine production and T-cell activation (CD25+, CD38+) and negative correlation ( p<0.05) between cytokine production and number of bulk memory CD4+T-cells (CD45RO+). Thus, these results suggest that T-cell dysfunction can be conditioned by cytokine dysbalance and could lead to development of disease after long-duration space flights.

Preparation of Cytokine-activated NK Cells for Use in Adoptive Cell Therapy in Cancer Patients: Protocol Optimization and Therapeutic Potential.

PubMed

van Ostaijen-ten Dam, Monique M; Prins, Henk-Jan; Boerman, Gerharda H; Vervat, Carly; Pende, Daniela; Putter, Hein; Lankester, Arjan; van Tol, Maarten J D; Zwaginga, Jaap J; Schilham, Marco W

2016-01-01

Cell-based immunotherapy using donor-derived natural killer (NK) cells after allogeneic hematopoietic stem cell transplantation may be an attractive treatment of residual leukemia. This study aimed to optimize clinical grade production of a cytokine-activated NK-cell product. NK cells were isolated either by double depletion (CD3(-), CD19(-)) or by sequential depletion and enrichment (CD3(-,) CD56(+)) via CliniMACS from leukapheresis material and cultured in vitro with interleukin (IL)-2 or IL-15. Both NK cell isolation procedures yielded comparable recovery of NK cells and levels of T-cell contamination. After culture with cytokines, the CD3(-)CD56(+) procedure resulted in NK cells of higher purity, that is, less T cells and monocytes, higher viability, and a slightly higher yield than the CD3(-)CD19- procedure. CD69, NKp44, and NKG2A expression were higher on CD3(-)CD56(+) products, whereas lysis of Daudi cells was comparable. Five days of culture led to higher expression of CD69, NKp44, and NKp30 and lysis of K562 and Daudi cell lines. Although CD69 expression and lysis of Daudi cells were slightly higher in cultures with IL-2, T-cell contamination was lower with IL-15. Therefore, further experiments were performed with CD3(-)CD56(+) products cultured with IL-15. Cryopreservation of IL-15-activated NK cells resulted in a loss of cytotoxicity (>92%), whereas thawing of isolated, uncultured NK cells followed by culture with IL-15 yielded cells with about 43% of the original lytic activity. Five-day IL-15-activated NK cells lysed tumor target cell lines and primary leukemic blasts, providing the basis for NK cell–based immunotherapeutic strategies in a clinical setting.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Getachew, Yonas, E-mail: yonas.getachew@utsouthwestern.edu; Cusimano, Frank A.; James, Laura P.

The role of the immune system, specifically NK, NKT and CD3 cells, in acetaminophen (APAP) induced liver injury remains inconsistently defined. In the present study, wild type (C57BL/6J) mice and granzyme B deficient (GrB −/−) mice were treated with acetaminophen to assess the role of the immune system in acute liver injury. Doses of acetaminophen that induced sub lethal liver injury in wild type mice unexpectedly produced fatal hepatotoxicity in granzyme B deficient (GrB −/−) mice. Analysis revealed that GrB −/− mice had an increased population of intrahepatic CD3 (+), CD4 (−), and CD8 (−) lymphocytes expressing the CD69 activationmore » marker and Fas ligand. Depletion of these cells in the GrB −/− and wild type mice made them less susceptible to APAP injury, while depletion of NK1.1 (+) cells or both CD4 (+) and CD8 (+) T cells failed to provide the same hepatoprotection. Transfer of the GrB −/− IHLs further exacerbated liver injury and increased mortality in wild type mice but not in LRP/LPR mice, lacking fas expression. Conclusions: Acetaminophen toxicity is enhanced by the presence of activated, FasL expressing intrahepatic CD3 (+), CD4 (−), CD8 (−), NK1.1 (−) T cells. Depletion of these cells from GrB −/− mice and wild type mice greatly reduces mortality and improves the course of liver injury recovery. - Highlights: • Intrahepatic lymphocytes (IHLs) from GrB −/− mice harbor activated DNT cells. • IHLs from GrB −/− mice exhibit enhanced Fas ligand expression. • Acetaminophen toxicity is enhanced by activated, FasL expressing DNT cells.« less

ESAT-6 and HspX Improve the Effectiveness of BCG to Induce Human Dendritic Cells-Dependent Th1 and NK Cells Activation

PubMed Central

Marongiu, Laura; Donini, Marta; Toffali, Lara; Zenaro, Elena; Dusi, Stefano

2013-01-01

The limited efficacy of the BCG vaccine against tuberculosis is partly due to the missing expression of immunogenic proteins. We analyzed whether the addition to BCG of ESAT-6 and HspX, two Mycobacterium tuberculosis (Mtb) antigens, could enhance its capacity to activate human dendritic cells (DCs). BCG showed a weak ability to induce DC maturation, cytokine release, and CD4+ lymphocytes and NK cells activation. The addition of ESAT-6 or HspX alone to BCG-stimulated DC did not improve these processes, whereas their simultaneous addition enhanced BCG-dependent DC maturation and cytokine release, as well as the ability of BCG-treated DCs to stimulate IFN-γ release and CD69 expression by CD4+ lymphocytes and NK cells. Addition of TLR2-blocking antibody decreased IL-12 release by BCG-stimulated DCs incubated with ESAT-6 and HspX, as well as IFN-γ secretion by CD4+ lymphocytes co-cultured with these cells. Moreover, HspX and ESAT-6 improved the capacity of BCG-treated DCs to induce the expression of memory phenotype marker CD45RO in naïve CD4+ T cells. Our results indicate that ESAT-6 and HspX cooperation enables BCG-treated human DCs to induce T lymphocyte and NK cell-mediated immune responses through TLR2-dependent IL-12 secretion. Therefore ESAT-6 and HspX represent good candidates for improving the effectiveness of BCG vaccination. PMID:24130733

An evaluation of serum soluble CD30 levels and serum CD26 (DPPIV) enzyme activity as markers of type 2 and type 1 cytokines in HIV patients receiving highly active antiretroviral therapy

PubMed Central

Keane, N M; Price, P; Lee, S; Stone, S F; French, M A

2001-01-01

This study evaluates serum CD26 (dipeptidyl peptidase IV, DPPIV) enzyme activity and serum levels of soluble CD30 as markers of T1 and T2 cytokine environments in HIV patients who achieved immune reconstitution after highly active antiretroviral therapy (HAART). Patients who had experienced inflammatory disease associated with pre-existent opportunistic infections after HAART (immune restoration diseases, IRD) were considered separately. Serum sCD30 levels and CD26 (DPPIV) enzyme activity were compared with IFN-γ production by PBMC cultured with cytomegalovirus (CMV) antigen in controls and patient groups. High sCD30 levels were associated with low IFN-γ production after antigenic stimulation in control subjects and, to a lesser extent, in immune reconstituted HIV patients. There was no association between serum CD26 (DPPIV) enzyme activity and IFN-γ production or sCD30 levels. Serum sCD30 levels and CD26 (DPPIV) enzyme activity were significantly increased in immune reconstituted patients with high HIV viral loads. Patients who had experienced CMV retinitis as an IRD had significantly higher sCD30 levels than all other patient groups. Hence, high sCD30 levels may be a marker of a T2 cytokine environment in HIV patients with immune reconstitution and are associated with higher HIV viral loads and a history of CMV associated IRD. PMID:11678906

Dendritic Cell-Based Genetic Immunotherapy for Ovarian Cancer

DTIC Science & Technology

2007-12-01

CAR. CD40 is a surface marker expressed by DCs that plays a crucial role in their maturation and subsequent stimulation of T cells. DC infection with... surface . CD40 is a cell surface marker expressed by DCs, is crucial for their maturation and the subsequent activation of the immune system by the DCs...cell surface . CD40 is a cell surface marker expressed by DCs, is crucial for their maturation and the subsequent activation of the immune system by the

Exhaustion of Activated CD8 T Cells Predicts Disease Progression in Primary HIV-1 Infection

PubMed Central

Hickling, Stephen; Hurst, Jacob; Meyerowitz, Jodi; Willberg, Christian B.; Robinson, Nicola; Brown, Helen; Kinloch, Sabine; Babiker, Abdel; Nwokolo, Nneka; Fox, Julie; Fidler, Sarah; Phillips, Rodney; Frater, John

2016-01-01

The rate at which HIV-1 infected individuals progress to AIDS is highly variable and impacted by T cell immunity. CD8 T cell inhibitory molecules are up-regulated in HIV-1 infection and associate with immune dysfunction. We evaluated participants (n = 122) recruited to the SPARTAC randomised clinical trial to determine whether CD8 T cell exhaustion markers PD-1, Lag-3 and Tim-3 were associated with immune activation and disease progression. Expression of PD-1, Tim-3, Lag-3 and CD38 on CD8 T cells from the closest pre-therapy time-point to seroconversion was measured by flow cytometry, and correlated with surrogate markers of HIV-1 disease (HIV-1 plasma viral load (pVL) and CD4 T cell count) and the trial endpoint (time to CD4 count <350 cells/μl or initiation of antiretroviral therapy). To explore the functional significance of these markers, co-expression of Eomes, T-bet and CD39 was assessed. Expression of PD-1 on CD8 and CD38 CD8 T cells correlated with pVL and CD4 count at baseline, and predicted time to the trial endpoint. Lag-3 expression was associated with pVL but not CD4 count. For all exhaustion markers, expression of CD38 on CD8 T cells increased the strength of associations. In Cox models, progression to the trial endpoint was most marked for PD-1/CD38 co-expressing cells, with evidence for a stronger effect within 12 weeks from confirmed diagnosis of PHI. The effect of PD-1 and Lag-3 expression on CD8 T cells retained statistical significance in Cox proportional hazards models including antiretroviral therapy and CD4 count, but not pVL as co-variants. Expression of ‘exhaustion’ or ‘immune checkpoint’ markers in early HIV-1 infection is associated with clinical progression and is impacted by immune activation and the duration of infection. New markers to identify exhausted T cells and novel interventions to reverse exhaustion may inform the development of novel immunotherapeutic approaches. PMID:27415828

Molecular cloning and characterization of markers and cytokines for equid myeloid cells.

PubMed

Steinbach, Falko; Stark, Robert; Ibrahim, Sherif; Gawad, Eman Abd-El; Ludwig, Hanns; Walter, Jakob; Commandeur, Ulrich; Mauel, Susanne

2005-10-18

The myeloid cell system comprises of monocytes, macrophages (MPhi), dendritic cells (DC), Kupffer cells, osteoclasts or microglia and is also known as the mononuclear phagocytic system (MPS). Essential cytokines to differentiate or activate these cells include GM-CSF or IL-4. Important markers for characterization include CD1, CD14, CD68, CD163 and CD206. All these markers, however, were not cloned or further characterized in equids by use of monoclonal antibodies earlier. To overcome this problem with the present study, two approaches were used. First, we cloned equine cytokines and markers, and second we analyzed cross-reactivity of human homologues or anti-human monoclonal antibodies. For cloning of equine cytokines and markers, we used degenerate primers delineated from other species, or equine-specific primers based on previous information in Genbank. Flow cytometry was used to determine the expression of markers on myeloid cells. Cross-reactivity could be shown for anti-human CD14, CD163 and mannose receptor (CD206) mAbs. Surface markers such as CD1 and CD68 that distinguish MPhi and DC were cloned and sequenced. According to blast homology, equine CD1a and CD1b could be identified and distinguished. With the resulting information, dendritic cells and macrophages of horses may be characterized.

Immunophenotyping does not improve predictivity of the local lymph node assay in mice.

PubMed

Strauss, Volker; Kolle, Susanne N; Honarvar, Naveed; Dammann, Martina; Groeters, Sibylle; Faulhammer, Frank; Landsiedel, Robert; van Ravenzwaay, Bennard

2015-04-01

The local lymph node assay (LLNA) is a regulatory accepted test for the identification of skin sensitizing substances by measuring radioactive thymidine incorporation into the lymph node. However, there is evidence that LLNA is overestimating the sensitization potential of certain substance classes in particular those exerting skin irritation. Some reports describe the additional use of flow cytometry-based immunophenotyping to better discriminate irritants from sensitizing irritants in LLNA. In the present study, the 22 performance standards plus 8 surfactants were assessed using the radioactive LLNA method. In addition, lymph node cells were immunophenotyped to evaluate the specificity of the lymph node response using cell surface markers such as B220 or CD19, CD3, CD4, CD8, I-A(κ) and CD69 with the aim to allow a better discrimination above all between irritants and sensitizers, but also non-irritating sensitizers and non-sensitizers. However, the markers assessed in this study do not sufficiently differentiate between irritants and irritant sensitizers and therefore did not improve the predictive capacity of the LLNA. Copyright © 2014 John Wiley & Sons, Ltd.

Clinical relevance of miR-mediated HLA-G regulation and the associated immune cell infiltration in renal cell carcinoma.

PubMed

Jasinski-Bergner, Simon; Stoehr, Christine; Bukur, Juergen; Massa, Chiara; Braun, Juliane; Hüttelmaier, Stefan; Spath, Verena; Wartenberg, Roland; Legal, Wolfgang; Taubert, Helge; Wach, Sven; Wullich, Bernd; Hartmann, Arndt; Seliger, Barbara

2015-06-01

In human tumors of distinct origin including renal cell carcinoma (RCC), the non-classical human leukocyte antigen G (HLA-G) is frequently expressed, thereby inhibiting the cytotoxic activity of T and natural killer (NK) cells. Recent studies demonstrated a strong post-transcriptional gene regulation of the HLA-G by miR-152, -148A, -148B and -133A. Standard methods were applied to characterize the expression and function of HLA-G, HLA-G-regulatory microRNAs (miRs) and the immune cell infiltration in 453 RCC lesions using a tissue microarray and five RCC cell lines linking these results to clinical parameters. Direct interactions with HLA-G regulatory miRs and the HLA-G 3' untranslated region (UTR) were detected and the affinities of these different miRs to the HLA-G 3'-UTR compared. qPCR analyses and immunohistochemical staining revealed an inverse expression of miR-148A and -133A with the HLA-G protein in situ and in vitro . Stable miR overexpression caused a downregulation of HLA-G protein enhancing the NK and LAK cell-mediated cytotoxicity in in vitro CD107a activation assays revealing a HLA-G-dependent cytotoxic activity of immune effector cells. A significant higher frequency of CD3 + /CD8 + T cell lymphocytes, but no differences in the activation markers CD69, CD25 or in the presence of CD56 + , FoxP3 + and CD4 + immune cells were detected in HLA-G + compared to HLA-G - RCC lesions. This could be associated with higher WHO grade, but not with a disease-specific survival. These data suggest a miR-mediated control of HLA-G expression in RCC, which is associated with a distinct pattern of immune cell infiltration.

Differential Dynamics of CD4+ and CD8+ T-Lymphocyte Proliferation and Activation in Acute Simian Immunodeficiency Virus Infection

PubMed Central

Kaur, Amitinder; Hale, Corrina L.; Ramanujan, Saroja; Jain, Rakesh K.; Johnson, R. Paul

2000-01-01

Although lymphocyte turnover in chronic human immunodeficiency virus and simian immunodeficiency virus (SIV) infection has been extensively studied, there is little information on turnover in acute infection. We carried out a prospective kinetic analysis of lymphocyte proliferation in 13 rhesus macaques inoculated with pathogenic SIV. A short-lived dramatic increase in circulating Ki-67+ lymphocytes observed at 1 to 4 weeks was temporally related to the onset of SIV replication. A 5- to 10-fold increase in Ki-67+ CD8+ T lymphocytes and a 2- to 3-fold increase in Ki-67+ CD3− CD8+ natural killer cells accounted for >85% of proliferating lymphocytes at peak proliferation. In contrast, there was little change in the percentage of Ki-67+ CD4+ T lymphocytes during acute infection, although transient increases in Ki-67− and Ki-67+ CD4+ T lymphocytes expressing CD69, Fas, and HLA-DR were observed. A two- to fourfold decline in CD4+ T lymphocytes expressing CD25 and CD69 was seen later in SIV infection. The majority of Ki-67+ CD8+ T lymphocytes were phenotypically CD45RA− CD49dhi Fashi CD25− CD69− CD28− HLA-DR− and persisted at levels twofold above baseline 6 months after SIV infection. Increased CD8+ T-lymphocyte proliferation was associated with cell expansion, paralleled the onset of SIV-specific cytotoxic T-lymphocyte activity, and had an oligoclonal component. Thus, divergent patterns of proliferation and activation are exhibited by CD4+ and CD8+ T lymphocytes in early SIV infection and may determine how these cells are differentially affected in AIDS. PMID:10954541

Acidic conditions induce the suppression of CD86 and CD54 expression in THP-1 cells.

PubMed

Mitachi, Takafumi; Mezaki, Minori; Yamashita, Kunihiko; Itagaki, Hiroshi

2018-01-01

To evaluate the sensitization potential of chemicals in cosmetics, using non-animal methods, a number of in vitro safety tests have been designed. Current assays are based on the expression of cell surface markers, such as CD86 and CD54, which are associated with the activation of dendritic cells, in skin sensitization tests. However, these markers are influenced by culture conditions through activating danger signals. In this study, we investigated the relationship between extracellular pH and the expression of the skin sensitization test human cell line activation test (h-CLAT) markers CD86 and CD54. We measured expression levels after THP-1 cells were exposed to representative contact allergens, i.e., 2,4-dinitrochlorobenzene and imidazolidinyl urea, under acidic conditions. These conditions were set by exposure to hydrochloric acid, lactic acid, and citric acid. An acidic extracellular pH (6-7) suppressed the augmentation of CD86 and CD54 levels by the sensitizer. Additionally, when the CD86/CD54 expression levels were suppressed, a reduction in the intracellular pH was confirmed. Furthermore, we observed that Na + /H + exchanger 1 (NHE-1), a protein that contributes to the regulation of extracellular/intracellular pH, is involved in CD86 and CD54 expression. These findings suggest that the extracellular/intracellular pH has substantial effects on in vitro skin sensitization markers and should be considered in evaluations of the safety of mixtures and commercial products in the future.

Fetal bovine bone marrow is a rich source of CD34+ hematopoietic progenitors with myelo-monocytic colony-forming activity.

PubMed

Pessa-Morikawa, Tiina; Niku, Mikael; Iivanainen, Antti

2012-03-01

The CD34 glycoprotein is an important marker of hematopoietic stem cells. We used a polyclonal rabbit anti-bovine CD34 antibody to stain fetal and adult bovine bone marrow cells. Flow cytometry revealed a low side scatter (SSC(low)) population of cells that were CD34(+) but negative for leukocyte lineage markers CD11b, CD14 or CD2. Hematopoietic colony assays with CD34(+) and CD34(-) bone marrow cells suggested that the colony-forming potential in SSC(low) bone marrow cells was confined to the CD34(+) fraction. In contrast, this population was not enriched for cells expressing high aldehyde dehydrogenase activity, a metabolic marker that has been used to characterize hematopoietic stem cells. Thus, the CD34 antigen can be used to identify and isolate bovine bone marrow cells exhibiting clonogenic potential in vitro. Moreover, the proportion of CD34(+) cells is very high in fetal bovine bone marrow, indicating it as a rich source of hematopoietic progenitors. Copyright © 2011 Elsevier Ltd. All rights reserved.

MHC Class II and CD9 in human eosinophils localize to detergent-resistant membrane microdomains.

PubMed

Akuthota, Praveen; Melo, Rossana C N; Spencer, Lisa A; Weller, Peter F

2012-02-01

Eosinophils function in murine allergic airways inflammation as professional antigen-presenting cells (APCs). In murine professional APC cell types, optimal functioning of MHC Class II depends on its lateral association in plasma membranes and colocalization with the tetraspanin CD9 into detergent-resistant membrane microdomains (DRMs). With human eosinophils, we evaluated the localization of MHC Class II (HLA-DR) to DRMs and the functional significance of such localization. In granulocyte-macrophage colony-stimulating factor-stimulated human eosinophils, antibody cross-linked HLA-DR colocalized by immunofluorescence microscopy focally on plasma membranes with CD9 and the DRM marker ganglioside GM1. In addition, HLA-DR coimmunoprecipitates with CD9 after chemical cross-linking of CD9. HLA-DR and CD9 were localized by Western blotting in eosinophil DRM subcellular fractions. DRM disruption with the cholesterol-depleting agent methyl-β-cyclodextrin decreased eosinophil surface expression of HLA-DR and CD9. We show that CD9 is abundant on the surface of eosinophils, presenting the first electron microscopy data of the ultrastructural immunolocalization of CD9 in human eosinophils. Disruption of HLA-DR-containing DRMs decreased the ability of superantigen-loaded human eosinophils to stimulate CD4(+) T-cell activation (CD69 expression), proliferation, and cytokine production. Our results, which demonstrate that eosinophil MHC Class II localizes to DRMs in association with CD9 in a functionally significant manner, represent a novel insight into the organization of the antigen presentation complex of human eosinophils.

MHC Class II and CD9 in Human Eosinophils Localize to Detergent-Resistant Membrane Microdomains

PubMed Central

Akuthota, Praveen; Melo, Rossana C. N.; Spencer, Lisa A.

2012-01-01

Eosinophils function in murine allergic airways inflammation as professional antigen-presenting cells (APCs). In murine professional APC cell types, optimal functioning of MHC Class II depends on its lateral association in plasma membranes and colocalization with the tetraspanin CD9 into detergent-resistant membrane microdomains (DRMs). With human eosinophils, we evaluated the localization of MHC Class II (HLA-DR) to DRMs and the functional significance of such localization. In granulocyte-macrophage colony-stimulating factor–stimulated human eosinophils, antibody cross-linked HLA-DR colocalized by immunofluorescence microscopy focally on plasma membranes with CD9 and the DRM marker ganglioside GM1. In addition, HLA-DR coimmunoprecipitates with CD9 after chemical cross-linking of CD9. HLA-DR and CD9 were localized by Western blotting in eosinophil DRM subcellular fractions. DRM disruption with the cholesterol-depleting agent methyl-β-cyclodextrin decreased eosinophil surface expression of HLA-DR and CD9. We show that CD9 is abundant on the surface of eosinophils, presenting the first electron microscopy data of the ultrastructural immunolocalization of CD9 in human eosinophils. Disruption of HLA-DR–containing DRMs decreased the ability of superantigen-loaded human eosinophils to stimulate CD4+ T-cell activation (CD69 expression), proliferation, and cytokine production. Our results, which demonstrate that eosinophil MHC Class II localizes to DRMs in association with CD9 in a functionally significant manner, represent a novel insight into the organization of the antigen presentation complex of human eosinophils. PMID:21885678

LMW Heparin Prevents Increased Kidney Expression of Proinflammatory Mediators in (NZBxNZW)F1 Mice

PubMed Central

Kanapathippillai, Premasany; Rekvig, Ole Petter; Fenton, Kristin Andreassen

2013-01-01

We have previously demonstrated that continuous infusion of low molecular weight (LMW) heparin delays autoantibody production and development of lupus nephritis in (NZBxNZW)F1 (B/W) mice. In this study we investigated the effect of LMW heparin on renal cytokine and chemokine expression and on nucleosome-mediated activation of nucleosome-specific splenocytes. Total mRNA extracted from kidneys of heparin-treated or -untreated B/W mice was analysed by qPCR for the expression of several cytokines, chemokines, and Toll-like receptors. Splenocytes taken from B/W mice were stimulated with nucleosomes with or without the presence of heparin. Splenocyte cell proliferation as thymidine incorporation and the expression of costimulatory molecules and cell activation markers were measured. Heparin treatment of B/W mice reduced the in vivo expression of CCR2, IL1β, and TLR7 compared to untreated B/W mice. Nucleosome-induced cell proliferation of splenocytes was not influenced by heparin. The expression of CD80, CD86, CD69, CD25, CTLA-4, and TLR 2, 7, 8, and 9 was upregulated upon stimulation by nucleosomes, irrespective of whether heparin was added to the cell culture or not. In conclusion, treatment with heparin lowers the kidney expression of proinflammatory mediators in B/W mice but does not affect nucleosomal activation of splenocytes. PMID:24151519

Monitoring non-immediate allergic reactions to iodine contrast media

PubMed Central

Torres, M J; Mayorga, C; Cornejo-Garcia, J A; Lopez, S; Chaves, P; Rondon, C; Fernandez, T; Blanca, M

2008-01-01

Non-immediate reactions to iodine contrast media (ICM) affect 2–5% of patients receiving these agents. We studied the immunological mechanisms involved in patients with a confirmed non-immediate reaction, maculopapular exanthema, after administration of ICM. The diagnosis was carried out by skin testing or drug provocation test. The immunological study was performed in sequential peripheral blood mononuclear cells taken from the onset of the reaction by flow cytometry and in skin biopsy by immunohistochemistry, with specific recognition by the lymphocyte transformation test (LTT) with different ICM. Flow cytometry showed an increase in the different activation markers [CD69, CD25 and human leucocyte antigen D-related (HLA-DR)] and the skin homing receptor [cutaneous lymphocyte-associated antigen (CLA)] in CD4 lymphocytes, whereas perforin was higher in the CD8 lymphocytes. The skin biopsy showed a perivascular mononuclear infiltrate composed of CD4 lymphocytes, expressing CD25, HLA-DR and CLA, with eosinophils. Intradermal skin tests and the LTT were positive to several ICM, including the culprit agent in four and three patients, respectively, with negative results in all 10 tolerant controls. We showed that a specific immunological mechanism was implicated in patients with non-immediate reactions to ICM. Moreover, the positive results in skin tests and lymphocyte proliferation tests indicated that an important cross-reactivity exists. PMID:18341616

Impairment of Circulating CD4⁺CD25⁺GARP⁺ regulatory T cells in patients with acute coronary syndrome.

PubMed

Meng, Kai; Zhang, Wei; Zhong, Yucheng; Mao, Xiaobo; Lin, Yingzhong; Huang, Ying; Lang, Mingjian; Peng, Yudong; Zhu, Zhengfeng; Liu, Yuzhou; Zhao, Xiaoqi; Yu, Kunwu; Wu, Bangwei; Ji, Qingwei; Zeng, Qiutang

2014-01-01

Atherosclerosis (AS) is an inflammatory and immune disease. Regulatory T cells (Tregs) suppress the activation of T cells and have been shown to play a protective role during the pathogenesis of AS. However, specific markers for Tregs are lacking. Recently, glycoprotein A repetitions predominant (GARP) was discovered as a specific marker of activated Tregs, and we therefore utilized GARP as a specific surface marker for Tregs in the current study. To assess whether GARP(+) Tregs are downregulated in patients with acute coronary syndrome (ACS), we examined CD4(+)CD25(+)GARP(+) T cell frequencies as well as their associated cytokines and suppressive function. Additionally, we compared GARP expression to that of FOXP3, which may be more sensitive as a marker of activated Tregs in patients with ACS. Patients with ACS demonstrated a significant decrease in circulating CD4(+)CD25(+)GARP(+) Tregs. Moreover, the suppressive function of Tregs and levels of related cytokines were also impaired in ACS patients compared to those with stable angina (SA) or normal coronary artery (NCA). Additionally, after TCR stimulation, peripheral blood mononuclear cells (PBMCs) from patients with ACS exhibited a decrease in CD4(+)CD25(+)GARP(+) Tregs. These fnding indicate that circulating CD4(+)CD25(+)GARP(+) Tregs are impaired in patients withACS. Thus, targeting GARP may promote the protective function of Tregs in ACS. © 2014 S. Karger AG, Basel.

Evaluation of interleukin 12 and CD56+ lymphocyte cells in pediatric hematopoietic stem cell transplantation for early diagnosis of acute graft versus host disease.

PubMed

Tootoonchian, Raziyeh; Pak, Fatemeh; Ardekani, Ali M; Sehati, Nasrin; Abedi-Valugerdi, Manuchehr; Kokhaei, Parviz

2016-11-01

The present study tried to explain CD56+ lymphocyte cells activities and possible prognostic role of these cells in Graft-Versus-Host-Disease (GVHD). The role of IL-12 activation and function is of interest in this study. Peripheral blood samples of 51 Hematopoietic Stem Cell Transplantation (HSCT) recipients collected at before (day -8) and after (days 7 and 14). PBMC were collected by Ficoll separation and analyzed by Flow Cytometry using triple antibody (CD45-PerCP, CD56-FITC, and CD69-PE staining and control antibody. Levels of the cytokine IL-12 in the patient's serum were evaluated by ELISA. Percentage of CD56+ lymphocytes (CD56+ bright ) cells was significantly increased at day 14 in patients with acute GVHD and percentage of lymphocytes expressing CD69 was significantly increased at days 7 and 14 posts HSCT in patients with acute GVHD in comparison to those in non-GVHD patients. Baseline serum IL-12 levels (pre-HSCT, day -8) were significantly higher in those HSCT recipients who did not develop GVHD. This study showed that post-transplant CD56+ lymphocytes and pre-transplant serum levels of IL-12 play significant roles in the induction of and protection against GVHD, respectively. The increase in the percentage of CD69+ cells indicates the activation of lymphocyte in acute GVHD group. Copyright © 2016 Elsevier B.V. All rights reserved.

Altered Cytokine Production By Specific Human Peripheral Blood Cell Subsets Immediately Following Spaceflight

NASA Technical Reports Server (NTRS)

Crucian, Brian E.; Cubbage, Michael L.; Sams, Clarence F.

1999-01-01

In this study, we have attempted to combine standard immunological assays with the cellular resolving power of the flow cytometer to positively identify the specific cell types involved in spaceflight-induced immune alterations. We have obtained whole blood samples from 27 astronauts collected at three timepoints (L-10, R+0 and R+3) surrounding four recent space shuttle missions. The duration of these missions ranged from 10 to 18 days. Assays performed included serum/urine cortisol, comprehensive subset phenotyping, assessment of cellular activation markers and intracellular cytokine production following mitogenic stimulation. Absolute levels of peripheral granulocytes were significantly elevated following spaceflight, but the levels of circulating lymphocytes and monocytes were unchanged. Lymphocyte subset analysis demonstrated trends towards a decreased percentage of T cells and an increased percentage of B cells. Nearly all of the astronauts exhibited an increased CD4:CD8 ratio, which was dramatic in some individuals. Assessment of memory (CD45RA+) vs. naive (CD45RO+) CD4+ T cell subsets was more ambiguous, with subjects tending to group more as a flight crew. All subjects from one mission demonstrated an increased CD45RA:CD45RO ratio, while all subjects from another Mission demonstrated a decreased ratio. While no significant trend was seen in the monocyte population as defined by scatter, a decreased percentage of the CD14+ CD16+ monocyte subset was seen following spaceflight in all subjects tested. In general, most of the cellular changes described above which were assessed at R+O and compared to L-10 trended to pre-flight levels by R+3. Although no significant differences were seen in the expression of the cellular activation markers CD69 and CD25 following exposure to microgravity, significant alterations were seen in cytokine production in response to mitogenic activation for specific subsets. T cell (CD3+) production of IL-2 was significantly decreased after at R+O as was IL-2 production by both CD4+ and CD8+ T cell subsets for most subjects. Production of IFN(sub gamma) did not appear to be affected by microgravity exposure in either T cells in general or in the CD8+ T cell subset. There was a spaceflight-induced decrease in IFN(sub gamma) production in the CD4+ T cell subset, however it did not reach statistical significance. Serum and urine stress-hormone analysis indicated significant physiologic stresses in astronauts following spaceflight. In summary, these results demonstrate alterations in the peripheral immune system of astronauts immediately after spaceflight of 10 to 18 days duration and support continued research regarding microgravity and immunology (including in-flight sampling) prior to routine long-term spaceflight for astronauts.

Effect of dim and bright light exposure on some immunological parameters measured under thermal neutral conditions.

PubMed

Hyun, Ki-Ja; Kondo, Masayuki; Koh, Taichin; Tokura, Hiromi; Tamotsu, Satoshi; Oishi, Tadashi

2005-01-01

This study assesses the effects of ambient light conditions, under a thermoneutral environment, on selected immunological parameters of 7 healthy young women (aged 19 to 22 yrs). Subjects entered the bioclimatic chamber at 11: 00 h, controlled at 26 degrees C and 60% relative humidity, a "neutral climate". They lead a well-regulated life in the climatic chamber (pre-condition) while exposed to dim (200 lux) or, on the next day, bright (5000 lux) light between 06 : 00 to 12 : 00 h. Just before the end of each period of light exposure, a blood sample was taken for later immunological assay of white blood cell count (WBC), phagocytosis, interferon-gamma (IFN-gamma), interleukin-4 (IL-4), CD69 T cells (CD69), CD4+CD25+ T cells (CD4+CD25+), and transforming growth factor-beta 1 (TGF-beta1). The results, when compared with the pre-condition, were as follows: 1) CD69 and IFN-gamma increased during normal conditions without thermal stress under dim light; 2) WBC increased and IL-4 decreased under bright light; 3) as shown by the highly significant decrease of TGF-beta1, the immune system was activated under bright light; 4) phagocytosis tended to increase under bright light exposure; 5) CD69 and IFN-gamma were significantly higher, and CD4+CD25+ tended to decrease under bright light; 6) phagocytosis tended to be lower and TGF-beta1 significantly higher under dim light, indicating a decline of immune system function. Taken together, this preliminary single time-point sampling study infers that some parameters are activated (CD69) while others are attenuated (phagocytosis, TGF-beta1) according to the environmental light intensity, dim vs. bright, in women adhering to a standardized routine in the absence of thermal stress. These findings are discussed in terms of inhibition of the sympathetic and excitation of the parasympathetic nervous system under the influence of life-style regularity and daytime bright light exposure.

Epigenetic Metabolite Acetate Inhibits Class I/II Histone Deacetylases, Promotes Histone Acetylation, and Increases HIV-1 Integration in CD4+ T Cells.

PubMed

Bolduc, Jean-François; Hany, Laurent; Barat, Corinne; Ouellet, Michel; Tremblay, Michel J

2017-08-15

In this study, we investigated the effect of acetate, the most concentrated short-chain fatty acid (SCFA) in the gut and bloodstream, on the susceptibility of primary human CD4 + T cells to HIV-1 infection. We report that HIV-1 replication is increased in CD3/CD28-costimulated CD4 + T cells upon acetate treatment. This enhancing effect correlates with increased expression of the early activation marker CD69 and impaired class I/II histone deacetylase (HDAC) activity. In addition, acetate enhances acetylation of histones H3 and H4 and augments HIV-1 integration into the genome of CD4 + T cells. Thus, we propose that upon antigen presentation, acetate influences class I/II HDAC activity that transforms condensed chromatin into a more relaxed structure. This event leads to a higher level of viral integration and enhanced HIV-1 production. In line with previous studies showing reactivation of latent HIV-1 by SCFAs, we provide evidence that acetate can also increase the susceptibility of primary human CD4 + T cells to productive HIV-1 infection. IMPORTANCE Alterations in the fecal microbiota and intestinal epithelial damage involved in the gastrointestinal disorder associated with HIV-1 infection result in microbial translocation that leads to disease progression and virus-related comorbidities. Indeed, notably via production of short-chain fatty acids, bacteria migrating from the lumen to the intestinal mucosa could influence HIV-1 replication by epigenetic regulatory mechanisms, such as histone acetylation. We demonstrate that acetate enhances virus production in primary human CD4 + T cells. Moreover, we report that acetate impairs class I/II histone deacetylase activity and increases integration of HIV-1 DNA into the host genome. Therefore, it can be postulated that bacterial metabolites such as acetate modulate HIV-1-mediated disease progression. Copyright © 2017 American Society for Microbiology.

Donor Cell Composition and Reactivity Predict Risk of Acute Graft-versus-Host Disease after Allogeneic Hematopoietic Stem Cell Transplantation.

PubMed

Sairafi, Darius; Stikvoort, Arwen; Gertow, Jens; Mattsson, Jonas; Uhlin, Michael

2016-01-01

Background . Graft-versus-host disease (GVHD) is a serious complication after allogeneic hematopoietic stem cell transplantation (HSCT). We designed a functional assay for assessment of individual risk for acute GVHD. Study Design and Methods . Blood samples were collected from patients and donors before HSCT. Two groups of seven patients each were selected, one in which individuals developed acute GVHD grades II-IV and one in which none showed any clinical signs of GVHD. Peripheral blood mononuclear cells (PBMCs) isolated from donors were incubated in mixed lymphocyte cultures (MLCs) with recipient PBMCs. The cells were characterized by flow cytometry before and after MLC. Results . Samples from donors in the GVHD group contained significantly lower frequencies of naïve γδ T-cells and T-cells expressing NK-cell markers CD56 and CD94. Donor samples in this group also exhibited lower frequencies of naïve CD95 + T-cells compared to controls. After MLC, there were dissimilarities in the CD4/CD8 T-cell ratio and frequency of CD69 + T-cells between the two patient groups, with the non-GVHD group showing higher frequencies of CD8 + and CD69 + T-cells. Conclusion . We conclude that a thorough flow cytometric analysis of donor cells for phenotype and allogeneic reactivity may be of value when assessing pretransplant risk for severe acute GVHD.

Lipopolysaccharide-binding protein, lipopolysaccharide, and soluble CD14 in sepsis of critically ill neonates and children.

PubMed

Pavcnik-Arnol, Maja; Hojker, Sergej; Derganc, Metka

2007-06-01

To compare the diagnostic accuracy of lipopolysaccharide-binding protein (LBP) for sepsis in critically ill neonates and children with the two markers participating in the same inflammatory pathway, lipopolysaccharide and soluble CD14. Prospective, observational study in a multidisciplinary neonatal and pediatric intensive care unit. 47 critically ill neonates and 49 critically ill children with systemic inflammatory response syndrome (SIRS) and suspected sepsis, classified into two groups: those with and those without sepsis. Serum LBP, lipopolysaccharide, soluble CD14, C-reactive protein, and procalcitonin were measured on 2 consecutive days. The area under the receiver operating characteristic curve (AUC), sensitivity, specificity, and predictive values were evaluated. AUC for LBP on the first day of suspected infection was 0.97 in neonates aged under 48 h, 0.93 in neonates over 48 h and 0.82 in children. AUCs for lipopolysaccharide and soluble CD14 were 0.77 and 0.74 in neonates under 48 h, 0.53 and 0.76 in neonates over 48 h, and 0.72 and 0.53 in children. AUCs for procalcitonin and C-reactive protein were 0.65 and 0.89 in neonates under 48 h, 0.65 and 0.91 in neonates over 48 h, and 0.76 and 0.69 in children. In critically ill neonates and children LBP concentration on the first day of suspected sepsis is a better marker of sepsis than lipopolysaccharide, soluble CD14, procalcitonin, and in neonates younger than 48 h and children, also a better marker than C-reactive protein. Lipopolysaccharide and soluble CD14 are not suitable markers for the differentiation of infectious and noninfectious SIRS.

T Follicular Helper Cell-Germinal Center B Cell Interaction Strength Regulates Entry into Plasma Cell or Recycling Germinal Center Cell Fate.

PubMed

Ise, Wataru; Fujii, Kentaro; Shiroguchi, Katsuyuki; Ito, Ayako; Kometani, Kohei; Takeda, Kiyoshi; Kawakami, Eiryo; Yamashita, Kazuo; Suzuki, Kazuhiro; Okada, Takaharu; Kurosaki, Tomohiro

2018-04-17

Higher- or lower-affinity germinal center (GC) B cells are directed either to plasma cell or GC recycling, respectively; however, how commitment to the plasma cell fate takes place is unclear. We found that a population of light zone (LZ) GC cells, Bcl6 lo CD69 hi expressing a transcription factor IRF4 and higher-affinity B cell receptors (BCRs) or Bcl6 hi CD69 hi with lower-affinity BCRs, favored the plasma cell or recycling GC cell fate, respectively. Mechanistically, CD40 acted as a dose-dependent regulator for Bcl6 lo CD69 hi cell formation. Furthermore, we found that expression of intercellular adhesion molecule 1 (ICAM-1) and signaling lymphocytic activation molecule (SLAM) in Bcl6 lo CD69 hi cells was higher than in Bcl6 hi CD69 hi cells, thereby affording more stable T follicular helper (Tfh)-GC B cell contacts. These data support a model whereby commitment to the plasma cell begins in the GC and suggest that stability of Tfh-GC B cell contacts is key for plasma cell-prone GC cell formation. Copyright © 2018. Published by Elsevier Inc.

Transcriptional Networks in Single Perivascular Cells Sorted from Human Adipose Tissue Reveal a Hierarchy of Mesenchymal Stem Cells.

PubMed

Hardy, W Reef; Moldovan, Nicanor I; Moldovan, Leni; Livak, Kenneth J; Datta, Krishna; Goswami, Chirayu; Corselli, Mirko; Traktuev, Dmitry O; Murray, Iain R; Péault, Bruno; March, Keith

2017-05-01

Adipose tissue is a rich source of multipotent mesenchymal stem-like cells, located in the perivascular niche. Based on their surface markers, these have been assigned to two main categories: CD31 - /CD45 - /CD34 + /CD146 - cells (adventitial stromal/stem cells [ASCs]) and CD31 - /CD45 - /CD34 - /CD146 + cells (pericytes [PCs]). These populations display heterogeneity of unknown significance. We hypothesized that aldehyde dehydrogenase (ALDH) activity, a functional marker of primitivity, could help to better define ASC and PC subclasses. To this end, the stromal vascular fraction from a human lipoaspirate was simultaneously stained with fluorescent antibodies to CD31, CD45, CD34, and CD146 antigens and the ALDH substrate Aldefluor, then sorted by fluorescence-activated cell sorting. Individual ASCs (n = 67) and PCs (n = 73) selected from the extremities of the ALDH-staining spectrum were transcriptionally profiled by Fluidigm single-cell quantitative polymerase chain reaction for a predefined set (n = 429) of marker genes. To these single-cell data, we applied differential expression and principal component and clustering analysis, as well as an original gene coexpression network reconstruction algorithm. Despite the stochasticity at the single-cell level, covariation of gene expression analysis yielded multiple network connectivity parameters suggesting that these perivascular progenitor cell subclasses possess the following order of maturity: (a) ALDH br ASC (most primitive); (b) ALDH dim ASC; (c) ALDH br PC; (d) ALDH dim PC (least primitive). This order was independently supported by specific combinations of class-specific expressed genes and further confirmed by the analysis of associated signaling pathways. In conclusion, single-cell transcriptional analysis of four populations isolated from fat by surface markers and enzyme activity suggests a developmental hierarchy among perivascular mesenchymal stem cells supported by markers and coexpression networks. Stem Cells 2017;35:1273-1289. © 2017 AlphaMed Press.

Involvement of mast cells by the malignant process in patients with Philadelphia chromosome negative myeloproliferative neoplasms.

PubMed

Wang, J; Ishii, T; Zhang, W; Sozer, S; Dai, Y; Mascarenhas, J; Najfeld, V; Zhao, Z J; Hoffman, R; Wisch, N; Xu, M

2009-09-01

The Philadelphia chromosome negative myeloproliferative neoplasms (MPNs) are clonal hematologic malignancies frequently characterized by a mutation in JAK2 (JAK2V617F). Peripheral blood (PB) CD34(+) cells from patients with polycythemia vera (PV) and primary myelofibrosis (PMF) generated in vitro significantly fewer mast cells (MCs) than normal PB CD34(+) cells. The numbers of MC progenitors assayed from MPN CD34(+) cells were, however, similar to that assayed from normal CD34(+) cells. A higher percentage of the cultured MPN MCs expressed FcvarepsilonRIalpha, CD63 and CD69 than normal MCs, suggesting that cultured MPN MCs are associated with an increased state of MC activation. Further analysis showed that a higher proportion of cultured PV and PMF MCs underwent apoptosis in vitro. By using JAK2V617F, MplW515L and chromosomal abnormalities as clonality markers, we showed that the malignant process involved MPN MCs. JAK2V617F-positive MC colonies were assayable from the PB CD34(+) cells of each of the 17 JAK2V617F positive MPN patients studied. Furthermore, erlotinib, a JAK2 inhibitor, was able to inhibit JAK2V617F-positive PV MC progenitor cells, indicating that malignant MC progenitor cells are a potential cellular target for such JAK2 inhibitor-directed therapy.

IL-2high tissue-resident T cells in the human liver: Sentinels for hepatotropic infection

PubMed Central

Davies, Jessica; Hansi, Navjyot; Easom, Nicholas J.W.; Burton, Alice R.; Stegmann, Kerstin A.; Schurich, Anna; Swadling, Leo; Male, Victoria; Luong, TuVinh; Davidson, Brian R.; Kennedy, Patrick T.F.

2017-01-01

The liver provides a tolerogenic immune niche exploited by several highly prevalent pathogens as well as by primary and metastatic tumors. We have sampled healthy and hepatitis B virus (HBV)–infected human livers to probe for a subset of T cells specialized to overcome local constraints and mediate immunity. We characterize a population of T-betloEomesloBlimp-1hiHobitlo T cells found within the intrahepatic but not the circulating memory CD8 T cell pool expressing liver-homing/retention markers (CD69+CD103+ CXCR6+CXCR3+). These tissue-resident memory T cells (TRM) are preferentially expanded in patients with partial immune control of HBV infection and can remain in the liver after the resolution of infection, including compartmentalized responses against epitopes within all major HBV proteins. Sequential IL-15 or antigen exposure followed by TGFβ induces liver-adapted TRM, including their signature high expression of exhaustion markers PD-1 and CD39. We suggest that these inhibitory molecules, together with paradoxically robust, rapid, cell-autonomous IL-2 and IFNγ production, equip liver CD8 TRM to survive while exerting local noncytolytic hepatic immunosurveillance. PMID:28526759

Combined Inhibition of Complement and CD14 Attenuates Bacteria-Induced Inflammation in Human Whole Blood More Efficiently Than Antagonizing the Toll-like Receptor 4–MD2 Complex

PubMed Central

Gustavsen, Alice; Nymo, Stig; Landsem, Anne; Christiansen, Dorte; Ryan, Liv; Husebye, Harald; Lau, Corinna; Pischke, Søren E.; Lambris, John D.; Espevik, Terje; Mollnes, Tom E.

2016-01-01

Background. Single inhibition of the Toll-like receptor 4 (TLR4)–MD2 complex failed in treatment of sepsis. CD14 is a coreceptor for several TLRs, including TLR4 and TLR2. The aim of this study was to investigate the effect of single TLR4-MD2 inhibition by using eritoran, compared with the effect of CD14 inhibition alone and combined with the C3 complement inhibitor compstatin (Cp40), on the bacteria-induced inflammatory response in human whole blood. Methods. Cytokines were measured by multiplex technology, and leukocyte activation markers CD11b and CD35 were measured by flow cytometry. Results. Lipopolysaccharide (LPS)–induced inflammatory markers were efficiently abolished by both anti-CD14 and eritoran. Anti-CD14 was significantly more effective than eritoran in inhibiting LPS-binding to HEK-293E cells transfected with CD14 and Escherichia coli–induced upregulation of monocyte activation markers (P < .01). Combining Cp40 with anti-CD14 was significantly more effective than combining Cp40 with eritoran in reducing E. coli–induced interleukin 6 (P < .05) and monocyte activation markers induced by both E. coli (P < .001) and Staphylococcus aureus (P < .01). Combining CP40 with anti-CD14 was more efficient than eritoran alone for 18 of 20 bacteria-induced inflammatory responses (mean P < .0001). Conclusions. Whole bacteria–induced inflammation was inhibited more efficiently by anti-CD14 than by eritoran, particularly when combined with complement inhibition. Combined CD14 and complement inhibition may prove a promising treatment strategy for bacterial sepsis. PMID:26977050

Altered CD19/CD22 balance in Egyptian children and adolescents with systemic lupus erythematosus.

PubMed

El-Sayed, Zeinab A; Ragab, Seham M; Khalifa, Khaled A; El Ashmawy, Ramy A

2009-01-01

B cells from systemic lupus erythematosus (SLE) patients display signalling defects that may underlie disease pathogenesis activity.CD19 and CD22 play a major role as regulators of B-cell response. The aim of this study was to clarify the relationship between B cell surface markers namely CD19, CD20 and CD22 expression and clinical and laboratory indices of SLE activity. The study included 33 SLE patients and 20 healthy children and adolescents as controls. Flowcytometric assay of dual markers, CD19/CD20, and CD20/CD22 was done. SLE disease activity was assessed by SLEDAI score. CD22% was significantly higher while CD20% was significantly lower in the study compared to the control group. No significant difference was observed in both groups with respect to CD19% or CD19/CD22% ratio. The level of CD22 expression was significantly lower in high and very high active cases than in mild and moderate cases and negatively correlated with SLDEAI score and ESR. Results obtained showed that, B cell surface receptors CD20 and CD22 are significantly affected in patients with SLE, pointing to their possible involvement in the aetiopathogenesis of the disease and in the regulatory mechanisms in response to the immune disturbance.

Diversity of trends of viremia and T-cell markers in experimental acute feline immunodeficiency virus infection.

PubMed

Roche, Sylvain; El Garch, Hanane; Brunet, Sylvie; Poulet, Hervé; Iwaz, Jean; Ecochard, René; Vanhems, Philippe

2013-01-01

The early events of human immunodeficiency virus infection seem critical for progression toward disease and antiretroviral therapy initiation. We wanted to clarify some still unknown prognostic relationships between inoculum size and changes in various immunological and virological markers. Feline immunodeficiency virus infection could be a helpful model. Viremia and T-cell markers (number of CD4, CD8, CD8β(low)CD62L(neg) T-cells, CD4/CD8 ratio, and percentage of CD8β(low)CD62L(neg) cells among CD8 T-cells) were measured over 12 weeks in 102 cats infected with different feline immunodeficiency virus strains and doses. Viremia and T-cell markers trajectory groups were determined and the dose-response relationships between inoculum titres and trajectory groups investigated. Cats given the same inoculum showed different patterns of changes in viremia and T-cell markers. A statistically significant positive dose-response relationship was observed between inoculum titre and i) viremia trajectory-groups (r = 0.80, p<0.01), ii) CD8β(low)CD62L(neg) cell-fraction trajectory-groups (r = 0.56, p<0.01). Significant correlations were also found between viremia and the CD4/CD8 ratio and between seven out of ten T-cell markers. In cats, the infectious dose determines early kinetics of viremia and initial CD8+ T-cell activation. An expansion of the CD8β(low)CD62L(neg) T-cells might be an early predictor of progression toward disease. The same might be expected in humans but needs confirmation.

Age-associated changes in monocyte and innate immune activation markers occur more rapidly in HIV infected women.

PubMed

Martin, Genevieve E; Gouillou, Maelenn; Hearps, Anna C; Angelovich, Thomas A; Cheng, Allen C; Lynch, Fiona; Cheng, Wan-Jung; Paukovics, Geza; Palmer, Clovis S; Novak, Richard M; Jaworowski, Anthony; Landay, Alan L; Crowe, Suzanne M

2013-01-01

Aging is associated with immune dysfunction and the related development of conditions with an inflammatory pathogenesis. Some of these immune changes are also observed in HIV infection, but the interaction between immune changes with aging and HIV infection are unknown. Whilst sex differences in innate immunity are recognized, little research into innate immune aging has been performed on women. This cross-sectional study of HIV positive and negative women used whole blood flow cytometric analysis to characterize monocyte and CD8(+) T cell subsets. Plasma markers of innate immune activation were measured using standard ELISA-based assays. HIV positive women exhibited elevated plasma levels of the innate immune activation markers CXCL10 (p<0.001), soluble CD163 (sCD163, p = 0.001), sCD14 (p = 0.022), neopterin (p = 0.029) and an increased proportion of CD16(+) monocytes (p = 0.009) compared to uninfected controls. Levels of the innate immune aging biomarkers sCD163 and the proportion of CD16(+) monocytes were equivalent to those observed in HIV negative women aged 14.5 and 10.6 years older, respectively. CXCL10 increased with age at an accelerated rate in HIV positive women (p = 0.002) suggesting a synergistic effect between HIV and aging on innate immune activation. Multivariable modeling indicated that age-related increases in innate immune biomarkers CXCL10 and sCD163 are independent of senescent changes in CD8(+) T lymphocytes. Quantifying the impact of HIV on immune aging reveals that HIV infection in women confers the equivalent of a 10-14 year increase in the levels of innate immune aging markers. These changes may contribute to the increased risk of inflammatory age-related diseases in HIV positive women.

Unique ζ-chain motifs mediate a direct TCR-actin linkage critical for immunological synapse formation and T-cell activation.

PubMed

Klieger, Yair; Almogi-Hazan, Osnat; Ish-Shalom, Eliran; Pato, Aviad; Pauker, Maor H; Barda-Saad, Mira; Wang, Lynn; Baniyash, Michal

2014-01-01

TCR-mediated activation induces receptor microclusters that evolve to a defined immune synapse (IS). Many studies showed that actin polymerization and remodeling, which create a scaffold critical to IS formation and stabilization, are TCR mediated. However, the mechanisms controlling simultaneous TCR and actin dynamic rearrangement in the IS are yet not fully understood. Herein, we identify two novel TCR ζ-chain motifs, mediating the TCR's direct interaction with actin and inducing actin bundling. While T cells expressing the ζ-chain mutated in these motifs lack cytoskeleton (actin) associated (cska)-TCRs, they express normal levels of non-cska and surface TCRs as cells expressing wild-type ζ-chain. However, such mutant cells are unable to display activation-dependent TCR clustering, IS formation, expression of CD25/CD69 activation markers, or produce/secrete cytokine, effects also seen in the corresponding APCs. We are the first to show a direct TCR-actin linkage, providing the missing gap linking between TCR-mediated Ag recognition, specific cytoskeleton orientation toward the T-cell-APC interacting pole and long-lived IS maintenance. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Lactococcus lactis and Lactobacillus salivarius differently modulate early immunological response of Wistar rats co-administered with Listeria monocytogenes.

PubMed

Lukic, J; Jancic, I; Mirkovic, N; Bufan, B; Djokic, J; Milenkovic, M; Begovic, J; Strahinic, I; Lozo, J

2017-10-13

In the light of the increasing resistance of bacterial pathogens to antibiotics, one of the main global strategies in applied science is development of alternative treatments, which would be safe both for the host and from the environmental perspective. Accordingly, the aim of this study was to test whether two lactic acid bacteria (LAB) strains, Lactococcus lactis BGBU1-4 and Lactobacillus salivarius BGHO1, could be applied as safe supplements for Listeria infection. Two major research objectives were set: to compare the effects of BGBU1-4 and BGHO1 on early immune response in gut tissue of Wistar rats co-administered with Listeria monocytogenes ATCC19111 and next, to test how this applies to their usage as therapeutics in acute ATCC19111 infection. Intestinal villi (IV), Peyer's patches (PP) and mesenteric lymph nodes (MLN) were used for the analysis. The results showed that BGHO1 increased the mRNA expression of innate immune markers CD14, interleukin (IL)-1β and tumour necrosis factor (TNF)-α in PP and IV, and, in parallel, caused a decrease of listeriolysin O (LLO) mRNA expression in same tissues. In MLN of BGHO1 treated rats, LLO expression was increased, along with an increase of the expression of OX-62 mRNA and CD69, pointing to the activation of adaptive immunity. On the other hand, in BGBU1-4 treated rats, there was no reduction of LLO mRNA expression and no induction of innate immunity markers in intestinal tissue. Additionally, CD14 and IL-1β, as well as LLO, but not OX-62 mRNA and CD69 expression, were elevated in MLN of BGBU1-4 treated rats. However, when applied therapeutically, both, BGBU1-4 and BGHO1, lowered Listeria count in spleens of infected rats. Our results not only reveal the potential of LAB to ameliorate Listeria infections, but suggest different immunological effects of two different LAB strains, both of which could be effective in Listeria elimination.

Honokiol Increases CD4+ T Cell Activation and Decreases TNF but Fails to Improve Survival Following Sepsis.

PubMed

Klingensmith, Nathan J; Chen, Ching-Wen; Liang, Zhe; Burd, Eileen M; Farris, Alton B; Arbiser, Jack L; Ford, Mandy L; Coopersmith, Craig M

2017-10-11

Honokiol is a biphenolic isolate extracted from the bark of the magnolia tree that has been used in traditional Chinese and Japanese medicine, and has more recently been investigated for its anti-inflammatory and anti-bacterial properties. Honokiol has previously been demonstrated to improve survival in sepsis models that have rapid 100% lethality. The purpose of this study was to determine the impact of Honokiol on the host response in a model of sepsis that more closely approximates human disease. Male and female C57BL/6 mice underwent cecal ligation and puncture (CLP) to induce polymicrobial intraabdominal sepsis. Mice were then randomized to receive an injection of either Honokiol (120 mg/kg/day) or vehicle and were sacrificed after 24 hours for functional studies or followed 7 days for survival. Honokiol treatment after sepsis increased the frequency of CD4 T cells and increased activation of CD4 T cells as measured by the activation marker CD69. Honokiol also increased splenic dendritic cells. Honokiol simultaneously decreased frequency and number of CD8 T cells. Honokiol decreased systemic TNF without impacting other systemic cytokines. Honokiol did not have a detectable effect on kidney function, lung physiology, liver function or intestinal integrity. In contrast to prior studies of Honokiol in a lethal model of sepsis, Honokiol did not alter survival at seven days (70% mortality for Honokiol vs. 60% mortality for vehicle). Honokiol is thus effective in modulating the host immune response and inflammation following a clinically relevant model of sepsis but is not sufficient to alter survival.

Photon manipulation in silicon nanophotonic circuits

NASA Astrophysics Data System (ADS)

Elshaari, Ali Wanis

2011-12-01

CD8+ T cells are the branch of the adaptive immune system responsible for recognizing and killing tumor cells or cells infected with intracellular pathogens, such as Listeria monocytogenes (LM). However, when CD8+ T cells target our own tissues, they can cause autoimmune diseases, such as type I diabetes, rheumatoid arthritis. For CD8+ T cells to fulfill these functions, the T cell receptors (TCRs) on CD8+ T cells must recognize pathogens or antigens presented on the surface of target cells. TCR ligation triggers multiple signaling pathways that lead to the activation and proliferation of CD8+ T cells. The goal of our research is to define the TCR-proximal signaling events that regulate CD8+ T cell-mediated immunity. To accomplish this goal, we are focusing on an adaptor protein Gads, which is critical for optimal TCR-mediated calcium mobilization. We reported the first analysis of the function of Gads in peripheral naive CD8+ T cells. To examine the function of Gads in CD8+ T cell mediated immune responses, we crossed Gads-/- mice with mice expressing an MHC class I-restricted transgenic TCR recognizing ovalbumin (OVA). The transgenic mice are called ovalbumin-specific T cell receptor-major histocompatibility complex class I restricted (OT-I) mice. We investigated the effect of Gads on the proliferation of CD8+ T cells following stimulation with peptide antigen in vivo and in vitro. We stimulated splenocytes from Gads+/+ OT-I and Gads -/- OT-I mice with the peptide agonist. The experiments revealed that Gads is required for optimal proliferation of CD8+ T cells. The regulation of Gads is most evident at the early time points of proliferation. Then we demonstrated that Gads-/- CD8+ T cells have impaired TCR-mediated exit from G0 phase of the cell cycle. In addition, Gads-/- CD8+ T cells have delayed expression of c-myc and the activation markers CD69 and CD25, upon stimulation with peptide antigen. Next, we investigated how Gads affects CD8+ T cell-mediated immunity in the context of infection with LM. We adoptively transferred naive CD8+ T cells from Gads+/+ OT-I mice and/or Gads -/- OT-I mice into congenic wild-type hosts. Then the recipient mice were infected with recombinant LM expressing ovalbumin (rLM-OVA). The CD8 + T cells from OT-I mice recognize and respond to the ovalbumin provided by this strain of LM. By using this system, we investigated how Gads regulates the activation of antigen-specific CD8+ T cells as well as the expansion and memory phases of CD8+ T cell-mediated immune responses following infection with rLM-OVA. We also examined the recall response of CD8+ T cells after the secondary encounter with the same pathogen. Our data demonstrated that Gads regulates the expression of activation markers CD69 and CD25 of antigen-specific CD8+ T cells but Gads is not required for the onset of accumulation of antigen-specific CD8 + T cells following infection. However, Gads is critical to sustain the expansion of CD8+ T cell-mediated immune response following infection. Although the differentiation of naive CD8+ T cells into memory cells is independent of Gads, Gads is required for an optimal recall response. Our data indicating that Gads regulates the initiation of proliferation of CD8+ T cells upon TCR ligation by peptide antigen seemed to contradict with our in vivo infection data showing that Gads is not required for the initiation of expansion of CD8+ T cell population. In order to explain the "discrepancy", we hypothesized that the homotypic interactions among CD8+ T cells compensate for Gads deficiency at the initial stage of accumulation of antigen-specific CD8+ T cells upon infection. Our data indicated that the need for Gads in cell cycle progression of CD8+ T cells when total splenocytes were stimulated could be overcome by stimulating purified CD8 + T cells. These data suggested that the homotypic interactions among CD8+ T cells facilitate the TCR signaling so as to compensate for Gads deficiency in promoting cell cycle entry and proliferation. To conclude, the role of Gads in TCR-mediated activation and proliferation of CD8+ T cells is dependent on the interactions of CD8 + T cells and their partners. Interestingly, if CD8+ T cells interact with non-CD8+ T cells, Gads regulates the kinetics of cell cycle entry; however, if CD8+ T cells interact with other CD8+ T cells, Gads is dispensable for cell cycle entry of CD8+ T cells. Overall, these studies will help us better understand how TCR-proximal signaling regulates the activation of CD8 + T cells.

The XC chemokine receptor 1 is a conserved selective marker of mammalian cells homologous to mouse CD8α+ dendritic cells

PubMed Central

Crozat, Karine; Guiton, Rachel; Contreras, Vanessa; Feuillet, Vincent; Dutertre, Charles-Antoine; Ventre, Erwan; Vu Manh, Thien-Phong; Baranek, Thomas; Storset, Anne K.; Marvel, Jacqueline; Boudinot, Pierre; Hosmalin, Anne; Schwartz-Cornil, Isabelle

2010-01-01

Human BDCA3+ dendritic cells (DCs) were suggested to be homologous to mouse CD8α+ DCs. We demonstrate that human BDCA3+ DCs are more efficient than their BDCA1+ counterparts or plasmacytoid DCs (pDCs) in cross-presenting antigen and activating CD8+ T cells, which is similar to mouse CD8α+ DCs as compared with CD11b+ DCs or pDCs, although with more moderate differences between human DC subsets. Yet, no specific marker was known to be shared between homologous DC subsets across species. We found that XC chemokine receptor 1 (XCR1) is specifically expressed and active in mouse CD8α+, human BDCA3+, and sheep CD26+ DCs and is conserved across species. The mRNA encoding the XCR1 ligand chemokine (C motif) ligand 1 (XCL1) is selectively expressed in natural killer (NK) and CD8+ T lymphocytes at steady-state and is enhanced upon activation. Moreover, the Xcl1 mRNA is selectively expressed at high levels in central memory compared with naive CD8+ T lymphocytes. Finally, XCR1−/− mice have decreased early CD8+ T cell responses to Listeria monocytogenes infection, which is associated with higher bacterial loads early in infection. Therefore, XCR1 constitutes the first conserved specific marker for cell subsets homologous to mouse CD8α+ DCs in higher vertebrates and promotes their ability to activate early CD8+ T cell defenses against an intracellular pathogenic bacteria. PMID:20479118

Vaginal Immunization to Elicit Primary T-Cell Activation and Dissemination

PubMed Central

Pettini, Elena; Prota, Gennaro; Ciabattini, Annalisa; Boianelli, Alessandro; Fiorino, Fabio; Pozzi, Gianni; Vicino, Antonio; Medaglini, Donata

2013-01-01

Primary T-cell activation at mucosal sites is of utmost importance for the development of vaccination strategies. T-cell priming after vaginal immunization, with ovalbumin and CpG oligodeoxynucleotide adjuvant as model vaccine formulation, was studied in vivo in hormone-synchronized mice and compared to the one induced by the nasal route. Twenty-four hours after both vaginal or nasal immunization, antigen-loaded dendritic cells were detected within the respective draining lymph nodes. Vaginal immunization elicited a strong recruitment of antigen-specific CD4+ T cells into draining lymph nodes that was more rapid than the one observed following nasal immunization. T-cell clonal expansion was first detected in iliac lymph nodes, draining the genital tract, and proliferated T cells disseminated towards distal lymph nodes and spleen similarly to what observed following nasal immunization. T cells were indeed activated by the antigen encounter and acquired homing molecules essential to disseminate towards distal lymphoid organs as confirmed by the modulation of CD45RB, CD69, CD44 and CD62L marker expression. A multi-type Galton Watson branching process, previously used for in vitro analysis of T-cell proliferation, was applied to model in vivo CFSE proliferation data in draining lymph nodes 57 hours following immunization, in order to calculate the probabilistic decision of a cell to enter in division, rest in quiescence or migrate/die. The modelling analysis indicated that the probability of a cell to proliferate was higher following vaginal than nasal immunization. All together these data show that vaginal immunization, despite the absence of an organized mucosal associated inductive site in the genital tract, is very efficient in priming antigen-specific CD4+ T cells and inducing their dissemination from draining lymph nodes towards distal lymphoid organs. PMID:24349003

The Polyunsaturated Fatty Acids Arachidonic Acid and Docosahexaenoic Acid Induce Mouse Dendritic Cells Maturation but Reduce T-Cell Responses In Vitro

PubMed Central

Carlsson, Johan A.; Wold, Agnes E.; Sandberg, Ann-Sofie; Östman, Sofia M.

2015-01-01

Long-chain polyunsaturated fatty acids (PUFAs) might regulate T-cell activation and lineage commitment. Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells. Spleen DCs from BALB/c mice were cultured in vitro with ovalbumin (OVA) with 50 μM fatty acids; α-linolenic acid, arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid or oleic acid and thereafter OVA-specific DO11.10 T cells were added to the cultures. Fatty acids were taken up by the DCs, as shown by gas chromatography analysis. After culture with arachidonic acid or DHA CD11c+ CD11b+ and CD11c+ CD11bneg DCs expressed more CD40, CD80, CD83, CD86 and PDL-1, while IAd remained unchanged. However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violetlow) and expressed CD69 or CD25, while more were necrotic (7AAD+). We noted an increased proportion of T cells with a regulatory T cell (Treg) phenotype, i.e., when gating on CD4+ FoxP3+ CTLA-4+, CD4+ FoxP3+ Helios+ or CD4+ FoxP3+ PD-1+, in co-cultures with arachidonic acid- or DHA-primed DCs relative to control cultures. The proportion of putative Tregs was inversely correlated to T-cell proliferation, indicating a suppressive function of these cells. With arachidonic acid DCs produced higher levels of prostaglandin E2 while T cells produced lower amounts of IL-10 and IFNγ. In conclusion arachidonic acid and DHA induced up-regulation of activation markers on DCs. However arachidonic acid- and DHA-primed DCs reduced T-cell proliferation and increased the proportion of T cells expressing FoxP3, indicating that these fatty acids can promote induction of regulatory T cells. PMID:26619195

Arthroscopic Harvest of Adipose-Derived Mesenchymal Stem Cells From the Infrapatellar Fat Pad.

PubMed

Dragoo, Jason L; Chang, Wenteh

2017-11-01

The successful isolation of adipose-derived mesenchymal stem cells (ADSCs) from the arthroscopically harvested infrapatellar fat pad (IFP) would provide orthopaedic surgeons with an autologous solution for regenerative procedures. To demonstrate the quantity and viability of the mesenchymal stem cell population arthroscopically harvested from the IFP as well as the surrounding synovium. Descriptive laboratory study. The posterior border of the IFP, including the surrounding synovial tissue, was harvested arthroscopically from patients undergoing anterior cruciate ligament reconstruction. Tissue was then collected in an AquaVage adipose canister, followed by fat fractionization using syringe emulsification and concentration with an AdiPrep device. In the laboratory, the layers of tissue were separated and then digested with 0.3% type I collagenase. The pelleted stromal vascular fraction (SVF) cells were then immediately analyzed for viability, mesenchymal cell surface markers by fluorescence-activated cell sorting, and clonogenic capacity. After culture expansion, the metabolic activity of the ADSCs was assessed by an AlamarBlue assay, and the multilineage differentiation capability was tested. The transition of surface antigens from the SVF toward expanded ADSCs at passage 2 was further evaluated. SVF cells were successfully harvested with a mean yield of 4.86 ± 2.64 × 10 5 cells/g of tissue and a mean viability of 69.03% ± 10.75%, with ages ranging from 17 to 52 years (mean, 35.14 ± 13.70 years; n = 7). The cultured ADSCs composed a mean 5.85% ± 5.89% of SVF cells with a mean yield of 0.33 ± 0.42 × 10 5 cells/g of tissue. The nonhematopoietic cells (CD45 - ) displayed the following surface antigens as a percentage of the viable population: CD44 + (52.21% ± 4.50%), CD73 + CD90 + CD105 + (19.20% ± 17.04%), and CD44 + CD73 + CD90 + CD105 + (15.32% ± 15.23%). There was also a significant increase in the expression of ADSC markers CD73 (96.97% ± 1.72%; P < .01), CD10 (84.47% ± 15.46%; P < .05), and CD166 (11.63% ± 7.84%; P < .005) starting at passage 2 compared with freshly harvested SVF cells. The clonogenic efficiency of SVF cells was determined at a mean 3.21% ± 1.52% for layer 1 and 1.51% ± 0.55% for layer 2. Differentiation into cartilage, fat, and bone tissue was demonstrated by tissue-specific staining and quantitative polymerase chain reaction. SVF cells from the IFP and adjacent synovial tissue were successfully harvested using an arthroscopic technique and produced ADSCs with surface markers that meet criteria for defined mesenchymal stem cells. An autologous source of stem cells can now be harvested using a simple arthroscopic technique that will allow orthopaedic surgeons easier access to progenitor cells for regenerative procedures.

Soluble CD30 serum level in HCV-positive chronic active hepatitis: A surrogate marker of disease activity?

PubMed

Foschi, F G; Gramenzi, A; Castelli, E; Cursaro, C; Pagani, S; Margotti, M; D'Errico, A; Andreone, P; Stefanini, G F; Bernardi, M

2000-06-01

In the present study, high levels of CD30s, a glycoprotein preferentially expressed and released by T lymphocytes producing Th(2)-type cytokines, were seen in the sera of patients with chronic hepatitis C, and a correlation with histological activity of the disease was found. CD30s levels were assayed in the sera of 29 HCV RNA-positive patients with histologically proven chronic active hepatitis and in 30 healthy blood donors. Thirteen of 29 (45%) HCV patients had CD30s serum levels above the normal range (>20 U/ml). Mean CD30s serum levels were significantly higher in HCV patients than in controls (P<0.0005). A positive correlation was found between serum CD30s levels and both the histological activity index (r=0.59, P=0.001) and ALT serum levels (r=0.5; P=0.006). The raised CD30s level found in more severe HCV liver disease indirectly suggests activation and expansion of Th(2)cells. CD30s levels could represent a useful surrogate marker of activity in chronic HCV infections. Copyright 2000 Academic Press.

Capturing the systemic immune signature of a norovirus infection: an n-of-1 case study within a clinical trial

PubMed Central

Cutler, Antony J.; Oliveira, Joao; Ferreira, Ricardo C.; Challis, Ben; Walker, Neil M.; Caddy, Sarah; Lu, Jia; Stevens, Helen E.; Smyth, Deborah J.; Pekalski, Marcin L.; Kennet, Jane; Hunter, Kara M.D.; Goodfellow, Ian; Wicker, Linda S.; Todd, John A.; Waldron-Lynch, Frank

2017-01-01

Background: The infection of a participant with norovirus during the adaptive study of interleukin-2 dose on regulatory T cells in type 1 diabetes (DILT1D) allowed a detailed insight into the cellular and cytokine immune responses to this prevalent gastrointestinal pathogen. Methods:  Serial blood, serum and peripheral blood mononuclear cell (PBMC) samples were collected pre-, and post-development of the infection. To differentiate between the immune response to norovirus and to control for the administration of a single dose of aldesleukin (recombinant interleukin-2, rIL-2) alone, samples from five non-infected participants administered similar doses were analysed in parallel. Results: Norovirus infection was self-limited and resolved within 24 hours, with the subsequent development of anti-norovirus antibodies. Serum pro- and anti-inflammatory cytokine levels, including IL-10, peaked during the symptomatic period of infection, coincident with increased frequencies of monocytes and neutrophils. At the same time, the frequency of regulatory CD4 + T cell (Treg), effector T cell (Teff) CD4 + and CD8 + subsets were dynamically reduced, rebounding to baseline levels or above at the next sampling point 24 hours later.  NK cells and NKT cells transiently increased CD69 expression and classical monocytes expressed increased levels of CD40, HLA-DR and SIGLEC-1, biomarkers of an interferon response. We also observed activation and mobilisation of Teffs, where increased frequencies of CD69 + and Ki-67 + effector memory Teffs were followed by the emergence of memory CD8 + Teff expressing the mucosal tissue homing markers CD103 and β7 integrin. Treg responses were coincident with the innate cell, Teff and cytokine response. Key Treg molecules FOXP3, CTLA-4, and CD25 were upregulated following infection, alongside an increase in frequency of Tregs with the capacity to home to tissues. Conclusions:  The results illustrate the innate, adaptive and counter-regulatory immune responses to norovirus infection. Low-dose IL-2 administration induces many of the Treg responses observed during infection. PMID:28815218

Suppressed PHA Activation of T Lymphocytes in Simulated Microgravity Is Restored by Direct Activation of Protein Kinase C with Phorbol Ester

NASA Technical Reports Server (NTRS)

Cooper, David; Pellis, Neal R.

1997-01-01

Various aspects of spaceflight, including microgravity, cosmic radiation, and physiological stress, may perturb immune function. We sought to understand the impact of microgravity alone on the cellular mechanisms critical to immunity. We utilized clinostatic RWV bioreactors that simulate aspects of microgravity to analyze the response of human PBMC to polyclonal activation. PHA responsiveness in the RWV was almost completely diminished. IL-2 and IFN-gamma secretion was reduced whereas IL- 1 beta and IL-6 secretion was increased, suggesting that monocytes may not be as adversely affected by simulated microgravity as T cells. Activation marker expression (CD25, CD69, CD71) was significantly reduced in RWV cultures. Furthermore, addition of exogenous IL-2 to these cultures did not restore proliferation. Reduced cell-cell and cell-substratum interactions may play a role in the loss of PHA responsiveness. However, PHA activation in Teflon culture bags that limit cell-substratum interactions did not suppress PHA activation. Furthermore, increasing cell density and, therefore, cell-cell interactions in the RWV cultures did not help restore PHA activation. However, placing PBMC within small collagen beads did partially restore PHA responsiveness. Activation of both PBMC and purified T cells with PMA and ionomycin was unaffected by RWV culture, indicating that signaling mechanisms downstream of PKC activation and calcium flux are not sensitive to simulated microgravity. Furthermore, submitogenic doses of PMA alone but not ionomycin alone restored PHA responsiveness of PBMC in RWV culture. Thus, our data indicate that during polyclonal activation the signaling pathways upstream of PKC activation are sensitive to simulated microgravity.

Dendritic cell co-stimulatory and co-inhibitory markers in chronic HCV: An Egyptian study

PubMed Central

Fouad, Hanan; Raziky, Maissa Saeed El; Aziz, Rasha Ahmed Abdel; Sabry, Dina; Aziz, Ghada Mahmoud Abdel; Ewais, Manal; Sayed, Ahmed Reda

2013-01-01

AIM: To assess co-stimulatory and co-inhibitory markers of dendritic cells (DCs) in hepatitis C virus (HCV) infected subjects with and without uremia. METHODS: Three subject groups were included in the study: group 1 involved 50 control subjects, group 2 involved 50 patients with chronic HCV infection and group 3 involved 50 HCV uremic subjects undergoing hemodialysis. CD83, CD86 and CD40 as co-stimulatory markers and PD-L1 as a co-inhibitory marker were assessed in peripheral blood mononuclear cells by real-time polymerase chain reaction. Interleukin-10 (IL-10) and hyaluronic acid (HA) levels were also assessed. All findings were correlated with disease activity, viral load and fibrogenesis. RESULTS: There was a significant decrease in co-stimulatory markers; CD83, CD86 and CD40 in groups 2 and 3 vs the control group. Co-stimulatory markers were significantly higher in group 3 vs group 2. There was a significant elevation in PD-L1 in both HCV groups vs the control group. PD-L1 was significantly lower in group 3 vs group 2. There was a significant elevation in IL-10 and HA levels in groups 2 and 3, where IL-10 was higher in group 3 and HA was lower in group 3 vs group 2. HA level was significantly correlated with disease activity and fibrosis grade in group 2. IL-10 was significantly correlated with fibrosis grade in group 2. There were significant negative correlations between co-stimulatory markers and viral load in groups 2 and 3, except CD83 in dialysis patients. There was a significant positive correlation between PD-L1 and viral load in both HCV groups. CONCLUSION: A significant decrease in DC co-stimulatory markers and a significant increase in a DC co-inhibitory marker were observed in HCV subjects and to a lesser extent in dialysis patients. PMID:24282359

Monocyte Activation Is Associated With Worse Cognitive Performance in HIV-Infected Women With Virologic Suppression

PubMed Central

Imp, Brandon M.; Rubin, Leah H.; Tien, Phyllis C.; Plankey, Michael W.; Golub, Elizabeth T.; French, Audrey L.; Valcour, Victor G.

2017-01-01

Background. Cognitive impairment persists despite suppression of plasma human immunodeficiency virus (HIV) RNA. Monocyte-related immune activation is a likely mechanism. We examined immune activation and cognition in a cohort of HIV-infected and uninfected women from the Women's Interagency HIV Study (WIHS). Methods. Blood levels of activation markers, soluble CD163 (sCD163), soluble CD14 (sCD14), CRP, IL-6, and a gut microbial translocation marker (intestinal fatty acid binding protein (I-FABP)) were measured in 253 women (73% HIV-infected). Markers were compared to concurrent (within ± one semiannual visit) neuropsychological testing performance. Results. Higher sCD163 levels were associated with worse overall performance and worse verbal learning, verbal memory, executive function, psychomotor speed, and fine motor skills (P < .05 for all comparisons). Higher sCD14 levels were associated with worse verbal learning, verbal memory, executive function, and psychomotor speed (P < .05 for all comparisons). Among women with virological suppression, sCD163 remained associated with overall performance, verbal memory, psychomotor speed, and fine motor skills, and sCD164 remained associated with executive function (P < .05 for all comparisons). CRP, IL-6, and I-FABP were not associated with worse cognitive performance. Conclusions. Monocyte activation was associated with worse cognitive performance, and associations persisted despite viral suppression. Persistent inflammatory mechanisms related to monocytes correlate to clinically pertinent brain outcomes. PMID:27789726

Pre-immunotherapy serum CA27.29 (MUC-1) mucin level and CD69+ lymphocytes correlate with effects of Theratope sialyl-Tn-KLH cancer vaccine in active specific immunotherapy.

PubMed

Reddish, M A; MacLean, G D; Poppema, S; Berg, A; Longenecker, B M

1996-06-01

Patients with metastatic breast, colorectal or ovarian cancers received active specific immunotherapy (ASI) with Theratope sialyl-Tn-KLH (keyhole limpet hemocyanin) cancer vaccine emulsified in Detox adjuvant. The median log2 anti-STn IgG titer generated by ASI, estimated by enzyme-linked immunosorbent assay with solid-phase ovine submaxillary mucin, was 5.322 (range = 0 - 9.322). Following ASI, 51 patients who generated titers higher than the median value for anti-STn+ mucin IgG survived longer than 46 patients who generated lower titers below the median. 38 of the patients were phenotyped for CD69 prior to ASI. The patients with lower numbers of CD69+ peripheral blood lymphocytes prior to immunotherapy (pre-ASI) also had low serum CA27.29 cancer antigen (MUC-1) levels, and had longer times to disease progression and improved survival following ASI. Elevated pre-ASI serum CA27.29 tumor antigen levels were associated with higher numbers of CD69+ PBL, with decreased anti-STn antibody production and decreased survival following ASI. The data are compatible with the hypothesis that elevated serum MUC-1 mucin is specifically immunosuppressive.

Circulating activated innate lymphoid cells and mucosal-associated invariant T cells are associated with airflow limitation in patients with asthma.

PubMed

Ishimori, Ayako; Harada, Norihiro; Chiba, Asako; Harada, Sonoko; Matsuno, Kei; Makino, Fumihiko; Ito, Jun; Ohta, Shoichiro; Ono, Junya; Atsuta, Ryo; Izuhara, Kenji; Takahashi, Kazuhisa; Miyake, Sachiko

2017-04-01

A variety of innate subsets of lymphoid cells such as natural killer (NK) cells, several populations of innate lymphoid cells (ILCs), and mucosal-associated invariant T (MAIT) cells as innate-like T lymphocytes are involved in asthma and may have important effector functions in asthmatic immune responses. In the present study, we investigated whether NK cells, ILCs, and MAIT cells in the peripheral blood of patients with asthma would be associated with clinical asthma parameters. We recruited 75 adult patients with mild to severe asthma. The peripheral blood mononuclear cells in peripheral venous blood samples from the patients were purified and stained with different combinations of appropriate antibodies. The cells were analyzed by flow cytometry. The percentage of activated (i.e., CD69 + ) NK cells in the total NK cell population was negatively correlated with FEV 1 % which is calculated by the forced expiratory volume in 1 s (FEV 1 )/the forced vital capacity (FVC). The percentages of CD69 + ILC1s and ILC2s were negatively correlated with FEV 1 % and %FEV 1 . The percentage of CD69 + ILC3s was positively correlated with BMI, and the percentage of CD69 + MAIT cells was negatively correlated with FEV 1 %. Moreover, the percentage of CD69 + NK cells, ILC1s, ILC2s, ILC3s, and MAIT cells were positively correlated with each other. For the first time, our data showed that activated NK cells, ILC1s, ILC2s, ILC3s, and MAIT cells were positively correlated with each other and may be associated with airflow limitation in patients with asthma. Copyright © 2016 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.

Clinical relevance of miR-mediated HLA-G regulation and the associated immune cell infiltration in renal cell carcinoma

PubMed Central

Jasinski-Bergner, Simon; Stoehr, Christine; Bukur, Juergen; Massa, Chiara; Braun, Juliane; Hüttelmaier, Stefan; Spath, Verena; Wartenberg, Roland; Legal, Wolfgang; Taubert, Helge; Wach, Sven; Wullich, Bernd; Hartmann, Arndt; Seliger, Barbara

2015-01-01

In human tumors of distinct origin including renal cell carcinoma (RCC), the non-classical human leukocyte antigen G (HLA-G) is frequently expressed, thereby inhibiting the cytotoxic activity of T and natural killer (NK) cells. Recent studies demonstrated a strong post-transcriptional gene regulation of the HLA-G by miR-152, −148A, −148B and −133A. Standard methods were applied to characterize the expression and function of HLA-G, HLA-G-regulatory microRNAs (miRs) and the immune cell infiltration in 453 RCC lesions using a tissue microarray and five RCC cell lines linking these results to clinical parameters. Direct interactions with HLA-G regulatory miRs and the HLA-G 3′ untranslated region (UTR) were detected and the affinities of these different miRs to the HLA-G 3′-UTR compared. qPCR analyses and immunohistochemical staining revealed an inverse expression of miR-148A and −133A with the HLA-G protein in situ and in vitro. Stable miR overexpression caused a downregulation of HLA-G protein enhancing the NK and LAK cell-mediated cytotoxicity in in vitro CD107a activation assays revealing a HLA-G-dependent cytotoxic activity of immune effector cells. A significant higher frequency of CD3+/CD8+ T cell lymphocytes, but no differences in the activation markers CD69, CD25 or in the presence of CD56+, FoxP3+ and CD4+ immune cells were detected in HLA-G+ compared to HLA-G− RCC lesions. This could be associated with higher WHO grade, but not with a disease-specific survival. These data suggest a miR-mediated control of HLA-G expression in RCC, which is associated with a distinct pattern of immune cell infiltration. PMID:26155421

B cell subset distribution is altered in patients with severe periodontitis.

PubMed

Demoersman, Julien; Pochard, Pierre; Framery, Camille; Simon, Quentin; Boisramé, Sylvie; Soueidan, Assem; Pers, Jacques-Olivier

2018-01-01

Several studies have recently highlighted the implication of B cells in physiopathogenesis of periodontal disease by showing that a B cell deficiency leads to improved periodontal parameters. However, the detailed profiles of circulating B cell subsets have not yet been investigated in patients with severe periodontitis (SP). We hypothesised that an abnormal distribution of B cell subsets could be detected in the blood of patients with severe periodontal lesions, as already reported for patients with chronic inflammatory diseases as systemic autoimmune diseases. Fifteen subjects with SP and 13 subjects without periodontitis, according to the definition proposed by the CDC periodontal disease surveillance work group, were enrolled in this pilot observational study. Two flow cytometry panels were designed to analyse the circulating B and B1 cell subset distribution in association with the RANKL expression. A significantly higher percentage of CD27+ memory B cells was observed in patients with SP. Among these CD27+ B cells, the proportion of the switched memory subset was significantly higher. At the same time, human B1 cells, which were previously associated with a regulatory function (CD20+CD69-CD43+CD27+CD11b+), decreased in SP patients. The RANKL expression increased in every B cell subset from the SP patients and was significantly greater in activated B cells than in the subjects without periodontitis. These preliminary results demonstrate the altered distribution of B cells in the context of severe periodontitis. Further investigations with a larger cohort of patients can elucidate if the analysis of the B cell compartment distribution can reflect the periodontal disease activity and be a reliable marker for its prognosis (clinical trial registration number: NCT02833285, B cell functions in periodontitis).

B cell subset distribution is altered in patients with severe periodontitis

PubMed Central

Demoersman, Julien; Pochard, Pierre; Framery, Camille; Simon, Quentin; Boisramé, Sylvie; Soueidan, Assem

2018-01-01

Several studies have recently highlighted the implication of B cells in physiopathogenesis of periodontal disease by showing that a B cell deficiency leads to improved periodontal parameters. However, the detailed profiles of circulating B cell subsets have not yet been investigated in patients with severe periodontitis (SP). We hypothesised that an abnormal distribution of B cell subsets could be detected in the blood of patients with severe periodontal lesions, as already reported for patients with chronic inflammatory diseases as systemic autoimmune diseases. Fifteen subjects with SP and 13 subjects without periodontitis, according to the definition proposed by the CDC periodontal disease surveillance work group, were enrolled in this pilot observational study. Two flow cytometry panels were designed to analyse the circulating B and B1 cell subset distribution in association with the RANKL expression. A significantly higher percentage of CD27+ memory B cells was observed in patients with SP. Among these CD27+ B cells, the proportion of the switched memory subset was significantly higher. At the same time, human B1 cells, which were previously associated with a regulatory function (CD20+CD69-CD43+CD27+CD11b+), decreased in SP patients. The RANKL expression increased in every B cell subset from the SP patients and was significantly greater in activated B cells than in the subjects without periodontitis. These preliminary results demonstrate the altered distribution of B cells in the context of severe periodontitis. Further investigations with a larger cohort of patients can elucidate if the analysis of the B cell compartment distribution can reflect the periodontal disease activity and be a reliable marker for its prognosis (clinical trial registration number: NCT02833285, B cell functions in periodontitis). PMID:29447240

Surveillance on the status of immune cells after Echinnococcus granulosus protoscoleces infection in Balb/c mice.

PubMed

Pan, Wei; Zhou, He-Jun; Shen, Yu-Juan; Wang, Ying; Xu, Yu-Xin; Hu, Yuan; Jiang, Yan-Yan; Yuan, Zhong-Ying; Ugwu, Chidiebere E; Cao, Jian-Ping

2013-01-01

Cystic echinococcosis is a global parasitic disease caused by infection with Echinococcus granulosus larvae with potentially life-threatening complications in humans. To date, the status of the immune cells believed to be associated with the pathogenicity of E. granulosus infection has not been demonstrated clearly. In this study, we developed a multiplex flow cytometry assay to investigate the systemic immune status of innate and adaptive immunity at 30, 180, 360 days post-infection (dpi) in mice infected with E. granulousus. At 30 dpi, an increase in the number of CD11b(+) and CD11c(+) antigen-presenting cells (APCs) was observed. This was accompanied by the slight down-regulated expression of the co-stimulatory molecule MHC-II, indicating the impairment of APCs in early infection through the release of secretory-excretory products. In all infected groups, we observed a significant increase in innate immune cells, including APCs and GR-1(+) cells, and a dramatic increase in the myeloid-derived suppressor cells (MDSC) expressing CD11b(+)/GR-1(+). Moreover, the upregulation of the activated markers CD69, CD44, CD40L, and the downregulation of CD62L were observed in the CD4(+) and CD8(+) T cells following infection. Regulatory T cells expressing CD4(+)/CD25(+)/FoxP3 (+) increased significantly over the course of infection. Our findings demonstrate that the microenvironment in the peripheral immune system after E. granulosus infection changes in subtle but detectably ways, especially during the persistent period of infection. We found that T cells were activated following infection, but observed that the significant increase of immunosuppressive cells such as MDSC and Treg cells could inhibit T cell response to E. granulosus antigens. We suggest these cells may play a neglected but key role in the downregulation of the immune response in long-term parasitic infection. Understanding the basic functions and temporal interactions of these immunosuppressive cells will pave the way for new strategies of parasite vaccine design.

Comprehensive Mass Cytometry Analysis of Cell Cycle, Activation, and Coinhibitory Receptors Expression in CD4 T Cells from Healthy and HIV-Infected Individuals.

PubMed

Corneau, Aurélien; Cosma, Antonio; Even, Sophie; Katlama, Christine; Le Grand, Roger; Frachet, Véronique; Blanc, Catherine; Autran, Brigitte

2017-01-01

Mass cytometry allows large multiplex analysis of cell cycle stages together with differentiation, activation, and exhaustion markers, allowing further assessment of the quiescence status of resting CD4 T cells. Peripheral blood CD4 T lymphocytes from 8 individuals, 4 healthy donors, and 4 HIV-infected on antiretroviral treatment (T) were stained with the same 26 monoclonal antibodies and dyes targeting surface and intracellular markers of differentiation, activation, exhaustion, and cell cycle stages. Samples were run on a CYTOF-2. Patterns of naïve [TN] CD4 T cells strongly differed from all other memory subsets central-memory (CM), transitional-memory (TM), effector-memory (EM), and terminally differentiated RA-expressing (TEMRA) subsets, while stem-cell memory (SCM) and T follicular-helper cells (TfH) were close to CM and TM cells with the highest percentages in cell cycle. EM and TEMRA were the most altered by HIV infection, with an increased frequency of activated and cycling cells. Activation markers and coinhibitory receptor expression differed among cell cycle stages, with HLA-DR fitting better than CD25 or CD38 with cycle, and opposite PD-1 gradients along differentiation and cell cycle. "Resting" DR-CD25- CD4+ T cells contained similar amounts of cells in G1 than the activated DR ± CD25± ones but three fold lower cells in S-G2-M. This broad multiplex mass cytometry analysis demonstrates some subsets of the so-called "resting" CD25-DR- CD4+ T cells contain noticeable amounts of cells into cycle or expressing coinhibitory receptors, opening new avenues for a redefinition of resting peripheral blood CD4 T cells harboring the HIV reservoirs. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

Immune-Regulatory Molecule CD69 Controls Peritoneal Fibrosis

PubMed Central

Liappas, Georgios; González-Mateo, Guadalupe Tirma; Sánchez-Díaz, Raquel; Lazcano, Juan José; Lasarte, Sandra; Matesanz-Marín, Adela; Zur, Rafal; Ferrantelli, Evelina; Ramírez, Laura García; Aguilera, Abelardo; Fernández-Ruiz, Elena; Beelen, Robert H.J.; Selgas, Rafael; Sánchez-Madrid, Francisco

2016-01-01

Patients with ESRD undergoing peritoneal dialysis develop progressive peritoneal fibrosis, which may lead to technique failure. Recent data point to Th17-mediated inflammation as a key contributor in peritoneal damage. The leukocyte antigen CD69 modulates the setting and progression of autoimmune and inflammatory diseases by controlling the balance between Th17 and regulatory T cells (Tregs). However, the relevance of CD69 in tissue fibrosis remains largely unknown. Thus, we explored the role of CD69 in fibroproliferative responses using a mouse model of peritoneal fibrosis induced by dialysis fluid exposure under either normal or uremic status. We found that cd69−/− mice compared with wild-type (WT) mice showed enhanced fibrosis, mesothelial to mesenchymal transition, IL-17 production, and Th17 cell infiltration in response to dialysis fluid treatment. Uremia contributed partially to peritoneal inflammatory and fibrotic responses. Additionally, antibody–mediated CD69 blockade in WT mice mimicked the fibrotic response of cd69−/− mice. Finally, IL-17 blockade in cd69−/− mice decreased peritoneal fibrosis to the WT levels, and mixed bone marrow from cd69−/− and Rag2−/−γc−/− mice transplanted into WT mice reproduced the severity of the response to dialysis fluid observed in cd69−/− mice, showing that CD69 exerts its regulatory function within the lymphocyte compartment. Overall, our results indicate that CD69 controls tissue fibrosis by regulating Th17-mediated inflammation. PMID:27151919

Inflammatory biomarkers for persistent fatigue in breast cancer survivors.

PubMed

Collado-Hidalgo, Alicia; Bower, Julienne E; Ganz, Patricia A; Cole, Steve W; Irwin, Michael R

2006-05-01

This study seeks to define immunologic and inflammatory variables associated with persistent post-treatment fatigue in breast cancer survivors. Leukocyte subsets, plasma inflammatory markers, and ex vivo proinflammatory cytokine production were assessed in 50 fatigued and nonfatigued breast cancer survivors recruited > or = 2 years after successful primary therapy. Multivariate statistical analyses were used to define a composite immunologic biomarker of fatigue risk. Fatigued breast cancer survivors were distinguished from nonfatigued survivors by increased ex vivo monocyte production of interleukin (IL)-6 and tumor necrosis factor-alpha following lipopolysaccharide stimulation, elevated plasma IL-1ra and soluble IL-6 receptor (sIL-6R/CD126), decreased monocyte cell-surface IL-6R, and decreased frequencies of activated T lymphocytes and myeloid dendritic cells in peripheral blood (all P < 0.05). An inverse correlation between sIL-6R and cell-surface IL-6R was consistent with inflammation-mediated shedding of IL-6R, and in vitro studies confirmed that proinflammatory cytokines induced such shedding. Multivariate linear discriminant function analysis identified two immunologic markers, the ratio of sIL-6R to monocyte-associated IL-6R and decreased circulating CD69+ T lymphocytes, as highly diagnostic of fatigue (P = 0.0005), with cross-validation estimates indicating 87% classification accuracy (sensitivity = 0.83; specificity = 0.83). These results extend links between fatigue and inflammatory markers to show a functional alteration in proinflammatory cytokine response to lipopolysaccharide and define a prognostic biomarker of behavioral fatigue.

Food supplement 20070721-GX may increase CD34+ stem cells and telomerase activity.

PubMed

Lin, Po-Cheng; Chiou, Tzyy-Wen; Liu, Po-Yen; Chen, Shee-Ping; Wang, Hsin-I; Huang, Pi-Chun; Lin, Shinn-Zong; Harn, Horng-Jyh

2012-01-01

Few rejuvenation and antiaging markers are used to evaluate food supplements. We measured three markers in peripheral blood to evaluate the antiaging effects of a food supplement containing placental extract. Samples were evaluated for CD34(+) cells, insulin-like growth factor 1 (IGF1), and telomerase activity, which are all markers related to aging. To control the quality of this food supplement, five active components were monitored. In total, we examined 44 individuals who took the food supplement from 1.2 months to 23 months; the average number of CD34(+) cells was almost 6-fold higher in the experimental group compared with the control group. Food supplement intake did not change serum IGF1 levels significantly. Finally, the average telomerase activity was 30% higher in the subjects taking this food supplement. In summary, our results suggest that the placental extract in the food supplement might contribute to rejuvenation and antiaging.

Serum Adipocytokine Levels as Surrogate Markers for Disease Activity of Crohn's Disease.

PubMed

Kim, Su Hwan; Jang, Seung Hyeon; Kim, Ji Won; Kim, Byeong Gwan; Lee, Kook Lae; Kim, You Sun; Han, Dong Soo; Kim, Joo Sung

2017-05-01

Determining inflammatory activity is crucial for assessing disease activity and for tailoring therapy in patients with Crohn׳s disease (CD). This study aimed to evaluate adipocytokine levels in patients with CD and to determine whether they can serve as surrogate markers for disease activity. Serum samples and information regarding the clinical features of patients in the CD Network Project registry were collected from March 2009 to February 2012. Patients with CD and disease duration of at least 2 years were enrolled in this study. Fasting serum leptin, adiponectin, obestatin and ghrelin levels were measured, and their correlation with clinical features of the patients was analyzed. Serum adipocytokine levels were evaluated according to disease activity as determined by CD activity index score. A total of 153 patients with CD were included. Serum ghrelin levels negatively correlated with patient age (P = 0.041) and age at diagnosis (P = 0.017), and positively correlated with C-reactive protein (CRP) levels (P = 0.017). Multiple regression analysis showed that serum ghrelin levels were related only to CRP levels (P = 0.032). Like ghrelin, serum leptin levels were also related to CRP levels (P < 0.001). Obestatin and adiponectin levels were not related to CRP levels. Serum adipocytokine levels did not significantly differ across different disease locations or behaviors. Serum ghrelin levels were significantly lower in patients with CD with a history of surgery than in those without (P = 0.007). Serum ghrelin and leptin levels may be useful as surrogate markers for disease activity in patients with CD. Copyright © 2017 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

Comparison of oxidative stress & leukocyte activation in patients with severe sepsis & burn injury

PubMed Central

Mühl, Diana; Woth, Gábor; Drenkovics, Livia; Varga, Adrienn; Ghosh, Subhamay; Csontos, Csaba; Bogár, Lajos; Wéber, György; Lantos, János

2011-01-01

Background & objectives: We evaluated pro- and anti-oxidant disturbances in sepsis and non-sepsis burn patients with systemic inflammatory response syndrome (SIRS). Adhesion molecules and inflammation markers on leukocytes were also analyzed. We hypothesized that oxidative stress and leukocyte activation markers can lead to the severity of sepsis. Methods: In 28 severe sepsis and 27 acute burn injury patients blood samples were collected at admission and 4 days consecutively. Oxidative stress markers: production of reactive oxygen species (ROS), myeloperoxidase, malondialdehyde and endogenous antioxidants: plasma protein sulphydryl groups, reduced glutathione, superoxide dismutase and catalase were measured. Flow cytometry was used to determine CD11a, CD14, CD18, CD49d and CD97 adhesion molecules on leukocytes. Procalcitonin, C-reactive protein, fibrinogen, platelet count and lactate were also analyzed. Results: Pro-oxidant parameters were significantly elevated in sepsis patients at admission, ROS intensity increased in burn patients until the 5th day. Endogenous antioxidant levels except catalase showed increased levels after burn trauma compared to sepsis. Elevated granulocyte activation and suppressed lymphocyte function were found at admission and early activation of granulocytes caused by increasing activation/migration markers in sepsis. Leukocyte adhesion molecule expression confirmed the suppressed lymphocyte and monocyte function in sepsis. Interpretation & conclusions: Severe sepsis is accompanied by oxidative stress and pathological leukocyte endothelial cell interactions. The laboratory parameters used for the evaluation of sepsis and several markers of pro- and antioxidant status were different between sepsis and non-sepsis burn patients. The tendency of changes in these parameters may refer to major oxidative stress in sepsis and developing SIRS in burns. PMID:21808137

Ex vivo tetramer staining and cell surface phenotyping for early activation markers CD38 and HLA-DR to enumerate and characterize malaria antigen-specific CD8+ T-cells induced in human volunteers immunized with a Plasmodium falciparum adenovirus-vectored malaria vaccine expressing AMA1.

PubMed

Schwenk, Robert; Banania, Glenna; Epstein, Judy; Kim, Yohan; Peters, Bjoern; Belmonte, Maria; Ganeshan, Harini; Huang, Jun; Reyes, Sharina; Stryhn, Anette; Ockenhouse, Christian F; Buus, Soren; Richie, Thomas L; Sedegah, Martha

2013-10-29

Malaria is responsible for up to a 600,000 deaths per year; conveying an urgent need for the development of a malaria vaccine. Studies with whole sporozoite vaccines in mice and non-human primates have shown that sporozoite-induced CD8+ T cells targeting liver stage antigens can mediate sterile protection. There is a need for a direct method to identify and phenotype malaria vaccine-induced CD8+ T cells in humans. Fluorochrome-labelled tetramers consisting of appropriate MHC class I molecules in complex with predicted binding peptides derived from Plasmodium falciparum AMA-1 were used to label ex vivo AMA-1 epitope specific CD8+ T cells from research subjects responding strongly to immunization with the NMRC-M3V-Ad-PfCA (adenovirus-vectored) malaria vaccine. The identification of these CD8+ T cells on the basis of their expression of early activation markers was also investigated. Analyses by flow cytometry demonstrated that two of the six tetramers tested: TLDEMRHFY: HLA-A*01:01 and NEVVVKEEY: HLA-B*18:01, labelled tetramer-specific CD8+ T cells from two HLA-A*01:01 volunteers and one HLA-B*18:01 volunteer, respectively. By contrast, post-immune CD8+ T cells from all six of the immunized volunteers exhibited enhanced expression of the CD38 and HLA-DRhi early activation markers. For the three volunteers with positive tetramer staining, the early activation phenotype positive cells included essentially all of the tetramer positive, malaria epitope- specific CD8+ T cells suggesting that the early activation phenotype could identify all malaria vaccine-induced CD8+ T cells without prior knowledge of their exact epitope specificity. The results demonstrated that class I tetramers can identify ex vivo malaria vaccine antigen-specific CD8+ T cells and could therefore be used to determine their frequency, cell surface phenotype and transcription factor usage. The results also demonstrated that vaccine antigen-specific CD8+ T cells could be identified by activation markers without prior knowledge of their antigen-specificity, using a subunit vaccine for proof-of-concept. Whether, whole parasite or adjuvanted protein vaccines will also induce {CD38 and HLA-DRhi}+ CD8+ T cell populations reflective of the antigen-specific response will the subject of future investigations.

Biomarker and Histopathology Evaluation of Patients with Recurrent Glioblastoma Treated with Galunisertib, Lomustine, or the Combination of Galunisertib and Lomustine

PubMed Central

Capper, David; von Deimling, Andreas; Brandes, Alba A.; Carpentier, Antoine F.; Kesari, Santosh; Sepulveda-Sanchez, Juan M.; Wheeler, Helen R.; Chinot, Olivier; Cher, Lawrence; Steinbach, Joachim P.; Specenier, Pol; Rodon, Jordi; Cleverly, Ann; Smith, Claire; Gueorguieva, Ivelina; Miles, Colin; Guba, Susan C.; Desaiah, Durisala; Estrem, Shawn T.; Lahn, Michael M.; Wick, Wolfgang

2017-01-01

Galunisertib, a Transforming growth factor-βRI (TGF-βRI) kinase inhibitor, blocks TGF-β-mediated tumor growth in glioblastoma. In a three-arm study of galunisertib (300 mg/day) monotherapy (intermittent dosing; each cycle =14 days on/14 days off), lomustine monotherapy, and galunisertib plus lomustine therapy, baseline tumor tissue was evaluated to identify markers associated with tumor stage (e.g., histopathology, Ki67, glial fibrillary acidic protein) and TGF-β-related signaling (e.g., pSMAD2). Other pharmacodynamic assessments included chemokine, cytokine, and T cell subsets alterations. 158 patients were randomized to galunisertib plus lomustine (n = 79), galunisertib (n = 39) and placebo+lomustine (n = 40). In 127 of these patients, tissue was adequate for central pathology review and biomarker work. Isocitrate dehydrogenase (IDH1) negative glioblastoma patients with baseline pSMAD2+ in cytoplasm had median overall survival (OS) 9.5 months vs. 6.9 months for patients with no tumor pSMAD2 expression (p = 0.4574). Eight patients were IDH1 R132H+ and had a median OS of 10.4 months compared to 6.9 months for patients with negative IDH1 R132H (p = 0.5452). IDH1 status was associated with numerically higher plasma macrophage-derived chemokine (MDC/CCL22), higher whole blood FOXP3, and reduced tumor CD3+ T cell counts. Compared to the baseline, treatment with galunisertib monotherapy preserved CD4+ T cell counts, eosinophils, lymphocytes, and the CD4/CD8 ratio. The T-regulatory cell compartment was associated with better OS with MDC/CCL22 as a prominent prognostic marker. PMID:28481241

Immunosenescence-Related Transcriptomic and Immunologic Changes in Older Individuals Following Influenza Vaccination

PubMed Central

Kennedy, Richard B.; Ovsyannikova, Inna G.; Haralambieva, Iana H.; Oberg, Ann L.; Zimmermann, Michael T.; Grill, Diane E.; Poland, Gregory A.

2016-01-01

The goal of annual influenza vaccination is to reduce mortality and morbidity associated with this disease through the generation of protective immune responses. The objective of the current study was to examine markers of immunosenescence and identify immunosenescence-related differences in gene expression, gene regulation, cytokine secretion, and immunologic changes in an older study population receiving seasonal influenza A/H1N1 vaccination. Surprisingly, prior studies in this cohort revealed weak correlations between immunosenescence markers and humoral immune response to vaccination. In this report, we further examined the relationship of each immunosenescence marker (age, T cell receptor excision circle frequency, telomerase expression, percentage of CD28− CD4+ T cells, percentage of CD28− CD8+ T cells, and the CD4/CD8 T cell ratio) with additional markers of immune response (serum cytokine and chemokine expression) and measures of gene expression and/or regulation. Many of the immunosenescence markers indeed correlated with distinct sets of individual DNA methylation sites, miRNA expression levels, mRNA expression levels, serum cytokines, and leukocyte subsets. However, when the individual immunosenescence markers were grouped by pathways or functional terms, several shared biological functions were identified: antigen processing and presentation pathways, MAPK, mTOR, TCR, BCR, and calcium signaling pathways, as well as key cellular metabolic, proliferation and survival activities. Furthermore, the percent of CD4+ and/or CD8+ T cells lacking CD28 expression also correlated with miRNAs regulating clusters of genes known to be involved in viral infection. Integrated (DNA methylation, mRNA, miRNA, and protein levels) network biology analysis of immunosenescence-related pathways and genesets identified both known pathways (e.g., chemokine signaling, CTL, and NK cell activity), as well as a gene expression module not previously annotated with a known function. These results may improve our ability to predict immune responses to influenza and aid in new vaccine development, and highlight the need for additional studies to better define and characterize immunosenescence. PMID:27853459

Serum neopterin and soluble CD163 as markers of macrophage activation in paracetamol (acetaminophen)-induced human acute liver injury.

PubMed

Craig, D G; Lee, P; Pryde, E A; Hayes, P C; Simpson, K J

2013-12-01

Macrophage activation is implicated in the pathogenesis of the systemic inflammatory response syndrome (SIRS) following paracetamol (acetaminophen) overdose (POD). Neopterin is synthesised from macrophages and reflects the intensity of monocyte/macrophage activation. Soluble CD163 (sCD163) is a marker of alternatively activated M2 macrophages. To examine neopterin and sCD163 levels in a cohort of acute liver injury patients. Consecutive patients (n = 41, (18 (43.9%) male) with acute liver injury were enrolled. Neopterin and sCD163 levels were measured by ELISA. A total of 24/33 (72.7%) POD patients developed hepatic encephalopathy (HE), and therefore acute liver failure. Both neopterin and sCD163 levels were significantly higher in PODs compared with chronic liver disease (neopterin P < 0.001, sCD163 P = 0.038) and healthy (both P < 0.001) controls. Admission neopterin levels were significantly higher in PODs: with HE (P = 0.001); with the SIRS (P = 0.005); who required renal replacement therapy (P = 0.003); who died or required liver transplantation (P = 0.006; AUROC 78.6% (95% CI 62.2-94.9%). Serum sCD163 levels were significantly higher in those PODs with the SIRS (P = 0.033) on admission, and were higher in those PODs who died or required OLT (P = 0.024). Both admission neopterin and sCD163 levels in PODs correlated with organ failure scores but not with serum ALT. There was no significant correlation between neopterin and sCD163 values. Both serum neopterin and sCD163 levels are significantly elevated following paracetamol overdose, and reflect the degree of macrophage activation in this condition. Serum neopterin in particular may have value as an early proxy marker of macrophage activation following paracetamol overdose. © 2013 John Wiley & Sons Ltd.

Expression of lactate transporters MCT1, MCT2 and CD147 in the red blood cells of three horse breeds: Finnhorse, Standardbred and Thoroughbred.

PubMed

Mykkänen, A K; Pösö, A R; McGowan, C M; McKane, S A

2010-11-01

In exercising horses, up to 50% of blood lactate is taken up into red blood cells (RBCs). Lactate transporter proteins MCT1, MCT2 and CD147 (an ancillary protein for MCT1) are expressed in the equine RBC membrane. In Standardbreds (SB), lactate transport activity is bimodally distributed and correlates with the amount of MCT1 and CD147. About 75% of SB studied have high lactate transport activity in RBCs. In other breeds, the distribution of lactate transport activity is unknown. To study whether similar bimodal distribution of MCT1 and CD147 is present also in the racing Finnhorse (FH) and Thoroughbred (TB) as in the SB and to study the distribution of MCT2 in all 3 breeds and to determine if there is a connection between MCT expression and performance markers in TB racehorses. Venous blood samples were taken from 118 FHs, 98 TBs and 44 SBs. Red blood cell membranes were purified and MCT1, MCT2 and CD147 measured by western blot. The amount of transporters was compared with TB performance markers. In TBs, the distribution of MCT1 was bimodal and in all breeds distribution of MCT2 unimodal. The amount of CD147 was clearly bimodal in FH and SB, with 85 and 82% expressing high amounts of CD147. In TBs, 88% had high expression of CD147 and 11% low expression, but one horse showed intermediate expression not apparent in FH or SB. Performance markers did not correlate with the amount of MCT1, MCT2 or CD147. High lactate transport activity was present in all 3 racing breeds, with the greatest proportion in the TB, followed by the racing FH, then SB. There was no significant statistical correlation found between lactate transporters in RBC membrane and markers of racing performance in the TB. © 2010 EVJ Ltd.

Conditions in blood sampling procedures that extend the ex vivo stability of eosinophil activity markers in peripheral blood from allergic patients and healthy controls.

PubMed

Halldén, G; Nopp, A; Ihre, E; Peterson, C; Lundahl, J

1999-11-01

Serum-ECP, EG2-epitope on intracellular ECP and surface expression of CD9 and CD11b in peripheral blood eosinophils (PBE) are considered to be markers that mirror clinical parameters in allergic inflammation. The aim was to investigate the impact of the blood sampling procedure on PBE markers and to identify optimal conditions for extended pre-analysis storage. Blood, from healthy individuals and patients with allergic rhinitis/asthma, was collected in tubes with EDTA, citrate, or without anti-coagulant. The expression of EG2-epitope, CD9, and CD11b were analyzed in eosinophils and neutrophils after 1, 5, and 24 hours of storage at +4 degrees C, according to the FOG-method and flow cytometry. In vitro stimulation with fMLP/PMA was used for metabolic activity analysis and CD11b mobilization. Following a 1-hour clotting period at +20 to 22 degrees C, samples were stored at +4 degrees C and serum-ECP levels were measured. The EG2-epitope, serum-ECP, and CD9 were stable in samples from both healthy controls and allergic patients at all storage conditions. The EG2-epitope, serum-ECP and PBE count were significantly increased in the patient group, whereas no differences were observed in the expression of CD9 or CD11b. Both granulocytes and monocytes retained their metabolic activity for 24 hours. Neutrophils in citrate-blood increased their ability to respond to fMLP, as compared with EDTA-blood. In vitro analysis of selected activity markers and functional tests could be performed on granulocytes from both healthy individuals and allergic patients after 24 hours storage at +4 degrees C. The anticoagulant citrate seems to be preferable to EDTA when monocytes or CD11b expression are analyzed.

Clinical-Grade Human Multipotent Adult Progenitor Cells Block CD8+ Cytotoxic T Lymphocytes

PubMed Central

Dekimpe, Emily; Van Woensel, Matthias; Roobrouck, Valerie D.; Bullens, Dominique M.; Pinxteren, Jef; Verfaillie, Catherine M.; Van Gool, Stefaan W.

2016-01-01

MultiStem cells are clinical-grade multipotent adult bone marrow-derived progenitor cells (MAPCs), with extensive replication potential and broader differentiation capacity compared with mesenchymal stem cells. Human MAPCs suppress T-cell proliferation induced by alloantigens and mutually interact with allogeneic natural killer cells. In this study, the interaction between MultiStem and CD8+ cytotoxic T lymphocytes (CTLs) was addressed for the first time. In an in vitro setting, the immunogenicity of MultiStem, the susceptibility of MultiStem toward CTL-mediated lysis, and its effects on CTL function were investigated. MultiStem was nonimmunogenic for alloreactive CTL induction and was—even after major histocompatibility complex class I upregulation—insensitive to alloantigen-specific CTL-mediated lysis. Furthermore, MultiStem reduced CTL proliferation and significantly decreased perforin expression during the T-cell activation phase. As a consequence, MultiStem dose-dependently impaired the induction of CTL function. These effects of MultiStem were mediated predominantly through contact-dependent mechanisms. Moreover, MultiStem cells considerably influenced the expression of T-cell activation markers CD25, CD69, and human leukocyte antigen-DR. The MultiStem-induced CD8−CD69+ T-cell population displayed a suppressive effect on the induction of CTL function during a subsequent mixed-lymphocyte culture. Finally, the killer activity of activated antigen-specific CTLs during their cytolytic effector phase was also diminished in the presence of MultiStem. This study confirms that these clinical-grade MAPCs are an immune-modulating population that inhibits CTL activation and effector responses and are, consequently, a highly valuable cell population for adoptive immunosuppressive therapy in diseases where damage is induced by CTLs. Significance Because multipotent adult progenitor cells (MAPCs) are among the noteworthy adult mesenchymal stem cell populations for immune therapy and have the advantage over mesenchymal stem cells (MSCs) of large-scale manufacturing and banking potential and thus prompt availability, it is important to understand how MAPCs interact with immune cells to validate their widespread therapeutic applicability. Cytotoxic immune effector cells play a crucial role in immune homeostasis and in the pathogenesis of some autoimmune diseases. This study assessed for the first time the in vitro influence of a clinical-grade human MAPC product (MultiStem) on the cytotoxic function of CD8+ T cells (CTLs) by evaluating the immunogenicity of MAPCs and the susceptibility of MAPCs toward CTL-mediated lysis and by analyzing the mechanism of MAPC-mediated modulation of CTL functionality. These results may represent a highly relevant contribution to the current knowledge and, in combination with the results of future phase II/III trials using MultiStem, could lead to an intriguing continuation of stem cell-based research for immunotherapy. PMID:27465071

Clinical-Grade Human Multipotent Adult Progenitor Cells Block CD8+ Cytotoxic T Lymphocytes.

PubMed

Plessers, Jeroen; Dekimpe, Emily; Van Woensel, Matthias; Roobrouck, Valerie D; Bullens, Dominique M; Pinxteren, Jef; Verfaillie, Catherine M; Van Gool, Stefaan W

2016-12-01

: MultiStem cells are clinical-grade multipotent adult bone marrow-derived progenitor cells (MAPCs), with extensive replication potential and broader differentiation capacity compared with mesenchymal stem cells. Human MAPCs suppress T-cell proliferation induced by alloantigens and mutually interact with allogeneic natural killer cells. In this study, the interaction between MultiStem and CD8 + cytotoxic T lymphocytes (CTLs) was addressed for the first time. In an in vitro setting, the immunogenicity of MultiStem, the susceptibility of MultiStem toward CTL-mediated lysis, and its effects on CTL function were investigated. MultiStem was nonimmunogenic for alloreactive CTL induction and was-even after major histocompatibility complex class I upregulation-insensitive to alloantigen-specific CTL-mediated lysis. Furthermore, MultiStem reduced CTL proliferation and significantly decreased perforin expression during the T-cell activation phase. As a consequence, MultiStem dose-dependently impaired the induction of CTL function. These effects of MultiStem were mediated predominantly through contact-dependent mechanisms. Moreover, MultiStem cells considerably influenced the expression of T-cell activation markers CD25, CD69, and human leukocyte antigen-DR. The MultiStem-induced CD8 - CD69 + T-cell population displayed a suppressive effect on the induction of CTL function during a subsequent mixed-lymphocyte culture. Finally, the killer activity of activated antigen-specific CTLs during their cytolytic effector phase was also diminished in the presence of MultiStem. This study confirms that these clinical-grade MAPCs are an immune-modulating population that inhibits CTL activation and effector responses and are, consequently, a highly valuable cell population for adoptive immunosuppressive therapy in diseases where damage is induced by CTLs. Because multipotent adult progenitor cells (MAPCs) are among the noteworthy adult mesenchymal stem cell populations for immune therapy and have the advantage over mesenchymal stem cells (MSCs) of large-scale manufacturing and banking potential and thus prompt availability, it is important to understand how MAPCs interact with immune cells to validate their widespread therapeutic applicability. Cytotoxic immune effector cells play a crucial role in immune homeostasis and in the pathogenesis of some autoimmune diseases. This study assessed for the first time the in vitro influence of a clinical-grade human MAPC product (MultiStem) on the cytotoxic function of CD8 + T cells (CTLs) by evaluating the immunogenicity of MAPCs and the susceptibility of MAPCs toward CTL-mediated lysis and by analyzing the mechanism of MAPC-mediated modulation of CTL functionality. These results may represent a highly relevant contribution to the current knowledge and, in combination with the results of future phase II/III trials using MultiStem, could lead to an intriguing continuation of stem cell-based research for immunotherapy. ©AlphaMed Press.

The Role of CD56 in Thyroid Fine Needle Aspiration Cytology: A Pilot Study Performed on Liquid Based Cytology

PubMed Central

Bizzarro, Tommaso; Martini, Maurizio; Marrocco, Carla; D’Amato, Donato; Traini, Emanuela; Lombardi, Celestino Pio; Pontecorvi, Alfredo; Fadda, Guido; Larocca, Luigi Maria; Rossi, Esther Diana

2015-01-01

Background Fine needle aspiration Cytology (FNAC) fulfills a reliable role in the evaluation of thyroid lesions. Although the majority of nodules are quite easily diagnosed as benign or malignant, 30% of them represent an indeterminate category whereby the application of ancillary techniques (i.e. immunocytochemistry-ICC and molecular testing) has been encouraged. The search for a specific immunomarker of malignancy sheds light on a huge number of ICC stains although none of them attempt to yield 100% conclusive results. Our aim was to define in a pilot study on thyroid FNAC whether CD56 might be a valid marker also in comparison with HBME-1 and Galectin-3. Materials and Methods Inasmuch as this is the largest pilot study using only liquid based cytology (LBC), we selected all the cases only in the categories of benign nodules (BN) and positive for malignancy (PM) for validation purposes. Eighty-five consecutive (including 50 PM and 35 BN) out of 950 thyroid FNACs had surgical follow-up. The ICC panel (HBME-1, Galectin-3 and CD56) was carried out on LBC and histology. Results All BNs and PMs were histological confirmed. CD56 was negative in 96% of the PM while 68.5% of the BNs showed cytoplasmic positivity for this marker, with an overall high sensitivity (96%) but lower specificity (69%). In specific, our 96% of the PMs did not show any follicular cell with CD56 expression. Different ICC combinations were evaluated showing that the panel made up of CD56 plus HBME-1 and Galectin-3 had the highest sensitivity (98%) and specificity (86%). Conclusions Our pilot study suggests that CD56 may be a good marker for ruling out PTC and its variants. The low specificity suggests that an immunopanel including also HBME-1 and Galectin-3 could obtain the highest diagnostic accuracy in thyroid lesions. Our results suggest that CD56 may be a feasible additional marker for identifying malignancies also in the FNs and SMs. PMID:26186733

Autonomic Dysfunction Correlates with Clinical and Inflammatory Activity in Patients with Crohn's Disease.

PubMed

Engel, Tal; Ben-Horin, Shomron; Beer-Gabel, Marc

2015-10-01

Autonomic dysfunction has been implicated in Crohn's disease (CD). We aimed to investigate heart rate variability (HRV) as a marker of possible autonomic imbalance in patients with CD. Thirty patients with CD and 30 age- and gender-matched healthy controls were enrolled in a prospective cohort study and underwent HRV study. Anxiety level was scored using the STAI questionnaire and CD clinical activity was assessed by Harvey-Bradshaw index. Blood tests including inflammatory markers were obtained for all participants. CD subjects had lower mean blood pressure (85.51 ± 11.07 mm Hg, 91.51 ± 6.99, P = 0.015) and albumin and significantly higher CRP and IL-6 compared with controls (P < 0.002 for all comparisons). Mean HRV values of very low-frequency power and low-frequency power components were significantly lower among CD subjects (P = 0.038 and 0.027, respectively), implying a predominant sympathetic tone. Anxiety level scores were significantly higher among patients with CD for both state anxiety (P = 0.001) and trait anxiety (P < 0.0001). However, patients with active disease had similar anxiety scores as patients in remission, yet had a significantly lower BMI, lower albumin level, and higher CRP and IL-6 levels (P < 0.05 for all comparisons). Moreover, despite similar anxiety scores, patients with active disease had higher pulse rate (P = 0.02) and lower HRV indexes, which correlated with albumin levels(r = 0.7, P = 0.001). Although patients with CD have higher anxiety levels compared with controls, they exhibit depressed HRV independent of this anxiety state and in direct correlation with disease activity and inflammatory markers. These observations suggest an inherent imbalance of autonomic function associated with active inflammation.

Herpes simplex virus type 2 serostatus is not associated with inflammatory or metabolic markers in antiretroviral therapy-treated HIV.

PubMed

Tan, Darrell H S; Raboud, Janet M; Szadkowski, Leah; Yi, Tae Joon; Shannon, Brett; Kaul, Rupert; Liles, W Conrad; Walmsley, Sharon L

2015-03-01

Systemic inflammation and immune activation may persist in HIV-infected persons on suppressive combination antiretroviral therapy (cART) and contribute to adverse health outcomes. We compared markers of immune activation, inflammation, and abnormal glucose and lipid metabolism in HIV-infected adults according to herpes simplex virus type 2 (HSV-2) serostatus in a 6-month observational cohort study in Toronto, Canada. HIV-infected adults on suppressive (viral load <50 copies/ml) cART were categorized as HSV-2 seropositive or seronegative using the HerpeSelect ELISA, and underwent study visits at baseline, 3 months, and 6 months. The primary outcome was the median percentage of activated (CD38(+)HLADR(+)) CD8 T cells. Secondary outcome measures included additional immune (activated CD4, regulatory T cells) and inflammatory (hsCRP, D-dimer, IL-1b, IL-6, MCP-1, TNF, sICAM-1, sVCAM-1, Ang1/Ang2 ratio) markers. Metabolic outcomes included the proportion with impaired fasting glucose/impaired glucose tolerance/diabetes, insulin sensitivity (calculated using the Matsuda index), insulin resistance (homeostasis model assessment of insulin resistance), and fasting lipids. The impact of HSV-2 on each outcome was estimated using generalized estimating equation regression models. Of 84 participants, 38 (45%) were HSV-2 seropositive. HSV signs and symptoms were uncommon. Aside from D-dimer, which was more often detectable in HSV-2 seropositives (adjusted odds ratio=3.58, 95% CI=1.27, 10.07), HSV-2 serostatus was not associated with differences in any other immune, inflammatory cytokine, acute phase reactant, endothelial activation, or metabolic markers examined in univariable or multivariable models. During the study, CD8 and CD4 T cell activation declined by 0.16% and 0.08% per month, respectively, while regulatory T cells increased by 0.05% per month. HSV-2 serostatus was not consistently associated with immune activation, inflammatory, or lipid and glucose metabolic markers in this cohort of HIV-infected adults on suppressive cART.

EMMPRIN (CD147) is induced by C/EBPβ and is differentially expressed in ALK+ and ALK- anaplastic large-cell lymphoma.

PubMed

Schmidt, Janine; Bonzheim, Irina; Steinhilber, Julia; Montes-Mojarro, Ivonne A; Ortiz-Hidalgo, Carlos; Klapper, Wolfram; Fend, Falko; Quintanilla-Martínez, Leticia

2017-09-01

Anaplastic lymphoma kinase-positive (ALK+) anaplastic large-cell lymphoma (ALCL) is characterized by expression of oncogenic ALK fusion proteins due to the translocation t(2;5)(p23;q35) or variants. Although genotypically a T-cell lymphoma, ALK+ ALCL cells frequently show loss of T-cell-specific surface antigens and expression of monocytic markers. C/EBPβ, a transcription factor constitutively overexpressed in ALK+ ALCL cells, has been shown to play an important role in the activation and differentiation of macrophages and is furthermore capable of transdifferentiating B-cell and T-cell progenitors to macrophages in vitro. To analyze the role of C/EBPβ for the unusual phenotype of ALK+ ALCL cells, C/EBPβ was knocked down by RNA interference in two ALK+ ALCL cell lines, and surface antigen expression profiles of these cell lines were generated using a Human Cell Surface Marker Screening Panel (BD Biosciences). Interesting candidate antigens were further analyzed by immunohistochemistry in primary ALCL ALK+ and ALK- cases. Antigen expression profiling revealed marked changes in the expression of the activation markers CD25, CD30, CD98, CD147, and CD227 after C/EBPβ knockdown. Immunohistochemical analysis confirmed a strong, membranous CD147 (EMMPRIN) expression in ALK+ ALCL cases. In contrast, ALK- ALCL cases showed a weaker CD147 expression. CD274 or PD-L1, an immune inhibitory receptor ligand, was downregulated after C/EBPβ knockdown. PD-L1 also showed stronger expression in ALK+ ALCL compared with ALK- ALCL, suggesting an additional role of C/EBPβ in ALK+ ALCL in generating an immunosuppressive environment. Finally, no expression changes of T-cell or monocytic markers were detected. In conclusion, surface antigen expression profiling demonstrates that C/EBPβ plays a critical role in the activation state of ALK+ ALCL cells and reveals CD147 and PD-L1 as important downstream targets. The multiple roles of CD147 in migration, adhesion, and invasion, as well as T-cell activation and proliferation suggest its involvement in the pathogenesis of ALCL.

Upregulation of CD147 Promotes Metastasis of Cholangiocarcinoma by Modulating the Epithelial-to-Mesenchymal Transitional Process.

PubMed

Dana, Paweena; Kariya, Ryusho; Vaeteewoottacharn, Kulthida; Sawanyawisuth, Kanlayanee; Seubwai, Wunchana; Matsuda, Kouki; Okada, Seiji; Wongkham, Sopit

2017-08-07

CD147 is a transmembrane protein that can induce the expression and activity of matrix metalloproteinases (MMPs). Expression of CD147 has been shown to potentiate cell migration, invasion, and metastasis of cancer. In this study, the critical role of CD147 in metastasis was elucidated using CD147-overexpressing cholangiocarcinoma (CCA) cells in vitro and in vivo. The molecular mechanism, demonstrated herein, supported the hypothesis that metastasis increased in CD147-overexpressing cells. Five CD147-overexpressing clones (Ex-CD147) were established from a low CD147-expressing CCA cell line, KKU-055, using lentivirus containing pReceiver-Lenti-CD147. The metastatic capability was determined using the tail vein injection mouse model and an in vitro 3D invasion assay. Liver colonization was assessed using anti-HLA class I immunohistochemistry. Adhesion abilities, cytoskeletal arrangements, MMP activities, the expressions of adhesion molecules, and epithelial-mesenchymal transitional markers were analyzed. All Ex-CD147 clones exhibited a high CD147 expression and high liver colonization in the tail vein-injected mouse model, whereas parental cells lacked this ability. Ex-CD147 clones exhibited metastatic phenotypes (i.e., an increase in F-actin rearrangement) and cell invasion and a decrease in cell adhesion. The molecular mechanisms were shown to be via the induction of MMP-2 activity and enhancement of epithelial-mesenchymal transitions. An increase in mesenchymal markers Slug, vimentin, and N-cadherin, and a decrease in epithelial markers E-cadherin and claudin-1, together with suppression of the adhesion molecule ICAM-1, were observed in the Ex-CD147 clones. Moreover, suppression of CD147 expression using siCD147 in two CCA cell lines with high CD147 expression significantly decreased cell migration and invasion of these CCA cells. These findings emphasize the essential role of CD147 in CCA metastasis and suggest CD147 as a promising target for the effective treatment of CCA.

Pre-transplant immune state defined by serum markers and alloreactivity predicts acute rejection after living donor kidney transplantation.

PubMed

Vondran, Florian W R; Timrott, Kai; Kollrich, Sonja; Steinhoff, Ann-Kristin; Kaltenborn, Alexander; Schrem, Harald; Klempnauer, Juergen; Lehner, Frank; Schwinzer, Reinhard

2014-09-01

Acute rejection (AR) remains a major cause for long-term kidney allograft failure. Reliable immunological parameters suitable to define the pre-transplant immune state and hence the individual risk of graft rejection are highly desired to preferably adapt the immunosuppressive regimen in advance. Donor and third party alloreactivities were determined by mixed lymphocyte cultures. Soluble forms of CD25, CD30, and CD44 were detected in patients' serum by ELISA. Various lymphocyte subpopulations were measured using flow cytometry. All patients received triple immunosuppression (tacrolimus/mycophenolate mofetil/steroids) and were grouped according to biopsy results within the first year: rejection-free (RF, n = 13), borderline (BL, n = 5), or acute rejection (AR, n = 7). Patients with AR showed the highest pre-transplant alloreactivities and serum levels (sCD25/sCD30/sCD44) according to the pattern RF < BL < AR. Relying on serum analysis only, multivariate logistic regression (logit link function) yielded a prognostic score for prediction of rejection with 75.0% sensitivity and 69.2% specificity. Patients with rejection showed markedly higher pre-transplant frequencies of CD4(+) /CD8(+) T cells lacking CD28, but lower numbers of CD8(+) CD161(bright) T cells and NK cells than RF individuals. Pre-transplant immune state defined by alloreactivity, serum markers, and particular lymphocyte subsets seems to correlate with occurrence of graft rejection after kidney transplantation. A prognostic score based on pre-transplant serum levels has shown great potential for prediction of rejection episodes and should be further evaluated. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

HIV turns plasmacytoid dendritic cells (pDC) into TRAIL-expressing killer pDC and down-regulates HIV coreceptors by Toll-like receptor 7-induced IFN-alpha.

PubMed

Hardy, Andrew W; Graham, David R; Shearer, Gene M; Herbeuval, Jean-Philippe

2007-10-30

Plasmacytoid dendritic cells (pDC) are key players in viral immunity and produce IFN-alpha after HIV-1 exposure, which in turn regulates TNF-related apoptosis-inducing ligand (TRAIL) expression by CD4(+) T cells. We show here that infectious and noninfectious HIV-1 virions induce activation of pDC into TRAIL-expressing IFN-producing killer pDC (IKpDC). IKpDC expressed high levels of activation markers (HLA-DR, CD80, CD83, and CD86) and the migration marker CCR7. Surprisingly, CXCR4 and CCR5 were down-regulated on IKpDC. We also show that HIV-1-induced IKpDC depended on Toll-like receptor 7 (TLR7) activation. HIV-1 or TLR7 agonistexposed IKpDC induced apoptosis of the CD4(+) T cell line SupT1 via the TRAIL pathway. Furthermore, IFN-alpha produced after HIV-induced TLR7 stimulation was responsible for TRAIL expression and the down-regulation of both CXCR4 and CCR5 by IKpDC. In contrast, activation and migration markers were not regulated by IFN-alpha. Finally, IFN-alpha increased the survival of IKpDC. We characterized a subset of pDC with a killer activity that is activated by endosomal-associated viral RNA and not by infection.

Comprehensive intestinal T helper cell profiling reveals specific accumulation of IFN-γ+IL-17+coproducing CD4+ T cells in active inflammatory bowel disease.

PubMed

Globig, Anna-Maria; Hennecke, Nadine; Martin, Bianca; Seidl, Maximilian; Ruf, Günther; Hasselblatt, Peter; Thimme, Robert; Bengsch, Bertram

2014-12-01

Skewed T helper (TH) cell responses and specific functions of TH1, TH2, TH17, and Treg cells have been implicated in the pathogenesis of inflammatory bowel disease (IBD) that led to the establishment of the pathogenic TH1/TH2 and TH17/Treg cell imbalance paradigms. However, the relevant TH cell population driving mucosal inflammation is still unknown. We performed a comprehensive TH cell profiling of circulating and intestinal lymphocytes isolated from patients with Crohn's disease (CD; n = 69) and ulcerative colitis (UC; n = 41) undergoing endoscopy or surgical resection and compared them with healthy controls (n = 45). Mucosal inflammation was assessed endoscopically and histologically. TH cells were analyzed by flow cytometric evaluation of cytokine production and differentiation marker expression. Specialized TH cell populations were enriched in the intestinal mucosa compared with peripheral blood. Specifically, we observed a concomitant upregulation of TH17 cells and Tregs in active inflammatory lesions in patients with both CD and UC compared with quiescent/mildly inflamed lesions and healthy tissue. Of note, interferon γ+ interleukin (IL)-17+coproducing CD4+ T cells with high expression of T-bet, CD26, and IL-22 resembling recently described pathogenic TH17 cells were specifically enriched in the inflamed mucosal tissue. Our results argue against the controversial TH1/TH2 or TH17/Treg paradigms. In contrast, they suggest that a subpopulation of TH17 cells sharing a TH1 signature may be specifically involved in intestinal inflammation in CD and UC. These findings provide a better understanding of IBD pathogenesis and may help explain the efficacy of anti-IL-12p40/IL-23 and failure of anti-IL-17A therapies despite the enrichment of TH17 cells.

Human muscle cells express a B7-related molecule, B7-H1, with strong negative immune regulatory potential: a novel mechanism of counterbalancing the immune attack in idiopathic inflammatory myopathies.

PubMed

Wiendl, Heinz; Mitsdoerffer, Meike; Schneider, Dagmar; Chen, Lieping; Lochmüller, Hanns; Melms, Arthur; Weller, Michael

2003-10-01

B7-H1 is a novel B7 family protein attributed to costimulatory and immune regulatory functions. Here we report that human myoblasts cultured from control subjects and patients with inflammatory myopathies as well as TE671 muscle rhabdomyosarcoma cells express high levels of B7-H1 after stimulation with the inflammatory cytokine IFN-gamma. Coculture experiments of MHC class I/II-positive myoblasts with CD4 and CD8 T cells in the presence of antigen demonstrated the functional consequences of muscle-related B7-H1 expression: production of inflammatory cytokines, IFN-gamma and IL-2, by CD4 as well CD8 T cells was markedly enhanced in the presence of a neutralizing anti-B7-H1 antibody. This observation was paralleled by an augmented expression of the T cell activation markers CD25, ICOS, and CD69, thus showing B7-H1-mediated inhibition of T cell activation. Further, we investigated 23 muscle biopsy specimens from patients with polymyositis (PM), inclusion body myositis (IBM), dermatomyositis (DM), and nonmyopathic controls for B7-H1 expression by immunohistochemistry: B7-H1 was expressed in PM, IBM, and DM specimens but not in noninflammatory and nonmyopathic controls. Staining was predominantly localized to areas of strong inflammation and to muscle cells as well as mononuclear cells. These data highlight the immune regulatory properties of muscle cells and suggest that B7-H1 expression represents an inhibitory mechanism induced upon inflammatory stimuli and aimed at protecting muscle fibers from immune aggression.

Hypergravity-induced immunomodulation in a rodent model: lymphocytes and lymphoid organs

NASA Technical Reports Server (NTRS)

Gridley, Daila S.; Pecaut, Michael J.; Green, Lora M.; Miller, Glen M.; Nelson, Gregory A.

2002-01-01

The major goal of this study was to quantify changes in lymphoid organs and cells over time due to centrifugation-induced hypergravity. C57BL/6 mice were exposed to 1, 2 and 3 G and the following assays were performed on days 1, 4, 7, 10, and 21: spleen, thymus, lung, and liver masses; total leukocyte, lymphocyte, monocyte/macrophage, and granulocyte counts; level of splenocyte apoptosis; enumeration of CD3+ T, CD3+/CD4+ T helper, CD3+/CD8+ T cytotoxic, B220+ B, and NK1.1+ natural killer cells; and quantification of cells expressing CD25, CD69, and CD71 activation markers. The data show that increased gravity resulted in decreased body, spleen, thymus, and liver, but not lung, mass. Significant reductions were noted in all three major leukocyte populations (lymphocytes, granulocytes, monocyte/macrophages) [correction of macrphages] with increased gravity; persistent depletion was noted in blood but not spleen. Among the various lymphocyte populations, the CD3+/CD8+ T cells and B220+ B cells were the most affected and NK1.1+ NK cells the least affected. Overall, the changes were most evident during the first week, with a greater influence noted for cells in the spleen. A linear relationship was found between some of the measurements and the level of gravity, especially on day 4. These findings indicate that hypergravity profoundly alters leukocyte number and distribution in a mammalian model and that some aberrations persisted throughout the three weeks of the study. In certain cases, the detected changes were similar to those observed after whole-body irradiation. In future investigations we hope to combine hypergravity with low-dose rate irradiation and immune challenge.

Glucagon-like peptide-1 receptor expression on human eosinophils and its regulation of eosinophil activation.

PubMed

Mitchell, P D; Salter, B M; Oliveria, J P; El-Gammal, A; Tworek, D; Smith, S G; Sehmi, R; Gauvreau, G M; Butler, M; O'Byrne, P M

2017-03-01

Glucagon-like peptide-1 (GLP-1) and its receptor are part of the incretin family of hormones that regulate glucose metabolism. GLP-1 also has immune modulatory roles. To measure the expression of the GLP-1 receptor (GLP-1R) on eosinophils and neutrophils in normal and asthmatic subjects and evaluate effects of a GLP-1 analog on eosinophil function. Peripheral blood samples were taken from 10 normal and 10 allergic asthmatic subjects. GLP-1R expression was measured on eosinophils and neutrophils. Subsequently, the asthmatic subjects underwent allergen and diluent inhalation challenges, and GLP-1R expression was measured. Purified eosinophils, collected from mild asthmatic subjects, were stimulated with lipopolysaccharide (LPS) and a GLP-1 analog to evaluate eosinophil cell activation markers CD11b and CD69 and cytokine (IL-4, IL-5, IL-8 and IL-13) production. Glucagon-like peptide-1 receptor is expressed on human eosinophils and neutrophils. Eosinophil, but not neutrophil, expression of GLP-1R is significantly higher in normal controls compared to allergic asthmatics. The expression of GLP-1R did not change on either eosinophils or neutrophils following allergen challenge. A GLP-1 analog significantly decreased the expression of eosinophil-surface activation markers following LPS stimulation and decreased eosinophil production of IL-4, IL-8 and IL-13, but not IL-5. Glucagon-like peptide-1 receptor is expressed on human eosinophils and neutrophils. A GLP-1 analog attenuates LPS-stimulated eosinophil activation. GLP-1 agonists may have additional adjunctive indications in treating persons with concomitant type 2 diabetes mellitus and asthma. © 2016 John Wiley & Sons Ltd.

Canonical Wnt Signaling as a Specific Marker for Normal and Tumorigenic Mammary Stem Cells

DTIC Science & Technology

2012-02-01

These cells were identified by  flow   cytometry  to detect cells that were positive for CD24 and CD49f.   2. We have established that activation of Wnt...09-1-0072 TITLE: Canonical Wnt Signaling as a Specific marker for Normal and Tumorigenic Mammary Stem Cells PRINCIPAL...activation of canonical Wnt signaling may be a very specific marker for mammary stem cells and be a target for transformation that results in the

Role of Lymphocyte Activation Gene-3 (Lag-3) in Conventional and Regulatory T Cell Function in Allogeneic Transplantation

PubMed Central

Sega, Emanuela I.; Leveson-Gower, Dennis B.; Florek, Mareike; Schneidawind, Dominik; Luong, Richard H.; Negrin, Robert S.

2014-01-01

Lag-3 has emerged as an important molecule in T cell biology. We investigated the role of Lag-3 in conventional T cell (Tcon) and regulatory T cell (Treg) function in murine GVHD with the hypothesis that Lag-3 engagement diminishes alloreactive T cell responses after bone marrow transplantation. We demonstrate that Lag-3 deficient Tcon (Lag-3−/− Tcon) induce significantly more severe GVHD than wild type (WT) Tcon and that the absence of Lag-3 on CD4 but not CD8 T cells is responsible for exacerbating GVHD. Lag-3−/− Tcon exhibited increased activation and proliferation as indicated by CFSE and bioluminescence imaging analyses and higher levels of activation markers such as CD69, CD107a, granzyme B, and Ki-67 as well as production of IL-10 and IFN-g early after transplantation. Lag-3−/− Tcon were less responsive to suppression by WT Treg as compared to WT Tcon. The absence of Lag-3, however, did not impair Treg function as both Lag-3−/− and WT Treg equally suppress the proliferation of Tcon in vitro and in vivo and protect against GVHD. Further, we demonstrate that allogeneic Treg acquire recipient MHC class II molecules through a process termed trogocytosis. As MHC class II is a ligand for Lag-3, we propose a novel suppression mechanism employed by Treg involving the acquisition of host MHC-II followed by the engagement of Lag-3 on T cells. These studies demonstrate for the first time the biologic function of Lag-3 expression on conventional and regulatory T cells in GVHD and identify Lag-3 as an important regulatory molecule involved in alloreactive T cell proliferation and activation after bone marrow transplantation. PMID:24475140

Increased CD69 Expression on Peripheral Eosinophils from Patients with Food Protein-Induced Enterocolitis Syndrome.

PubMed

Wada, Taizo; Matsuda, Yusuke; Toma, Tomoko; Koizumi, Eiko; Okamoto, Hiroyuki; Yachie, Akihiro

2016-01-01

Food protein-induced enterocolitis syndrome (FPIES) is an uncommon, non-IgE-mediated food allergy. We recently described a significant increase in fecal eosinophil-derived neurotoxin (EDN) after ingestion of the causative food. However, little is known about the activation status of circulating eosinophils in patients with an acute FPIES reaction. Surface CD69 expression was assessed by flow cytometry on peripheral eosinophils from 5 patients with FPIES before and after ingestion of the causative food. Fecal EDN was measured by enzyme-linked immunosorbent assay. No eosinophil activation was observed before ingestion; however, a significant increase in CD69 expression on eosinophils after an acute FIPES reaction was demonstrated in all of the patients. There was no significant change in absolute eosinophil counts in the peripheral blood. The levels of fecal EDN increased on the day after ingestion of the causative food in all patients. These results suggest that circulating eosinophils as well as eosinophils in the intestinal mucosal tissue are activated in acute FPIES reactions and might be associated with systemic immune events in FPIES. © 2016 S. Karger AG, Basel.

Antigen-Presenting Intratumoral B Cells Affect CD4+ TIL Phenotypes in Non-Small Cell Lung Cancer Patients.

PubMed

Bruno, Tullia C; Ebner, Peggy J; Moore, Brandon L; Squalls, Olivia G; Waugh, Katherine A; Eruslanov, Evgeniy B; Singhal, Sunil; Mitchell, John D; Franklin, Wilbur A; Merrick, Daniel T; McCarter, Martin D; Palmer, Brent E; Kern, Jeffrey A; Slansky, Jill E

2017-10-01

Effective immunotherapy options for patients with non-small cell lung cancer (NSCLC) are becoming increasingly available. The immunotherapy focus has been on tumor-infiltrating T cells (TILs); however, tumor-infiltrating B cells (TIL-Bs) have also been reported to correlate with NSCLC patient survival. The function of TIL-Bs in human cancer has been understudied, with little focus on their role as antigen-presenting cells and their influence on CD4 + TILs. Compared with other immune subsets detected in freshly isolated primary tumors from NSCLC patients, we observed increased numbers of intratumoral B cells relative to B cells from tumor-adjacent tissues. Furthermore, we demonstrated that TIL-Bs can efficiently present antigen to CD4 + TILs and alter the CD4 + TIL phenotype using an in vitro antigen-presentation assay. Specifically, we identified three CD4 + TIL responses to TIL-Bs, which we categorized as activated, antigen-associated, and nonresponsive. Within the activated and antigen-associated CD4 + TIL population, activated TIL-Bs (CD19 + CD20 + CD69 + CD27 + CD21 + ) were associated with an effector T-cell response (IFNγ + CD4 + TILs). Alternatively, exhausted TIL-Bs (CD19 + CD20 + CD69 + CD27 - CD21 - ) were associated with a regulatory T-cell phenotype (FoxP3 + CD4 + TILs). Our results demonstrate a new role for TIL-Bs in NSCLC tumors in their interplay with CD4 + TILs in the tumor microenvironment, establishing them as a potential therapeutic target in NSCLC immunotherapy. Cancer Immunol Res; 5(10); 898-907. ©2017 AACR . ©2017 American Association for Cancer Research.

Phenotypic Changes and Impaired Function of Peripheral γδ T Cells in Patients With Sepsis.

PubMed

Liao, Xue-Lian; Feng, Ting; Zhang, Jiang-Qian; Cao, Xing; Wu, Qi-Hong; Xie, Zhi-Chao; Kang, Yan; Li, Hong

2017-09-01

Recent studies demonstrated the significant loss of gamma delta T (γδ T) cells in patients with sepsis. Given the distinct functions of γδ T cells in human anti-infection immunity, we are interested in evaluating the phenotype and function of peripheral γδ T cells in septic patients and determining their prognostic implication. This prospective study has been conducted in three intensive care units of a university hospital. During the period from October 2014 to June 2015, we enrolled 107 patients who were consecutively admitted and diagnosed with severe sepsis or septic shock (excluding previous immunosuppression) and 45 healthy controls. Using flow cytometry, we analyzed the in vivo percentage of γδ T cells in cluster of differentiation (CD)3 cells from peripheral blood mononuclear cells as well as their expression of surface markers (CD69, natural-killer group 2 member D [NKG2D], programmed death receptor 1 [PD-1]) and intracellular cytokines (interferon-γ [IFN-γ], interleukin [IL]-17, IL-10, transforming growth factor-β [TGF-β]). Then we further evaluated the different responses of γδ T cells after the antigen stimulation ex vivo by measuring CD69 and IFN-γ expression. Lastly, we conducted the multiple logistic regressions to analyze the risk factor for prognosis. Compared with control group, γδ T cells in septic patients displayed a decrease in percentage, increase in CD69, decrease in NKG2D, and increase in cytokine expression (pro-inflammatory IFN-γ, IL-17, anti-inflammatory IL-10, TGF-β) in vivo. After the antigen stimulation ex vivo, both CD69 and IFN-γ expression in γδ T cells were significantly lower in septic patients than control group. Importantly, the decrease in CD69 and IFN-γ expression was more pronounced in non-survivors than survivors. Multiple logistic regression analysis revealed that lower expression of IFN-γ after stimulation is a dependent risk factor that associated with patient 28-day death in septic patients (OR: 0.908 [95% CI: 0.853-0.966]). Septic patients showed altered phenotype and function of γδ T cells. The impaired IFN-γ expression by γδ T cells after the antigen stimulation is associated with mortality in septic patients.

Protective effect of an aphrodisiac herb Tribulus terrestris Linn on cadmium-induced testicular damage

PubMed Central

Rajendar, B.; Bharavi, K.; Rao, G. S.; Kishore, P.V.S; Kumar, P. Ravi; Kumar, C.S.V Satish; Patel, T. Pankaj

2011-01-01

Aim: The aim of the present study was to investigate whether Tribulus terrestris Linn (TT) could protect the cadmium (Cd)-induced testicular tissue peroxidation in rats and to explore the underlying mechanism of the same. Materials and Methods: In vitro and in vivo studies were conducted to know the protective effect of ethanolic extract of TT (eTT) in Cd toxicity. In in vitro studies, total antioxidant and ferrous metal ion chelating activity of TT was studied. In vivo studies were conducted in rats. A total of 40 Wistar strain adult male rats were divided into four groups. Group 1 served as control, while group 2 to 4 received CdCl2 (3 mg/kg b. wt. s/c once a week). In addition to Cd, group 3 and 4 rats also received eTT (5 mg/kg b.wt. daily as oral gavage) and α-tocopherol (75 mg/kg daily by oral gavage), respectively. At the end of 6th week, all the rats were sacrificed and the separated testes were weighted and processed for estimation of tissue peroxidation markers, antioxidant markers, functional markers, and Cd concentration. The testes were also subjected to histopathological screening. Results: In in vitro studies, the percentage of metal ion chelating activity of 50 μg/ml of eTT and α-tocopherol were 2.76 and 9.39, respectively, and the antioxidant capacity of eTT was equivalent to 0.063 μg of α-tocopherol/μg of eTT. In in vivo studies, administration of Cd significantly reduced the absolute and relative testicular weight, antioxidant markers such as superoxide dismutase and glutathione, and functional markers such as LDH and ALP, along with significant increase in peroxidation markers such as malondialdehyde and protein carbonyls in testicular tissue. Testes of Cd only-treated group showed histological insults like necrotic changes in seminiferous tubules and interstitium, shrunken tubules with desquamated basal lamina, vacuolization and destruction of sertoli cells, and degenerating Leydig cells. This group also had higher Cd levels in testicular tissue. Co-treatment with eTT and α-tocopherol significantly reduced the Cd burden in the testes along with reversal of the Cd-induced changes. Conclusions: eTT exhibited protective effect against Cd-induced testicular damage. The protective effect appears to be mediated through inhibition of testicular tissue peroxidation by antioxidant and metal chelator activity and also, may be indirectly by stimulating the testosterone production from Leydig cells. PMID:22022002

CD24 can be used to isolate Lgr5+ putative colonic epithelial stem cells in mice

PubMed Central

King, Jeffrey B.; von Furstenberg, Richard J.; Smith, Brian J.; McNaughton, Kirk K.; Galanko, Joseph A.

2012-01-01

A growing body of evidence has implicated CD24, a cell-surface protein, as a marker of colorectal cancer stem cells and target for antitumor therapy, although its presence in normal colonic epithelium has not been fully characterized. Previously, our group showed that CD24-based cell sorting can be used to isolate a fraction of murine small intestinal epithelial cells enriched in actively cycling stem cells. Similarly, we hypothesized that CD24-based isolation of colonic epithelial cells would generate a fraction enriched in actively cycling colonic epithelial stem cells (CESCs). Immunohistochemistry performed on mouse colonic tissue showed CD24 expression in the bottom half of proximal colon crypts and the crypt base in the distal colon. This pattern of distribution was similar to enhanced green fluorescent protein (EGFP) expression in Lgr5-EGFP mice. Areas expressing CD24 contained actively proliferating cells as determined by ethynyl deoxyuridine (EdU) incorporation, with a distinct difference between the proximal colon, where EdU-labeled cells were most frequent in the midcrypt, and the distal colon, where they were primarily at the crypt base. Flow cytometric analyses of single epithelial cells, identified by epithelial cell adhesion molecule (EpCAM) positivity, from mouse colon revealed an actively cycling CD24+ fraction that contained the majority of Lgr5-EGFP+ putative CESCs. Transcript analysis by quantitative RT-PCR confirmed enrichment of active CESC markers [leucine-rich-repeat-containing G protein-coupled receptor 5 (Lgr5), ephrin type B receptor 2 (EphB2), and CD166] in the CD24+EpCAM+ fraction but also showed enrichment of quiescent CESC markers [leucine-rich repeats and immunoglobin domains (Lrig), doublecortin and calmodulin kinase-like 1 (DCAMKL-1), and murine telomerase reverse transcriptase (mTert)]. We conclude that CD24-based sorting in wild-type mice isolates a colonic epithelial fraction highly enriched in actively cycling and quiescent putative CESCs. Furthermore, the presence of CD24 expression in normal colonic epithelium may have important implications for the use of anti-CD24-based colorectal cancer therapies. PMID:22723265

The plasma levels of soluble ST2 as a marker of gut mucosal damage in early HIV infection

PubMed Central

Mehraj, Vikram; Jenabian, Mohammad-Ali; Ponte, Rosalie; Lebouché, Bertrand; Costiniuk, Cecilia; Thomas, Réjean; Baril, Jean-Guy; LeBlanc, Roger; Cox, Joseph; Tremblay, Cécile; Routy, Jean-Pierre

2016-01-01

Objective: Following tissue barrier breaches, interleukin-33 (IL-33) is released as an ‘alarmin’ to induce inflammation. Soluble suppression of tumorigenicity 2 (sST2), as an IL-33 decoy receptor, contributes to limit inflammation. We assessed the relationship between the IL-33/ST2 axis and markers of gut mucosal damage in patients with early (EHI) and chronic HIV infection (CHI) and elite controllers. Design: Analyses on patients with EHI and CHI were conducted to determine IL-33/sST2 changes over time. Methods: IL-33 and sST2 levels were measured in plasma. Correlations between sST2 levels and plasma viral load, CD4+ and CD8+ T-cell counts, expression of T-cell activation/exhaustion markers, gut mucosal damage, microbial translocation and inflammation markers, as well as kynurenine/tryptophan ratio were assessed. Results: Plasma sST2 levels were elevated in EHI compared with untreated CHI and uninfected controls, whereas IL-33 levels were comparable in all groups. In EHI, sST2 levels were positively correlated with the CD8+ T-cell count and the percentage of T cells expressing activation and exhaustion markers, but not with viral load or CD4+ T-cell count. Plasma sST2 levels also correlated with plasma levels of gut mucosal damage, microbial translocation and kynurenine/tryptophan ratio and for some markers of inflammation. Prospective analyses showed that early antiretroviral therapy had no impact on sST2 levels, whereas longer treatment duration initiated during CHI normalized sST2. Conclusion: As sST2 levels were elevated in EHI and were correlated with CD8+ T-cell count, immune activation, and microbial translocation, sST2 may serve as a marker of disease progression, gut damage and may directly contribute to HIV pathogenesis. PMID:27045377

Advanced Glycation End Products Enhance Macrophages Polarization into M1 Phenotype through Activating RAGE/NF-κB Pathway

PubMed Central

Jin, Xian; Yao, Tongqing; Zhou, Zhong'e; Zhu, Jian; Zhang, Song; Hu, Wei; Shen, Chengxing

2015-01-01

Atherosclerotic lesions are accelerated in patients with diabetes. M1 (classically activated in contrast to M2 alternatively activated) macrophages play key roles in the progression of atherosclerosis. Since advanced glycation end products (AGEs) are major pathogenic factors and active inflammation inducers in diabetes mellitus, this study assessed the effects of AGEs on macrophage polarization. The present study showed that AGEs significantly promoted macrophages to express IL-6 and TNF-α. M1 macrophage markers such as iNOS and surface markers including CD11c and CD86 were significantly upregulated while M2 macrophage markers such as Arg1 and CD206 remained unchanged after AGEs stimulation. AGEs significantly increased RAGE expression in macrophages and activated NF-κB pathway, and the aforementioned effects were partly abolished by administration of anti-RAGE antibody or NF-κB inhibitor PDTC. In conclusion, our results suggest that AGEs enhance macrophage differentiation into proinflammatory M1 phenotype at least partly via RAGE/NF-κB pathway activation. PMID:26114112

Immunophenotyping and activation status of maternal peripheral blood leukocytes during pregnancy and labour, both term and preterm.

PubMed

Zhang, Jianhong; Shynlova, Oksana; Sabra, Sally; Bang, Annie; Briollais, Laurent; Lye, Stephen J

2017-10-01

The onset of labour in rodents and in humans is associated with physiological inflammation which is manifested by infiltration of activated maternal peripheral leukocytes (mPLs) into uterine tissues. Here, we used flow cytometry to immunophenotype mPLs throughout gestation and labour, both term and preterm. Peripheral blood was collected from non-pregnant women and pregnant women in the 1st, 2nd and 3rd trimesters. Samples were also collected from women in active labour at term (TL) or preterm (PTL) and compared with women term not-in-labour (TNIL) and preterm not-in-labour (PTNIL). Different leukocyte populations were identified by surface markers such as CD45, CD14, CD15, CD3, CD4, CD8, CD19 and CD56. Their activation status was measured by the expression levels of CD11b, CD44, CD55, CD181 and CD192 proteins. Of all circulating CD45+ leukocytes, we detected significant increases in CD15+ granulocytes (i) in pregnant women versus non-pregnant; (ii) in TL women versus TNIL and versus pregnant women in the 1st/2nd/3rd trimester; (iii) in PTL women versus PTNIL. TL was characterized by (iv) increased expressions of CD11b, CD55 and CD192 on granulocytes; (v) increased mean fluorescent intensity (MFI) of CD55 and CD192 on monocytes; (vi) increased CD44 MFI on CD3+ lymphocytes as compared to late gestation. In summary, we have identified sub-populations of mPLs that are specifically activated in association with gestation (granulocytes) or with the onset of labour (granulocytes, monocytes and lymphocytes). Additionally, beta regression analysis created a set of reference values to rank this association between immune markers of pregnancy and to identify activation status with potential prognostic and diagnostic capability. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Sexual Dimorphic Responses in Lymphocytes of Healthy Individuals after Carica papaya Consumption

PubMed Central

Jumat, Nur Ramziahrazanah; Chong, Mun Yee; Seman, Zainina; Jamaluddin, Rosita; Wong, Nyet Kui; Abdullah, Maha

2017-01-01

Sexual dimorphism in immune response is widely recognized, but few human studies have observed this distinction. Food with endo-immunomodulatory potential may reveal novel sex-biased in vivo interactions. Immunomodulatory effects of Carica papaya were compared between healthy male and female individuals. Volunteers were given fixed meals supplemented with papaya for 2 days. Changes in blood immune profiles and hormone levels were determined. In females, total natural killer (NK) cell percentages decreased (12.7 ± 4.4 vs 14.6 ± 5.8%, p = 0.018, n = 18) while B cells increased (15.2 ± 5.5 vs 14.5 ± 5.0, p = 0.037, n = 18) after papaya consumption. Increased 17β-estradiol (511.1 ± 579.7 vs 282.7 ± 165.0 pmol/l, p = 0.036, n = 9) observed in females may be crucial to this change. Differentiation markers (CD45RA, CD69, CD25) analyzed on lymphocytes showed naïve (CD45RA+) non-CD4+ lymphocytes were reduced in females (40.7 ± 8.1 vs 46.8 ± 5.4%, p = 0.012, n = 8) but not males. A general suppressive effect of papaya on CD69+ cells, and higher percentage of CD69+ populations in females and non-CD4 lymphocytes, may be relevant. CD107a+ NK cells were significantly increased in males (16.8 ± 7.0 vs 14.7 ± 4.8, p = 0.038, n = 9) but not females. Effect in females may be disrupted by the action of progesterone, which was significantly correlated with this population (R = 0.771, p = 0.025, n = 8) after papaya consumption. In males, total T helper cells were increased (33.4 ± 6.4 vs 32.4 ± 6.1%, p = 0.040, n = 15). Strong significant negative correlation between testosterone and CD25+CD4+ lymphocytes, may play a role in the lower total CD4+ T cells reported in males. Thus, dissimilar immune profiles were elicited in the sexes after papaya consumption and may have sex hormone influence. PMID:28649252

Membrane receptor-mediated apoptosis and caspase activation in the differentiated EoL-1 eosinophilic cell line.

PubMed

Al-Rabia, Mohammed W; Blaylock, Morgan G; Sexton, Darren W; Walsh, Garry M

2004-06-01

Caspases are key molecules in the control of apoptosis, but relatively little is known about their contribution to eosinophil apoptosis. We examined caspase-3, -8, and -9 activities in receptor ligation-dependent apoptosis induction in the differentiated human eosinophilic cell line EoL-1. Differentiated EoL-1 exhibited bi-lobed nuclei, eosinophil-associated membrane receptors, and basic granule proteins. Annexin-V fluorescein isothiocyanate binding to EoL-1 revealed significant (P<0.01) apoptosis induction in cells cultured for 20 h with monoclonal antibodies (mAb) specific for CD45 (71%+/-4.3), CD45RA (58%+/-2.3), CD45RB (68%+/-2.4), CD95 (47%+/-2.6), and CD69 (52%+/-2.1) compared with control (23%+/-1.6) or CD45RO mAb (27%+/-3.9). The pan-caspase inhibitor Z-Val-Ala-Asp-fluoromethylketone (fmk) and inhibitors of caspase-8 (Z-Ile-Glu-Thr-Asp-fmk) and caspase-9 (Z-Leu-Glu-His-Asp-fmk) significantly inhibited mAb-induced apoptosis of EoL-1 but had no effect on constitutive (baseline) apoptosis at 16 and 20 h. Caspase activity was analyzed using the novel CaspaTag trade mark technique and flow cytometry. EoL-1 treated with pan-CD45, CD45RA, CD45RB, and CD95 mAb exhibited caspase-3 and -9 activation at 12 h post-treatment, which increased at 16 and 20 h. Activated caspase-8 was detected 12 and 16 h after ligation with CD45, CD45RA, CD45RB, and CD95 mAb followed by a trend toward basal levels at 20 h. CD69 ligation resulted in caspase-3 activation, a modest but significant activation of caspase-8, and a loss in mitochondrial transmembrane potential but had no significant effect on activation of caspase-9. Thus, the intrinsic and extrinsic caspase pathways are involved in controlling receptor ligation-mediated apoptosis induction in human eosinophils, findings that may aid the development of a more targeted, anti-inflammatory therapy for asthma.

A polysaccharide isolated from Agaricus blazei Murill (ABP-AW1) as a potential Th1 immunity-stimulating adjuvant

PubMed Central

CUI, LIRAN; SUN, YONGXU; XU, HAO; XU, HUIYU; CONG, HUAN; LIU, JICHENG

2013-01-01

In the present study, a low molecular weight polysaccharide, ABP-AW1, isolated from Agaricus blazei Murill was assessed for its potential adjuvant activity. ABP-AW1 is considered to create a ‘depot’ of antigen at a subcutaneous injection site. ICR mice were immunized with 100 μg ovalbumin (OVA) alone or with 100 μg OVA formulated in 0.9% saline containing 200 μg aluminum (alum) or ABP-AW1 (50, 100 and 200 μg) on days 1 and 15. Two weeks after the secondary immunization, splenocyte proliferation, the expression of surface markers, cytokine production and the OVA-specific antibody levels in the serum were determined. The OVA/ABP-AW1 vaccine, in comparison with OVA alone, markedly increased the proliferation of splenic lymphocytes and elicited greater antigen-specific CD4+ T cell activation, as determined by splenic CD4+CD69+ T cells and Th1 cytokine interferon (IFN)-γ release. The combination of ABP-AW1 and OVA also enhanced IgG2b antibody responses to OVA. In conclusion, these data indicated that ABP-AW1 significantly enhanced the humoral and cellular immune responses against OVA in the mice, suggesting that ABP-AW1 stimulated Th1-type immunity. We suggest that ABP-AW1 may serve as a new adjuvant. PMID:24137460

BRAF and MEK Inhibitors Influence the Function of Reprogrammed T Cells: Consequences for Adoptive T-Cell Therapy.

PubMed

Dörrie, Jan; Babalija, Lek; Hoyer, Stefanie; Gerer, Kerstin F; Schuler, Gerold; Heinzerling, Lucie; Schaft, Niels

2018-01-18

BRAF and MEK inhibitors (BRAFi/MEKi), the standard treatment for patients with BRAF V600 mutated melanoma, are currently explored in combination with various immunotherapies, notably checkpoint inhibitors and adoptive transfer of receptor-transfected T cells. Since two BRAFi/MEKi combinations with similar efficacy are approved, potential differences in their effects on immune cells would enable a rational choice for triple therapies. Therefore, we characterized the influence of the clinically approved BRAFi/MEKi combinations dabrafenib (Dabra) and trametinib (Tram) vs. vemurafenib (Vem) and cobimetinib (Cobi) on the activation and functionality of chimeric antigen receptor (CAR)-transfected T cells. We co-cultured CAR-transfected CD8⁺ T cells and target cells with clinically relevant concentrations of the inhibitors and determined the antigen-induced cytokine secretion. All BRAFi/MEKi reduced this release as single agents, with Dabra having the mildest inhibitory effect, and Dabra + Tram having a clearly milder inhibitory effect than Vem + Cobi. A similar picture was observed for the upregulation of the activation markers CD25 and CD69 on CAR-transfected T cells after antigen-specific stimulation. Most importantly, the cytolytic capacity of the CAR-T cells was significantly inhibited by Cobi and Vem + Cobi, whereas the other kinase inhibitors showed no effect. Therefore, the combination Dabra + Tram would be more suitable for combining with T-cell-based immunotherapy than Vem + Cobi.

A polysaccharide isolated from Agaricus blazei Murill (ABP-AW1) as a potential Th1 immunity-stimulating adjuvant.

PubMed

Cui, Liran; Sun, Yongxu; Xu, Hao; Xu, Huiyu; Cong, Huan; Liu, Jicheng

2013-10-01

In the present study, a low molecular weight polysaccharide, ABP-AW1, isolated from Agaricus blazei Murill was assessed for its potential adjuvant activity. ABP-AW1 is considered to create a 'depot' of antigen at a subcutaneous injection site. ICR mice were immunized with 100 μg ovalbumin (OVA) alone or with 100 μg OVA formulated in 0.9% saline containing 200 μg aluminum (alum) or ABP-AW1 (50, 100 and 200 μg) on days 1 and 15. Two weeks after the secondary immunization, splenocyte proliferation, the expression of surface markers, cytokine production and the OVA-specific antibody levels in the serum were determined. The OVA/ABP-AW1 vaccine, in comparison with OVA alone, markedly increased the proliferation of splenic lymphocytes and elicited greater antigen-specific CD4 + T cell activation, as determined by splenic CD4 + CD69 + T cells and Th1 cytokine interferon (IFN)-γ release. The combination of ABP-AW1 and OVA also enhanced IgG2b antibody responses to OVA. In conclusion, these data indicated that ABP-AW1 significantly enhanced the humoral and cellular immune responses against OVA in the mice, suggesting that ABP-AW1 stimulated Th1-type immunity. We suggest that ABP-AW1 may serve as a new adjuvant.

[Activation of peripheral T lymphocytes in children with epilepsy and production of cytokines].

PubMed

Yang, Jie; Hu, Chongkang; Jiang, Xun

2016-09-01

Objective To study the state of peripheral T lymphocytes and cytokine levels in children with epilepsy. Methods Twenty children with epilepsy and 20 healthy age-matched children were recruited and their peripheral blood was collected. The activation of T lymphocytes was evaluated by detecting the expressions of CD25, CD69 and cytotoxic T lymphocyte-assicated antigen 4 (CTLA4). The function of T lymphocytes was evaluated by detecting the expressions of interferon γ (IFN-γ), tumor necrosis factor α (TNF-α), IL-17A and IL-6. The activation of regulatory T cells (Tregs) was evaluated by detecting the expression of IL-10. Results Children with epilepsy had higher expressions of CD25, CD69 and CTLA-4 in T lymphocytes than the controls did. The expressions of IFN-γ, TNF-α, IL-17A and IL-6 in T lymphocytes of children with epilepsy were higher than those of the controls. Frequency of Tregs producing IL-10 was higher in children with epilepsy as compared with the controls. Conclusion Peripheral T lymphocytes of children with epilepsy are activated and produce cytokines.

Increased endothelial microparticles and oxidative stress at extreme altitude.

PubMed

Pichler Hefti, Jacqueline; Leichtle, Alexander; Stutz, Monika; Hefti, Urs; Geiser, Thomas; Huber, Andreas R; Merz, Tobias M

2016-04-01

Hypoxia and oxidative stress affect endothelial function. Endothelial microparticles (MP) are established measures of endothelial dysfunction and influence vascular reactivity. To evaluate the effects of hypoxia and antioxidant supplementation on endothelial MP profiles, a double-blind, placebo-controlled trial, during a high altitude expedition was performed. 29 participants were randomly assigned to a treatment group (n = 14), receiving vitamin E, C, A, and N-acetylcysteine daily, and a control group (n = 15), receiving placebo. Blood samples were obtained at 490 m (baseline), 3530, 4590, and 6210 m. A sensitive tandem mass spectrometry method was used to measure 8-iso-prostaglandin F2α and hydroxyoctadecadienoic acids as markers of oxidative stress. Assessment of MP profiles including endothelial activation markers (CD62+MP and CD144+MP) and cell apoptosis markers (phosphatidylserine+MP and CD31+MP) was performed using a standardized flow cytometry-based protocol. 15 subjects reached all altitudes and were included in the final analysis. Oxidative stress increased significantly at altitude. No statistically significant changes were observed comparing baseline to altitude measurements of phosphatidylserine expressing MP (p = 0.1718) and CD31+MP (p = 0.1305). Compared to baseline measurements, a significant increase in CD62+MP (p = 0.0079) and of CD144+MP was detected (p = 0.0315) at high altitudes. No significant difference in any MP level or oxidative stress markers were found between the treatment and the control group. Hypobaric hypoxia is associated with increased oxidative stress and induces a significant increase in CD62+ and CD144+MP, whereas phosphatidylserine+MP and CD31+MP remain unchanged. This indicates that endothelial activation rather than an apoptosis is the primary factor of hypoxia induced endothelial dysfunction.

High Cellular Monocyte Activation in People Living With Human Immunodeficiency Virus on Combination Antiretroviral Therapy and Lifestyle-Matched Controls Is Associated With Greater Inflammation in Cerebrospinal Fluid

PubMed Central

Booiman, Thijs; Wit, Ferdinand W.; Maurer, Irma; De Francesco, Davide; Sabin, Caroline A.; Harskamp, Agnes M.; Prins, Maria; Garagnani, Paolo; Pirazzini, Chiara; Franceschi, Claudio; Fuchs, Dietmar; Gisslén, Magnus; Winston, Alan; Reiss, Peter; Reiss, P.; Wit, F. W. N. M.; Schouten, J.; Kooij, K. W.; van Zoest, R. A.; Elsenga, B. C.; Janssen, F. R.; Heidenrijk, M.; Zikkenheiner, W.; van der Valk, M.; Kootstra, N. A.; Booiman, T.; Harskamp-Holwerda, A. M.; Boeser-Nunnink, B.; Maurer, I.; Mangas Ruiz, M. M.; Girigorie, A. F.; Villaudy, J.; Frankin, E.; Pasternak, A.; Berkhout, B.; van der Kuyl, T.; Portegies, P.; Schmand, B. A.; Geurtsen, G. J.; ter Stege, J. A.; Klein Twennaar, M.; Majoie, C. B. L. M.; Caan, M. W. A.; Su, T.; Weijer, K.; Bisschop, P. H. L. T.; Kalsbeek, A.; Wezel, M.; Visser, I.; Ruhé, H. G.; Franceschi, C.; Garagnani, P.; Pirazzini, C.; Capri, M.; Dall’Olio, F.; Chiricolo, M.; Salvioli, S.; Hoeijmakers, J.; Pothof, J.; Prins, M.; Martens, M.; Moll, S.; Berkel, J.; Totté, M.; Kovalev, S.; Gisslén, M.; Fuchs, D.; Zetterberg, H.; Winston, A.; Underwood, J.; McDonald, L.; Stott, M.; Legg, K.; Lovell, A.; Erlwein, O.; Doyle, N.; Kingsley, C.; Sharp, D. J.; Leech, R.; Cole, J. H.; Zaheri, S.; Hillebregt, M. M. J.; Ruijs, Y. M. C.; Benschop, D. P.; Burger, D.; de Graaff-Teulen, M.; Guaraldi, G.; Bürkle, A.; Sindlinger, T.; Moreno-Villanueva, M.; Keller, A.; Sabin, C.; de Francesco, D.; Libert, C.; Dewaele, S.

2017-01-01

Abstract Background. Increased monocyte activation and intestinal damage have been shown to be predictive for the increased morbidity and mortality observed in treated people living with human immunodeficiency virus (PLHIV). Methods. A cross-sectional analysis of cellular and soluble markers of monocyte activation, coagulation, intestinal damage, and inflammation in plasma and cerebrospinal fluid (CSF) of PLHIV with suppressed plasma viremia on combination antiretroviral therapy and age and demographically comparable HIV-negative individuals participating in the Comorbidity in Relation to AIDS (COBRA) cohort and, where appropriate, age-matched blood bank donors (BBD). Results. People living with HIV, HIV-negative individuals, and BBD had comparable percentages of classical, intermediate, and nonclassical monocytes. Expression of CD163, CD32, CD64, HLA-DR, CD38, CD40, CD86, CD91, CD11c, and CX3CR1 on monocytes did not differ between PLHIV and HIV-negative individuals, but it differed significantly from BBD. Principal component analysis revealed that 57.5% of PLHIV and 62.5% of HIV-negative individuals had a high monocyte activation profile compared with 2.9% of BBD. Cellular monocyte activation in the COBRA cohort was strongly associated with soluble markers of monocyte activation and inflammation in the CSF. Conclusions. People living with HIV and HIV-negative COBRA participants had high levels of cellular monocyte activation compared with age-matched BBD. High monocyte activation was predictive for inflammation in the CSF. PMID:28680905

Stromal cell markers are differentially expressed in the synovial tissue of patients with early arthritis.

PubMed

Choi, Ivy Y; Karpus, Olga N; Turner, Jason D; Hardie, Debbie; Marshall, Jennifer L; de Hair, Maria J H; Maijer, Karen I; Tak, Paul P; Raza, Karim; Hamann, Jörg; Buckley, Christopher D; Gerlag, Danielle M; Filer, Andrew

2017-01-01

Previous studies have shown increased expression of stromal markers in synovial tissue (ST) of patients with established rheumatoid arthritis (RA). Here, ST expression of stromal markers in early arthritis in relationship to diagnosis and prognostic outcome was studied. ST from 56 patients included in two different early arthritis cohorts and 7 non-inflammatory controls was analysed using immunofluorescence to detect stromal markers CD55, CD248, fibroblast activation protein (FAP) and podoplanin. Diagnostic classification (gout, psoriatic arthritis, unclassified arthritis (UA), parvovirus associated arthritis, reactive arthritis and RA), disease outcome (resolving vs persistent) and clinical variables were determined at baseline and after follow-up, and related to the expression of stromal markers. We observed expression of all stromal markers in ST of early arthritis patients, independent of diagnosis or prognostic outcome. Synovial expression of FAP was significantly higher in patients developing early RA compared to other diagnostic groups and non-inflammatory controls. In RA FAP protein was expressed in both lining and sublining layers. Podoplanin expression was higher in all early inflammatory arthritis patients than controls, but did not differentiate diagnostic outcomes. Stromal marker expression was not associated with prognostic outcomes of disease persistence or resolution. There was no association with clinical or sonographic variables. Stromal cell markers CD55, CD248, FAP and podoplanin are expressed in ST in the earliest stage of arthritis. Baseline expression of FAP is higher in early synovitis patients who fulfil classification criteria for RA over time. These results suggest that significant fibroblast activation occurs in RA in the early window of disease.

CD4/CD8 Ratio Predicts Yellow Fever Vaccine-Induced Antibody Titers in Virologically Suppressed HIV-Infected Patients.

PubMed

Avelino-Silva, Vivian Iida; Miyaji, Karina Takesaki; Mathias, Augusto; Costa, Dayane Alves; de Carvalho Dias, Juliana Zanatta; Lima, Sheila Barbosa; Simoes, Marisol; Freire, Marcos S; Caiaffa-Filho, Helio H; Hong, Marisa A; Lopes, Marta H; Sartori, Ana M; Kallas, Esper G

2016-02-01

Yellow fever vaccine (YFV) induces weaker immune responses in HIV-infected individuals. However, little is known about YFV responses among antiretroviral-treated patients and potential immunological predictors of YFV response in this population. We enrolled 34 antiretroviral therapy (ART)-treated HIV-infected and 58 HIV-uninfected adults who received a single YFV dose to evaluate antibody levels and predictors of immunity, focusing on CD4(+) T-cell count, CD4(+)/CD8(+) ratio, and Human Pegivirus (GBV-C) viremia. Participants with other immunosuppressive conditions were excluded. Median time since YFV was nonsignificantly shorter in HIV-infected participants than in HIV-uninfected participants (42 and 69 months, respectively, P = 0.16). Mean neutralizing antibody (NAb) titers was lower in HIV-infected participants than HIV-uninfected participants (3.3 vs. 3.6 log10mIU/mL, P = 0.044), a difference that remained significant after adjustment for age, sex, and time since vaccination (P = 0.024). In HIV-infected participants, lower NAb titers were associated with longer time since YFV (rho: -0.38, P = 0.027) and lower CD4(+)/CD8(+) ratio (rho: 0.42, P = 0.014), but not CD4(+) T-cell count (P = 0.52). None of these factors were associated with NAb titers in HIV-uninfected participant. GBV-C viremia was not associated with difference in NAb titers overall or among HIV-infected participants. ART-treated HIV-infected individuals seem to have impaired and/or less durable responses to YFV than HIV-uninfected individuals, which were associated with lower CD4(+)/CD8(+) ratio, but not with CD4(+) T-cell count. These results supports the notion that low CD4(+)/CD8(+) ratio, a marker linked to persistent immune activation, is a better indicator of functional immune disturbance than CD4(+) T-cell count in patients with successful ART.

Differential effect on cell-mediated immunity in human volunteers after intake of different lactobacilli

PubMed Central

Rask, C; Adlerberth, I; Berggren, A; Ahrén, I L; Wold, A E

2013-01-01

Probiotics are live microorganisms which have beneficial effects on the host when ingested in adequate amounts. Probiotic bacteria may stimulate immune effector functions in a strain-specific manner. In this blind placebo-controlled trial, we investigated the effects on the immune system following daily intake of six different strains of lactobacilli or the Gram-negative bacterium Pseudomonas lundensis for 2 or 5 weeks. Blood lymphocyte subsets were quantified by fluorescence activated cell sorter and the expression of activation and memory markers was determined. The bacterial strains were also examined for their capacity to adhere to human intestinal cells and to be phagocytosed by human peripheral blood mononuclear cells. Intake of Lactobacillus plantarum strain 299v increased the expression of the activation marker CD25 (P = 0·01) on CD8+ T cells and the memory cell marker CD45RO on CD4+ T cells (P = 0·03), whereas intake of L. paracasei tended to expand the natural killer T (NK T) cell population (P = 0·06). The phagocytic activity of granulocytes was increased following intake of L. plantarum 299v, L. plantarum HEAL, L. paracasei or L. fermentum. In contrast, ingestion of L. rhamnosus decreased the expression of CD25 and CD45RO significantly within the CD4+ cell population. The observed immune effects after in-vivo administration of the probiotic bacteria could not be predicted by either their adherence capacity or the in-vitro-induced cytokine production. The stimulation of CD8+ T cells and NK T cells suggests that intake of probiotic bacteria may enhance the immune defence against, e.g. viral infections or tumours. PMID:23574328

Differential effect on cell-mediated immunity in human volunteers after intake of different lactobacilli.

PubMed

Rask, C; Adlerberth, I; Berggren, A; Ahrén, I L; Wold, A E

2013-05-01

Probiotics are live microorganisms which have beneficial effects on the host when ingested in adequate amounts. Probiotic bacteria may stimulate immune effector functions in a strain-specific manner. In this blind placebo-controlled trial, we investigated the effects on the immune system following daily intake of six different strains of lactobacilli or the Gram-negative bacterium Pseudomonas lundensis for 2 or 5 weeks. Blood lymphocyte subsets were quantified by fluorescence activated cell sorter and the expression of activation and memory markers was determined. The bacterial strains were also examined for their capacity to adhere to human intestinal cells and to be phagocytosed by human peripheral blood mononuclear cells. Intake of Lactobacillus plantarum strain 299v increased the expression of the activation marker CD25 (P = 0·01) on CD8(+) T cells and the memory cell marker CD45RO on CD4(+) T cells (P = 0·03), whereas intake of L. paracasei tended to expand the natural killer T (NK T) cell population (P = 0·06). The phagocytic activity of granulocytes was increased following intake of L. plantarum 299v, L. plantarum HEAL, L. paracasei or L. fermentum. In contrast, ingestion of L. rhamnosus decreased the expression of CD25 and CD45RO significantly within the CD4(+) cell population. The observed immune effects after in-vivo administration of the probiotic bacteria could not be predicted by either their adherence capacity or the in-vitro-induced cytokine production. The stimulation of CD8(+) T cells and NK T cells suggests that intake of probiotic bacteria may enhance the immune defence against, e.g. viral infections or tumours. © 2012 British Society for Immunology.

Altered Innate and Lymphocytic Immune Responses in Mouse Splenocytes Post-Flight

NASA Technical Reports Server (NTRS)

Hwang, ShenAn; Crucian, Brian E.; Sams, Clarence F.; Actor, Jeffrey K.

2011-01-01

Space flight is known to affect immune responses of astronauts and animals, decreasing lymphocytic responses to mitogenic stimuli, delayed typed hypersensitivity reactions, and T-cell activation. Despite changes in immune suppression, there are no reports of consistent adverse clinical events post flight. To further investigate the spectrum of affected immune responses, murine splenocytes were stimulated immediately post-shuttle flight (14 days on STS-135) with T-cell stimulators or toll-like receptor agonists. Comparisons were made to ground control splenocytes from age-matched mice. Cell phenotypes were assessed, as well as activation markers and associated cytokine production. The CD4+ population decreased with no concurrent decrease in CD8+ cells from shuttle mice post flight compared to ground controls. Regarding antigen presenting cell populations, the number of CD11c+ cells were slightly elevated post flight, compared to ground controls, with increased MHC Class I expression (I-A(sup b)) and no change in Class II expression (H-2K(sup b)). CD86+ populations were also significantly diminished. However, the decreased markers did not correlate with activity. Stimulation of splenocytes post flight showed significant increase in bead uptake, increased Class I expression, increased TNF-alpha and IL-6 production in response to TLR-2 (zymosan) and TLR-4 (LPS) agonists. While most activated (ConA or anti-CD3/anti-CD28) CD4+ cells showed markedly diminished responses (reduced IL-2 production), non-specific T cell responses to superantigen (SEA/SEB) increased post flight as determined by expression of early activation markers. Production of additional cytokines was also dysregulated postflight. Overall, persistent immune changes during space flight could represent unique clinical risks for exploration class missions. The consequences of pathogenic encounter remain an important concern that should be addressed.

Hepatic dendritic cell subsets in the mouse.

PubMed

Jomantaite, Ieva; Dikopoulos, Nektarios; Kröger, Andrea; Leithäuser, Frank; Hauser, Hansjörg; Schirmbeck, Reinhold; Reimann, Jörg

2004-02-01

The CD11c(+) cell population in the non-parenchymal cell population of the mouse liver contains dendritic cells (DC), NK cells, B cells and T cells. In the hepatic CD11c(+) DC population from immunocompetent or immunodeficient [recombinase-activating gene-1 (RAG1)(-/-)] C57BL/6 mice (rigorously depleted of T cells, B cells and NK cells), we identified a B220(+) CD11c(int) subset of 'plasmacytoid' DC, and a B220(-) CD11c(+) DC subset. The latter DC population could be subdivided into a major, immature (CD40(lo) CD80(lo) CD86(lo) MHC class II(lo)) CD11c(int) subset, and a minor, mature (CD40(hi) CD80(hi) CD86(hi) MHC class II(hi)) CD11c(hi) subset. Stimulated B220(+) but not B220(-) DC produced type I interferon. NKT cell activation in vivo increased the number of liver B220(-) DC three- to fourfold within 18 h post-injection, and up-regulated their surface expression of activation marker, while it contracted the B220(+) DC population. Early in virus infection, the hepatic B220(+) DC subset expanded, and both, the B220(+) as well as B220(-) DC populations in the liver matured. In vitro, B220(-) but not B220(+) DC primed CD4(+) or CD8(+)T cells. Expression of distinct marker profiles and functions, and distinct early reaction to activation signals hence identify two distinct B220(+) and B220(-) subsets in CD11c(+) DC populations freshly isolated from the mouse liver.

Decoupling activation and exhaustion of B cells in spontaneous controllers of HIV infection

PubMed Central

Sciaranghella, Gaia; Tong, Neath; Mahan, Alison E.; Suscovich, Todd J.; Alter, Galit

2013-01-01

Objective To define the impact of chronic viremia and associated immune activation on B-cell exhaustion in HIV infection. Design Progressive HIV infection is marked by B-cell anergy and exhaustion coupled with dramatic hypergammaglobulinemia. Although both upregulation of CD95 and loss of CD21 have been used as markers of infection-associated B-cell dysfunction, little is known regarding the specific profiles of dysfunctional B cells and whether persistent viral replication and its associated immune activation play a central role in driving B-cell dysfunction. Methods Multiparameter flow cytometry was used to define the profile of dysfunctional B cells. The changes in the expression of CD21 and CD95 were tracked on B-cell subpopulations in patients with differential control of viral replication. Results Although the emergence of exhausted, CD21low tissue-like memory B cells followed similar patterns in both progressors and controllers, the frequency of CD21low activated memory B cells was lower in spontaneous controllers. Conclusion Our results suggest that the loss of CD21 and the upregulation of CD95 occur as separate events during the development of B-cell dysfunction. The loss of CD21 is a marker of B-cell exhaustion induced in the absence of appreciable viral replication, whereas the upregulation of CD95 is tightly linked to persistent viral replication and its associated immune activation. Thus, these dysfunctional profiles potentially represent two functionally distinct states within the B-cell compartment. PMID:23135171

Donor-specific antibodies require preactivated immune system to harm renal transplant.

PubMed

Süsal, Caner; Döhler, Bernd; Ruhenstroth, Andrea; Morath, Christian; Slavcev, Antonij; Fehr, Thomas; Wagner, Eric; Krüger, Bernd; Rees, Margaret; Balen, Sanja; Živčić-Ćosić, Stela; Norman, Douglas J; Kuypers, Dirk; Emonds, Marie-Paule; Pisarski, Przemyslaw; Bösmüller, Claudia; Weimer, Rolf; Mytilineos, Joannis; Scherer, Sabine; Tran, Thuong H; Gombos, Petra; Schemmer, Peter; Zeier, Martin; Opelz, Gerhard

2016-07-01

It is an unresolved issue why some kidney transplant recipients with pretransplant donor-specific HLA antibodies (DSA) show a high transplant failure rate, whereas in other patients DSA do not harm the graft. We investigated whether help from preactivated T-cells might be necessary for DSA to exert a deleterious effect. The impact of pretransplant DSA and immune activation marker soluble CD30 (sCD30) on 3-year graft survival was analyzed in 385 presensitized kidney transplant recipients. A deleterious influence of pretransplant DSA on graft survival was evident only in patients who were positive for the immune activation marker sCD30. In the absence of sCD30 positivity, 3-year graft survival was virtually identical in patients with or without DSA (83.1±3.9% and 84.3±2.8%, P=0.81). A strikingly lower 3-year graft survival rate of 62.1±6.4% was observed in patients who were both sCD30 and DSA positive (HR 2.92, P<0.001). Even in the presence of strong DSA with ≥5000 MFI, the 3-year graft survival rate was high if the recipients were sCD30 negative. Pretransplant DSA have a significantly deleterious impact on graft survival only in the presence of high pretransplant levels of the activation marker sCD30. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

[Comparative immunophenotypic characterization of human and monkey permanent lymphoid culture cells].

PubMed

Agrba, V Z; Lapin, B A; Medvedeva, N M; Ignatova, I E; Karal-Ogly, D D

2007-01-01

The aim of the study was to define the comparative immunophenotypic characteristics ofwidely spread lymphoid cell cultures, derived from Burkett's lymphoma named as Raji and P3HR-1 in comparison with analogous monkey cultures. It has been shown that P3HR-1 culture consists of similar type cells - activated B-lymphocytes CD23 with k phenotype, which demonstrates its monoclonality. Raji culture includes cells with markers of immature B-lymphocytes CD10 and CD24, as well as elements expressing CD10 antigens. T-cell markers were found in none of the cultures. In contrast to human cells, monkey lymphoid culture expressed both B- and T-cell markers. Moreover, in one of them, obtained from a green monkey, T-cells of suppressor type (CD8) prevailed. The immunophenotypic characteristics of primate lymphoid cell cultures, revealed by the study, are of great importance for their proper application to medical and biological studies.

A distinct plasmablast and naïve B-cell phenotype in primary immune thrombocytopenia

PubMed Central

Flint, Shaun M.; Gibson, Adele; Lucas, Geoff; Nandigam, Raghava; Taylor, Louise; Provan, Drew; Newland, Adrian C.; Savage, Caroline O.; Henderson, Robert B.

2016-01-01

Primary immune thrombocytopenia is an autoimmune disorder in which platelet destruction is a consequence of both B- and T-cell dysregulation. Flow cytometry was used to further characterize the B- and T-cell compartments in a cross-sectional cohort of 26 immune thrombocytopenia patients including antiplatelet antibody positive (n=14) and negative (n=12) patients exposed to a range of therapies, and a cohort of matched healthy volunteers. Markers for B-cell activating factor and its receptors, relevant B-cell activation markers (CD95 and CD21) and markers for CD4+ T-cell subsets, including circulating T-follicular helper-like cells, were included. Our results indicate that an expanded population of CD95+ naïve B cells correlated with disease activity in immune thrombocytopenia patients regardless of treatment status. A population of CD21-naïve B cells was specifically expanded in autoantibody-positive immune thrombocytopenia patients. Furthermore, the B-cell maturation antigen, a receptor for B-cell activating factor, was consistently and strongly up-regulated on plasmablasts from immune thrombocytopenia patients. These observations have parallels in other autoantibody-mediated diseases and suggest that loss of peripheral tolerance in naïve B cells may be an important component of immune thrombocytopenia pathogenesis. Moreover, the B-cell maturation antigen represents a potential target for plasma cell directed therapies in immune thrombocytopenia. PMID:26969086

A distinct plasmablast and naïve B-cell phenotype in primary immune thrombocytopenia.

PubMed

Flint, Shaun M; Gibson, Adele; Lucas, Geoff; Nandigam, Raghava; Taylor, Louise; Provan, Drew; Newland, Adrian C; Savage, Caroline O; Henderson, Robert B

2016-06-01

Primary immune thrombocytopenia is an autoimmune disorder in which platelet destruction is a consequence of both B- and T-cell dysregulation. Flow cytometry was used to further characterize the B- and T-cell compartments in a cross-sectional cohort of 26 immune thrombocytopenia patients including antiplatelet antibody positive (n=14) and negative (n=12) patients exposed to a range of therapies, and a cohort of matched healthy volunteers. Markers for B-cell activating factor and its receptors, relevant B-cell activation markers (CD95 and CD21) and markers for CD4(+) T-cell subsets, including circulating T-follicular helper-like cells, were included. Our results indicate that an expanded population of CD95(+) naïve B cells correlated with disease activity in immune thrombocytopenia patients regardless of treatment status. A population of CD21-naïve B cells was specifically expanded in autoantibody-positive immune thrombocytopenia patients. Furthermore, the B-cell maturation antigen, a receptor for B-cell activating factor, was consistently and strongly up-regulated on plasmablasts from immune thrombocytopenia patients. These observations have parallels in other autoantibody-mediated diseases and suggest that loss of peripheral tolerance in naïve B cells may be an important component of immune thrombocytopenia pathogenesis. Moreover, the B-cell maturation antigen represents a potential target for plasma cell directed therapies in immune thrombocytopenia. Copyright© Ferrata Storti Foundation.

Regulatory role of periodontal ligament fibroblasts for innate immune cell function and differentiation.

PubMed

Konermann, Anna; Stabenow, Dirk; Knolle, Percy A; Held, Stefanie A E; Deschner, James; Jäger, Andreas

2012-10-01

Innate immunity is crucial for an effective host defense against pathogenic microorganisms in periodontal tissues. As periodontal ligament (PDL) cells synthesize immunomodulatory cytokines, the aim of this in vitro study was to investigate whether these cells can interact with innate immune cells. Resting and inflammatory primed (IL-1β, TNF-α, HMGB1) human PDL cells were co-cultured with human monocyte-derived dendritic cells or macrophages. Migration, phenotypic maturation and modulation of phagocytosis of Porphyromonas gingivalis by immune cells were investigated upon co-culture with PDL cells and/or their released soluble factors. PDL cells interacted with immune cells under both non-inflammatory and inflammatory conditions. Immune cell migration was significantly enhanced by co-culture with PDL cells, which also affected their phenotypic maturation both through cell-cell contact and through released soluble mediators. The dendritic cell maturation markers CD83 and CD86 were upregulated as much as both 'alternatively activated' M2 macrophage maturation markers CD23 and CD163. In contrast, the 'classically activated' M1 macrophage maturation marker CD64 was downregulated. Finally, PDL cells significantly enhanced the phagocytosis of Porphyromonas gingivalis by immune cells. Our experiments revealed that PDL cells are not only structural elements of the periodontium, but actively influence immune responses by interaction with innate immune cells.

Characteristics of HIV target CD4 T cells collected using different sampling methods from the genital tract of HIV seronegative women.

PubMed

Iyer, Smita S; Sabula, Michael J; Mehta, C Christina; Haddad, Lisa B; Brown, Nakita L; Amara, Rama R; Ofotokun, Igho; Sheth, Anandi N

2017-01-01

Understanding the immune profile of CD4 T cells, the primary targets for HIV, in the female genital tract (FGT) is critical for evaluating and developing effective biomedical HIV prevention strategies in women. However, longitudinal investigation of HIV susceptibility markers expressed by FGT CD4 T cells has been hindered by low cellular yield and risk of sampling-associated trauma. We investigated three minimally invasive FGT sampling methods to characterize and compare CD4 T cell yield and phenotype with the goal of establishing feasible sampling strategies for immune profiling of mucosal CD4 T cells. FGT samples were collected bimonthly from 12 healthy HIV negative women of reproductive age in the following order: 1) Cervicovaginal lavage (CVL), 2) two sequential endocervical flocked swabs (FS), and 3) two sequential endocervical cytobrushes (CB1, CB2). Cells were isolated and phentoyped via flow cytometry. CD4 T cell recovery was highest from each individual CB compared to either CVL or FS (p < 0.0001). The majority of CD4 T cells within the FGT, regardless of sampling method, expressed CCR5 relative to peripheral blood (p < 0.01). Within the CB, CCR5+ CD4 T cells expressed significantly higher levels of α4β7, CD69, and low levels of CD27 relative to CCR5- CD4 T cells (all p < 0.001). We also identified CD4 Treg lineage cells expressing CCR5 among CB samples. Using three different mucosal sampling methods collected longitudinally we demonstrate that CD4 T cells within the FGT express CCR5 and α4β7 and are highly activated, attributes which could act in concert to facilitate HIV acquisition. FS and CB sampling methods can allow for investigation of strategies to reduce HIV target cells in the FGT and could inform the design and interpretation microbicide and vaccine studies in women.

An innovative cascade system for simultaneous separation of multiple cell types.

PubMed

Pierzchalski, Arkadiusz; Mittag, Anja; Bocsi, Jozsef; Tarnok, Attila

2013-01-01

Isolation of different cell types from one sample by fluorescence activated cell sorting is standard but expensive and time consuming. Magnetic separation is more cost effective and faster by but requires substantial effort. An innovative pluriBead-cascade cell isolation system (pluriSelect GmbH, Leipzig, Germany) simultaneously separates two or more different cell types. It is based on antibody-mediated binding of cells to beads of different size and their isolation with sieves of different mesh-size. For the first time, we validated the pluriSelect system for simultaneous separation of CD4+- and CD8+-cells from human EDTA-blood samples. Results were compared with those obtained by magnetic activated cell sorting (MACS; two steps -first isolation of CD4+, then restaining of the residual cell suspension with anti-human CD8+ MACS antibody followed by the second isolation). pluriSelect separation was done in whole blood, MACS separation on density gradient isolated mononuclear cells. Isolated and residual cells were immunophenotyped by 7-color 9-marker panel (CD3; CD16/56; CD4; CD8; CD14; CD19; CD45; HLA-DR) using flow cytometry. Cell count, purity, yield and viability (7-AAD exclusion) were determined. There were no significant differences between both systems regarding purity (MACS (median[range]: 92.4% [91.5-94.9] vs. pluriSelect 95% [94.9-96.8])) of CD4+ cells, however CD8+ isolation showed lower purity by MACS (74.8% [67.6-77.9], pluriSelect 89.9% [89.0-95.7]). Yield was not significantly different for CD4 (MACS 58.5% [54.1-67.5], pluriSelect 67.9% [56.8-69.8]) and for CD8 (MACS 57.2% [41.3-72.0], pluriSelect 67.2% [60.0-78.5]). Viability was slightly higher with MACS for CD4+ (98.4% [97.8-99.0], pluriSelect 94.1% [92.1-95.2]) and for CD8+-cells (98.8% [98.3-99.1], pluriSelect 86.7% [84.2-89.9]). pluriSelect separation was substantially faster than MACS (1h vs. 2.5h) and no pre-enrichment steps were necessary. In conclusion, pluriSelect is a fast, simple and gentle system for efficient simultaneous separation of two and more cell subpopulation directly from whole blood and provides a simple alternative to magnetic separation.

Increased HIV-1 transcriptional activity and infectious burden in peripheral blood and gut-associated CD4+ T cells expressing CD30

PubMed Central

Carvidi, Alexander B.; Smith, Louis C. B.; Khan, Shahzada; Trapecar, Martin; Stoddart, Cheryl A.; Kuritzkes, Daniel R.

2018-01-01

HIV-1-infected cells persist indefinitely despite the use of combination antiretroviral therapy (ART), and novel therapeutic strategies to target and purge residual infected cells in individuals on ART are urgently needed. Here, we demonstrate that CD4+ T cell-associated HIV-1 RNA is often highly enriched in cells expressing CD30, and that cells expressing this marker considerably contribute to the total pool of transcriptionally active CD4+ lymphocytes in individuals on suppressive ART. Using in situ RNA hybridization studies, we show co-localization of CD30 with HIV-1 transcriptional activity in gut-associated lymphoid tissues. We also demonstrate that ex vivo treatment with brentuximab vedotin, an antibody-drug conjugate (ADC) that targets CD30, significantly reduces the total amount of HIV-1 DNA in peripheral blood mononuclear cells obtained from infected, ART-suppressed individuals. Finally, we observed that an HIV-1-infected individual, who received repeated brentuximab vedotin infusions for lymphoma, had no detectable virus in peripheral blood mononuclear cells. Overall, CD30 may be a marker of residual, transcriptionally active HIV-1 infected cells in the setting of suppressive ART. Given that CD30 is only expressed on a small number of total mononuclear cells, it is a potential therapeutic target of persistent HIV-1 infection. PMID:29470552

Calcium, zinc and vitamin E ameliorate cadmium-induced renal oxidative damage in albino Wistar rats.

PubMed

Adi, Pradeepkiran Jangampalli; Burra, Siva Prasad; Vataparti, Amardev Rajesh; Matcha, Bhaskar

2016-01-01

This study was aimed to examine the protective effects of supplementation with calcium + zinc (Ca + Zn) or vitamin E (Vit-E) on Cd-induced renal oxidative damage. Young albino Wistar rats (180 ± 10 g) (n = 6) control rats, Cd, Cd + Ca + Zn, and Cd + Vit-E experimental groups and the experimental period was 30 days. Rats were exposed to Cd (20 mg/kg body weight) alone treated as Cd treated group and the absence or presence of Ca + Zn (2 mg/kg each) or Vit-E (20 mg/kg body weight) supplementation treated as two separate groups. The activities of the stress marker enzymes superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), glutathione-S-transferase (GST) and lipid peroxidase (LPx) were determined in renal mitochondrial fractions of experimental rats. We observed quantitative changes in SOD isoenzymatic patterns by non-denaturing PAGE analysis, and quantified band densities. These results showed that Cd exposure leads to decreases in SOD, CAT, GR, and GPx activities and a concomitant increase in LPx and GST activities. Ca + Zn and Vit-E administration with Cd significantly reversed Cd-induced perturbations in oxidative stress marker enzymes. However, Vit-E showed more inhibitory activity against Cd than did Ca + Zn, and it protected against Cd-induced nephrotoxicity.

RBP4 activates antigen-presenting cells leading to adipose tissue inflammation and systemic insulin resistance

PubMed Central

Moraes-Vieira, Pedro M.; Yore, Mark M.; Dwyer, Peter M.; Syed, Ismail; Aryal, Pratik; Kahn, Barbara B.

2014-01-01

Insulin resistance is a major cause of diabetes and is highly associated with adipose tissue (AT) inflammation in obesity. RBP4, a retinol-transporter, is elevated in insulin resistance and contributes to increased diabetes risk. We aimed to determine the mechanisms for RBP4-induced insulin resistance. Here we show that RBP4 elevation causes AT inflammation by activating innate immunity which elicits an adaptive immune-response. RBP4-overexpressing mice (RBP4-Ox) are insulin-resistant and glucose-intolerant and have increased AT macrophage and CD4 T-cell infiltration. In RBP4-Ox, AT CD206+ macrophages express pro-inflammatory markers and activate CD4 T-cells while maintaining alternatively-activated macrophage markers. These effects result from direct activation of AT antigen-presenting cells (APCs) by RBP4 through a JNK-dependent pathway. Transfer of RBP4-activated APCs into normal mice is sufficient to induce AT inflammation, insulin resistance and glucose intolerance. Thus, RBP4 causes insulin resistance, at least partly, by activating AT APCs which induce CD4 T-cell Th1 polarization and AT inflammation. PMID:24606904

Impact of monotherapy on HIV-1 reservoir, immune activation, and co-infection with Epstein-Barr virus

PubMed Central

Petrara, Maria Raffaella; Cattelan, Anna Maria; Sasset, Lolita; Freguja, Riccardo; Carmona, Francesco; Sanavia, Silvia; Zanchetta, Marisa; Del Bianco, Paola

2017-01-01

Objectives Although monotherapy (mART) effectiveness in maintaining viral suppression and CD4 cell count has been extensively examined in HIV-1-infected patients, its impact on HIV-1 reservoir, immune activation, microbial translocation and co-infection with Epstein-Barr Virus (EBV) is unclear. Methods This retrospective study involved 32 patients who switched to mART; patients were studied at baseline, 48 and 96 weeks after mART initiation. Thirty-two patients who continued combined antiretroviral therapy (cART) over the same period of time were included in the study. Markers of HIV-1 reservoir (HIV-1 DNA and intracellular HIV-1 RNA) were quantified by real-time PCR. Markers of T-(CD3+CD8+CD38+) and B-(CD19+CD80/86+ and CD19+CD10-CD21lowCD27+) cell activation were evaluated by flow cytometry. Plasma levels of microbial translocation markers were quantified by real-time PCR (16S ribosomal DNA and mitochondrial [mt]DNA) or by ELISA (LPS and sCD14). EBV was typed and quantified by multiplex real-time PCR. Results At baseline, no differences were found between mART and cART groups. Three (10%) mART-treated patients had a virological failure vs none in the cART group. Levels of HIV-1 DNA, intracellular HIV-1 RNA and EBV-DNA remained stable in the mART group, while decreased significantly in the cART group. Percentages of T- and B-activated cells significantly increased in the mART-treated patients, while remained at low levels in the cART-treated ones (p = 0.014 and p<0.001, respectively). Notably, levels of mtDNA remained stable in the cART group, but significantly rose in the mART one (p<0.001). Conclusions Long-term mART is associated with higher levels of T- and B-cell activation and, conversely to cART, does not reduce the size of HIV-1 reservoir and EBV co-infection. PMID:28926641

Ex-vivo characterization of regulatory T cells in pulmonary tuberculosis patients, latently infected persons, and healthy endemic controls.

PubMed

Zewdie, Martha; Howe, Rawleigh; Hoff, Søren T; Doherty, T Mark; Getachew, Nahom; Tarekegne, Azeb; Tessema, Bamlak; Yamuah, Lawrence; Aseffa, Abraham; Abebe, Markos

2016-09-01

Regulatory T cells (Treg) are an essential arm of adaptive immunity not only in tolerance and autoimmunity but also in infectious diseases. In Tuberculosis (TB), it has been suggested that the frequency of Tregs is higher in the blood of TB patients when compared to healthy controls with subsequent decline after treatment. However, with the discovery that FOXP3, the hallmark marker of Tregs, is not exclusive to Tregs and the lack of specific markers for Tregs, it has been a challenge to fully understand the role of Tregs in TB. We isolated PBMC from smear positive TB patients (TB, N = 13) before and after treatment, latent TB infected participants (LTBI, N = 8), and healthy endemic controls (EC, N = 9) and evaluated the frequency of different populations of Tregs and expression of FOXP3 by flowcytometry using six markers. The findings in this study showed that the association of Treg frequency with TB disease depends on the phenotypic markers used. While the frequency of CD4(+)CD25(+/hi) T cells was higher in TB patients compared to LTBI individuals, there was no difference in the frequency of CD4(+)CD25(+)FOXP3(+)CD127(lo) Treg among TB, LTBI, or EC. However, delineation of Tregs into active and naïve subsets revealed a significant increase in FOXP3 expression in active primed Tregs (CD4(+)CD25(+)FOXP3(+)CD127(lo)CD45RO(+)Ki-67(+)) of TB patients compared to LTBI and EC; and a significantly higher frequency of resting primed (CD45RO(+)Ki-67(-)) Treg in QuantiFERON negative EC compared to TB patients. After treatment completion, there was a significant decline in the frequency of active primed Treg, median (IQR) from 12.4% (9.5-21.9) of Tregs to 9.3% (7.0-12.2); P = 0.003 Wilcoxon signed rank test. We conclude that Treg subsets may be differentially regulated and expressed in TB disease, cure, and infection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Ex-vivo characterization of regulatory T cells in pulmonary tuberculosis patients, latently infected persons, and healthy endemic controls

PubMed Central

Zewdie, Martha; Howe, Rawleigh; Hoff, Søren T.; Doherty, T. Mark; Getachew, Nahom; Tarekegne, Azeb; Tessema, Bamlak; Yamuah, Lawrence; Aseffa, Abraham; Abebe, Markos

2016-01-01

SUMMARY Background Regulatory T cells (Treg) are an essential arm of adaptive immunity not only in tolerance and autoimmunity but also in infectious diseases. In Tuberculosis (TB), it has been suggested that the frequency of Tregs is higher in the blood of TB patients when compared to healthy controls with subsequent decline after treatment. However, with the discovery that FOXP3, the hallmark marker of Tregs, is not exclusive to Tregs and the lack of specific markers for Tregs, it has been a challenge to fully understand the role of Tregs in TB. Method We isolated PBMC from smear positive TB patients (TB, N = 13) before and after treatment, latent TB infected participants (LTBI, N = 8), and healthy endemic controls (EC, N = 9) and evaluated the frequency of different populations of Tregs and expression of FOXP3 by flowcytometry using six markers. Results The findings in this study showed that the association of Treg frequency with TB disease depends on the phenotypic markers used. While the frequency of CD4+CD25+/hi T cells was higher in TB patients compared to LTBI individuals, there was no difference in the frequency of CD4+CD25+FOXP3+CD127lo Treg among TB, LTBI, or EC. However, delineation of Tregs into active and naïve subsets revealed a significant increase in FOXP3 expression in active primed Tregs (CD4+CD25+FOXP3+CD127loCD45RO+Ki-67+) of TB patients compared to LTBI and EC; and a significantly higher frequency of resting primed (CD45RO+Ki-67−) Treg in QuantiFERON negative EC compared to TB patients. After treatment completion, there was a significant decline in the frequency of active primed Treg, median (IQR) from 12.4% (9.5–21.9) of Tregs to 9.3% (7.0–12.2); P = 0.003 Wilcoxon signed rank test. We conclude that Treg subsets may be differentially regulated and expressed in TB disease, cure, and infection. PMID:27553411

VISA is Required for B Cell Expression of TLR7

PubMed Central

Xu, Liang-Guo; Jin, Lei; Zhang, Bi-Cheng; Akerlund, Janie L.; Shu, Hong-Bing; Cambier, John C.

2011-01-01

B cells play a critical role in the initialization and development of the Systemic Lupus Erythematosus (SLE) that is dependent on the expression of the endosomal ssRNA receptor TLR7. Previous studies have established that B cell expression of TLR7 is controlled by the Type I IFN secreted by Plasmacytoid Dendritic Cells (PDC). Here we report that VISA, also known as MAVS, IPS-1 and CardIf, essential for RIG-I/MDA5-mediated signaling following sensing of cytosolic RNA, regulate B cell expression of TLR7 and CD23. We found that B cells from VISA−/− mouse express reduced TLR7, but normal basal levels of Type I IFN. We also show that while IFNβ and TLR7 agonists synergize to promote TLR7 expression in VISA−/− B cells, they do not fully complement the defect seen in VISA−/− cells. Cell transfer experiments revealed that the observed effects of VISA−/− are B cell intrinsic. The reduced TLR7 expression in B cells is correlated with impaired TLR7 agonist-induced up-regulation of activation markers CD69 and CD86, cell proliferation, production of IFNα, TNF, IL-12 and NF-κB activation. Finally, studies indicate that genetic background may influence the observed phenotype of our VISA−/− mice, since VISA−/− B cells differ in CD23 and TLR7 expression when on C57BL/6 vs 129Sv-C57BL/6 background. Thus, our findings suggest an unexpected link between VISA-mediated cytosolic RLR signaling and autoimmunity. PMID:22105994

VISA is required for B cell expression of TLR7.

PubMed

Xu, Liang-Guo; Jin, Lei; Zhang, Bi-Cheng; Akerlund, Linda J; Shu, Hong-Bing; Cambier, John C

2012-01-01

B cells play a critical role in the initialization and development of the systemic lupus erythematosus that is dependent on the expression of the endosomal ssRNA receptor TLR7. Previous studies have established that B cell expression of TLR7 is controlled by the type I IFN secreted by plasmacytoid dendritic cells. In this article, we report that VISA, also known as MAVS, IPS-1, and CardIf, essential for RIG-I/MDA5-mediated signaling following sensing of cytosolic RNA, regulate B cell expression of TLR7 and CD23. We found that B cells from a VISA(-/-) mouse express reduced TLR7 but normal basal levels of type I IFN. We also show that although IFN-β and TLR7 agonists synergize to promote TLR7 expression in VISA(-/-) B cells, they do not fully complement the defect seen in VISA(-/-) cells. Cell transfer experiments revealed that the observed effects of VISA(-/-) are B cell intrinsic. The reduced TLR7 expression in B cells is correlated with impaired TLR7 agonist-induced upregulation of activation markers CD69 and CD86, cell proliferation, production of IFN-α, TNF, and IL-12, and NF-κB activation. Finally, studies indicate that genetic background may influence the observed phenotype of our VISA(-/-) mice, because VISA(-/-) B cells differ in CD23 and TLR7 expression when on C57BL/6 versus 129Sv-C57BL/6 background. Thus, our findings suggest an unexpected link between VISA-mediated cytosolic RLR signaling and autoimmunity.

Biological effects of low-dose ionizing radiation exposure on interventional cardiologists.

PubMed

Zakeri, F; Hirobe, T; Akbari Noghabi, K

2010-09-01

Interventional cardiologists (ICs) are likely to receive high radiation exposure as a result of procedures they undertake. To assess the effects of low-dose X-ray radiation exposure on chromosomal damage and on selected indices of cellular and humoral immunity in ICs. The study population consisted of 37 ICs and 37 clinical physicians as the control group with similar age, sex and duration of employment, without any work-related exposure to ionizing radiation. Cytogenetic studies were performed by chromosome aberration analysis and immunological studies by flow cytometry, enzyme-linked immunosorbent assay and immunodiffusion techniques. The frequencies of aberrant cells, chromosome breaks and dicentrics plus centric rings were significantly higher in the exposed group compared to the control group (P < 0.05; P < 0.01; P < 0.001, respectively), without positive correlation between the frequency of dicentric and centric ring aberrations and the cumulative doses of the ICs (r = 0.24, not significant). A significant increase was observed in the expression of activation marker CD69 on TCD4(+) stimulated cells in serum immunoglobulin G and interleukin (IL)-2 (P < 0.05) and a significant decrease in serum IL-10 (P < 0.05) in the ICs compared with that of the control group. There was no statistical difference between the two groups in terms of number of white blood cells and lymphocytes, CD3(+), CD4(+) and CD8(+) T cells, CD19(+) and CD16(+) 56(+) cells and concentrations of interferon (IFN)-gamma, IL-4, IL-6 and IL-8 cytokines. While cytogenetic results show higher chromosomal damage, some immune responses are stimulated or modulated immunologically in ICs.

Downregulation of Melanoma Cell Adhesion Molecule (MCAM/CD146) Accelerates Cellular Senescence in Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells.

PubMed

Jin, Hye Jin; Kwon, Ji Hye; Kim, Miyeon; Bae, Yun Kyung; Choi, Soo Jin; Oh, Wonil; Yang, Yoon Sun; Jeon, Hong Bae

2016-04-01

Therapeutic applications of mesenchymal stem cells (MSCs) for treating various diseases have increased in recent years. To ensure that treatment is effective, an adequate MSC dosage should be determined before these cells are used for therapeutic purposes. To obtain a sufficient number of cells for therapeutic applications, MSCs must be expanded in long-term cell culture, which inevitably triggers cellular senescence. In this study, we investigated the surface markers of human umbilical cord blood-derived MSCs (hUCB-MSCs) associated with cellular senescence using fluorescence-activated cell sorting analysis and 242 cell surface-marker antibodies. Among these surface proteins, we selected the melanoma cell adhesion molecule (MCAM/CD146) for further study with the aim of validating observed expression differences and investigating the associated implications in hUCB-MSCs during cellular senescence. We observed that CD146 expression markedly decreased in hUCB-MSCs following prolonged in vitro expansion. Using preparative sorting, we found that hUCB-MSCs with high CD146 expression displayed high growth rates, multilineage differentiation, expression of stemness markers, and telomerase activity, as well as significantly lower expression of the senescence markers p16, p21, p53, and senescence-associated β-galactosidase, compared with that observed in hUCB-MSCs with low-level CD146 expression. In contrast, CD146 downregulation with small interfering RNAs enhanced the senescence phenotype. In addition, CD146 suppression in hUCB-MSCs caused downregulation of other cellular senescence regulators, including Bmi-1, Id1, and Twist1. Collectively, our results suggest that CD146 regulates cellular senescence; thus, it could be used as a therapeutic marker to identify senescent hUCB-MSCs. One of the fundamental requirements for mesenchymal stem cell (MSC)-based therapies is the expansion of MSCs during long-term culture because a sufficient number of functional cells is required. However, long-term growth inevitably induces cellular senescence, which potentially causes poor clinical outcomes by inducing growth arrest and the loss of stem cell properties. Thus, the identification of markers for evaluating the status of MSC senescence during long-term culture may enhance the success of MSC-based therapy. This study provides strong evidence that CD146 is a novel and useful marker for predicting senescence in human umbilical cord blood-derived MSCs (hUCB-MSCs), and CD146 can potentially be applied in quality-control assessments of hUCB-MSC-based therapy. ©AlphaMed Press.

The PD-1: PD-L1 pathway promotes development of brain-resident memory T cells following acute viral encephalitis.

PubMed

Prasad, Sujata; Hu, Shuxian; Sheng, Wen S; Chauhan, Priyanka; Singh, Amar; Lokensgard, James R

2017-04-13

Previous work from our laboratory has demonstrated that during acute viral brain infection, glial cells modulate antiviral T cell effector responses through the PD-1: PD-L1 pathway, thereby limiting the deleterious consequences of unrestrained neuroinflammation. Here, we evaluated the PD-1: PD-L1 pathway in development of brain-resident memory T cells (bT RM ) following murine cytomegalovirus (MCMV) infection. Flow cytometric analysis of immune cells was performed at 7, 14, and 30 days post-infection (dpi) to assess the shift of brain-infiltrating CD8 + T cell populations from short-lived effector cells (SLEC) to memory precursor effector cells (MPEC), as well as generation of bT RMs . In wild-type (WT) animals, we observed a switch in the phenotype of brain-infiltrating CD8 + T cell populations from KLRG1 + CD127 - (SLEC) to KLRG1 - CD127 + (MPEC) during transition from acute through chronic phases of infection. At 14 and 30 dpi, the majority of CD8 + T cells expressed CD127, a marker of memory cells. In contrast, fewer CD8 + T cells expressed CD127 within brains of infected, PD-L1 knockout (KO) animals. Notably, in WT mice, a large population of CD8 + T cells was phenotyped as CD103 + CD69 + , markers of bT RM , and differences were observed in the numbers of these cells when compared to PD-L1 KOs. Immunohistochemical studies revealed that brain-resident CD103 + bT RM cells were localized to the parenchyma. Higher frequencies of CXCR3 were also observed among WT animals in contrast to PD-L1 KOs. Taken together, our results indicate that bT RMs are present within the CNS following viral infection and the PD-1: PD-L1 pathway plays a role in the generation of this brain-resident population.

Influence of molecular weight on in vitro immunostimulatory properties of instant coffee.

PubMed

Passos, Cláudia P; Cepeda, Márcio R; Ferreira, Sónia S; Nunes, Fernando M; Evtuguin, Dmitry V; Madureira, Pedro; Vilanova, Manuel; Coimbra, Manuel A

2014-10-15

Instant coffee was prepared and fractionated into higher (>100kDa), medium (5-10, 10-30, 30-100kDa) and lower (1-5, <1kDa) molecular weight fractions. Sugars and linkage composition characteristics of arabinogalactans and galactomannans were recovered in all fractions. Also, amino acid analysis performed after hydrolysis showed similar compositions in all fractions. On the contrary, free chlorogenic acids and caffeine were only detected in the lowest molecular weight fraction (<1kDa). A direct relationship between the melanoidins browning index and the molecular weight was observed. The fractions obtained were incubated in vitro with murine spleen lymphocytes in order to evaluate their possible immunostimulatory abilities. The surface expression of CD69 (early activation marker) on different lymphocyte sub-populations showed that the fraction with 1-5kDa was able to induce activation of B-lymphocytes. This was the only fraction to induce B-lymphocyte activation, since all the other fractions failed, even when higher concentrations were used. Copyright © 2014 Elsevier Ltd. All rights reserved.

High Cellular Monocyte Activation in People Living With Human Immunodeficiency Virus on Combination Antiretroviral Therapy and Lifestyle-Matched Controls Is Associated With Greater Inflammation in Cerebrospinal Fluid.

PubMed

Booiman, Thijs; Wit, Ferdinand W; Maurer, Irma; De Francesco, Davide; Sabin, Caroline A; Harskamp, Agnes M; Prins, Maria; Garagnani, Paolo; Pirazzini, Chiara; Franceschi, Claudio; Fuchs, Dietmar; Gisslén, Magnus; Winston, Alan; Reiss, Peter; Kootstra, Neeltje A

2017-01-01

Increased monocyte activation and intestinal damage have been shown to be predictive for the increased morbidity and mortality observed in treated people living with human immunodeficiency virus (PLHIV). A cross-sectional analysis of cellular and soluble markers of monocyte activation, coagulation, intestinal damage, and inflammation in plasma and cerebrospinal fluid (CSF) of PLHIV with suppressed plasma viremia on combination antiretroviral therapy and age and demographically comparable HIV-negative individuals participating in the Comorbidity in Relation to AIDS (COBRA) cohort and, where appropriate, age-matched blood bank donors (BBD). People living with HIV, HIV-negative individuals, and BBD had comparable percentages of classical, intermediate, and nonclassical monocytes. Expression of CD163, CD32, CD64, HLA-DR, CD38, CD40, CD86, CD91, CD11c, and CX3CR1 on monocytes did not differ between PLHIV and HIV-negative individuals, but it differed significantly from BBD. Principal component analysis revealed that 57.5% of PLHIV and 62.5% of HIV-negative individuals had a high monocyte activation profile compared with 2.9% of BBD. Cellular monocyte activation in the COBRA cohort was strongly associated with soluble markers of monocyte activation and inflammation in the CSF. People living with HIV and HIV-negative COBRA participants had high levels of cellular monocyte activation compared with age-matched BBD. High monocyte activation was predictive for inflammation in the CSF. © The Author 2017. Published by Oxford University Press on behalf of Infectious Diseases Society of America.

Apigenin inhibited hypoxia induced stem cell marker expression in a head and neck squamous cell carcinoma cell line.

PubMed

Ketkaew, Yuwaporn; Osathanon, Thanaphum; Pavasant, Prasit; Sooampon, Sireerat

2017-02-01

Cancer stem cells contribute to tumor recurrence, and a hypoxic environment is critical for maintaining cancer stem cells. Apigenin is a natural product with anticancer activity. However, the effect of apigenin on cancer stem cells remains unclear. Our aim was to investigate the effect of apigenin on cancer stem cell marker expression in head and neck squamous cell carcinoma cells under hypoxia. We used three head and neck squamous cell carcinoma cell lines; HN-8, HN-30, and HSC-3. The mRNA expression of cancer stem cell markers was determined by semiquantitative RT-PCR and Real-time PCR. The cytotoxic effect of apigenin was determined by MTT colorimetric assay. Flow cytometry was used to reveal the number of cells expressing cancer stem cell surface markers. HN-30 cells, a cancer cell line from the pharynx, showed the greatest response to hypoxia by increasing their expression of CD44, CD105, NANOG, OCT-4, REX-1, and VEGF. Apigenin significantly decreased HN-30 cell viability in dose- and time-dependent manners. In addition, 40μM apigenin significantly down-regulated the mRNA expression of CD44, NANOG, and CD105. Consistent with these results, the hypoxia-induced increase in CD44 + cells, CD105 + cells, and STRO-1 + cells was significantly abolished by apigenin. Apigenin suppresses cancer stem cell marker expression and the number of cells expressing cell surface markers under hypoxia. Copyright © 2016 Elsevier Ltd. All rights reserved.

The role of CD133 in normal human prostate stem cells and malignant cancer-initiating cells.

PubMed

Vander Griend, Donald J; Karthaus, Wouter L; Dalrymple, Susan; Meeker, Alan; DeMarzo, Angelo M; Isaacs, John T

2008-12-01

Resolving the specific cell of origin for prostate cancer is critical to define rational targets for therapeutic intervention and requires the isolation and characterization of both normal human prostate stem cells and prostate cancer-initiating cells (CIC). Single epithelial cells from fresh normal human prostate tissue and prostate epithelial cell (PrEC) cultures derived from them were evaluated for the presence of subpopulations expressing stem cell markers and exhibiting stem-like growth characteristics. When epithelial cell suspensions containing cells expressing the stem cell marker CD133+ are inoculated in vivo, regeneration of stratified human prostate glands requires inductive prostate stromal cells. PrEC cultures contain a small subpopulation of CD133+ cells, and fluorescence-activated cell sorting-purified CD133+ PrECs self-renew and regenerate cell populations expressing markers of transit-amplifying cells (DeltaNp63), intermediate cells (prostate stem cell antigen), and neuroendocrine cells (CD56). Using a series of CD133 monoclonal antibodies, attachment and growth of CD133+ PrECs requires surface expression of full-length glycosylated CD133 protein. Within a series of androgen receptor-positive (AR+) human prostate cancer cell lines, CD133+ cells are present at a low frequency, self-renew, express AR, generate phenotypically heterogeneous progeny negative for CD133, and possess an unlimited proliferative capacity, consistent with CD133+ cells being CICs. Unlike normal adult prostate stem cells, prostate CICs are AR+ and do not require functional CD133. This suggests that (a) AR-expressing prostate CICs are derived from a malignantly transformed intermediate cell that acquires "stem-like activity" and not from a malignantly transformed normal stem cell and (b) AR signaling pathways are a therapeutic target for prostate CICs.

Treatment of mice with fenbendazole attenuates allergic airways inflammation and Th2 cytokine production in a model of asthma.

PubMed

Cai, Yeping; Zhou, Jiansheng; Webb, Dianne C

2009-01-01

Mouse models have provided a significant insight into the role of T-helper (Th) 2 cytokines such as IL-5 and IL-13 in regulating eosinophilia and other key features of asthma. However, the validity of these models can be compromised by inadvertent infection of experimental mouse colonies with pathogens such as oxyurid parasites (pinworms). While the benzimidazole derivative, fenbendazole (FBZ), is commonly used to treat such outbreaks, the effects of FBZ on mouse models of Th2 disease are largely unknown. In this investigation, we show that mice fed FBZ-supplemented food during the in utero and post-weaning period developed attenuated lung eosinophilia, antigen-specific IgG1 and Th2 cytokine responses in a model of asthma. Treatment of the mediastinal lymph node cells from allergic mice with FBZ in vitro attenuated cell proliferation, IL-5 and IL-13 production and expression of the early lymphocyte activation marker, CD69 on CD4(+) T cells and CD19(+) B cells. In addition, eosinophilia and Th2 responses remained attenuated after a 4-week withholding period in allergic mice treated preweaning with FBZ. Thus, FBZ modulates the amplitude of Th2 responses both in vivo and in vitro.

Reconstitution of Intestinal CD4 and Th17 T Cells in Antiretroviral Therapy Suppressed HIV-Infected Subjects: Implication for Residual Immune Activation from the Results of a Clinical Trial

PubMed Central

d'Ettorre, Gabriella; Baroncelli, Silvia; Micci, Luca; Ceccarelli, Giancarlo; Andreotti, Mauro; Sharma, Prachi; Fanello, Gianfranco; Fiocca, Fausto; Cavallari, Eugenio Nelson; Giustini, Noemi; Mallano, Alessandra; Galluzzo, Clementina M.; Vella, Stefano; Mastroianni, Claudio M.; Silvestri, Guido; Paiardini, Mirko; Vullo, Vincenzo

2014-01-01

Introduction During HIV infection the severe depletion of intestinal CD4+ T-cells is associated with microbial translocation, systemic immune activation, and disease progression. This study examined intestinal and peripheral CD4+ T-cell subsets reconstitution under combined antiretroviral therapy (cART), and systemic immune activation markers. Methods This longitudinal single-arm pilot study evaluates CD4+ T cells, including Th1 and Th17, in gut and blood and soluble markers for inflammation in HIV-infected individuals before (M0) and after eight (M8) months of cART. From January 2010 to December 2011, 10 HIV-1 naïve patients were screened and 9 enrolled. Blood and gut CD4+ T-cells subsets and cellular immune activation were determined by flow-cytometry and plasma soluble CD14 by ELISA. CD4+ Th17 cells were detected in gut biopsies by immunohistochemistry. Microbial translocation was measured by limulus-amebocyte-lysate assay to detect bacterial lipopolysaccharide (LPS) and PCR Real Time to detect plasma bacterial 16S rDNA. Results Eight months of cART increased intestinal CD4+ and Th17 cells and reduced levels of T-cell activation and proliferation. The magnitude of intestinal CD4+ T-cell reconstitution correlated with the reduction of plasma LPS. Importantly, the magnitude of Th17 cells reconstitution correlated directly with blood CD4+ T-cell recovery. Conclusion Short-term antiretroviral therapy resulted in a significant increase in the levels of total and Th17 CD4+ T-cells in the gut mucosa and in decline of T-cell activation. The observation that pre-treatment levels of CD4+ and of CD8+ T-cell activation are predictors of the magnitude of Th17 cell reconstitution following cART provides further rationale for an early initiation of cART in HIV-infected individuals. Trial Registration ClinicalTrials.gov NCT02097381 PMID:25340778

Analysis of immune activation and clinical events in acute infectious mononucleosis.

PubMed

Williams, Hilary; Macsween, Karen; McAulay, Karen; Higgins, Craig; Harrison, Nadine; Swerdlow, Anthony; Britton, Kate; Crawford, Dorothy

2004-07-01

The symptoms of infectious mononucleosis (IM) are thought to be caused by T cell activation and cytokine production. Surface lymphocyte activation marker (SLAM)-associated protein (SAP) regulates lymphocyte activation via signals from cell-surface CD244 (2B4) and SLAM (CD150). We followed T cell activation via this SAP/SLAM/CD244 pathway in IM and analyzed whether the results were associated with clinical severity. At diagnosis, SAP, SLAM, and CD244 were significantly up-regulated on CD4 and CD8 T cells; expression decreased during IM, but CD244 and SLAM levels remained higher on CD8 cells 40 days later. There were significantly more lymphocytes expressing CD8 and CD244/CD8 in patients with severe sore throat. The expression of CD8 alone and CD244 on CD8 cells correlated with increased virus load. We suggest that T cells expressing CD244 and SLAM are responsible for the clinical features of IM but that the control of activation is maintained by parallel increased expression of SAP.

[Association between expression of lectin type receptors by natural killers and intensity of liver fibrosis during chronic hepatitis C].

PubMed

Malova, E S; Balmasova, I P; Iuschuk, N D; Shmeleva, E V; Eremina, O F

2010-01-01

To study functional activity of natural killers on different stages of fibrosis during chronic hepatitis C. Functional activity of CD3-/CD56+/CD16+ lymphocytes measured as expression of natural killers receptors (NKR) and natural cytotoxicity receptors (NCR) was assessed by flow cytometry. At stage I of fibrosis, decrease of number of CD3-/CD56+/NKG2D+ cells was observed, whereas at precirrhotic stage III--sharp decrease of CD3-/CD56+/CD94+ and CD3-/ CD56+/NKG2D+ populations, and at cirrhotic stage--decrease of number of CD3-/CD56+/ NKG2D+ cells and increase of cytolytic activity of natural killers carrying CD107a marker compared to precirrhotic stage. Obtained data demonstrate that natural killers during chronic hepatitis C receive regulatory signals mainly through lectin type receptors (CD94 and NKG2D).

Markers of activated inflammatory cells correlate with severity of liver damage in children with nonalcoholic fatty liver disease.

PubMed

De Vito, Rita; Alisi, Anna; Masotti, Andrea; Ceccarelli, Sara; Panera, Nadia; Citti, Arianna; Salata, Michele; Valenti, Luca; Feldstein, Ariel E; Nobili, Valerio

2012-07-01

Concomitantly to the obesity epidemic, nonalcoholic fatty liver disease (NAFLD) has become the leading cause of liver disease in children. NAFLD encompasses a spectrum of histological damage ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), with possible progression to cirrhosis. There is growing evidence that the immune system plays a pivotal role in the initiation and progression to NASH but the cellular nature of the hepatic inflammation is still unknown. The present study includes 34 children with biopsy-proven NAFLD. Liver damage was evaluated by the NAFLD activity score (NAS), and the inflammatory infiltrate was characterized by immunohistochemistry for CD45, CD3 and CD163 which are markers of leukocytes, T cells and activated Kupffer cells/macrophages, respectively. Our results have shown that CD45+ (P<0.0001) and CD163+ (P<0.0001) cells were markedly increased in children with severe histological activity (NAS≥5) compared to children with lower activity (NAS<5), whereas CD3+ cells were significantly lower (P<0.01) in children with severe histological activity. There was a significant association between the numbers of CD45+, CD3+ and CD163+ cells, regarding both the portal tract and liver lobule, and the severity of steatosis, ballooning and fibrosis (P<0.01). These data suggest that the severity and composition of the inflammatory infiltrate correlate with steatosis and the severity of disease in children with NAFLD. Moreover, a decrease in CD3+ cells may be involved in the pathogenesis of liver damage. Future studies should evaluate whether it can predict the progression of liver disease independently of established histological scores.

Duodenal intraepithelial T lymphocytes in patients with functional dyspepsia

PubMed Central

Gargala, Gilles; Lecleire, Stéphane; François, Arnaud; Jacquot, Serge; Déchelotte, Pierre; Ballet, Jean Jacques; Favennec, Loic; Ducrotté, Philippe

2007-01-01

AIM: To quantify the intraepithelial lymphocytes (IELs) and to document the membrane expression of CD4, CD8, TCRγδ and adhesion and/or activation-associated molecules (CD103, CD28, CD44, CD69, HLA-DR, CD95/Fas) in the duodenal mucosa of patients with functional dyspepsia (FD) in order to provide arguments for an immunological process in FD. METHODS: Twenty-six FD patients according to Rome II criteria (20 were H pylori negative) were studied and compared to 12 healthy adults. IELs were isolated from five duodenal biopsy samples, then quantified by microscopy and flow cytometry while the membrane phenotypes were determined by cytofluorometry. RESULTS: Duodenal histological examination was normal. In H pylori negative patients, the number of IELs was not different from that in healthy controls. Median percentage expression of CD4, CD8, or TCRγδ and CD103, CD44, CD28, CD69 on CD3+ IELs, among the adhesion/activation associated molecules tested, was not different from that in healthy controls. In contrast, the median percentage expression of CD95/Fas [22 (9-65) vs 45 (19-88), P = 0.03] and HLA-DR expressing CD3+ IELs [4 (0-30) vs 13 (4-42), P = 0.04] was significantly lower in the H pylori negative FD group than in healthy controls, respectively. The number of IELs was significantly greater in H pylori positive FD patients than in healthy controls [median ratiofor 100 enterocytes 27.5 (6.7-62.5) vs 10.8 (3-33.3), P = 0.02] due to a higher number of CD8+ CD3+ IELs. CONCLUSION: In H pylori negative FD patients, the phenotypic characterization of IELs suggests that we cannot exclude a role of IELs in FD. PMID:17511033

Functional Analysis of CD28/B7 and CD40/CD40L Costimulation During the in vivo Type 2 Immune Response

DTIC Science & Technology

1995-10-06

these activation markers on B cells and changes in B cell size (forward light scatter) were analyzed by flow cytometry (Figure 7). B cell surface B7...activation ofnaive CD4+ Th cells requires two signals delivered from antigen presenting cells (APes). The engagement ofthe T cell surface receptor...shown that T cell surface ii molecule CD28, and its homologue CTLA-4, can provide costimulatory signals to 10 cells when they interact with their ligands

In the absence of its cytosolic domain, the CD28 molecule still contributes to T cell activation

PubMed Central

Morin, Stéphanie; Giroux, Valentin; Favre, Cédric; Bechah, Yassina; Auphan-Anezin, Nathalie; Roncagalli, Romain; Mège, Jean-Louis; Olive, Daniel; Malissen, Marie; Nunes, Jacques

2015-01-01

The CD28 costimulatory receptor has a pivotal role in T cell biology as this molecule amplifies T cell receptor (TCR) signals to provide an efficient immune T cell response. There is a large debate about how CD28 mediates these signals. Here, we designed a CD28 gene targeted knock-in mouse strain lacking the cytoplasmic tail of CD28. As is the case in CD28-deficient (CD28 knock-out) mice, regulatory T cell homeostasis and T cell activation are altered in these CD28 knock-in mice. Unexpectedly, the presence of a CD28 molecule deprived of its cytoplasmic tail could partially induce some early activation events in T cells such as signaling events or expression of early activation markers. These results unravel a new mechanism of T cell costimulation by CD28, independent of its cytoplasmic tail. PMID:25725801

Spontaneous apoptosis, oxidative status and immunophenotype markers in blood lymphocytes of AIDS patients.

PubMed

Losa, G A; Graber, R

2000-01-01

Peripheral blood mononuclear cells (PBMC) from 251 HIV-positive drug abusers of known clinical stage and from 40 healthy donors were tested for conventional immunologic markers (CD3, CD4, CD8, CD19, CD14, CD16/CD56, CD45 and HLA-DR). Additional cell parameters and the occurrence of spontaneous apoptosis (programmed cell death) were investigated on freshly isolated PBMC by flow cytometric measurement of either annexin-V bound to plasma membrane phosphatidylserine or propidium iodide uptake. The activity of gamma-glutamyltransferase (gamma-GT), an ectoenzyme contributing to the synthesis of the intracellular antioxidant glutathione (GSH) and involved in early apoptosis, was also determined in these cells. Immunocompetent T-cell counts were lower in HIV+ patients, with the exception of CD8+ and HLA-DR+ lymphocytes. The external binding of annexin-V was significantly higher in HIV+ PBMC and occurred in both CD8+ and CD4+ T-lymphocyte subsets. The activity of gamma-GT, was significantly lower in the PBMC from HIV+ patients, indicating that the redox status of PBMC may be affected in HIV+ individuals. Finally, the most dominant features characterising patients receiving antiretroviral therapy were greater long-term stability in the distribution of various cell parameters excepted the level of apoptosis.

Spontaneous Apoptosis, Oxidative Status and Immunophenotype Markers in Blood Lymphocytes of AIDS Patients

PubMed Central

Losa, Gabriele A.; Graber, Riccardo

2000-01-01

Peripheral blood mononuclear cells (PBMC) from 251 HIV‐positive drug abusers of known clinical stage and from 40 healthy donors were tested for conventional immunologic markers (CD3, CD4, CD8, CD19, CD14, CD16/CD56, CD45 and HLA‐DR). Additional cell parameters and the occurrence of spontaneous apoptosis (programmed cell death) were investigated on freshly isolated PBMC by flow cytometric measurement of either annexin‐V bound to plasma membrane phosphatidylserine or propidium iodide uptake. The activity of γ‐glutamyltransferase (γ‐GT), an ectoenzyme contributing to the synthesis of the intracellular antioxidant glutathione (GSH) and involved in early apoptosis, was also determined in these cells. Immunocompetent T‐cell counts were lower in HIV+ patients, with the exception of CD8+ and HLA‐DR+ lymphocytes. The external binding of annexin‐V was significantly higher in HIV+ PBMC and occurred in both CD8+ and CD4+ T‐lymphocyte subsets. The activity of γ‐GT, was significantly lower in the PBMC from HIV+ patients, indicating that the redox status of PBMC may be affected in HIV+ individuals. Finally, the most dominant features characterising patients receiving antiretroviral therapy were greater long‐term stability in the distribution of various cell parameters excepted the level of apoptosis. PMID:11254221

Genetic Variation Affects C-Reactive Protein Elevations in Crohn's Disease.

PubMed

Moran, Christopher J; Kaplan, Jess L; Winter, Harland S

2018-04-28

C-reactive protein (CRP) is a serum marker that is used to measure disease activity in Crohn's disease (CD). However, a subset of CD patients have normal CRP during flares. In rheumatoid arthritis and lupus, genetic variants can restrict CRP elevations during flares. This study sought to determine if common CRP genetic variants affect CRP values during active CD. Subjects with CD who participated in the Partners HealthCare BioBank were genotyped for 5 common CRP genetic variants (rs2794520, rs3122012, rs3093077, rs2808635, and rs1800947). Medical records were reviewed to determine disease activity and the highest CRP value during active CD. CRP values during active infection or malignancy at the time of the test were excluded. CRP values were compared by genotype using the Mann-Whitney test. The study included 199 subjects with active CD (21 to 86 years of age). Subjects with the rs2794520 TT genotype had a lower CRP than subjects with the CC genotype (58.3 mg/L vs 28.4 mg/L, P = 0.008). Subjects with the rs1800947 CG genotype had a lower CRP than those with the CC genotype (54.3 mg/L vs 22.4 mg/L, P < 0.0001); 41.6% of TT subjects had a normal CRP compared with 24.1% of CT subjects and 16.5% of CC subjects (P = 0.041). This study demonstrates that rs2794520 and rs1800947 are associated with a restriction of CRP elevations during active CD. While CRP is typically a reliable biomarker in CD, there is a subset of CD patients with a genetically determined restriction of CRP in whom other disease markers should be utilized.

Adoptive cell therapy for lymphoma with CD4 T cells depleted of CD137-expressing regulatory T cells.

PubMed

Goldstein, Matthew J; Kohrt, Holbrook E; Houot, Roch; Varghese, Bindu; Lin, Jack T; Swanson, Erica; Levy, Ronald

2012-03-01

Adoptive immunotherapy with antitumor T cells is a promising novel approach for the treatment of cancer. However, T-cell therapy may be limited by the cotransfer of regulatory T cells (T(reg)). Here, we explored this hypothesis by using 2 cell surface markers, CD44 and CD137, to isolate antitumor CD4 T cells while excluding T(regs). In a murine model of B-cell lymphoma, only CD137(neg)CD44(hi) CD4 T cells infiltrated tumor sites and provided protection. Conversely, the population of CD137(pos)CD44hi CD4 T cells consisted primarily of activated T(regs). Notably, this CD137(pos) T(reg) population persisted following adoptive transfer and maintained expression of FoxP3 as well as CD137. Moreover, in vitro these CD137(pos) cells suppressed the proliferation of effector cells in a contact-dependent manner, and in vivo adding the CD137(pos)CD44(hi) CD4 cells to CD137(neg)CD44(hi) CD4 cells suppressed the antitumor immune response. Thus, CD137 expression on CD4 T cells defined a population of activated T(regs) that greatly limited antitumor immune responses. Consistent with observations in the murine model, human lymphoma biopsies also contained a population of CD137(pos) CD4 T cells that were predominantly CD25(pos)FoxP3(pos) T(regs). In conclusion, our findings identify 2 surface markers that can be used to facilitate the enrichment of antitumor CD4 T cells while depleting an inhibitory T(reg) population.

Sexual Dimorphic Responses in Lymphocytes of Healthy Individuals after Carica papaya Consumption.

PubMed

Jumat, Nur Ramziahrazanah; Chong, Mun Yee; Seman, Zainina; Jamaluddin, Rosita; Wong, Nyet Kui; Abdullah, Maha

2017-01-01

Sexual dimorphism in immune response is widely recognized, but few human studies have observed this distinction. Food with endo-immunomodulatory potential may reveal novel sex-biased in vivo interactions. Immunomodulatory effects of Carica papaya were compared between healthy male and female individuals. Volunteers were given fixed meals supplemented with papaya for 2 days. Changes in blood immune profiles and hormone levels were determined. In females, total natural killer (NK) cell percentages decreased (12.7 ± 4.4 vs 14.6 ± 5.8%, p  = 0.018, n  = 18) while B cells increased (15.2 ± 5.5 vs 14.5 ± 5.0, p  = 0.037, n  = 18) after papaya consumption. Increased 17β-estradiol (511.1 ± 579.7 vs 282.7 ± 165.0 pmol/l, p  = 0.036, n  = 9) observed in females may be crucial to this change. Differentiation markers (CD45RA, CD69, CD25) analyzed on lymphocytes showed naïve (CD45RA + ) non-CD4 + lymphocytes were reduced in females (40.7 ± 8.1 vs 46.8 ± 5.4%, p  = 0.012, n  = 8) but not males. A general suppressive effect of papaya on CD69 + cells, and higher percentage of CD69 + populations in females and non-CD4 lymphocytes, may be relevant. CD107a + NK cells were significantly increased in males (16.8 ± 7.0 vs 14.7 ± 4.8, p  = 0.038, n  = 9) but not females. Effect in females may be disrupted by the action of progesterone, which was significantly correlated with this population ( R  = 0.771, p  = 0.025, n  = 8) after papaya consumption. In males, total T helper cells were increased (33.4 ± 6.4 vs 32.4 ± 6.1%, p  = 0.040, n  = 15). Strong significant negative correlation between testosterone and CD25 + CD4 + lymphocytes, may play a role in the lower total CD4 + T cells reported in males. Thus, dissimilar immune profiles were elicited in the sexes after papaya consumption and may have sex hormone influence.

Characterization of pancreatic stem cells derived from adult human pancreas ducts by fluorescence activated cell sorting.

PubMed

Lin, Han-Tso; Chiou, Shih-Hwa; Kao, Chung-Lan; Shyr, Yi-Ming; Hsu, Chien-Jen; Tarng, Yih-Wen; Ho, Larry L-T; Kwok, Ching-Fai; Ku, Hung-Hai

2006-07-28

To isolate putative pancreatic stem cells (PSCs) from human adult tissues of pancreas duct using serum-free, conditioned medium. The characterization of surface phenotype of these PSCs was analyzed by flow cytometry. The potential for pancreatic lineage and the capability of beta-cell differentiation in these PSCs were evaluated as well. By using serum-free medium supplemented with essential growth factors, we attempted to isolate the putative PSCs which has been reported to express nestin and pdx-1. The Matrigel(TM) was employed to evaluate the differential capacity of isolated cells. Dithizone staining, insulin content/secretion measurement, and immunohistochemistry staining were used to monitor the differentiation. Fluorescence activated cell sorting (FACS) was used to detect the phenotypic markers of putative PSCs. A monolayer of spindle-like cells was cultivated. The putative PSCs expressed pdx-1 and nestin. They were also able to differentiate into insulin-, glucagon-, and somatostatin-positive cells. The spectrum of phenotypic markers in PSCs was investigated; a similarity was revealed when using human bone marrow-derived stem cells as the comparative experiment, such as CD29, CD44, CD49, CD50, CD51, CD62E, PDGFR-alpha, CD73 (SH2), CD81, CD105(SH3). In this study, we successfully isolated PSCs from adult human pancreatic duct by using serum-free medium. These PSCs not only expressed nestin and pdx-1 but also exhibited markers attributable to mesenchymal stem cells. Although work is needed to elucidate the role of these cells, the application of these PSCs might be therapeutic strategies for diabetes mellitus.

Early pediatric atopic dermatitis shows only a cutaneous lymphocyte antigen (CLA)(+) TH2/TH1 cell imbalance, whereas adults acquire CLA(+) TH22/TC22 cell subsets.

PubMed

Czarnowicki, Tali; Esaki, Hitokazu; Gonzalez, Juana; Malajian, Dana; Shemer, Avner; Noda, Shinji; Talasila, Sreya; Berry, Adam; Gray, Jayla; Becker, Lauren; Estrada, Yeriel; Xu, Hui; Zheng, Xiuzhong; Suárez-Fariñas, Mayte; Krueger, James G; Paller, Amy S; Guttman-Yassky, Emma

2015-10-01

Identifying differences and similarities between cutaneous lymphocyte antigen (CLA)(+) polarized T-cell subsets in children versus adults with atopic dermatitis (AD) is critical for directing new treatments toward children. We sought to compare activation markers and frequencies of skin-homing (CLA(+)) versus systemic (CLA(-)) "polar" CD4 and CD8 T-cell subsets in patients with early pediatric AD, adults with AD, and control subjects. Flow cytometry was used to measure CD69/inducible costimulator/HLA-DR frequency in memory cell subsets, as well as IFN-γ, IL-13, IL-9, IL-17, and IL-22 cytokines, defining TH1/cytotoxic T (TC) 1, TH2/TC2, TH9/TC9, TH17/TC17, and TH22/TC22 populations in CD4 and CD8 cells, respectively. We compared peripheral blood from 19 children less than 5 years old and 42 adults with well-characterized moderate-to-severe AD, as well as age-matched control subjects (17 children and 25 adults). Selective inducible costimulator activation (P < .001) was seen in children. CLA(+) TH2 T cells were markedly expanded in both children and adults with AD compared with those in control subjects, but decreases in CLA(+) TH1 T-cell numbers were greater in children with AD (17% vs 7.4%, P = .007). Unlike in adults, no imbalances were detected in CLA(-) T cells from pediatric patients with AD nor were there altered frequencies of TH22 T cells within the CLA(+) or CLA(-) compartments. Adults with AD had increased frequencies of IL-22-producing CD4 and CD8 T cells within the skin-homing population, compared with controls (9.5% vs 4.5% and 8.6% vs 2.4%, respectively; P < .001), as well as increased HLA-DR activation (P < .01). These data suggest that TH2 activation within skin-homing T cells might drive AD in children and that reduced counterregulation by TH1 T cells might contribute to excess TH2 activation. TH22 "spreading" of AD is not seen in young children and might be influenced by immune development, disease chronicity, or recurrent skin infections. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Impact of HIV Infection and Anti-Retroviral Therapy on the Immune Profile of and Microbial Translocation in HIV-Infected Children in Vietnam.

PubMed

Bi, Xiuqiong; Ishizaki, Azumi; Nguyen, Lam Van; Matsuda, Kazunori; Pham, Hung Viet; Phan, Chung Thi Thu; Ogata, Kiyohito; Giang, Thuy Thi Thanh; Phung, Thuy Thi Bich; Nguyen, Tuyen Thi; Tokoro, Masaharu; Pham, An Nhat; Khu, Dung Thi Khanh; Ichimura, Hiroshi

2016-08-02

CD4⁺ T-lymphocyte destruction, microbial translocation, and systemic immune activation are the main mechanisms of the pathogenesis of human immunodeficiency virus type 1 (HIV) infection. To investigate the impact of HIV infection and antiretroviral therapy (ART) on the immune profile of and microbial translocation in HIV-infected children, 60 HIV vertically infected children (31 without ART: HIV(+) and 29 with ART: ART(+)) and 20 HIV-uninfected children (HIV(-)) aged 2-12 years were recruited in Vietnam, and their blood samples were immunologically and bacteriologically analyzed. Among the HIV(+) children, the total CD4⁺-cell and their subset (type 1 helper T-cell (Th1)/Th2/Th17) counts were inversely correlated with age (all p < 0.05), whereas regulatory T-cell (Treg) counts and CD4/CD8 ratios had become lower, and the CD38⁺HLA (human leukocyte antigen)-DR⁺CD8⁺- (activated CD8⁺) cell percentage and plasma soluble CD14 (sCD14, a monocyte activation marker) levels had become higher than those of HIV(-) children by the age of 2 years; the CD4/CD8 ratio was inversely correlated with the plasma HIV RNA load and CD8⁺-cell activation status. Among the ART(+) children, the total CD4⁺-cell and Th2/Th17/Treg-subset counts and the CD4/CD8 ratio gradually increased, with estimated ART periods of normalization being 4.8-8.3 years, whereas Th1 counts and the CD8⁺-cell activation status normalized within 1 year of ART initiation. sCD14 levels remained high even after ART initiation. The detection frequency of bacterial 16S/23S ribosomal DNA/RNA in blood did not differ between HIV-infected and -uninfected children. Thus, in children, HIV infection caused a rapid decrease in Treg counts and the early activation of CD8⁺ cells and monocytes, and ART induced rapid Th1 recovery and early CD8⁺-cell activation normalization but had little effect on monocyte activation. The CD4/CD8 ratio could therefore be an additional marker for ART monitoring.

Combining FoxP3 and Helios with GARP/LAP markers can identify expanded Treg subsets in cancer patients.

PubMed

Abd Al Samid, May; Chaudhary, Belal; Khaled, Yazan S; Ammori, Basil J; Elkord, Eyad

2016-03-22

Regulatory T cells (Tregs) comprise numerous heterogeneous subsets with distinct phenotypic and functional features. Identifying Treg markers is critical to investigate the role and clinical impact of various Treg subsets in pathological settings, and also for developing more effective immunotherapies. We have recently shown that non-activated FoxP3-Helios+ and activated FoxP3+/-Helios+ CD4+ T cells express GARP/LAP immunosuppressive markers in healthy donors. In this study we report similar observations in the peripheral blood of patients with pancreatic cancer (PC) and liver metastases from colorectal cancer (LICRC). Comparing levels of different Treg subpopulations in cancer patients and controls, we report that in PC patients, and unlike LICRC patients, there was no increase in Treg levels as defined by FoxP3 and Helios. However, defining Tregs based on GARP/LAP expression showed that FoxP3-LAP+ Tregs in non-activated and activated settings, and FoxP3+Helios+GARP+LAP+ activated Tregs were significantly increased in both groups of patients, compared with controls. This work implies that a combination of Treg-specific markers could be used to more accurately determine expanded Treg subsets and to understand their contribution in cancer settings. Additionally, GARP-/+LAP+ CD4+ T cells made IL-10, and not IFN-γ, and levels of IL-10-secreting CD4+ T cells were elevated in LICRC patients, especially with higher tumor staging. Taken together, our results indicate that investigations of Treg levels in different cancers should consider diverse Treg-related markers such as GARP, LAP, Helios, and others and not only FoxP3 as a sole Treg-specific marker.

Tuberculin-Specific T Cells Are Reduced in Active Pulmonary Tuberculosis Compared to LTBI or Status Post BCG Vaccination

PubMed Central

Streitz, Mathias; Fuhrmann, Stephan; Powell, Fiona; Quassem, Ali; Nomura, Laurel; Maecker, Holden; Martus, Peter; Volk, Hans-Dieter

2011-01-01

Functional characteristics of tuberculosis (TB)–specific CD4 T cells were studied in clinically active pulmonary TB (n = 21) and high TB exposure including LTBI (n = 17). Following tuberculin stimulation, activated CD4 T cells were identified by flow-cytometry (CD154 up-regulation, degranulation, interferon γ [IFN-γ], tumor necrosis factor α [TNF-α], and interleukin 2 [IL-2\\ production). Interestingly, CD154 up-regulation accounted for ∼80% of activated CD4 T cells in the active TB group but just 40% in the controls, whereas IFN-γ accounted for only ∼50% of activated cells in each group. The frequencies of CD4 T cells displaying at least 1 activation marker discriminated better between the groups than those displaying degranulation or IFN-γ production alone. PMID:21186260

Soluble CD14 in cerebrospinal fluid is associated with markers of inflammation and axonal damage in untreated HIV-infected patients: a retrospective cross-sectional study.

PubMed

Jespersen, Sofie; Pedersen, Karin Kæreby; Anesten, Birgitta; Zetterberg, Henrik; Fuchs, Dietmar; Gisslén, Magnus; Hagberg, Lars; Trøseid, Marius; Nielsen, Susanne Dam

2016-04-21

HIV-associated cognitive impairment has declined since the introduction of combination antiretroviral treatment (cART). However, milder forms of cognitive impairment persist. Inflammation in the cerebrospinal fluid (CSF) has been associated with cognitive impairment, and CSF neurofilament light chain protein (NFL) and CSF neopterin concentrations are increased in those patients. Microbial translocation in HIV infection has been suggested to contribute to chronic inflammation, and lipopolysaccharide (LPS) and soluble CD14 (sCD14) are markers of microbial translocation and the resulting monocyte activation, respectively. We hypothesised that microbial translocation contributes to inflammation and axonal damage in the central nervous system (CNS) in untreated HIV infection. We analyzed paired samples of plasma and CSF from 62 HIV-infected, untreated patients without cognitive symptoms from Sahlgrenska University Hospital, Gothenburg, Sweden. Measurements of neopterin and NFL in CSF were available from previous studies. Plasma and CSF sCD14 was measured using ELISA (R&D, Minneapolis, MN), and plasma and CSF LPS was measured using LAL colorimetric assay (Lonza, Walkersville, MD, USA). Univariate and multivariate regression analyses were performed. LPS in plasma was associated with plasma sCD14 (r = 0.31, P = 0.015), and plasma sCD14 was associated with CSF sCD14 (r = 0.32, P = 0.012). Furthermore, CSF sCD14 was associated with NFL (r = 0.32, P = 0.031) and neopterin (r = 0.32, P = 0.012) in CSF. LPS was not detectable in CSF. In a multivariate regression model CSF sCD14 remained associated with NFL and neopterin after adjusting for age, CD4+ cell count, and HIV RNA in CSF. In a group of untreated, HIV-infected patients LPS was associated with sCD14 in plasma, and plasma sCD14 was associated CSF sCD14. CSF sCD14 were associated with markers of CNS inflammation and axonal damage. This suggest that microbial translocation might be a driver of systemic and CNS inflammation. However, LPS was not detectable in the CSF, and since sCD14 is a marker of monocyte activation sCD14 may be increased due to other causes than microbial translocation. Further studies regarding cognitive impairment and biomarkers are warranted to fully understand causality.

The CD4/CD8 Ratio is Inversely Associated with Carotid Intima-Media Thickness Progression in Human Immunodeficiency Virus-Infected Patients on Antiretroviral Treatment.

PubMed

Bernal Morell, Enrique; Serrano Cabeza, José; Muñoz, Ángeles; Marín, Irene; Masiá, Mar; Gutiérrez, Félix; Cano, Alfredo

2016-07-01

Inversion of the CD4/CD8 ratio (<1) has been identified as a surrogate marker of immunosenescence and an independent predictor of AIDS events in HIV-infected patients and mortality in the general population. We aimed to assess the association between the CD4/CD8 ratio and carotid intima-media thickness (cIMT) progression in treated HIV-infected patients as a marker of coronary heart disease. A longitudinal study was conducted during 3 years in 96 virally suppressed HIV-infected patients receiving antiretroviral treatment (ART). We analyzed the associations between the CD4/CD8 ratio, cardiovascular risk factors, and antiretroviral treatment (ART) and progression of subclinical atherosclerosis assessed using cIMT at baseline and after 3 years. Finally, 96 patients completed the study. Seventy six (79.1%) patients were male, aged 44 ± 10 years; 39 (40.6%) were on treatment with protease inhibitors; 49 (51.04%) with nonnucleoside reverse transcriptase inhibitors, 6 (6.25%) with integrase inhibitors, 3 (3.12%) with maraviroc, and 2 (2.08%) just with nucleoside reverse transcriptase inhibitors. The mean of ART exposition was 6.9 ± 5.9 years. Twenty six (27%) patients had family history of ischemic heart disease, 51 (53.12%) were smokers, 12 (12.5%) were hypertensive, 4 (4.16%) had type 2 diabetes, 23 (23.9%) had dyslipidemia, and 31 (32.3%) were infected with hepatitis C virus. Baseline cIMT was significantly associated with age (rho = 0.497; p < .001), basal glucemia (rho = 0.323; p = .001), triglycerides (rho = 0.232; p = .023), Framingham risk score (rho = 0.324; p = .001), CD4/CD8 ratio (rho = -0.176; p = .05), and dyslipidemia (0.72 ± 0.16 mm vs. 0.63 ± 0.11 mm; p = .029). In multivariable analysis where cardiovascular risk factor and ART were included, cIMT progression was inversely associated with CD4/CD8 ratio [odds ratio (OR) = 0.283; confidence interval (95% CI) 0.099-0.809; p = .019]. In conclusion, the inversion of CD4/CD8 ratio in treated HIV-infected patients is independently associated with cIMT progression, a marker of coronary heart disease. Therefore, it might be clinically useful as predictor of cardiovascular events.

Aggressive Phenotype of Cells Disseminated via Hematogenous and Lymphatic Route in Breast Cancer Patients.

PubMed

Markiewicz, Aleksandra; Nagel, Anna; Szade, Jolanta; Majewska, Hanna; Skokowski, Jaroslaw; Seroczynska, Barbara; Stokowy, Tomasz; Welnicka-Jaskiewicz, Marzena; Zaczek, Anna J

2018-06-01

Intratumoral heterogeneity of breast cancer remains a major challenge in successful treatment. Failure of cancer therapies can also be accredited to inability to systemically eradicate cancer stem cells (CSCs). Recent evidence points to the role of epithelial-mesenchymal transition (EMT) in expanding the pool of tumor cells with CSCs features. Thus, we assessed expression level as well as heterogeneity of CSCs markers in primary tumors (PT), lymph node metastasis (LNM), and circulating tumor cells (CTCs)-enriched blood fractions in order to correlate them with signs of EMT activation as well as clinicopathological data of breast cancer patients. Level of CSCs markers (ALDH1, CD44, CD133, OCT-4, NANOG) and EMT markers was quantified in PT (N=107), LNM (N=56), and CTCs-enriched blood fractions (N=85). Heterogeneity of CSCs markers expression within each PT and LNM was assessed by calculating Gini Index. Percentage of ALDH1-positive cells was elevated in PT in comparison to LNM (P = .005). However, heterogeneity of the four CSCs markers: ALDH1 (P = .019), CD133 (P = .009), OCT-4 (P = .027), and CD44 (P < .001) was decreased in LNM. Samples classified as mesenchymal (post-EMT) showed elevated expression of CSCs markers (OCT-4 and CD44 in PT; OCT-4 in LNM; ALDH1, OCT-4, NANOG, CD44 in CTCs). Patients with mesenchymal-like CTCs had worse prognosis than patients with epithelial-like or no CTCs (P = .0025). CSCs markers are enriched in PT, LNM, and CTCs with mesenchymal features, but their heterogeneity is decreased in metastatic lymph nodes. Mesenchymal CTCs phenotype correlates with poor prognosis of the patients. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Mass Cytometry Identifies Distinct Lung CD4+ T Cell Patterns in Löfgren’s Syndrome and Non-Löfgren’s Syndrome Sarcoidosis

PubMed Central

Kaiser, Ylva; Lakshmikanth, Tadepally; Chen, Yang; Mikes, Jaromir; Eklund, Anders; Brodin, Petter; Achour, Adnane; Grunewald, Johan

2017-01-01

Sarcoidosis is a granulomatous disorder of unknown etiology, characterized by accumulation of activated CD4+ T cells in the lungs. Disease phenotypes Löfgren’s syndrome (LS) and “non-LS” differ in terms of clinical manifestations, genetic background, HLA association, and prognosis, but the underlying inflammatory mechanisms largely remain unknown. Bronchoalveolar lavage fluid cells from four HLA-DRB1*03+ LS and four HLA-DRB1*03− non-LS patients were analyzed by mass cytometry, using a panel of 33 unique markers. Differentially regulated CD4+ T cell populations were identified using the Citrus algorithm, and t-stochastic neighborhood embedding was applied for dimensionality reduction and single-cell data visualization. We identified 19 individual CD4+ T cell clusters differing significantly in abundance between LS and non-LS patients. Seven clusters more frequent in LS patients were characterized by significantly higher expression of regulatory receptors CTLA-4, PD-1, and ICOS, along with low expression of adhesion marker CD44. In contrast, 12 clusters primarily found in non-LS displayed elevated expression of activation and effector markers HLA-DR, CD127, CD39, as well as CD44. Hierarchical clustering further indicated functional heterogeneity and diverse origins of T cell receptor Vα2.3/Vβ22-restricted cells in LS. Finally, a near-complete overlap of CD8 and Ki-67 expression suggested larger influence of CD8+ T cell activity on sarcoid inflammation than previously appreciated. In this study, we provide detailed characterization of pulmonary T cells and immunological parameters that define separate disease pathways in LS and non-LS. With direct association to clinical parameters, such as granuloma persistence, resolution, or chronic inflammation, these results provide a valuable foundation for further exploration and potential clinical application. PMID:28955342

[CD69 expression on T cell surface in patients with obstructive sleep apnea hypopnea syndrome].

PubMed

Chen, Y H; Wang, P F; Wang, H J; Hu, X; Li, Z Y; Xiong, H Q

2017-02-20

Objective: To investigate the detection and significance of T cell CD69 expression in peripheral blood of patients with obstructive sleep apnea hypopnea syndrome (OSAHS). Method: According to AHI, 81 OSAHS patients diagnosed by PSG were divided into 3 groups: light, medium and heavy, with 27 cases in each group; 27 patients without OSAHS as control group. Flow cytometry was used to detect the expression rate of CD69 in T cells, to analyze the correlation between the expression rate of CD69 on T cells and the gender, age, BMI, and PSG index. Result: ①The CD69 expression rate of T cells in peripheral blood of OSAHS patients with snoring degree increases gradually ( P < 0.05); Comparison between the two shows that there was no significant difference in CD69 expression rate on T cells between the control group and the mild group ( t = 1.649, P > 0.05); there were significant differences between the other groups ( P < 0.05). ②The CD69 expression rate of T cells in peripheral blood of OSAHS patients has no correlation with BMI, age and gender ( P > 0.05), were positively correlated with AHI, negatively correlated with LSaO₂ ( P < 0.01). ③The CD69 expression rate of T cells and AHI in 27 cases of severe OSAHS patients with a comprehensive treatment has significantly reduced, LSaO₂ increased significantly ( P < 0.01). Conclusion: Increased expression of CD69 in peripheral blood T cells may be one of the mechanisms of OSAHS complicated with cardiovascular disease. Detection of CD69 expression rate in T cells for reflecting the degree of disease in patients with OSAHS, assessment of risk of cardiovascular damage, have certain clinical significance. Copyright© by the Editorial Department of Journal of Clinical Otorhinolaryngology Head and Neck Surgery.

Monocyte-lymphocyte fusion induced by the HIV-1 envelope generates functional heterokaryons with an activated monocyte-like phenotype.

PubMed

Martínez-Méndez, David; Rivera-Toledo, Evelyn; Ortega, Enrique; Licona-Limón, Ileana; Huerta, Leonor

2017-03-01

Enveloped viruses induce cell-cell fusion when infected cells expressing viral envelope proteins interact with target cells, or through the contact of cell-free viral particles with adjoining target cells. CD4 + T lymphocytes and cells from the monocyte-macrophage lineage express receptors for HIV envelope protein. We have previously reported that lymphoid Jurkat T cells expressing the HIV-1 envelope protein (Env) can fuse with THP-1 monocytic cells, forming heterokaryons with a predominantly myeloid phenotype. This study shows that the expression of monocytic markers in heterokaryons is stable, whereas the expression of lymphoid markers is mostly lost. Like THP-1 cells, heterokaryons exhibited FcγR-dependent phagocytic activity and showed an enhanced expression of the activation marker ICAM-1 upon stimulation with PMA. In addition, heterokaryons showed morphological changes compatible with maturation, and high expression of the differentiation marker CD11b in the absence of differentiation-inducing agents. No morphological change nor increase in CD11b expression were observed when an HIV-fusion inhibitor blocked fusion, or when THP-1 cells were cocultured with Jurkat cells expressing a non-fusogenic Env protein, showing that differentiation was not induced merely by cell-cell interaction but required cell-cell fusion. Inhibition of TLR2/TLR4 signaling by a TIRAP inhibitor greatly reduced the expression of CD11b in heterokaryons. Thus, lymphocyte-monocyte heterokaryons induced by HIV-1 Env are stable and functional, and fusion prompts a phenotype characteristic of activated monocytes via intracellular TLR2/TLR4 signaling. Copyright © 2017 Elsevier Inc. All rights reserved.

CD137 is a Useful Marker for Identifying CD4+ T Cell Responses to Mycobacterium tuberculosis.

PubMed

Yan, Z-H; Zheng, X-F; Yi, L; Wang, J; Wang, X-J; Wei, P-J; Jia, H-Y; Zhou, L-J; Zhao, Y-L; Zhang, H-T

2017-05-01

Upregulation of CD137 on recently activated CD8 + T cells has been used to identify rare viral and tumour antigen-specific T cells from the peripheral blood. We aimed to evaluate the accuracy of CD137 for identifying Mycobacterium tuberculosis (Mtb)-reactive CD4 + T cells in the peripheral blood of infected individuals by flow cytometry and to investigate the characteristics of these CD137 + CD4 + T cells. We initially enrolled 31 active tuberculosis (TB) patients, 31 individuals with latent TB infection (LTBI) and 25 healthy donors. The intracellular CD137 and interferon-γ (IFN-γ) production by CD4 + T cells was simultaneously detected under unstimulated and CFP10-stimulated (culture filtrate protein 10, a Mtb-specific antigen) conditions. In unstimulated CD4 + T cells, we found that the CD137 expression in the TB group was significantly higher than that in the LTBI group. Stimulation with CFP10 largely increased the CD4 + T cell CD137 expression in both the TB and LTBI groups. After CFP10 stimulation, the frequency of CD137 + CD4 + T cells was higher than that of IFN-γ + CD4 + T cells in both the TB and LTBI groups. Most of the CFP10-activated IFN-γ-secreting cells were CD137-positive, but only a small fraction of the CD137-positive cells expressed IFN-γ. An additional 20 patients with TB were enrolled to characterize the CD45RO + CCR7 + , CD45RO + CCR7 - and CD45RO - subsets in the CD137 + CD4 + T cell populations. The Mtb-specific CD137 + CD4 + T cells were mainly identified as having an effector memory phenotype. In conclusion, CD137 is a useful marker that can be used for identifying Mtb-reactive CD4 + T cells by flow cytometry. © 2017 The Foundation for the Scandinavian Journal of Immunology.

Ultrafine particles and platelet activation in patients with coronary heart disease – results from a prospective panel study

PubMed Central

Rückerl, Regina; Phipps, Richard P; Schneider, Alexandra; Frampton, Mark; Cyrys, Josef; Oberdörster, Günther; Wichmann, H Erich; Peters, Annette

2007-01-01

Background Epidemiological studies on health effects of air pollution have consistently shown adverse cardiovascular effects. Toxicological studies have provided evidence for thrombogenic effects of particles. A prospective panel study in a susceptible population was conducted in Erfurt, Germany, to study the effects of daily changes in ambient particles on various blood cells and soluble CD40ligand (sCD40L, also known as CD154), a marker for platelet activation that can cause increased coagulation and inflammation. Blood cells and plasma sCD40L levels were repeatedly measured in 57 male patients with coronary heart disease (CHD) during winter 2000/2001. Fixed effects linear regression models were applied, adjusting for trend, weekday and meteorological parameters. Hourly data on ultrafine particles (UFP, number concentration of particles from 0.01 to 0.1 μm), mass concentration of particles less than 10 and 2.5 μm in diameter (PM10, PM2.5), accumulation mode particle counts (AP, 0.1–1.0 μm), elemental and organic carbon, gaseous pollutants and meteorological data were collected at central monitoring sites. Results An immediate increase in plasma sCD40L was found in association with UFP and AP (% change from geometric mean: 7.1; CI: [0.1, 14.5] and 6.9; CI: [0.5, 13.8], respectively). Platelet counts decreased in association with UFP showing an immediate, a three days delayed (lag 3) and a 5-day average response (% change from the mean: -1.8; CI: [-3.4,-0.2]; -2.4; CI: [-4.5,-0.3] and -2.2; CI: [-4.0,-0.3] respectively). Conclusion The increased plasma sCD40L levels support the hypothesis that higher levels of ambient air pollution lead to an inflammatory response in patients with CHD thus providing a possible explanation for the observed association between air pollution and cardiovascular morbidity and mortality in susceptible parts of the population. PMID:17241467

Mucosal CCR1 gene expression as a marker of molecular activity in Crohn's disease: preliminary data.

PubMed

Dobre, Maria; Mănuc, Teodora Ecaterina; Milanesi, Elena; Pleşea, Iancu Emil; Ţieranu, Eugen Nicolae; Popa, Caterina; Mănuc, Mircea; Preda, Carmen Monica; Ţieranu, Ioana; Diculescu, Mihai Mircea; Ionescu, Elena Mirela; Becheanu, Gabriel

2017-01-01

A series of mechanisms of immune response, inflammation and apoptosis have been demonstrated to contribute to the appearance and evolution of Crohn's disease (CD) through the overexpression of several cytokines and chemokines in a susceptible host. The aim of this study was to identify the differences in gene expression profiles analyzing a panel of candidate genes in the mucosa from patients with active CD (CD-A), patients in remission (CD-R), and normal controls. Nine individuals were enrolled in the study: six CD patients (three with active lesions, three with mucosal healing) and three controls without inflammatory bowel disease (IBD) seen on endoscopy. All the individuals underwent mucosal biopsy during colonoscopy. Gene expression levels of 84 genes previously associated with CD were evaluated by polymerase chain reaction (PCR) array. Ten genes out of 84 were found significantly differentially expressed in CD-A (CCL11, CCL25, DEFA5, GCG, IL17A, LCN2, REG1A, STAT3, MUC1, CCR1) and eight genes in CD-R (CASP1, IL23A, STAT1, STAT3, TNF, CCR1, CCL5, and HSP90B1) when compared to controls. A quantitative gene expression analysis revealed that CCR1 gene was more expressed in CD-A than in CD-R. Our data suggest that CCR1 gene may be a putative marker of molecular activity of Crohn's disease. Following these preliminary data, a confirmation in larger cohort studies could represent a useful method in order to identify new therapeutic targets.

Improved Cognitive Performance and Reduced Monocyte Activation in Virally Suppressed Chronic HIV Following Dual CCR2 and CCR5 Antagonism.

PubMed

D'Antoni, Michelle L; Paul, Robert H; Mitchell, Brooks I; Kohorn, Lindsay; Fischer, Laurent; Lefebvre, Eric; Seyedkazemi, Star; Nakamoto, Beau K; Walker, Maegen; Kallianpur, Kalpana J; Ogata-Arakaki, Debra; Ndhlovu, Lishomwa C; Shikuma, Cecilia

2018-05-16

To evaluate changes in neuropsychological (NP) performance and in plasma and cell surface markers of peripheral monocyte activation/migration following treatment with cenicriviroc (CVC), a dual C-C chemokine receptor type 2 (CCR2) and type 5 (CCR5) antagonist, in treatment-experienced, HIV-infected individuals. Single-arm, 24-week, open-label clinical trial. HIV-infected individuals on antiretroviral therapy (ART) >1 year with plasma HIV RNA <50 copies/ml and below-normal cognitive performance [defined as age, gender and education-adjusted NP performance (NPZ) <-0.5 in a single cognitive domain or in global performance] were enrolled. Changes over 24 weeks were assessed for global and domain-specific NPZ scores, plasma markers of monocyte/macrophage activation [neopterin, soluble (s)CD14 and sCD163] quantified by ELISA, and CCR2 and CCR5 expression on monocytes and T cells measured by flow cytometry. Seventeen of 20 enrolled participants completed the study. Improvements over 24 weeks were observed in global NPZ [median change (Δ)=0.24; p=0.008], and in cognitive domains of attention (Δ0.23; p=0.011) and working memory (Δ0.44; p=0.017). Plasma levels of sCD163, sCD14, and neopterin decreased significantly (p's<0.01). CCR2 and CCR5 monocyte expression remained unchanged; however, CCR5 levels on CD4 and CD8 T cells and CCR2 expression on CD4 T cells increased (p's<0.01). CVC given over 24 weeks was associated with improved NP test performance and decreased plasma markers of monocyte immune activation in virally-suppressed, HIV-infected participants. These data potentially link changes in monocyte activation to cognitive performance. Further study of CVC for HIV cognitive impairment in a randomized controlled study is warranted.

Burn-injury affects gut-associated lymphoid tissues derived CD4+ T cells.

PubMed

Fazal, Nadeem; Shelip, Alla; Alzahrani, Alhusain J

2013-01-01

After scald burn-injury, the intestinal immune system responds to maintain immune balance. In this regard CD4+T cells in Gut-Associated Lymphoid Tissues (GALT), like mesenteric lymph nodes (MLN) and Peyer's patches (PP) respond to avoid immune suppression following major injury such as burn. Therefore, we hypothesized that the gut CD4+T cells become dysfunctional and turn the immune homeostasis towards depression of CD4+ T cell-mediated adaptive immune responses. In the current study we show down regulation of mucosal CD4+ T cell proliferation, IL-2 production and cell surface marker expression of mucosal CD4+ T cells moving towards suppressive-type. Acute burn-injury lead to up-regulation of regulatory marker (CD25+), down regulation of adhesion (CD62L, CD11a) and homing receptor (CD49d) expression, and up-regulation of negative co-stimulatory (CTLA-4) molecule. Moreover, CD4+CD25+ T cells of intestinal origin showed resistance to spontaneous as well as induced apoptosis that may contribute to suppression of effector CD4+ T cells. Furthermore, gut CD4+CD25+ T cells obtained from burn-injured animals were able to down-regulate naïve CD4+ T cell proliferation following adoptive transfer of burn-injured CD4+CD25+ T cells into sham control animals, without any significant effect on cell surface activation markers. Together, these data demonstrate that the intestinal CD4+ T cells evolve a strategy to promote suppressive CD4+ T cell effector responses, as evidenced by enhanced CD4+CD25+ T cells, up-regulated CTLA-4 expression, reduced IL-2 production, tendency towards diminished apoptosis of suppressive CD4+ T cells, and thus lose their natural ability to regulate immune homeostasis following acute burn-injury and prevent immune paralysis.

Malnutrition in HIV-Infected Children Is an Indicator of Severe Disease with an Impaired Response to Antiretroviral Therapy.

PubMed

Muenchhoff, Maximilian; Healy, Michael; Singh, Ravesh; Roider, Julia; Groll, Andreas; Kindra, Chirjeev; Sibaya, Thobekile; Moonsamy, Angeline; McGregor, Callum; Phan, Michelle Q; Palma, Alejandro; Kloverpris, Henrik; Leslie, Alasdair; Bobat, Raziya; LaRussa, Philip; Ndung'u, Thumbi; Goulder, Philip; Sobieszczyk, Magdalena E; Archary, Mohendran

2018-01-01

This observational study aimed to describe immunopathogenesis and treatment outcomes in children with and without severe acute malnutrition (SAM) and HIV-infection. We studied markers of microbial translocation (16sDNA), intestinal damage (iFABP), monocyte activation (sCD14), T-cell activation (CD38, HLA-DR) and immune exhaustion (PD1) in 32 HIV-infected children with and 41 HIV-infected children without SAM prior to initiation of antiretroviral therapy (ART) and cross-sectionally compared these children to 15 HIV-uninfected children with and 19 HIV-uninfected children without SAM. We then prospectively measured these markers and correlated them to treatment outcomes in the HIV-infected children at 48 weeks following initiation of ART. Plasma levels of 16sDNA, iFABP and sCD14 were measured by quantitative real time PCR, ELISA and Luminex, respectively. T cell phenotype markers were measured by flow cytometry. Multiple regression analysis was performed using generalized linear models (GLMs) and the least absolute shrinkage and selection operator (LASSO) approach for variable selection. Microbial translocation, T cell activation and exhaustion were increased in HIV-uninfected children with SAM compared to HIV-uninfected children without SAM. In HIV-infected children microbial translocation, immune activation, and exhaustion was strongly increased but did not differ by SAM-status. SAM was associated with increased mortality rates early after ART initiation. Malnutrition, age, microbial translocation, monocyte, and CD8 T cell activation were independently associated with decreased rates of CD4% immune recovery after 48 weeks of ART. SAM is associated with increased microbial translocation, immune activation, and immune exhaustion in HIV-uninfected children and with worse prognosis and impaired immune recovery in HIV-infected children on ART.

Malnutrition in HIV-Infected Children Is an Indicator of Severe Disease with an Impaired Response to Antiretroviral Therapy

PubMed Central

Healy, Michael; Singh, Ravesh; Roider, Julia; Groll, Andreas; Kindra, Chirjeev; Sibaya, Thobekile; Moonsamy, Angeline; McGregor, Callum; Phan, Michelle Q.; Palma, Alejandro; Kloverpris, Henrik; Leslie, Alasdair; Bobat, Raziya; LaRussa, Philip; Ndung'u, Thumbi; Goulder, Philip; Sobieszczyk, Magdalena E.; Archary, Mohendran

2018-01-01

Abstract This observational study aimed to describe immunopathogenesis and treatment outcomes in children with and without severe acute malnutrition (SAM) and HIV-infection. We studied markers of microbial translocation (16sDNA), intestinal damage (iFABP), monocyte activation (sCD14), T-cell activation (CD38, HLA-DR) and immune exhaustion (PD1) in 32 HIV-infected children with and 41 HIV-infected children without SAM prior to initiation of antiretroviral therapy (ART) and cross-sectionally compared these children to 15 HIV-uninfected children with and 19 HIV-uninfected children without SAM. We then prospectively measured these markers and correlated them to treatment outcomes in the HIV-infected children at 48 weeks following initiation of ART. Plasma levels of 16sDNA, iFABP and sCD14 were measured by quantitative real time PCR, ELISA and Luminex, respectively. T cell phenotype markers were measured by flow cytometry. Multiple regression analysis was performed using generalized linear models (GLMs) and the least absolute shrinkage and selection operator (LASSO) approach for variable selection. Microbial translocation, T cell activation and exhaustion were increased in HIV-uninfected children with SAM compared to HIV-uninfected children without SAM. In HIV-infected children microbial translocation, immune activation, and exhaustion was strongly increased but did not differ by SAM-status. SAM was associated with increased mortality rates early after ART initiation. Malnutrition, age, microbial translocation, monocyte, and CD8 T cell activation were independently associated with decreased rates of CD4% immune recovery after 48 weeks of ART. SAM is associated with increased microbial translocation, immune activation, and immune exhaustion in HIV-uninfected children and with worse prognosis and impaired immune recovery in HIV-infected children on ART. PMID:28670966

T Cell Activation Thresholds are Affected by Gravitational

NASA Technical Reports Server (NTRS)

Adams, Charley; Gonzalez, M.; Nelman-Gonzalez, M.

1999-01-01

T cells stimulated in space flight by various mitogenic signals show a dramatic reduction in proliferation and expression of early activation markers. Similar results are also obtained in a ground based model of microgravity, clinorotation, which provides a vector-averaged reduction of the apparent gravity on cells without significant shear force. Here we demonstrate that T cell inhibition is due to an increase in the required threshold for activation. Dose response curves indicate that cells activated during clinorotation require higher stimulation to achieve the same level of activation, as measured by CD69 expression. Interleukin 2 receptor expression, and DNA synthesis. The amount of stimulation necessary for 50% activation is 5 fold in the clinostat relative to static. Correlation of TCR internalization with activation also exhibit a dramatic right shift in clinorotation, demonstrating unequivocally that signal transduction mechanism independent of TCR triggering account for the increased activation threshold. Previous results from space flight experiments are consistent with the dose response curves obtained for clinorotation. Activation thresholds are important aspects of T cell memory, autoimmunity and tolerance Clinorotation is a useful, noninvasive tool for the study of cellular and biochemical event regulating T cell activation threshold and the effects of gravitation forces on these systems.

Activation of Latent HIV Using Drug-loaded Nanoparticles

NASA Astrophysics Data System (ADS)

Kovochich, Michael

Antiretroviral therapy is currently only capable of controlling human immunodeficiency virus (HIV) replication, rather than completely eradicating virus from patients. This is due in part to the establishment of a latent virus reservoir in resting CD4+ T-cells, which persists even in the presence of highly active antiretroviral therapy (HAART). It is thought that forced activation of latently infected cells could induce virus production, allowing targeting of the cell by the immune response. A variety of molecules are able to stimulate HIV from latency. However, no tested purging strategy has proven capable of eliminating the infection completely or preventing viral rebound if therapy is stopped. Hence, novel latency activation approaches are required. Nanoparticles can offer several advantages over more traditional drug delivery methods, including improved drug solubility, stability, and the ability to simultaneously target multiple different molecules to particular cell or tissue types. Here we describe the development of a novel lipid nanoparticle with the protein kinase C activator bryostatin-2 incorporated (LNP-Bry). These particles can target, activate primary human CD4+ T-cells, and stimulate latent virus production from human T-cell lines in vitro and from latently infected cells in a humanized mouse model ex vivo. This activation was synergistically enhanced by the histone deacetylase inhibitor (HDACi) sodium butyrate. Furthermore, LNP-Bry can also be loaded with the protease inhibitor nelfinavir (LNP-Bry-Nel), producing a particle capable of both activating latent virus and inhibiting viral spread. LNP-Bry was further tested for its in vivo biodistribution in both wild type mice (C57 black 6), as well as humanized mice (SCID-hu Thy/Liv, and bone marrow-liver-thymus [BLT]). LNP-Bry accumulated in the spleen and induced the early activation marker CD69 in wild type mice. Taken together, these data demonstrate the ability of nanotechnological approaches to provide improved methods for activating latent HIV and provide key proof-of-principle experiments showing how novel delivery systems may enhance future HIV therapy.

[The Influence of UV-Light on the Sub-Populational Composition and Expression of Membrane Markers of Lymphocytes of Donor Blood].

PubMed

Artyukhov, V G; Basharina, O V; Zemchenkova, O V; Ryazantsev, S V

2016-01-01

The influence of UV-light (240-390 nm) at dozes of 151 and 755 J/m2 on the content of membrane markers of lymphocytes using the method of flow cytometry was investigated. It was demonstrated that during incubation of UV-irradiated lymphocytes the change of their populational and sub-populational composition occurs. Expression of complexes of CD3, CD 19,.CD8, CD 16, CD25 and CD95 increased. This increase was caused mainly by de novo synthesis. UV-light had immunostimulating effect on CD8+ T-lymphocyte population. Together with the increase of cytotoxic cells and NK-cells, activation of lymphocytes (increased amount of CD25+ and CD95+ cells) took place. Amount of cells undergone apoptosis or necrosis increased proportionally to the dosage. These changes were more expressed during incubation of lymphocytes in nutrition medium without autological blood serum, e.g. under deficiency of growth factors and antioxidants.

Elevated soluble CD30 correlates with development of bronchiolitis obliterans syndrome following lung transplantation.

PubMed

Fields, Ryan C; Bharat, Ankit; Steward, Nancy; Aloush, Aviva; Meyers, Brian F; Trulock, Elbert P; Chapman, William C; Patterson, G Alexander; Mohanakumar, Thalachallour

2006-12-27

The long-term function of lung transplants is limited by chronic rejection (bronchiolitis obliterans syndrome, BOS). Due to lack of specific markers, BOS is diagnosed clinically. Because there is strong evidence that alloimmunity plays a significant role in the pathogenesis of BOS, we investigated whether soluble CD30 (sCD30), a T-cell activation marker, would correlate with BOS. Sera collected serially from BOS+ (n = 20) and matched BOS- (n = 20) lung transplant (LT) patients were analyzed for sCD30 by enzyme-linked immunosorbent assay. Pretransplant sera and sera from normal donors were also analyzed. PreLT levels were comparable to normal subjects. However, posttransplant there was a significant elevation in sCD30 levels during BOS development in all BOS+ patients, compared to BOS- (mean 139.8+/-10.7 vs. 14.8+/-2.7 U/ml, P < 0.001). sCD30 levels declined in the BOS+ patients but were still elevated compared to BOS- (48.52+/-5.04 vs. 7.19+/-2.9, P < 0.0001). We conclude that sCD30 may represent a novel marker to monitor the development of BOS.

Neutrophil CD64 expression, procalcitonin and presepsin are useful to differentiate infections from flares in SLE patients with SIRS.

PubMed

Echeverri, A; Naranjo-Escobar, J; Posso-Osorio, I; Aguirre-Valencia, D; Zambrano, D; Castaño, G L; Martínez, J D; Cañas, C A; Tobón, G J

2018-06-01

Background/Objective Differentiating systemic lupus erythematosus (SLE) activity from infections in febrile patients is difficult because of similar initial clinical presentation. The aim of this study is to evaluate the usefulness of a number of biomarkers for differentiating infections from activity in SLE patients admitted with systemic inflammatory response (SIRS). Methods Patients with SLE and SIRS admitted to the emergency room were included in this study. Measurements of different markers including procalcitonin, neutrophil CD64 expression and presepsin, were performed. Infection was considered present when positive cultures and/or polymerase chain reaction were obtained. Sensitivity and specificity were calculated for all biomarkers. Results Twenty-seven patients were admitted, 23 women (82.5%), mean age 33.2 years. An infectious disease was confirmed in 12 cases. Markers for SLE activity including anti-DNA titers by IIF ( p = 0.041) and enzyme-linked immunosorbent assay ( p = 0.009) were used for differentiating SLE flares from infection. On the contrary, increased procalcitonin ( p = 0.047), neutrophil CD64 expression by flow cytometry ( p = 0.037) and presepsin ( p = 0.037) levels were observed in infected SLE patients. Conclusions High neutrophil CD64 expression, presepsin and procalcitonin levels are useful to differentiate infections from activity in SLE patients. In most cases, a positive bioscore that includes these three markers demonstrate the presence of an infectious disease.

Licorice infusion: Chemical profile and effects on the activation and the cell cycle progression of human lymphocytes

PubMed Central

Cheel, José; Onofre, Gabriela; Vokurkova, Doris; Tůmová, Lenka; Neugebauerová, Jarmila

2010-01-01

A licorice infusion (LI) and its major constituents were investigated for their capacity to stimulate the activation and the cell cycle progression of human lymphocytes, measured by the CD69 expression and DNA content, respectively. The chemical profile of LI was determined by high-performance liquid chromatography-diode array detection (HPLC-DAD). Results: Two major components of LI were identified as liquiritin (1) and glycyrrhizin (2); total flavones and flavonols were shown as its minor constituents. The LI (100-800 μg/ml) stimulated the expression of CD69 on lymphocytes in a concentration-independent manner. Values of the activation index (AI) of total lymphocytes treated with LI (100-800 μg/ml) did not differ significantly among them (P < 0.05), but were 50% lower than the AI value exhibited by cells treated with phytohemagglutinin (PHA). The LI showed a similar effect on T cells, but on a lower scale. Compounds 1 and 2 (12-100 μg/ml) did not stimulate the CD69 expression on lymphocytes. The LI, 1 and 2 showed no meaningful effect on cell cycle progression of lymphocytes. The experimental data indicates that LI stimulates the activation of lymphocytes as a result of a proliferation-independent process. This finding suggests that LI could be considered as a potential specific immune stimulator. PMID:20548933

Action of the photosensitizer QLT0074 upon human T lymphocytes in vitro

NASA Astrophysics Data System (ADS)

Hunt, David W. C.; Jiang, Huijun; Salmon, Ruth A.; Granville, David J.; North, John R.; Richter, Anna M.

2001-04-01

A new photosensitizer, presently designated QLT0074, may be useful for the treatment of skin conditions, particularly those mediated by T lymphocytes, with photodynamic therapy (PDT). QLT0074 was tested against human peripheral blood T cells and Jurkat T lymphoma cells. Low concentrations of QLT0074 and blue light were sufficient to induce apoptosis in peripheral blood T cells or Jurkat T lymphoma cells as indicated by expression of the apoptosis-associated mitochondrial 7A6 marker, annexin-V labeling or activation of capsase-3 and cleavage of the capsase substrate poly(ADP-ribose)polymerase (PARP). Flow cytometry studies performed following PDT showed that peripheral blood T cells with high expression of the interleukin-2 receptor (CD25) took up greater amounts of QLT0074 and were eliminated to a greater degree than T cells with low CD25 levels. This effect of PDT was also shown by the reduction in the percentage of T cells that expressed other activation-associated markers such as very late activation antigen-4 (CD49d), human leukocyte antigen DR (HLA-DR), intercellular adhesion molecule-1 (CD54) and Fas (CD95). In the case of T cells that remained viable following PDT, CD25 expression was lower while CD54, CD95 and HLA-DR levels were unchanged. Thus, PDT with QLT0074 has selective, dose-dependent effects on T cells in vitro.

Monocyte-lymphocyte fusion induced by the HIV-1 envelope generates functional heterokaryons with an activated monocyte-like phenotype

DOE Office of Scientific and Technical Information (OSTI.GOV)

Martínez-Méndez, David; Rivera-Toledo, Evelyn; Ortega, Enrique

Enveloped viruses induce cell-cell fusion when infected cells expressing viral envelope proteins interact with target cells, or through the contact of cell-free viral particles with adjoining target cells. CD4{sup +} T lymphocytes and cells from the monocyte-macrophage lineage express receptors for HIV envelope protein. We have previously reported that lymphoid Jurkat T cells expressing the HIV-1 envelope protein (Env) can fuse with THP-1 monocytic cells, forming heterokaryons with a predominantly myeloid phenotype. This study shows that the expression of monocytic markers in heterokaryons is stable, whereas the expression of lymphoid markers is mostly lost. Like THP-1 cells, heterokaryons exhibited FcγR-dependentmore » phagocytic activity and showed an enhanced expression of the activation marker ICAM-1 upon stimulation with PMA. In addition, heterokaryons showed morphological changes compatible with maturation, and high expression of the differentiation marker CD11b in the absence of differentiation-inducing agents. No morphological change nor increase in CD11b expression were observed when an HIV-fusion inhibitor blocked fusion, or when THP-1 cells were cocultured with Jurkat cells expressing a non-fusogenic Env protein, showing that differentiation was not induced merely by cell-cell interaction but required cell-cell fusion. Inhibition of TLR2/TLR4 signaling by a TIRAP inhibitor greatly reduced the expression of CD11b in heterokaryons. Thus, lymphocyte-monocyte heterokaryons induced by HIV-1 Env are stable and functional, and fusion prompts a phenotype characteristic of activated monocytes via intracellular TLR2/TLR4 signaling. - Highlights: • Jurkat T cells expressing the HIV-1 envelope fuse with THP-1 monocytes. • Heterokaryons display a dominant myeloid phenotype and monocyte function. • Heterokaryons exhibit activation features in the absence of activation agents. • Activation is not due to cell-cell interaction but requires cell-cell fusion. • The activated monocyte-like phenotype is mediated by TLR2/TLR4 signaling.« less

Comparison of quantitative flow cytometric data provided by panels with lower and increased color number

NASA Astrophysics Data System (ADS)

Bocsi, József; Mittag, Anja; Pierzchalski, Arkadiusz; Baumgartner, Adolf; Dähnert, Ingo; Tárnok, Attila

2012-03-01

To date the flow cytometry (FCM) industry is booming with new generations of commercial clinical instruments. Long-term clinical studies have the dilemma that moving to new instruments being capable of more complex cell-analysis makes it difficult to compare new data with those obtained on older instruments with less complex analysis panels. Since 15 years we conduct follow-up studies on children with congenital heart diseases. In this period we moved from 2- to 3- and now to 10-color FCM immunophenotyping panels. Questions arise how to compare and transfer data from lower to higher level of complexity. Two comparable antibody panels for leukocyte immunophenotyping (12-tube 2-colors, and 9-tube 4-colors) were measured on a BD FACScalibur FCM (calibration: Spherotech beads) in 19 blood samples from children with congenital heart disease. This increase of colors was accompanied by moving antibodies that were in the 2-color panel either FITC or PE labeled to red dyes such as PerCP or APC. Algorithms were developed for bridging data for quantitative characterization of antigen expression (mean fluorescence intensity) and frequency of different cell subpopulations in combination with rainbow bead standard data. This approach worked for the most relevant antibodies (CD3, CD4, CD8 etc.) well, but rendered substantial uncertainty for activation markers (CD69 etc.). Our techniques are particularly well suited to the analysis in long-term studies and have the potential to compare older and recent results in a standardized way.

The rat macrophage scavenger receptor CD163: expression, regulation and role in inflammatory mediator production.

PubMed

Polfliet, Machteld M J; Fabriek, Babs O; Daniëls, Wouter P; Dijkstra, Christine D; van den Berg, Timo K

2006-01-01

The monoclonal antibody ED2 is widely used to define macrophages (mphi) in the rat. We have recently identified the ED2 antigen as the rat CD163 glycoprotein. CD163 is a member of the scavenger receptor cysteine-rich group B (SRCR-B) family and functions as a scavenger receptor for hemoglobin-haptoglobin complexes. Moreover, CD163 has also been indicated as a marker for alternatively activated mphi. In the current study, we identify rat CD163/ED2-antigen as a marker for mature tissue mphi. Rat CD163 is constitutively expressed on most subpopulations of mature tissue mphi, including splenic red pulp mphi, thymic cortical mphi, Kupffer cells in the liver, resident bone marrow mphi and central nervous system perivascular and meningeal mphi, but is apparently absent from monocytes. Rat CD163 expression can be promoted by glucocorticoids, and this can be further enhanced by IL4. Finally, engagement of rat CD163 on peritoneal mphi induces the production of pro-inflammatory mediators, including NO, IL-1beta, IL-6 and TNF-alpha. Collectively, our findings identify rat CD163 as a broadly expressed macrophage scavenger receptor that may play a role in the activation of mphi during hemolytic and/or inflammatory conditions.

CD44-positive cells are candidates for astrocyte precursor cells in developing mouse cerebellum.

PubMed

Cai, Na; Kurachi, Masashi; Shibasaki, Koji; Okano-Uchida, Takayuki; Ishizaki, Yasuki

2012-03-01

Neural stem cells are generally considered to be committed to becoming precursor cells before terminally differentiating into either neurons or glial cells during neural development. Neuronal and oligodendrocyte precursor cells have been identified in several areas in the murine central nervous system. The presence of astrocyte precursor cells (APCs) is not so well understood. The present study provides several lines of evidence that CD44-positive cells are APCs in the early postnatal mouse cerebellum. In developing mouse cerebellum, CD44-positive cells, mostly located in the white matter, were positive for the markers of the astrocyte lineage, but negative for the markers of mature astrocytes. CD44-positive cells were purified from postnatal cerebellum by fluorescence-activated cell sorting and characterized in vitro. In the absence of any signaling molecule, many cells died by apoptosis. The surviving cells gradually expressed glial fibrillary acidic protein, a marker for mature astrocytes, indicating that differentiation into mature astrocytes is the default program for these cells. The cells produced no neurospheres nor neurons nor oligodendrocytes under any condition examined, indicating these cells are not neural stem cells. Leukemia inhibitory factor greatly promoted astrocytic differentiation of CD44-positive cells, whereas bone morphogenetic protein 4 (BMP4) did not. Fibroblast growth factor-2 was a potent mitogen for these cells, but was insufficient for survival. BMP4 inhibited activation of caspase-3 and greatly promoted survival, suggesting a novel role for BMP4 in the control of development of astrocytes in cerebellum. We isolated and characterized only CD44 strongly positive large cells and discarded small and/or CD44 weakly positive cells in this study. Further studies are necessary to characterize these cells to help determine whether CD44 is a selective and specific marker for APCs in the developing mouse cerebellum. In conclusion, we succeeded in preparing APC candidates from developing mouse cerebellum, characterized them in vitro, and found that BMPs are survival factors for these cells.

Reviewing and Updating the Major Molecular Markers for Stem Cells

PubMed Central

Calloni, Raquel; Cordero, Elvira Alicia Aparicio; Henriques, João Antonio Pêgas

2013-01-01

Stem cells (SC) are able to self-renew and to differentiate into many types of committed cells, making SCs interesting for cellular therapy. However, the pool of SCs in vivo and in vitro consists of a mix of cells at several stages of differentiation, making it difficult to obtain a homogeneous population of SCs for research. Therefore, it is important to isolate and characterize unambiguous molecular markers that can be applied to SCs. Here, we review classical and new candidate molecular markers that have been established to show a molecular profile for human embryonic stem cells (hESCs), mesenchymal stem cells (MSCs), and hematopoietic stem cells (HSCs). The commonly cited markers for embryonic ESCs are Nanog, Oct-4, Sox-2, Rex-1, Dnmt3b, Lin-28, Tdgf1, FoxD3, Tert, Utf-1, Gal, Cx43, Gdf3, Gtcm1, Terf1, Terf2, Lefty A, and Lefty B. MSCs are primarily identified by the expression of CD13, CD29, CD44, CD49e, CD54, CD71, CD73, CD90, CD105, CD106, CD166, and HLA-ABC and lack CD14, CD31, CD34, CD45, CD62E, CD62L, CD62P, and HLA-DR expression. HSCs are mainly isolated based on the expression of CD34, but the combination of this marker with CD133 and CD90, together with a lack of CD38 and other lineage markers, provides the most homogeneous pool of SCs. Here, we present new and alternative markers for SCs, along with microRNA profiles, for these cells. PMID:23336433

The FGL2/fibroleukin prothrombinase is involved in alveolar macrophage activation in COPD through the MAPK pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Yanling; Xu, Sanpeng; Xiao, Fei

2010-05-28

Fibrinogen-like protein 2 (FGL2)/fibroleukin has been reported to play a vital role in the pathogenesis of some critical inflammatory diseases by possessing immunomodulatory activity through the mediation of 'immune coagulation' and the regulation of maturation and proliferation of immune cells. We observed upregulated FGL2 expression in alveolar macrophages from peripheral lungs of chronic obstructive pulmonary disease (COPD) patients and found a correlation between FGL2 expression and increased macrophage activation markers (CD11b and CD14). The role of FGL2 in the activation of macrophages was confirmed by the detection of significantly decreased macrophage activation marker (CD11b, CD11c, and CD71) expression as wellmore » as the inhibition of cell migration and inflammatory cytokine (IL-8 and MMP-9) production in an LPS-induced FGL2 knockdown human monocytic leukemia cell line (THP-1). Increased FGL2 expression co-localized with upregulated phosphorylated p38 mitogen-activated protein kinase (p38-MAPK) in the lung tissues from COPD patients. Moreover, FGL2 knockdown in THP-1 cells significantly downregulated LPS-induced phosphorylation of p38-MAPK while upregulating phosphorylation of c-Jun N-terminal kinase (JNK). Thus, we demonstrate that FGL2 plays an important role in macrophage activation in the lungs of COPD patients through MAPK pathway modulation.« less

Decreased non-MHC-restricted (CD56+) killer cell cytotoxicity after spaceflight

NASA Technical Reports Server (NTRS)

Mehta, S. K.; Kaur, I.; Grimm, E. A.; Smid, C.; Feeback, D. L.; Pierson, D. L.

2001-01-01

Cytotoxic activity of non-major histocompatibility complex-restricted (CD56+) (NMHC) killer cells and cell surface marker expression of peripheral blood mononuclear cells were determined before and after spaceflight. Ten astronauts (9 men, 1 woman) from two space shuttle missions (9- and 10-day duration) participated in the study. Blood samples were collected 10 days before launch, within 3 h after landing, and 3 days after landing. All peripheral blood mononuclear cell preparations were cryopreserved and analyzed simultaneously in a 4-h cytotoxicity (51)Cr release assay using K562 target cells. NMHC killer cell lytic activity was normalized per 1,000 CD56+ cells. When all 10 subjects were considered as one study group, NMHC killer cell numbers did not change significantly during the three sampling periods, but at landing lytic activity had decreased by approximately 40% (P < 0.05) from preflight values. Nine of ten astronauts had decreased lytic activity immediately after flight. NMHC killer cell cytotoxicity of only three astronauts returned toward preflight values by 3 days after landing. Consistent with decreased NMHC killer cell cytotoxicity, urinary cortisol significantly increased after landing compared with preflight levels. Plasma cortisol and ACTH levels at landing were not significantly different from preflight values. No correlation of changes in NMHC killer cell function or hormone levels with factors such as age, gender, mission, or spaceflight experience was found. After landing, expression of the major lymphocyte surface markers (CD3, CD4, CD8, CD14, CD16, CD56), as determined by flow cytometric analysis, did not show any consistent changes from measurements made before flight.

Mass Cytometry Analysis Reveals the Landscape and Dynamics of CD32a+ CD4+ T Cells From Early HIV Infection to Effective cART.

PubMed

Coindre, Sixtine; Tchitchek, Nicolas; Alaoui, Lamine; Vaslin, Bruno; Bourgeois, Christine; Goujard, Cecile; Avettand-Fenoel, Veronique; Lecuroux, Camille; Bruhns, Pierre; Le Grand, Roger; Beignon, Anne-Sophie; Lambotte, Olivier; Favier, Benoit

2018-01-01

CD32a has been proposed as a specific marker of latently HIV-infected CD4 + T cells. However, CD32a was recently found to be expressed on CD4 + T cells of healthy donors, leading to controversy on the relevance of this marker in HIV persistence. Here, we used mass cytometry to characterize the landscape and variation in the abundance of CD32a + CD4 + T cells during HIV infection. To this end, we analyzed CD32a + CD4 + T cells in primary HIV infection before and after effective combination antiretroviral therapy (cART) and in healthy donors. We found that CD32a + CD4 + T cells include heterogeneous subsets that are differentially affected by HIV infection. Our analysis revealed that naive ( N ), central memory ( CM ), and effector/memory ( Eff/Mem ) CD32a + CD4 + T-cell clusters that co-express LILRA2- and CD64-activating receptors were more abundant in primary HIV infection and cART stages. Conversely, LILRA2 - CD32a + CD4 + T-cell clusters of either the T N , T CM , or T Eff/Mem phenotype were more abundant in healthy individuals. Finally, an activated CD32a + CD4 + T Eff/Mem cell cluster co-expressing LILRA2, CD57, and NKG2C was more abundant in all HIV stages, particularly during primary HIV infection. Overall, our data show that multiple abundance modifications of CD32a + CD4 + T-cell subsets occur in the early phase of HIV infection, and some of which are conserved after effective cART. Our study brings a better comprehension of the relationship between CD32a expression and CD4 + T cells during HIV infection.

Liver-specific gene expression in cultured human hematopoietic stem cells.

PubMed

Fiegel, Henning C; Lioznov, Michael V; Cortes-Dericks, Lourdes; Lange, Claudia; Kluth, Dietrich; Fehse, Boris; Zander, Axel R

2003-01-01

Hematopoietic and hepatic stem cells share characteristic markers such as CD34, c-kit, and Thy1. Based on the recent observations that hepatocytes may originate from bone marrow, we investigated the potential of CD34(+) bone marrow cells to differentiate into hepatocytic cells in vitro. CD34(+) and CD34(-) human bone marrow cells were separated by magnetic cell sorting. Cells were cultured on a collagen matrix in a defined medium containing hepatocyte growth factor. Cell count and size were measured by flow cytometry, and reverse transcription polymerase chain reaction was carried out for the liver-specific markers CK-19 and albumin. During cell culture, CD34(+) cells showed an increasing cell number and proliferative activity as assessed by Ki-67 staining. Under the specified culture conditions, CD34(+) cells expressed albumin RNA and CK-19 RNA after 28 days, whereas CD34(-) cells did not show liver-specific gene expression. The results indicate that CD34(+) adult human bone marrow stem cells can differentiate into hepatocytic cells in vitro.

The Biological Function and Clinical Utilization of CD147 in Human Diseases: A Review of the Current Scientific Literature

PubMed Central

Xiong, Lijuan; Edwards, Carl K.; Zhou, Lijun

2014-01-01

CD147 or EMMPRIN is a member of the immunoglobulin superfamily in humans. It is widely expressed in human tumors and plays a central role in the progression of many cancers by stimulating the secretion of matrix metalloproteinases (MMPs) and cytokines. CD147 regulates cell proliferation, apoptosis, and tumor cell migration, metastasis and differentiation, especially under hypoxic conditions. CD147 is also important to many organ systems. This review will provide a detailed overview of the discovery, characterization, molecular structure, diverse biological functions and regulatory mechanisms of CD147 in human physiological and pathological processes. In particular, recent studies have demonstrated the potential application of CD147 not only as a phenotypic marker of activated regulatory T cells but also as a potential diagnostic marker for early-stage disease. Moreover, CD147 is recognized as an effective therapeutic target for hepatocellular carcinoma (HCC) and other cancers, and exciting clinical progress has been made in HCC treatment using CD147-directed monoclonal antibodies. PMID:25268615

Reproducible diagnosis of chronic lymphocytic leukemia by flow cytometry: An European Research Initiative on CLL (ERIC) & European Society for Clinical Cell Analysis (ESCCA) Harmonisation project

PubMed Central

Kreuzer, Karl‐Anton; Soosapilla, Asha; Spacek, Martin; Stehlikova, Olga; Gambell, Peter; McIver‐Brown, Neil; Villamor, Neus; Psarra, Katherina; Arroz, Maria; Milani, Raffaella; de la Serna, Javier; Cedena, M. Teresa; Jaksic, Ozren; Nomdedeu, Josep; Moreno, Carol; Rigolin, Gian Matteo; Cuneo, Antonio; Johansen, Preben; Johnsen, Hans E.; Rosenquist, Richard; Niemann, Carsten Utoft; Kern, Wolfgang; Westerman, David; Trneny, Marek; Mulligan, Stephen; Doubek, Michael; Pospisilova, Sarka; Hillmen, Peter; Oscier, David; Hallek, Michael; Ghia, Paolo; Montserrat, Emili

2018-01-01

The diagnostic criteria for CLL rely on morphology and immunophenotype. Current approaches have limitations affecting reproducibility and there is no consensus on the role of new markers. The aim of this project was to identify reproducible criteria and consensus on markers recommended for the diagnosis of CLL. ERIC/ESCCA members classified 14 of 35 potential markers as “required” or “recommended” for CLL diagnosis, consensus being defined as >75% and >50% agreement, respectively. An approach to validate “required” markers using normal peripheral blood was developed. Responses were received from 150 participants with a diagnostic workload >20 CLL cases per week in 23/150 (15%), 5–20 in 82/150 (55%), and <5 cases per week in 45/150 (30%). The consensus for “required” diagnostic markers included: CD19, CD5, CD20, CD23, Kappa, and Lambda. “Recommended” markers potentially useful for differential diagnosis were: CD43, CD79b, CD81, CD200, CD10, and ROR1. Reproducible criteria for component reagents were assessed retrospectively in 14,643 cases from 13 different centers and showed >97% concordance with current approaches. A pilot study to validate staining quality was completed in 11 centers. Markers considered as “required” for the diagnosis of CLL by the participants in this study (CD19, CD5, CD20, CD23, Kappa, and Lambda) are consistent with current diagnostic criteria and practice. Importantly, a reproducible approach to validate and apply these markers in individual laboratories has been identified. Finally, a consensus “recommended” panel of markers to refine diagnosis in borderline cases (CD43, CD79b, CD81, CD200, CD10, and ROR1) has been defined and will be prospectively evaluated. © 2017 International Clinical Cytometry Society PMID:29024461

Reproducible diagnosis of chronic lymphocytic leukemia by flow cytometry: An European Research Initiative on CLL (ERIC) & European Society for Clinical Cell Analysis (ESCCA) Harmonisation project.

PubMed

Rawstron, Andy C; Kreuzer, Karl-Anton; Soosapilla, Asha; Spacek, Martin; Stehlikova, Olga; Gambell, Peter; McIver-Brown, Neil; Villamor, Neus; Psarra, Katherina; Arroz, Maria; Milani, Raffaella; de la Serna, Javier; Cedena, M Teresa; Jaksic, Ozren; Nomdedeu, Josep; Moreno, Carol; Rigolin, Gian Matteo; Cuneo, Antonio; Johansen, Preben; Johnsen, Hans E; Rosenquist, Richard; Niemann, Carsten Utoft; Kern, Wolfgang; Westerman, David; Trneny, Marek; Mulligan, Stephen; Doubek, Michael; Pospisilova, Sarka; Hillmen, Peter; Oscier, David; Hallek, Michael; Ghia, Paolo; Montserrat, Emili

2018-01-01

The diagnostic criteria for CLL rely on morphology and immunophenotype. Current approaches have limitations affecting reproducibility and there is no consensus on the role of new markers. The aim of this project was to identify reproducible criteria and consensus on markers recommended for the diagnosis of CLL. ERIC/ESCCA members classified 14 of 35 potential markers as "required" or "recommended" for CLL diagnosis, consensus being defined as >75% and >50% agreement, respectively. An approach to validate "required" markers using normal peripheral blood was developed. Responses were received from 150 participants with a diagnostic workload >20 CLL cases per week in 23/150 (15%), 5-20 in 82/150 (55%), and <5 cases per week in 45/150 (30%). The consensus for "required" diagnostic markers included: CD19, CD5, CD20, CD23, Kappa, and Lambda. "Recommended" markers potentially useful for differential diagnosis were: CD43, CD79b, CD81, CD200, CD10, and ROR1. Reproducible criteria for component reagents were assessed retrospectively in 14,643 cases from 13 different centers and showed >97% concordance with current approaches. A pilot study to validate staining quality was completed in 11 centers. Markers considered as "required" for the diagnosis of CLL by the participants in this study (CD19, CD5, CD20, CD23, Kappa, and Lambda) are consistent with current diagnostic criteria and practice. Importantly, a reproducible approach to validate and apply these markers in individual laboratories has been identified. Finally, a consensus "recommended" panel of markers to refine diagnosis in borderline cases (CD43, CD79b, CD81, CD200, CD10, and ROR1) has been defined and will be prospectively evaluated. © 2017 International Clinical Cytometry Society. © 2017 The Authors. Cytometry Part B: Clinical Cytometry published by Wiley Periodicals, Inc. on behalf of International Clinical Cytometry Society.

Comprehensive Astronaut Immune Assessment Following a Short-Duration Space Flight

NASA Technical Reports Server (NTRS)

Crucian, Brian; Stowe, Raymond; Yetman, Deborah; Pierson, Duane; Sams, Clarence

2006-01-01

Immune system dysregulation has been demonstrated to occur during spaceflight and has the potential to cause serious health risks to crewmembers participating in exploration class missions. As a part of an ongoing NASA flight experiment assessing viral immunity (DSO-500), a generalized immune assessment was performed on 3 crewmembers who participated in the recent STS-114 Space Shuttle mission. The following assays were performed: (1) comprehensive immunophenotype analysis; (2) T cell function/intracellular cytokine profiles; (4) secreted Th1/Th2 cytokine profiles via cytometric bead array. Immunophenotype analysis included a leukocyte differential, lymphocyte subsets, T cell subsets, cytotoxic/effector CD8+ T cells, memory/naive T cell subsets and constitutively activated T cells. Study timepoints were L-180, L-65, L-10, R+0, R+3 and R+14. Detailed data are presented in the poster text. As expected from a limited number of human subjects, data tended to vary with respect to most parameters. Specific post-flight alterations were as follows (subject number in parentheses): Granulocytosis (2/3), reduced NK cells (3/3), elevated CD4/CD8 ratio (3/3), general CD8+ phenotype shift to a less differentiated phenotype (3/3), elevated levels of memory CD4+ T cells (3/3), loss of L-selectin on T cell subsets (3/3), increased levels of activated T cells (2/3), reduced IL-2 producing T cell subsets (3/3), levels of IFNg producing T cells were unchanged. CD8+ T cell expression of the CD69 activation markers following whole blood stimulation with SEA+SEB were dramatically reduced postflight (3/3), whereas other T cell function assessments were largely unchanged. Cytometric bead array assessment of secreted T cell cytokines was performed, following whole blood stimulation with either CD3/CD28 antibodies or PMA+ionomycin for 48 hours. Specific cytokines assessed were IFNg, TNFa, IL-2, IL-4, IL-5, IL-10. Following CD3/CD28 stimulation, all three crewmembers had a mission-associated reduction in the levels of secreted IFNg. One crewmember had a post-flight inversion in the IFNg/IL-10 ratio postflight, which trended back to baseline by R+14. Detailed cytokine data are presented in the poster text. This testing regimen was designed to correlate immunophenotype changes (thought to correspond to specific in-vivo immune responses or pathogenesis), against altered leukocyte function and cytokine profiles. In-flight studies are required to determine if post-flight alterations are reflective of the in-flight condition, or are a response to landing and readaptation.

Post-Spaceflight (STS-135) Mouse Splenocytes Demonstrate Altered Activation Properties and Surface Molecule Expression

PubMed Central

Crucian, Brian; Sams, Clarence

2015-01-01

Alterations in immune function have been documented during or post-spaceflight and in ground based models of microgravity. Identification of immune parameters that are dysregulated during spaceflight is an important step in mitigating crew health risks during deep space missions. The in vitro analysis of leukocyte activity post-spaceflight in both human and animal species is primarily focused on lymphocytic function. This report completes a broader spectrum analysis of mouse lymphocyte and monocyte changes post 13 days orbital flight (mission STS-135). Analysis includes an examination in surface markers for cell activation, and antigen presentation and co-stimulatory molecules. Cytokine production was measured after stimulation with T-cell mitogen or TLR-2, TLR-4, or TLR-5 agonists. Splenocyte surface marker analysis immediate post-spaceflight and after in vitro culture demonstrated unique changes in phenotypic populations between the flight mice and matched treatment ground controls. Post-spaceflight splenocytes (flight splenocytes) had lower expression intensity of CD4+CD25+ and CD8+CD25+ cells, lower percentage of CD11c+MHC II+ cells, and higher percentage of CD11c+MHC I+ populations compared to ground controls. The flight splenocytes demonstrated an increase in phagocytic activity. Stimulation with ConA led to decrease in CD4+ population but increased CD4+CD25+ cells compared to ground controls. Culturing with TLR agonists led to a decrease in CD11c+ population in splenocytes isolated from flight mice compared to ground controls. Consequently, flight splenocytes with or without TLR-agonist stimulation showed a decrease in CD11c+MHC I+, CD11c+MHC II+, and CD11c+CD86+ cells compared to ground controls. Production of IFN-γ was decreased and IL-2 was increased from ConA stimulated flight splenocytes. This study demonstrated that expression of surface molecules can be affected by conditions of spaceflight and impaired responsiveness persists under culture conditions in vitro. PMID:25970640

Does the liposuction method influence the phenotypic characteristic of human adipose-derived stem cells?

PubMed

Bajek, Anna; Gurtowska, Natalia; Gackowska, Lidia; Kubiszewska, Izabela; Bodnar, Magdalena; Marszałek, Andrzej; Januszewski, Rafał; Michalkiewicz, Jacek; Drewa, Tomasz

2015-05-14

Adipose-derived stem cells (ASCs) possess a high differentiation and proliferation potential. However, the phenotypic characterization of ASCs is still difficult. Until now, there is no extensive analysis of ASCs markers depending on different liposuction methods. Therefore, the aim of the present study was to analyse 242 surface markers and determine the differences in the phenotypic pattern between ASCs obtained during mechanical and ultrasound-assisted liposuction. ASCs were isolated from healthy donors, due to mechanical and ultrasound-assisted liposuction and cultured in standard medium to the second passage. Differentiation potential and markers expression was evaluated to confirm the mesenchymal nature of cells. Then, the BD LyoplateTM Human Cell Surface Marker Screening Panel was used. Results shown that both population of ASCs are characterized by high expression of markers specific for ASCs: cluster of differentiation (CD)9, CD10, CD34, CD44, CD49d, CD54, CD55, CD59, CD71 and low expression of CD11a, CD11c and CD144. Moreover, we have noticed significant differences in antigen expression in 58 markers from the 242 studied. Presented study shows for the first time that different liposuction methods are not a significant factor which can influence the expression of human ASCs surface markers. © 2015 The Authors.

Cellular Profile and Expression of Immunologic Markers in Chronic Apical Periodontitis from HIV-infected Patients Undergoing Highly Active Antiretroviral Therapy.

PubMed

Gama, Túlio Gustavo Veiga; Pires, Fabio Ramoa; Armada, Luciana; Gonçalves, Lucio Souza

2016-06-01

This study tested the hypothesis that the inflammatory cell profile (CD3-, CD4-, CD8-, CD20-, and CD68-positive cells) and the expression of immunologic markers (tumor necrosis factor α, interferon-γ, interleukin-6, and interleukin-18) in chronic apical periodontitis are the same between non-HIV-infected patients and HIV-infected patients undergoing highly active antiretroviral therapy (HAART). Thirty-four surgically excised chronic apical periodontitis lesions were sampled from 34 patients (17 HIV-infected and 17 non-HIV-infected). The lesions were extracted from teeth with no previous endodontic treatment. All HIV-infected patients were undergoing HAART. The specimens were submitted to histopathologic and immunohistochemical analyses by using an optical microscope. Immunoexpression was graded into 2 levels, focal to weak and moderate to strong. The χ(2), Fisher exact, and Mann-Whitney tests were used to analyze all significant differences between groups. Periapical cysts represented 70.6% and 52.9% of the lesions in the HIV-infected and non-HIV-infected groups, respectively; however, no statistically significant difference was observed (P = .481). There were no statistically significant differences between groups for the inflammatory cell profile and for any of the immunologic markers (P > .05). There are no statistically significant differences of the cellular profile and expression of immunologic markers in chronic apical periodontitis between non-HIV-infected patients and HIV-infected patients undergoing HAART. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Posttransplant sCD30 as a biomarker to predict kidney graft outcome.

PubMed

Süsal, Caner; Opelz, Gerhard

2012-09-08

In current clinical praxis, monitoring of immunosuppressive agents in organ transplantation is restricted to measurement of drug blood levels and does not consider the drug's variable effect on the individual patient's immune system. Establishment of biological markers that measure the biological effect of immunosuppressive drugs is desirable and would enable the identification of patients who are at risk of developing rejection, or patients who are suitable for minimization or weaning of immunosuppressive therapy. Several studies demonstrated that the technically simple posttransplant measurement in serum of the T cell activation marker soluble CD30 (sCD30) allows prediction of subsequent graft loss in kidney transplant recipients. sCD30 is a relatively large molecule and therefore an attractive biological marker which is resistant to repeated thawing cycles and temperature differences and easily determined using commercial ELISA. Whether sCD30-based prospective adjustment of immunosuppressive therapy can prevent irreversible graft damage and improve long-term graft outcome awaits evaluation in randomized controlled trials. Copyright © 2011 Elsevier B.V. All rights reserved.

Sequential CD34 cell fractionation by magnetophoresis in a magnetic dipole flow sorter.

PubMed

Schneider, Thomas; Karl, Stephan; Moore, Lee R; Chalmers, Jeffrey J; Williams, P Stephen; Zborowski, Maciej

2010-01-01

Cell separation and fractionation based on fluorescent and magnetic labeling procedures are common tools in contemporary research. These techniques rely on binding of fluorophores or magnetic particles conjugated to antibodies to target cells. Cell surface marker expression levels within cell populations vary with progression through the cell cycle. In an earlier work we showed the reproducible magnetic fractionation (single pass) of the Jurkat cell line based on the population distribution of CD45 surface marker expression. Here we present a study on magnetic fractionation of a stem and progenitor cell (SPC) population using the established acute myelogenous leukemia cell line KG-1a as a cell model. The cells express a CD34 cell surface marker associated with the hematopoietic progenitor cell activity and the progenitor cell lineage commitment. The CD34 expression level is approximately an order of magnitude lower than that of the CD45 marker, which required further improvements of the magnetic fractionation apparatus. The cells were immunomagnetically labeled using a sandwich of anti-CD34 antibody-phycoerythrin (PE) conjugate and anti-PE magnetic nanobead and fractionated into eight components using a continuous flow dipole magnetophoresis apparatus. The CD34 marker expression distribution between sorted fractions was measured by quantitative PE flow cytometry (using QuantiBRITE PE calibration beads), and it was shown to be correlated with the cell magnetophoretic mobility distribution. A flow outlet addressing scheme based on the concept of the transport lamina thickness was used to control cell distribution between the eight outlet ports. The fractional cell distributions showed good agreement with numerical simulations of the fractionation based on the cell magnetophoretic mobility distribution in the unsorted sample.

Endothelial microparticles and vascular parameters in subjects with and without arterial hypertension and coronary artery disease.

PubMed

Sansone, Roberto; Baaken, Maximilian; Horn, Patrick; Schuler, Dominik; Westenfeld, Ralf; Amabile, Nicolas; Kelm, Malte; Heiss, Christian

2018-08-01

Endothelial microparticles (EMPs) are markers of endothelial injury and activation. The role of EMPs in arterial hypertension is not well understood and EMPs are increased both in arterial hypertension and coronary artery disease (CAD). The data presented here show EMPs as defined by CD31 + /41 - , CD62e + , and CD144 + surface markers and vascular hemodynamic parameters including office and central blood pressure, heart rate, aortic augmentation index, pulse wave velocity, flow-mediated dilation, nitroglycerin-mediated dilation, brachial artery diameter, hyperemic wall shear stress, and laser Doppler perfusion of the cutaneous microcirculation of normotensives and hypertensives with and without CAD.

Characterization of cancer stem cell properties of CD24 and CD26-positive human malignant mesothelioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yamazaki, Hiroto; Naito, Motohiko; Ghani, Farhana Ishrat

2012-03-16

Highlights: Black-Right-Pointing-Pointer We focused on CD24 and CD26 for further analysis of CSC properties in MM. Black-Right-Pointing-Pointer Their expressions were correlated with chemoresistance, cell growth, and invasion. Black-Right-Pointing-Pointer Their expressions were also correlated with several cancer related genes. Black-Right-Pointing-Pointer The expression of each marker was correlated with different CSC property in Meso1. Black-Right-Pointing-Pointer Phosphorylation of ERK by EGF was regulated by expression of CD26, but not CD24. -- Abstract: Malignant mesothelioma (MM) is an asbestos-related malignancy characterized by rapid growth and poor prognosis. In our previous study, we have demonstrated that several cancer stem cell (CSC) markers correlated with CSCmore » properties in MM cells. Among these markers, we focused on two: CD24, the common CSC marker, and CD26, the additional CSC marker. We further analyzed the CSC properties of CD24 and CD26-positve MM cells. We established RNAi-knockdown cells and found that these markers were significantly correlated with chemoresistance, proliferation, and invasion potentials in vitro. Interestingly, while Meso-1 cells expressed both CD24 and CD26, the presence of each of these two markers was correlated with different CSC property. In addition, downstream signaling of these markers was explored by microarray analysis, which revealed that their expressions were correlated with several cancer-related genes. Furthermore, phosphorylation of ERK by EGF stimulation was significantly affected by the expression of CD26, but not CD24. These results suggest that CD24 and CD26 differentially regulate the CSC potentials of MM and could be promising targets for CSC-oriented therapy.« less

Efficacy of T Regulatory Cells, Th17 Cells and the Associated Markers in Monitoring Tuberculosis Treatment Response

PubMed Central

Agrawal, Sonali; Parkash, Om; Palaniappan, Alangudi Natarajan; Bhatia, Ashok Kumar; Kumar, Santosh; Chauhan, Devendra Singh; Madhan Kumar, M.

2018-01-01

Treatment monitoring is an essential aspect for tuberculosis (TB) disease management. Sputum smear microscopy is the only available tool for monitoring, but it suffers from demerits. Therefore, we sought to evaluate markers and cellular subsets of T regulatory (Treg) cells and T helper (Th) 17 cells in pulmonary TB patients (PTB) for TB treatment monitoring. Peripheral blood mononuclear cells (PBMCs) were stimulated in vitro (with purified protein derivative (PPD)) overnight which was followed by a polychromatic flow cytometry approach to study Treg and Th17 markers and cellular subsets in PTB (n = 12) undergoing antituberculous treatment (ATT). The baseline levels of these markers and cellular subsets were evaluated in normal healthy subjects (NHS). We observed a significant decrease in the expression of CD25 (p<0.01) marker and percentage of T-cell subsets like CD4+CD25+ (p<0.001) and CD4+CD25+CD39+ (p<0.05) at the end of intensive phase (IP) as well as in the continuation phase (CP) of ATT. A decrease in CD25 marker expression and percentage of CD4+CD25+ T cell subset showed a positive correlation to sputum conversion both in high and low sputum positive PTB. In eight PTB with cavitary lesions, only CD4+CD25+FoxP3 Treg subset manifested a significant decrease at the end of CP. Thus, results of this study show that CD25 marker and CD4+CD25+ T cells can serve as better markers for monitoring TB treatment efficacy. The Treg subset CD4+CD25+FoxP3 may be useful for prediction of favorable response in PTB with extensive lung lesions. However, these findings have to be evaluated in a larger patient cohort. PMID:29472922

Differentiation and activation of equine monocyte-derived dendritic cells are not correlated with CD206 or CD83 expression

PubMed Central

Moyo, Nathifa A; Marchi, Emanuele; Steinbach, Falko

2013-01-01

Dendritic cells (DC) are the main immune mediators inducing primary immune responses. DC generated from monocytes (MoDC) are a model system to study the biology of DC in vitro, as they represent inflammatory DC in vivo. Previous studies on the generation of MoDC in horses indicated that there was no distinct difference between immature and mature DC and that the expression profile was distinctly different from humans, where CD206 is expressed on immature MoDC whereas CD83 is expressed on mature MoDC. Here we describe the kinetics of equine MoDC differentiation and activation, analysing both phenotypic and functional characteristics. Blood monocytes were first differentiated with equine granulocyte–macrophage colony-stimulating factor and interleukin-4 generating immature DC (iMoDC). These cells were further activated with a cocktail of cytokines including interferon-γ) but not CD40 ligand to obtain mature DC (mMoDC). To determine the expression of a broad range of markers for which no monoclonal antibodies were available to analyse the protein expression, microarray and quantitative PCR analysis were performed to carry out gene expression analysis. This study demonstrates that equine iMoDC and mMoDC can be distinguished both phenotypically and functionally but the expression pattern of some markers including CD206 and CD83 is dissimilar to the human system. PMID:23461413

Soluble CD163, a marker of Kupffer cell activation, is related to portal hypertension in patients with liver cirrhosis.

PubMed

Grønbaek, H; Sandahl, T D; Mortensen, C; Vilstrup, H; Møller, H J; Møller, S

2012-07-01

Activation of Kupffer cells may be involved in the pathogenesis of portal hypertension by release of vasoconstrictive substances and fibrosis due to co-activation of hepatic stellate cells. To study soluble plasma (s) CD163, a specific marker of activated macrophages, as a biomarker for portal hypertension in patients with liver cirrhosis. We measured sCD163 concentration and the hepatic venous pressure gradient (HVPG) by liver vein catheterisation in 81 cirrhosis patients (Child-Pugh CP-A: n = 26, CP-B: n = 29, CP-C: n = 26) and 22 healthy subjects. We also measured their cardiac output (CO), cardiac index and systemic vascular resistance (SVR). Liver status was examined by Child-Pugh and MELD-score. In cirrhosis, sCD163 concentration was nearly three times higher than in controls (4.7 ± 2.5 vs. 1.6 ± 0.5 mg/L, P < 0.001). sCD163 was also higher, as measured in steps by CP-score (P < 0.001). The HVPG rose steeply to an asymptote of 22 mmHg with sCD163 up to about 5 mg/L and not to higher values with higher sCD163. In a multivariate analysis, sCD163 was the only independent predictor of the HVPG but did not predict any of the systemic circulatory findings. sCD163 > 3.95 mg/L (upper normal limit) predicted HVPG ≥ 10 mmHg with a positive predictive value of 0.99. Circulating sCD163 originating from activated Kupffer cells is increased in cirrhosis with increasing Child-Pugh score and with increasing HVPG, and it is an independent predictor for HVPG. These findings support a primary role of macrophage activation in portal hypertension, and may indicate a target for biological intervention. © 2012 Blackwell Publishing Ltd.

Thymic pathological examination of non-thymomatous myasthenia gravis patients: A pilot study for prediction of outcome

PubMed Central

Peimani, Zeinab; Banihashemi, Mohammad Amin; Namazi, Niloofar; Monabati, Ahmad; Mojallal, Mehra; Borhani-Haghighi, Afshin

2014-01-01

Background Myasthenia gravis (MG) is an autoimmune disorder characterized by weakness and fatigability of skeletal muscles. The aim of this study was to determine if pathological characteristics in non-thymomatous patients of MG would correlate with prognosis in a three year follow up. Methods Patients who had had their thymectomy at least three years prior to the study were selected from three hospitals and were followed for 3 years. Prognosis was assessed via a devised prognostic scoring system. A pathological exam of the specimen from the thymus was done using the following immunohistochemical markers: Bcl2, CD 3, CD 4, CD 5, CD 7, CD 10, CD 20cy, CD 23, CD 43, and Ki67. Results Fifteen patients fulfilled the inclusion criteria and had a complete follow-up. This included 3 males and 12 females with a mean age of 36.6 years at the start of the study. The dominant cell population was T lymphocytes. All T cells expressed CD 3, CD 43, CD 5, and Bcl-2. In 2 patients, CD 10 marker was positive in T cells. B cells were negative for activation marker CD 23, except for germinal center dendritic cells. Due to the limited number of patients in the study, the power of the study would not allow for an analysis to assess correlation between histopathological data and prognosis. Conclusion This pilot study was an attempt to discover any prognostic indices from the histopathological examination of the resected thymic tissue in the patients with myasthenia gravis. PMID:24800046

LAG-3 Represents a Marker of CD4+ T Cells with Regulatory Activity in Patients with Bone Fracture.

PubMed

Wang, Jun; Ti, Yunfan; Wang, Yicun; Guo, Guodong; Jiang, Hui; Chang, Menghan; Qian, Hongbo; Zhao, Jianning; Sun, Guojing

2018-04-19

The lymphocyte activation gene 3 (LAG-3) is a CD4 homolog with binding affinity to MHC class II molecules. It is thought that LAG-3 exerts a bimodal function, such that co-ligation of LAG-3 and CD3 could deliver an inhibitory signal in conventional T cells, whereas, on regulatory T cells, LAG-3 expression could promote their inhibitory function. In this study, we investigated the role of LAG-3 expression on CD4 + T cells in patients with long bone fracture. We found that LAG-3 + cells represented approximately 13% of peripheral blood CD4 + T cells on average. Compared to LAG-3 - CD4 + T cells, LAG-3 + CD4 + T cells presented significantly higher Foxp3 and CTLA-4 expression. Directly ex vivo or with TCR stimulation, LAG-3 + CD4 + T cells expressed significantly higher levels of IL-10 and TGF-β than LAG-3 - CD4 + T cells. Interestingly, blocking the LAG-3-MHC class II interaction actually increased the IL-10 expression by LAG-3 + CD4 + T cells. The frequency of LAG-3 + CD4 + T cell was positively correlated with restoration of healthy bone function in long bone fracture patients. These results together suggested that LAG-3 is a marker of CD4 + T cells with regulatory function; at the same time, LAG-3 might have limited the full suppressive potential of Treg cells.

CD147-mediated chemotaxis of CD4+CD161+ T cells may contribute to local inflammation in rheumatoid arthritis.

PubMed

Lv, Minghua; Miao, Jinlin; Zhao, Peng; Luo, Xing; Han, Qing; Wu, Zhenbiao; Zhang, Kui; Zhu, Ping

2018-01-01

CD161 is used as a surrogate marker for Th17 cells, which are implicated in the pathogenesis of rheumatoid arthritis (RA). In this study, we evaluated the percentage, clinical significance, and CD98 and CD147 expression of CD4 + CD161 + T cells. The potential role of CD147 and CD98 in cyclophilin A-induced chemotaxis of CD4 + CD161 + T cells was analyzed. Thirty-seven RA patients, 15 paired synovial fluid (SF) of RA, and 22 healthy controls were recruited. The cell populations and surface expression of CD98 and CD147 were analyzed by flow cytometry. Spearman's rank correlation coefficient and multiple linear regression were applied to calculate the correlations. Chemotaxis assay was used to investigate CD4 + CD161 + T cell migration. We found that the percentage of CD4 + CD161 + T cells and their expression of CD147 and CD98 in SF were higher than in the peripheral blood of RA patients. Percentage of SF CD4 + CD161 + T cells was positively correlated with 28-Joint Disease Activity Score (DAS28). CD147 monoclonal antibody (HAb18) attenuated the chemotactic ability of CD4 + CD161 + T cells. An increased CD4 + CD161 + T cell percentage and expression of CD147 and CD98 were shown in RA SF. Percentage of SF CD4 + CD161 + T cells can be used as a predictive marker of disease activity in RA. CD147 block significantly decreased the chemotactic index of CD4 + CD161 + cells induced by cyclophilin A (CypA). These results imply that the accumulation of CD4 + CD161 + T cells in SF and their high expression of CD147 may be associated with CypA-mediated chemotaxis and contribute to local inflammation in RA.

Lymphocyte Redox Imbalance and Reduced Proliferation after a Single Session of High Intensity Interval Exercise.

PubMed

Tossige-Gomes, Rosalina; Costa, Karine Beatriz; Ottone, Vinícius de Oliveira; Magalhães, Flávio de Castro; Amorim, Fabiano Trigueiro; Rocha-Vieira, Etel

2016-01-01

This study investigated whether an acute session of high-intensity interval training (HIIT) is sufficient to alter lymphocyte function and redox status. Sixteen young healthy men underwent a HIIT session on a cycloergometer, consisting of eight bouts of 1 min at 90-100% of peak power, with 75 seconds of active recovery at 30 W between bouts. Venous blood was collected before, immediately after, and 30 minutes after the HIIT session. In response to Staphylococcus aureus superantigen B (SEB) stimulation, lymphocyte proliferation decreased and the IL-2 concentration increased after the HIIT session. However, the HIIT session had no effect on lymphocyte proliferation or IL-2 response to phytohemagglutinin stimulation. The HIIT session also induced lymphocyte redox imbalance, characterized by an increase in the concentration of thiobarbituric acid reactive substances and a decrease in the activity of the antioxidant enzyme catalase. Lymphocyte viability was not affected by the HIIT session. The frequencies of CD25+ and CD69+ T helper and B lymphocytes in response to superantigen stimulation were lower after exercise, suggesting that superantigen-induced lymphocyte activation was reduced by HIIT. However, HIIT also led to a reduction in the frequency of CD4+ and CD19+ cells, so the frequencies of CD25+ and CD69+ cells within the CD4 and CD19 cell populations were not affected by HIIT. These data indicate that the reduced lymphocyte proliferation observed after HIIT is not due to reduced early lymphocyte activation by superantigen. Our findings show that an acute HIIT session promotes lymphocyte redox imbalance and reduces lymphocyte proliferation in response to superantigenic, but not to mitogenic stimulation. This observation cannot be explained by alteration of the early lymphocyte activation response to superantigen. The manner in which lymphocyte function modulation by an acute HIIT session can affect individual immunity and susceptibility to infection is important and requires further investigation.

Lymphocyte Redox Imbalance and Reduced Proliferation after a Single Session of High Intensity Interval Exercise

PubMed Central

Tossige-Gomes, Rosalina; Costa, Karine Beatriz; Ottone, Vinícius de Oliveira; Magalhães, Flávio de Castro; Amorim, Fabiano Trigueiro; Rocha-Vieira, Etel

2016-01-01

This study investigated whether an acute session of high-intensity interval training (HIIT) is sufficient to alter lymphocyte function and redox status. Sixteen young healthy men underwent a HIIT session on a cycloergometer, consisting of eight bouts of 1 min at 90–100% of peak power, with 75 seconds of active recovery at 30 W between bouts. Venous blood was collected before, immediately after, and 30 minutes after the HIIT session. In response to Staphylococcus aureus superantigen B (SEB) stimulation, lymphocyte proliferation decreased and the IL-2 concentration increased after the HIIT session. However, the HIIT session had no effect on lymphocyte proliferation or IL-2 response to phytohemagglutinin stimulation. The HIIT session also induced lymphocyte redox imbalance, characterized by an increase in the concentration of thiobarbituric acid reactive substances and a decrease in the activity of the antioxidant enzyme catalase. Lymphocyte viability was not affected by the HIIT session. The frequencies of CD25+ and CD69+ T helper and B lymphocytes in response to superantigen stimulation were lower after exercise, suggesting that superantigen-induced lymphocyte activation was reduced by HIIT. However, HIIT also led to a reduction in the frequency of CD4+ and CD19+ cells, so the frequencies of CD25+ and CD69+ cells within the CD4 and CD19 cell populations were not affected by HIIT. These data indicate that the reduced lymphocyte proliferation observed after HIIT is not due to reduced early lymphocyte activation by superantigen. Our findings show that an acute HIIT session promotes lymphocyte redox imbalance and reduces lymphocyte proliferation in response to superantigenic, but not to mitogenic stimulation. This observation cannot be explained by alteration of the early lymphocyte activation response to superantigen. The manner in which lymphocyte function modulation by an acute HIIT session can affect individual immunity and susceptibility to infection is important and requires further investigation. PMID:27096389

Beneficial effect of pistachio consumption on glucose metabolism, insulin resistance, inflammation, and related metabolic risk markers: a randomized clinical trial.

PubMed

Hernández-Alonso, Pablo; Salas-Salvadó, Jordi; Baldrich-Mora, Mònica; Juanola-Falgarona, Martí; Bulló, Mònica

2014-11-01

To examine whether a pistachio-rich diet reduces the prediabetes stage and improves its metabolic risk profile. Prediabetic subjects were recruited to participate in this Spanish randomized clinical trial between 20 September 2011 and 4 February 2013. In a crossover manner, 54 subjects consumed two diets, each for 4 months: a pistachio-supplemented diet (PD) and a control diet (CD). A 2-week washout period separated study periods. Diets were isocaloric and matched for protein, fiber, and saturated fatty acids. A total of 55% of the CD calories came from carbohydrates and 30% from fat, whereas for the PD, these percentages were 50 and 35%, respectively (including 57 g/day of pistachios). Fasting glucose, insulin, and HOMA of insulin resistance decreased significantly after the PD compared with the CD. Other cardiometabolic risk markers such as fibrinogen, oxidized LDL, and platelet factor 4 significantly decreased under the PD compared with the CD (P < 0.05), whereas glucagon-like peptide-1 increased. Interleukin-6 mRNA and resistin gene expression decreased by 9 and 6%, respectively, in lymphocytes after the pistachio intervention (P < 0.05, for PD vs. CD). SLC2A4 expression increased by 69% in CD (P = 0.03, for PD vs. CD). Cellular glucose uptake by lymphocytes decreased by 78.78% during the PD (P = 0.01, PD vs. CD). Chronic pistachio consumption is emerging as a useful nutritional strategy for the prediabetic state. Data suggest that pistachios have a glucose- and insulin-lowering effect, promote a healthier metabolic profile, and reverse certain metabolic deleterious consequences of prediabetes. © 2014 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

P-wave and QT dispersion in patients with conversion disorder.

PubMed

Izci, Filiz; Hocagil, Hilal; Izci, Servet; Izci, Vedat; Koc, Merve Iris; Acar, Rezzan Deniz

2015-01-01

The aim of this study was to investigate QT dispersion (QTd), which is the noninvasive marker of ventricular arrhythmia and sudden cardiac death, and P-wave dispersion, which is the noninvasive marker of atrial arrhythmia, in patients with conversion disorder (CD). A total of 60 patients with no known organic disease who were admitted to outpatient emergency clinic and were diagnosed with CD after psychiatric consultation were included in this study along with 60 healthy control subjects. Beck Anxiety Inventory and Beck Depression Scale were administered to patients and 12-lead electrocardiogram measurements were obtained. Pd and QTd were calculated by a single blinded cardiologist. There was no statistically significant difference in terms of age, sex, education level, socioeconomic status, weight, height, and body mass index between CD patients and controls. Beck Anxiety Inventory scores (25.2±10.8 and 3.8±3.2, respectively, P<0.001) and Beck Depression Scale scores (11.24±6.15 and 6.58±5.69, respectively, P<0.01) were significantly higher in CD patients. P-wave dispersion measurements did not show any significant differences between conversion patients and control group (46±5.7 vs 44±5.5, respectively, P=0.156). Regarding QTc and QTd, there was a statistically significant increase in all intervals in conversion patients (416±10 vs 398±12, P<0.001, and 47±4.8 vs 20±6.1, P<0.001, respectively). A similar relation to that in literature between QTd and anxiety and somatoform disorders was also observed in CD patients. QTc and QTd were significantly increased compared to the control group in patients with CD. These results suggest a possibility of increased risk of ventricular arrhythmia resulting from QTd in CD patients. Larger samples are needed to evaluate the clinical course and prognosis in terms of arrhythmia risk in CD patients.

Accumulation of CD69+ tissue‑resident memory T cells in the nasal polyps of patients with chronic rhinosinusitis.

PubMed

Ickrath, Pascal; Kleinsasser, Norbert; Ding, Xin; Ginzkey, Christian; Beyersdorf, Niklas; Hagen, Rudolf; Kerkau, Thomas; Hackenberg, Stephan

2018-08-01

In patients with chronic rhinosinusitis with nasal polyps (CRSwNP), a relative accumulation of cluster of differentiation (CD)8+ T cells over CD4+ T cells occurs in nasal polyps compared with the peripheral blood. Nasal CD8+ T cells and CD4+ T cells predominantly present an effector memory phenotype. Immunological studies have reported that memory T cells recirculate from the tissues to the peripheral blood and a high percentage of these T cells persist within the tissue. The aim of the present study was to characterize CD69+ sphingosine‑1‑phosphate receptor 1 (S1PR1)‑ tissue resident memory T cells (Trm) in the polyps of patients with CRSwNP. Tissue and blood samples were collected from 10 patients undergoing nasal sinus surgery. Expression of specific extra‑ and intracellular molecules were analyzed using multicolor flow cytometry. A significantly higher level of CD8+ T cells than CD4+ T cells was present in nasal polyps, while significantly more CD4+ T cells than CD8+ T cells were detected in the peripheral blood of patients with CRSwNP. The frequency of CD69+ T cells was significantly higher in CD8+ and CD4+ T cells in nasal polyps compared with the peripheral blood. The frequency of CD69+ S1PR1‑ Trm was also significantly higher in CD4+ and CD8+ T cells from nasal polyps compared with the peripheral blood. Within polyps, the frequency of CD69+ S1PR1‑ Trm was again significantly higher in CD8+ compared with CD4+ T cells. In summary, a significantly higher frequency of CD69+ S1PR1‑ T cells was observed in the nasal polyps compared with the peripheral blood in patients with CRSwNP. The results of the present study suggest that local regulation of the immune response occurs within nasal polyps. As such, Trm should be considered a potential stimulus in the pathogenesis of nasal polyps. However, the role of Trm in nasal polyps as a pathogenic trigger of the local inflammatory reaction requires further investigation.

СD44+/CD24- markers of cancer stem cells in patients with breast cancer of different molecular subtypes.

PubMed

Chekhun, S V; Zadvorny, T V; Tymovska, Yu O; Anikusko, M F; Novak, O E; Polishchuk, L Z

2015-03-01

To determine frequency of tumors with immunohistochemical markers of cancer stem cells (CSC) CD44+/CD24- in patients with breast cancer (BC) of different molecular subtype and to evaluate their prognostic value. Surgical material of 132 patients with BC stage I-II, age from 23 to 75 years, mean age - 50.2 ± 3.1 years was studied. Clinical, immunohistochemical (expression CD44+/CD24-), morphological, statistical. BC is characterized by heterogeneity of molecular subtypes and expression of markers (CD44+/CD24-). Immunohistochemical study of expression of CSC markers in surgical material has detected their expression in 34 (25.4%) patients with BC of different molecular subtypes. The highest frequency of cells with expression of CSC marker was observed in patients with basal molecular subtype (44.8% patients). Most of BC patients with phenotype CD44+/CD24 had stage I of tumor process (34.3%). Statistical processing of data has showen that Yule colligation coefficient equaled 0.28 (р > 0.05) that argues poor correlation between stage of tumor process and number of tumors with positive expression of CSC markers. Statistical processing of data has showen high correlation between presence of cells with expression of CSC markers and metastases of BC in regional lymph nodes (Yule colligation coefficient equals 0.943; р < 0.5). Difference in overall survival of patients with BC of basal molecular subtype depending on expression of CSC CD44+/CD24- markers was detected. Survival of patients with basal BC was reliably higher at lack in tumors of cells with CSC markers CD44+/CD24- and, correspondingly, lower at presence of such cells (р < 0.05). In patients with BC of luminal (A and B), HER-2-positive subtypes, significant change in survival of patients depending on expression of CSC markers was not determined (р > 0.05). Significance of tumor cells with markers CD44+/CD24- within the limits of molecular subtype of BC may be additional criterion for advanced biological characteristic of BC, and in patients with BC of basal molecular subtype - for predictive evaluation of individual potential of tumor to aggressive clinical course.

Flow analysis of individual blood extracellular vesicles in acute coronary syndrome.

PubMed

Vagida, Murad; Arakelyan, Anush; Lebedeva, Anna; Grivel, Jean-Charles; Shpektor, Alexander; Vasilieva, Elena; Margolis, Leonid

2017-03-01

A diverse population of small extracellular vesicles (EVs) that are released by various cells has been characterized predominantly in bulk, a procedure whereby the individual characteristics of EVs are lost. Here, we used a new nanotechnology-based flow cytometric analysis to characterize the antigenic composition of individual EVs in patients with acute coronary syndrome (ACS). Plasma EVs were captured with 15-nm magnetic nanoparticles coupled to antibodies against CD31 (predominantly an endothelial marker), CD41a (a marker for platelets), and CD63 or MHC class I (common EV markers). The total amounts of EVs were higher in the ACS patients than in the controls, predominantly due to the contribution of patients with acute myocardial infarction. For all captured fractions, the differences in the EV amounts were restricted to CD41a + EVs. The increase in the numbers of EVs in the ACS patients, predominantly of platelet origin, probably reflects platelet activation and may indicate disease progression.

The requirement for freshly isolated human colorectal cancer (CRC) cells in isolating CRC stem cells.

PubMed

Fan, F; Bellister, S; Lu, J; Ye, X; Boulbes, D R; Tozzi, F; Sceusi, E; Kopetz, S; Tian, F; Xia, L; Zhou, Y; Bhattacharya, R; Ellis, L M

2015-02-03

Isolation of colorectal cancer (CRC) cell populations enriched for cancer stem cells (CSCs) may facilitate target identification. There is no consensus regarding the best methods for isolating CRC stem cells (CRC-SCs). We determined the suitability of various cellular models and various stem cell markers for the isolation of CRC-SCs. Established human CRC cell lines, established CRC cell lines passaged through mice, patient-derived xenograft (PDX)-derived cells, early passage/newly established cell lines, and cells directly from clinical specimens were studied. Cells were FAC-sorted for the CRC-SC markers CD44, CD133, and aldehyde dehydrogenase (ALDH). Sphere formation and in vivo tumorigenicity studies were used to validate CRC-SC enrichment. None of the markers studied in established cell lines, grown either in vitro or in vivo, consistently enriched for CRC-SCs. In the three other cellular models, CD44 and CD133 did not reliably enrich for stemness. In contrast, freshly isolated PDX-derived cells or early passage/newly established CRC cell lines with high ALDH activity formed spheres in vitro and enhanced tumorigenicity in vivo, whereas cells with low ALDH activity did not. PDX-derived cells, early passages/newly established CRC cell lines and cells from clinical specimen with high ALDH activity can be used to identify CRC-SC-enriched populations. Established CRC cell lines should not be used to isolate CSCs.

[Immunological aspects of a piloted mission to Mars].

PubMed

Morukov, B V; Rykova, M P; Antropova, E N; Berendeeva, T A; Morukov, I B; Ponomarev, S A

2013-01-01

The paper deals with the results of the effects of 520-day isolation and confinement modeling some elements of a mission to Mars on the immune system. Longitudinal analyses revealed that the mechanisms of adaptive response of the human immune system to the conditions of extremely long isolation led to a change of the parameters, characterizing innate and adaptive immunity. Among them the most important are: changes in the signaling PRRs--TLR, manifested in the reduction of the percentage of circulating monocytes and granulocytes expressed on its own surfaces TLR2, TLR4 and TLR6, decreases early NK cell activation potential, increases in the percentage T- and B-lymphocytes, that expressed early activation marker CD69 after adequate stimulation, and in production of cytokines in response to PHA stimulation. The active mobilization of the mechanisms of adaptive immunity, the implementation of the function of the level of immunity to a qualitatively different level, apparently, should be taken as a sign of adaptive adjustment of an organism in response to the complex influence of unfavorable factors, aimed at the preservation of immune homeostasis.

[Inhibitory effect and mechanism of tofacitinib on the secretion of cytokines by T cells in human peripheral blood].

PubMed

Wu, Kunlun; Zhao, Jun; Wu, Qiongli; Wu, Changyou

2017-11-01

Objective To study the inhibitory effect of tofacitinib on the production of cytokines by T cells in human peripheral blood and its mechanism. Methods Peripheral blood mononuclear cells (PBMCs) and purified T cells were cultured and stimulated with anti-CD3 plus anti-CD28 antibodies in the presence or absence of tofacitinib (0.5 μmol/L). The levels of interferon γ (IFN-γ), tumor necrosis factor α (TNF-α) and interleukin 2 (IL-2) in the culture supernatants were detected by ELISA, and the expressions of activated molecules CD69 and CD25 on the surface of CD4 + and CD8 + T cells, the production of cytokines and the phosphorylation of signal transducers and transcriptional activators STAT1, STAT3, STAT4 in T cells were examined by flow cytometry. At the same time, the proliferation and apoptosis of T cells were observed by 5- (and 6-) carboxyfluorescein diacetate succinimidyl ester (CFSE) staining and the flow cy tometry with annexin V-FITC/PI, respectively. Results Tofacitinib inhibited the production of IFN-γ, TNF-α and the expression of CD25 on T cells from the peripheral blood. In addition, the proliferation and the phosphorylation of STAT1, STAT3, STAT4 by T cells were also depressed. However, tofacitinib had no effect on the secretion of IL-2, the expression of CD69 and the apoptosis of T cells. Conclusion Tofacitinib can inhibit the secretion of IFN-γ and TNF-α by T cells in the peripheral blood, and its mechanism might be related to the inhibitory effect of tofacitinib on the activation, proliferation and signal transduction in T cells.

Functional heterogeneity of side population cells in skeletal muscle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Uezumi, Akiyoshi; Ojima, Koichi; Fukada, So-ichiro

2006-03-17

Skeletal muscle regeneration has been exclusively attributed to myogenic precursors, satellite cells. A stem cell-rich fraction referred to as side population (SP) cells also resides in skeletal muscle, but its roles in muscle regeneration remain unclear. We found that muscle SP cells could be subdivided into three sub-fractions using CD31 and CD45 markers. The majority of SP cells in normal non-regenerating muscle expressed CD31 and had endothelial characteristics. However, CD31{sup -}CD45{sup -} SP cells, which are a minor subpopulation in normal muscle, actively proliferated upon muscle injury and expressed not only several regulatory genes for muscle regeneration but also somemore » mesenchymal lineage markers. CD31{sup -}CD45{sup -} SP cells showed the greatest myogenic potential among three SP sub-fractions, but indeed revealed mesenchymal potentials in vitro. These SP cells preferentially differentiated into myofibers after intramuscular transplantation in vivo. Our results revealed the heterogeneity of muscle SP cells and suggest that CD31{sup -}CD45{sup -} SP cells participate in muscle regeneration.« less

Cellular and Molecular Level Responses After Radiofrequency Radiation Exposure, Alone or in Combination with X-rays or Chemicals

DTIC Science & Technology

1992-07-27

cell surface marker CD22 , which plays a role in early B-cell activation, is present within the cytoplasm of all B- cells, but expressed only on the...surface of a subpopulation of those cells. CD22 is an activation receptor associated with cell proliferation of small resting B-cells, and acts as an...adhesion molecule mediating the binding of B-cells to other hematopoietic cells (Stamenkovic & Seed, 1990). The CD22 surface glycoprotein is the putative

Universal monitoring of minimal residual disease in acute myeloid leukemia.

PubMed

Coustan-Smith, Elaine; Song, Guangchun; Shurtleff, Sheila; Yeoh, Allen Eng-Juh; Chng, Wee Joo; Chen, Siew Peng; Rubnitz, Jeffrey E; Pui, Ching-Hon; Downing, James R; Campana, Dario

2018-05-03

Optimal management of acute myeloid leukemia (AML) requires monitoring of treatment response, but minimal residual disease (MRD) may escape detection. We sought to identify distinctive features of AML cells for universal MRD monitoring. We compared genome-wide gene expression of AML cells from 157 patients with that of normal myeloblasts. Markers encoded by aberrantly expressed genes, including some previously associated with leukemia stem cells, were studied by flow cytometry in 240 patients with AML and in nonleukemic myeloblasts from 63 bone marrow samples. Twenty-two (CD9, CD18, CD25, CD32, CD44, CD47, CD52, CD54, CD59, CD64, CD68, CD86, CD93, CD96, CD97, CD99, CD123, CD200, CD300a/c, CD366, CD371, and CX3CR1) markers were aberrantly expressed in AML. Leukemia-associated profiles defined by these markers extended to immature CD34+CD38- AML cells; expression remained stable during treatment. The markers yielded MRD measurements matching those of standard methods in 208 samples from 52 patients undergoing chemotherapy and revealed otherwise undetectable MRD. They allowed MRD monitoring in 129 consecutive patients, yielding prognostically significant results. Using a machine-learning algorithm to reduce high-dimensional data sets to 2-dimensional data, the markers allowed a clear visualization of MRD and could detect 1 leukemic cell among more than 100,000 normal cells. The markers uncovered in this study allow universal and sensitive monitoring of MRD in AML. In combination with contemporary analytical tools, the markers improve the discrimination between leukemic and normal cells, thus facilitating data interpretation and, hence, the reliability of MRD results. National Cancer Institute (CA60419 and CA21765); American Lebanese Syrian Associated Charities; National Medical Research Council of Singapore (1299/2011); Viva Foundation for Children with Cancer, Children's Cancer Foundation, Tote Board & Turf Club, and Lee Foundation of Singapore.

Oral manifestations of human immunodeficiency virus/acquired immunodeficiency syndrome and their correlation to cluster of differentiation lymphocyte count in population of North-East India in highly active antiretroviral therapy era.

PubMed

Nayak, Sarat Kumar; Das, Bijay Kumar; Das, Surya Narayan; Mohapatra, Namita; Nayak, Suryakanti; Bhuyan, Lipsa

2016-01-01

The human immunodeficiency virus (HIV) infection which manifests as acquired immunodeficiency syndrome (AIDS) is a disease involving the defects of the T-lymphocyte arm of the immune system. Certain laboratory parameters such as the cluster of differentiation (CD4) count and clinical parameters have long been used as markers of disease progression. In industrialized countries, many studies show a highly correlation between the incidence of oral lesions and immunosuppression and hence, can be used as a marker of immunosuppression. This might not be applicable to a developing country like India. In this study, efforts have been made to supplement the present knowledge on various aspects of oral manifestations in HIV patients in the Indian subcontinent. To correlate the oral manifestations in HIV/AIDS patients to the level of circulating CD4+ T-lymphocyte count and their effect in anti-retroviral therapy (ART). A total of 104 HIV positive patients were examined for oral lesions. The CD4 count estimated on the same day by fluorescent activated cell sort count machine was then correlated with various oral lesions. Oral manifestations appeared when CD4 count decreased below 500 cells/mm 3 . Moreover, oral lesions found at different stages showed very strong correlation to their respective CD4 count. Furthermore, there was considerable decline in the incidence of oral manifestations in patients undergoing highly active ART. Oral manifestations are highly predictive markers of severe immune deterioration and disease progression in HIV patients.

A novel kefir product (PFT) activates dendritic cells to induce CD4+T and CD8+T cell responses in vitro.

PubMed

Ghoneum, Mamdooh; Felo, Nouran; Agrawal, Sudhanshu; Agrawal, Anshu

2015-12-01

Lactobacilli have been widely studied for their probiotic effects and have been reported to function as antiviral and anticancer agents. However, the underlying mechanisms via immune modulation are poorly understood. PFT is a freeze dried compound of Lactobacillus kefiri P-IF with a unique composition and functionality. In this study, we examined the potential stimulatory effects of two concentrations (50 µg and 100 µg/mL) of PFT on human monocyte-derived dendritic cell (DC) function in vitro. Results showed that PFT upregulated the expression of DC surface co-stimulatory and maturation markers CD80, CD86, and HLADR in a concentration dependent manner. PFT at 100 µg/mL markedly increased the secretion of IL-6, IL-10, TNF-α, and IL-1β by DCs. This concentration of PFT also stimulated the production of antiviral cytokines, IFN-α and IFN-λ(IL29) in DCs. Additionally, PFT at 100 µg/mL activated moDCs prime CD4(+)T cells and significantly increased the levels of IL-10, IFN-γ, and TNF-α by 1.7, four, three-fold, respectively. Furthermore PFT-stimulated DCs were also effective in enhancing the cytotoxic potential of CD8(+)T cells via the induction of Granzyme-B and upregulation of CD107a, and CD103 expression, a marker of resident/regulatory CD8(+)T cells. These data suggest that PFT functions as a natural adjuvant for DC activation and thus may be used in DC-based vaccine strategies against viral infections and cancer. © The Author(s) 2015.

CD25 is expressed by canine cutaneous mast cell tumors but not by cutaneous connective tissue mast cells.

PubMed

Meyer, A; Gruber, A D; Klopfleisch, R

2012-11-01

Canine cutaneous mast cell tumors (MCT) of different histological grades have distinct biological behaviors. However, little is known about underlying molecular mechanisms that lead to tumor development and increasing malignancy with higher tumor grade. Recent studies have identified the interleukin-2 receptor (IL-2R) subunits CD25 and CD2 as markers that distinguish nonneoplastic from neoplastic mast cells in human systemic mastocytosis. In this study, their potential as a marker for canine MCT and their possible impact on MCT carcinogenesis were evaluated. mRNA expression levels of both genes were compared between grade 1 (n = 12) and grade 3 (n = 8) MCT, and protein expression levels of CD25 were compared in 90 MCT of different tumor grades. mRNA expression levels of both CD25 and CD2 were upregulated in grade 3 MCT. In contrast, CD25 protein was expressed by fewer tumor cells and at decreased levels in grade 3 tumors, while most grade 1 MCT had strong CD25 protein expression. Moreover, CD25 was not expressed by nonneoplastic, resting cutaneous mast cells, while few presumably activated mast cells in tissue samples from dogs with allergic dermatitis had weak CD25 expression. Taken together, these findings suggest that CD25 may play a critical role in early MCT development and may be a stimulatory factor in grade 1 MCT, while grade 3 MCT seem to be less dependent on CD25. Because of the low number of CD25-positive tumor cells in high-grade tumors, the usefulness of CD25 as a tumor marker is, however, questionable.

T cell activation responses are differentially regulated during clinorotation and in spaceflight

NASA Technical Reports Server (NTRS)

Hashemi, B. B.; Penkala, J. E.; Vens, C.; Huls, H.; Cubbage, M.; Sams, C. F.

1999-01-01

Studies of T lymphocyte activation with mitogenic lectins during spaceflight have shown a dramatic inhibition of activation as measured by DNA synthesis at 72 h, but the mechanism of this inhibition is unknown. We have investigated the progression of cellular events during the first 24 h of activation using both spaceflight microgravity culture and a ground-based model system that relies on the low shear culture environment of a rotating clinostat (clinorotation). Stimulation of human peripheral blood mononuclear cells (PBMCs) with soluble anti-CD3 (Leu4) in clinorotation and in microgravity culture shows a dramatic reduction in surface expression of the receptor for IL-2 (CD25) and CD69. An absence of bulk RNA synthesis in clinorotation indicates that stimulation with soluble Leu4 does not induce transition of T cells from G0 to the G1 stage of the cell cycle. However, internalization of the TCR by T cells and normal levels of IL-1 synthesis by monocytes indicate that intercellular interactions that are required for activation occur during clinorotation. Complementation of TCR-mediated signaling by phorbol ester restores the ability of PBMCs to express CD25 in clinorotation, indicating that a PKC-associated pathway may be compromised under these conditions. Bypassing the TCR by direct activation of intracellular pathways with a combination of phorbol ester and calcium ionophore in clinorotation resulted in full expression of CD25; however, only partial expression of CD25 occurred in microgravity culture. Though stimulation of purified T cells with Bead-Leu4 in microgravity culture resulted in the engagement and internalization of the TCR, the cells still failed to express CD25. When T cells were stimulated with Bead-Leu4 in microgravity culture, they were able to partially express CD69, a receptor that is constitutively stored in intracellular pools and can be expressed in the absence of new gene expression. Our results suggest that the inhibition of T cell proliferative response in microgravity culture is a result of alterations in signaling events within the first few hours of activation, which are required for the expression of important regulatory molecules.

The macrophage activation marker CD163 is associated with IL28B genotype and hepatic inflammation in chronic hepatitis C virus infected patients.

PubMed

Dultz, G; Gerber, L; Zeuzem, S; Sarrazin, C; Waidmann, O

2016-04-01

Recent data highlighted the association of the macrophage activation marker CD163 with histological inflammation and fibrosis in chronic hepatitis C virus (HCV) infection. The aim of this study was to investigate the influence of successful antiviral treatment and IL28B genotypes on macrophage activation reflected by CD163 levels in HCV infected patients. In a retrospective cohort study, serum sCD163 levels were correlated with results of liver histopathology, IL28B genotyping and clinical parameters in 329 patients with HCV infection, 15 healthy controls and in 161 patients who achieved a sustained virologic response after antiviral treatment. sCD163 levels were significantly higher in patients with chronic HCV infection in comparison to healthy controls (5202 vs 896 ng/mL, P < 0.001). In the multivariate logistic regression analyses, sCD163 was independently associated with histologically determined inflammation (P = 0.043) but not with fibrosis (P = 0.091). sCD163 dropped significantly after successful antiviral treatment in comparison to baseline values (5202 vs 3093 ng/mL, P < 0.001). In the univariate analyses, sCD163 was significantly associated with IL28B genotype (C/C vs C/T+T/T) with higher values in the C/C group (6098 vs 4812 ng/mL, P = 0.003). In the multivariate logistic regression model, sCD163 levels were significantly associated with IL28B genotype (P = 0.003) and sustained virologic response (SVR) (P < 0.001). Our data support the association of activated liver macrophages with hepatic necroinflammation in chronic HCV infection as sCD163 levels drop rapidly after SVR. The irresponsiveness of IL28B minor genotypes to interferon might be related to a lower level of macrophage activation in these patients. © 2015 John Wiley & Sons Ltd.

Isolation and characterisation of human gingival margin-derived STRO-1/MACS+ and MACS− cell populations

PubMed Central

El-Sayed, Karim M Fawzy; Paris, Sebastian; Graetz, Christian; Kassem, Neemat; Mekhemar, Mohamed; Ungefroren, Hendrick; Fändrich, Fred; Dörfer, Christof

2015-01-01

Recently, gingival margin-derived stem/progenitor cells isolated via STRO-1/magnetic activated cell sorting (MACS) showed remarkable periodontal regenerative potential in vivo. As a second-stage investigation, the present study's aim was to perform in vitro characterisation and comparison of the stem/progenitor cell characteristics of sorted STRO-1-positive (MACS+) and STRO-1-negative (MACS−) cell populations from the human free gingival margin. Cells were isolated from the free gingiva using a minimally invasive technique and were magnetically sorted using anti-STRO-1 antibodies. Subsequently, the MACS+ and MACS− cell fractions were characterized by flow cytometry for expression of CD14, CD34, CD45, CD73, CD90, CD105, CD146/MUC18 and STRO-1. Colony-forming unit (CFU) and multilineage differentiation potential were assayed for both cell fractions. Mineralisation marker expression was examined using real-time polymerase chain reaction (PCR). MACS+ and MACS− cell fractions showed plastic adherence. MACS+ cells, in contrast to MACS− cells, showed all of the predefined mesenchymal stem/progenitor cell characteristics and a significantly higher number of CFUs (P<0.01). More than 95% of MACS+ cells expressed CD105, CD90 and CD73; lacked the haematopoietic markers CD45, CD34 and CD14, and expressed STRO-1 and CD146/MUC18. MACS− cells showed a different surface marker expression profile, with almost no expression of CD14 or STRO-1, and more than 95% of these cells expressed CD73, CD90 and CD146/MUC18, as well as the haematopoietic markers CD34 and CD45 and CD105. MACS+ cells could be differentiated along osteoblastic, adipocytic and chondroblastic lineages. In contrast, MACS− cells demonstrated slight osteogenic potential. Unstimulated MACS+ cells showed significantly higher expression of collagen I (P<0.05) and collagen III (P<0.01), whereas MACS− cells demonstrated higher expression of osteonectin (P<0.05; Mann–Whitney). The present study is the first to compare gingival MACS+ and MACS− cell populations demonstrating that MACS+ cells, in contrast to MACS− cells, harbour stem/progenitor cell characteristics. This study also validates the effectiveness of the STRO-1/MACS+ technique for the isolation of gingival stem/progenitor cells. Human free gingival margin-derived STRO-1/MACS+ cells are a unique renewable source of multipotent stem/progenitor cells. PMID:25257881

Brief Report: CD14brightCD16- monocytes and sCD14 level negatively associate with CD4-memory T-cell frequency and predict HCV-decline on therapy.

PubMed

Judge, Chelsey J; Sandberg, Johan K; Funderburg, Nicholas T; Sherman, Kenneth E; Butt, Adeel A; Kang, Minhee; Landay, Alan L; Lederman, Michael M; Anthony, Donald D

2016-11-01

During HIV+ hepatitis C virus (HCV)+ coinfection CD14CD16 monocytes produce soluble immune-activation markers that predict disease progression and poor response to interferon (IFN)-α treatment. We evaluated relationships among immune activation, monocyte phenotype, CD4-memory T cells, and HCV-, cytomegalovirus-, and cytomegalovirus/Epstein-Barr virus/influenza-specific IFN-γ-response before and during IFN-α treatment. Effector-memory and central-memory CD4 T-cell frequencies were lower in HCV+ HIV+ donors than in uninfected donors and correlated negatively with HCV level, CD14CD16 monocytes, and plasma sCD14. sCD14 and CD14CD16 monocytes negatively correlated with IFN-α-dependent HCV decline. CD4 effector-memory T cells positively associated with cytomegalovirus/Epstein-Barr virus/influenza(CEF)-specific IFN-γ response, while sCD14 negatively associated with both CD4 effector-memory T cells and CEF-specific IFN-γ response. These data support a role for memory-CD4 T cells in HCV containment and link immune activation and CD14CD16-monocyte frequency to the failure of IFN-dependent HCV clearance.

Expression of the myeloid-associated marker CD33 is not an exclusive factor for leukemic plasmacytoid dendritic cells.

PubMed

Garnache-Ottou, Francine; Chaperot, Laurence; Biichle, Sabeha; Ferrand, Christophe; Remy-Martin, Jean-Paul; Deconinck, Eric; de Tailly, Patrick Darodes; Bulabois, Bénédicte; Poulet, Jacqueline; Kuhlein, Emilienne; Jacob, Marie-Christine; Salaun, Véronique; Arock, Michel; Drenou, Bernard; Schillinger, Françoise; Seilles, Estelle; Tiberghien, Pierre; Bensa, Jean-Claude; Plumas, Joel; Saas, Philippe

2005-02-01

A new entity of acute leukemia coexpressing CD4(+)CD56(+) markers without any other lineage-specific markers has been identified recently as arising from lymphoid-related plasmacytoid dendritic cells (pDCs). In our laboratory, cells from a patient with such CD4(+)CD56(+) lineage-negative leukemia were unexpectedly found to also express the myeloid marker CD33. To confirm the diagnosis of pDC leukemia despite the CD33 expression, we demonstrated that the leukemic cells indeed exhibited pDC phenotypic and functional properties. In 7 of 8 other patients with CD4(+)CD56(+) pDC malignancies, we were able to confirm that the tumor cells expressed CD33 although with variable expression levels. CD33 expression was shown by flow cytometry, reverse transcriptase-polymerase chain reaction, and immunoblot analysis. Furthermore, CD33 monoclonal antibody stimulation of purified CD4(+)CD56(+) leukemic cells led to cytokine secretion, thus confirming the presence of a functional CD33 on these leukemic cells. Moreover, we found that circulating pDCs in healthy individuals also weakly express CD33. Overall, our results demonstrate that the expression of CD33 on CD4(+)CD56(+) lineage-negative cells should not exclude the diagnosis of pDC leukemia and underline that pDC-specific markers should be used at diagnosis for CD4(+)CD56(+) malignancies.

Aberrant expression of cancer stem cell markers (CD44, CD90, and CD133) contributes to disease progression and reduced survival in hepatoblastoma patients: 4-year survival data.

PubMed

Bahnassy, Abeer A; Fawzy, Mohamed; El-Wakil, Mohamed; Zekri, Abdel-Rahman N; Abdel-Sayed, Ahmed; Sheta, Marwa

2015-03-01

Hepatoblastoma (HB) is an embryonal tumor of the liver in children. Prognosis and response to treatment in HB are highly variable. Cancer stem cells (CSCs) constitute a population of cells, which contribute to the development and progression of many tumors. However, their role in HB is not well defined yet. We assessed the prognostic and predictive values of some CSC markers in HB patients. Protein and messenger RNA expressions of the CSC markers CD133, CD90, and CD44 were assessed in 43 HB patients and 20 normal hepatic tissues using immunohistochemistry and quantitative real-time polymerase chain reaction. The expression levels of these markers were correlated to standard prognostic factors, patients' response to treatment, overall survival (OS), and disease-free survival (DFS). CD44, CD90, and CD133 proteins were detected in 48.8%, 32.6%, and 48.8% compared with 46.5%, 41.7%, and 58.1% RNA, respectively (concordance, 77.8%-96%). None of the normal tissue samples was positive for any of the markers. Significant correlations were reported between α-fetoprotein and both CD44 and CD133 (P = 0.02) as well as between tumor types CD90 and CD133 (P = 0.009). Reduced OS correlated with CD44, CD90, and CD133 expressions (P < 0.001), advanced stage (P < 0.001), response to treatment (P < 0.001), and total excision of the tumor. Reduced DFS correlated with CD44 and CD133 expressions (P < 0.001) only. In conclusion, CD133, CD44, and CD90 could be used as prognostic and predictive markers in HB. High expression of these markers is significantly associated with poor response to treatment and reduced survival. Moreover, complete surgical resection and systemic chemotherapy are essential to achieve good response and prolonged survival, especially in early stage patients. Copyright © 2015 Elsevier Inc. All rights reserved.

Increased expression of activation antigens on CD8+ T lymphocytes in Myalgic Encephalomyelitis/chronic fatigue syndrome: inverse associations with lowered CD19+ expression and CD4+/CD8+ ratio, but no associations with (auto)immune, leaky gut, oxidative and nitrosative stress biomarkers.

PubMed

Maes, Michael; Bosmans, Eugene; Kubera, Marta

2015-01-01

There is now evidence that specific subgroups of patients with Myalgic Encephalomyelitis / chronic fatigue syndrome (ME/CFS) suffer from a neuro-psychiatric-immune disorder. This study was carried out to delineate the expression of the activation markers CD38 and human leukocyte antigen (HLA) DR on CD4+ and CD8+ peripheral blood lymphocytes in ME/CFS. Proportions and absolute numbers of peripheral lymphocytes expressing CD3+, CD19+, CD4+, CD8+, CD38+ and HLA-DR+ were measured in ME/CFS (n=139), chronic fatigue (CF, n=65) and normal controls (n=40). The proportions of CD3+, CD8+, CD8+CD38+ and CD8+HLA-DR+ were significantly higher in ME/CFS patients than controls, while CD38+, CD8+CD38+, CD8+HLA-DR+ and CD38+HLA-DR+ were significantly higher in ME/CFS than CF. The percentage of CD19+ cells and the CD4+/CD8+ ratio were significantly lower in ME/CFS and CF than in controls. There were highly significant inverse correlations between the increased expression of CD38+, especially that of CD8+CD38+, and the lowered CD4+/CD8+ ratio and CD19+ expression. There were no significant associations between the flow cytometric results and severity or duration of illness and peripheral blood biomarkers of oxidative and nitrosative stress (O&NS, i.e. IgM responses to O&N modified epitopes), leaky gut (IgM or IgA responses to LPS of gut commensal bacteria), cytokines (interleukin-1, tumor necrosis factor-α), neopterin, lysozyme and autoimmune responses to serotonin. The results support that a) increased CD38 and HLA-DR expression on CD8+ T cells are biomarkers of ME/CFS; b) increased CD38 antigen expression may contribute to suppression of the CD4+/CD8+ ratio and CD19+ expression; c) there are different immune subgroups of ME/CFS patients, e.g. increased CD8+ activation marker expression versus inflammation or O&NS processes; and d) viral infections or reactivation may play a role in a some ME/CFS patients.

Routine detection of Epstein-Barr virus specific T-cells in the peripheral blood by flow cytometry

NASA Technical Reports Server (NTRS)

Crucian, B. E.; Stowe, R. P.; Pierson, D. L.; Sams, C. F.

2001-01-01

The ability to detect cytomegalovirus-specific T-cells (CD4(+)) in the peripheral blood by flow cytometry has been recently described by Picker et al. In this method, cells are incubated with viral antigen and responding (cytokine producing) T-cells are then identified by flow cytometry. To date, this technique has not been reliably used to detect Epstein-Barr virus (EBV)-specific T-cells primarily due to the superantigen/mitogenic properties of the virus which non-specifically activate T-cells. By modifying culture conditions under which the antigens are presented, we have overcome this limitation and developed an assay to detect and quantitate EBV-specific T-cells. The detection of cytokine producing T-cells by flow cytometry requires an extremely strong signal (such as culture in the presence of PMA and ionomycin). Our data indicate that in modified culture conditions (early removal of viral antigen) the non-specific activation of T-cells by EBV is reduced, but antigen presentation will continue uninhibited. Using this method, EBV-specific T-cells may be legitimately detected using flow cytometry. No reduction in the numbers of antigen-specific T-cells was observed by the early removal of target antigen when verified using cytomegalovirus antigen (a virus with no non-specific T-cell activation properties). In EBV-seropositive individuals, the phenotype of the EBV-specific cytokine producing T-cells was evaluated using four-color flow cytometry and found to be CD45(+), CD3(+), CD4(+), CD45RA(-), CD69(+), CD25(-). This phenotype indicates the stimulation of circulating previously unactivated memory T-cells. No cytokine production was observed in CD4(+) T-cells from EBV-seronegative individuals, confirming the specificity of this assay. In addition, the use of four color cytometry (CD45, CD3, CD69, IFNgamma/IL-2) allows the total quantitative assessment of EBV-specific T-cells while monitoring the interference of EBV non-specific mitogenic activity. This method may have significant utility for the monitoring of the immune response to latent virus infection/reactivation.

Impact of antiretroviral therapy (ART) timing on chronic immune activation/inflammation and end-organ damage.

PubMed

Rajasuriar, Reena; Wright, Edwina; Lewin, Sharon R

2015-01-01

The purpose of this review was to summarize recent studies on the effect of early antiretroviral therapy (ART) in HIV-infected patients on markers of immune activation/inflammation, viral persistence and serious non-AIDS events. Early ART, initiated within days to months of HIV infection, was associated with marked reduction in T-cell activation often reaching levels observed in HIV-uninfected individuals. However, the impact of early ART on markers of innate immune activation, microbial translocation and inflammation/coagulation was less clear. Early ART has also been associated with a significant reduction in the frequency of latently infected cells, which was greater if ART was initiated within days to weeks rather than months following infection. However, few studies have evaluated the relationship between immune activation and viral reservoirs, specifically following early ART. Early ART may potentially reduce serious non-AIDS events and associated mortality, but most of these studies have extrapolated from changes in surrogate markers, such as CD4 : CD8 ratio. Early ART was associated with beneficial effects on multiple markers of immune activation, inflammation and viral persistence. Longer term prospective studies are still needed to determine whether early ART translates to a significant reduction in serious non-AIDS events and mortality.

Profiling of normal and malignant breast tissue show CD44high/CD24low phenotype as a predominant stem/progenitor marker when used in combination with Ep-CAM/CD49f markers

PubMed Central

2013-01-01

Background Accumulating evidence supports cancer to initiate and develop from a small population of stem-like cells termed as cancer stem cells (CSC). The exact phenotype of CSC and their counterparts in normal mammary gland is not well characterized. In this study our aim was to evaluate the phenotype and function of stem/progenitor cells in normal mammary epithelial cell populations and their malignant counterparts. Methods Freshly isolated cells from both normal and malignant human breasts were sorted using 13 widely used stem/progenitor cell markers individually or in combination by multi-parametric (up to 9 colors) cell sorting. The sorted populations were functionally evaluated by their ability to form colonies and mammospheres, in vitro. Results We have compared, for the first time, the stem/progenitor markers of normal and malignant breasts side-by-side. Amongst all markers tested, we found CD44high/CD24low cell surface marker combination to be the most efficient at selecting normal epithelial progenitors. Further fractionation of CD44high/CD24low positive cells showed that this phenotype selects for luminal progenitors within Ep-CAMhigh/CD49f + cells, and enriches for basal progenitors within Ep-CAM-/low/CD49f + cells. On the other hand, primary breast cancer samples, which were mainly luminal Ep-CAMhigh, had CD44high/CD24low cells among both CD49fneg and CD49f + cancer cell fractions. However, functionally, CSC were predominantly CD49f + proposing the use of CD44high/CD24low in combination with Ep-CAM/CD49f cell surface markers to further enrich for CSC. Conclusion Our study clearly demonstrates that both normal and malignant breast cells with the CD44high/CD24low phenotype have the highest stem/progenitor cell ability when used in combination with Ep-CAM/CD49f reference markers. We believe that this extensive characterization study will help in understanding breast cancer carcinogenesis, heterogeneity and drug resistance. PMID:23768049

Biomarkers of disease activity in vitiligo: A systematic review.

PubMed

Speeckaert, R; Speeckaert, M; De Schepper, S; van Geel, N

2017-09-01

The pathophysiology of vitiligo is complex although recent research has discovered several markers which are linked to vitiligo and associated with disease activity. Besides providing insights into the driving mechanisms of vitiligo, these findings could reveal potential biomarkers. Activity markers can be used to monitor disease activity in clinical trials and may also be useful in daily practice. The aim of this systematic review was to document which factors have been associated with vitiligo activity in skin and blood. A second goal was to determine how well these factors are validated in terms of sensitivity and specificity as biomarkers to determine vitiligo activity. Both in skin (n=43) as in blood (n=66) an adequate number of studies fulfilled the predefined inclusion criteria. These studies used diverse methods and investigated a broad range of plausible biomarkers. Unfortunately, sensitivity and specificity analyses were scarce. In skin, simple histopathology with or without supplemental CD4 and CD8 stainings can still be considered as the gold standard, although more recently chemokine (C-X-C motif) ligand (CXCL) 9 and NLRP1 have demonstrated a good and possibly even better association with progressive disease. Regarding circulating biomarkers, cytokines (IL-1β, IL-17, IFN-γ, TGF-β), autoantibodies, oxidative stress markers, immune cells (Tregs), soluble CDs (sCD25, sCD27) and chemokines (CXCL9, CXCL10) are still competing. However, the two latter may be preferable as both chemokines and soluble CDs are easy to measure and the available studies display promising results. A large multicenter study could make more definitive statements regarding their sensitivity and specificity. Copyright © 2017 Elsevier B.V. All rights reserved.

Relationship Between Markers of Platelet Activation and Inflammation with Disease Activity in Wegener’s Granulomatosis

PubMed Central

TOMASSON, GUNNAR; LAVALLEY, MICHAEL; TANRIVERDI, KAHRAMAN; FINKIELMAN, JAVIER D.; DAVIS, JOHN C.; HOFFMAN, GARY S.; McCUNE, W. JOSEPH; St. CLAIR, E. WILLIAM; SPECKS, ULRICH; SPIERA, ROBERT; STONE, JOHN H.; FREEDMAN, JANE E.; MERKEL, PETER A.

2013-01-01

Objective There remains a need for biomarkers to guide therapy in antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis. Our objective was to determine whether measures of platelet activation or inflammation are associated with disease activity in Wegener’s granulomatosis (WG). Methods Study subjects were participants in a clinical trial. Soluble CD40 ligand (sCD40L), C-reactive protein, interleukin 6 (IL-6), IL-8, monocyte chemoattractant protein 1 (MCP-1), P-selectin, vascular endothelial growth factor, and proteinase 3 (PR3)-specific ANCA were measured by ELISA using plasma samples obtained at baseline (active disease), at remission, and prior to, during, and after first flares. Disease activity was assessed by the Birmingham Vasculitis Activity Score for WG (BVAS/WG). Association of biomarkers with disease activity was determined with conditional logistic and linear regression. Results Over a mean followup of 27 months, 180 subjects underwent 2044 visits; markers were measured in 563 samples. Longitudinally, all markers other than IL-6 were associated with disease activity. The strongest associations for active disease at baseline versus remission were observed for sCD40L (OR 4.72, 95% CI 2.47–9.03), P-selectin (OR 6.26, 95% CI 2.78–14.10), PR3-ANCA (OR 9.41, 4.03–21.99), and inversely for MCP-1 (OR 0.36, 95% CI 0.22–0.57). BVAS/WG increased by 0.80 (95% CI 0.44–1.16), 0.83 (95% CI 0.42–1.25), and 0.81 (95% CI 0.48–1.15) per unit-increase in PR3-ANCA, sCD40L, and P-selectin, respectively; and decreased by 1.54 (95% CI 0.96–2.12) per unit-increase in MCP-1. Conclusion Cytokines arising from within the circulation, including those of platelet activation, correlate with disease activity in WG. PMID:21411717

Activin receptor-like kinase 5 inhibition reverses impairment of endothelial cell viability by endogenous islet mesenchymal stromal cells.

PubMed

Clarkin, Claire E; King, Aileen J; Dhadda, Paramjeet; Chagastelles, Pedro; Nardi, Nance; Wheeler-Jones, Caroline P; Jones, Peter M

2013-03-01

Following islet transplantation, islet graft revascularization is compromised due to loss of endothelial cells (ECs) during islet culture. TGF-β signaling pathways are essential for vascular homeostasis but their importance for islet EC function is unclear. We have identified a population of multipotent mesenchymal stromal cells (MSCs) within islets and investigated how modulation of TGF-β signaling by these cells influences islet EC viability. Cultured islets exhibited reduced expression of EC markers (VEGFR2, VE-cadherin and CD31), which was associated with diminished but sustained expression of endoglin a marker of both ECs and MSCs. Double fluorescent labeling of islets in situ with the EC marker CD31 disclosed a population of CD31-negative cells which were positive for endoglin. In vitro coculture of microvascular ECs with endoglin-positive, CD31-negative islet MSCs reduced VEGFR2 protein expression, disrupted EC angiogenic behavior, and increased EC detachment. Medium conditioned by islet MSCs significantly decreased EC viability and increased EC caspase 3/7 activity. EC:MSC cocultures showed enhanced Smad2 phosphorylation consistent with altered ALK5 signaling. Pharmacological inhibition of ALK5 activity with SB431542 (SB) improved EC survival upon contact with MSCs, and SB-treated cultured islets retained EC marker expression and sensitivity to exogenous VEGF164 . Thus, endoglin-expressing islet MSCs influence EC ALK5 signaling in vitro, which decreases EC viability, and changes in ALK5 activity in whole cultured islets contribute to islet EC loss. Modifying TGF-β signaling may enable maintenance of islet ECs during islet isolation and thus improve islet graft revascularization post-transplantation. Copyright © 2013 AlphaMed Press.

Widespread Non-Hematopoietic Tissue Distribution by Transplanted Human Progenitor Cells with High Aldehyde Dehydrogenase Activity

PubMed Central

Hess, David A.; Craft, Timothy P.; Wirthlin, Louisa; Hohm, Sarah; Zhou, Ping; Eades, William C.; Creer, Michael H.; Sands, Mark S.; Nolta, Jan A.

2011-01-01

Transplanted adult progenitor cells distribute to peripheral organs and can promote endogenous cellular repair in damaged tissues. However, development of cell-based regenerative therapies has been hindered by the lack of pre-clinical models to efficiently assess multiple organ distribution and difficulty defining human cells with regenerative function. After transplantation into beta-glucuronidase (GUSB)-deficient NOD/SCID/MPSVII mice, we characterized the distribution of lineage depleted human umbilical cord blood-derived cells purified by selection using high aldehyde dehydrogenase activity (ALDH) with CD133 co-expression. ALDHhi or ALDHhiCD133+ cells produced robust hematopoietic reconstitution, and variable levels of tissue distribution in multiple organs. GUSB+ donor cells that co-expressed human (HLA-A,B,C) and hematopoietic (CD45+) cell surface markers were the primary cell phenotype found adjacent to the vascular beds of several tissues, including islet and ductal regions of mouse pancreata. In contrast, variable phenotypes were detected in the chimeric liver, with HLA+/CD45+ cells demonstrating robust GUSB expression adjacent to blood vessels, and CD45−/HLA− cells with diluted GUSB expression predominant in the liver parenchyma. However, true non-hematopoietic human (HLA+/CD45−) cells were rarely detected in other peripheral tissues, suggesting that these GUSB+/HLA−/CD45− cells in the liver were a result of downregulated human surface marker expression in vivo, not widespread seeding of non-hematopoietic cells. However, relying solely on continued expression of cell surface markers, as employed in traditional xenotransplantation models, may underestimate true tissue distribution. ALDH-expressing progenitor cells demonstrated widespread and tissue-specific distribution of variable cellular phenotypes, indicating that these adult progenitor cells should be explored in transplantation models of tissue damage. PMID:18055447

CD44 Interacts with HIF-2α to Modulate the Hypoxic Phenotype of Perinecrotic and Perivascular Glioma Cells.

PubMed

Johansson, Elinn; Grassi, Elisa S; Pantazopoulou, Vasiliki; Tong, Bei; Lindgren, David; Berg, Tracy J; Pietras, Elin J; Axelson, Håkan; Pietras, Alexander

2017-08-15

Hypoxia-inducible factors enhance glioma stemness, and glioma stem cells have an amplified hypoxic response despite residing within a perivascular niche. Still, little is known about differential HIF regulation in stem versus bulk glioma cells. We show that the intracellular domain of stem cell marker CD44 (CD44ICD) is released at hypoxia, binds HIF-2α (but not HIF-1α), enhances HIF target gene activation, and is required for hypoxia-induced stemness in glioma. In a glioma mouse model, CD44 was restricted to hypoxic and perivascular tumor regions, and in human glioma, a hypoxia signature correlated with CD44. The CD44ICD was sufficient to induce hypoxic signaling at perivascular oxygen tensions, and blocking CD44 cleavage decreased HIF-2α stabilization in CD44-expressing cells. Our data indicate that the stem cell marker CD44 modulates the hypoxic response of glioma cells and that the pseudo-hypoxic phenotype of stem-like glioma cells is achieved by stabilization of HIF-2α through interaction with CD44, independently of oxygen. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Investigation of Functional Activity of Cells in Granulomatous Inflammatory Lesions from Mice with Latent Tuberculous Infection in the New Ex Vivo Model

PubMed Central

2013-01-01

The new ex vivo model system measuring functional input of individual granuloma cells to formation of granulomatous inflammatory lesions in mice with latent tuberculous infection has been developed and described in the current study. Monolayer cultures of cells that migrated from individual granulomas were established in the proposed culture settings for mouse spleen and lung granulomas induced by in vivo exposure to BCG vaccine. The cellular composition of individual granulomas was analyzed. The expression of the leukocyte surface markers such as phagocytic receptors CD11b, CD11c, CD14, and CD16/CD32 and the expression of the costimulatory molecules CD80, CD83, and CD86 were tested as well as the production of proinflammatory cytokines (IFNγ and IL-1α) and growth factors (GM-CSF and FGFb) for cells of individual granulomas. The colocalization of the phagocytic receptors and costimulatory molecules in the surface microdomains of granuloma cells (with and without acid-fast BCG-mycobacteria) has also been detected. It was found that some part of cytokine macrophage producers have carried acid-fast mycobacteria. Detected modulation in dynamics of production of pro-inflammatory cytokines, growth factors, and leukocyte surface markers by granuloma cells has indicated continued processes of activation and deactivation of granuloma inflammation cells during the latent tuberculous infection progress in mice. PMID:24198843

Helicobacter pylori cagA+ Is Associated with Milder Duodenal Histological Changes in Chilean Celiac Patients.

PubMed

Lucero, Yalda; Oyarzún, Amaya; O'Ryan, Miguel; Quera, Rodrigo; Espinosa, Nelly; Valenzuela, Romina; Simian, Daniela; Alcalde, Elisa; Arce, Claudio; Farfán, Mauricio J; Vergara, Alejandra F; Gajardo, Iván; Mendez, Jocelyn; Carrasco, Jorge; Errázuriz, Germán; Gonzalez, Mónica; Ossa, Juan C; Maiza, Eduardo; Perez-Bravo, Francisco; Castro, Magdalena; Araya, Magdalena

2017-01-01

HIGHLIGHTS What is already known about this subject?Celiac disease (CD) has a high clinical and histological diversity and the mechanisms underlying this phenomenon remain elusive. H. pylori is a bacterium that chronically infect gastric and duodenal mucosa activating both a Th1/Th17 and T-reg pathways.The role of H. pylori (and the effect of their virulence factors) in CD have not yet completely elucidated.What are the new findings? cagA + H. pylori strains are associated to milder histological damage in infected CD patients.In active-CD patients the presence of cagA + H. pylori is associated to an increase in T-reg markers, contrasting with a downregulation in cagA + infected potential-CD individuals.How might it impact on clinical practice in the foreseeable future?The identification of microbiological factors that could modulate inflammation and clinical expression of CD may be used in the future as preventive strategies or as supplementary treatment in patients that cannot achieve complete remission, contributing to the better care of these patients. Background: Mechanisms underlying the high clinical and histological diversity of celiac disease (CD) remain elusive. Helicobacter pylori (Hp) chronically infects gastric and duodenal mucosa and has been associated with protection against some immune-mediated conditions, but its role (specifically of cagA + strains) in CD is unclear. Objective: To assess the relationship between gastric Hp infection ( cagA + strains) and duodenal histological damage in patients with CD. Design: Case-control study including patients with active-CD, potential-CD and non-celiac individuals. Clinical presentation, HLA genotype, Hp/ cagA gene detection in gastric mucosa, duodenal histology, Foxp3 positive cells and TGF-β expression in duodenal lamina propria were analyzed. Results: We recruited 116 patients, 29 active-CD, 37 potential-CD, and 50 non-CD controls. Hp detection was similar in the three groups (~30-40%), but cagA + strains were more common in infected potential-CD than in active-CD (10/11 vs. 4/10; p = 0.020) and non-CD (10/20; p = 0.025). Among active-CD patients, Foxp3 positivity was significantly higher in subjects with cagA + Hp+ compared to cagA- Hp+ ( p < 0.01) and Hp- ( p < 0.01). In cagA+ Hp+ individuals, Foxp3 positivity was also higher comparing active- to potential-CD ( p < 0.01). TGF-β expression in duodenum was similar in active-CD with cagA + Hp+ compared to Hp- and was significantly downregulated in cagA + potential-CD subjects compared to other groups. Conclusion: Hp infection rates were similar among individuals with/without CD, but infection with cagA + strains was associated with milder histological damage in celiac patients infected by Hp, and in active-CD cases with higher expression of T-reg markers. Results suggest that infection by cagA + Hp may be protective for CD progression, or conversely, that these strains are prone to colonize intestinal mucosa with less severe damage.

Increased frequencies of CD8+CD57+ T cells are associated with antibody neutralization breadth against HIV in viraemic controllers

PubMed Central

Palmer, Christine D; Romero-Tejeda, Marisol; Scully, Eileen P; Lockhart, Ainsley; Seaman, Michael S; Goldenthal, Ariel; Piechocka-Trocha, Alicja; Walker, Bruce D; Chibnik, Lori B; Jost, Stephanie; Porichis, Filippos

2016-01-01

Introduction An effective prophylactic vaccine against HIV will need to elicit antibody responses capable of recognizing and neutralizing rapidly evolving antigenic regions. The immunologic milieu associated with development of neutralizing antibody breadth remains to be fully defined. In this study, we sought to identify immunological signatures associated with neutralization breadth in HIV controllers. We applied an immune monitoring approach to analyze markers of T cell and myeloid cell activation by flow cytometry, comparing broad neutralizers with low- and non-neutralizers using multivariate and univariate analyses. Methods Antibody neutralization breadth was determined, and cryopreserved peripheral blood mononuclear cells were stained for T cell and myeloid cell activation markers. Subjects were grouped according to neutralization breadth, and T cell and myeloid cell activation was analyzed by partial least squares discriminant analysis to determine immune signatures associated with high neutralization breadth. Results We show that neutralization breadth in HIV viraemic controllers (VC) was strongly associated with increased frequencies of CD8+CD57+ T cells and that this association was independent of viral load, CD4 count and time since HIV diagnosis. Conclusions Our data show elevated frequencies of CD8+CD57+ T cells in VC who develop neutralization breadth against HIV. This immune signature could serve as a potential biomarker of neutralization breadth and should be further investigated in other HIV-positive cohorts and in HIV vaccine trials. PMID:27938646

Peripheral T-Cell Reactivity to Heat Shock Protein 70 and Its Cofactor GrpE from Tropheryma whipplei Is Reduced in Patients with Classical Whipple's Disease

PubMed Central

Trotta, Lucia; Weigt, Kathleen; Schinnerling, Katina; Geelhaar-Karsch, Anika; Oelkers, Gerrit; Biagi, Federico; Corazza, Gino Roberto; Allers, Kristina; Schneider, Thomas; Erben, Ulrike

2017-01-01

ABSTRACT Classical Whipple's disease (CWD) is characterized by the lack of specific Th1 response toward Tropheryma whipplei in genetically predisposed individuals. The cofactor GrpE of heat shock protein 70 (Hsp70) from T. whipplei was previously identified as a B-cell antigen. We tested the capacity of Hsp70 and GrpE to elicit specific proinflammatory T-cell responses. Peripheral mononuclear cells from CWD patients and healthy donors were stimulated with T. whipplei lysate or recombinant GrpE or Hsp70 before levels of CD40L, CD69, perforin, granzyme B, CD107a, and gamma interferon (IFN-γ) were determined in T cells by flow cytometry. Upon stimulation with total bacterial lysate or recombinant GrpE or Hsp70 of T. whipplei, the proportions of activated effector CD4+ T cells, determined as CD40L+ IFN-γ+, were significantly lower in patients with CWD than in healthy controls; CD8+ T cells of untreated CWD patients revealed an enhanced activation toward unspecific stimulation and T. whipplei-specific degranulation, although CD69+ IFN-γ+ CD8+ T cells were reduced upon stimulation with T. whipplei lysate and recombinant T. whipplei-derived proteins. Hsp70 and its cofactor GrpE are immunogenic in healthy individuals, eliciting effective responses against T. whipplei to control bacterial spreading. The lack of specific T-cell responses against these T. whipplei-derived proteins may contribute to the pathogenesis of CWD. PMID:28559404

Peripheral T-Cell Reactivity to Heat Shock Protein 70 and Its Cofactor GrpE from Tropheryma whipplei Is Reduced in Patients with Classical Whipple's Disease.

PubMed

Trotta, Lucia; Weigt, Kathleen; Schinnerling, Katina; Geelhaar-Karsch, Anika; Oelkers, Gerrit; Biagi, Federico; Corazza, Gino Roberto; Allers, Kristina; Schneider, Thomas; Erben, Ulrike; Moos, Verena

2017-08-01

Classical Whipple's disease (CWD) is characterized by the lack of specific Th1 response toward Tropheryma whipplei in genetically predisposed individuals. The cofactor GrpE of heat shock protein 70 (Hsp70) from T. whipplei was previously identified as a B-cell antigen. We tested the capacity of Hsp70 and GrpE to elicit specific proinflammatory T-cell responses. Peripheral mononuclear cells from CWD patients and healthy donors were stimulated with T. whipplei lysate or recombinant GrpE or Hsp70 before levels of CD40L, CD69, perforin, granzyme B, CD107a, and gamma interferon (IFN-γ) were determined in T cells by flow cytometry. Upon stimulation with total bacterial lysate or recombinant GrpE or Hsp70 of T. whipplei , the proportions of activated effector CD4 + T cells, determined as CD40L + IFN-γ + , were significantly lower in patients with CWD than in healthy controls; CD8 + T cells of untreated CWD patients revealed an enhanced activation toward unspecific stimulation and T. whipplei -specific degranulation, although CD69 + IFN-γ + CD8 + T cells were reduced upon stimulation with T. whipplei lysate and recombinant T. whipplei -derived proteins. Hsp70 and its cofactor GrpE are immunogenic in healthy individuals, eliciting effective responses against T. whipplei to control bacterial spreading. The lack of specific T-cell responses against these T. whipplei -derived proteins may contribute to the pathogenesis of CWD. Copyright © 2017 American Society for Microbiology.

CCR5+ CD8 T-cell levels and monocyte activation precede the onset of acute coronary syndrome in HIV-infected patients on antiretroviral therapy.

PubMed

Tarancon-Diez, Laura; De Pablo-Bernal, Rebeca S; Álvarez-Rios, Ana I; Rosado-Sánchez, Isaac; Dominguez-Molina, Beatriz; Genebat, Miguel; Pacheco, Yolanda M; Jiménez, José Luis; Muñoz-Fernández, M Ángeles; Ruiz-Mateos, Ezequiel; Leal, Manuel

2017-06-02

Acute coronary syndrome (ACS) is nowadays one of the leading causes of morbid-mortality in HIV-infected population, but innate and adaptive immune mechanisms preceding this event are unknown. In this work we comprehensively and longitudinally observed, by multiparametric flow cytometry and following a case-control design, increased CCR5 + CD8 + T-cells levels and monocytes expressing activation and adhesion markers in HIV-infected patients who are going to suffer ACS. In addition, we found direct associations between activated CD8 + T-cells and myeloid cells that were only statistically significant in the group of patients with ACS and in the follow up time point just before the ACS. Our data highlight the important role of CCR5 in the onset of ACS and suggest this receptor as a marker of cardiovascular risk and potential therapeutic target to prevent the development of such non-AIDS-related event in HIV-infected patients.

[Expression of embryonic markers in pterygium derived mesenchymal cells].

PubMed

Pascual, G; Montes, M A; Pérez-Rico, C; Pérez-Kohler, B; Bellón, J M; Buján, J

2010-12-01

Destruction of the limbal epithelium barrier is the most important mechanism of pterygium formation (conjunctiva proliferation, encroaching onto the cornea). It is thought to arise from activated and proliferating limbal epithelial stem cells. The objective of this study is to evaluate the presence of undifferentiated mesenchymal cells (stem cells) in cultured cells extracted from human pterygium. Cells from 6 human pterygium were isolated by explantation and placed in cultures with amniomax medium. Once the monolayer was reached the cells were seeded onto 24 well microplates. The cells were studied in the second sub-culture. The immunohistochemical expression of different embryonic stem cell markers, OCT3/4 and CD9, was analysed. The differentiated phenotypes were characterised with the monoclonal antibodies anti-CD31, α-actin and vimentin. All the cell populations obtained from pterygium showed vimentin expression. Less than 1% of the cells were positive for CD31 and α-actin markers. The majority of the cell population was positive for OCT3/4 and CD9. The cell population obtained from pterygium expressed mesenchymal cell phenotype and embryonic markers, such us OCT3/4 and CD9. This undifferentiated population could be involved in the large recurrence rate of this type of tissue after surgery. Copyright © 2010 Sociedad Española de Oftalmología. Published by Elsevier Espana. All rights reserved.

Temporal Characterization of Microglia/Macrophage Phenotypes in a Mouse Model of Neonatal Hypoxic-Ischemic Brain Injury

PubMed Central

Hellström Erkenstam, Nina; Smith, Peter L. P.; Fleiss, Bobbi; Nair, Syam; Svedin, Pernilla; Wang, Wei; Boström, Martina; Gressens, Pierre; Hagberg, Henrik; Brown, Kelly L.; Sävman, Karin; Mallard, Carina

2016-01-01

Immune cells display a high degree of phenotypic plasticity, which may facilitate their participation in both the progression and resolution of injury-induced inflammation. The purpose of this study was to investigate the temporal expression of genes associated with classical and alternative polarization phenotypes described for macrophages and to identify related cell populations in the brain following neonatal hypoxia-ischemia (HI). HI was induced in 9-day old mice and brain tissue was collected up to 7 days post-insult to investigate expression of genes associated with macrophage activation. Using cell-markers, CD86 (classic activation) and CD206 (alternative activation), we assessed temporal changes of CD11b+ cell populations in the brain and studied the protein expression of the immunomodulatory factor galectin-3 in these cells. HI induced a rapid regulation (6 h) of genes associated with both classical and alternative polarization phenotypes in the injured hemisphere. FACS analysis showed a marked increase in the number of CD11b+CD86+ cells at 24 h after HI (+3667%), which was coupled with a relative suppression of CD11b+CD206+ cells and cells that did not express neither CD86 nor CD206. The CD11b+CD206+ population was mixed with some cells also expressing CD86. Confocal microscopy confirmed that a subset of cells expressed both CD86 and CD206, particularly in injured gray and white matter. Protein concentration of galectin-3 was markedly increased mainly in the cell population lacking CD86 or CD206 in the injured hemisphere. These cells were predominantly resident microglia as very few galectin-3 positive cells co-localized with infiltrating myeloid cells in Lys-EGFP-ki mice after HI. In summary, HI was characterized by an early mixed gene response, but with a large expansion of mainly the CD86 positive population during the first day. However, the injured hemisphere also contained a subset of cells expressing both CD86 and CD206 and a large population that expressed neither activation marker CD86 nor CD206. Interestingly, these cells expressed the highest levels of galectin-3 and were found to be predominantly resident microglia. Galectin-3 is a protein involved in chemotaxis and macrophage polarization suggesting a novel role in cell infiltration and immunomodulation for this cell population after neonatal injury. PMID:28018179

Bi-directional activation between mesenchymal stem cells and CLL B-cells: implication for CLL disease progression.

PubMed

Ding, Wei; Nowakowski, Grzegorz S; Knox, Traci R; Boysen, Justin C; Maas, Mary L; Schwager, Susan M; Wu, Wenting; Wellik, Linda E; Dietz, Allan B; Ghosh, Asish K; Secreto, Charla R; Medina, Kay L; Shanafelt, Tait D; Zent, Clive S; Call, Timothy G; Kay, Neil E

2009-11-01

It was hypothesized that contact between chronic lymphocytic leukaemia (CLL) B-cells and marrow stromal cells impact both cell types. To test this hypothesis, we utilized a long-term primary culture system from bone biopsies that reliably generates a mesenchymal stem cell (MSC). Co-culture of MSC with CLL B-cells protected the latter from both spontaneous apoptosis and drug-induced apoptosis. The CD38 expression in previously CD38 positive CLL B-cells was up-regulated with MSC co-culture. Upregulation of CD71, CD25, CD69 and CD70 in CLL B-cells was found in the co-culture. CD71 upregulation was more significantly associated with high-risk CLL, implicating CD71 regulation in the microenvironment predicting disease progression. In MSC, rapid ERK and AKT phosphorylation (within 30 min) were detected when CLL B-cells and MSC were separated by transwell; indicating that activation of MSC was mediated by soluble factors. These findings support a bi-directional activation between bone marrow stromal cells and CLL B-cells.

Release of soluble CD30 after allogeneic stimulation is mediated by memory T cells and regulated by IFN-γ and IL-2.

PubMed

Velásquez, Sonia Y; García, Luis F; Opelz, Gerhard; Alvarez, Cristiam M; Süsal, Caner

2013-07-27

Membrane CD30 is an important costimulatory molecule for activated T lymphocytes, and serum level of soluble CD30 (sCD30) is considered a marker for predicting outcome in kidney transplantation. We investigated the kinetics of CD30 expression on CD4 and CD8 T-cell populations and the source of sCD30 during alloimmune responses in vitro. The effect of neutralizing antibodies against interferon (IFN)-γ and other cytokines on sCD30 release and the involvement of metalloproteinases ADAM10 and ADAM17/TACE that are responsible for sCD30 shedding were also assessed. Memory phenotypes and CD30 expression on allostimulated CD3 lymphocytes were evaluated in dialysis patients and matched controls. Allogeneic stimulation resulted in conversion of naive responder cells to central memory CD4 cells (P<0.001 at 96 hr) and effector CD8 cells (P<0.01 at 120 hr), which was accompanied by increased CD30 expression. Release of sCD30 was attributed mainly to central memory cells, and neutralization of IFN-γ (P<0.001) and interleukin (IL)-2 (P<0.001) impaired the release of sCD30 during allostimulation but did not alter the levels of ADAM10 and ADAM17/TACE. CD30 expression was modulated in dialysis patients in a similar way as in healthy controls. Allostimulation results in the up-regulation of the T-cell activation marker CD30 on CD4 as well as CD8 memory T cells and increased release of sCD30 from these cells in an IFN-γ- and IL-2-dependent manner. These results may explain clinical findings on the suitability of sCD30 and IFN-γ- and IL-2-producing T cells for immune monitoring of kidney transplant recipients before and after transplantation.

CD26 expression and adenosine deaminase activity in regulatory T cells (Treg) and CD4+ T effector cells in patients with head and neck squamous cell carcinoma

PubMed Central

Mandapathil, Magis; Szczepanski, Miroslaw; Harasymczuk, Malgorzata; Ren, Jin; Cheng, Dongmei; Jackson, Edwin K.; Gorelik, Elieser; Johnson, Jonas; Lang, Stephan; Whiteside, Theresa L

2012-01-01

Adenosine deaminase (ADA) is responsible for the deamination of immunosuppressive adenosine to inosine. In human T lymphocytes, ADA is associated with dipeptidyl peptidase IV (CD26). ADA expression and activity were evaluated in regulatory T cells (Treg) and CD4+ T effector cells (Teff) of patients with head and neck squamous cell cancer (HNSCC). CD4+CD39+ and CD4+CD39neg T cells were isolated by single-cell sorting from the peripheral blood of 15 HNSCC patients and 15 healthy donors (NC). CD26/ADA expression in these cells was studied by multicolor flow cytometry, confocal microscopy, RT-PCR and immunohistochemistry in tumor tissues. ADA activity was evaluated by mass spectrometry, suppression of Teff proliferation in CFSE assays and cytokine production by Luminex. CD4+CD39+ Treg had low and CD4+CD39neg Teff high CD26/ADA expression and ADA activity in NC or HNSCC. The frequency and suppressor activity of CD39+CD26neg Treg were elevated in patients relative to NC (p < 0.01). However, ADA activity in patients’ CD4+CD39neg Teff was decreased (p < 0.05), resulting in extracellular adenosine accumulation. Also, patients’ Teff were more sensitive to inhibitory signals delivered via adenosine receptors. IL-2, IL12 and INFγ upregulated ADA expression and activity in CD4+CD39neg Teff, whereas IL-10, PGE2 and CADO downregulated it. The differentially expressed CD26/ADA can serve as surface markers for functionally-active CD39+CD26neg Treg. PMID:22934258

CD8+ T Lymphocyte Expansion, Proliferation and Activation in Dengue Fever

PubMed Central

de Matos, Andréia Manso; Carvalho, Karina Inacio; Rosa, Daniela Santoro; Villas-Boas, Lucy Santos; da Silva, Wanessa Cardoso; Rodrigues, Célia Luiza de Lima; Oliveira, Olímpia Massae Nakasone Peel Furtado; Levi, José Eduardo; Araújo, Evaldo Stanislau Affonso; Pannuti, Claudio Sergio; Luna, Expedito José Albuquerque; Kallas, Esper George

2015-01-01

Dengue fever induces a robust immune response, including massive T cell activation. The level of T cell activation may, however, be associated with more severe disease. In this study, we explored the level of CD8+ T lymphocyte activation in the first six days after onset of symptoms during a DENV2 outbreak in early 2010 on the coast of São Paulo State, Brazil. Using flow cytometry we detected a progressive increase in the percentage of CD8+ T cells in 74 dengue fever cases. Peripheral blood mononuclear cells from 30 cases were thawed and evaluated using expanded phenotyping. The expansion of the CD8+ T cells was coupled with increased Ki67 expression. Cell activation was observed later in the course of disease, as determined by the expression of the activation markers CD38 and HLA-DR. This increased CD8+ T lymphocyte activation was observed in all memory subsets, but was more pronounced in the effector memory subset, as defined by higher CD38 expression. Our results show that most CD8+ T cell subsets are expanded during DENV2 infection and that the effector memory subset is the predominantly affected sub population. PMID:25675375

Accessory cells with a veiled morphology and movement pattern generated from monocytes after avoidance of plastic adherence and of NADPH oxidase activation. A comparison with GM-CSF/IL-4-induced monocyte-derived dendritic cells.

PubMed

Ruwhof, Cindy; Canning, Martha O; Grotenhuis, Kristel; de Wit, Harm J; Florencia, Zenovia Z; de Haan-Meulman, Meeny; Drexhage, Hemmo A

2002-07-01

Veiled cells (VC) present in afferent lymph transport antigen from the periphery to the draining lymph nodes. Although VC in lymph form a heterogeneous population, some of the cells clearly belong on morphological grounds to the Langerhans cell (LC)/ dendritic cell (DC) series. Here we show that culturing monocytes for 24 hrs while avoiding plastic adherence (polypropylene tubes) and avoiding the activation of NADPH oxidase (blocking agents) results in the generation of a population of veiled accessory cells. The generated VC were actively moving cells like lymph-borne VC in vivo. The monocyte (mo)-derived VC population existed of CD14(dim/-) and CD14(brighT) cells. Of these the CD14(dim/-) VC were as good in stimulating allogeneic T cell proliferation as immature DC (iDC) obtained after one week of adherent culture of monocytes in granulocyte-macrophage-colony stimulating factor (GM-CSF)/interleukin (IL)-4. This underscores the accessory cell function of the mo-derived CD14(dim/-) VC. Although the CD14(dim/-)VC had a modest expression of the DC-specific marker CD83 and were positive for S100, expression of the DC-specific markers CD1a, Langerin, DC-SIGN, and DC-LAMP were absent. This indicates that the here generated CD14(dim/-) VC can not be considered as classical LC/DC. It was also impossible to turn the CD14(dim/-) mo-derived VC population into typical DC by culture for one week in GM-CSF/IL-4 or LPS. In fact the cells died tinder such circumstances, gaining some macrophage characteristics before dying. The IL-12 production from mo-derived CD14(dim/-) VC was lower, whereas the production of IL-10 was higher as compared to iDC. Consequently the T cells that were stimulated by these mo-derived VC produced less IFN-gamma as compared with T cells stimulated by iDC. Our data indicate that it is possible to rapidly generate a population of CD14(dim/-) veiled accessory cells from monocytes. The marker pattern and cytokine production of these VC indicate that this population is not a classical DC population. The cells might earlier be related to the veiled macrophage-like cells also earlier described in afferent lymph.

Co-Expression of Putative Cancer Stem Cell Markers CD44 and CD133 in Prostate Carcinomas.

PubMed

Kalantari, Elham; Asgari, Mojgan; Nikpanah, Seyedehmoozhan; Salarieh, Naghme; Asadi Lari, Mohammad Hossein; Madjd, Zahra

2017-10-01

Cancer stem cells (CSCs) are the main players of prostate tumorigenesis thus; characterization of CSCs can pave the way for understanding the early detection, drug resistance, metastasis and relapse. The current study was conducted to evaluate the expression level and clinical significance of the potential CSC markers CD44 and CD133 in a series of prostate tissues. One hundred and forty eight prostate tissues composed of prostate cancer (PCa), high-grade prostatic intraepithelial neoplasia (HGPIN), and benign prostate hyperplasia (BPH) were immunostained for the putative CSC markers CD44 and CD133. Subsequently, the correlation between the expression of these markers and the clinicopathological variables was examined. A higher level of CD44 expression was observed in 42% of PCa, 57% of HGPIN, and 42% BPH tissues. In the case of CD133 expression PCa, HGPIN, and BPH samples demonstrated high immunoreactivity in 46%, 43%, and 42% of cells, respectively. Statistical analysis showed an inverse significant correlation between CD44 expression with Gleason score of PCa (P = 0.02), while no significant correlation was observed between CD133 expression and clinicopathological parameters. A significant reciprocal correlation was observed between the expression of two putative CSC markers CD44 and CD133 in PCa specimens while not indicating clinical significance. Further clinical investigation is required to consider these markers as targets of new therapeutic strategies for PCa.

Reduced CD5(+) CD24(hi) CD38(hi) and interleukin-10(+) regulatory B cells in active anti-neutrophil cytoplasmic autoantibody-associated vasculitis permit increased circulating autoantibodies.

PubMed

Aybar, L T; McGregor, J G; Hogan, S L; Hu, Y; Mendoza, C E; Brant, E J; Poulton, C J; Henderson, C D; Falk, R J; Bunch, D O

2015-05-01

Pathogenesis of anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis is B cell-dependent, although how particular B cell subsets modulate immunopathogenesis remains unknown. Although their phenotype remains controversial, regulatory B cells (Bregs ), play a role in immunological tolerance via interleukin (IL)-10. Putative CD19(+) CD24(hi) CD38(hi) and CD19(+) CD24(hi) CD27(+) Bregs were evaluated in addition to their CD5(+) subsets in 69 patients with ANCA-associated vasculitis (AAV). B cell IL-10 was verified by flow cytometry following culture with CD40 ligand and cytosine-phosphate-guanosine (CpG) DNA. Patients with active disease had decreased levels of CD5(+) CD24(hi) CD38(hi) B cells and IL-10(+) B cells compared to patients in remission and healthy controls (HCs). As IL-10(+) and CD5(+) CD24(hi) CD38(hi) B cells normalized in remission within an individual, ANCA titres decreased. The CD5(+) subset of CD24(hi) CD38(hi) B cells decreases in active disease and rebounds during remission similarly to IL-10-producing B cells. Moreover, CD5(+) B cells are enriched in the ability to produce IL-10 compared to CD5(neg) B cells. Together these results suggest that CD5 may identify functional IL-10-producing Bregs . The malfunction of Bregs during active disease due to reduced IL-10 expression may thus permit ANCA production. © 2014 British Society for Immunology.

Evaluation of CD44 and CD133 as markers of liver cancer stem cells in Egyptian patients with HCV-induced chronic liver diseases versus hepatocellular carcinoma

PubMed Central

Rozeik, Mohammed Saeed; Hammam, Olfat Ali; Ali, Ali Ibrahim; Magdy, Mona; Khalil, Heba; Anas, Amgad; Abo el Hassan, Ahmed Abdelaleem; Rahim, Ali Abdel; El-Shabasy, Ahmed Ibrahim

2017-01-01

Background Cancer stem cells (CSCs) play a critical role in tumor development, progression, metastasis and recurrence. Aim To evaluate hepatic expression of CD44 and CD133 in Egyptian patients with HCV-induced chronic liver diseases and hepatocellular carcinomas (HCCs), and to assess its correlation with inflammatory activity scores, stages of fibrosis (in chronic hepatitis with or without cirrhosis) and grades of HCC. Methods This prospective case-control study was conducted on eighty subjects who attended the Tropical Diseases Department, Al-Azhar University Hospital, and in collaboration with Theodor Bilharz Research Institute (2014–2016). They were divided as follows: A) Control healthy group: Ten individuals with serologically negative HCV-Ab and HBsAg, and histopathologically normal liver, B) Seventy patients subdivided into 3 groups; Twenty subjects each, as: HCV-Ab+ non-cirrhotic, HCV-Ab+ cirrhotic and HCC. Necroinflammatory activity and fibrosis in non-neoplastic liver biopsies were scored according to the METAVIR scoring system. CD44 and CD133 immunostaining was evaluated in all groups semi-quantitatively using H score. Statistical analysis was performed by SPSS version 22, using independent-samples t-test. Results Our study showed a significant increase of mean CD44 & CD133 expression values with disease progression among the groups (p<0.05). Their expressions increased significantly with the inflammatory activity scores and stages of fibrosis, reaching the highest values in A3F4 score compared to A1F1 (p<0.05). Moreover, there was a significant increase of their expressions across HCC grades (p<0.05), however with no significant correlation with focal lesions size. Conclusion CSCs clusters exhibiting CD133+ and/or CD44+ profiles were identified in chronic hepatitis, liver cirrhosis and HCC. CD133 and CD44 expressions significantly corresponded to the increased inflammatory activity, fibrosis stages and higher tumor grades. Therefore, evaluation of CD44 and CD133 expression profiles as CSCs markers in non-neoplastic liver and HCCs can help in development of novel therapeutic agents for HCC targeting and prevention. PMID:28894525

Rapid and preferential distribution of blood-borne αCD3εAb to the liver is followed by local stimulation of T cells and natural killer T cells

PubMed Central

Wingender, Gerhard; Schumak, Beatrix; Schurich, Anna; Gessner, J Engelbert; Endl, Elmar; Limmer, Andreas; Knolle, Percy A

2006-01-01

Dissemination of soluble molecules or antigens via the blood stream is considered to lead to a uniform distribution in the various organs of the body, but organ-specific microarchitecture and vascularization may influence this. Following intravenous injection of αCD3ε antibody (αCD3εAb) we observed clear differences in antibody binding to Fcγ receptor (FcγR)+ antigen-presenting cells (APCs) or T lymphocytes in different organs. Significant binding of blood-borne αCD3εAb was only detected in the spleen and liver and not in the thymus or lymph node. In the spleen, only 10% of dendritic cells/macrophages and 40% of T-cell receptor (TCR)-β+ cells were positive for αCD3εAb, and, dependent on FcγR-mediated cross-linking of αCD3εAb, a similar percentage of splenic TCR-β+ cells were stimulated and became CD69+. Stimulation of TCR-β+ cells in the liver was at least as efficient as in the spleen, but almost all T cells and all scavenger liver sinusoidal endothelial cells bound αCD3εAb. In contrast to CD69 up-regulation, only CD4+ natural killer T (NKT) cells and CD11ahigh CD8+ T cells were activated by αCD3εAb and expressed interferon (IFN)-γ. Again, IFN-γ release from NKT/T cells was at least as efficient in the liver as in the spleen. Taken together, our results support the notion that the combination of extensive hepatic vascularization and very high scavenger activity allows the liver to fulfill its metabolic tasks and to promote stimulation of the large but widely distributed hepatic population of NKT/T cells. PMID:16423047

Helios, and not FoxP3, is the marker of activated Tregs expressing GARP/LAP.

PubMed

Elkord, Eyad; Abd Al Samid, May; Chaudhary, Belal

2015-08-21

Regulatory T cells (Tregs) are key players of immune regulation/dysregulation both in physiological and pathophysiological settings. Despite significant advances in understanding Treg function, there is still a pressing need to define reliable and specific markers that can distinguish different Treg subpopulations. Herein we show for the first time that markers of activated Tregs [latency associated peptide (LAP) and glycoprotein A repetitions predominant (GARP, or LRRC32)] are expressed on CD4+FoxP3- T cells expressing Helios (FoxP3-Helios+) in the steady state. Following TCR activation, GARP/LAP are up-regulated on CD4+Helios+ T cells regardless of FoxP3 expression (FoxP3+/-Helios+). We show that CD4+GARP+/-LAP+ Tregs make IL-10 immunosuppressive cytokine but not IFN-γ effector cytokine. Further characterization of FoxP3/Helios subpopulations showed that FoxP3+Helios+ Tregs proliferate in vitro significantly less than FoxP3+Helios- Tregs upon TCR stimulation. Unlike FoxP3+Helios- Tregs, FoxP3+Helios+ Tregs secrete IL-10 but not IFN-γ or IL-2, confirming they are bona fide Tregs with immunosuppressive characteristics. Taken together, Helios, and not FoxP3, is the marker of activated Tregs expressing GARP/LAP, and FoxP3+Helios+ Tregs have more suppressive characteristics, compared with FoxP3+Helios- Tregs. Our work implies that therapeutic modalities for treating autoimmune and inflammatory diseases, allergies and graft rejection should be designed to induce and/or expand FoxP3+Helios+ Tregs, while therapies against cancers or infectious diseases should avoid such expansion/induction.

Full-length soluble CD147 promotes MMP-2 expression and is a potential serological marker in detection of hepatocellular carcinoma

PubMed Central

2014-01-01

Background As a surface glycoprotein, CD147 is capable of stimulating the production of matrix metalloproteinases (MMPs) from neighboring fibroblasts. The aim of the present study is to explore the role of soluble CD147 on MMPs secretion from hepatocellular carcinoma (HCC) cells, and to investigate the diagnostic value of serum soluble CD147 in the HCC detection. Methods We identified the form of soluble CD147 in cell culture supernate of HCC cells and serum of patients with HCC, and explored the role of soluble CD147 on MMPs secretion. Serum CD147 levels were detected by the enzyme-linked immunosorbent assay, and the value of soluble CD147 as a marker in HCC detection was analyzed. Results Full length soluble CD147 was presented in the culture medium of HCC cells and serum of patients with HCC. The extracellular domain of soluble CD147 promoted the expression of CD147 and MMP-2 from HCC cells. Knockdown of CD147 markedly diminished the up-regulation of CD147 and MMP-2 which induced by soluble CD147. Soluble CD147 activated ERK, FAK, and PI3K/Akt pathways, leading to the up-regulation of MMP-2. The level of soluble CD147 in serum of patients with HCC was significantly elevated compared with healthy individuals (P < 0.001). Soluble CD147 levels were found to be associated with HCC tumor size (P = 0.007) and Child-Pugh grade (P = 0.007). Moreover, soluble CD147 showed a better performance in distinguishing HCC compared with alpha-fetoprotein. Conclusions The extracellular domain of soluble CD147 enhances the secretion of MMP-2 from HCC cells, requiring the cooperation of membrane CD147 and activation of ERK, FAK, and PI3K/Akt signaling. The measurement of soluble CD147 may offer a useful approach in diagnosis of HCC. PMID:24996644

Full-length soluble CD147 promotes MMP-2 expression and is a potential serological marker in detection of hepatocellular carcinoma.

PubMed

Wu, Jiao; Hao, Zhi-Wei; Zhao, You-Xu; Yang, Xiang-Min; Tang, Hao; Zhang, Xin; Song, Fei; Sun, Xiu-Xuan; Wang, Bin; Nan, Gang; Chen, Zhi-Nan; Bian, Huijie

2014-07-04

As a surface glycoprotein, CD147 is capable of stimulating the production of matrix metalloproteinases (MMPs) from neighboring fibroblasts. The aim of the present study is to explore the role of soluble CD147 on MMPs secretion from hepatocellular carcinoma (HCC) cells, and to investigate the diagnostic value of serum soluble CD147 in the HCC detection. We identified the form of soluble CD147 in cell culture supernate of HCC cells and serum of patients with HCC, and explored the role of soluble CD147 on MMPs secretion. Serum CD147 levels were detected by the enzyme-linked immunosorbent assay, and the value of soluble CD147 as a marker in HCC detection was analyzed. Full length soluble CD147 was presented in the culture medium of HCC cells and serum of patients with HCC. The extracellular domain of soluble CD147 promoted the expression of CD147 and MMP-2 from HCC cells. Knockdown of CD147 markedly diminished the up-regulation of CD147 and MMP-2 which induced by soluble CD147. Soluble CD147 activated ERK, FAK, and PI3K/Akt pathways, leading to the up-regulation of MMP-2. The level of soluble CD147 in serum of patients with HCC was significantly elevated compared with healthy individuals (P < 0.001). Soluble CD147 levels were found to be associated with HCC tumor size (P = 0.007) and Child-Pugh grade (P = 0.007). Moreover, soluble CD147 showed a better performance in distinguishing HCC compared with alpha-fetoprotein. The extracellular domain of soluble CD147 enhances the secretion of MMP-2 from HCC cells, requiring the cooperation of membrane CD147 and activation of ERK, FAK, and PI3K/Akt signaling. The measurement of soluble CD147 may offer a useful approach in diagnosis of HCC.

Effector cell signature in peripheral blood following nasal allergen challenge in grass pollen allergic individuals.

PubMed

Shamji, M H; Bellido, V; Scadding, G W; Layhadi, J A; Cheung, D K M; Calderon, M A; Asare, A; Gao, Z; Turka, L A; Tchao, N; Togias, A; Phippard, D; Durham, S R

2015-02-01

Several studies have demonstrated the time course of inflammatory mediators in nasal fluids following nasal allergen challenge (NAC), whereas the effects of NAC on cells in the periphery are unknown. We examined the time course of effector cell markers (for basophils, dendritic cells and T cells) in peripheral blood after nasal grass pollen allergen challenge. Twelve participants with seasonal allergic rhinitis underwent a control (diluent) challenge followed by NAC after an interval of 14 days. Nasal symptoms and peak nasal inspiratory flow (PNIF) were recorded along with peripheral basophil, T-cell and dendritic cell responses (flow cytometry), T-cell proliferative responses (thymidine incorporation), and cytokine expression (FluoroSpot assay). Robust increases in nasal symptoms and decreases in PNIF were observed during the early (0-1 h) response and modest significant changes during the late (1-24 h) response. Sequential peaks in peripheral blood basophil activation markers were observed (CD107a at 3 h, CD63 at 6 h, and CD203c(bright) at 24 h). T effector/memory cells (CD4(+) CD25(lo) ) were increased at 6 h and accompanied by increases in CD80(+) and CD86(+) plasmacytoid dendritic cells (pDCs). Ex vivo grass antigen-driven T-cell proliferative responses and the frequency of IL-4(+) CD4(+) T cells were significantly increased at 6 h after NAC when compared to the control day. Basophil, T-cell, and dendritic cell activation increased the frequency of allergen-driven IL-4(+) CD4(+) T cells, and T-cell proliferative responses are detectable in the periphery after NAC. These data confirm systemic cellular activation following a local nasal provocation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Oxidative stress induced by cadmium in the plasma, erythrocytes and lymphocytes of rats: Attenuation by grape seed proanthocyanidins.

PubMed

Nazima, B; Manoharan, V; Miltonprabu, S

2016-04-01

The present study has been designed to investigate the ameliorative effect of grape seed proanthocyanidins (GSP) on cadmium (Cd)-induced oxidative damage in rat erythrocytes. Twenty four male Wistar rats were divided into four groups: control, GSP-treated group (100 mg kg(-1) body weight (BW)), Cd-treated group (cadmium chloride, 5 mg kg(-1) BW), and GSP + Cd-treated group in which GSP was orally pre-administered 90 min before Cd intoxication for 4 weeks. At the end of the experimental period, blood samples were collected by cardiac puncture and were processed for various biochemical estimations. The extent of oxidative damage in isolated rat erythrocyte membrane was assessed by measuring lipid peroxidation, enzymatic and non-enzymatic content, calcium ion (Ca(2+))/magnesium ion (Mg(2+))-ATPase and sodium ion (Na(+))/potassium ion (K(+))-ATPase activities, free iron, calcium, hydrogen peroxide (H2O2) concentration, and osmotic fragility. Our results unveiled that Cd intoxication significantly increased the erythrocyte lipid peroxidation markers and decreased the activity of enzymatic and non-enzymatic markers in erythrocytes. Conversely, GSP pretreatment significantly prevented the decrease in the activities of antioxidant enzymes and membrane-bound ATPases. GSP also restored the levels of iron, calcium, and H2O2 in Cd-treated rats. Conformational changes in erythrocytes of various groups were also determined using morphological and ultrastructural electron microscopic analysis. The findings of our study clearly revealed that GSP affords superior protection against Cd-induced reactive oxygen species generation, lipid peroxidation, and free radical generation in Cd-treated rats, which presumably reflects the ability of this flavonoid to protect erythrocytes and lymphocytes of rats from the toxic effects of Cd. © The Author(s) 2015.

An abundant tissue macrophage population in the adult murine heart with a distinct alternatively-activated macrophage profile.

PubMed

Pinto, Alexander R; Paolicelli, Rosa; Salimova, Ekaterina; Gospocic, Janko; Slonimsky, Esfir; Bilbao-Cortes, Daniel; Godwin, James W; Rosenthal, Nadia A

2012-01-01

Cardiac tissue macrophages (cTMs) are a previously uncharacterised cell type that we have identified and characterise here as an abundant GFP(+) population within the adult Cx(3)cr1(GFP/+) knock-in mouse heart. They comprise the predominant myeloid cell population in the myocardium, and are found throughout myocardial interstitial spaces interacting directly with capillary endothelial cells and cardiomyocytes. Flow cytometry-based immunophenotyping shows that cTMs exhibit canonical macrophage markers. Gene expression analysis shows that cTMs (CD45(+)CD11b(+)GFP(+)) are distinct from mononuclear CD45(+)CD11b(+)GFP(+) cells sorted from the spleen and brain of adult Cx(3)cr1(GFP/+) mice. Gene expression profiling reveals that cTMs closely resemble alternatively-activated anti-inflammatory M2 macrophages, expressing a number of M2 markers, including Mrc1, CD163, and Lyve-1. While cTMs perform normal tissue macrophage homeostatic functions, they also exhibit a distinct phenotype, involving secretion of salutary factors (including IGF-1) and immune modulation. In summary, the characterisation of cTMs at the cellular and molecular level defines a potentially important role for these cells in cardiac homeostasis.

CD147 (Basigin/Emmprin) identifies FoxP3+CD45RO+CTLA4+-activated human regulatory T cells.

PubMed

Solstad, Therese; Bains, Simer Jit; Landskron, Johannes; Aandahl, Einar Martin; Thiede, Bernd; Taskén, Kjetil; Torgersen, Knut Martin

2011-11-10

Human CD4(+)FoxP3(+) T cells are functionally and phenotypically heterogeneous providing plasticity to immune activation and regulation. To better understand the functional dynamics within this subset, we first used a combined strategy of subcellular fractionation and proteomics to describe differences at the protein level between highly purified human CD4(+)CD25(+) and CD4(+)CD25(-) T-cell populations. This identified a set of membrane proteins highly expressed on the cell surface of human regulatory T cells (Tregs), including CD71, CD95, CD147, and CD148. CD147 (Basigin or Emmprin) divided CD4(+)CD25(+) cells into distinct subsets. Furthermore, CD147, CD25, FoxP3, and in particular CTLA-4 expression correlated. Phenotypical and functional analyses suggested that CD147 marks the switch between resting (CD45RA(+)) and activated (CD45RO(+)) subsets within the FoxP3(+) T-cell population. Sorting of regulatory T cells into CD147(-) and CD147(+) populations demonstrated that CD147 identifies an activated and highly suppressive CD45RO(+) Treg subset. When analyzing CD4(+) T cells for their cytokine producing potential, CD147 levels grouped the FoxP3(+) subset into 3 categories with different ability to produce IL-2, TNF-α, IFN-γ, and IL-17. Together, this suggests that CD147 is a direct marker for activated Tregs within the CD4(+)FoxP3(+) subset and may provide means to manipulate cells important for immune homeostasis.

Expansion and productive HIV-1 infection of Foxp3 positive CD4 T cells at pleural sites of HIV/TB co-infection

PubMed Central

Hirsch, Christina S; Baseke, Joy; Kafuluma, John Lusiba; Nserko, Mary; Mayanja-Kizza, Harriet; Toossi, Zahra

2016-01-01

Background CD4 T-cells expressing Foxp3 are expanded systemically during active tuberculosis (TB) regardless of HIV-1 co-infection. Foxp3+ CD4 T cells are targets of HIV-1 infection. However, expansion of HIV-1 infected Foxp3+ CD4 T cells at sites of HIV/TB co-infection, and whether they contribute to promotion of HIV-1 viral activity is not known. Methods Pleural fluid mononuclear cells (PFMC) from HIV/TB co-infected patients with pleural TB were characterized by immune-staining and FACS analysis for surface markers CD4, CD127, CCR5, CXCR4, HLA-DR and intracellular expression of Foxp3, HIVp24, IFN-γ and Bcl-2. Whole PFMC and bead separated CD4+CD25+CD127− T cells were assessed for HIV-1 LTR strong stop (SS) DNA by real-time PCR, which represents viral DNA post cell entry and initiation of reverse transcription. Results High numbers of HIV-1 p24 positive Foxp3+ and Foxp3+CD127− CD4 T cells were identified in PFMC from HIV/TB co-infected subjects. CD4+Foxp3+CD127− T cells displayed high expression of the cellular activation marker, HLA-DR. Further, expression of the HIV-1 co-receptors, CCR5 and CXCR4, were higher on CD4+Foxp3+T cells compared to CD4+Foxp3− T cells. Purified CD4+CD25+CD127− T cells isolated from PFMC of HIV/TB co-infected patients, were over 90% CD4+Foxp3+T cells, and exhibited higher HIV-1 SS DNA as compared to whole PFMC, and as compared to CD4+CD25+CD127− T cells from an HIV-infected subject with pleural mesothelioma. HIV-1 p24+ Foxp3+ CD4+T cells from HIV/TB patients higher in Bcl-2 expression as compared to both HIV-1 p24+ Foxp3− CD4 T cells, and Foxp3+ CD4+T cells without HIV-p24 expression. Conclusion Foxp3+ CD4 T cells in PFMC from HIV/TB co-infected subjects are predisposed to productive HIV-1 infection and have survival advantage as compared to Foxp3 negative CD4 T cells. PMID:28124031

T-lymphocyte populations following a period of high volume training in female soccer players.

PubMed

Brown, F F; Bigley, A B; Ross, J C; LaVoy, E C; Simpson, R J; Galloway, S D R

2015-12-01

To investigate the T-lymphocyte response to a period of increased training volume in trained females compared to habitual activity in female controls. Thirteen trained female (19.8 ± 1.9 yrs) soccer players were monitored during a two-week long high volume training period (increased by 39%) and thirteen female untrained (20.5 ± 2.2 yrs) controls were monitored during two-weeks of habitual activity. Blood lymphocytes, collected at rest, were isolated before and after the two-week period. Isolated lymphocytes were assessed for the cell surface expression of the co-receptor CD28, a marker of T-lymphocyte naivety, and CD57 a marker used to identify highly-differentiated T-lymphocytes. Co-expression of these markers was identified on helper CD4(+) and cytotoxic CD8(+) T-lymphocytes. In addition a further population of γδ(+) T-lymphocytes were identified. Plasma was used to determine Cytomegalovirus (CMV) serostatus. No difference was observed in the T-lymphocyte populations following the two-week period of increased volume training. At baseline the number of total CD3(+), cytotoxic CD8(+), naïve (CD8(+) CD28(+) CD57(-)), intermediate (CD8(+) CD28(+) CD57(+)) T-lymphocytes and the number and proportion of γδ(+) T-lymphocytes were greater in the trained compared to the untrained females (p<0.05). The proportion of CD4(+)T-lymphocytes was greater in the untrained compared to the trained (p<0.05), in turn the CD4(+):CD8(+) ratio was also greater in the untrained females (p<0.05). Inclusion of percentage body fat as a covariate removed the main effect of training status in all T-lymphocyte sub-populations, with the exception of the γδ(+) T-lymphocyte population. 8% of the untrained group was defined as positive for CMV whereas 23% of the trained group was positive for CMV. However, CMV was not a significant covariate in the analysis of T-lymphocyte proportions. The period of high volume training had no effect on T-lymphocyte populations in trained females. However, baseline training status differences were evident between groups. This indicates that long-term exercise training, as opposed to short-term changes in exercise volume, appears to elicit discernible changes in the composition of the blood T-lymphocyte pool. Copyright © 2015 Elsevier Inc. All rights reserved.

Detection and functional analysis of tumor infiltrating T-lymphocytes (TIL) in liver metastases from colorectal cancer.

PubMed

Wagner, Philipp; Koch, Moritz; Nummer, Daniel; Palm, Sylvia; Galindo, Luis; Autenrieth, Daniel; Rahbari, Nuh; Schmitz-Winnenthal, Friedrich H; Schirrmacher, Volker; Büchler, Markus W; Beckhove, Philipp; Weitz, Jürgen

2008-08-01

Tumor-infiltrating T lymphocytes (TIL) play an important role in primary colorectal cancer, but their activity in liver metastases has not yet been investigated. The aim of this study was to examine whether tumor-selective infiltration, activation, and cytotoxic activity of TIL can be demonstrated in situ in colorectal liver metastases. TIL were obtained from liver metastases and corresponding normal liver tissue of 16 patients with colorectal liver metastases. Characterization of TIL in situ was performed by multicolor flowcytometric analysis. Presence of tumor antigen-reactive T cells was evaluated by interferon gamma Elispot analysis. TIL in colorectal liver metastases responding against tumor antigens were present in most patients. Although the proportions of CD3(+) T cells were comparable in liver metastasis and normal liver tissue, metastases contained significantly enhanced proportions of CD4(+) cells (49% vs. 22%, P < .001). Among all CD4(+) T helper cells, the proportion of activated (CD4(+)CD25(+)) effector cells was significantly increased in liver metastases (15.0% vs. 7.8%, P = .003). Metastases showed significantly higher proportions of activated (CD69(+) [70.1% vs. 49.8%, P = .02] and CD25(+) [4.1% vs. .6%, P = .06]) and cytotoxically active (CD107a(+)) CD8(+) TIL (3.2% vs. 1.3%, P = .03). Importantly, the presence of activated T helper cells correlated with the frequencies of cytotoxic T lymphocytes that exerted cytotoxic activity in situ (P = .02). CD4(+) and CD8(+) TIL are selectively activated in liver metastases, and cytotoxic T lymphocytes exert tumor-selective cytotoxic activity in situ in the presence of activated T helper cells, suggesting the requirement of in-situ-activated T helper cells for efficient cytotoxic T lymphocytes effector function.

Increased CD56(bright) NK cells in HIV-HCV co-infection and HCV mono-infection are associated with distinctive alterations of their phenotype.

PubMed

Bhardwaj, Suvercha; Ahmad, Fareed; Wedemeyer, Heiner; Cornberg, Marcus; Schulze Zur Wiesch, Julian; van Lunzen, Jan; Sarin, Shiv K; Schmidt, Reinhold E; Meyer-Olson, Dirk

2016-04-18

HIV-HCV co-infection is associated with accelerated progression to hepatic fibrosis, cirrhosis and hepatocellular carcinoma than HCV mono-infection. The contribution of innate immunity during HIV-HCV co-infection has been a relatively under-investigated area. Natural killer (NK) cells are pivotal sentinels of innate immunity against viruses and tumour cells. In this study we evaluated the effect of HIV-HCV co-infection on peripheral blood NK cell subsets with emphasis on the phenotype of CD56(bright) NK cells. Sixty patients were included in the study; HIV mono-infected (n = 12), HCV mono-infected (n = 15), HCV-HIV co-infected (n = 21) and healthy controls (n = 16). PBMCs were isolated and immunophenotyping of NK cells was performed by flowcytometry. We observed an expansion of CD56(bright) NK cell subset in HIV-HCV co-infection as compared to healthy controls and HIV mono-infected group. All the infected groups had an upregulated expression of the activating receptor NKG2D on CD56(bright) NK cells in comparison to healthy controls while not differing amongst themselves. The expression of NKp46 in HIV-HCV co-infected group was significantly upregulated as compared to both HIV as well as HCV mono-infections while NKp30 expression in the HIV-HCV co-infected group significantly differed as compared to HIV mono-infection. The CD56(bright) NK cell subset was activated in HIV-HCV co-infection as assessed by the expression of CD69 as compared to healthy controls but was significantly downregulated in comparison to HIV mono-infection. CD95 expression on CD56(bright) NK cells followed the same pattern where there was an increased expression of CD95 in HIV mono-infection and HIV-HCV co-infection as compared to healthy controls. In contrast to CD69 expression, CD95 expression in HCV mono-infection was decreased when compared to HIV mono-infection and HIV-HCV co-infection. Finally, expression of CXCR3 on CD56(bright) NK cells was increased in HIV-HCV co-infection in comparison to HIV mono-infection while remaining similar to HCV mono-infection. Thus, HIV-HCV co-infection is able to modulate the phenotype of CD56(bright) NK cell subset in a unique way such that NKp46 and CXCR3 expressions are distinct for co-infection while both mono-infections have an additive effect on CD56(bright), CD69 with CD95 expressions. HCV mono-infection has a dominant effect on NKp30 expression while NKG2D and CD127 expressions remained same in all the groups.

Association of Glioblastoma Multiforme Stem Cell Characteristics, Differentiation, and Microglia Marker Genes with Patient Survival

PubMed Central

Balz, Ellen; Herzog, Susann; Plantera, Laura; Vogelgesang, Silke; Seifert, Carolin; Bialke, Angela; Venugopal, Chitra; Singh, Sheila K.; Hoffmann, Wolfgang; Schroeder, Henry W. S.

2018-01-01

Patients with glioblastoma multiforme (GBM) are at high risk to develop a relapse despite multimodal therapy. Assumedly, glioma stem cells (GSCs) are responsible for treatment resistance of GBM. Identification of specific GSC markers may help to develop targeted therapies. Here, we performed expression analyses of stem cell (ABCG2, CD44, CD95, CD133, ELF4, Nanog, and Nestin) as well as differentiation and microglia markers (GFAP, Iba1, and Sparc) in GBM compared to nonmalignant brain. Furthermore, the role of these proteins for patient survival and their expression in LN18 stem-like neurospheres was analyzed. At mRNA level, ABCG2 and CD95 were reduced, GFAP was unchanged; all other investigated markers were increased in GBM. At protein level, CD44, ELF4, Nanog, Nestin, and Sparc were elevated in GBM, but only CD133 and Nestin were strongly associated with survival time. In addition, ABCG2 and GFAP expression was decreased in LN18 neurospheres whereas CD44, CD95, CD133, ELF4, Nanog, Nestin, and Sparc were upregulated. Altogether only CD133 and Nestin were associated with survival rates. This raises concerns regarding the suitability of the other target structures as prognostic markers, but makes both CD133 and Nestin candidates for GBM therapy. Nevertheless, a search for more specific marker proteins is urgently needed. PMID:29535786

Nocardia rubra cell-wall skeleton promotes CD4+ T cell activation and drives Th1 immune response.

PubMed

Wang, Guangchuan; Wu, Jie; Miao, Miao; Dou, Heng; Nan, Ning; Shi, Mingsheng; Yu, Guang; Shan, Fengping

2017-08-01

Several lines of evidences have shown that Nocardia rubra cell wall skeleton (Nr-CWS) has immunoregulatory and anti-tumor activities. However, there is no information about the effect of Nr-CWS on CD4 + T cells. The aim of this study was to explore the effect of Nr-CWS on the phenotype and function of CD4 + T cells. Our results of in vitro experiments showed that Nr-CWS could significantly up-regulate the expression of CD69 and CD25 on CD4 + T cells, promote the proliferation of CD4 + T cells, increase the production of IFN-γ, TNF-α and IL-2 in the supernatants, but has no significant effect on the apoptosis and death of CD4 + T cells. Results of in vivo experiments showed that Nr-CWS could promote the proliferation of CD4 + T cells, and increase the production of IL-2, IFN-γ and TNF-α (Th1 type cytokines). These data suggest that Nr-CWS can enhance the activation of CD4 + T cells, promote the proliferation of CD4 + T cells and the differentiation of CD4 + T cells to Th1 cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Serum Neutrophil Gelatinase B-associated Lipocalin and Matrix Metalloproteinase-9 Complex as a Surrogate Marker for Mucosal Healing in Patients with Crohn's Disease.

PubMed

de Bruyn, Magali; Arijs, Ingrid; De Hertogh, Gert; Ferrante, Marc; Van Assche, Gert; Rutgeerts, Paul; Vermeire, Séverine; Opdenakker, Ghislain

2015-12-01

Although costly and uncomfortable for the patient, the current standard to assess mucosal healing in Crohn's disease [CD] patients is endoscopy. The aim of this study was to evaluate NGAL-MMP-9 as surrogate marker for mucosal healing in CD patients. Serum NGAL-MMP-9 levels were determined with sandwich enzyme-linked immunosorbent assay before and up to 5 years after first infliximab infusion in 108 active CD patients [median age at first infliximab 36 years, 57% female] and 43 healthy controls [HC, median age 27 years, 60% female]. Serum samples were matched to the time of endoscopy and complete endoscopic healing was defined as absence of ulcerations. Histological healing was defined as absence of epithelial damage [D'Haens score]. At baseline, median [interquartile range] NGAL-MMP-9 levels were significantly higher in active CD patients vs HC (77.6 [36.9-141.0] vs 25.5 [17.8-42.8] ng/ml; p < 0.001). After treatment, NGAL-MMP-9 levels significantly decreased in completely healed CD patients [n = 38] (84.5 [36.7-138.4] to 23.4 [7.4-42.5] ng/ml; p < 0.001) and--to a lesser extent--in non-healed CD patients [n = 36] (100.9 [43.4-152.6] to 43.8 [27.0-96.8] ng/ml; p = 0.001). Receiver operating characteristic analysis defined a NGAL-MMP-9 cut-off level of 45 ng/ml corresponding to complete endoscopic healing (area under the curve [AUC] = 0.79, 82% sensitivity, 65% specificity) and histological healing [AUC = 0.72, 79% sensitivity, 53% specificity]. At baseline, C-reactive protein [CRP] was not elevated in 33% of active CD patients, whereas 53% of these patients did have elevated NGAL-MMP-9 levels. In the search for surrogate markers to assess mucosal healing in inflammatory bowel disease, NGAL-MMP-9 supplements and outperforms CRP in both ulcerative colitis and CD patients. Copyright © 2015 European Crohn’s and Colitis Organisation (ECCO). Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Murine T cell activation is regulated by surfen (bis-2-methyl-4-amino-quinolyl-6-carbamide)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Warford, Jordan, E-mail: jordan.warford@dal.ca; Doucette, Carolyn D., E-mail: carolyn.doucette@dal.ca; Hoskin, David W., E-mail: d.w.hoskin@dal.ca

2014-01-10

Highlights: •Surfen is the first inhibitor of glycosaminoglycan function to be studied in murine T cells. •Surfen reduces T cell proliferation stimulated in vitro and in vivo. •Surfen reduces CD25 expression in T cells activated in vivo but not in vitro. •Surfen increases T cell proliferation when T cell receptor activation is bypassed. •Surfen’s effects are blocked by co-administration of heparin sulfate. -- Abstract: Surfen (bis-2-methyl-4-amino-quinolyl-6-carbamide) binds to glycosaminoglycans (GAGs) and has been shown to influence their function, and the function of proteoglycans (complexes of GAGs linked to a core protein). T cells synthesize, secrete and express GAGs and proteoglycansmore » which are involved in several aspects of T cell function. However, there are as yet no studies on the effect of GAG-binding agents such as surfen on T cell function. In this study, surfen was found to influence murine T cell activation. Doses between 2.5 and 20 μM produced a graduated reduction in the proliferation of T cells activated with anti-CD3/CD28 antibody-coated T cell expander beads. Surfen (20 mg/kg) was also administered to mice treated with anti-CD3 antibody to activate T cells in vivo. Lymphocytes from surfen-treated mice also showed reduced proliferation and lymph node cell counts were reduced. Surfen reduced labeling with a cell viability marker (7-ADD) but to a much lower extent than its effect on proliferation. Surfen also reduced CD25 (the α-subunit of the interleukin (IL)-2 receptor) expression with no effect on CD69 expression in T cells treated in vivo but not in vitro. When receptor activation was bypassed by treating T cells in vitro with phorbyl myristate acetate (10 ng/ml) and ionomycin (100 ng/ml), surfen treatment either increased proliferation (10 μM) or had no effect (2.5, 5 and 20 μM). In vitro treatment of T cells with surfen had no effect on IL-2 or interferon-γ synthesis and did not alter proliferation of the IL-2 dependent cell line CTLL-2. The effect of surfen was antagonized dose-dependently by co-treatment with heparin sulfate. We conclude that surfen inhibits T cell proliferation in vitro and in vivo. When T cell receptor-driven activation is bypassed surfen had a neutral or stimulatory effect on T cell proliferation. The results imply that endogenous GAGs and proteoglycans play a complex role in promoting or inhibiting different aspects of T cell activation.« less

Soluble CD26/CD30 levels in visceral leishmaniasis: markers of disease activity

PubMed Central

Ajdary, S; Riazi-Rad, F; Jafari-Shakib, R; Mohebbali, M

2006-01-01

Leishmania infantum is the causative agent of zoonotic visceral leishmaniasis (VL). If untreated the disease could be fatal; however, in some cases the infection can run a subclinical course. In subclinical infections a Th1-response predominates, while Th2-responses and/or probably Treg cells are related to unfavourable outcome of the disease in active VL. In the present study we determined the levels of soluble (s) CD26 and CD30 co-stimulatory molecules in sera from patients with active VL, asymptomatic individuals and healthy volunteers. Results showed a significant difference in both sCD26 and sCD30 between infected cases and normal individuals (P ≤ 0·001). However, there was no significant difference in sCD26 levels between asymptomatic cases and patients, although the difference was not significant. sCD30 levels were significantly higher in VL patients than asymptomatic cases (P ≤ 0·001). These findings suggest a possible association between sCD26 and sCD30 levels and the clinical manifestation of L. infantum infection. PMID:16792672

Microglia activation in Niemann-Pick disease, type C1 is amendable to therapeutic intervention.

PubMed

Cougnoux, Antony; Drummond, Rebecca A; Collar, Amanda L; Iben, James R; Salman, Alexander; Westgarth, Harrison; Wassif, Christopher A; Cawley, Niamh X; Farhat, Nicole Y; Ozato, Keiko; Lionakis, Michail S; Porter, Forbes D

2018-06-15

Niemann-Pick disease, type C1 (NPC1) is a neurodegenerative disorder with limited treatment options. NPC1 is associated with neuroinflammation; however, attempts to therapeutically target neuroinflammation in NPC1 have had mixed success. We show here that NPC1 neuroinflammation is characterized by an atypical microglia activation phenotype. Specifically, Npc1-/- microglia demonstrated altered morphology, reduced levels of lineage markers and a shift toward glycolytic metabolism. Treatment with 2-hydroxypropyl-β-cyclodextrin (HPβCD), a drug currently being studied in a phase 2b/3 clinical trial, reversed all microglia-associated defects in Npc1-/- animals. In addition, impairing microglia mediated neuroinflammation by genetic deletion of IRF8 led to decreased symptoms and increased lifespan. We identified CD22 as a marker of dysregulated microglia in Npc1 mutant mice and subsequently demonstrated that elevated cerebrospinal fluid levels of CD22 in NPC1 patients responds to HPβCD administration. Collectively, these data provide the first in-depth analysis of microglia function in NPC1 and suggest possible new therapeutic approaches.

The PBX1 lupus susceptibility gene regulates CD44 expression

PubMed Central

Niu, Yuxin; Sengupta, Mayami; Titov, Anton A.; Choi, Seung-Chul; Morel, Laurence

2017-01-01

PBX1-d is novel splice isoform of pre-B-cell leukemia homeobox 1 (PBX1) that lacks its DNA-binding and Hox-binding domains, and functions as a dominant negative. We have shown that PBX1-d expression in CD4+ T cells is associated with systemic lupus erythematosus (SLE) in a mouse model as well as in human subjects. More specifically, PBX1-d expression leads to the production of autoreactive activated CD4+ T cells, a reduced frequency and function of Foxp3+ regulatory T (Treg) cells and an expansion of follicular helper T (Tfh) cells. Very little is known about the function of PBX1 in T cells, except that it directly regulates the expression of miRNAs associated with Treg and Tfh homeostasis. In the present study, we show that PBX1 directly regulated the expression of CD44, a marker of T cell activation. Two PBX1 binding sites in the promoter directly regulated CD44 expression, with PBX1-d driving a higher expression than the normal isoform PBX1-b. In addition, mutations in each of the two binding sites had different effects of PBX1-b and PBX1-d. Finally, we showed that an enhanced recruitment of co-factor MEIS by PBX1-d over PBX1-b, while there was no difference for co-factor PREP1 recruitment. Therefore, this study demonstrates that the lupus-associated PBX1-d isoform directly transactivates CD44, a marker of CD44 activation and memory, and that it has different DNA binding and co-factor recruitment relative to the normal isoform. Taken together, these results confirm that PBX1 directly regulates genes related to T cell activation and show that the lupus-associated isoform PBX1-d has unique molecular functions. PMID:28257976

The CD14+CD16+ Inflammatory Monocyte Subset Displays Increased Mitochondrial Activity and Effector Function During Acute Plasmodium vivax Malaria

PubMed Central

Antonelli, Lis R. V.; Leoratti, Fabiana M. S.; Costa, Pedro A. C.; Rocha, Bruno C.; Diniz, Suelen Q.; Tada, Mauro S.; Pereira, Dhelio B.; Teixeira-Carvalho, Andrea; Golenbock, Douglas T.; Gonçalves, Ricardo; Gazzinelli, Ricardo T.

2014-01-01

Infection with Plasmodium vivax results in strong activation of monocytes, which are important components of both the systemic inflammatory response and parasite control. The overall goal of this study was to define the role of monocytes during P. vivax malaria. Here, we demonstrate that P. vivax–infected patients display significant increase in circulating monocytes, which were defined as CD14+CD16− (classical), CD14+CD16+ (inflammatory), and CD14loCD16+ (patrolling) cells. While the classical and inflammatory monocytes were found to be the primary source of pro-inflammatory cytokines, the CD16+ cells, in particular the CD14+CD16+ monocytes, expressed the highest levels of activation markers, which included chemokine receptors and adhesion molecules. Morphologically, CD14+ were distinguished from CD14lo monocytes by displaying larger and more active mitochondria. CD14+CD16+ monocytes were more efficient in phagocytizing P. vivax-infected reticulocytes, which induced them to produce high levels of intracellular TNF-α and reactive oxygen species. Importantly, antibodies specific for ICAM-1, PECAM-1 or LFA-1 efficiently blocked the phagocytosis of infected reticulocytes by monocytes. Hence, our results provide key information on the mechanism by which CD14+CD16+ cells control parasite burden, supporting the hypothesis that they play a role in resistance to P. vivax infection. PMID:25233271

CD147, CD44, and the epidermal growth factor receptor (EGFR) signaling pathway cooperate to regulate breast epithelial cell invasiveness.

PubMed

Grass, G Daniel; Tolliver, Lauren B; Bratoeva, Momka; Toole, Bryan P

2013-09-06

The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777-788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer.

Equine Mesenchymal Stromal Cells Retain a Pericyte-Like Phenotype

PubMed Central

Sheldrake, Tara A.; Dawson, Lucy; Menghini, Timothy; Rink, Burgunde Elisabeth; Amilon, Karin; Khan, Nusrat; Péault, Bruno; Donadeu, Francesc Xavier

2017-01-01

Mesenchymal stem/stromal cells (MSCs) have been used in human and equine regenerative medicine, and interest in exploiting their potential has increased dramatically over the years. Despite significant effort to characterize equine MSCs, the actual origin of these cells and how much of their native phenotype is maintained in culture have not been determined. In this study, we investigated the relationship between MSCs, derived from adipose tissue (AT) and bone marrow (BM), and pericytes in the horse. Both pericyte (CD146, NG2, and αSMA) and MSC (CD29, CD90, and CD73) markers were detected in equine AT and colocalized around blood vessels. Importantly, as assessed by flow cytometry, both pericyte (CD146, NG2, and αSMA) and MSC (CD29, CD44, CD90, and CD105) markers were present in a majority (≥90%) of cells in cultures of AT-MSCs and BM-MSCs; however, levels of pericyte markers were variable within each of those populations. Moreover, the expression of pericyte markers was maintained for at least eight passages in both AT-MSCs and BM-MSCs. Hematopoietic (CD45) and endothelial (CD144) markers were also detected at low levels in MSCs by quantitative polymerase chain reaction (qPCR). Finally, in coculture experiments, AT-MSCs closely associated with networks produced by endothelial cells, resembling the natural perivascular location of pericytes in vivo. Our results indicate that equine MSCs originate from perivascular cells and moreover maintain a pericyte-like phenotype in culture. Therefore, we suggest that, in addition to classical MSC markers, pericyte markers such as CD146 could be used when assessing and characterizing equine MSCs. PMID:28376684

Equine Mesenchymal Stromal Cells Retain a Pericyte-Like Phenotype.

PubMed

Esteves, Cristina L; Sheldrake, Tara A; Dawson, Lucy; Menghini, Timothy; Rink, Burgunde Elisabeth; Amilon, Karin; Khan, Nusrat; Péault, Bruno; Donadeu, Francesc Xavier

2017-07-01

Mesenchymal stem/stromal cells (MSCs) have been used in human and equine regenerative medicine, and interest in exploiting their potential has increased dramatically over the years. Despite significant effort to characterize equine MSCs, the actual origin of these cells and how much of their native phenotype is maintained in culture have not been determined. In this study, we investigated the relationship between MSCs, derived from adipose tissue (AT) and bone marrow (BM), and pericytes in the horse. Both pericyte (CD146, NG2, and αSMA) and MSC (CD29, CD90, and CD73) markers were detected in equine AT and colocalized around blood vessels. Importantly, as assessed by flow cytometry, both pericyte (CD146, NG2, and αSMA) and MSC (CD29, CD44, CD90, and CD105) markers were present in a majority (≥90%) of cells in cultures of AT-MSCs and BM-MSCs; however, levels of pericyte markers were variable within each of those populations. Moreover, the expression of pericyte markers was maintained for at least eight passages in both AT-MSCs and BM-MSCs. Hematopoietic (CD45) and endothelial (CD144) markers were also detected at low levels in MSCs by quantitative polymerase chain reaction (qPCR). Finally, in coculture experiments, AT-MSCs closely associated with networks produced by endothelial cells, resembling the natural perivascular location of pericytes in vivo. Our results indicate that equine MSCs originate from perivascular cells and moreover maintain a pericyte-like phenotype in culture. Therefore, we suggest that, in addition to classical MSC markers, pericyte markers such as CD146 could be used when assessing and characterizing equine MSCs.

Clinical Significance of PD-L1+ Exosomes in Plasma of Head and Neck Cancer Patients.

PubMed

Theodoraki, Marie-Nicole; Yerneni, Saigopalakrishna S; Hoffmann, Thomas K; Gooding, William E; Whiteside, Theresa L

2018-02-15

Purpose: The microenvironment of head and neck squamous cell carcinomas (HNSCC) is highly immunosuppressive. HNSCCs expressing elevated levels of PD-L1 have especially poor outcome. Exosomes that carry PD-L1 and suppress T-cell functions have been isolated from plasma of patients with HNSCC. The potential contributions of PD-L1 + exosomes to immune suppression and disease activity are evaluated. Experimental Design: Exosomes isolated from plasma of 40 HNSCC patients by size exclusion chromatography were captured on beads using anti-CD63 Abs, stained for PD-1 and PD-L1 and analyzed by flow cytometry. The percentages and mean fluorescence intensities (MFI) of PD-L1 + and PD-1 + exosome/bead complexes were correlated with the patients' clinicopathologic data. PD-L1 high or PD-L1 low exosomes were incubated with activated CD69 + human CD8 + T cells ± PD-1 inhibitor. Changes in CD69 expression levels on T cells were measured. Patients' plasma was tested for soluble PD-L1 (sPD-L1) by ELISA. Results: Levels of PD-L1 carried by exosomes correlated with patients' disease activity, the UICC stage and the lymph node status ( P = 0.0008-0.013). In contrast, plasma levels of sPD-L1 or exosome PD-1 levels did not correlate with any clinicopathologic parameters. CD69 expression levels were inhibited ( P < 0.03) by coincubation with PD-L1 high but not by PD-L1 low exosomes. Blocking of PD-L1 + exosome signaling to PD-1 + T cells attenuated immune suppression. Conclusions: PD-L1 levels on exosomes, but not levels of sPD-L1, associated with disease progression in HNSCC patients. Circulating PD-L1 + exosomes emerge as useful metrics of disease and immune activity in HNSCC patients. Circulating PD-L1 high exosomes in HNC patients' plasma but not soluble PD-L1 levels associate with disease progression. Clin Cancer Res; 24(4); 896-905. ©2017 AACR . ©2017 American Association for Cancer Research.

Helicobacter pylori cagA+ Is Associated with Milder Duodenal Histological Changes in Chilean Celiac Patients

PubMed Central

Lucero, Yalda; Oyarzún, Amaya; O'Ryan, Miguel; Quera, Rodrigo; Espinosa, Nelly; Valenzuela, Romina; Simian, Daniela; Alcalde, Elisa; Arce, Claudio; Farfán, Mauricio J.; Vergara, Alejandra F.; Gajardo, Iván; Mendez, Jocelyn; Carrasco, Jorge; Errázuriz, Germán; Gonzalez, Mónica; Ossa, Juan C.; Maiza, Eduardo; Perez-Bravo, Francisco; Castro, Magdalena; Araya, Magdalena

2017-01-01

HIGHLIGHTS What is already known about this subject?Celiac disease (CD) has a high clinical and histological diversity and the mechanisms underlying this phenomenon remain elusive.H. pylori is a bacterium that chronically infect gastric and duodenal mucosa activating both a Th1/Th17 and T-reg pathways.The role of H. pylori (and the effect of their virulence factors) in CD have not yet completely elucidated.What are the new findings?cagA+ H. pylori strains are associated to milder histological damage in infected CD patients.In active-CD patients the presence of cagA+ H. pylori is associated to an increase in T-reg markers, contrasting with a downregulation in cagA+ infected potential-CD individuals.How might it impact on clinical practice in the foreseeable future?The identification of microbiological factors that could modulate inflammation and clinical expression of CD may be used in the future as preventive strategies or as supplementary treatment in patients that cannot achieve complete remission, contributing to the better care of these patients. Background: Mechanisms underlying the high clinical and histological diversity of celiac disease (CD) remain elusive. Helicobacter pylori (Hp) chronically infects gastric and duodenal mucosa and has been associated with protection against some immune-mediated conditions, but its role (specifically of cagA+ strains) in CD is unclear. Objective: To assess the relationship between gastric Hp infection (cagA+ strains) and duodenal histological damage in patients with CD. Design: Case-control study including patients with active-CD, potential-CD and non-celiac individuals. Clinical presentation, HLA genotype, Hp/cagA gene detection in gastric mucosa, duodenal histology, Foxp3 positive cells and TGF-β expression in duodenal lamina propria were analyzed. Results: We recruited 116 patients, 29 active-CD, 37 potential-CD, and 50 non-CD controls. Hp detection was similar in the three groups (~30–40%), but cagA+ strains were more common in infected potential-CD than in active-CD (10/11 vs. 4/10; p = 0.020) and non-CD (10/20; p = 0.025). Among active-CD patients, Foxp3 positivity was significantly higher in subjects with cagA+ Hp+ compared to cagA- Hp+ (p < 0.01) and Hp- (p < 0.01). In cagA+ Hp+ individuals, Foxp3 positivity was also higher comparing active- to potential-CD (p < 0.01). TGF-β expression in duodenum was similar in active-CD with cagA+ Hp+ compared to Hp- and was significantly downregulated in cagA+ potential-CD subjects compared to other groups. Conclusion: Hp infection rates were similar among individuals with/without CD, but infection with cagA+ strains was associated with milder histological damage in celiac patients infected by Hp, and in active-CD cases with higher expression of T-reg markers. Results suggest that infection by cagA+ Hp may be protective for CD progression, or conversely, that these strains are prone to colonize intestinal mucosa with less severe damage. PMID:28879170

The Prognostic Role of Cancer Stem Cell Markers for Long-term Outcome After Resection of Colonic Liver Metastases.

PubMed

Spelt, Lidewij; Sasor, Agata; Ansari, Daniel; Hilmersson, Katarzyna Said; Andersson, Roland

2018-01-01

To assess the expression of cancer stem cell (CSC) markers CD44, CD133 and CD24 in colon cancer liver metastases and analyse their predictive value for overall survival (OS) and disease-free survival (DFS) after liver resection. Patients operated on for colon cancer liver metastases were included. CSC marker expression was determined through immunohistochemistry analysis. OS and DFS were compared between marker-positive and marker-negative patients. Multivariate analysis was performed to select predictive variables for OS and DFS. CD133-positive patients had a worse DFS than CD133-negative patients, with a median DFS of 12 and 25 months (p=0.051). Multivariate analysis selected CD133 expression as a significant predictor for DFS. CD44 and CD24 were not found to predict OS or DFS. CD133 expression in colonic liver metastases is a negative prognostic factor for DFS after liver resection. In the future, CD133 could be used as a biomarker for risk stratification, and possibly for developing novel targeted therapy. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Effect of astragalus injection on serious abdominal traumatic patients' cellular immunity.

PubMed

Wu, Jian; Wang, Yi-xin; Su, Wen-li; Zhu, Wen-xian; Lu, Jing-wei; Li, Zhen-kai

2006-03-01

To explore the change of serious abdominal traumatic patients' cellular immunity and the effect of Astragalus Injection (AI) on it. Sixty-three serious abdominal traumatic patients were randomly assigned into two groups, the conventional group and the treated group, patients in the conventional group were given conventional treatment, while others in the treated group were given conventional treatment as the basis, with AI 20 ml was added into 250 ml of 5% glucose solution given through intravenous dripping, and then on the first day and 14th day, their T cell activated antigens as well as that of 10 healthy subjects were monitored. On the first day, in the conventional group and treated group, the levels of CD(3)(+), CD(4)(+), CD(4)(+)/CD(8)(+), CD(16)(+), CD(69)(+) and CD(3)(+)/homologous leucocytic antigen-DR (HLA-DR(+)) were apparently lower than those in the healthy group (P < 0.05), while the CD(8)(+) was significantly higher than that in the healthy group (P < 0.05), and there was no significant difference between the conventional group and the treated group (P > 0.05); on the 14th days, the levels of CD(3)(+), CD(4)(+), CD(4)(+)/CD(8)(+), CD(16)(+), CD(69)(+) and CD(3)(+)/HLA-DR(+) of the treated group got closed to healthy subject value, and got even higher than those of conventional group (P < 0.05); CD(8)(+) got close to that of healthy subjects, while obviously lower than that of conventional group (P < 0.05). After serious abdominal trauma, cellular immunity lowered, auxiliary use of AI was beneficial to the restoration of cellular immunity.

Outer membrane protein A of Acinetobacter baumannii induces differentiation of CD4+ T cells toward a Th1 polarizing phenotype through the activation of dendritic cells.

PubMed

Lee, Jun Sik; Lee, Je Chul; Lee, Chang-Min; Jung, In Duk; Jeong, Young-Il; Seong, Eun-Young; Chung, Hae-Young; Park, Yeong-Min

2007-06-30

Acinetobacter baumannii is an increasing hospital-acquired pathogen that causes a various type of infections, but little is known about the protective immune response to this microorganism. Outer membrane protein A of A. baumannii (AbOmpA) is a major porin protein and plays an important role in pathogenesis. We analyzed interaction between AbOmpA and dendritic cells (DCs) to characterize the role of this protein in promoting innate and adaptive immune responses. AbOmpA functionally activates bone marrow-derived DCs by augmenting expression of the surface markers, CD40, CD54, B7 family (CD80 and CD86) and major histocompatibility complex class I and II. AbOmpA induces production of Th1-promoting interleukin-12 from DCs and augments the syngeneic and allogeneic immunostimulatory capacity of DCs. AbOmpA stimulates production of interferon-gamma from T cells in mixed lymphocyte reactions, which suggesting Th1-polarizing capacity. CD4(+) T cells stimulated by AbOmpA-stimulated DCs show a Th1-polarizing cytokine profile. The expression of surface markers on DCs is mediated by both mitogen-activated protein kinases and NF-kappaB pathways. Our findings suggest that AbOmpA induces maturation of DCs and drives Th1 polarization, which are important properties for determining the nature of immune response against A. baumannii.

Sitagliptin but not alpha glucosidase inhibitor reduced the serum soluble CD163, a marker for activated macrophage, in individuals with type 2 diabetes mellitus.

PubMed

Hattori, Akiko; Takemoto, Minoru; Tokuyama, Hirotake; Koshizaka, Masaya; Yokote, Koutaro

2017-04-01

Dipeptidyl peptidase-4 inhibitor (DPP-4i) is commonly used worldwide for the treatment of type 2 diabetes mellitus. In addition to its hypoglycemic activity, DPP-4i might have anti-inflammatory effects. In this study we examined the effects of DPP-4i on the serum levels of soluble CD163 (sCD163), a marker for activated macrophages, in individuals with type 2 diabetes mellitus. We compared these anti-inflammatory effects with those of α glucosidase inhibitor (αGI). Japanese patients with type 2 diabetes mellitus who were stably maintained on ≤2mg/day glimepiride alone were recruited and randomly assigned to receive additional sitagliptin (n=37) or αGI (n=37). Levels of sCD163 were measured before the addition and after a 24-week treatment period. Addition of sitagliptin significantly reduced the serum sCD163 (632 vs. 575ng/mL, p<0.05), while αGI did not display this effect (624 vs. 607ng/mL). The changes in levels of sCD163 were not related to changes in either HbA1c or body mass index (BMI). Our results suggested that DPP-4i might exert anti-inflammatory effects in individuals with type 2 diabetes mellitus, which are independent of its effects on glycemia and BMI. Copyright © 2017 Elsevier B.V. All rights reserved.

B and T Lymphocyte Attenuator Down-regulation by HIV-1 Depends on Type I Interferon and Contributes to T-Cell Hyperactivation

PubMed Central

Zhang, Zheng; Xu, Xiangsheng; Lu, Jiyun; Zhang, Shuye; Gu, Lanlan; Fu, Junliang; Jin, Lei; Li, Haiying; Zhao, Min; Zhang, Jiyuan; Wu, Hao; Su, Lishan; Fu, Yang-Xin

2011-01-01

Background. Nonspecific T-cell hyperactivation is the main driving force for human immunodeficiency virus (HIV)–1 disease progression, but the reasons why the excess immune response is not properly shut off are poorly defined. Methods. Eighty-five HIV-1–infected individuals were enrolled to characterize B and T lymphocyte attenuator (BTLA) expression and function. Infection and blockade assays were used to dissect the factors that influenced BTLA signaling in vitro. Results. BTLA expression on overall CD4+ and CD8+ T cells was progressively decreased in HIV-1 infection, which was directly correlated with disease progression and CD4+ T-cell differentiation and activation. BTLA+CD4+ T cells from HIV-1–infected patients also displayed an altered immune status, which was indicated by reduced expression of naive markers but increased activation and exhaustion markers. Cross-linking of BTLA can substantially decrease CD4+ T-cell activation in vitro. This responsiveness of CD4+ T cells to BTLA-mediated inhibitory signaling was further found to be impaired in HIV-1–infected patients. Furthermore, HIV-1 NL4-3 down-regulated BTLA expression on CD4+ T cells dependent on plasmacytoid dendritic cell (pDC)-derived interferon (IFN)-α. Blockade of IFN-α or depletion of pDCs prevents HIV-1-induced BTLA down-regulation. Conclusions. HIV-1 infection potentially impairs BTLA-mediated signaling dependent on pDC-derived IFN-α, which may contribute to broad T-cell hyperactivation induced by chronic HIV-1 infection. PMID:21592997

Effects of DHA-enriched fish oil on monocyte/macrophage activation marker sCD163, asymmetric dimethyl arginine, and insulin resistance in type 2 diabetic patients.

PubMed

Toupchian, Omid; Sotoudeh, Gity; Mansoori, Anahita; Nasli-Esfahani, Ensieh; Djalali, Mahmoud; Keshavarz, Seyyed Ali; Koohdani, Fariba

2016-01-01

The beneficial effects of n-3 polyunsaturated fatty acids on reducing cardiovascular risks are well documented. However, the relative effect on some markers of macrophage activation and vascular function is unclear. The primary objective of this study was to investigate the effects of docosahexaenoic acid (DHA)-enriched fish oil on the marker of monocyte/macrophage activation factor soluble CD163, asymmetric dimethyl arginine (ADMA), and insulin resistance in type 2 diabetic patients. In this double-blind randomized controlled trial, 72 type 2 diabetic patients with an age between 30-70 years and body mass index (BMI) of 18.5 to 40 kg/m(2) were randomly assigned to receive 2.4-g DHA-enriched fish oil or placebo per day for 8 weeks. Anthropometric measurements, biochemical, and body composition analyses were assessed at baseline and end of study. Analysis of covariance (ANCOVA) was conducted by controlling for possible confounders to assess between-group differences. Serum levels of sCD163, triglycerides, waist circumference (WC), and weight to height ratio (WHtR) decreased significantly in the fish oil group when compared with the control group. Serum ADMA concentration decreased in the fish oil group with no significant between-group differences. Controlling for confounders revealed that the differences observed in sCD163, triglycerides, WC, and WHtR remained statistically significant. Short-time fish oil supplementation decreased serum sCD163, triglycerides levels, WC, and WHtR in T2DM patients. Because of the positive relationship between sCD163 levels and some T2DM and obesity-related complications, it seems that DHA can be considered as a key intervention in obesity and T2DM. Copyright © 2016 National Lipid Association. Published by Elsevier Inc. All rights reserved.

Differences of serum interleukin-6 levels in normotensive and preeclampsia women

NASA Astrophysics Data System (ADS)

Lumbanraja, S. N.; Junitasari, R.; Pasaribu, H. P.

2018-03-01

Red cell distribution width (RDW) has been shown higher with the increase of inflammatory activity. As CD4 count is not available in all cities in Indonesia, RDW was hoped to be the next promising marker to predict the progression of HIV infection. This study aims to find the association of RDW with aCD4 count in HIV-positive reproductive women in Indonesia. It was a prospective cross sectional study enrolled 37 consented outpatients HIV-positive reproductive women in Haji Adam Malik General Hospital, Medan, Indonesia. A 10 mL blood was taken, separated into two tubes for complete blood count and CD4 count. Data were collected and analyzed with SPSS 19.Mean CD4 absolute was lowest in >14.5% RDW group but without statistical significance. By classify CD4 absolute count, CD4 absolute ≤350/mm3 showed significantly higher RDW than CD4 absolute >350/mm3 (13.98±2.10% vs 12.79±0.88% p=0.02). No correlation was between RDW and CD4 absolute, as well as, with CD4%.In conclusion, high RDW can be a promising marker to predict the low CD4 count in HIV-positive reproductive women within antiretroviral therapy.

Various types of stem cells, including a population of very small embryonic-like stem cells, are mobilized into peripheral blood in patients with Crohn's disease.

PubMed

Marlicz, Wojciech; Zuba-Surma, Ewa; Kucia, Magda; Blogowski, Wojciech; Starzynska, Teresa; Ratajczak, Mariusz Z

2012-09-01

Developmentally early cells, including hematopoietic stem progenitor cells (HSPCs), mesenchymal stem cells (MSCs), endothelial progenitor cells (EPCs), and very small embryonic-like stem cells (VSELs), are mobilized into peripheral blood (PB) in response to tissue/organ injury. We sought to determine whether these cells are mobilized into PB in patients with Crohn's disease (CD). Twenty-five patients with active CD, 20 patients in clinical remission, and 25 age-matched controls were recruited and PB samples harvested. The circulating CD133+/Lin-/CD45+ and CD34+/Lin-/CD45+ cells enriched for HSPCs, CD105+/STRO-1+/CD45- cells enriched for MSCs, CD34+/KDR+/CD31+/CD45-cells enriched for EPCs, and small CXCR4+CD34+CD133+ subsets of Lin-CD45- cells that correspond to the population of VSELs were counted by fluorescence-activated cell sorting (FACS) and evaluated by direct immunofluorescence staining for pluripotency embryonic markers and by reverse-transcription polymerase chain reaction (RT-PCR) for expression of messenger (m)RNAs for a panel of genes expressed in intestine epithelial stem cells. The serum concentration of factors involved in stem cell trafficking, such as stromal derived factor-1 (SDF-1), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) were measured by enzyme-linked immunosorbent assay (ELISA). Our data indicate that cells expressing markers for MSCs, EPCs, and small Oct-4+Nanog+SSEA-4+CXCR4+lin-CD45- VSELs are mobilized into PB in CD. The mobilized cells also expressed at the mRNA level genes playing a role in development and regeneration of gastrointestinal epithelium. All these changes were accompanied by increased serum concentrations of VEGF and HGF. CD triggers the mobilization of MSCs, EPCs, and VSELs, while the significance and precise role of these mobilized cells in repair of damaged intestine requires further study. Copyright © 2012 Crohn's & Colitis Foundation of America, Inc.

Positive association between concentration of phthalate metabolites in urine and microparticles in adolescents and young adults.

PubMed

Lin, Chien-Yu; Hsieh, Chia-Jung; Lo, Shyh-Chyi; Chen, Pau-Chung; Torng, Pao-Ling; Hu, Anren; Sung, Fung-Chang; Su, Ta-Chen

2016-01-01

Di-(2-ethylhexyl) phthalate (DEHP) has been used worldwide in various products for many years. In vitro studies have shown that exposure to DEHP and its metabolite mono(2-ethylhexyl) phthalate (MEHP) induces endothelial cell apoptosis. Moreover, exposure to DEHP had been linked to cardiovascular risk factors and cardiovascular diseases in epidemiological studies. Circulating microparticles have been known to be indicators of vascular injury. However, whether DEHP or its metabolites are independently associated with microparticles in humans remains unknown. From 2006 to 2008, we recruited 793 subjects (12-30years) from a population-based sample to participate in this cardiovascular disease prevention examination. Each participant was subjected to interviews and biological sample collection to determine the relationship between concentrations of DEHP metabolites MEHP, mono(ethyl-5-hydroxyhexyl) phthalate, and mono(2-ethly-5-oxoheyl) phthalate in urine and concentrations of endothelial microparticles (CD62E and CD31+/CD42a-), platelet microparticles (CD62P and CD31+/CD42a+), and CD14 in serum. Multiple linear regression analysis revealed that an ln-unit increase in MEHP concentration in urine was positively associated with an increase in serum microparticle counts/μL of 0.132 (±0.016) in CD31+/CD42a- (endothelial apoptosis marker), 0.117 (±0.023) in CD31+/CD42a+ (platelet apoptosis marker), and 0.026 (±0.007) in CD14 (monocyte, macrophage, and neutrophil activation marker). There was no association between DEHP metabolite concentration and CD62E or CD62P. In conclusion, a higher MEHP concentration in urine was associated with an increase in endothelial and platelet microparticles in this cohort of adolescents and young adults. Further studies are warranted to clarify the causal relationship between exposure to DEHP and atherosclerosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Vaccination against CD99 inhibits atherogenesis in low-density lipoprotein receptor-deficient mice.

PubMed

van Wanrooij, Eva J A; de Vos, Paula; Bixel, M Gabriele; Vestweber, Dietmar; van Berkel, Theo J C; Kuiper, Johan

2008-06-01

Murine CD99 was recently found to be expressed on leukocytes and endothelial cells, where it is concentrated at inter-endothelial contacts. Blockade of CD99 by specific antibodies inhibits leukocyte extravasation to inflamed sites in vivo. The aim of the present study is to show the role of CD99 in atherosclerosis using a CD99 vaccination protocol to block the function of CD99 during atherosclerosis. We constructed a DNA vaccine against CD99 by cloning the extracellular domain of murine CD99 into pcDNA3. Vaccination was performed by oral administration of attenuated Salmonella typhimurium transformed with pcDNA3-CD99. This vaccination results in a CD99-specific, CD8-mediated cytotoxic response and subsequent reduction of CD99-expressing cells. We showed that CD99 is expressed on vascular endothelium overlying atherosclerotic plaques and found that CD99 expression is upregulated during western-type diet feeding. CD99 vaccination induced the formation of CD8-positive T cells that were cytotoxic against cells transfected with pcDNA3-CD99. Activation of CD8(+) T cells was demonstrated by a 30% increase in CD8(+)CD69(+) double-positive T cells in spleen and mediastinal lymph nodes. Furthermore, lymphocytes isolated from CD99-vaccinated mice specifically lysed CD99-expressing cells. More importantly, vaccination against CD99 attenuated atherosclerotic lesion formation in the aortic valve leaflets by 38% and in the carotid artery by 69% compared with mice that were vaccinated with a control vector. Furthermore, a lower number of cells were found in atherosclerotic lesions, implying that fewer leukocytes were recruited to these sites. These observations were accompanied by a decrease in CD99 expression on leukocytes. We conclude that vaccination against CD99 decreases atherogenesis by the selective removal of CD99-expressing cells, which could reduce leukocyte recruitment into atherosclerotic lesions and attenuate atherogenesis.

Immunologic reconstitution during PEG-ADA therapy in an unusual mosaic ADA deficient patient.

PubMed

Liu, Ping; Santisteban, Ines; Burroughs, Lauri M; Ochs, Hans D; Torgerson, Troy R; Hershfield, Michael S; Rawlings, David J; Scharenberg, Andrew M

2009-02-01

We report detailed genetic and immunologic studies in a patient diagnosed with adenosine deaminase (ADA) deficiency and combined immune deficiency at age 5 years. At the time of diagnosis, although all other lymphocyte subsets were depleted, circulating CD8(+) T cells with a terminally differentiated phenotype were abundant and expressed normal ADA activity due to a reversion mutation in a CD8(+) T cell or precursor. Over the first 9 months of replacement therapy with PEG-ADA, the patient steadily accumulated mature naïve CD4(+) and CD8(+) T cells, as well as CD4(+)/FOXP3(+) regulatory T cells, consistent with restoration of a functional cellular immune system. While CD19(+) naïve B cells also accumulated in response to PEG-ADA therapy, a high proportion of these B cells exhibited an immature surface marker phenotype even after 9 months, and immunization with neoantigen bacteriophage varphiX174 demonstrated a markedly subnormal humoral immune response. Our observations in this single patient have important implications for gene therapy of human ADA deficiency, as they indicate that ADA expression within even a large circulating lymphocyte population may not be sufficient to support adequate immune reconstitution. They also suggest that an immature surface marker phenotype of the peripheral B cell compartment may be a useful surrogate marker for incomplete humoral immune reconstitution during enzyme replacement, and possibly other forms of hematopoietic cell therapies.

CD81 Controls Sustained T Cell Activation Signaling and Defines the Maturation Stages of Cognate Immunological Synapses

PubMed Central

Rocha-Perugini, V.; Zamai, M.; González-Granado, J. M.; Barreiro, O.; Tejera, E.; Yañez-Mó, M.; Caiolfa, V. R.

2013-01-01

In this study, we investigated the dynamics of the molecular interactions of tetraspanin CD81 in T lymphocytes, and we show that CD81 controls the organization of the immune synapse (IS) and T cell activation. Using quantitative microscopy, including fluorescence recovery after photobleaching (FRAP), phasor fluorescence lifetime imaging microscopy-Föster resonance energy transfer (phasorFLIM-FRET), and total internal reflection fluorescence microscopy (TIRFM), we demonstrate that CD81 interacts with ICAM-1 and CD3 during conjugation between T cells and antigen-presenting cells (APCs). CD81 and ICAM-1 exhibit distinct mobilities in central and peripheral areas of early and late T cell-APC contacts. Moreover, CD81–ICAM-1 and CD81-CD3 dynamic interactions increase over the time course of IS formation, as these molecules redistribute throughout the contact area. Therefore, CD81 associations unexpectedly define novel sequential steps of IS maturation. Our results indicate that CD81 controls the temporal progression of the IS and the permanence of CD3 in the membrane contact area, contributing to sustained T cell receptor (TCR)-CD3-mediated signaling. Accordingly, we find that CD81 is required for proper T cell activation, regulating CD3ζ, ZAP-70, LAT, and extracellular signal-regulated kinase (ERK) phosphorylation; CD69 surface expression; and interleukin-2 (IL-2) secretion. Our data demonstrate the important role of CD81 in the molecular organization and dynamics of the IS architecture that sets the signaling threshold in T cell activation. PMID:23858057

Comparison of basophil activation tests using CD63 or CD203c expression in patients with insect venom allergy.

PubMed

Eberlein-König, B; Varga, R; Mempel, M; Darsow, U; Behrendt, H; Ring, J

2006-09-01

Flow cytometric basophil activation tests have been developed as cellular tests for in vitro diagnosis of IgE-mediated reactions. Different activation markers (CD63 or CD203c) with distinct ways of regulation have been used after stimulation with various allergens. It was the aim of the present study to compare basophil activation tests by measuring both CD63 and CD203c upregulation in patients with insect venom allergy. 43 patients with a history of insect venom anaphylaxis were examined. A careful allergy history was taken, and skin tests and determination of specific IgE-antibodies were performed. Basophil activation tests (BAT) using CD63 or CD203c expression were done after stimulation with different concentrations of bee and wasp venom extracts. 25 healthy subjects with negative history of insect venom allergy were studied as controls. The CD203c protocol showed a slightly higher sensitivity than the CD63 protocol (97% vs. 89%) with regard to patients' history. The magnitude of basophil response was higher with CD203c in comparison to CD63 for both insect venoms. Specificity was 100% for the CD63 protocol and 89% for the CD203c protocol with regard to controls with negative history and negative RAST. These results support the reliability of basophil activation tests using either CD63 or CD203c as cellular tests in the in vitro diagnosis of patients with bee or wasp venom allergy with a slightly higher sensitivity for the CD203c protocol.

The NOD2 p.Leu1007fsX1008 mutation (rs2066847) is a stronger predictor of the clinical course of Crohn's disease than the FOXO3A intron variant rs12212067.

PubMed

Schnitzler, Fabian; Friedrich, Matthias; Wolf, Christiane; Angelberger, Marianne; Diegelmann, Julia; Olszak, Torsten; Beigel, Florian; Tillack, Cornelia; Stallhofer, Johannes; Göke, Burkhard; Glas, Jürgen; Lohse, Peter; Brand, Stephan

2014-01-01

Very recently, a sub-analysis of genome-wide association scans revealed that the non-coding single nucleotide polymorphism (SNP) rs12212067 in the FOXO3A gene is associated with a milder course of Crohn's disease (CD) (Cell 2013;155:57-69). The aim of our study was to evaluate the clinical value of the SNP rs12212067 in predicting the severity of CD by correlating CD patient genotype status with the most relevant complications of CD such as stenoses, fistulas, and CD-related surgery. We genotyped 550 CD patients for rs12212067 (FOXO3A) and the three common CD-associated NOD2 mutations rs2066844, rs2066847, and rs2066847 and performed genotype-phenotype analyses. No significant phenotypic differences were found between the wild-type genotype TT of the FOXO3A SNP rs12212067 and the minor genotypes TG and GG independently from NOD2 variants. The allele frequency of the minor G allele was 12.7%. Age at diagnosis, disease duration, body mass index, surgery rate, stenoses, fistula, need for immunosuppressive therapy, and disease course were not significantly different. In contrast, the NOD2 mutant p.Leu1007fsX1008 (rs2066847) was highly associated with penetrating CD (p = 0.01), the development of fistulas (p = 0.01) and stenoses (p = 0.01), and ileal disease localization (p = 0.03). Importantly, the NOD2 SNP rs2066847 was a strong separator between an aggressive and a mild course of CD (p = 2.99×10(-5)), while the FOXO3A SNP rs12212067 did not separate between mild and aggressive CD behavior in our cohort (p = 0.35). 96.2% of the homozygous NOD2 p.Leu1007fsX1008 carriers had an aggressive disease behavior compared to 69.3% of the patients with the NOD2 wild-type genotype (p = 0.007). In clinical practice, the NOD2 variant p.Leu1007fsX1008 (rs2066847), in particular in homozygous form, is a much stronger marker for a severe clinical phenotype than the FOXO3A rs12212067 SNP for a mild disease course on an individual patient level despite its important impact on the inflammatory response of monocytes.

Porcine cluster of differentiation (CD) markers 2018 update.

PubMed

Dawson, Harry D; Lunney, Joan K

2018-06-01

Pigs are a major source of food worldwide; preventing and treating their infectious diseases is essential, requiring a thorough understanding of porcine immunity. The use of pigs as models for human physiology is a growing area; progress in this area has been limited because the immune toolkit is not robust. The international community has established cluster of differentiation (CD) markers for assessing cells involved in immunity as well as characterizing numerous other cells like stem cells. Overall, for humans 419 proteins have been designated as CD markers, each reacting with a defined set of antibodies (Abs). This paper summarizes current knowledge of swine CD markers and identifies 359 corresponding CD proteins in pigs. A broad-based literature and vendor search was conducted to identify defined sets of monoclonal (mAbs) and polyclonal Abs (pAbs) reacting with porcine CD markers along with other reagents (fusion proteins, ELISAs, PCR assays, and gene edited cell and pig models). This process identified over 800 reagents that are reportedly reactive with 266 pig CD markers. Despite this number, there is a great need to develop and characterize additional CD marker reagents, particularly mAbs, for pig research. There are numerous high priority targets: reagents for the characterization of porcine innate lymphoid cells, polarized macrophages and T regulatory cells and for the detection of porcine CD45 isoforms. Overall, improved technologies and genomics have contributed to dramatic increases in our knowledge of the pig, its immune system, disease and vaccine responses, and utility as a biomedical model. The development of more CD reagents will clearly advance these initiatives. Published by Elsevier Ltd.

Cysteine-rich Domain 1 of CD40 Mediates Receptor Self-assembly*

PubMed Central

Smulski, Cristian R.; Beyrath, Julien; Decossas, Marion; Chekkat, Neila; Wolff, Philippe; Estieu-Gionnet, Karine; Guichard, Gilles; Speiser, Daniel; Schneider, Pascal; Fournel, Sylvie

2013-01-01

The activation of CD40 on B cells, macrophages, and dendritic cells by its ligand CD154 (CD40L) is essential for the development of humoral and cellular immune responses. CD40L and other TNF superfamily ligands are noncovalent homotrimers, but the form under which CD40 exists in the absence of ligand remains to be elucidated. Here, we show that both cell surface-expressed and soluble CD40 self-assemble, most probably as noncovalent dimers. The cysteine-rich domain 1 (CRD1) of CD40 participated to dimerization and was also required for efficient receptor expression. Modelization of a CD40 dimer allowed the identification of lysine 29 in CRD1, whose mutation decreased CD40 self-interaction without affecting expression or response to ligand. When expressed alone, recombinant CD40-CRD1 bound CD40 with a KD of 0.6 μm. This molecule triggered expression of maturation markers on human dendritic cells and potentiated CD40L activity. These results suggest that CD40 self-assembly modulates signaling, possibly by maintaining the receptor in a quiescent state. PMID:23463508

CD34 Expression by Hair Follicle Stem Cells Is Required for Skin Tumor Development in Mice

PubMed Central

Trempus, Carol S.; Morris, Rebecca J.; Ehinger, Matthew; Elmore, Amy; Bortner, Carl D.; Ito, Mayumi; Cotsarelis, George; Nijhof, Joanne G.W.; Peckham, John; Flagler, Norris; Kissling, Grace; Humble, Margaret M.; King, Leon C.; Adams, Linda D.; Desai, Dhimant; Amin, Shantu; Tennant, Raymond W.

2007-01-01

The cell surface marker CD34 marks mouse hair follicle bulge cells, which have attributes of stem cells, including quiescence and multipotency. Using a CD34 knockout (KO) mouse, we tested the hypothesis that CD34 may participate in tumor development in mice because hair follicle stem cells are thought to be a major target of carcinogens in the two-stage model of mouse skin carcinogenesis. Following initiation with 200 nmol 7,12-dimethylbenz(a)anthracene (DMBA), mice were promoted with 12-O-tetradecanoylphorbol-13-acetate (TPA) for 20 weeks. Under these conditions, CD34KO mice failed to develop papillomas. Increasing the initiating dose of DMBA to 400 nmol resulted in tumor development in the CD34KO mice, albeit with an increased latency and lower tumor yield compared with the wild-type (WT) strain. DNA adduct analysis of keratinocytes from DMBA-initiated CD34KO mice revealed that DMBA was metabolically activated into carcinogenic diol epoxides at both 200 and 400 nmol. Chronic exposure to TPA revealed that CD34KO skin developed and sustained epidermal hyperplasia. However, CD34KO hair follicles typically remained in telogen rather than transitioning into anagen growth, confirmed by retention of bromodeoxyuridine-labeled bulge stem cells within the hair follicle. Unique localization of the hair follicle progenitor cell marker MTS24 was found in interfollicular basal cells in TPA-treated WT mice, whereas staining remained restricted to the hair follicles of CD34KO mice, suggesting that progenitor cells migrate into epidermis differently between strains. These data show that CD34 is required for TPA-induced hair follicle stem cell activation and tumor formation in mice. PMID:17483328

Role of beta-cell autoantibodies as a predictor marker in diabetic patients and their relationship to glycemic control.

PubMed

Ali, Naglaa A; Swelam, Enas; AI Banna, Ehab A; Showkry, Amira

2012-01-01

To evaluate glutamic acid decarboxylase autoantibodies (GAD65), islet cell autoantibodies (ICA) and insulin autoantibodies (IAA) as disease markers and their relationship to certain residual beta-cell function as well as glycemic control among patients with diabetes mellitus. Also, to evaluate of the level of CD4+CD25+(Treg) out of CD4 cells among patients with immune mediated diabetes mellitus (DM). The study included 80 individuals divided into: 40 diabetic patients (group A) and 20 risk siblings (group B) of diabetic father or mother or both. 20 healthy individuals enrolled as control group (group C) all were with no family history of DM. GAD, ICA, IAA autoantibodies and C-peptide were determined by ELISA. HbA1 by ion exchange chromatography and measurement of the expression of CD4+CD25+ (T reg) by flowcytometry. The most frequently encountered antibody in adult and children groups was GAD65, followed by ICA. But in risk group the most frequently antibody was ICA, followed by GAD. In the risk group, there was no statistical difference in the level of CD4+CD25+ in comparison with control group. There was significant decrease in the percentage of CD4+CD25+ in adult and children patients groups with positive autoantibodies than those with negative autoantibodies. In conclusions, at the time of diagnosis the majority of patients with type I diabetes have autoantibodies that are reactive to islet antigens. GAD, ICA, IAA are of value for predicting IDDM in sibling of diabetic parents type I. CD4+CD25+ Treg cells may actively suppress activation of the immune system and prevent pathological self-reactivity.

Invariant NKT Cell Response to Dengue Virus Infection in Human

PubMed Central

Matangkasombut, Ponpan; Chan-in, Wilawan; Opasawaschai, Anunya; Pongchaikul, Pisut; Tangthawornchaikul, Nattaya; Vasanawathana, Sirijitt; Limpitikul, Wannee; Malasit, Prida; Duangchinda, Thaneeya; Screaton, Gavin; Mongkolsapaya, Juthathip

2014-01-01

Background Dengue viral infection is a global health threat without vaccine or specific treatment. The clinical outcome varies from asymptomatic, mild dengue fever (DF) to severe dengue hemorrhagic fever (DHF). While adaptive immune responses were found to be detrimental in the dengue pathogenesis, the roles of earlier innate events remain largely uninvestigated. Invariant natural killer T (iNKT) cells represent innate-like T cells that could dictate subsequent adaptive response but their role in human dengue virus infection is not known. We hypothesized that iNKT cells play a role in human dengue infection. Methods Blood samples from a well-characterized cohort of children with DF, DHF, in comparison to non-dengue febrile illness (OFI) and healthy controls at various time points were studied. iNKT cells activation were analyzed by the expression of CD69 by flow cytometry. Their cytokine production was then analyzed after α-GalCer stimulation. Further, the CD1d expression on monocytes, and CD69 expression on conventional T cells were measured. Results iNKT cells were activated during acute dengue infection. The level of iNKT cell activation associates with the disease severity. Furthermore, these iNKT cells had altered functional response to subsequent ex vivo stimulation with α-GalCer. Moreover, during acute dengue infection, monocytic CD1d expression was also upregulated and conventional T cells also became activated. Conclusion iNKT cells might play an early and critical role in the pathogenesis of severe dengue viral infection in human. Targeting iNKT cells and CD1d serve as a potential therapeutic strategy for severe dengue infection in the future. PMID:24945350

Novel Adult Stem Cells for Peripheral Nerve Regeneration

DTIC Science & Technology

2012-09-01

were also positive for MSC surface marker CD29 and CD44 (Fig. 1F-G). However, CD29 and CD44 are also expressed in SMCs, so we will not use these non...tubulin. In addition, MVSCs were negative for perivascular MSC marker CD146 (Fig. 1H) and SMC progenitor marker Sca-1 (Fig. 1I). MVSCs were also...University of California, Berkeley, California 94720, USA. 2 UC Berkeley-UCSF Graduate Program in Bioengineering, Berkeley, California 94720, USA. 3

Helios, and not FoxP3, is the marker of activated Tregs expressing GARP/LAP

PubMed Central

Elkord, Eyad; Abd Al Samid, May; Chaudhary, Belal

2015-01-01

Regulatory T cells (Tregs) are key players of immune regulation/dysregulation both in physiological and pathophysiological settings. Despite significant advances in understanding Treg function, there is still a pressing need to define reliable and specific markers that can distinguish different Treg subpopulations. Herein we show for the first time that markers of activated Tregs [latency associated peptide (LAP) and glycoprotein A repetitions predominant (GARP, or LRRC32)] are expressed on CD4+FoxP3− T cells expressing Helios (FoxP3−Helios+) in the steady state. Following TCR activation, GARP/LAP are up-regulated on CD4+Helios+ T cells regardless of FoxP3 expression (FoxP3+/−Helios+). We show that CD4+GARP+/−LAP+ Tregs make IL-10 immunosuppressive cytokine but not IFN-γ effector cytokine. Further characterization of FoxP3/Helios subpopulations showed that FoxP3+Helios+ Tregs proliferate in vitro significantly less than FoxP3+Helios− Tregs upon TCR stimulation. Unlike FoxP3+Helios− Tregs, FoxP3+Helios+ Tregs secrete IL-10 but not IFN-γ or IL-2, confirming they are bona fide Tregs with immunosuppressive characteristics. Taken together, Helios, and not FoxP3, is the marker of activated Tregs expressing GARP/LAP, and FoxP3+Helios+ Tregs have more suppressive characteristics, compared with FoxP3+Helios− Tregs. Our work implies that therapeutic modalities for treating autoimmune and inflammatory diseases, allergies and graft rejection should be designed to induce and/or expand FoxP3+Helios+ Tregs, while therapies against cancers or infectious diseases should avoid such expansion/induction. PMID:26343373

Elevated pretransplantation soluble CD30 is associated with decreased early allograft function after human lung transplantation.

PubMed

Shah, Ashish S; Leffell, M Sue; Lucas, Donna; Zachary, Andrea A

2009-02-01

Early allograft function after lung transplantation is variable. Clinical criteria have limited predictive value for early graft function. Recipient immunologic state before LTx may affect early lung function. We investigated the association between pretransplantation soluble CD30 (sCD30), a marker of Th2-type T-cell activation, and early clinical parameters of allograft function. Between September 2002 and January 2007, a total of 80 transplantations were performed at Johns Hopkins Hospital. Of the patients, 43 had a pretransplantation sCD30 level determined. Pre- and postoperative patient variables were collected, and patients were stratified into two groups: sCD30 <20 (low sCD30) and >20 (high sCD30). High sCD30 (n = 26) and low sCD30 (n = 17) groups were similar in age, gender, and ischemia time. In the high sCD30 group, a higher percentage of patients had pulmonary fibrosis and a lower percentage had emphysema. Oxygenation at 48 hours was significantly worse in the high sCD30 group as compared with the low sCD30 (p = 0.003). Moreover, prolonged intubation and 90-day mortality were greater in the high sCD30 group. This represents the first report of the use of sCD30 as a marker for early allograft function in human lung transplanation. Increased pretransplantation recipient sCD30 appears to be associated with decreased early post-transplantation gas exchange, prolonged intubation, and early mortality.

Identification of cancer stem cell markers in human malignant mesothelioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ghani, Farhana Ishrat; Yamazaki, Hiroto; Iwata, Satoshi

2011-01-14

Research highlights: {yields} We performed serial transplantation of surgical samples and established new cell lines of malignant mesothelioma. {yields} SP cell and expressions of CD9/CD24/CD26 were often observed in mesothelioma cell lines. {yields} SP and CD24{sup +} cells proliferated by asymmetric cell division-like manner. CD9{sup +} and CD24{sup +} cells have higher potential to generate spheroid colony. {yields} The marker-positive cells have clear tendency to generate larger tumors in mice. -- Abstract: Malignant mesothelioma (MM) is an aggressive and therapy-resistant neoplasm arising from the pleural mesothelial cells and usually associated with long-term asbestos exposure. Recent studies suggest that tumors containmore » cancer stem cells (CSCs) and their stem cell characteristics are thought to confer therapy-resistance. However, whether MM cell has any stem cell characteristics is not known. To understand the molecular basis of MM, we first performed serial transplantation of surgical samples into NOD/SCID mice and established new cell lines. Next, we performed marker analysis of the MM cell lines and found that many of them contain SP cells and expressed several putative CSC markers such as CD9, CD24, and CD26. Interestingly, expression of CD26 closely correlated with that of CD24 in some cases. Sorting and culture assay revealed that SP and CD24{sup +} cells proliferated by asymmetric cell division-like manner. In addition, CD9{sup +} and CD24{sup +} cells have higher potential to generate spheroid colony than negative cells in the stem cell medium. Moreover, these marker-positive cells have clear tendency to generate larger tumors in mouse transplantation assay. Taken together, our data suggest that SP, CD9, CD24, and CD26 are CSC markers of MM and could be used as novel therapeutic targets.« less

Metal availability, soil nutrient, and enzyme activity in response to application of organic amendments in Cd-contaminated soil.

PubMed

Yang, Zhanbiao; Liu, Lixia; Lv, Yanfeng; Cheng, Zhang; Xu, Xiaoxun; Xian, Junren; Zhu, Xuemei; Yang, Yuanxiang

2018-01-01

The study investigated the effects of organic amendments: green tea amendment (GTA) and oil cake amendment (OCA) on Cd bioavailability, soil nutrients, and soil enzyme activity in Cd-contaminated soil. The amendments were added to the soil at the doses of 1, 3, and 5% and were incubated for 45 days. Then, pakchoi cabbage was planted to test the remediation effect of the above two organic amendments. The diethylenetriaminepentaacetic acid (DTPA)-extractable Cd in GTA and OCA treatments was reduced by 14.69-27.51 and 13.75-68.77%, respectively, compared to no amendment-applied treatment. The application of GTA and OCA notably decreased the proportion of exchangeable fraction of Cd, but increased the percentage of oxide and organic-bound fraction of Cd, thereby suppressing the uptake by pakchoi cabbage. Cd concentration of aboveground parts decreased by 8.21-18.05 and 7.77-35.89% in GTA and OCA treatments, respectively. Relative to the no amendment-applied treatment, both GTA and OCA had enhanced soil nutrients and enzyme activities largely. Redundancy analysis showed that organic matter, total P, available N, and DTPA-extractable Cd significantly affected the enzyme activities. Furthermore, the application of OCA at the dose of 5% was more effective in reducing bioavailable Cd, enhancing soil available nutrients and urease and catalase activities in contaminated soil. These results indicated that oil cake should be used to immobilize metal and improve fertility and quality of Cd-contaminated soil.

Group 2 Innate Lymphoid Cells Exhibit a Dynamic Phenotype in Allergic Airway Inflammation

PubMed Central

Li, Bobby W. S.; Stadhouders, Ralph; de Bruijn, Marjolein J. W.; Lukkes, Melanie; Beerens, Dior M. J. M.; Brem, Maarten D.; KleinJan, Alex; Bergen, Ingrid; Vroman, Heleen; Kool, Mirjam; van IJcken, Wilfred F. J.; Rao, Tata Nageswara; Fehling, Hans Jörg; Hendriks, Rudi W.

2017-01-01

Group 2 innate lymphoid cells (ILC2) are implicated in allergic asthma as an early innate source of the type 2 cytokines IL-5 and IL-13. However, their induction in house dust mite (HDM)-mediated airway inflammation additionally requires T cell activation. It is currently unknown whether phenotypic differences exist between ILC2s that are activated in a T cell-dependent or T cell-independent fashion. Here, we compared ILC2s in IL-33- and HDM-driven airway inflammation. Using flow cytometry, we found that surface expression levels of various markers frequently used to identify ILC2s were dependent on their mode of activation, highly variable over time, and differed between tissue compartments, including bronchoalveolar lavage (BAL) fluid, lung, draining lymph nodes, and spleen. Whereas in vivo IL-33-activated BAL fluid ILC2s exhibited an almost uniform CD25+CD127+T1/ST2+ICOS+KLRG1+ phenotype, at a comparable time point after HDM exposure BAL fluid ILC2s had a very heterogeneous surface marker phenotype. A major fraction of HDM-activated ILC2s were CD25lowCD127+T1/ST2low ICOSlowKLRG1low, but nevertheless had the capacity to produce large amounts of type 2 cytokines. HDM-activated CD25low ILC2s in BAL fluid and lung rapidly reverted to CD25high ILC2s upon in vivo stimulation with IL-33. Genome-wide transcriptional profiling of BAL ILC2s revealed ~1,600 differentially expressed genes: HDM-stimulated ILC2s specifically expressed genes involved in the regulation of adaptive immunity through B and T cell interactions, whereas IL-33-stimulated ILC2s expressed high levels of proliferation-related and cytokine genes. In both airway inflammation models ILC2s were present in the lung submucosa close to epithelial cells, as identified by confocal microscopy. In chronic HDM-driven airway inflammation ILC2s were also found inside organized cellular infiltrates near T cells. Collectively, our findings show that ILC2s are phenotypically more heterogeneous than previously thought, whereby their surface marker and gene expression profile are highly dynamic. PMID:29250067

An Epstein-Barr virus-associated superantigen

PubMed Central

1996-01-01

More than 90% of adults are latently infected with Epstein-Barr virus (EBV), the causative agent of infectious mononucleosis, a self-limiting lymphoproliferative disease characterized by extensive T cell activation. Reactivation of this herpesvirus during immunosuppression is often associated with oncogenesis. These considerations led us to analyze the early events that occur after exposure of the immune system to EBV. Strong major histocompatibility complex (MHC) class II- dependent but not MHC-restricted, T cell proliferation was observed in vitro in response to autologous, lytically infected EBV-transformed B cells. By measuring the appearance of the early activation marker CD69 on individual T cell V beta subsets, we could demonstrate selective activation of human V beta 13- T cells. This was confirmed with murine T cell hybridomas expressing various human BV genes. While EBV- Burkitt's lymphoma cells were nonstimulatory, they induced V beta- restricted T cell activation after EBV infection. EBV specific activation was also demonstrated in cord blood cells, excluding a recall-antigen response. Thus, all of the characteristics of a superantigen-stimulated response are seen, indicating that induction of the EBV lytic cycle is associated with the expression of a superantigen in B cells. A model is presented proposing a role for the superantigen in infection, latency, and oncogenesis. PMID:9064357

A novel three-dimensional heterotypic spheroid model for the assessment of the activity of cancer immunotherapy agents.

PubMed

Herter, Sylvia; Morra, Laura; Schlenker, Ramona; Sulcova, Jitka; Fahrni, Linda; Waldhauer, Inja; Lehmann, Steffi; Reisländer, Timo; Agarkova, Irina; Kelm, Jens M; Klein, Christian; Umana, Pablo; Bacac, Marina

2017-01-01

The complexity of the tumor microenvironment is difficult to mimic in vitro, particularly regarding tumor-host interactions. To enable better assessment of cancer immunotherapy agents in vitro, we developed a three-dimensional (3D) heterotypic spheroid model composed of tumor cells, fibroblasts, and immune cells. Drug targeting, efficient stimulation of immune cell infiltration, and specific elimination of tumor or fibroblast spheroid areas were demonstrated following treatment with a novel immunocytokine (interleukin-2 variant; IgG-IL2v) and tumor- or fibroblast-targeted T cell bispecific antibody (TCB). Following treatment with IgG-IL2v, activation of T cells, NK cells, and NKT cells was demonstrated by increased expression of the activation marker CD69 and enhanced cytokine secretion. The combination of TCBs with IgG-IL2v molecules was more effective than monotherapy, as shown by enhanced effects on immune cell infiltration; activation; increased cytokine secretion; and faster, more efficient elimination of targeted cells. This study demonstrates that the 3D heterotypic spheroid model provides a novel and versatile tool for in vitro evaluation of cancer immunotherapy agents and allows for assessment of additional aspects of the activity of cancer immunotherapy agents, including analysis of immune cell infiltration and drug targeting.

Elevated Endomyocardial Biopsy Macrophage-Related Markers in Intractable Myocardial Diseases.

PubMed

Hayashi, Yuka; Hanawa, Haruo; Jiao, Shuang; Hasegawa, Go; Ohno, Yukako; Yoshida, Kaori; Suzuki, Tomoyasu; Kashimura, Takeshi; Obata, Hiroaki; Tanaka, Komei; Watanabe, Tohru; Minamino, Tohru

2015-12-01

Tissue macrophages can be activated by endogenous danger signals released from cells that are stressed or injured, leading to infiltration of inflammatory macrophages and neutrophils. We postulated that macrophage-related markers might be closely associated with the existence of endogenous danger signals, reflecting ongoing tissue injury in the absence of foreign substances. This study was designed to assess the ability of macrophage-related markers in endomyocardial biopsies to predict ongoing cardiac injury in non-inflammatory myocardial diseases. We examined levels of macrophage-related markers (CD68, CD163, CD45) in endomyocardial biopsies from patients (n = 86) with various myocardial diseases by quantitative reverse transcription-polymerase chain reaction (n = 78) and immunohistochemistry (n = 56). Thirty-three patients without inflammatory cardiac disease such as myocarditis and sarcoidosis were classified as "improved" or "non-improved" defined as a 10% increase in left ventricular ejection fraction by echocardiograph and a value greater than 30% at the time of follow-up. All macrophage-related (MacR) markers levels were not higher in non-improved dilated cardiomyopathy (DCM) patients than improved patients. However, patients with cardiac amyloidosis, cardiac Fabry disease, mitochondrial cardiomyopathy, and biventricular arrhythmogenic right ventricular cardiomyopathy (ARVC), which were categorized as "non-improvement diseases," had elevated macrophage-related markers compared to improved patients. Macrophage-related markers levels were increased in endomyocardial biopsy samples of patients with intractable myocardial diseases such as amyloidosis, mitochondrial disease, Fabry disease, and biventricular ARVC.

The CD117 immunohistochemistry tissue microarray survey for quality assurance and interlaboratory comparison: a College of American Pathologists Cell Markers Committee Study.

PubMed

Dorfman, David M; Bui, Marilyn M; Tubbs, Raymond R; Hsi, Eric D; Fitzgibbons, Patrick L; Linden, Michael D; Rickert, Robert R; Roche, Patrick C

2006-06-01

We have developed tissue microarray-based surveys to allow laboratories to compare their performance in staining predictive immunohistochemical markers, including proto-oncogene CD117 (c-kit), which is characteristically expressed in gastrointestinal stromal tumors (GISTs). GISTs exhibit activating mutations in the c-kit proto-oncogene, which render them amenable to treatment with imatinib mesylate. Consequently, correct identification of c-Kit expression is important for the diagnosis and treatment of GISTs. To analyze CD117 immunohistochemical staining performance by a large number of clinical laboratories. A mechanical device was used to construct tissue microarrays consisting of 3 x 1-mm cores of 10 tumor samples, which can be used to generate hundreds of tissue sections from the arrayed cases, suitable for large-scale interlaboratory comparison of immunohistochemical staining. An initial survey of 63 laboratories and a second survey of 90 laboratories, performed in 2004 and 2005, exhibited >81% concordance for 7 of 10 cores, including all 4 GIST cases, which were immunoreactive for CD117 with >95% staining concordance. Three of the cores achieved less than 81% concordance of results, possibly due to the presence of foci of necrosis in one core and CD117-positive mast cells in 2 cores of CD117-negative neoplasms. There was good performance among a large number of laboratories performing CD117 immunohistochemical staining, with consistently higher concordance of results for CD117-positive GIST cases than for nonimmunoreactive cases. Tissue microarrays for CD117 and other predictive markers should be useful for interlaboratory comparisons, quality assurance, and education of participants regarding staining nuances such as the expression of CKIT by nonneoplastic mast cells.

Single-Stage Preparation of Human Cartilage Grafts Generated from Bone Marrow-Derived CD271+ Mononuclear Cells.

PubMed

Petters, Oliver; Schmidt, Christian; Henkelmann, Ralf; Pieroh, Philipp; Hütter, Gero; Marquass, Bastian; Aust, Gabriela; Schulz, Ronny M

2018-04-15

Due to the limited self-healing capacity of articular cartilage, innovative, regenerative approaches are of particular interest. The use of two-stage procedures utilizing in vitro-expanded mesenchymal stromal cells (MSCs) from various cell sources requires good manufacturing practice-compliant production, a process with high demands on time, staffing, and financial resources. In contrast, one- stage procedures are directly available, but need a safe enrichment of potent MSCs. CD271 is a surface marker known to marking the majority of native MSCs in bone marrow (BM). In this study, the feasibility of generating a single-stage cartilage graft of enriched CD271 + BM-derived mononuclear cells (MNCs) without in vitro monolayer expansion from eight healthy donors was investigated. Cartilage grafts were generated by magnetic-activated cell sorting and separated cells were directly transferred into collagen type I hydrogels, followed by 3D proliferation and differentiation period of CD271 + , CD271 - , or unseparated MNCs. CD271 + MNCs showed the highest proliferation rate, cell viability, sulfated glycosaminoglycan deposition, and cartilage marker expression compared to the CD271 - or unseparated MNC fractions in 3D culture. Analysis according to the minimal criteria of the International Society for Cellular Therapy highlighted a 66.8-fold enrichment of fibroblast colony-forming units in CD271 + MNCs and the only fulfillment of the MSC marker profile compared to unseparated MNCs. In summary, CD271 + MNCs are capable of generating adequate articular cartilage grafts presenting high cell viability and notable chondrogenic matrix deposition in a CE-marked collagen type I hydrogel, which can obviate the need for an initial monolayer expansion.

Identification and validation of multiple cell surface markers of clinical-grade adipose-derived mesenchymal stromal cells as novel release criteria for good manufacturing practice-compliant production.

PubMed

Camilleri, Emily T; Gustafson, Michael P; Dudakovic, Amel; Riester, Scott M; Garces, Catalina Galeano; Paradise, Christopher R; Takai, Hideki; Karperien, Marcel; Cool, Simon; Sampen, Hee-Jeong Im; Larson, A Noelle; Qu, Wenchun; Smith, Jay; Dietz, Allan B; van Wijnen, Andre J

2016-08-11

Clinical translation of mesenchymal stromal cells (MSCs) necessitates basic characterization of the cell product since variability in biological source and processing of MSCs may impact therapeutic outcomes. Although expression of classical cell surface markers (e.g., CD90, CD73, CD105, and CD44) is used to define MSCs, identification of functionally relevant cell surface markers would provide more robust release criteria and options for quality control. In addition, cell surface expression may distinguish between MSCs from different sources, including bone marrow-derived MSCs and clinical-grade adipose-derived MSCs (AMSCs) grown in human platelet lysate (hPL). In this work we utilized quantitative PCR, flow cytometry, and RNA-sequencing to characterize AMSCs grown in hPL and validated non-classical markers in 15 clinical-grade donors. We characterized the surface marker transcriptome of AMSCs, validated the expression of classical markers, and identified nine non-classical markers (i.e., CD36, CD163, CD271, CD200, CD273, CD274, CD146, CD248, and CD140B) that may potentially discriminate AMSCs from other cell types. More importantly, these markers exhibit variability in cell surface expression among different cell isolates from a diverse cohort of donors, including freshly prepared, previously frozen, or proliferative state AMSCs and may be informative when manufacturing cells. Our study establishes that clinical-grade AMSCs expanded in hPL represent a homogeneous cell culture population according to classical markers,. Additionally, we validated new biomarkers for further AMSC characterization that may provide novel information guiding the development of new release criteria. Use of Autologous Bone Marrow Aspirate Concentrate in Painful Knee Osteoarthritis (BMAC): Clinicaltrials.gov NCT01931007 . Registered August 26, 2013. MSC for Occlusive Disease of the Kidney: Clinicaltrials.gov NCT01840540 . Registered April 23, 2013. Mesenchymal Stem Cell Therapy in Multiple System Atrophy: Clinicaltrials.gov NCT02315027 . Registered October 31, 2014. Efficacy and Safety of Adult Human Mesenchymal Stem Cells to Treat Steroid Refractory Acute Graft Versus Host Disease. Clinicaltrials.gov NCT00366145 . Registered August 17, 2006. A Dose-escalation Safety Trial for Intrathecal Autologous Mesenchymal Stem Cell Therapy in Amyotrophic Lateral Sclerosis. Clinicaltrials.gov NCT01609283 . Registered May 18, 2012.

Evaluation of circulating markers of hepatic apoptosis and inflammation in obese children with and without obstructive sleep apnea.

PubMed

Alkhouri, Naim; Kheirandish-Gozal, Leila; Matloob, Ammar; Alonso-Álvarez, María Luz; Khalyfa, Abdelnaby; Terán-Santos, Joaquin; Okwu, Vera; Lopez, Rocio; Gileles-Hillel, Alex; Dweik, Raed; Gozal, David

2015-09-01

Hepatocyte apoptosis and macrophage activation contribute to the disease progression of nonalcoholic fatty liver disease (NAFLD). Obstructive sleep apnea (OSA) in obese children is associated with the severity of NAFLD. The aim of this study was to evaluate plasma levels of soluble Fas (sFas), soluble Fas ligand (sFasL), cytokeratin 18 (CK18) (markers of apoptosis), and soluble CD163 (sCD163) (marker of macrophage activation) in obese children with and without OSA. Consecutive obese children who were evaluated for OSA were recruited. The diagnosis of OSA was made using overnight polysomnography (PSG). Fasting blood samples were used to determine plasma CK18, sFas, sFasL, and sCD163 levels using specific sandwich enzyme-linked immunosorbent assay (ELISA). Fifty-eight subjects were included in the analysis with a mean age of 8.9 ± 3.2 years and a mean body mass index (BMI) z-score of 2.4 ± 0.49. Circulating sFas and sFasL levels were significantly lower in subjects with mild and severe OSA compared with those without OSA (p < 0.005 for both). In addition, sCD163 levels increased with an increasing severity of OSA (no OSA = 1.6 ± 0.25 mg/L, mild OSA = 2.3 ± 0.45, and severe OSA = 3.0 ± 0.52; p < 0.001), and they correlated with the apnea-hypopnea index (AHI) [rho (95% confidence interval, CI) of 0.71 (0.41, 1.00), p-value <0.001]. In six patients with severe OSA from whom samples were taken before and after tonsillectomy, the sCD163 level decreased significantly after treatment, and there was a trend toward an increase in sFasL. Markers of apoptosis and macrophage activation are altered in obese children with OSA, indicating increased apoptotic and inflammatory pressures. Copyright © 2015 Elsevier B.V. All rights reserved.

Effectiveness of Vascular Markers (Immunohistochemical Stains) in Soft Tissue Sarcomas.

PubMed

Naeem, Namra; Mushtaq, Sajid; Akhter, Noreen; Hussain, Mudassar; Hassan, Usman

2018-05-01

To ascertain the effectiveness of IHC markers of vascular origin like CD31, CD34, FLI1 and ERG in vascular soft tissue sarcomas including angiosarcomas, Kaposi sarcomas, epithelioid hemangioendothelioma and a non-vascular soft tissue sarcoma (Epithelioid sarcoma). Descriptive study. Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore, from 2011 to 2017. Diagnosed cases of angiosarcomas (n=48), epithelioid hemangioendothelioma (n=9), Kaposi sarcoma (n=9) and epithelioid sarcoma (n=20) were selected. Immunohistochemical staining as performed on formalin fixed paraffin embedded sections. The sections were stained for the following markers: CD34 (VENTANA clone Q Bend 10), CD31 (Leica clone 1 A 10), FLI1 (CELL MARQUE clone MRQ-1) and ERG (CELL MARQUE clone EP111). A complete panel of CD34, CD31 and ERG was applied on 8/48 cases of angiosarcomas with triple positivity in 6 cases. Eight cases showed positivity for only CD31 and ERG and 2 cases showed positivity for only ERG. A complete panel of CD34, CD31 and ERG was applied on 3/9 cases of epithelioid hemangioendothelioma with positivity for all markers in 2 cases. Combined positivity for ERG and CD34 was seen in 2 cases and on 4 cases only CD31 immunohistochemical was solely applied with 100% positivity. FLI1 was not applied on any case. Among 9 cases of Kaposi sarcoma, ERG, CD34 and CD31 in combination were applied on only 1 case with triple positivity. Remaining cases show positivity for either CD34, CD31 or FLI1. Majority of cases of epithelioid sarcomas were diagnosed on the basis of cytokeratin and CD34 positivity with loss of INI1. The other vascular markers showed negativity in all cases. Among these four markers, ERG immunohistochemical stain is highly effective for endothelial differentiation due to its specific nuclear staining pattern in normal blood vessel endothelial cells (internal control) as well as neoplastic cells of vascular tumors and lack of background staining.

FcγRIIb expression in early stage chronic lymphocytic leukemia.

PubMed

Bosch, Rosa; Mora, Alba; Vicente, Eva Puy; Ferrer, Gerardo; Jansà, Sonia; Damle, Rajendra; Gorlatov, Sergey; Rai, Kanti; Montserrat, Emili; Nomdedeu, Josep; Pratcorona, Marta; Blanco, Laura; Saavedra, Silvana; Garrido, Ana; Esquirol, Albert; Garcia, Irene; Granell, Miquel; Martino, Rodrigo; Delgado, Julio; Sierra, Jorge; Chiorazzi, Nicholas; Moreno, Carol

2017-11-01

In normal B-cells, B-cell antigen receptor (BCR) signaling can be negatively regulated by the low-affinity receptor FcγRIIb (CD32b). To better understand the role of FcγRIIb in chronic lymphocytic leukemia (CLL), we correlated its expression on 155 samples from newly-diagnosed Binet A patients with clinical characteristics and outcome. FcγRIIb expression was similar in normal B-cells and leukemic cells, this being heterogenous among patients and within CLL clones. FcγRIIb expression did not correlate with well known prognostic markers [disease stage, serum beta-2 microglobulin (B2M), IGHV mutational status, expression of ZAP-70 and CD38, and cytogenetics] except for a weak concordance with CD49d. Moreover, patients with low FcγRIIb expression (69/155, 44.5%) required therapy earlier than those with high FcγRIIb expression (86/155, 55.5%) (median 151.4 months vs. not reached; p=.071). These results encourage further investigation on the role of FcγRIIb in CLL biology and prognostic significance in larger series of patients.

Modulation of the immune response by infection with Cryptosporidium spp. in children with allergic diseases.

PubMed

Guangorena-Gómez, J O; Maravilla-Domínguez, A; García-Arenas, G; Cervantes-Flores, M; Meza-Velázquez, R; Rivera-Guillén, M; Acosta-Saavedra, L C; Goytia-Acevedo, R C

2016-08-01

It has been demonstrated that the allergic response can be ameliorated by the administration of pathogen derivatives that activate Toll-like receptors and induce a Th1-type immune response (IR). Cryptosporidium is a parasite that promotes an IR via Toll-like receptors and elicits the production of Th1-type cytokines, which limit cryptosporidiosis. The aim of this study was to investigate allergy-related immune markers in children naturally infected with Cryptosporidium. In a cross-sectional study, 49 children with or without clinical diagnosis of allergies, oocysts of Cryptosporidium spp. in the faeces were screened microscopically. We microscopically screened for leucocytes, examined T and B cells for allergy-related activation markers using flow cytometry and evaluated serum for total IgE using chemiluminescence. Children with allergies and Cryptosporidium in the faeces had significantly lower levels of total IgE, B cells, CD19(+) CD23(+) and CD19(+) CD124(+) cells as well as a greater percentage of interferon-gamma (IFN-γ(+) ) and IL-4(+) CD4(+) cells than children with allergies without Cryptosporidium. This is the first description of the modulation of the IR in children with allergic diseases in the setting of natural Cryptosporidium infection. Our findings suggest the involvement of CD4(+) cells producing IL-4 and IFN-γ in the IR to Cryptosporidium in naturally infected children. © 2016 John Wiley & Sons Ltd.

Bifidobacterium infantis NLS Super Strain Reduces the Expression of α-Defensin-5, a Marker of Innate Immunity, in the Mucosa of Active Celiac Disease Patients.

PubMed

Pinto-Sánchez, Maria I; Smecuol, Edgardo C; Temprano, Maria P; Sugai, Emilia; González, Andrea; Moreno, María L; Huang, Xianxi; Bercik, Premysl; Cabanne, Ana; Vázquez, Horacio; Niveloni, Sonia; Mazure, Roberto; Mauriño, Eduardo; Verdú, Elena F; Bai, Julio C

2017-10-01

We have previously shown a reduction of gastrointestinal symptoms after the oral administration of Bifidobacterium infantis Natren Life Start super strain (NLS-SS) in untreated celiac disease (CD) patients. The symptomatic improvement was not associated with changes in intestinal permeability or serum levels of cytokines, chemokines, or growth factors. Therefore, we hypothesized that the beneficial symptomatic effect observed previously in patients with CD treated with B. infantis may be related to the modulation of innate immunity. To investigate the potential mechanisms of a probiotic B. infantis Natren Life Start super strain on the mucosal expression of innate immune markers in adult patients with active untreated CD compared with those treated with B. infantis×6 weeks and after 1 year of gluten-free diet (GFD). Numbers of macrophages and Paneth cells and α-defensin-5 expression were assessed by immunohistochemistry in duodenal biopsies. We showed that GFD decreases duodenal macrophage counts in CD patients more effectively than B. infantis. In contrast, B. infantis decreases Paneth cell counts and expression of α-defensin-5 in CD (P<0.001). The results identify differential innate immune effects of treatment with B. infantis compared with 1 year of GFD. Further studies are needed to investigate synergistic effects of GFD and B. infantis supplementation in CD.

Characterization of resident lymphocytes in human pancreatic islets

PubMed Central

Radenkovic, M.; Uvebrant, K.; Skog, O.; Sarmiento, L.; Avartsson, J.; Storm, P.; Vickman, P.; Bertilsson, P.‐A.; Fex, M.; Korgsgren, O.

2016-01-01

Summary The current view of type 1 diabetes (T1D) is that it is an immune‐mediated disease where lymphocytes infiltrate the pancreatic islets, promote killing of beta cells and cause overt diabetes. Although tissue resident immune cells have been demonstrated in several organs, the composition of lymphocytes in human healthy pancreatic islets have been scarcely studied. Here we aimed to investigate the phenotype of immune cells associated with human islets of non‐diabetic organ donors. A flow cytometry analysis of isolated islets from perfused pancreases (n = 38) was employed to identify alpha, beta, T, natural killer (NK) and B cells. Moreover, the expression of insulin and glucagon transcripts was evaluated by RNA sequencing. Up to 80% of the lymphocytes were CD3+ T cells with a remarkable bias towards CD8+ cells. Central memory and effector memory phenotypes dominated within the CD8+ and CD4+ T cells and most CD8+ T cells were positive for CD69 and up to 50–70% for CD103, both markers of resident memory cells. The frequency of B and NK cells was low in most islet preparations (12 and 3% of CD45+ cells, respectively), and the frequency of alpha and beta cells varied between donors and correlated clearly with insulin and glucagon mRNA expression. In conclusion, we demonstrated the predominance of canonical tissue resident memory CD8+ T cells associated with human islets. We believe that these results are important to understand more clearly the immunobiology of human islets and the disease‐related phenotypes observed in diabetes. PMID:27783386

Comparative characterization of stem cell marker expression, metabolic activity and resistance to doxorubicin in adherent and spheroid cells derived from the canine prostate adenocarcinoma cell line CT1258.

PubMed

Liu, Wen; Moulay, Mohammed; Willenbrock, Saskia; Roolf, Catrin; Junghanss, Christian; Ngenazahayo, Anaclet; Nolte, Ingo; Murua Escobar, Hugo

2015-04-01

Canine prostate cancer represents a spontaneous animal model for the human counterpart. Cells with stem cell-like character are considered to play a major role in therapeutic resistance and tumor relapse. Thus, the identification of markers allowing for recognition and characterization of these cells is essential. Expression of 12 stem cell marker genes in the canine prostate cancer cell line CT1258 and spheroid cells generated from these was analyzed by quantitative real-time PCR. In CT1258 and the generated spheroid cells, CD44 and CD133 expression was analyzed by flow cytometry, as well as proliferation and doxorubicin resistance. Integrin alpha-6 (ITGA6) expression and metabolic activity were significantly up-regulated in CT1258-derived spheroid cells, while doxorubicin resistance remained comparable. ITGA6 de-regulation and metabolic activity appear to be characteristic of the generated spheres, indicating potential intervention targets. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Increased expression of CD44 is associated with more aggressive behavior in clear cell renal cell carcinoma.

PubMed

Zanjani, Leili Saeednejad; Madjd, Zahra; Abolhasani, Maryam; Rasti, Arezoo; Fodstad, Oystein; Andersson, Yvonne; Asgari, Mojgan

2018-01-01

Although CD44 has been suggested as a prognostic marker in renal cell carcinoma (RCC), the prognostic significance of this marker in three main subtypes of RCC is still unclear. Thus, the present study was conducted to evaluate the expression and prognostic significance of CD44 as a cancer stem cell marker in different histological subtypes of RCC. Methodology & results: CD44 expression was evaluated in 206 well-defined renal tumor samples using immunohistochemistry on tissue microarrays. Higher CD44 expression was associated with more aggressive behavior, tumor progression and worse prognosis in clear cell RCC (ccRCC) but not in papillary and chromophobe RCC subtypes. Cancer stem cell marker CD44 may be a promising target for cancer treatment only in ccRCC.

Plasma markers of B-cell activation and clonality in pediatric liver and hematopoietic stem cell transplant recipients

PubMed Central

Engels, Eric A.; Savoldo, Barbara; Pfeiffer, Ruth M.; Costello, Rene; Zingone, Adriana; Heslop, Helen E.; Landgren, Ola

2012-01-01

Introduction Transplant recipients are at risk of post-transplant lymphoproliferative disease (PTLD). Methods: Thirty-six pediatric transplant recipients were evaluated (18 hematopoietic stem cell and 18 liver recipients; 12 had PTLD). We studied 207 longitudinal plasma samples from these recipients for three markers of B-cell activation or clonality: immunoglobulin free light chains (FLCs), soluble CD30 (sCD30), and monoclonal immunoglobulins (M-proteins). Results Kappa FLCs, lambda FLCs, and sCD30 were elevated in 20.8%, 28.0%, and 94.2% of plasma specimens, respectively. FLC and sCD30 levels increased significantly 1.18–1.82 fold per log10 Epstein Barr virus (EBV) load in peripheral blood. Five PTLD cases manifested elevated FLCs with an abnormal kappa/lambda ratio, suggesting monoclonal FLC production. M-proteins were present in 91% of PTLD cases, vs. 50–67% of other recipients with high or low EBV loads (p=0.13). Concordance of FLCs, M-proteins, and PTLD tumor light chain restriction was imperfect. For example, one PTLD case with an IgG lambda M-protein had a tumor that was kappa restricted, and another case with an M-protein had a T-cell PTLD. In an additional case, an IgM kappa M-protein and excess kappa FLCs were both detected in plasma at PTLD diagnosis; while the tumor was not restricted at diagnosis, kappa restriction was present 5 years later when the PTLD relapsed. Discussion Plasma markers of B-cell dysfunction are frequent following transplantation and associated with poor EBV control. These abnormal markers may be produced by oligoclonal B-cell populations or PTLD tumor cells, and could potentially help identify recipients at high risk of PTLD. PMID:23222884

CD147, CD44, and the Epidermal Growth Factor Receptor (EGFR) Signaling Pathway Cooperate to Regulate Breast Epithelial Cell Invasiveness*

PubMed Central

Grass, G. Daniel; Tolliver, Lauren B.; Bratoeva, Momka; Toole, Bryan P.

2013-01-01

The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777–788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer. PMID:23888049

MMP‐2 and MMP‐14 Silencing Inhibits VEGFR2 Cleavage and Induces the Differentiation of Porcine Adipose‐Derived Mesenchymal Stem Cells to Endothelial Cells

PubMed Central

Almalki, Sami G.; Llamas Valle, Yovani

2017-01-01

Abstract The molecular mechanisms that control the ability of adipose‐derived mesenchymal stem cells (AMSCs) to remodel three‐dimensional extracellular matrix barriers during differentiation are not clearly understood. Herein, we studied the expression of matrix metalloproteinases (MMPs) during the differentiation of AMSCs to endothelial cells (ECs) in vitro. MSCs were isolated from porcine abdominal adipose tissue, and characterized by immunopositivity to CD44, CD90, CD105, and immunonegativity to CD14 and CD45. Plasticity of AMSCs was confirmed by multilineage differentiation. The mRNA transcripts for MMPs and Tissue Inhibitor of Metalloproteinases (TIMPs), and protein expression of EC markers were analyzed. The enzyme activity and protein expression were analyzed by gelatin zymography, enzyme‐linked immunosorbent assay (ELISA), and Western blot. The differentiation of AMSCs to ECs was confirmed by mRNA and protein expressions of the endothelial markers. The mRNA transcripts for MMP‐2 and MMP‐14 were significantly increased during the differentiation of MSCs into ECs. Findings revealed an elevated MMP‐14 and MMP‐2 expression, and MMP2 enzyme activity. Silencing of MMP‐2 and MMP‐14 significantly increased the expression of EC markers, formation of capillary tubes, and acetylated‐low‐density lipoprotein uptake, and decreased the cleavage of vascular endothelial growth factor receptor type 2 (VEGFR2). Inhibition of VEGFR2 significantly decreased the expression of EC markers. These novel findings demonstrate that the upregulation of MMP2 and MMP14 has an inhibitory effect on the differentiation of AMSCs to ECs, and silencing these MMPs inhibit the cleavage of VEGFR2 and stimulate the differentiation of AMSCs to ECs. These findings provide a potential mechanism for the regulatory role of MMP‐2 and MMP‐14 in the re‐endothelialization of coronary arteries following intervention. Stem Cells Translational Medicine 2017;6:1385–1398 PMID:28213979

Postthymic maturation influences the CD8 T cell response to antigen.

PubMed

Makaroff, Lydia E; Hendricks, Deborah W; Niec, Rachel E; Fink, Pamela J

2009-03-24

Complete T cell development requires postthymic maturation, and we investigated the influence of this ontological period on the CD8 T cell response to infection by comparing responses of mature CD8 T cells with those of recent thymic emigrants (RTEs). When activated with a noninflammatory stimulus or a bacterial or viral pathogen, CD8 RTEs generated a lower proportion of cytokine-producing effector cells and long-lived memory precursors compared with their mature counterparts. Although peripheral T cell maturation is complete within several weeks after thymic egress, RTE-derived memory cells continued to express inappropriate levels of memory cell markers and display an altered pattern of cytokine production, even 8 weeks after infection. When rechallenged, RTE-derived memory cells generated secondary effector cells that were phenotypically and functionally equivalent to those generated by their mature counterparts. The defects at the effector and memory stages were not associated with differences in the expression of T cell receptor-, costimulation-, or activation-associated cell surface markers yet were associated with lower Ly6C expression levels at the effector stage. This work demonstrates that the stage of postthymic maturation influences cell fate decisions and cytokine profiles of stimulated CD8 T cells, with repercussions that are apparent long after cells have progressed from the RTE compartment.

Morphologic, phenotypic, and transcriptomic characterization of classically and alternatively activated canine blood-derived macrophages in vitro

PubMed Central

Heinrich, Franziska; Lehmbecker, Annika; Raddatz, Barbara B.; Kegler, Kristel; Tipold, Andrea; Stein, Veronika M.; Kalkuhl, Arno; Deschl, Ulrich; Baumgärtner, Wolfgang; Ulrich, Reiner

2017-01-01

Macrophages are a heterogeneous cell population playing a pivotal role in tissue homeostasis and inflammation, and their phenotype strongly depends on the micromilieu. Despite its increasing importance as a translational animal model for human diseases, there is a considerable gap of knowledge with respect to macrophage polarization in dogs. The present study comprehensively investigated the morphologic, phenotypic, and transcriptomic characteristics of unstimulated (M0), M1- (GM-CSF, LPS, IFNγ-stimulated) and M2- (M-CSF, IL-4-stimulated)-polarized canine blood-derived macrophages in vitro. Scanning electron microscopy revealed distinct morphologies of polarized macrophages with formation of multinucleated cells in M2-macrophages, while immunofluorescence employing literature-based prototype-antibodies against CD16, CD32, iNOS, MHC class II (M1-markers), CD163, CD206, and arginase-1 (M2-markers) demonstrated that only CD206 was able to discriminate M2-macrophages from both other phenotypes, highlighting this molecule as a promising marker for canine M2-macrophages. Global microarray analysis revealed profound changes in the transcriptome of polarized canine macrophages. Functional analysis pointed out that M1-polarization was associated with biological processes such as “respiratory burst”, whereas M2-polarization was associated with processes such as “mitosis”. Literature-based marker gene selection revealed only minor overlaps in the gene sets of the dog compared to prototype markers of murine and human macrophages. Biomarker selection using supervised clustering suggested latexin (LXN) and membrane-spanning 4-domains, subfamily A, member 2 (MS4A2) to be the most powerful predicting biomarkers for canine M1- and M2-macrophages, respectively. Immunofluorescence for both markers demonstrated expression of both proteins by macrophages in vitro but failed to reveal differences between canine M1 and M2-macrophages. The present study provides a solid basis for future studies upon the role of macrophage polarization in spontaneous diseases of the dog, a species that has emerging importance for translational research. PMID:28817687

Early Virological and Immunological Events in Asymptomatic Epstein-Barr Virus Infection in African Children

PubMed Central

Jayasooriya, Shamanthi; de Silva, Thushan I.; Njie-jobe, Jainaba; Sanyang, Chilel; Leese, Alison M.; Bell, Andrew I.; McAulay, Karen A.; Yanchun, Peng; Long, Heather M.; Dong, Tao; Whittle, Hilton C.; Rickinson, Alan B.; Rowland-Jones, Sarah L.; Hislop, Andrew D.; Flanagan, Katie L.

2015-01-01

Epstein-Barr virus (EBV) infection often occurs in early childhood and is asymptomatic. However, if delayed until adolescence, primary infection may manifest as acute infectious mononucleosis (AIM), a febrile illness characterised by global CD8+ T-cell lymphocytosis, much of it reflecting a huge expansion of activated EBV-specific CD8+ T-cells. While the events of AIM have been intensely studied, little is known about how these relate to asymptomatic primary infection. Here Gambian children (14–18 months old, an age at which many acquire the virus) were followed for the ensuing six months, monitoring circulating EBV loads, antibody status against virus capsid antigen (VCA) and both total and virus-specific CD8+ T-cell numbers. Many children were IgG anti-VCA-positive and, though no longer IgM-positive, still retained high virus loads comparable to AIM patients and had detectable EBV-specific T-cells, some still expressing activation markers. Virus loads and the frequency/activation status of specific T-cells decreased over time, consistent with resolution of a relatively recent primary infection. Six children with similarly high EBV loads were IgM anti-VCA-positive, indicating very recent infection. In three of these donors with HLA types allowing MHC-tetramer analysis, highly activated EBV-specific T-cells were detectable in the blood with one individual epitope response reaching 15% of all CD8+ T-cells. That response was culled and the cells lost activation markers over time, just as seen in AIM. However, unlike AIM, these events occurred without marked expansion of total CD8+ numbers. Thus asymptomatic EBV infection in children elicits a virus-specific CD8+ T-cell response that can control the infection without over-expansion; conversely, in AIM it appears the CD8 over-expansion, rather than virus load per se, is the cause of disease symptoms. PMID:25816224

Longitudinal characterization of dysfunctional T cell-activation during human acute Ebola infection

PubMed Central

Agrati, C; Castilletti, C; Casetti, R; Sacchi, A; Falasca, L; Turchi, F; Tumino, N; Bordoni, V; Cimini, E; Viola, D; Lalle, E; Bordi, L; Lanini, S; Martini, F; Nicastri, E; Petrosillo, N; Puro, V; Piacentini, M; Di Caro, A; Kobinger, G P; Zumla, A; Ippolito, G; Capobianchi, M R

2016-01-01

Data on immune responses during human Ebola virus disease (EVD) are scanty, due to limitations imposed by biosafety requirements and logistics. A sustained activation of T-cells was recently described but functional studies during the acute phase of human EVD are still missing. Aim of this work was to evaluate the kinetics and functionality of T-cell subsets, as well as the expression of activation, autophagy, apoptosis and exhaustion markers during the acute phase of EVD until recovery. Two EVD patients admitted to the Italian National Institute for Infectious Diseases, Lazzaro Spallanzani, were sampled sequentially from soon after symptom onset until recovery and analyzed by flow cytometry and ELISpot assay. An early and sustained decrease of CD4 T-cells was seen in both patients, with an inversion of the CD4/CD8 ratio that was reverted during the recovery period. In parallel with the CD4 T-cell depletion, a massive T-cell activation occurred and was associated with autophagic/apoptotic phenotype, enhanced expression of the exhaustion marker PD-1 and impaired IFN-gamma production. The immunological impairment was accompanied by EBV reactivation. The association of an early and sustained dysfunctional T-cell activation in parallel to an overall CD4 T-cell decline may represent a previously unknown critical point of Ebola virus (EBOV)-induced immune subversion. The recent observation of late occurrence of EBOV-associated neurological disease highlights the importance to monitor the immuno-competence recovery at discharge as a tool to evaluate the risk of late sequelae associated with resumption of EBOV replication. Further studies are required to define the molecular mechanisms of EVD-driven activation/exhaustion and depletion of T-cells. PMID:27031961

Longitudinal characterization of dysfunctional T cell-activation during human acute Ebola infection.

PubMed

Agrati, C; Castilletti, C; Casetti, R; Sacchi, A; Falasca, L; Turchi, F; Tumino, N; Bordoni, V; Cimini, E; Viola, D; Lalle, E; Bordi, L; Lanini, S; Martini, F; Nicastri, E; Petrosillo, N; Puro, V; Piacentini, M; Di Caro, A; Kobinger, G P; Zumla, A; Ippolito, G; Capobianchi, M R

2016-03-31

Data on immune responses during human Ebola virus disease (EVD) are scanty, due to limitations imposed by biosafety requirements and logistics. A sustained activation of T-cells was recently described but functional studies during the acute phase of human EVD are still missing. Aim of this work was to evaluate the kinetics and functionality of T-cell subsets, as well as the expression of activation, autophagy, apoptosis and exhaustion markers during the acute phase of EVD until recovery. Two EVD patients admitted to the Italian National Institute for Infectious Diseases, Lazzaro Spallanzani, were sampled sequentially from soon after symptom onset until recovery and analyzed by flow cytometry and ELISpot assay. An early and sustained decrease of CD4 T-cells was seen in both patients, with an inversion of the CD4/CD8 ratio that was reverted during the recovery period. In parallel with the CD4 T-cell depletion, a massive T-cell activation occurred and was associated with autophagic/apoptotic phenotype, enhanced expression of the exhaustion marker PD-1 and impaired IFN-gamma production. The immunological impairment was accompanied by EBV reactivation. The association of an early and sustained dysfunctional T-cell activation in parallel to an overall CD4 T-cell decline may represent a previously unknown critical point of Ebola virus (EBOV)-induced immune subversion. The recent observation of late occurrence of EBOV-associated neurological disease highlights the importance to monitor the immuno-competence recovery at discharge as a tool to evaluate the risk of late sequelae associated with resumption of EBOV replication. Further studies are required to define the molecular mechanisms of EVD-driven activation/exhaustion and depletion of T-cells.

CD6 as a potential target for treating multiple sclerosis

PubMed Central

Singer, Nora G.; Whitbred, Joy; Bowen, Michael A.; Lin, Feng

2017-01-01

CD6 was established as a marker of T cells more than three decades ago, and recent studies have identified CD6 as a risk gene for multiple sclerosis (MS), a disease in which autoreactive T cells are integrally involved. Nevertheless, the precise role of CD6 in regulating T-cell responses is controversial and its significance in the pathogenesis of various diseases remains elusive, partly due to the lack of animals engineered to alter expression of the CD6 gene. In this report, we found that CD6 KO mice showed decreased pathogenic T-cell responses, reduced spinal cord T-cell infiltration, and attenuated disease severity in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. CD6-deficient T cells exhibited augmented activation, but also significantly reduced survival and proliferation after activation, leading to overall decreased Th1 and Th17 polarization. Activated CD6-deficient T cells also showed impaired infiltration through brain microvascular endothelial cell monolayers. Furthermore, by developing CD6 humanized mice, we identified a mouse anti-human CD6 monoclonal antibody that is highly effective in treating established EAE without depleting T cells. These results suggest that (i) CD6 is a negative regulator of T-cell activation, (ii) at the same time, CD6 is a positive regulator of activated T-cell survival/proliferation and infiltration; and (iii) CD6 is a potential new target for treating MS and potentially other T-cell–driven autoimmune conditions. PMID:28209777

Intestinal double-positive CD4+CD8+ T cells are highly activated memory cells with an increased capacity to produce cytokines.

PubMed

Pahar, Bapi; Lackner, Andrew A; Veazey, Ronald S

2006-03-01

Peripheral blood and intestinal CD4+CD8+ double-positive (DP) T cells have been described in several species including humans, but their function and immunophenotypic characteristics are still not clearly understood. Here we demonstrate that DP T cells are abundant in the intestinal lamina propria of normal rhesus macaques (Macaca mulatta). Moreover, DP T cells have a memory phenotype and are capable of producing different and/or higher levels of cytokines and chemokines in response to mitogen stimulation compared to CD4+ single-positive T cells. Intestinal DP T cells are also highly activated and have higher expression of CCR5, which makes them preferred targets for simian immunodeficiency virus/HIV infection. Increased levels of CD69, CD25 and HLA-DR, and lower CD62L expression were found on intestinal DP T cells populations compared to CD4+ single-positive T cells. Collectively, these findings demonstrate that intestinal and peripheral blood DP T cells are effector cells and may be important in regulating immune responses, which distinguishes them from the immature DP cells found in the thymus. Finally, these intestinal DP T cells may be important target cells for HIV infection and replication due to their activation, memory phenotype and high expression of CCR5.

CD30 expression in follicular lymphoma.

PubMed

Gardner, L J; Polski, J M; Evans, H L; Perkins, S L; Dunphy, C H

2001-08-01

CD30(+) anaplastic large cell lymphomas were originally described as being of T-cell, null cell, and B-cell origin. CD30, however, is not a specific marker of anaplastic large cell lymphoma and has been found to be expressed in reactive as well as neoplastic populations as a probable activation marker. In addition, CD30(+) cells have also been described in both diffuse large B-cell and follicular lymphomas (FLs), resembling the pattern seen in reactive tonsils and lymph nodes. We report an index case of FL with CD30 expression, which on initial touch preparations and flow cytometric immunophenotyping revealed a prominent population of CD30(+) cells with marked cellular pleomorphism (anaplasia) in a background of typical FL. Immunohistochemistry of the paraffin section for CD30 in our index case confirmed unequivocal CD30(+) pleomorphic cells in the malignant nodules in occasional clusters. This case prompted a study of additional cases of FL for pattern of immunoreactivity with CD30 on paraffin sections. Twenty-two additional cases of FL (grades 1-3) were retrieved for CD30 immunoperoxidase staining as in the index case. This study demonstrated 32% of the additional cases of FL had definitive CD30(+), large, pleomorphic malignant cells by paraffin immunohistochemistry. In 2 cases (9%), the pattern of immunoreactivity with CD30 showed clustering and variable staining of large cells, as our index case. This study underscores the morphologic and immunophenotypic spectrum of FL that includes CD30 staining and cellular pleomorphism.

DDX4 (DEAD box polypeptide 4) colocalizes with cancer stem cell marker CD133 in ovarian cancers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Ki Hyung; Biomedical Research Institute and Pusan Cancer Center, Pusan National University Hospital, Busan; Kang, Yun-Jeong

Highlights: • Germ cell marker DDX4 was significantly increased in ovarian cancer. • Ovarian cancer stem cell marker CD133 was significantly increased in ovarian cancer. • DDX4 and CD133 were mostly colocalized in various types of ovarian cancer tissues. • CD133 positive ovarian cancer cells also express DDX4 whereas CD133-negative cells did not possess DDX4. • Germ cell marker DDX4 has the potential of ovarian cancer stem cell marker. - Abstract: DDX4 (DEAD box polypeptide 4), characterized by the conserved motif Asp-Glu-Ala-Asp (DEAD), is an RNA helicase which is implicated in various cellular processes involving the alteration of RNA secondarymore » structure, such as translation initiation, nuclear and mitochondrial splicing, and ribosome and spliceosome assembly. DDX4 is known to be a germ cell-specific protein and is used as a sorting marker of germline stem cells for the production of oocytes. A recent report about DDX4 in ovarian cancer showed that DDX4 is overexpressed in epithelial ovarian cancer and disrupts a DNA damage-induced G2 checkpoint. We investigated the relationship between DDX4 and ovarian cancer stem cells by analyzing the expression patterns of DDX4 and the cancer stem cell marker CD133 in ovarian cancers via tissue microarray. Both DDX4 and CD133 were significantly increased in ovarian cancer compared to benign tumors, and showed similar patterns of expression. In addition, DDX4 and CD133 were mostly colocalized in various types of ovarian cancer tissues. Furthermore, almost all CD133 positive ovarian cancer cells also express DDX4 whereas CD133-negative cells did not possess DDX4, suggesting a strong possibility that DDX4 plays an important role in cancer stem cells, and/or can be used as an ovarian cancer stem cell marker.« less

The Fungal Quorum-Sensing Molecule Farnesol Activates Innate Immune Cells but Suppresses Cellular Adaptive Immunity

PubMed Central

Leonhardt, Ines; Spielberg, Steffi; Weber, Michael; Albrecht-Eckardt, Daniela; Bläss, Markus; Claus, Ralf; Barz, Dagmar; Scherlach, Kirstin; Hertweck, Christian; Löffler, Jürgen; Hünniger, Kerstin

2015-01-01

ABSTRACT Farnesol, produced by the polymorphic fungus Candida albicans, is the first quorum-sensing molecule discovered in eukaryotes. Its main function is control of C. albicans filamentation, a process closely linked to pathogenesis. In this study, we analyzed the effects of farnesol on innate immune cells known to be important for fungal clearance and protective immunity. Farnesol enhanced the expression of activation markers on monocytes (CD86 and HLA-DR) and neutrophils (CD66b and CD11b) and promoted oxidative burst and the release of proinflammatory cytokines (tumor necrosis factor alpha [TNF-α] and macrophage inflammatory protein 1 alpha [MIP-1α]). However, this activation did not result in enhanced fungal uptake or killing. Furthermore, the differentiation of monocytes to immature dendritic cells (iDC) was significantly affected by farnesol. Several markers important for maturation and antigen presentation like CD1a, CD83, CD86, and CD80 were significantly reduced in the presence of farnesol. Furthermore, farnesol modulated migrational behavior and cytokine release and impaired the ability of DC to induce T cell proliferation. Of major importance was the absence of interleukin 12 (IL-12) induction in iDC generated in the presence of farnesol. Transcriptome analyses revealed a farnesol-induced shift in effector molecule expression and a down-regulation of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor during monocytes to iDC differentiation. Taken together, our data unveil the ability of farnesol to act as a virulence factor of C. albicans by influencing innate immune cells to promote inflammation and mitigating the Th1 response, which is essential for fungal clearance. PMID:25784697

Immunological abnormalities as potential biomarkers in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis.

PubMed

Brenu, Ekua W; van Driel, Mieke L; Staines, Don R; Ashton, Kevin J; Ramos, Sandra B; Keane, James; Klimas, Nancy G; Marshall-Gradisnik, Sonya M

2011-05-28

Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is characterised by severe prolonged fatigue, and decreases in cognition and other physiological functions, resulting in severe loss of quality of life, difficult clinical management and high costs to the health care system. To date there is no proven pathomechanism to satisfactorily explain this disorder. Studies have identified abnormalities in immune function but these data are inconsistent. We investigated the profile of markers of immune function (including novel markers) in CFS/ME patients. We included 95 CFS/ME patients and 50 healthy controls. All participants were assessed on natural killer (NK) and CD8(+) T cell cytotoxic activities, Th1 and Th2 cytokine profile of CD4(+) T cells, expression of vasoactive intestinal peptide receptor 2 (VPACR2), levels of NK phenotypes (CD56(bright) and CD56(dim)) and regulatory T cells expressing FoxP3 transcription factor. Compared to healthy individuals, CFS/ME patients displayed significant increases in IL-10, IFN-γ, TNF-α, CD4(+)CD25(+) T cells, FoxP3 and VPACR2 expression. Cytotoxic activity of NK and CD8(+) T cells and NK phenotypes, in particular the CD56(bright) NK cells were significantly decreased in CFS/ME patients. Additionally granzyme A and granzyme K expression were reduced while expression levels of perforin were significantly increased in the CFS/ME population relative to the control population. These data suggest significant dysregulation of the immune system in CFS/ME patients. Our study found immunological abnormalities which may serve as biomarkers in CFS/ME patients with potential for an application as a diagnostic tool.

Investigating Cell Surface Markers on Normal Hematopoietic Stem Cells in Three Different Niche Conditions

PubMed Central

Garg, Swati; Madkaikar, Manisha

2013-01-01

Hematopoietic stem cells are of therapeutic interest to the clinicians and researchers due to their promising assistance in management of malignant and inherited hematological conditions. Evaluation of cell surface markers using multiparametric flow cytometry is a well adapted qualitative measure of cells in question for many years. An artillery of these markers has been studied in hematological malignancies and related disorders. However, their role and differential expression on normal hematopoietic stem cells from clinically available sources is not always described carefully. In the present study, we attempted to evaluate expression of CD44, CD90, CD96 and CD123 in three clinically available sources of normal HSCs (Hematopoietic stem cells). Sources of HSCs in the present study involved umbilical cord blood (UCB), normal bone marrow (NBM) and bone marrow from idiopathic thrombocytopenic purpura (ITP) patients (IBM). CD44 is an important homing receptor while CD90 is involved in maintaining stem cell quiescent. CD96 is known to be leukemia specific marker and CD123 is involved in stem cell differentiation and survival. We observed a significant difference in expression CD44, CD90 and CD123 on normal HSCs derived from umbilical cord and ITP marrow. CD96 was highly expressed on HSCs obtained from ITP marrow. Investigating expression of these markers on normal HSCs in different niches will be helpful in correlating their function with niche condition and delineating their ‘abnormal’ expression from the normal. PMID:24386557

Investigating cell surface markers on normal hematopoietic stem cells in three different niche conditions.

PubMed

Garg, Swati; Madkaikar, Manisha; Ghosh, Kanjaksha

2013-11-01

Hematopoietic stem cells are of therapeutic interest to the clinicians and researchers due to their promising assistance in management of malignant and inherited hematological conditions. Evaluation of cell surface markers using multiparametric flow cytometry is a well adapted qualitative measure of cells in question for many years. An artillery of these markers has been studied in hematological malignancies and related disorders. However, their role and differential expression on normal hematopoietic stem cells from clinically available sources is not always described carefully. In the present study, we attempted to evaluate expression of CD44, CD90, CD96 and CD123 in three clinically available sources of normal HSCs (Hematopoietic stem cells). Sources of HSCs in the present study involved umbilical cord blood (UCB), normal bone marrow (NBM) and bone marrow from idiopathic thrombocytopenic purpura (ITP) patients (IBM). CD44 is an important homing receptor while CD90 is involved in maintaining stem cell quiescent. CD96 is known to be leukemia specific marker and CD123 is involved in stem cell differentiation and survival. We observed a significant difference in expression CD44, CD90 and CD123 on normal HSCs derived from umbilical cord and ITP marrow. CD96 was highly expressed on HSCs obtained from ITP marrow. Investigating expression of these markers on normal HSCs in different niches will be helpful in correlating their function with niche condition and delineating their 'abnormal' expression from the normal.

Phenotypic profile of expanded NK cells in chronic lymphoproliferative disorders: a surrogate marker for NK-cell clonality

PubMed Central

Bárcena, Paloma; Jara-Acevedo, María; Tabernero, María Dolores; López, Antonio; Sánchez, María Luz; García-Montero, Andrés C.; Muñoz-García, Noemí; Vidriales, María Belén; Paiva, Artur; Lecrevisse, Quentin; Lima, Margarida; Langerak, Anton W.; Böttcher, Sebastian; van Dongen, Jacques J.M.

2015-01-01

Currently, the lack of a universal and specific marker of clonality hampers the diagnosis and classification of chronic expansions of natural killer (NK) cells. Here we investigated the utility of flow cytometric detection of aberrant/altered NK-cell phenotypes as a surrogate marker for clonality, in the diagnostic work-up of chronic lymphoproliferative disorders of NK cells (CLPD-NK). For this purpose, a large panel of markers was evaluated by multiparametric flow cytometry on peripheral blood (PB) CD56low NK cells from 60 patients, including 23 subjects with predefined clonal (n = 9) and polyclonal (n = 14) CD56low NK-cell expansions, and 37 with CLPD-NK of undetermined clonality; also, PB samples from 10 healthy adults were included. Clonality was established using the human androgen receptor (HUMARA) assay. Clonal NK cells were found to show decreased expression of CD7, CD11b and CD38, and higher CD2, CD94 and HLADR levels vs. normal NK cells, together with a restricted repertoire of expression of the CD158a, CD158b and CD161 killer-associated receptors. In turn, NK cells from both clonal and polyclonal CLPD-NK showed similar/overlapping phenotypic profiles, except for high and more homogeneous expression of CD94 and HLADR, which was restricted to clonal CLPD-NK. We conclude that the CD94hi/HLADR+ phenotypic profile proved to be a useful surrogate marker for NK-cell clonality. PMID:26556869

Immune Consequences of Decreasing Tumor Vasculature with Antiangiogenic Tyrosine Kinase Inhibitors in Combination with Therapeutic Vaccines

PubMed Central

Farsaci, Benedetto; Donahue, Renee N.; Coplin, Michael A.; Grenga, Italia; Lepone, Lauren M.; Molinolo, Alfredo A.; Hodge, James W.

2014-01-01

This study investigated the effects on the tumor microenvironment of combining antiangiogenic tyrosine kinase inhibitors (TKI) with therapeutic vaccines, and in particular, how vascular changes affect tumor-infiltrating immune cells. We conducted studies using a TKI (sunitinib or sorafenib) in combination with recombinant vaccines in 2 murine tumor models: colon carcinoma (MC38-CEA) and breast cancer (4T1). Tumor vasculature was measured by immunohistochemistry using 3 endothelial cell markers: CD31 (mature), CD105 (immature/proliferating), and CD11b (monocytic). We assessed oxygenation, tight junctions, compactness, and pressure within tumors, along with the frequency and phenotype of tumor-infiltrating T lymphocytes (TIL), myeloid-derived suppressor cells (MDSC), and tumor-associated macrophages (TAM) following treatment with antiangiogenic TKIs alone, vaccine alone, or the combination of a TKI with vaccine. The combined regimen decreased tumor vasculature, compactness, tight junctions, and pressure, leading to vascular normalization and increased tumor oxygenation. This combination therapy also increased TILs, including tumor antigen-specific CD8 T cells, and elevated the expression of activation markers FAS-L, CXCL-9, CD31, and CD105 in MDSCs and TAMs, leading to reduced tumor volumes and an increase in the number of tumor-free animals. The improved antitumor activity induced by combining antiangiogenic TKIs with vaccine may be the result of activated lymphoid and myeloid cells in the tumor microenvironment, resulting from vascular normalization, decreased tumor-cell density, and the consequent improvement in vascular perfusion and oxygenation. Therapies that alter tumor architecture can thus have a dramatic impact on the effectiveness of cancer immunotherapy. PMID:25092771

Biomarkers for evaluation of mast cell and basophil activation.

PubMed

Kabashima, Kenji; Nakashima, Chisa; Nonomura, Yumi; Otsuka, Atsushi; Cardamone, Chiara; Parente, Roberta; De Feo, Giulia; Triggiani, Massimo

2018-03-01

Mast cells and basophils play a pathogenetic role in allergic, inflammatory, and autoimmune disorders. These cells have different development, anatomical location and life span but share many similarities in mechanisms of activation and type of mediators. Mediators secreted by mast cells and basophils correlate with clinical severity in asthma, chronic urticaria, anaphylaxis, and other diseases. Therefore, effective biomarkers to measure mast cell and basophil activation in vivo could potentially have high diagnostic and prognostic values. An ideal biomarker should be specific for mast cells or basophils, easily and reproducibly detectable in blood or biological fluids and should be metabolically stable. Markers of mast cell and basophil include molecules secreted by stimulated cells and surface molecules expressed upon activation. Some markers, such as histamine and lipid mediators are common to mast cells and basophils whereas others, such as tryptase and other proteases, are relatively specific for mast cells. The best surface markers of activation expressed on mast cells and basophils are CD63 and CD203. While these mediators and surface molecules have been associated to a variety of diseases, none of them fulfills requirements for an optimal biomarker and search for better indicators of mast cell/basophil activation in vivo is ongoing. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Expression of CD163 in the liver of patients with viral hepatitis.

PubMed

Hiraoka, Atsushi; Horiike, Norio; Akbar, Sk Md Fazle; Michitaka, Kojiro; Matsuyama, Takami; Onji, Morikazu

2005-01-01

CD163 is a marker of activated macrophages, and increased levels of soluble CD163 have been detected in sera obtained from patients with hepatitis. The aim of this study was to detect the expression of CD163 in the liver from patients with viral hepatitis. Frozen sections of liver specimens were obtained from 5 patients with acute viral hepatitis (AH) and from 23 patients with chronic viral hepatitis (CH). The expression of CD163 in the liver was determined immunohistochemically using monoclonal antibody to human CD163. Double immunostaining was done to assess those cell types that express CD163 in the liver. The frequencies of CD163-positive cells were significantly higher both in the portal areas and in the hepatic lobules in the liver of patients with AH compared to those with CH (p < 0.05). Double immunostaining revealed that most of the CD163-positive cells were macrophages and Kupffer cells, because they expressed CD68. The expression of CD163 was very low in endothelial cells and liver stellate cells. This study shows that macrophages are activated in hepatitis liver.

Ligation of CD8α on human natural killer cells prevents activation-induced apoptosis and enhances cytolytic activity

PubMed Central

Addison, Elena G; North, Janet; Bakhsh, Ismail; Marden, Chloe; Haq, Sumaira; Al-Sarraj, Samia; Malayeri, Reza; Wickremasinghe, R Gitendra; Davies, Jeffrey K; Lowdell, Mark W

2005-01-01

It has been previously shown that the subset of human natural killer (NK) cells which express CD8 in a homodimeric α/α form are more cytotoxic than their CD8– counterparts but the mechanisms behind this differential cytolytic activity remained unknown. Target cell lysis by CD8– NK cells is associated with high levels of effector cell apoptosis, which is in contrast to the significantly lower levels found in the CD8α+ cells after lysis of the same targets. We report that cross-linking of the CD8α chains on NK cells induces rapid rises in intracellular Ca2+ and increased expression of CD69 at the cell surface by initiating the influx of extracellular Ca2+ ions. We demonstrate that secretion of cytolytic enzymes initiates NK-cell apoptosis from which CD8α+ NK cells are protected by an influx of exogenous calcium following ligation of CD8 on the NK-cell surface. This ligation is through interaction with fellow NK cells in the cell conjugate and can occur when the target cells lack major histocompatibility complex (MHC) Class I expression. Protection from apoptosis is blocked by preincubation of the NK cells with anti-MHC Class I antibody. Thus, in contrast to the CD8– subset, CD8α+ NK cells are capable of sequential lysis of multiple target cells. PMID:16236125

Respiratory Syncytial Virus (RSV) Infects CD4+ T Cells: Frequency of Circulating CD4+ RSV+ T Cells as a Marker of Disease Severity in Young Children.

PubMed

Raiden, Silvina; Sananez, Inés; Remes-Lenicov, Federico; Pandolfi, Julieta; Romero, Cecilia; De Lillo, Leonardo; Ceballos, Ana; Geffner, Jorge; Arruvito, Lourdes

2017-04-01

Although human airway epithelial cells are the main target of respiratory syncytial virus (RSV), it also infects immune cells, such as macrophages and B cells. Whether T cells are permissive to RSV infection is unknown. We sought to analyze the permissiveness of CD4+ T cells to RSV infection. CD4+ and CD8+ T cells from cord blood, healthy young children, and adults were challenged by RSV or cocultured with infected HEp-2 cells. Infection, phenotype, and cytokine production by T cells were analyzed by flow cytometry or enzyme-linked immunosorbent assay. Expression of RSV antigens by circulating CD4+ T cells from infected children was analyzed by flow cytometry, and disease severity was defined by standard criteria. CD4+ and CD8+ T cells were productively infected by RSV. Infection decreased interleukin 2 and interferon γ production as well as the expression of CD25 and Ki-67 by activated CD4+ T cells. Respiratory syncytial virus antigens were detected in circulating CD4+ and CD8+ T cells during severe RSV infection of young children. Interestingly, the frequency of CD4+ RSV+ T cells positively correlated with disease severity. Respiratory syncytial virus infects CD4+ and CD8+ T cells and compromises T-cell function. The frequency of circulating CD4+ RSV+ T cells might represent a novel marker of severe infection. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Evidence for Anti-Inflammatory Effects of Exercise in CKD

PubMed Central

Kosmadakis, George C.; Watson, Emma L.; Bevington, Alan; Feehally, John; Bishop, Nicolette C.; Smith, Alice C.

2014-01-01

CKD is associated with a complex state of immune dysfunction characterized by immune depression, predisposing patients to infections, and immune activation, resulting in inflammation that associates with higher risk of cardiovascular disease. Physical exercise may enhance immune function and exert anti-inflammatory effects, but such effects are unclear in CKD. We investigated the separate effects of acute and regular moderate-intensity aerobic exercise on neutrophil degranulation (elastase release), activation of T lymphocytes (CD69 expression) and monocytes (CD86 and HLA-DR expression), and plasma inflammatory markers (IL-6, IL-10, soluble TNF-receptors, and C-reactive protein) in patients with predialysis CKD. A single 30-minute (acute) bout of walking induced a normal pattern of leukocyte mobilization and had no effect on T-lymphocyte and monocyte activation but improved neutrophil responsiveness to a bacterial challenge in the postexercise period. Furthermore, acute exercise induced a systemic anti-inflammatory environment, evidenced by a marked increase in plasma IL-10 levels (peaked at 1 hour postexercise), that was most likely mediated by increased plasma IL-6 levels (peaked immediately postexercise). Six months of regular walking exercise (30 min/d for 5 times/wk) exerted anti-inflammatory effects (reduction in the ratio of plasma IL-6 to IL-10 levels) and a downregulation of T-lymphocyte and monocyte activation, but it had no effect on circulating immune cell numbers or neutrophil degranulation responses. Renal function, proteinuria, and BP were also unaffected. These findings provide compelling evidence that walking exercise is safe with regard to immune and inflammatory responses and has the potential to be an effective anti-inflammatory therapy in predialysis CKD. PMID:24700875

Metformin Inhibits Advanced Glycation End Products-Induced Inflammatory Response in Murine Macrophages Partly through AMPK Activation and RAGE/NFκB Pathway Suppression

PubMed Central

Zhou, Zhong'e; Tang, Yong; Chen, Chengjun; Lu, Yi; Liu, Liang

2016-01-01

Advanced glycation end products (AGEs) are major inflammatory mediators in diabetes, affecting atherosclerosis progression via macrophages. Metformin slows diabetic atherosclerosis progression through mechanisms that remain to be fully elucidated. The present study of murine bone marrow derived macrophages showed that (1) AGEs enhanced proinflammatory cytokines (interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α)) mRNA expression, RAGE expression, and NFκB activation; (2) metformin pretreatment inhibited AGEs effects and AGEs-induced cluster designation 86 (CD86) (M1 marker) expression, while promoting CD206 (M2 marker) surface expression and anti-inflammatory cytokine (IL-10) mRNA expression; and (3) the AMPK inhibitor, Compound C, attenuated metformin effects. In conclusion, metformin inhibits AGEs-induced inflammatory response in murine macrophages partly through AMPK activation and RAGE/NFκB pathway suppression. PMID:27761470

NK Cells with KIR2DS2 Immunogenotype Have a Functional Activation Advantage To Efficiently Kill Glioblastoma and Prolong Animal Survival

PubMed Central

Gras Navarro, Andrea; Kmiecik, Justyna; Leiss, Lina; Zelkowski, Mateusz; Engelsen, Agnete; Bruserud, Øystein; Zimmer, Jacques; Enger, Per Øyvind

2014-01-01

Glioblastomas (GBMs) are lethal brain cancers that are resistant to current therapies. We investigated the cytotoxicity of human allogeneic NK cells against patient-derived GBM in vitro and in vivo, as well as mechanisms mediating their efficacy. We demonstrate that KIR2DS2 immunogenotype NK cells were more potent killers, notwithstanding the absence of inhibitory killer Ig–like receptor (KIR)-HLA ligand mismatch. FACS-sorted and enriched KIR2DS2+ NK cell subpopulations retained significantly high levels of CD69 and CD16 when in contact with GBM cells at a 1:1 ratio and highly expressed CD107a and secreted more soluble CD137 and granzyme A. In contrast, KIR2DS2− immunogenotype donor NK cells were less cytotoxic against GBM and K562, and, similar to FACS-sorted or gated KIR2DS2− NK cells, significantly diminished CD16, CD107a, granzyme A, and CD69 when in contact with GBM cells. Furthermore, NK cell–mediated GBM killing in vitro depended upon the expression of ligands for the activating receptor NKG2D and was partially abrogated by Ab blockade. Treatment of GBM xenografts in NOD/SCID mice with NK cells from a KIR2DS2+ donor lacking inhibitory KIR-HLA ligand mismatch significantly prolonged the median survival to 163 d compared with vehicle controls (log-rank test, p = 0.0001), in contrast to 117.5 d (log-rank test, p = 0.0005) for NK cells with several inhibitory KIR-HLA ligand mismatches but lacking KIR2DS2 genotype. Significantly more CD56+CD16+ NK cells from a KIR2DS2+ donor survived in nontumor-bearing brains 3 wk after infusion compared with KIR2DS2− NK cells, independent of their proliferative capacity. In conclusion, KIR2DS2 identifies potent alloreactive NK cells against GBM that are mediated by commensurate, but dominant, activating signals. PMID:25381437

Soluble CD163 is increased in patients with acute pancreatitis independent of disease severity.

PubMed

Karrasch, Thomas; Brünnler, Tanja; Hamer, Okka W; Schmid, Karin; Voelk, Markus; Herfarth, Hans; Buechler, Christa

2015-10-01

Macrophages are crucially involved in the pathophysiology of acute pancreatitis. Soluble CD163 (sCD163) is specifically released from macrophages and systemic levels are increased in inflammatory diseases. Here, sCD163 was measured in serum of 50 patients with acute pancreatitis to find out possible associations with disease activity. Admission levels of systemic sCD163 were nearly three-fold higher in patients with acute pancreatitis compared to controls. In patients sCD163 did not correlate with C-reactive protein and leukocyte count as established markers of inflammation. Levels were not associated with disease severity assessed by the Schroeder score, Balthazar score, Acute Physiology, Age, and Chronic Health Evaluation (Apache) II score and peripancreatic necrosis score. Soluble CD163 was not related to complications of acute pancreatitis. These data show that serum sCD163 is increased in acute pancreatitis indicating activation of macrophages but is not associated with disease severity and outcome. Copyright © 2015 Elsevier Inc. All rights reserved.

Integrin αIIb (CD41) plays a role in the maintenance of hematopoietic stem cell activity in the mouse embryonic aorta

PubMed Central

Boisset, Jean-Charles; Clapes, Thomas; Van Der Linden, Reinier; Dzierzak, Elaine; Robin, Catherine

2013-01-01

Summary Integrins are transmembrane receptors that play important roles as modulators of cell behaviour through their adhesion properties and the initiation of signaling cascades. The αIIb integrin subunit (CD41) is one of the first cell surface markers indicative of hematopoietic commitment. αIIb pairs exclusively with β3 to form the αIIbβ3 integrin. β3 (CD61) also pairs with αv (CD51) to form the αvβ3 integrin. The expression and putative role of these integrins during mouse hematopoietic development is as yet unknown. We show here that hematopoietic stem cells (HSCs) differentially express αIIbβ3 and αvβ3 integrins throughout development. Whereas the first HSCs generated in the aorta at mid-gestation express both integrins, HSCs from the placenta only express αvβ3, and most fetal liver HSCs do not express either integrin. By using αIIb deficient embryos, we show that αIIb is not only a reliable HSC marker but it also plays an important and specific function in maintaining the HSC activity in the mouse embryonic aorta. PMID:23789102

Characterization of hepatic progenitors from human fetal liver during second trimester.

PubMed

Rao, Mekala-Subba; Khan, Aleem-Ahmed; Parveen, Nyamath; Habeeb, Mohammed-Aejaz; Habibullah, Chittoor-Mohammed; Pande, Gopal

2008-10-07

To enrich hepatic progenitors using epithelial cell adhesion molecule (EpCAM) as a marker from human fetal liver and investigate the expression of human leukocyte antigen (HLA) and their markers associated with hepatic progenitor cells. EpCAM +ve cells were isolated using magnetic cell sorting (MACS) from human fetuses (n = 10) at 15-25 wk gestation. Expression of markers for hepatic progenitors such as albumin, alpha-fetoprotein (AFP), CD29 (integrin beta1), CD49f (integrin alpha6) and CD90 (Thy 1) was studied by using flow cytometry, immunocytochemistry and RT-PCR; HLA class I (A, B, C) and class II (DR) expression was studied by flow cytometry only. FACS analysis indicated that EpCAM +ve cells were positive for CD29, CD49f, CD90, CD34, HLA class I, albumin and AFP but negative for HLA class II (DR) and CD45. RT PCR showed that EpCAM +ve cells expressed liver epithelial markers (CK18), biliary specific marker (CK19) and hepatic markers (albumin, AFP). On immunocytochemical staining, EpCAM +ve cells were shown positive signals for CK18 and albumin. Our study suggests that these EpCAM +ve cells can be used as hepatic progenitors for cell transplantation with a minimum risk of alloreactivity and these cells may serve as a potential source for enrichment of hepatic progenitor.

Immunologic reconstitution during PEG-ADA therapy in an unusual mosaic ADA-deficient patient

PubMed Central

Liu, Ping; Santisteban, Ines; Burroughs, Laurie M.; Ochs, Hans D.; Torgerson, Troy R.; Hershfield, Michael S.; Rawlings, David J.; Scharenberg, Andrew M.

2009-01-01

We report detailed genetic and immunologic studies in a patient diagnosed with adenosine deaminase (ADA) deficiency and combined immune deficiency at age 5 years. At the time of diagnosis, although all other lymphocyte subsets were depleted, circulating CD8+ T cells with a terminally differentiated phenotype were abundant and expressed normal ADA activity due to a reversion mutation in a CD8+ T cell or precursor. Over the first 9 months of replacement therapy with PEG-ADA, the patient steadily accumulated mature naïve CD4+ and CD8+ T cells, as well as CD4+/FOXP3+ regulatory T cells, consistent with restoration of a functional cellular immune system. While CD19+ naïve B cells also accumulated in response to PEG-ADA therapy, a high proportion of these B cells exhibited an immature surface marker phenotype even after 9 months, and immunization with neoantigen bacteriophage φX174 demonstrated a markedly subnormal humoral immune response. Our observations in this single patient have important implications for gene therapy of human ADA deficiency, as they indicate that ADA expression within even a large circulating lymphocyte population may not be sufficient to support adequate immune reconstitution. They also suggest that an immature surface marker phenotype of the peripheral B cell compartment may be a useful surrogate marker for incomplete humoral immune reconstitution during enzyme replacement, and possibly other forms of hematopoietic cell therapies. PMID:18952502

NP001 regulation of macrophage activation markers in ALS: A phase I clinical and biomarker study

PubMed Central

MILLER, ROBERT G.; ZHANG, RONGZHEN; BLOCK, GILBERT; KATZ, JONATHAN; BAROHN, RICHARD; KASARSKIS, EDWARD; FORSHEW, DALLAS; GOPALAKRISHNAN, VIDHYA; MCGRATH, MICHAEL S.

2017-01-01

This is a phase I, placebo-controlled, single ascending dose safety and tolerability study of NP001 in patients with ALS. NP001 is a novel regulator of inflammatory macrophages and monocytes. As ALS progression is thought to be related to neuroinflammation, an additional objective of the study was to assess the effects of NP001 administration on monocyte activation markers. Thirty-two ALS patients were enrolled and received either placebo (eight) or one of four (six at each dose) ascending single i.v. doses (0.2, 0.8, 1.6 and 3.2 mg/kg NP001). Patients were monitored for safety, and blood monocyte immune activation markers CD16 and HLA-DR were assessed pre- and 24 h post-dosing. Changes from baseline were calculated. Results showed that NP001 was generally safe and well tolerated. Importantly, a single dose of NP001 caused a dose-dependent reduction in expression of monocyte CD16, a marker of monocyte activation/inflammation. Additionally, monocyte HLA-DR expression was also decreased in those patients with elevated values at baseline. In conclusion, these data indicate that NP001 has an acute effect on inflammatory monocytes in ALS patient blood. The potential for modulation of inflammation in the context of ALS disease progression will require further study with long-term follow-up. PMID:25192333

Echinococcus multilocularis vesicular fluid inhibits activation and proliferation of natural killer cells.

PubMed

Bellanger, Anne-Pauline; Mougey, Valentine; Pallandre, Jean-Rene; Gbaguidi-Haore, Houssein; Godet, Yann; Millon, Laurence

2017-08-25

Alveolar echinococcosis is a severe chronic helminthic disease that mimics slow-growing liver cancer. The immune evasion strategy of Echinococcus multilocularis Leuckart, 1863 remains poorly understood. The aim of this study was to investigate in vitro the impact of E. multilocularis vesicular fluid (Em-VF) on peripheral blood mononuclear cells (PBMC) and on natural killer (NK) cells. PBMC and NK cells were exposed to Em-VF (1 µg/ml) during six days. The effect of Em-VF was assessed on CD69, viability and proliferation, and on and transforming growth factor β (TGF-β), interferon γ (IFN-γ), interleukin 17 (IL-17) and interleukin 10, using flow cytometry and ELISA, respectively. Exposure to Em-VF had no bearing on PBMC's viability, proliferation and expression of CD69. In contrast, higher levels of IL-17 at day three and of TGF-β at day six were observed in PBMC supernatant after exposure to Em-VF (p < 0.05, Wilcoxon signed-rank test). Exposure to Em-VF induced a significant decrease of CD69 expression of NK cells at day three and a significant decrease of proliferation of NK cells at day six (p < 0.05, Wilcoxon signed-rank test). In contrast, NK cells viability and levels of cytokines did not vary significantly over Em-VF stimulation. Exposure to Em-VF had a significant bearing on activation and proliferation of NK cells. NK cells may play an important role in the immune response of the host against E. multilocularis.

Characterization of CD8+ T-cell response in acute and resolved hepatitis A virus infection.

PubMed

Schulte, I; Hitziger, T; Giugliano, S; Timm, J; Gold, H; Heinemann, F M; Khudyakov, Y; Strasser, M; König, C; Castermans, E; Mok, J Y; van Esch, W J E; Bertoletti, A; Schumacher, T N; Roggendorf, M

2011-02-01

In contrast to the infection with other hepatotropic viruses, hepatitis A virus (HAV) always causes acute self-limited hepatitis, although the role for virus-specific CD8 T cells in viral containment is unclear. Herein, we analyzed the T cell response in patients with acute hepatitis by utilizing a set of overlapping peptides and predicted HLA-A2 binders from the polyprotein. A set of 11 predicted peptides from the HAV polyprotein, identified as potential binders, were synthesized. Peripheral blood mononuclear cells (PBMCs) from patients were tested for IFNγ secretion after stimulation with these peptides and ex vivo with HLA-A2 tetramers. Phenotyping was carried out by staining with the activation marker CD38 and the memory marker CD127. Eight out of 11 predicted HLA-A2 binders showed a high binding affinity and five of them were recognized by CD8+ T cells from patients with hepatitis A. There were significant differences in the magnitude of the responses to these five peptides. One was reproducibly immunodominant and the only one detectable ex vivo by tetramer staining of CD8+ T cells. These cells have an activated phenotype (CD38hi CD127lo) during acute infection. Three additional epitopes were identified in HLA-A2 negative patients, most likely representing epitopes restricted by other HLA-class I-alleles (HLA-A11, B35, B40). Patients with acute hepatitis A have a strong multi-specific T cell response detected by ICS. With the tetramer carrying the dominant HLA-A2 epitope, HAV-specific and activated CD8+ T cells could be detected ex vivo. This first description of the HAV specific CTL-epitopes will allow future studies on strength, breadth, and kinetics of the T-cell response in hepatitis A. Copyright © 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Comparing diagnostic yield of a novel pan-enteric video capsule endoscope with ileocolonoscopy in patients with active Crohn's disease: a feasibility study.

PubMed

Leighton, Jonathan A; Helper, Debra J; Gralnek, Ian M; Dotan, Iris; Fernandez-Urien, Ignacio; Lahat, Adi; Malik, Pramod; Mullin, Gerard E; Rosa, Bruno

2017-01-01

Crohn's disease (CD) is typically diagnosed with ileocolonoscopy (IC); however, when inflammation is localized solely in the small bowel, visualization of the entire small-bowel mucosa can be challenging. The aim of this study was to compare the diagnostic yield of a pan-enteric video capsule endoscope (small-bowel colon [SBC] capsule) versus IC in patients with active CD. This was a prospective, multicenter study. Patients with known active CD and proven bowel luminal patency underwent a standardized colon cleansing protocol followed by ingestion of the capsule. After passage of the capsule, IC was performed and recorded. Lesions indicative of active CD were assessed. One hundred fourteen subjects were screened; 66 subjects completed both endoscopic procedures. The per-subject diagnostic yield rate for active CD lesions was 83.3% for SBC and 69.7% for IC (yield difference, 13.6%; 95% confidence interval [CI], 2.6%-24.7%); 65% of subjects had active CD lesions identified by both modalities. Of the 12 subjects who were positive for active CD by SBC only, 5 subjects were found to have active CD lesions in the terminal ileum. Three subjects were positive for active CD by IC only. Three hundred fifty-five classifying bowel segments were analyzed; the per-segment diagnostic yield rate was 40.6% for SBC and 32.7% for IC (yield difference 7.9%; 95% CI, 3.3%-12.4%). This preliminary study shows that the diagnostic yields for SBC might be higher than IC; however, the magnitude of difference between the two is difficult to estimate. Further study is needed to confirm these findings. Copyright © 2017 American Society for Gastrointestinal Endoscopy. Published by Elsevier Inc. All rights reserved.

Expression pattern of pluripotent markers in different embryonic developmental stages of buffalo (Bubalus bubalis) embryos and putative embryonic stem cells generated by parthenogenetic activation.

PubMed

Singh, Karn P; Kaushik, Ramakant; Garg, Veena; Sharma, Ruchi; George, Aman; Singh, Manoj K; Manik, Radhey S; Palta, Prabhat; Singla, Suresh K; Chauhan, Manmohan S

2012-12-01

In this study, we describe the production of buffalo parthenogenetic blastocysts and subsequent isolation of parthenogenetic embryonic stem cell (PGESC)-like cells. PGESC colonies exhibited dome-shaped morphology and were clearly distinguishable from the feeder layer cells. Different stages of development of parthenogenetic embryos and derived embryonic stem cell (ESC)-like cells expressed key ESC-specific markers, including OCT-4, NANOG, SOX-2, FOXD3, REX-1, STAT-3, TELOMERASE, NUCLEOSTEMIN, and cMYC. Immunofluorescence-based studies revealed that the PGESCs were positive for surface-based pluripotent markers, viz., SSEA-3, SSEA-4, TRA 1-80, TRA 1-60, CD-9, and CD-90 and exhibited high alkaline phosphatase (ALP) activity. PGEC cell-like cells formed embryoid body (EB)-like structures in hanging drop cultures and when cultured for extended period of time spontaneously differentiated into derivatives of three embryonic germ layers as confirmed by RT-PCR for ectodermal (CYTOKERATIN8, NF-68), mesodermal (MSX1, BMP-4, ASA), and endodermal markers (AFP, HNF-4, GATA-4). Differentiation of PGESCs toward the neuronal lineage was successfully directed by supplementation of serum-containing media with retinoic acid. Our results indicate that the isolated ESC-like cells from parthenogenetic blastocyst hold properties of ESCs and express markers of pluripotency. The pluripotency markers were also expressed by early cleavage-stage of buffalo embryos.

IL‐12 and IL‐15 induce the expression of CXCR6 and CD49a on peripheral natural killer cells

PubMed Central

Hydes, Theresa; Noll, Angela; Salinas‐Riester, Gabriela; Abuhilal, Mohammed; Armstrong, Thomas; Hamady, Zaed; Primrose, John; Takhar, Arjun; Walter, Lutz

2017-01-01

Abstract Introduction Murine hepatic NK cells exhibit adaptive features, with liver‐specific adhesion molecules CXCR6 and CD49a acting as surface markers. Methods We investigated human liver‐resident CXCR6+ and CD49a+ NK cells using RNA sequencing, flow cytometry, and functional analysis. We further assessed the role of cytokines in generating NK cells with these phenotypes from the peripheral blood. Results Hepatic CD49a+ NK cells could be induced using cytokines and produce high quantities of IFNγ and TNFα, in contrast to hepatic CXCR6+ NK cells. RNA sequencing of liver‐resident CXCR6+ NK cells confirmed a tolerant immature phenotype with reduced expression of markers associated with maturity and cytotoxicity. Liver‐resident double‐positive CXCR6 + CD49a+ hepatic NK cells are immature but maintain high expression of Th1 cytokines as observed for single‐positive CD49a+ NK cells. We show that stimulation with activating cytokines can readily induce upregulation of both CD49a and CXCR6 on NK cells in the peripheral blood. In particular, IL‐12 and IL‐15 can generate CXCR6 + CD49a+ NK cells in vitro from NK cells isolated from the peripheral blood, with comparable phenotypic and functional features to liver‐resident CD49a+ NK cells, including enhanced IFNγ and NKG2C expression. Conclusion IL‐12 and IL‐15 may be key for generating NK cells with a tissue‐homing phenotype and strong Th1 cytokine profile in the blood, and links peripheral activation of NK cells with tissue‐homing. These findings may have important therapeutic implications for immunotherapy of chronic liver disease. PMID:28952190

IL-12 and IL-15 induce the expression of CXCR6 and CD49a on peripheral natural killer cells.

PubMed

Hydes, Theresa; Noll, Angela; Salinas-Riester, Gabriela; Abuhilal, Mohammed; Armstrong, Thomas; Hamady, Zaed; Primrose, John; Takhar, Arjun; Walter, Lutz; Khakoo, Salim I

2018-03-01

Murine hepatic NK cells exhibit adaptive features, with liver-specific adhesion molecules CXCR6 and CD49a acting as surface markers. We investigated human liver-resident CXCR6+ and CD49a+ NK cells using RNA sequencing, flow cytometry, and functional analysis. We further assessed the role of cytokines in generating NK cells with these phenotypes from the peripheral blood. Hepatic CD49a+ NK cells could be induced using cytokines and produce high quantities of IFNγ and TNFα, in contrast to hepatic CXCR6+ NK cells. RNA sequencing of liver-resident CXCR6+ NK cells confirmed a tolerant immature phenotype with reduced expression of markers associated with maturity and cytotoxicity. Liver-resident double-positive CXCR6 + CD49a+ hepatic NK cells are immature but maintain high expression of Th1 cytokines as observed for single-positive CD49a+ NK cells. We show that stimulation with activating cytokines can readily induce upregulation of both CD49a and CXCR6 on NK cells in the peripheral blood. In particular, IL-12 and IL-15 can generate CXCR6 + CD49a+ NK cells in vitro from NK cells isolated from the peripheral blood, with comparable phenotypic and functional features to liver-resident CD49a+ NK cells, including enhanced IFNγ and NKG2C expression. IL-12 and IL-15 may be key for generating NK cells with a tissue-homing phenotype and strong Th1 cytokine profile in the blood, and links peripheral activation of NK cells with tissue-homing. These findings may have important therapeutic implications for immunotherapy of chronic liver disease. © 2017 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd.

CD52, CD22, CD26, EG5 and IGF-1R expression in thymic malignancies.

PubMed

Remon, J; Abedallaa, N; Taranchon-Clermont, E; Bluthgen, V; Lindsay, C R; Besse, B; Thomas de Montpréville, V

2017-06-01

Thymic epithelial tumours are rare cancers for which new treatment options are required. Identification of putative predictive markers is important for developing clinical trials. We studied the expression of five putative predictive biomarkers, potentially actionable by approved experimental drugs. CD52, CD22, CD26, EG5, and IGF-1R expression were investigated by immunohistochemistry in formalin-fixed surgical samples of thymic epithelial tumour patients. All samples containing 10% positive epithelial tumour cells, independent of tumour cell intensity, were considered as positive. Correlation with histological subtype was performed. 106 surgical samples (89 thymomas, 12 thymic carcinoma, and 5 thymic neuroendocrine tumours) were evaluated. Overall, CD52, CD22, CD26, EG5 and IGF-1R expression was observed in 7%, 42%, 25%, 42% and 77% of samples, respectively. CD52 expression was more frequent in B2 and B3 thymoma. All TET subtypes stained for CD22, mainly AB thymoma (68%). CD26 expression also correlated with AB thymoma (68%), and A thymoma (50%) subtype, while IGFR1 was the most common marker expressed by thymic carcinoma samples (92%), followed by EG5 (60%). Only EG5 expression was significantly higher in thymic carcinomas than in thymomas (75% vs. 38%, p=0.026). Our data were consistent with a previous study of IGF-1R expression. Based on their expression, activity of agents targeting CD52, CD 22, CD26 and EG5 could be further explored in TET patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Breast implant-associated, ALK-negative, T-cell, anaplastic, large-cell lymphoma: establishment and characterization of a model cell line (TLBR-1) for this newly emerging clinical entity.

PubMed

Lechner, Melissa G; Lade, Stephen; Liebertz, Daniel J; Prince, H Miles; Brody, Garry S; Webster, Howard R; Epstein, Alan L

2011-04-01

Primary lymphomas of the breast are very rare (0.2-1.5% of breast malignancies) and the vast majority (95%) are of B-cell origin. Recently, 40 cases of clinically indolent anaplastic large-cell kinase (ALK)-negative, T-cell, anaplastic, non-Hodgkin lymphomas (T-ALCL) have been reported worldwide. A tumor biopsy specimen from a patient in this series was obtained for characterization. By using a human stromal feeder layer and IL-2, a novel cell line, TLBR-1, was established from this biopsy and investigated by using cytogenetics and various biomolecular methods. Immunoperoxidase staining of the tumor biopsy showed a CD30/CD8/CD4 coexpressing T-cell population that was epithelial membrane antigen (EMA)(+) and perforin(+) . Multiplex polymerase chain reaction (PCR) of TCRγ genes showed monoclonality that suggested a T-cell origin, yet pan-T markers CD2/5/7, anaplastic large-cell kinase (ALK)-1, pancytokeratins, CD20, CD56, and Epstein-Barr virus (EBV) by in situ hybridization (ISH) were negative. TLBR-1 is IL-2 dependent, has a relatively long doubling time (55 hours), and displays different cellular shapes in culture. Cytogenetic analysis of tumor and TLBR-1 cells confirmed a highly anaplastic cell population with a modal number of 47 chromosomes lacking t(2;5). PCR screens for EBV and human T-lymphotropic virus types 1 and 2 (HTLV-1/2) were negative. Fluorescence-activated cell-sorting (FACS) analysis showed strong positivity for CD4/8, CD30, CD71, and CD26 expression, and antigen presentation (HLA-DR(+) CD80(+) CD86(+) ), IL-2 signaling (CD25(+) CD122(+) ), and NK (CD56(+) ) markers, and Western blots demonstrated strong Notch1 expression. Severe combined immunodeficiency (SCID) mouse TLBR-1 heterotransplants recapitulated the histology and marker characteristics of the original tumor. TLBR-1, a novel ALK-negative, T-cell, anaplastic, large-cell lymphoma, closely resembles the original biopsy and represents an important tool for studying this newly recognized disease entity. Copyright © 2010 American Cancer Society.

Breast Implant-Associated, ALK-Negative, T-Cell, Anaplastic, Large-Cell Lymphoma: Establishment and Characterization of a Model Cell Line (TLBR-1) for This Newly Emerging Clinical Entity

PubMed Central

Lechner, Melissa G.; Lade, Stephen; Liebertz, Daniel J.; Prince, H. Miles; Brody, Garry S.; Webster, Howard R.; Epstein, Alan L.

2014-01-01

BACKGROUND Primary lymphomas of the breast are very rare (0.2–1.5% of breast malignancies) and the vast majority (95%) are of B-cell origin. Recently, 40 cases of clinically indolent anaplastic large-cell kinase (ALK)-negative, T-cell, anaplastic, non-Hodgkin lymphomas (T-ALCL) have been reported worldwide. METHODS A tumor biopsy specimen from a patient in this series was obtained for characterization. By using a human stromal feeder layer and IL-2, a novel cell line, TLBR-1, was established from this biopsy and investigated by using cytogenetics and various biomolecular methods. RESULTS Immunoperoxidase staining of the tumor biopsy showed a CD30/CD8/CD4 coexpressing T-cell population that was epithelial membrane antigen (EMA)+ and perforin+. Multiplex polymerase chain reaction (PCR) of TCRγ genes showed monoclonality that suggested a T-cell origin, yet pan-T markers CD2/5/7, anaplastic large-cell kinase (ALK)-1, pancytokeratins, CD20, CD56, and Epstein-Barr virus (EBV) by in situ hybridization (ISH) were negative. TLBR-1 is IL-2 dependent, has a relatively long doubling time (55 hours), and displays different cellular shapes in culture. Cytogenetic analysis of tumor and TLBR-1 cells confirmed a highly anaplastic cell population with a modal number of 47 chromosomes lacking t(2;5). PCR screens for EBV and human T-lymphotropic virus types 1 and 2 (HTLV-1/2) were negative. Fluorescence-activated cell-sorting (FACS) analysis showed strong positivity for CD4/8, CD30, CD71, and CD26 expression, and antigen presentation (HLA-DR+CD80+CD86+), IL-2 signaling (CD25+CD122+), and NK (CD56+) markers, and Western blots demonstrated strong Notch1 expression. Severe combined immunodeficiency (SCID) mouse TLBR-1 heterotransplants recapitulated the histology and marker characteristics of the original tumor. CONCLUSIONS TLBR-1, a novel ALK-negative, T-cell, anaplastic, large-cell lymphoma, closely resembles the original biopsy and represents an important tool for studying this newly recognized disease entity. PMID:21425149

Down-regulation of microRNA-451a facilitates the activation and proliferation of CD4+ T cells by targeting Myc in patients with dilated cardiomyopathy.

PubMed

Zeng, Zhipeng; Wang, Ke; Li, Yuanyuan; Xia, Ni; Nie, Shaofang; Lv, Bingjie; Zhang, Min; Tu, Xin; Li, Qianqian; Tang, Tingting; Cheng, Xiang

2017-04-07

CD4 + T cells are abnormally activated in patients with dilated cardiomyopathy (DCM) and might be associated with the immunopathogenesis of the disease. However, the underlying mechanisms of CD4 + T cell activation remain largely undefined. Our aim was to investigate whether the dysregulation of microRNAs (miRNAs) was associated with CD4 + T cell activation in DCM. CD4 + T cells from DCM patients showed increased expression levels of CD25 and CD69 and enhanced proliferation in response to anti-CD3/28, indicating an activated state. miRNA profiling analysis of magnetically sorted CD4 + T cells revealed a distinct pattern of miRNA expression in CD4 + T cells from DCM patients compared with controls. The level of miRNA-451a (miR-451a) was significantly decreased in the CD4 + T cells of DCM patients compared with that of the controls. The transfection of T cells with an miR-451a mimic inhibited their activation and proliferation, whereas an miR-451a inhibitor produced the opposite effects. Myc was directly inhibited by miR-451a via interaction with its 3'-UTR, thus identifying it as an miR-451a target in T cells. The knockdown of Myc suppressed the activation and proliferation of T cells, and the expression of Myc was significantly up-regulated at the mRNA level in CD4 + T cells from patients with DCM. A strong inverse correlation was observed between the Myc mRNA expression and miR-451a transcription level. Our data suggest that the down-regulation of miR-451a contributes to the activation and proliferation of CD4 + T cells by targeting the transcription factor Myc in DCM patients and may contribute to the immunopathogenesis of DCM. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Purification of human adipose-derived stem cells from fat tissues using PLGA/silk screen hybrid membranes.

PubMed

Chen, Da-Chung; Chen, Li-Yu; Ling, Qing-Dong; Wu, Meng-Hsueh; Wang, Ching-Tang; Suresh Kumar, S; Chang, Yung; Munusamy, Murugan A; Alarfajj, Abdullah A; Wang, Han-Chow; Hsu, Shih-Tien; Higuchi, Akon

2014-05-01

The purification of human adipose-derived stem cells (hADSCs) from human adipose tissue cells (stromal vascular fraction) was investigated using membrane filtration through poly(lactide-co-glycolic acid)/silk screen hybrid membranes. Membrane filtration methods are attractive in regenerative medicine because they reduce the time required to purify hADSCs (i.e., less than 30 min) compared with conventional culture methods, which require 5-12 days. hADSCs expressing the mesenchymal stem cell markers CD44, CD73, and CD90 were concentrated in the permeation solution from the hybrid membranes. Expression of the surface markers CD44, CD73, and CD99 on the cells in the permeation solution from the hybrid membranes, which were obtained using 18 mL of feed solution containing 50 × 10⁴ cells, was statistically significantly higher than that of the primary adipose tissue cells, indicating that the hADSCs can be purified in the permeation solution by the membrane filtration method. Cells expressing the stem cell-associated marker CD34 could be successfully isolated in the permeation solution, whereas CD34⁺ cells could not be purified by the conventional culture method. The hADSCs in the permeation solution demonstrated a superior capacity for osteogenic differentiation based on their alkali phosphatase activity, their osterix gene expression, and the results of mineralization analysis by Alizarin Red S and von Kossa staining compared with the cells from the suspension of human adipose tissue. These results suggest that the hADSCs capable of osteogenic differentiation preferentially permeate through the hybrid membranes. Copyright © 2014 Elsevier Ltd. All rights reserved.

Presence of Antigen-Experienced T Cells with Low Grade of Differentiation and Proliferative Potential in Chronic Chagas Disease Myocarditis

PubMed Central

Cabeza-Meckert, Patricia; Viotti, Rodolfo; Garelli, Fernando; Favaloro, Liliana E.; Favaloro, Roberto R.; Laguens, Rubén; Laucella, Susana A.

2014-01-01

Background The main consequence of chronic Trypanosoma cruzi infection is the development of myocarditis in approximately 20–30% of infected individuals but not until 10–20 years after the initial infection. We have previously shown that circulating interferon-γ-secreting T cells responsive to Trypanosoma cruzi antigens in chronic Chagas disease patients display a low grade of differentiation and the frequency of these T lymphocytes decreases along with the severity of heart disease. This study thought to explore the expression of inhibitory receptors, transcription factors of type 1 or regulatory T cells, and markers of T cell differentiation, immunosenescence or active cell cycle in cardiac explants from patients with advanced Chagas disease myocarditis. Methodology/Principal Findings The expression of different markers for T and B cells as well as for macrophages was evaluated by immunohistochemistry and immunofluorescence techniques in cardiac explants from patients with advanced chronic Chagas disease submitted to heart transplantation. Most infiltrating cells displayed markers of antigen-experienced T cells (CD3+, CD4+, CD8+, CD45RO+) with a low grade of differentiation (CD27+, CD57−, CD45RA−, PD-1−). A skewed T helper1/T cytotoxic 1 profile was supported by the expression of T-bet; whereas FOXP3+ cells were scarce and located only in areas of severe myocarditis. In addition, a significant proliferative capacity of CD3+ T cells, assessed by Ki67 staining, was found. Conclusions/Significance The quality of T cell responses and immunoregulatory mechanisms might determine the pattern of the cellular response and the severity of disease in chronic Trypanosoma cruzi infection. PMID:25144227

Progenitor Cells from Cartilage: Grade Specific Differences in Stem Cell Marker Expression

PubMed Central

Mazor, Marija; Cesaro, Annabelle; Ali, Mazen; Best, Thomas M.; Lespessaille, Eric; Toumi, Hechmi

2017-01-01

Recent research has confirmed the presence of Mesenchymal stem cell (MSC)-like progenitors (MPC) in both normal and osteoarthritic cartilage. However, there is only limited information concerning how MPC markers are expressed with osteoarthritis (OA) progression. The purpose of this study was to compare the prevalence of various MPC markers in different OA grades. Human osteoarthritic tibial plateaus were obtained from ten patients undergoing total knee replacement. Each sample had been classified into a mild or severe group according to OARSI scoring. Tissue was taken from each specimen and mRNA expression levels of CD105, CD166, Notch 1, Sox9, Acan and Col II A1 were measured at day 0 and day 14 (2 weeks in vitro). Furthermore, MSC markers: Nucleostemin, CD90, CD73, CD166, CD105 and Notch 1 were studied by immunofluorescence. mRNA levels of MSC markers did not differ between mild and severe OA at day 0. At day 14, protein analysis showed that proliferated cells from both sources expressed all 6 MSC markers. Only cells from the mild OA subjects resulted in a significant increase of mRNA CD105 and CD166 after in vitro expansion. Moreover, cells from the mild OA subjects showed significantly higher levels of CD105, Sox9 and Acan compared with those from severe OA specimens. Results confirmed the presence of MSC markers in mild and severe OA tissue at both mRNA and protein levels. We found significant differences between cells obtained from mild compared to severe OA specimens suggests that mild OA derived cells may have a greater MSC potential. PMID:28805694

Long-lived tissue resident HIV-1 specific memory CD8+ T cells are generated by skin immunization with live virus vectored microneedle arrays.

PubMed

Zaric, Marija; Becker, Pablo Daniel; Hervouet, Catherine; Kalcheva, Petya; Ibarzo Yus, Barbara; Cocita, Clement; O'Neill, Lauren Alexandra; Kwon, Sung-Yun; Klavinskis, Linda Sylvia

2017-12-28

The generation of tissue resident memory (T RM ) cells at the body surfaces to provide a front line defence against invading pathogens represents an important goal in vaccine development for a wide variety of pathogens. It has been widely assumed that local vaccine delivery to the mucosae is necessary to achieve that aim. Here we characterise a novel micro-needle array (MA) delivery system fabricated to deliver a live recombinant human adenovirus type 5 vaccine vector (AdHu5) encoding HIV-1 gag. We demonstrate rapid dissolution kinetics of the microneedles in skin. Moreover, a consequence of MA vaccine cargo release was the generation of long-lived antigen-specific CD8 + T cells that accumulate in mucosal tissues, including the female genital and respiratory tract. The memory CD8 + T cell population maintained in the peripheral mucosal tissues was attributable to a MA delivered AdHu5 vaccine instructing CD8 + T cell expression of CXCR3 + , CD103 +, CD49a + , CD69 + , CD127 + homing, retention and survival markers. Furthermore, memory CD8 + T cells generated by MA immunization significantly expanded upon locally administered antigenic challenge and showed a predominant poly-functional profile producing high levels of IFNγ and Granzyme B. These data demonstrate that skin vaccine delivery using microneedle technology induces mobilization of long lived, poly-functional CD8 + T cells to peripheral tissues, phenotypically displaying hallmarks of residency and yields new insights into how to design and deliver effective vaccine candidates with properties to exert local immunosurveillance at the mucosal surfaces. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

CD25 and CD69 induction by α4β1 outside-in signalling requires TCR early signalling complex proteins

PubMed Central

Cimo, Ann-Marie; Ahmed, Zamal; McIntyre, Bradley W.; Lewis, Dorothy E.; Ladbury, John E.

2013-01-01

Distinct signalling pathways producing diverse cellular outcomes can utilize similar subsets of proteins. For example, proteins from the TCR (T-cell receptor) ESC (early signalling complex) are also involved in interferon-α receptor signalling. Defining the mechanism for how these proteins function within a given pathway is important in understanding the integration and communication of signalling networks with one another. We investigated the contributions of the TCR ESC proteins Lck (lymphocyte-specific kinase), ZAP-70 (ζ-chain-associated protein of 70 kDa), Vav1, SLP-76 [SH2 (Src homology 2)-domain-containing leukocyte protein of 76 kDa] and LAT (linker for activation of T-cells) to integrin outside-in signalling in human T-cells. Lck, ZAP-70, SLP-76, Vav1 and LAT were activated by α4β1 outside-in signalling, but in a manner different from TCR signalling. TCR stimulation recruits ESC proteins to activate the mitogen-activated protein kinase ERK (extracellular-signal-regulated kinase). α4β1 outside-in-mediated ERK activation did not require TCR ESC proteins. However, α4β1 outside-in signalling induced CD25 and co-stimulated CD69 and this was dependent on TCR ESC proteins. TCR and α4β1 outside-in signalling are integrated through the common use of TCR ESC proteins; however, these proteins display functionally distinct roles in these pathways. These novel insights into the cross-talk between integrin outside-in and TCR signalling pathways are highly relevant to the development of therapeutic strategies to overcome disease associated with T-cell deregulation. PMID:23758320

Serum soluble CD163 levels in patients with influenza-associated encephalopathy.

PubMed

Hasegawa, Shunji; Matsushige, Takeshi; Inoue, Hirofumi; Takahara, Midori; Kajimoto, Madoka; Momonaka, Hiroshi; Ishida, Chiemi; Tanaka, Saya; Morishima, Tsuneo; Ichiyama, Takashi

2013-08-01

Influenza-associated encephalopathy (IE) is a serious complication during influenza viral infection. Common clinical symptoms of IE include seizures and progressive coma with high-grade fever. We previously reported that hypercytokinemia and monocyte/macrophage activation may play an important role in the pathogenesis of IE. CD163 is a scavenger receptor for hemoglobin-haptoglobin complexes and is expressed by monocytes/macrophages. Proteolytic cleavage of monocyte-bound CD163 by matrix metalloproteinases releases soluble CD163 (sCD163). However, there have been no reports regarding serum sCD163 levels in IE patients. We measured serum levels of sCD163 as a marker of monocyte/macrophage activation in IE patients with poor outcomes, those without neurological sequelae, influenza patients without IE, and control subjects. Serum sCD163 levels were significantly higher in IE patients with poor outcomes than in those without neurological sequelae. In particular, sCD163 levels in cases of death were significantly higher than those in other cases. Our results suggest that monocyte/macrophage activation is related to the pathogenesis of severe IE. Copyright © 2012 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

FOXP3 and GARP (LRRC32): the master and its minion.

PubMed

Probst-Kepper, Michael; Buer, Jan

2010-02-05

The transcription factor FOXP3 is essential for the development and function of CD4+CD25hiFOXP3+ regulatory T (T(reg)) cells, but also expressed in activated human helper T cells without acquisition of a regulatory phenotype. This comment focuses on glycoprotein-A repetitions predominant (GARP or LRRC32) recently identified as specific marker of activated human T(reg) cells, which may provide the missing link toward a better molecular definition of the regulatory phenotype.

Canonical Wnt Signaling as a Specific Marker of Normal and Tumorigenic Mammary Stem Cells

DTIC Science & Technology

2013-02-01

get enough sorted mammary cells for the transplantation experiments. We are currently working with our Flow Cytometry Core to sort Lin-/CD24+/CD49...activity our flow cytometry data suggests t here is a 2-fold increase in the number of FOG+ MEC’s in BATgal animals compared to contro ls which...this populat ion of cells is enriched for stem cell activity. Flow cytometry will determine the percentage of FOG+ cells within pre-neoplastic BATgai

Macrophages and prognosis of oral squamous cell carcinoma: A systematic review.

PubMed

Alves, Alessandro Menna; Diel, Leonardo Francisco; Lamers, Marcelo Lazzaron

2018-05-01

Oral Squamous Cell Carcinoma (OSCC) presents a tumor microenvironment rich in inflammatory cells. Depending on the stimulus, macrophages can polarize in M1 or M2 profile, where M1 acts as proinflammatory and antitumor, and M2 is anti-inflammatory and shows protumor activity. Several studies have shown that macrophages are important to the prognosis of patients with different types of cancer. Our aim was to conduct a systematic review to evaluate the role of macrophages in the prognosis of OSCC patients. A search in the Pubmed, Scopus, and ISI Web of Knowledge database was performed, and it was included only studies that evaluated the importance of macrophages in the prognosis of OSCC patients. From initial 286 articles, 14 fully attended the inclusion criteria. In the majority of the articles, it was evaluated only CD68, a panmacrophage marker, or CD163, a M2 marker. Only one article evaluated the M1 marker, CD11c. Besides, 5 articles analyzed the presence of macrophages in different areas of the tumor. Higher concentrations of CD68 and CD163 were associated with worse survival. In conclusion, macrophages are important to OSCC patients' prognosis; however, it is necessary to address in which tumor region the presence of polarized macrophage is more important to the outcome. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

On the reliability of the CD123-endowed basophil activation test (BAT) and its application in food allergy.

PubMed

Chirumbolo, Salvatore; Bjørklund, Geir; Vella, Antonio

2018-06-15

the recent paper by Appel et al., evaluated the use of the basophil activaion test (BAT) in sesame allergy and concluded that BAT should be preferentially used in association with high protein sesame extract-skin prick test (HSPE-SPT) for the diagnosis of sesame food allergy, so preventing any ethical issue regarding the possble risk associated with oral food challenge (OFC) 1 . The authors used a non commercial BAT approach endowed with the CD123 pos /HLADR neg phenotyping protocol and using CD63 and/or CD203c as activation markers 2,3 . Basophils were initially captured in flow cytometry (FC) from a not clearly separated side scatter (SSC)-CD123 pos . This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Coexpression of CD14 and CD326 discriminate hepatic precursors in the human fetal liver.

PubMed

Fomin, Marina E; Tai, Lung-Kuo; Bárcena, Alicia; Muench, Marcus O

2011-07-01

The molecular and cellular profile of liver cells during early human development is incomplete, complicating the isolation and study of hepatocytes, cholangiocytes, and hepatic stem cells from the complex amalgam of hepatic and hematopoietic cells, that is, the fetal liver. Epithelial cell adhesion molecule, CD326, has emerged as a marker of hepatic stem cells, and lipopolysaccharide receptor CD14 is known to be expressed on adult hepatocytes. Using flow cytometry, we studied the breadth of CD326 and CD14 expression in midgestation liver. Both CD45(+) hematopoietic and CD45(-) nonhematopoietic cells expressed CD326. Moreover, diverse cell types expressing CD326 were revealed among CD45(-) cells by costaining for CD14. Fluorescence-activated cell sorting was used to isolate nonhematopoietic cells distinguished by expression of high levels of CD326 and low CD14 (CD326(++)CD14(lo)), which were characterized for gene expression associated with liver development. CD326(++)CD14(lo) cells expressed the genes albumin, α-fetoprotein, hepatic nuclear factor 3α, prospero-related homeobox 1, cytochrome P450 3A7, α(1)-antitrypsin, and transferrin. Proteins expressed included cell-surface CD24, CD26, CD29, CD34, CD49f, CD243, and CD324 and, in the cytoplasm, cytokeratins-7/8 (CAM 5.2 antigen) and some cytokeratin-19. Cultured CD326(++)CD14(lo) cells yielded albumin(+) hepatocytes, cytokeratin-19(+) cholangiocytes, and hepatoblasts expressing both markers. Using epifluorescence microscopy we observed CD326 and CD14 expression on fetal hepatocytes comprising the liver parenchyma, as well as on cells associated with ductal plates and surrounding large vessels. These findings indicate that expression of CD14 and CD326 can be used to identify functionally distinct subsets of fetal liver cells, including CD326(++)CD14(lo) cells, representing a mixture of parenchymal cells, cholangiocytes, and hepatoblasts.

Ex vivo expansion of human umbilical cord blood-derived T-lymphocytes with homologous cord blood plasma.

PubMed

Kim, Yong-Man; Jung, Min-Hyung; Song, Ha-Young; Yang, Hyun Ok; Lee, Sung-Tae; Kim, Jong-Hyeok; Kim, Young-Tak; Nam, Joo-Hyun; Mok, Jung-Eun

2005-02-01

This study was designed to establish a more effective and safe culture system for adoptive immunotherapy by investigating the use of homologous cord blood plasma (HCBP) instead of fetal bovine serum (FBS), which has various limitations including ethical problems for the ex vivo expansion of human umbilical T lymphocytes. Fresh human umbilical mononuclear cell fractions were isolated by Ficoll-Hypaque density centrifugation. Nonadherent mononuclear cell fractions were cultured with anti-CD3 antibody (5 microg/ml), IL-2 (175 U/ml), and either 10% FBS or 10% HCBP. On day 8, the cellular proliferation rate and cell surface markers were assessed. There was no significant difference in proliferation when human umbilical cord blood T lymphocytes were grown in medium supplemented with FBS or HCBP (p > 0.05). In medium containing FBS, the proportion of CD3(+)CD4(+) (markers for helper T cell), CD3(+)CD8(+) (cytotoxic T cell), CD3(+)CD25(+) (activated T cell), CD3(+)CD38(+) (immature T cell), and CD3(+)CD45RO(+) (memory T cell) cells was significantly increased (p < 0.05), whereas proportion of CD3(+)CD45RA(+) (naive T cell) and CD16(+)CD56(+) (NK cell) cells was significantly decreased (p < 0.05). In HCBP supplemented medium, the proportion of CD3(+)CD8(+), CD3(+)CD25(+), CD3(+)CD45RA(+), and CD3(+)CD45RO(+) cells was significantly increased (p < 0.05). The proportion of CD3(+)CD4(+), CD3(+)CD45RO(+) and CD3(+)CD38(+) cells was significantly higher, but proportion of CD3(+)CD45RA(+) and CD3(+)CD8(+) cells was significantly lower in FBS compared with HCBP supplemented medium (p < 0.05). Our results support the feasibility of ex vivo expansion of human umbilical cord blood T lymphocytes in medium supplemented with HCBP for future adoptive cellular immunotherapy.

Elevated Levels of Serum Vascular Endothelial Growth Factor-A Are Not Related to NK Cell Parameters in Recurrent IVF Failure.

PubMed

Bansal, Rhea; Ford, Brian; Bhaskaran, Shree; Thum, Meenyau; Bansal, Amolak

2017-01-01

Vascular Endothelial Growth Factor and NK cells have an interrelated role in angiogenesis that is critical for placentation and success of in vitro fertilization. An attempt was made to assess a possible relationship between the two in this study. A case control study was performed comparing the serum levels of VEGF-A and its receptor VEGF-R1 with levels of NK cells, activated NK cells and NK cytotoxicity in 62 women with Repeated Implantation Failure (RIF). The healthy control group consisted of 72 women of similar age, without known issues in achieving pregnancy or evidence of autoimmunity. Levels of VEGF-A and VEGF-R1 were quantified by ELISA methods with standard curve interpolation. NK cell subsets were determined with flow cytometry using fluorescent-tagged anti-CD56, anti-CD16, anti-CD3 and anti-CD69. NK cytotoxicity was performed by incubating peripheral blood mononuclear cells and K562 cultured cells with propidium iodide, steroid, intralipid and intravenous immunoglobulin, using previously described methods. Statistical analysis involved Mann-Whitney-U and Spearman's rank correlation testing with p-values defined as <0.05. It was found that VEGF-A levels were significantly raised in women with RIF compared to healthy controls (362.9 vs . 171.6 pg/ml , p<0.0001), with no difference in VEGF-R1 levels between groups (1499 vs . 1202 pg/ml , p=0.4082). There was no correlation between VEGF-A or VEGF-R1 and the absolute levels of circulating NK cells, CD69 activated NK cells or NK cytotoxicity. The absence of correlation between VEGF-A or VEGF-R1 and NK cells suggests VEGF secretion and regulation is independent of NK cell activity in RIF.

Interferon-γ regulates the function of mesenchymal stem cells from oral lichen planus via indoleamine 2,3-dioxygenase activity.

PubMed

Zhang, Zhihui; Han, Ying; Song, Jiangyuan; Luo, Ruxi; Jin, Xin; Mu, Dongdong; Su, Sha; Ji, Xiaoli; Ren, Yan-Fang; Liu, Hongwei

2015-01-01

Little is known about mesenchymal stem cells (MSCs) in normal or inflammatory oral mucosal tissues, such as in oral lichen planus (OLP). Our objectives were to identify, isolate, and characterize MSCs from normal human oral mucosa and OLP lesions, and to evaluate indoleamine 2,3 dioxygenase (IDO) activity in mediating immunomodulation of MSCs from these tissues. Expressions of MSCs-related markers were examined in isolated cells by flow cytometry. Self-renewal and multilineage differentiations were studied to characterize these MSCs. Interferon-γ (IFN-γ), IDO, and STRO-1 were assessed by immunofluorescence. MSCs from oral mucosa and OLP or IFN-γ-pretreated MSCs were co-cultured with allogeneic mixed lymphocyte reaction assays (MLR). Proliferation and apoptosis of MLR or MSCs were detected by CCK8 and the annexin V-FITC apoptosis detection kit, respectively. IDO expression and activity were measured by real-time PCR, Western blotting, and high-performance liquid chromatography. Isolated cells from oral mucosa and OLP expressed MSC-related markers STRO-1, CD105, and CD90 but were absent for hematopoietic stem cell markers CD34. Besides, they all showed self-renewal and multilineage differentiation capacities. MSCs in OLP presented STRO-1/IDO+ phenotype by immunofluorescence. MSCs and IFN-γ-pretreated MSCs could inhibit lymphocyte proliferation via IDO activity, but not via cell apoptosis. Long-term IFN-γ could also inhibit MSC proliferation via IDO activity. Mesenchymal stem cells can be isolated from human oral mucosa and OLP tissues. Besides self-renewal and multilineage differentiation properties, these cells may participate in immunomodulation mediated by IFN-γ via IDO activity in human OLP. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Caspase-3 Is Transiently Activated without Cell Death during Early Antigen Driven Expansion of CD8+ T Cells In Vivo

PubMed Central

McComb, Scott; Mulligan, Rebecca; Sad, Subash

2010-01-01

Background CD8+ T cell responses develop rapidly during infection and are swiftly reduced during contraction, wherein >90% of primed CD8+ T cells are eliminated. The role of apoptotic mechanisms in controlling this rapid proliferation and contraction of CD8+ T cells remains unclear. Surprisingly, evidence has shown non-apoptotic activation of caspase-3 to occur during in vitro T-cell proliferation, but the relevance of these mechanisms to in vivo CD8+ T cell responses has yet to be examined. Methods and Findings We have evaluated the activity of caspase-3, a key downstream inducer of apoptosis, throughout the entirety of a CD8+ T cell response. We utilized two infection models that differ in the intensity, onset and duration of antigen-presentation and inflammation. Expression of cleaved caspase-3 in antigen specific CD8+ T cells was coupled to the timing and strength of antigen presentation in lymphoid organs. We also observed coordinated activation of additional canonical apoptotic markers, including phosphatidylserine exposure. Limiting dilution analysis directly showed that in the presence of IL7, very little cell death occurred in both caspase-3hi and caspase-3low CD8+ T cells. The expression of active caspase-3 peaked before effector phenotype (CD62Llow) CD8+ T cells emerged, and was undetectable in effector-phenotype cells. In addition, OVA-specific CD8+ cells remained active caspase-3low throughout the contraction phase. Conclusions Our results specifically implicate antigen and not inflammation in driving activation of apoptotic mechanisms without cell death in proliferating CD8+ T cells. Furthermore, the contraction of CD8+ T cell response following expansion is likely not mediated by the key downstream apoptosis inducer, caspase-3. PMID:21203525

Cross-sectional study of CD4: CD8 ratio recovery in young adults with perinatally acquired HIV-1 infection.

PubMed

Pollock, Katrina M; Pintilie, Hannah; Foster, Caroline; Fidler, Sarah

2018-02-01

Antiretroviral therapy (ART) has improved survival into adulthood for young people with perinatally acquired HIV-1 (yp-PaHIV), but long-term prognosis remains unclear. We hypothesized that on-going immune activation, reflected in the failure of CD4:CD8 ratio normalization would be observed in yp-PaHIV, despite ART.A cross-sectional study of routinely collected clinical data from a cohort of yp-PaHIV (≥16 years).Data were collected from records of individuals attending a specialist clinic for yp-PaHIV transitioning to adult care. CD4:CD8 ratio and proportion with CD4:CD8 ratio ≥1, demographic data and viral parameters, including HIV-1 viral load (VL) and human cytomegalovirus (CMV) IgG, were analyzed with IBM SPSS Statistics v22.A total of 115 yp-PaHIV, median (IQR) age 22.0 (20.0-24.0) years, were studied, of whom 59 were females, and the majority were Black African 75/115 (65.2%). Where measured, CMV antibodies were frequently detected (71/74, 95.9%) and CMV IgG titre was inversely associated with CD4:CD8 ratio, (Rho -0.383, P = .012). Of those taking ART, 69 out of 90 (76.7%) yp-PaHIV had suppressed HIV viremia (<50 RNA copies/mL) and recovery of CD4:CD8 ratio to ≥1 was seen in 26 out of 69 (37.7%) with suppressed HIV viremia. Persistence of low CD4:CD8 ratio was observed even in those with a CD4 count ≥500 cells/μL, where 28/52 (53.8%) had a CD4:CD8 ratio <1. Of those with suppressed viremia, the median (IQR) age for starting ART was 8.0 (5.0-12.8) years and CD4:CD8 ratio was inversely associated with age at ART start, Rho -0.348, (P = .028).In this cohort of yp-PaHIV, despite lifelong HIV infection and widespread CMV coinfection, CD4:CD8 ratio recovery rate was comparable to adults treated in acute infection. Where persistence of CD4:CD8 ratio abnormality was observed, on-going immune activation may have significance for non-AIDS outcomes. Taken together our findings indicate immune resilience to be a feature of these adult survivors of perinatally acquired HIV infection, which can be supported with early antiretroviral therapy.

A Multiepitope of XBP1, CD138 and CS1 Peptides Induces Myeloma-Specific Cytotoxic T lymphocytes in T cells of Smoldering Myeloma Patients

PubMed Central

Bae, Jooeun; Prabhala, Rao; Voskertchian, Annie; Brown, Andrew; Maguire, Craig; Richardson, Paul; Dranoff, Glen; Anderson, Kenneth C.; Munshi, Nikhil C.

2014-01-01

We evaluated a cocktail of HLA-A2-specific peptides including heteroclitic XBP1 US184-192 (YISPWILAV), heteroclitic XBP1 SP367-375 (YLFPQLISV), native CD138260-268 (GLVGLIFAV) and native CS1239-247 (SLFVLGLFL), for their ability to elicit multipeptide specific cytotoxic T lymphocytes (MP-CTL) using T cells from smoldering multiple myeloma (SMM) patients. Our results demonstrate that MP-CTL generated from SMM patients’ T cells show effective anti-MM responses including CD137 (4-1BB) upregulation, CTL proliferation, IFN-γ production, and degranulation (CD107a) in an HLA-A2-restricted and peptide-specific manner. Phenotypically, we observed increased total CD3+CD8+ T cells (>80%) and cellular activation (CD69+) within the memory SMM MP-CTL (CD45RO+/CD3+CD8+) subset after repeated multipeptide stimulation. Importantly, SMM patients could be categorized into distinct groups by their level of MP-CTL expansion and anti-tumor activity. In high responders, the effector memory (CCR7-CD45RO+/CD3+CD8+) T cell subset was enriched, while the remaining responders’ CTL contained a higher frequency of the terminal effector (CCR7-CD45RO-/CD3+CD8+) subset. These results suggest that this multipeptide cocktail has the potential to induce effective and durable memory MP-CTL in SMM patients. Therefore, our findings provide the rationale for clinical evaluation of a therapeutic vaccine to prevent or delay progression of SMM to active disease. PMID:24935722

Decreased NK-Cell Cytotoxicity after Short Flights on the Space Shuttle

NASA Technical Reports Server (NTRS)

Mehta, Satish K.; Grimm, Elizabeth A.; Smid, Christine; Kaur, Indreshpal; Feeback, Daniel L.; Pierson, Duane L.

2000-01-01

Cytotoxic activity of natural killer (NK) cells and cell surface marker expression of peripheral blood mononuclear cells (PBMCs) isolated from 11 U.S. astronauts on two different missions were determined before and after 9 or 10 days of spaceflight aboard the space shuttle. Blood samples were collected 10 and 3 days before launch, within 3 hours after landing, and 3 days after landing. All PBMC preparations were cryopreserved and analyzed simultaneously in a 4-hour cytotoxicity "Cr-release assay using NK-sensitive K-562 target cells. Compared to preflight values, NK-cell cytotoxicity (corrected for lymphopenia observed on landing day) was significantly decreased at landing (P < 0.0125). It then apparently began to recover and approached preflight values by 3 days after landing. Consistent with decreased NK-cell cytotoxicity, significant increases from preflight values were found in plasma adrenocorticotropic hormone at landing. Plasma and urinary cortisol levels did not change significantly from preflight values. Expression of major lymphocyte surface markers (CD3, CD4, CD8, CD14, CD16, CD56), determined by flow cytometric analysis, revealed no consistent phenotypic changes in relative percent of NK or other lymphoid cells after 10 days of spaceflight.

Functional and genomic analyses of FOXP3-transduced Jurkat-T cells as regulatory T (Treg)-like cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Joon-Young; Kim, Han-Jong; Hurt, Elaine M.

2007-10-12

FOXP3, a forkhead transcription factor is essential for the development and function of CD4{sup +}CD25{sup +} regulatory T cells (Tregs). In contrast to conversion of murine naive T cells to Tregs by transduction of Foxp3, it is controversial whether ectopic expression of FOXP3 in human CD4{sup +} T cells is sufficient for acquisition of suppressive activity. Here, we show that retroviral transduction of FOXP3 induces a Treg phenotype in human leukemic CD4{sup +} Jurkat-T cells, evidenced by increased expression of Treg-associated cell surface markers as well as inhibition of cytokine production. Furthermore, FOXP3-transduced Jurkat-T cells suppress the proliferation of humanmore » CD4{sup +}CD25{sup -} T cells. Additionally, DNA microarray analysis identifies Treg-related genes regulated by FOXP3. Our study demonstrates that enforced expression of FOXP3 confers Treg-like properties on Jurkat-T cells, which can be a convenient and efficient Treg-like cell model for further study to identify Treg cell surface markers and target genes regulated by FOXP3.« less

Granzyme B as a diagnostic marker of tuberculosis in patients with and without HIV coinfection.

PubMed

Sarkar, Pronoti; Mitra, Soumik; Pant, Priyannk; Kotwal, Aarti; Kakati, Barnali; Masih, Victor; Sindhwani, Girish; Biswas, Debasis

2016-05-01

Immunodiagnostic tests for tuberculosis (TB) are based on the estimation of interferon γ (IFN-γ) or IFN-γ-secreting CD4(+) T cells following ex vivo stimulation with ESAT6 and CFP-10. Sensitivity of these tests is likely to be compromised in CD4(+) T-cell-depleted situations, like HIV-TB coinfection. CD4(+) and CD8(+) T cells, isolated from 3 groups, viz., HIV-negative patients with active TB, HIV-TB coinfected patients, and healthy household contacts (HHCs) were cocultivated with autologous dendritic cells, and the cytokine response to rESAT6 stimulation was compared between groups in supernatants. While CD4(+) T-cell stimulation yielded significantly elevated levels of IFN-γ and interleukin 4 in HIV-negative TB patients, compared to HHCs, the levels of both these cytokines were nondiscriminatory between HIV-positive TB patients and HHCs. However, CD8(+) T-cell stimulation yielded significantly elevated granzyme B titers in both groups of patients, irrespective of HIV coinfection status. Hence, contrary to IFN-γ, granzyme B might be a useful diagnostic marker for Mycobacterium tuberculosis infection particularly in HIV coinfected patients. Copyright © 2016 Elsevier Inc. All rights reserved.

Flow cytometric assessment of activation of peripheral blood platelets in dogs with normal platelet count and asymptomatic thrombocytopenia.

PubMed

Żmigrodzka, M; Guzera, M; Winnicka, A

2016-01-01

Platelets play a crucial role in hemostasis. Their activation has not yet been evaluated in healthy dogs with a normal and low platelet count. The aim of this study was to determine the influence of activators on platelet activation in dogs with a normal platelet count and asymptomatic thrombocytopenia. 72 clinically healthy dogs were enrolled. Patients were allocated into three groups. Group 1 consisted of 30 dogs with a normal platelet count, group 2 included 22 dogs with a platelet count between 100 and 200×109/l and group 3 consisted of 20 dogs with a platelet count lower than 100×109/l. Platelet rich-plasma (PRP) was obtained from peripheral blood samples using tripotassium ethylenediaminetetraacetic acid (K3-EDTA) as anticoagulant. Next, platelets were stimulated using phorbol-12-myristate-13-acetate or thrombin, stabilized using procaine or left unstimulated. The expression of CD51 and CD41/CD61 was evaluated. Co-expression of CD41/CD61 and Annexin V served as a marker of platelet activation. The expression of CD41/CD61 and CD51 did not differ between the 3 groups. Thrombin-stimulated platelets had a significantly higher activity in dogs with a normal platelet count than in dogs with asymptomatic thrombocytopenia. Procaine inhibited platelet activity in all groups. In conclusion, activation of platelets of healthy dogs in vitro varied depending on the platelet count and platelet activator.

Chronic inflammation is a feature of Achilles tendinopathy and rupture.

PubMed

Dakin, Stephanie Georgina; Newton, Julia; Martinez, Fernando O; Hedley, Robert; Gwilym, Stephen; Jones, Natasha; Reid, Hamish A B; Wood, Simon; Wells, Graham; Appleton, Louise; Wheway, Kim; Watkins, Bridget; Carr, Andrew Jonathan

2018-03-01

Recent investigation of human tissue and cells from positional tendons such as the rotator cuff has clarified the importance of inflammation in the development and progression of tendon disease. These mechanisms remain poorly understood in disease of energy-storing tendons such as the Achilles. Using tissue biopsies from patients, we investigated if inflammation is a feature of Achilles tendinopathy and rupture. We studied Achilles tendon biopsies from symptomatic patients with either mid-portion tendinopathy or rupture for evidence of abnormal inflammatory signatures. Tendon-derived stromal cells from healthy hamstring and diseased Achilles were cultured to determine the effects of cytokine treatment on expression of inflammatory markers. Tendinopathic and ruptured Achilles highly expressed CD14+ and CD68+ cells and showed a complex inflammation signature, involving NF-κB, interferon and STAT-6 activation pathways. Interferon markers IRF1 and IRF5 were highly expressed in tendinopathic samples. Achilles ruptures showed increased PTGS2 and interleukin-8 expression. Tendinopathic and ruptured Achilles tissues expressed stromal fibroblast activation markers podoplanin and CD106. Tendon cells isolated from diseased Achilles showed increased expression of pro-inflammatory and stromal fibroblast activation markers after cytokine stimulation compared with healthy hamstring tendon cells. Tissue and cells derived from tendinopathic and ruptured Achilles tendons show evidence of chronic (non-resolving) inflammation. The energy-storing Achilles shares common cellular and molecular inflammatory mechanisms with functionally distinct rotator cuff positional tendons. Differences seen in the profile of ruptured Achilles are likely to be attributable to a superimposed phase of acute inflammation and neo-vascularisation. Strategies that target chronic inflammation are of potential therapeutic benefit for patients with Achilles tendon disease. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Chronic inflammation is a feature of Achilles tendinopathy and rupture

PubMed Central

Newton, Julia; Martinez, Fernando O; Hedley, Robert; Gwilym, Stephen; Jones, Natasha; Reid, Hamish A B; Wood, Simon; Wells, Graham; Appleton, Louise; Wheway, Kim; Watkins, Bridget; Carr, Andrew Jonathan

2018-01-01

Background Recent investigation of human tissue and cells from positional tendons such as the rotator cuff has clarified the importance of inflammation in the development and progression of tendon disease. These mechanisms remain poorly understood in disease of energy-storing tendons such as the Achilles. Using tissue biopsies from patients, we investigated if inflammation is a feature of Achilles tendinopathy and rupture. Methods We studied Achilles tendon biopsies from symptomatic patients with either mid-portion tendinopathy or rupture for evidence of abnormal inflammatory signatures. Tendon-derived stromal cells from healthy hamstring and diseased Achilles were cultured to determine the effects of cytokine treatment on expression of inflammatory markers. Results Tendinopathic and ruptured Achilles highly expressed CD14+ and CD68+ cells and showed a complex inflammation signature, involving NF-κB, interferon and STAT-6 activation pathways. Interferon markers IRF1 and IRF5 were highly expressed in tendinopathic samples. Achilles ruptures showed increased PTGS2 and interleukin-8 expression. Tendinopathic and ruptured Achilles tissues expressed stromal fibroblast activation markers podoplanin and CD106. Tendon cells isolated from diseased Achilles showed increased expression of pro-inflammatory and stromal fibroblast activation markers after cytokine stimulation compared with healthy hamstring tendon cells. Conclusions Tissue and cells derived from tendinopathic and ruptured Achilles tendons show evidence of chronic (non-resolving) inflammation. The energy-storing Achilles shares common cellular and molecular inflammatory mechanisms with functionally distinct rotator cuff positional tendons. Differences seen in the profile of ruptured Achilles are likely to be attributable to a superimposed phase of acute inflammation and neo-vascularisation. Strategies that target chronic inflammation are of potential therapeutic benefit for patients with Achilles tendon disease. PMID:29118051

Evaluation of pre- and posttransplantation serum interferon-gamma and soluble CD30 for predicting liver allograft rejection.

PubMed

Kim, K H; Oh, E-J; Jung, E-S; Park, Y-J; Choi, J Y; Kim, D-G; Lee, K Y; Kang, C S

2006-06-01

The aim of the present study was to identify whether the serum interferon-gamma (IFNgamma), a Th1 cytokine, or soluble CD30 (sCD30), a marker for activation of Th2 cytokine-producing T cells, predict acute cellular rejection episodes among liver graft patients. Pretransplant and posttransplant sera from 32 living donor liver transplant recipients obtained on days 1, 3, and 7 after surgery were tested for serum IFNgamma and sCD30 concentrations using commercial enzyme-linked immunosorbent assay kits. Recipients with an acute rejection episode (ARE) (n=14) displayed significantly higher IFNgamma concentrations pretransplant than did the patients with no ARE (n=18) (P<.05). The pretransplant serum levels of sCD30 were not different between the non-ARE and ARE groups. However, in comparison with the non-ARE group, who showed steadily decreasing serum sCD30 levels after transplantation, 12 among the 14 patients in the ARE group showed increasing sCD30 levels from day 1 to day 3 after transplantation (P<.05). These results suggest that the sCD30 increment during the early period after liver transplantation affects the immune response of rejection. This observation emphasizes the clinical relevance of serum sCD30, in addition to serum IFNgamma, as predictive markers for acute liver graft rejection.

TNFα and IFNγ but not perforin are critical for CD8 T cell-mediated protection against pulmonary Yersinia pestis infection.

PubMed

Szaba, Frank M; Kummer, Lawrence W; Duso, Debra K; Koroleva, Ekaterina P; Tumanov, Alexei V; Cooper, Andrea M; Bliska, James B; Smiley, Stephen T; Lin, Jr-Shiuan

2014-05-01

Septic pneumonias resulting from bacterial infections of the lung are a leading cause of human death worldwide. Little is known about the capacity of CD8 T cell-mediated immunity to combat these infections and the types of effector functions that may be most effective. Pneumonic plague is an acutely lethal septic pneumonia caused by the Gram-negative bacterium Yersinia pestis. We recently identified a dominant and protective Y. pestis antigen, YopE69-77, recognized by CD8 T cells in C57BL/6 mice. Here, we use gene-deficient mice, Ab-mediated depletion, cell transfers, and bone marrow chimeric mice to investigate the effector functions of YopE69-77-specific CD8 T cells and their relative contributions during pulmonary Y. pestis infection. We demonstrate that YopE69-77-specific CD8 T cells exhibit perforin-dependent cytotoxicity in vivo; however, perforin is dispensable for YopE69-77-mediated protection. In contrast, YopE69-77-mediated protection is severely impaired when production of TNFα and IFNγ by CD8 T cells is simultaneously ablated. Interestingly, TNFα is absolutely required at the time of challenge infection and can be provided by either T cells or non-T cells, whereas IFNγ provided by T cells prior to challenge appears to facilitate the differentiation of optimally protective CD8 T cells. We conclude that cytokine production, not cytotoxicity, is essential for CD8 T cell-mediated control of pulmonary Y. pestis infection and we suggest that assays detecting Ag-specific TNFα production in addition to antibody titers may be useful correlates of vaccine efficacy against plague and other acutely lethal septic bacterial pneumonias.

Posttransplant sCD30 as a predictor of kidney graft outcome.

PubMed

Süsal, Caner; Döhler, Bernd; Sadeghi, Mahmoud; Salmela, Kaija T; Weimer, Rolf; Zeier, Martin; Opelz, Gerhard

2011-06-27

Reliable markers for assessing the biological effect of immunosuppressive drugs and identification of transplant recipients at risk of developing rejection are not available. In a prospective multicenter study, we investigated whether posttransplant measurement of the T-cell activation marker soluble CD30 (sCD30) can be used for estimating the risk of graft loss in kidney transplant recipients. Pre- and posttransplant sera of 2322 adult deceased-donor kidney recipients were tested for serum sCD30 content using a commercial enzyme-linked immunosorbent assay. sCD30 decreased posttransplant and reached a nadir on day 30. Patients with a high sCD30 of more than or equal to 40 U/mL on day 30 showed a subsequent graft survival rate after 3 years of 78.3±4.1%, significantly lower than the 90.3±1.0% rate in recipients with a low sCD30 on day 30 of less than 40 U/mL (log-rank P<0.001; Cox hazard ratio 2.02, P<0.001). Although an association was found between pre- and posttransplant sCD30 levels, patients with high sCD30 on posttransplant day 30 demonstrated significantly lower 3-year graft survival irrespective of the pretransplant level. Our data suggest that posttransplant measurement of sCD30 on day 30 is a predictor of subsequent graft loss in kidney transplant recipients and that sCD30 may potentially serve as an indicator for adjustment of immunosuppressive medication.

The PBX1 lupus susceptibility gene regulates CD44 expression.

PubMed

Niu, Yuxin; Sengupta, Mayami; Titov, Anton A; Choi, Seung-Chul; Morel, Laurence

2017-05-01

PBX1-d is novel splice isoform of pre-B-cell leukemia homeobox 1 (PBX1) that lacks its DNA-binding and Hox-binding domains, and functions as a dominant negative. We have shown that PBX1-d expression in CD4 + T cells is associated with systemic lupus erythematosus (SLE) in a mouse model as well as in human subjects. More specifically, PBX1-d expression leads to the production of autoreactive activated CD4+ T cells, a reduced frequency and function of Foxp3+ regulatory T (Treg) cells and an expansion of follicular helper T (Tfh) cells. Very little is known about the function of PBX1 in T cells, except that it directly regulates the expression of miRNAs associated with Treg and Tfh homeostasis. In the present study, we show that PBX1 directly regulated the expression of CD44, a marker of T cell activation. Two PBX1 binding sites in the promoter directly regulated CD44 expression, with PBX1-d driving a higher expression than the normal isoform PBX1-b. In addition, mutations in each of the two binding sites had different effects of PBX1-b and PBX1-d. Finally, we showed that an enhanced recruitment of co-factor MEIS by PBX1-d over PBX1-b, while there was no difference for co-factor PREP1 recruitment. Therefore, this study demonstrates that the lupus-associated PBX1-d isoform directly transactivates CD44, a marker of CD44 activation and memory, and that it has different DNA binding and co-factor recruitment relative to the normal isoform. Taken together, these results confirm that PBX1 directly regulates genes related to T cell activation and shows that the lupus-associated isoform PBX1-d has unique molecular functions. Copyright © 2017 Elsevier Ltd. All rights reserved.

A novel vaccination strategy mediating the induction of lung-resident memory CD8 T cells confers heterosubtypic immunity against future pandemic influenza virus

PubMed Central

Lee, Yu-Na; Lee, Young-Tae; Kim, Min-Chul; Gewirtz, Andrew T.; Kang, Sang-Moo

2016-01-01

The currently used vaccine strategy to combat influenza A virus (IAV) aims to provide highly specific immunity to circulating seasonal IAV strains. However, the outbreak of 2009 influenza pandemic highlights the danger in this strategy. Here, we tested the hypothesis that universal vaccination that offers broader but weaker protection would result in cross protective T-cell responses after primary IAV infection, which would subsequently provide protective immunity against future pandemic strains. Specifically, we used tandem repeat M2e epitopes on virus-like particles (M2e5x VLP) that induced heterosubtypic immunity by eliciting antibodies to a conserved M2e epitope. M2e5x VLP was found to be superior to strain-specific current split vaccine in conferring heterosubtypic cross protection and in equipping the host with cross-protective lung-resident nucleoprotein-specific memory CD8+ T cell responses to a subsequent secondary infection with a new pandemic potential strain. Immune correlates for subsequent heterosubtypic immunity by M2e5x VLP vaccination were found to be virus-specific CD8+ T cells secreting IFN-γ and expressing lung-resident memory phenotypic markers CD69+ and CD103+ as well as M2e antibodies. Hence, vaccination with M2e5x VLP may be developable as a new strategy to combat future pandemic outbreaks. PMID:26864033

The CD11a and Endothelial Protein C Receptor Marker Combination Simplifies and Improves the Purification of Mouse Hematopoietic Stem Cells

PubMed Central

Karimzadeh, Alborz; Scarfone, Vanessa M.; Varady, Erika; Chao, Connie; Grathwohl, Karin; Fathman, John W.; Fruman, David A.; Serwold, Thomas

2018-01-01

Abstract Hematopoietic stem cells (HSCs) are the self‐renewing multipotent progenitors to all blood cell types. Identification and isolation of HSCs for study has depended on the expression of combinations of surface markers on HSCs that reliably distinguish them from other cell types. However, the increasing number of markers required to isolate HSCs has made it tedious, expensive, and difficult for newcomers, suggesting the need for a simpler panel of HSC markers. We previously showed that phenotypic HSCs could be separated based on expression of CD11a and that only the CD11a negative fraction contained true HSCs. Here, we show that CD11a and another HSC marker, endothelial protein C receptor (EPCR), can be used to effectively identify and purify HSCs. We introduce a new two‐color HSC sorting method that can highly enrich for HSCs with efficiencies comparable to the gold standard combination of CD150 and CD48. Our results demonstrate that adding CD11a and EPCR to the HSC biologist's toolkit improves the purity of and simplifies isolation of HSCs. stem cells translational medicine 2018;7:468–476 PMID:29543389

Decrease in T Cell Activation and Calcium Flux during Clinorotation

NASA Technical Reports Server (NTRS)

Sams, Clarence; Holtzclaw, J. David

2006-01-01

We investigated the effect of altered gravitational environments on T cell activation. We isolated human, naive T cells (CD3+CD14-CD19-CD16-CD56-CD25-CD69-CD45RA-) following IRB approved protocols. These purified T cells were then incubated with 6 mm polystyrene beads coated with OKT3 (Ortho Biotech, Raritan, NJ) and antiCD28 (Becton Dickinson (BD), San Jose, CA) at 37 C for 24 hours. Antibodies were at a 1:1 ratio and the bead-to-cell ratio was 2:1. Four incubation conditions existed: 1) static or "1g"; 2) centrifugation at 10 relative centrifugal force (RCF) or "10g"; 3) clinorotation at 25 RPM (functional weightlessness or "0g"); and 4) clinorotation at 80 RPM ("1g" plus net shear force approx.30 dynes/sq cm). Following incubation, T cells were stained for CD25 expression (BD) and intracellular calcium (ratio of Fluo4 to Fura Red, Molecular Probes, Eugene, OR) and analyzed by flow cytometry (Coulter EPICS XL, Miami, FL). Results: Static or "1g" T cells had the highest level of CD25 expression and intracellular calcium. T cells centrifuged at 10 RCF ("10g") had lower CD25 expression and calcium levels compared to the static control. However, cells centrifuged at 10 RCF had higher CD25 expression and calcium levels than those exposed to 24 RPM clinorotation ("0g"). T cells exposed to 24 RPM clinorotation had lower CD25 expression, but the approximately the same calcium levels than T cells exposed to 80 RPM clinorotation. These data suggest that stress-activated calcium channel exist in T cells and may play a role during T cell activation.

Gamma-glutamyltransferase activity in exosomes as a potential marker for prostate cancer.

PubMed

Kawakami, Kyojiro; Fujita, Yasunori; Matsuda, Yoko; Arai, Tomio; Horie, Kengo; Kameyama, Koji; Kato, Taku; Masunaga, Koichi; Kasuya, Yutaka; Tanaka, Masashi; Mizutani, Kosuke; Deguchi, Takashi; Ito, Masafumi

2017-05-05

Exosomes or extracellular vesicles have the potential as a diagnostic marker for various diseases including cancer. In order to identify novel exosomal markers for prostate cancer (PC), we performed proteomic analysis of exosomes isolated from PC cell lines and examined the usefulness of the marker in patients. Exosomes isolated by differential centrifugation from the culture medium of androgen-dependent LNCaP prostate cancer cell line and its sublines of partially androgen-independent C4, androgen-independent C4-2 and bone metastatic C4-2B were subjected to iTRAQ-based proteomic analysis. Exosomes were also isolated by immunocapture and separated by size exclusion chromatography and density gradient centrifugation. Protein expression was determined by Western blot analysis. GGT activity was measured using a fluorescent probe, γ-glutamyl hydroxymethyl rhodamine green (gGlu-HMRG). Immunohistochemical analysis of tissues was performed using anti-GGT1 antibody. Among proteins upregulated in C4-2 and C4-2B cells than in LNCaP cells, we focused on gamma-glutamyltransferase 1 (GGT1), a cell-surface enzyme that regulates the catabolism of extracellular glutathione. The levels of both GGT1 large and small subunits were elevated in exosomes isolated from C4-2 and C4-2B cells by differential centrifugation and by immunocapture with anti-CD9 or -prostate-specific membrane antigen (PSMA) antibody. In cell lysates and exosomes, GGT1 expression correlated with GGT activity. Size exclusion chromatography of human serum demonstrated the presence of GGT activity and GGT1 subunits in fractions positive for CD9. Density gradient centrifugation revealed the co-presence of GGT1 subunits with CD9 in exosomes isolated by differential centrifugation from human serum. Since GGT activity correlated with GGT1 expression in serum exosomes isolated by differential centrifugation, we measured serum exosomal GGT activity in patients. Unexpectedly, we found that serum exosomal GGT activity was significantly higher in PC patients than in benign prostatic hyperplasia (BPH) patients. In support of this finding, immunohistochemical analysis showed increased GGT1 expression in PC tissues compared with BPH tissues. Our results suggest that serum exosomal GGT activity could be a useful biomarker for PC.

CXCR6 marks a novel subset of T-betloEomeshi natural killer cells residing in human liver

PubMed Central

Stegmann, Kerstin A.; Robertson, Francis; Hansi, Navjyot; Gill, Upkar; Pallant, Celeste; Christophides, Theodoros; Pallett, Laura J.; Peppa, Dimitra; Dunn, Claire; Fusai, Giuseppe; Male, Victoria; Davidson, Brian R.; Kennedy, Patrick; Maini, Mala K.

2016-01-01

Natural killer cells (NK) are highly enriched in the human liver, where they can regulate immunity and immunopathology. We probed them for a liver-resident subset, distinct from conventional bone-marrow-derived NK. CXCR6+ NK were strikingly enriched in healthy and diseased liver compared to blood (p < 0.0001). Human hepatic CXCR6+ NK had an immature phenotype (predominantly CD56brightCD16−CD57−), and expressed the tissue-residency marker CD69. CXCR6+ NK produced fewer cytotoxic mediators and pro-inflammatory cytokines than the non-liver-specific CXCR6− fraction. Instead CXCR6+ NK could upregulate TRAIL, a key death ligand in hepatitis pathogenesis. CXCR6 demarcated liver NK into two transcriptionally distinct populations: T-bethiEomeslo(CXCR6−) and T-betloEomeshi(CXCR6+); the latter was virtually absent in the periphery. The small circulating CXCR6+ subset was predominantly T-bethiEomeslo, suggesting its lineage was closer to CXCR6− peripheral than CXCR6+ liver NK. These data reveal a large subset of human liver-resident T-betloEomeshi NK, distinguished by their surface expression of CXCR6, adapted for hepatic tolerance and inducible anti-viral immunity. PMID:27210614

CXCR6 marks a novel subset of T-bet(lo)Eomes(hi) natural killer cells residing in human liver.

PubMed

Stegmann, Kerstin A; Robertson, Francis; Hansi, Navjyot; Gill, Upkar; Pallant, Celeste; Christophides, Theodoros; Pallett, Laura J; Peppa, Dimitra; Dunn, Claire; Fusai, Giuseppe; Male, Victoria; Davidson, Brian R; Kennedy, Patrick; Maini, Mala K

2016-05-23

Natural killer cells (NK) are highly enriched in the human liver, where they can regulate immunity and immunopathology. We probed them for a liver-resident subset, distinct from conventional bone-marrow-derived NK. CXCR6+ NK were strikingly enriched in healthy and diseased liver compared to blood (p < 0.0001). Human hepatic CXCR6+ NK had an immature phenotype (predominantly CD56(bright)CD16-CD57-), and expressed the tissue-residency marker CD69. CXCR6+ NK produced fewer cytotoxic mediators and pro-inflammatory cytokines than the non-liver-specific CXCR6- fraction. Instead CXCR6+ NK could upregulate TRAIL, a key death ligand in hepatitis pathogenesis. CXCR6 demarcated liver NK into two transcriptionally distinct populations: T-bet(hi)Eomes(lo)(CXCR6-) and T-bet(lo)Eomes(hi)(CXCR6+); the latter was virtually absent in the periphery. The small circulating CXCR6+ subset was predominantly T-bet(hi)Eomes(lo), suggesting its lineage was closer to CXCR6- peripheral than CXCR6+ liver NK. These data reveal a large subset of human liver-resident T-bet(lo)Eomes(hi) NK, distinguished by their surface expression of CXCR6, adapted for hepatic tolerance and inducible anti-viral immunity.

Skeletal myogenic differentiation of human urine-derived cells as a potential source for skeletal muscle regeneration.

PubMed

Chen, Wei; Xie, Minkai; Yang, Bin; Bharadwaj, Shantaram; Song, Lujie; Liu, Guihua; Yi, Shanhong; Ye, Gang; Atala, Anthony; Zhang, Yuanyuan

2017-02-01

Stem cells are regarded as possible cell therapy candidates for skeletal muscle regeneration. However, invasive harvesting of those cells can cause potential harvest-site morbidity. The goal of this study was to assess whether human urine-derived stem cells (USCs), obtained through non-invasive procedures, can differentiate into skeletal muscle linage cells (Sk-MCs) and potentially be used for skeletal muscle regeneration. In this study, USCs were harvested from six healthy individuals aged 25-55. Expression profiles of cell-surface markers were assessed by flow cytometry. To optimize the myogenic differentiation medium, we selected two from four different types of myogenic differentiation media to induce the USCs. Differentiated USCs were identified with myogenic markers by gene and protein expression. USCs were implanted into the tibialis anterior muscles of nude mice for 1 month. The results showed that USCs displayed surface markers with positive staining for CD24, CD29, CD44, CD73, CD90, CD105, CD117, CD133, CD146, SSEA-4 and STRO-1, and negative staining for CD14, CD31, CD34 and CD45. After myogenic differentiation, a change in morphology was observed from 'rice-grain'-like cells to spindle-shaped cells. The USCs expressed specific Sk-MC transcripts and protein markers (myf5, myoD, myosin, and desmin) after being induced with different myogenic culture media. Implanted cells expressed Sk-MC markers stably in vivo. Our findings suggest that USCs are able to differentiate into the Sk-MC lineage in vitro and after being implanted in vivo. Thus, they might be a potential source for cell injection therapy in the use of skeletal muscle regeneration. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

Prognostic value of CD44 expression in penile squamous cell carcinoma: a pilot study.

PubMed

Minardi, Daniele; Lucarini, Guendalina; Filosa, Alessandra; Zizzi, Antonio; Simonetti, Oriana; Offidani, Anna Maria; d'Anzeo, Gianluca; Di Primio, Roberto; Montironi, Rodolfo; Muzzonigro, Giovanni

2012-10-01

Several studies have reported on the prognostic value of molecular markers for metastasis risk and survival in penile squamous cell carcinoma (SCC) patients. The usefulness of CD44 expression as such a marker has been studied in different tumors, but not in penile SCC. Our aim was to determine whether CD44 expression may serve as a prognostic marker for lymph node metastasis and survival in penile SCC patients. CD44 immunoistochemical expression was investigated in tissue specimens from 39 patients with penile SCC. CD44 cell positivity, staining intensity and distribution were analyzed and correlated with tumor stage, grade, lymph node status and disease-specific survival. CD44 expression was detected in epithelial cells of both intratumoral and normal tissues with different intensities and staining distributions. In normal tissues CD44 protein was mainly detected in cell membranes, whereas in the tumor compartments it was found in both the cell membranes and the cytoplasm. The intensities and percentages of CD44 expressing cells did not correlate with tumor stage and/or grade. Seventy-three percent of the patients with lymph node metastasis showed high intensities of CD44 staining, as compared to 44% of the patients without lymph node metastasis (P = 0.03). Lymph node-positive patients showed both cytoplasmic and membranous CD44 expression. High CD44 expression was found to be significantly correlated with a decreased 5 year overall survival (P = 0.01). CD44 levels and patterns of expression can be considered as markers for penile SCC aggressiveness and, in addition, may serve as predictive markers for lymph node metastasis, also in patients with clinically negative lymph nodes. CD44 expression may provide prognostic information for penile SCC patients, next to classical clinical-pathological factors.

In vivo immunotoxicity evaluation of Gd2O3 nanoprobes prepared by laser ablation in liquid for MRI preclinical applications

NASA Astrophysics Data System (ADS)

Tian, Xiumei; Guan, Xiaoying; Luo, Ningqi; Yang, Fanwen; Chen, Dihu; Peng, Ye; Zhu, Jixiang; He, Fupo; Li, Li; Chen, Xiaoming

2014-09-01

Gd2O3 nanoprobes prepared by laser ablation in liquid can be used as magnetic resonance imaging contrast agent. However, their immunotoxicity in vivo remains unknown. In this article, the in vitro biocompatibility of the Gd2O3 nanoprobe was evaluated in terms of cell uptake, cell viability, and apoptosis. In vivo immunotoxicity was detected by monitoring the levels of the immunity mediator, cluster of differentiation (CD) markers in Balb/c mice. The results show that no in vitro cytotoxicity was observed, and no significant changes in the expression levels of CD206 and CD69 between the nanoprobe-injected group and the Gd-DTPA group in mice were observed. Importantly, the immunotoxicity data revealed significant differences in the expression levels of CD40, CD80, CD11b, and reactive oxygen species. In addition, transmission electron microscopy images showed that few Gd2O3 nanoprobes were localized in phagosomes by the endocytic pathway. In conclusion, the toxic effects of our Gd2O3 nanoprobe may be due to endocytosis during which the microstructure or ultrastructure of cells is slightly damaged and induces the generation of an oxidative stress reaction that further stimulates the innate immune response. Therefore, it is important to use a sensitive assay for the in vivo immunotoxicity measurements to evaluate the risk assessment of Gd2O3-based biomaterials at the molecular level.

Activated effector and memory T cells contribute to circulating sCD30: potential marker for islet allograft rejection.

PubMed

Saini, D; Ramachandran, S; Nataraju, A; Benshoff, N; Liu, W; Desai, N; Chapman, W; Mohanakumar, T

2008-09-01

T-cell activation up-regulates CD30 resulting in an increase in serum soluble CD30 (sCD30). CD4+ T cells, a major source for sCD30, play a significant role in the pathogenesis of rejection. In this study, sCD30 was measured pre- and posttransplant in mouse islet allograft models and human islet allograft recipients. sCD30 was measured by ELISA in diabetic C57BL/6, CD4Knockout (KO) and CD8KO islet allograft recipients. sCD30 increased significantly prior to rejection (1.8 +/- 1 days) in 80% of allograft recipients. Sensitization with donor splenocytes, or a second graft, further increased sCD30 (282.5 +/- 53.5 for the rejecting first graft vs. 374.6 +/- 129 for the rejecting second graft) prior to rejection suggesting memory CD4+ T cells contribute to sCD30. CD4KO failed to reject islet allograft and did not demonstrate sCD30 increase. CD8KO showed elevated (227 +/- 107) sCD30 (1 day) prior to rejection. High pretransplant sCD30 (>20 U/ml) correlated with poor outcome in human islet allograft recipients. Further, increase in sCD30 posttransplant preceded (3-4 months) loss of islet function. We conclude that sCD30 is released from activated CD4 T cells prior to islet allograft rejection and monitoring sCD30 can be a valuable adjunct in the follow-up of islet transplant recipients.

Markers of apoptosis and proliferation related gene products as predictors of treatment outcome in childhood acute lymphoblastic leukemia.

PubMed

Hafez, Mohammad; Al-Tonbary, Youssef; El-Bayoumi, Mohammed A; Hatem, Nadia; Hawas, Samia; Mansour, Ahmed; Marzouk, Iman; Hafez, Mona M; Yahia, Sohier; Farahat, Nahla

2007-06-01

The aim of the study is to characterize markers of apoptosis in children with acute lymphoblastic leukemia (ALL) in relation to treatment outcome of the disease. The study was performed on 34 children with ALL and 39 healthy children as a control group. Apoptosis was assessed by cell morphology; DNA fragmentation; ELISA and RT-PCR for CD95, CD95L, BcL-2 and nuclear factor-kappa B (NF-kappaB); and flow cytometry for CD95, CD40, CD49d and CD11a. Apoptosis was significantly lower in patients than controls. Apoptosis detected by CD95 ligand was significantly lower in cases with no remission after treatment than those who achieved remission. Anti-apoptotic factors: CD40, BcL-2, and NF-kappaB were all found to be higher in cases than controls and in cases with no remission than those achieved remission. CD49d was significantly lower in cases than controls, and significantly lower in cases with who did not achieve remission. CD11a levels were similar in the various groups. Delayed apoptosis of ALL cells is genetically controlled either directly or indirectly by a network of oncogenes and tumor suppressor genes. CD40 appeared to stimulate both T and B lineage and is considered the most potent influencer and predictor of resistance to therapy. Inhibitors for the activity of CD40, Bcl-2 and NF-kappaB as well as stimulants to CD95 could have a potential therapeutic benefit.

Soluble CD30 is more relevant to disease activity of atopic dermatitis than soluble CD26

PubMed Central

Katoh, N; Hirano, S; Suehiro, M; Ikenaga, K; Yamashita, T; Sugawara, N; Yasuno, H

2000-01-01

It is suggested that CD30 and CD26 are surface molecules expressed on activated Th2 and Th1 cells, respectively. We examined plasma levels of soluble CD26 (sCD26) and sCD30 in patients with atopic dermatitis (AD) when their eruptions were aggravated and in non-atopic healthy controls, and then analysed the possible correlation between these values and the levels of several clinical markers. The plasma levels of both sCD30 and sCD26 were significantly higher in AD patients than in controls, both in exacerbation status and after conventional treatment. Multiple regression analyses showed that plasma sCD30 was a much better predictor of the levels of serum IgE, serum LDH and plasma sCD25, and the area and the score of AD eruption than sCD26, although elevated levels of both sCD30 and sCD26 are associated with these clinical predictors of AD. Importantly, sCD30 plasma levels decreased significantly in AD patients after conventional treatment, while no significant transition was noted in the concentration of sCD26. Moreover, a significant reduction of sCD30 levels was observed in the group of patients whose eruption score was reduced > 50%, whereas it was not in those < 50%. These findings provide evidence that the successful treatment of AD is associated with down-activation of Th2. PMID:10931130

Soluble CD163 is an indicator of liver inflammation and fibrosis in patients chronically infected with the hepatitis B virus.

PubMed

Dultz, G; Gerber, L; Farnik, H; Berger, A; Vermehren, J; Pleli, T; Zeuzem, S; Piiper, A; Kronenberger, B; Waidmann, O

2015-04-01

Soluble CD163 (sCD163), a marker for macrophage activation, was found to be associated with the severity of liver cirrhosis. The aim of the current study was to investigate whether serum sCD163 levels correlate with liver inflammation and fibrosis in patients with chronic hepatitis B virus (HBV) infection. In a retrospective cohort study, serum sCD163 levels were assessed by ELISA together with clinical and laboratory data in 186 patients with chronic HBV infection and 15 healthy controls. The relation between parameters for liver fibrosis and necroinflammation and sCD163 levels was analysed. Additionally, sCD163 was quantified in a subset of follow-up serum samples after initiation of antiviral treatment. sCD163 levels differed among phases of chronic HBV infection (P < 0.0001), and sCD163 concentrations were associated with inflammatory activity and fibrosis in the liver. sCD163 levels ≥ 1961 ng/l had a high specificity in the identification of subjects with substantial fibrosis (F ≥ 2). sCD163 concentrations decreased significantly after initiation of antiviral treatment. The correlation of sCD163 levels with necroinflammation and fibrosis and the sCD163 decline under treatment indicates that macrophage activation plays a role in HBV-related liver pathogenesis. © 2014 John Wiley & Sons Ltd.

The Low-Cost Compound Lignosulfonic Acid (LA) Exhibits Broad-Spectrum Anti-HIV and Anti-HSV Activity and Has Potential for Microbicidal Applications

PubMed Central

D’huys, Thomas; Petrova, Mariya I.; Lebeer, Sarah; Snoeck, Robert; Andrei, Graciela; Schols, Dominique

2015-01-01

Objectives Lignosulfonic acid (LA), a low-cost lignin-derived polyanionic macromolecule, was extensively studied for its anti-HIV and anti-HSV activity in various cellular assays, its mechanism of viral inhibition and safety profile as potential microbicide. Results LA demonstrated potent inhibitory activity of HIV replication against a wide range of R5 and X4 HIV strains and prevented the uptake of HIV by bystander CD4+ T cells from persistently infected T cells in vitro (IC50: 0.07 – 0.34 μM). LA also inhibited HSV-2 replication in vitro in different cell types (IC50: 0.42 – 1.1 μM) and in rodents in vivo. Furthermore, LA neutralized the HIV-1 and HSV-2 DC-SIGN-mediated viral transfer to CD4+ T cells (IC50: ∼1 μM). In addition, dual HIV-1/HSV-2 infection in T cells was potently blocked by LA (IC50: 0.71 μM). No antiviral activity was observed against the non-enveloped viruses Coxsackie type B4 and Reovirus type 1. LA is defined as a HIV entry inhibitor since it interfered with gp120 binding to the cell surface of T cells. Pretreatment of PBMCs with LA neither increased expression levels of cellular activation markers (CD69, CD25 and HLA-DR), nor enhanced HIV-1 replication. Furthermore, we found that LA had non-antagonistic effects with acyclovir, PRO2000 or LabyA1 (combination index (CI): 0.46 – 1.03) in its anti-HSV-2 activity and synergized with tenofovir (CI: 0.59) in its anti-HIV-1 activity. To identify mechanisms of LA resistance, we generated in vitro a mutant HIV-1 NL4.3LAresistant virus, which acquired seven mutations in the HIV-1 envelope glycoproteins: S160N, V170N, Q280H and R389T in gp120 and K77Q, N113D and H132Y in gp41. Additionally, HIV-1 NL4.3LAresistant virus showed cross-resistance with feglymycin, enfuvirtide, PRO2000 and mAb b12, four well-described HIV binding/fusion inhibitors. Importantly, LA did not affect the growth of vaginal Lactobacilli strains. Conclusion Overall, these data highlight LA as a potential and unique low-cost microbicide displaying broad anti-HIV and anti-HSV activity. PMID:26132818

Epithelial-mesenchymal transition in breast epithelial cells treated with cadmium and the role of Snail.

PubMed

Wei, Zhengxi; Shan, Zhongguo; Shaikh, Zahir A

2018-04-01

Epidemiological and experimental studies have implicated cadmium (Cd) with breast cancer. In breast epithelial MCF10A and MDA-MB-231 cells, Cd has been shown to promote cell growth. The present study examined whether Cd also promotes epithelial-mesenchymal transition (EMT), a hallmark of cancer progression. Human breast epithelial cells consisting of non-cancerous MCF10A, non-metastatic HCC 1937 and HCC 38, and metastatic MDA-MB-231 were treated with 1 or 3 μM Cd for 4 weeks. The MCF10A epithelial cells switched to a more mesenchymal-like morphology, which was accompanied by a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal markers N-cadherin and vimentin. In both non-metastatic HCC 1937 and HCC 38 cells, treatment with Cd decreased the epithelial marker claudin-1. In addition, E-cadherin also decreased in the HCC 1937 cells. Even the mesenchymal-like MDA-MB-231 cells exhibited an increase in the mesenchymal marker vimentin. These changes indicated that prolonged treatment with Cd resulted in EMT in both normal and cancer-derived breast epithelial cells. Furthermore, both the MCF10A and MDA-MB-231 cells labeled with Zcad, a dual sensor for tracking EMT, demonstrated a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal marker ZEB-1. Treatment of cells with Cd significantly increased the level of Snail, a transcription factor involved in the regulation of EMT. However, the Cd-induced Snail expression was completely abolished by actinomycin D. Luciferase reporter assay indicated that the expression of Snail was regulated by Cd at the promotor level. Snail was essential for Cd-induced promotion of EMT in the MDA-MB-231 cells, as knockdown of Snail expression blocked Cd-induced cell migration. Together, these results indicate that Cd promotes EMT in breast epithelial cells and does so by modulating the transcription of Snail. Copyright © 2018 Elsevier Inc. All rights reserved.

Intragraft expression of the IL-10 gene is up-regulated in renal protocol biopsies with early interstitial fibrosis, tubular atrophy, and subclinical rejection.

PubMed

Hueso, Miguel; Navarro, Estanis; Moreso, Francesc; O'Valle, Francisco; Pérez-Riba, Mercè; Del Moral, Raimundo García; Grinyó, Josep M; Serón, Daniel

2010-04-01

Grafts with subclinical rejection associated with interstitial fibrosis and tubular atrophy (SCR+IF/TA) show poorer survival than grafts with subclinical rejection without IF/TA (SCR). Aiming to detect differences among SCR+IF/TA and SCR, we immunophenotyped the inflammatory infiltrate (CD45, CD3, CD20, CD68) and used a low-density array to determine levels of T(H)1 (interleukin IL-2, IL-3, gamma-interferon, tumor necrosis factor-alpha, lymphotoxin-alpha, lymphotoxin-beta, granulocyte-macrophage colony-stimulating factor) and T(H)2 (IL-4, IL-5, IL-6, IL-10, and IL-13) transcripts as well as of IL-2R (as marker for T-cell activation) in 31 protocol biopsies of renal allografts. Here we show that grafts with early IF/TA and SCR can be distinguished from grafts with SCR on the basis of the activation of IL-10 gene expression and of an increased infiltration by B-lymphocytes in a cellular context in which the degree of T-cell activation is similar in both groups of biopsies, as demonstrated by equivalent levels of IL-2R mRNA. These results suggest that the up-regulation of the IL-10 gene expression, as well as an increased proportion of B-lymphocytes in the inflammatory infiltrates, might be useful as markers of early chronic lesions in grafts with SCR.

Intragraft Expression of the IL-10 Gene Is Up-Regulated in Renal Protocol Biopsies with Early Interstitial Fibrosis, Tubular Atrophy, and Subclinical Rejection

PubMed Central

Hueso, Miguel; Navarro, Estanis; Moreso, Francesc; O'Valle, Francisco; Pérez-Riba, Mercè; del Moral, Raimundo García; Grinyó, Josep M.; Serón, Daniel

2010-01-01

Grafts with subclinical rejection associated with interstitial fibrosis and tubular atrophy (SCR+IF/TA) show poorer survival than grafts with subclinical rejection without IF/TA (SCR). Aiming to detect differences among SCR+IF/TA and SCR, we immunophenotyped the inflammatory infiltrate (CD45, CD3, CD20, CD68) and used a low-density array to determine levels of TH1 (interleukin IL-2, IL-3, γ-interferon, tumor necrosis factor-α, lymphotoxin-α, lymphotoxin-β, granulocyte-macrophage colony-stimulating factor) and TH2 (IL-4, IL-5, IL-6, IL-10, and IL-13) transcripts as well as of IL-2R (as marker for T-cell activation) in 31 protocol biopsies of renal allografts. Here we show that grafts with early IF/TA and SCR can be distinguished from grafts with SCR on the basis of the activation of IL-10 gene expression and of an increased infiltration by B-lymphocytes in a cellular context in which the degree of T-cell activation is similar in both groups of biopsies, as demonstrated by equivalent levels of IL-2R mRNA. These results suggest that the up-regulation of the IL-10 gene expression, as well as an increased proportion of B-lymphocytes in the inflammatory infiltrates, might be useful as markers of early chronic lesions in grafts with SCR. PMID:20150436

Enhancing the Oncolytic Activity of CD133-Targeted Measles Virus: Receptor Extension or Chimerism with Vesicular Stomatitis Virus Are Most Effective

PubMed Central

Kleinlützum, Dina; Hanauer, Julia D. S.; Muik, Alexander; Hanschmann, Kay-Martin; Kays, Sarah-Katharina; Ayala-Breton, Camilo; Peng, Kah-Whye; Mühlebach, Michael D.; Abel, Tobias; Buchholz, Christian J.

2017-01-01

Therapy resistance and tumor recurrence are often linked to a small refractory and highly tumorigenic subpopulation of neoplastic cells, known as cancer stem cells (CSCs). A putative marker of CSCs is CD133 (prominin-1). We have previously described a CD133-targeted oncolytic measles virus (MV-CD133) as a promising approach to specifically eliminate CD133-positive tumor cells. Selectivity was introduced at the level of cell entry by an engineered MV hemagglutinin (H). The H protein was blinded for its native receptors and displayed a CD133-specific single-chain antibody fragment (scFv) as targeting domain. Interestingly, MV-CD133 was more active in killing CD133-positive tumors than the unmodified MV-NSe despite being highly selective for its target cells. To further enhance the antitumoral activity of MV-CD133, we here pursued arming technologies, receptor extension, and chimeras between MV-CD133 and vesicular stomatitis virus (VSV). All newly generated viruses including VSV-CD133 were highly selective in eliminating CD133-positive cells. MV-CD46/CD133 killed in addition CD133-negative cells being positive for the MV receptors. In an orthotopic glioma model, MV-CD46/CD133 and MVSCD-CD133, which encodes the super cytosine deaminase, were most effective. Notably, VSV-CD133 caused fatal neurotoxicity in this tumor model. Use of CD133 as receptor could be excluded as being causative. In a subcutaneous tumor model of hepatocellular cancer, VSV-CD133 revealed the most potent oncolytic activity and also significantly prolonged survival of the mice when injected intravenously. Compared to MV-CD133, VSV-CD133 infected a more than 104-fold larger area of the tumor within the same time period. Our data not only suggest new concepts and approaches toward enhancing the oncolytic activity of CD133-targeted oncolytic viruses but also raise awareness about careful toxicity testing of novel virus types. PMID:28695108

Reevaluation of immune activation in the era of cART and an aging HIV-infected population

PubMed Central

de Armas, Lesley R.; Pallikkuth, Suresh; George, Varghese; Rinaldi, Stefano; Pahwa, Rajendra; Arheart, Kristopher L.

2017-01-01

Biological aging is associated with immune activation (IA) and declining immunity due to systemic inflammation. It is widely accepted that HIV infection causes persistent IA and premature immune senescence despite effective antiretroviral therapy and virologic suppression; however, the effects of combined HIV infection and aging are not well defined. Here, we assessed the relationship between markers of IA and inflammation during biological aging in HIV-infected and -uninfected populations. Antibody response to seasonal influenza vaccination was implemented as a measure of immune competence and relationships between IA, inflammation, and antibody responses were explored using statistical modeling appropriate for integrating high-dimensional data sets. Our results show that markers of IA, such as coexpression of HLA antigen D related (HLA-DR) and CD38 on CD4+ T cells, exhibit strong associations with HIV infection but not with biological age. Certain variables that showed a strong relationship with aging, such as declining naive and CD38+ CD4 and CD8+ T cells, did so regardless of HIV infection. Interestingly, the variable of biological age was not identified in a predictive model as significantly impacting vaccine responses in either group, while distinct IA and inflammatory variables were closely associated with vaccine response in HIV-infected and -uninfected populations. These findings shed light on the most relevant and persistent immune defects during virological suppression with antiretroviral therapy. PMID:29046481

Reevaluation of immune activation in the era of cART and an aging HIV-infected population.

PubMed

de Armas, Lesley R; Pallikkuth, Suresh; George, Varghese; Rinaldi, Stefano; Pahwa, Rajendra; Arheart, Kristopher L; Pahwa, Savita

2017-10-19

Biological aging is associated with immune activation (IA) and declining immunity due to systemic inflammation. It is widely accepted that HIV infection causes persistent IA and premature immune senescence despite effective antiretroviral therapy and virologic suppression; however, the effects of combined HIV infection and aging are not well defined. Here, we assessed the relationship between markers of IA and inflammation during biological aging in HIV-infected and -uninfected populations. Antibody response to seasonal influenza vaccination was implemented as a measure of immune competence and relationships between IA, inflammation, and antibody responses were explored using statistical modeling appropriate for integrating high-dimensional data sets. Our results show that markers of IA, such as coexpression of HLA antigen D related (HLA-DR) and CD38 on CD4+ T cells, exhibit strong associations with HIV infection but not with biological age. Certain variables that showed a strong relationship with aging, such as declining naive and CD38+ CD4 and CD8+ T cells, did so regardless of HIV infection. Interestingly, the variable of biological age was not identified in a predictive model as significantly impacting vaccine responses in either group, while distinct IA and inflammatory variables were closely associated with vaccine response in HIV-infected and -uninfected populations. These findings shed light on the most relevant and persistent immune defects during virological suppression with antiretroviral therapy.

Immunophenotyping of posttraumatic neutrophils on a routine haematology analyser.

PubMed

Groeneveld, Kathelijne Maaike; Heeres, Marjolein; Leenen, Loek Petrus Hendrikus; Huisman, Albert; Koenderman, Leo

2012-01-01

Flow cytometry markers have been proposed as useful predictors for the occurrence of posttraumatic inflammatory complications. However, currently the need for a dedicated laboratory and the labour-intensive analytical procedures make these markers less suitable for clinical practice. We tested an approach to overcome these limitations. Neutrophils of healthy donors were incubated with antibodies commonly used in trauma research: CD11b (MAC-1), L-selectin (CD62L), FcγRIII (CD16), and FcγRII (CD32) in active form (MoPhab A27). Flow cytometric analysis was performed both on a FACSCalibur, a standard flow cytometer, and on a Cell-Dyn Sapphire, a routine haematology analyser. There was a high level of agreement between the two types of analysers, with 41% for FcγRIII, 80% for L-selectin, 98% for CD11b, and even a 100% agreement for active FcγRII. Moreover, analysis on the routine haematology analyser was possible in less than a quarter of the time in comparison to the flow cytometer. Analysis of neutrophil phenotype on the Cell-Dyn Sapphire leads to the same conclusion compared to a standard flow cytometer. The markedly reduced time necessary for analysis and reduced labour intensity constitutes a step forward in implementation of this type of analysis in clinical diagnostics in trauma research. Copyright © 2012 Kathelijne Maaike Groeneveld et al.

CXCR4(+)CD45(-) BMMNC subpopulation is superior to unfractionated BMMNCs for protection after ischemic stroke in mice.

PubMed

Wang, Jianping; Liu, Xi; Lu, Hong; Jiang, Chao; Cui, Xiaobing; Yu, Lie; Fu, Xiaojie; Li, Qian; Wang, Jian

2015-03-01

Cell-based therapy is considered to be a promising therapeutic strategy for stroke treatment. Although unfractionated bone marrow mononuclear cells (BMMNCs) have been tried in both preclinical and clinical trials, the effective subpopulations need to be identified. In this study, we used fluorescence-activated cell sorting to harvest the CXCR4(+)CD45(+) and CXCR4(+)CD45(-) BMMNC subpopulations from transgenic mice that express enhanced green fluorescent protein. We then allogeneically grafted unfractionated BMMNCs or a subpopulation into mice subjected to transient middle cerebral artery occlusion (tMCAO) and compared the effects on stroke outcomes. We found that CXCR4(+)CD45(-) BMMNCs, but not CXCR4(+)CD45(+) BMMNCs, more effectively reduced infarction volume and neurologic deficits than did unfractionated BMMNCs. Brain tissue from the ischemic hemisphere of mice treated with CXCR4(+)CD45(-) BMMNCs had higher levels of vascular endothelial growth factor and lower levels of TNF-α than did tissue from mice treated with unfractionated BMMNCs. In contrast, CXCR4(+)CD45(+) BMMNCs showed an increase in TNF-α. Additionally, CXCR4(+)CD45(+) and CXCR4(+)CD45(-) populations exhibited more robust migration into the lesion areas and were better able to express cell-specific markers of different linages than were the unfractionated BMMNCs. Endothelial and astrocyte cell markers did not colocalize with eGFP(+) cells in the brains of tMCAO mice that received CXCR4(+)CD45(+) BMMNCs. In vitro, the CXCR4(+)CD45(-) BMMNCs expressed significantly more Oct-4 and Nanog mRNA than did the unfractionated BMMNCs. However, we did not detect gene expression of these two pluripotent markers in CXCR4(+)CD45(+) BMMNCs. Taken together, our study shows for the first time that the CXCR4(+)CD45(-) BMMNC subpopulation is superior to unfractionated BMMNCs in ameliorating cerebral damage in a mouse model of tMCAO and could represent a new therapeutic approach for stroke treatment. Copyright © 2014 Elsevier Inc. All rights reserved.

Aurora A drives early signalling and vesicle dynamics during T-cell activation

PubMed Central

Blas-Rus, Noelia; Bustos-Morán, Eugenio; Pérez de Castro, Ignacio; de Cárcer, Guillermo; Borroto, Aldo; Camafeita, Emilio; Jorge, Inmaculada; Vázquez, Jesús; Alarcón, Balbino; Malumbres, Marcos; Martín-Cófreces, Noa B.; Sánchez-Madrid, Francisco

2016-01-01

Aurora A is a serine/threonine kinase that contributes to the progression of mitosis by inducing microtubule nucleation. Here we have identified an unexpected role for Aurora A kinase in antigen-driven T-cell activation. We find that Aurora A is phosphorylated at the immunological synapse (IS) during TCR-driven cell contact. Inhibition of Aurora A with pharmacological agents or genetic deletion in human or mouse T cells severely disrupts the dynamics of microtubules and CD3ζ-bearing vesicles at the IS. The absence of Aurora A activity also impairs the activation of early signalling molecules downstream of the TCR and the expression of IL-2, CD25 and CD69. Aurora A inhibition causes delocalized clustering of Lck at the IS and decreases phosphorylation levels of tyrosine kinase Lck, thus indicating Aurora A is required for maintaining Lck active. These findings implicate Aurora A in the propagation of the TCR activation signal. PMID:27091106

Different distribution patterns of lymphocytes and microglia in the hippocampus of patients with residual versus paranoid schizophrenia: further evidence for disease course-related immune alterations?

PubMed

Busse, Stefan; Busse, Mandy; Schiltz, Kolja; Bielau, Hendrik; Gos, Tomasz; Brisch, Ralf; Mawrin, Christian; Schmitt, Andrea; Jordan, Wolfgang; Müller, Ulf J; Bernstein, Hans-Gert; Bogerts, Bernhard; Steiner, Johann

2012-11-01

Certain cytokines have been identified in the peripheral blood as trait markers of schizophrenia, while others are considered relapse-related state markers. Furthermore, data from peripheral blood, cerebrospinal fluid (CSF) and nuclear imaging studies suggest that (1) blood-brain barrier (BBB) dysfunction (e.g., immigration of lymphocytes into brain tissue and intrathecal antibody production) correlates with the development of negative symptoms, while (2) the brain's mononuclear phagocyte system (microglial cells) is activated during acute psychosis. Based on these neuroinflammatory hypotheses, we have quantified the numerical density of immunostained CD3+ T-lymphocytes, CD20+ B-lymphocytes, and HLA-DR+ microglial cells in the posterior hippocampus of 17 schizophrenia patients and 11 matched controls. Disease course-related immune alterations were considered by a separate analysis of residual (prevailing negative symptoms, n=7) and paranoid (prominent positive symptoms, n=10) schizophrenia cases. Higher densities of CD3+ and CD20+ lymphocytes were observed in residual versus paranoid schizophrenia (CD 3: left: P=0.047, right: P=0.038; CD20: left: P=0.020, right: P=0.010) and controls (CD3: left: P=0.057, right: P=0.069; CD20: left: P=0.008, right: P=0.006). In contrast, HLA-DR+ microglia were increased in paranoid schizophrenia versus residual schizophrenia (left: P=0.030, right: P=0.012). A similar trend emerged when this group was compared to controls (left: P=0.090, right: P=0.090). BBB impairment and infiltration of T cells and B cells may contribute to the pathophysiology of residual schizophrenia, while microglial activation seems to play a role in paranoid schizophrenia. The identification of diverse immune endophenotypes may facilitate the development of distinct anti-inflammatory schizophrenia therapies to normalize BBB function, (auto)antibody production or microglial activity. Copyright © 2012 Elsevier Inc. All rights reserved.

Immunological abnormalities as potential biomarkers in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis

PubMed Central

2011-01-01

Background Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is characterised by severe prolonged fatigue, and decreases in cognition and other physiological functions, resulting in severe loss of quality of life, difficult clinical management and high costs to the health care system. To date there is no proven pathomechanism to satisfactorily explain this disorder. Studies have identified abnormalities in immune function but these data are inconsistent. We investigated the profile of markers of immune function (including novel markers) in CFS/ME patients. Methods We included 95 CFS/ME patients and 50 healthy controls. All participants were assessed on natural killer (NK) and CD8+T cell cytotoxic activities, Th1 and Th2 cytokine profile of CD4+T cells, expression of vasoactive intestinal peptide receptor 2 (VPACR2), levels of NK phenotypes (CD56bright and CD56dim) and regulatory T cells expressing FoxP3 transcription factor. Results Compared to healthy individuals, CFS/ME patients displayed significant increases in IL-10, IFN-γ, TNF-α, CD4+CD25+ T cells, FoxP3 and VPACR2 expression. Cytotoxic activity of NK and CD8+T cells and NK phenotypes, in particular the CD56bright NK cells were significantly decreased in CFS/ME patients. Additionally granzyme A and granzyme K expression were reduced while expression levels of perforin were significantly increased in the CFS/ME population relative to the control population. These data suggest significant dysregulation of the immune system in CFS/ME patients. Conclusions Our study found immunological abnormalities which may serve as biomarkers in CFS/ME patients with potential for an application as a diagnostic tool. PMID:21619669

The NOD2 p.Leu1007fsX1008 Mutation (rs2066847) Is a Stronger Predictor of the Clinical Course of Crohn's Disease than the FOXO3A Intron Variant rs12212067

PubMed Central

Wolf, Christiane; Angelberger, Marianne; Diegelmann, Julia; Olszak, Torsten; Beigel, Florian; Tillack, Cornelia; Stallhofer, Johannes; Göke, Burkhard; Glas, Jürgen; Lohse, Peter; Brand, Stephan

2014-01-01

Background Very recently, a sub-analysis of genome-wide association scans revealed that the non-coding single nucleotide polymorphism (SNP) rs12212067 in the FOXO3A gene is associated with a milder course of Crohn's disease (CD) (Cell 2013;155:57–69). The aim of our study was to evaluate the clinical value of the SNP rs12212067 in predicting the severity of CD by correlating CD patient genotype status with the most relevant complications of CD such as stenoses, fistulas, and CD-related surgery. Methodology/Principal Findings We genotyped 550 CD patients for rs12212067 (FOXO3A) and the three common CD-associated NOD2 mutations rs2066844, rs2066847, and rs2066847 and performed genotype-phenotype analyses. Results No significant phenotypic differences were found between the wild-type genotype TT of the FOXO3A SNP rs12212067 and the minor genotypes TG and GG independently from NOD2 variants. The allele frequency of the minor G allele was 12.7%. Age at diagnosis, disease duration, body mass index, surgery rate, stenoses, fistula, need for immunosuppressive therapy, and disease course were not significantly different. In contrast, the NOD2 mutant p.Leu1007fsX1008 (rs2066847) was highly associated with penetrating CD (p = 0.01), the development of fistulas (p = 0.01) and stenoses (p = 0.01), and ileal disease localization (p = 0.03). Importantly, the NOD2 SNP rs2066847 was a strong separator between an aggressive and a mild course of CD (p = 2.99×10−5), while the FOXO3A SNP rs12212067 did not separate between mild and aggressive CD behavior in our cohort (p = 0.35). 96.2% of the homozygous NOD2 p.Leu1007fsX1008 carriers had an aggressive disease behavior compared to 69.3% of the patients with the NOD2 wild-type genotype (p = 0.007). Conclusion/Significance In clinical practice, the NOD2 variant p.Leu1007fsX1008 (rs2066847), in particular in homozygous form, is a much stronger marker for a severe clinical phenotype than the FOXO3A rs12212067 SNP for a mild disease course on an individual patient level despite its important impact on the inflammatory response of monocytes. PMID:25365249

CRYAB modulates the activation of CD4+ T cells from relapsing-remitting multiple sclerosis patients.

PubMed

Quach, Que Lan; Metz, Luanne M; Thomas, Jenna C; Rothbard, Jonathan B; Steinman, Lawrence; Ousman, Shalina S

2013-12-01

Suppression of activation of pathogenic CD4(+) T cells is a potential therapeutic intervention in multiple sclerosis (MS). We previously showed that a small heat shock protein, CRYAB, reduced T cell proliferation, pro-inflammatory cytokine production and clinical signs of experimental allergic encephalomyelitis, a model of MS. We assessed whether the ability of CRYAB to reduce the activation of T cells translated to the human disease. CD4(+) T cells from healthy controls and volunteers with MS were activated in vitro in the presence or absence of a CRYAB peptide (residues 73-92). Parameters of activation (proliferation rate, cytokine secretion) and tolerance (anergy, activation-induced cell death, microRNAs) were evaluated. The secretion of pro-inflammatory cytokines by CD4(+) T cells was decreased in the presence of CRYAB in a subset of relapsing-remitting multiple sclerosis (RRMS) participants with mild disease severity while no changes were observed in healthy controls. Further, there was a correlation for higher levels of miR181a microRNA, a marker upregulated in tolerant CD8(+) T cells, in CD4(+) T cells of MS patients that displayed suppressed cytokine production (responders). CRYAB may be capable of suppressing the activation of CD4(+) T cells from a subset of RRMS patients who appear to have less disability but similar age and disease duration.

New Insight on a Combination of Policosanol and 10-Dehydrogingerdione Phytochemicals as Inhibitors for Platelet Activation Biomarkers and Atherogenicity Risk in Dyslipidemic Rabbits: Role of CETP and PCSK9 Inhibition.

PubMed

Elseweidy, Mohamed Mahmoud; Amin, Rawia Sarhan; Atteia, Hebatallah Husseini; El-Zeiky, Reham Raafat; Al-Gabri, Naif A

2018-05-09

Platelet markers [soluble p selectin (sP-selectin) and soluble CD40 ligand (sCD40L)] are associated with platelet activation and cardiovascular risk. Both policosanol and 10-dehydrogingerdione are natural products with proven CETP inhibitory and antiatherogenic effects. Present work aimed mainly to investigate the levels of platelet activation biomarkers in the serum of dyslipidemic rabbits and the potential of these phytochemicals either alone or in a combination form to protect against atherogenicity. Additionally, this work clarified their effect on PCSK9, a key player in atherosclerosis progression. Daily administration of policosanol and/or 10-dehydrogingerdione at a dose level 10 mg/kg bw resulted in a CETP inhibitory activity, increasing HDL-C level. This protective effect was associated with improvement in lipid profile components and a reduction in PCSK9 level. Interestingly, this combination strengthened the CETP inhibitory activity of these phytochemicals, leading to a greater increase in serum HDL-C level than monotherapy. However, this combination did not enhance the reduction in PCSK9 level. Both drugs also decreased platelet activation and inflammation markers such as sCD40L, sP-selectin, and interferon-gamma (IFN-γ), and their combination showed a synergistic effect. Therefore, such phytochemicals may be regarded as promising agents in the protection against atherothrombosis risk.

Assimilation of elements and digestion in grass shrimp pre-exposed to dietary mercury.

PubMed

Seebaugh, David R; Wallace, William G; L'amoreaux, William J; Stewart, Gillian M

2012-08-01

Grass shrimp Palaemonetes pugio were fed mercury (Hg)-contaminated oligochaetes for 15 days and analyzed for Hg, cadmium (Cd), and carbon assimilation efficiencies (AE) as well as toxicological end points related to digestion. Disproportionate increases in stable Hg concentrations in shrimp did not appear to be related to partitioning to trophically available Hg in worms. Hg AE by pre-exposed shrimp reached a plateau (approximately 53 %), whereas Cd AE varied (approximately 40-60 %) in a manner that was not dose-dependent. Carbon AE did not differ among treatments (approximately 69 %). Gut residence time was not impacted significantly by Hg pre-exposure (grand median approximately 465 min), however, there was a trend between curves showing percentages of individuals with markers in feces over time versus treatment. Feces-elimination rate did not vary with dietary pre-exposure. Extracellular protease activity varied approximately 1.9-fold but did not exhibit dose-dependency. pH increased over the range of Hg pre-exposures within the anterior (pH approximately 5.33-6.51) and posterior (pH approximately 5.29-6.25) regions of the cardiac proventriculus and Hg assimilation exhibited a negative relationship to hydrogen ion concentrations. The results of this study indicate that previous Hg ingestion can elicit post-assimilatory impacts on grass shrimp digestive physiology, which may, in turn, influence Hg assimilation during subsequent digestive cycles.

Aging, inflammation, and HIV infection.

PubMed

Aberg, Judith A

2012-01-01

Prolonged survival in HIV infection is accompanied by an increased frequency of non-HIV-related comorbidities. A number of age-related comorbidities occur earlier in HIV-infected patients than in individuals without HIV infection. This "accelerated aging" appears to be largely related to chronic inflammation, chronic immune activation, and immunosenescence in HIV infection. Levels of markers of inflammation and coagulopathy are elevated in HIV-infected patients, and elevations in markers such as high-sensitivity C-reactive protein, D-dimer, and interleukin 6 (IL-6) have been associated with increased risk for cardiovascular disease, opportunistic conditions, or all-cause mortality. In both HIV infection and aging, immunosenescence is marked by an increased proportion of CD28-, CD57+ memory CD8+ T cells with reduced capacity to produce interleukin 2 (IL-2), increased production of IL-6, resistance to apoptosis, and shortened telomeres. A number of AIDS Clinical Trials Group studies are under way to examine treatment aimed at reducing chronic inflammation and immune activation in HIV infection. This article summarizes a presentation by Judith A. Aberg, MD, at the IAS-USA live continuing medical education course held in New York City in October 2011.

Omega-3 Eicosapentaenoic Acid Decreases CD133 Colon Cancer Stem-Like Cell Marker Expression While Increasing Sensitivity to Chemotherapy

PubMed Central

De Carlo, Flavia; Witte, Theodore R.; Hardman, W. Elaine; Claudio, Pier Paolo

2013-01-01

Colorectal cancer is the third leading cause of cancer-related death in the western world. In vitro and in vivo experiments showed that omega-3 polyunsaturated fatty acids (n-3 PUFAs) can attenuate the proliferation of cancer cells, including colon cancer, and increase the efficacy of various anticancer drugs. However, these studies address the effects of n-3 PUFAs on the bulk of the tumor cells and not on the undifferentiated colon cancer stem-like cells (CSLCs) that are responsible for tumor formation and maintenance. CSLCs have also been linked to the acquisition of chemotherapy resistance and to tumor relapse. Colon CSLCs have been immunophenotyped using several antibodies against cellular markers including CD133, CD44, EpCAM, and ALDH. Anti-CD133 has been used to isolate a population of colon cancer cells that retains stem cells properties (CSLCs) from both established cell lines and primary cell cultures. We demonstrated that the n-3 PUFA, eicosapentaenoic acid (EPA), was actively incorporated into the membrane lipids of COLO 320 DM cells. 25 uM EPA decreased the cell number of the overall population of cancer cells, but not of the CD133 (+) CSLCs. Also, we observed that EPA induced down-regulation of CD133 expression and up-regulation of colonic epithelium differentiation markers, Cytokeratin 20 (CK20) and Mucin 2 (MUC2). Finally, we demonstrated that EPA increased the sensitivity of COLO 320 DM cells (total population) to both standard-of-care chemotherapies (5-Fluorouracil and oxaliplatin), whereas EPA increased the sensitivity of the CD133 (+) CSLCs to only 5-Fluorouracil. PMID:23874993

Accumulation of Multipotent Progenitor Cells on Polymethylpentene Membranes During Extracorporeal Membrane Oxygenation.

PubMed

Lehle, Karla; Friedl, Lucas; Wilm, Julius; Philipp, Alois; Müller, Thomas; Lubnow, Matthias; Schmid, Christof

2016-06-01

Multipotent progenitor cells were mobilized during pediatric extracorporeal membrane oxygenation (ECMO). We hypothesize that these cells also adhered onto polymethylpentene (PMP) fibers within the membrane oxygenator (MO) during adult ECMO support. Mononuclear cells were removed from the surface of explanted PMP-MOs (n = 16). Endothelial-like outgrowth and mesenchymal-like cells were characterized by flow cytometric analysis using different surface markers. Spindle-shaped attaching cells were identified early, but without proliferative activity. After long-term cultivation palisading type or cobblestone-type outgrowth cells with high proliferative activity appeared and were characterized as (i) leukocytoid CD45+/CD31+ (CD133+/VEGFR-II+/CD90+/CD14+/CD146dim/CD105dim); (ii) endothelial-like CD45-/CD31+ (VEGF-RII+/CD146+/CD105+/CD133-/CD14-/CD90-); and (iii) mesenchymal-like cells CD45-/CD31- (CD105+/CD90+/CD133dim/VEGFR-II-/CD146-/CD14-). The distribution of the cell populations depended on the MO and cultivation time. Endothelial-like cells formed capillary-like structures and did uptake Dil-acetylated low-density lipoprotein. Endothelial- and mesenchymal-like cells adhered on the surface of PMP-MOs. Further research is needed to identify the clinical relevance of these cells. Copyright © 2015 The Authors. Artificial Organs published by Wiley Periodicals, Inc. on behalf of International Center for Artificial Organs and Transplantation (ICAOT).

Massive infection and loss of CD4+ T cells occurs in the intestinal tract of neonatal rhesus macaques in acute SIV infection.

PubMed

Wang, Xiaolei; Rasmussen, Terri; Pahar, Bapi; Poonia, Bhawna; Alvarez, Xavier; Lackner, Andrew A; Veazey, Ronald S

2007-02-01

Rapid, profound, and selective depletion of memory CD4+ T cells has now been confirmed to occur in simian immunodeficiency virus (SIV)-infected adult macaques and human immunodeficiency virus (HIV)-infected humans. Within days of infection, marked depletion of memory CD4+ T cells occurs primarily in mucosal tissues, the major reservoir for memory CD4+ T cells in adults. However, HIV infection in neonates often results in higher viral loads and rapid disease progression, despite the paucity of memory CD4+ T cells in the peripheral blood. Here, we examined the immunophenotype of CD4+ T cells in normal and SIV-infected neonatal macaques to determine the distribution of naive and memory T-cell subsets in tissues. We demonstrate that, similar to adults, neonates have abundant memory CD4+ T cells in the intestinal tract and spleen and that these are selectively infected and depleted in primary SIV infection. Within 12 days of SIV infection, activated (CD69+), central memory (CD95+CD28+) CD4+ T cells are marked and persistently depleted in the intestine and other tissues of neonates compared with controls. The results in dicate that "activated" central memory CD4+ T cells are the major target for early SIV infection and CD4+ T cell depletion in neonatal macaques.

A simplified plastic embedding and immunohistologic technique for immunophenotypic analysis of human hematopoietic and lymphoid tissues.

PubMed Central

Casey, T. T.; Cousar, J. B.; Collins, R. D.

1988-01-01

Routine fixation and paraffin embedding destroys many hematopoietic and lymphoid differentiation antigens detected by flow cytometry or frozen section immunohistochemistry. On the other hand, morphologic evaluation is difficult in flow cytometric or frozen section studies. A simplified three-step plastic embedding system using acetone-fixed tissues embedded in glycol-methacrylate (GMA) resin has been found to provide both excellent morphologic and antigenic preservation. With our system, a wide variety of antigens are detected in plastic sections without trypsinization or prolonged embedding procedures; pan-B (CD19, CD22), pan-T (CD7, CD5, CD3, CD2), T-subset (CD4, CD8, CD1, CD25) markers as well as surface immunoglobulin and markers for myeloid and mononuclear-phagocyte cells are preserved. In summary, modifications of plastic embedding techniques used in this study simplify the procedure, apparently achieve excellent antigenic preservation, and facilitate evaluation of morphologic details in relation to immunocytochemical markers. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:3282442

Identification of myogenic-endothelial progenitor cells in the interstitial spaces of skeletal muscle.

PubMed

Tamaki, Tetsuro; Akatsuka, Akira; Ando, Kiyoshi; Nakamura, Yoshihiko; Matsuzawa, Hideyuki; Hotta, Tomomitsu; Roy, Roland R; Edgerton, V Reggie

2002-05-13

Putative myogenic and endothelial (myo-endothelial) cell progenitors were identified in the interstitial spaces of murine skeletal muscle by immunohistochemistry and immunoelectron microscopy using CD34 antigen. Enzymatically isolated cells were characterized by fluorescence-activated cell sorting on the basis of cell surface antigen expression, and were sorted as a CD34+ and CD45- fraction. Cells in this fraction were approximately 94% positive for Sca-1, and mostly negative (<3% positive) for CD14, 31, 49, 144, c-kit, and FLK-1. The CD34+/45- cells formed colonies in clonal cell cultures and colony-forming units displayed the potential to differentiate into adipocytes, endothelial, and myogenic cells. The CD34+/45- cells fully differentiated into vascular endothelial cells and skeletal muscle fibers in vivo after transplantation. Immediately after sorting, CD34+/45- cells expressed only c-met mRNA, and did not express any other myogenic cell-related markers such as MyoD, myf-5, myf-6, myogenin, M-cadherin, Pax-3, and Pax-7. However, after 3 d of culture, these cells expressed mRNA for all myogenic markers. CD34+/45- cells were distinct from satellite cells, as they expressed Bcrp1/ABCG2 gene mRNA (Zhou et al., 2001). These findings suggest that myo-endothelial progenitors reside in the interstitial spaces of mammalian skeletal muscles, and that they can potentially contribute to postnatal skeletal muscle growth.

Recognition of Naegleriae ameba surface protein epitopes by anti-human CD45 antibodies.

PubMed

Ravine, Terrence J; Polski, Jacek M; Jenkins, James

2010-04-01

Phagocytosis is a highly conserved mechanism exhibited by both free-living amebas and mammalian blood cells. Similarities demonstrated by either cell type during engulfment of the same bacterial species may imply analogous surface proteins involved in receptor-mediated endocytosis. The increased availability of anti-human leukocyte antibodies or clusters of differentiation (CD) markers used in conjunction with flow cytometric (FCM) and/or immunohistochemical (IHC) analysis provides investigators with a relatively easy method to screen different cell populations for comparable plasma membrane proteins. In this study, we incubated Naegleria and Acanthamoeba amebas with several directly conjugated anti-human leukocyte monoclonal antibodies (mAb) for similarly recognized amebic epitopes. CD marker selection was based upon a recognized role of each mAb in phagocyte activation and/or uptake of bacteria. These included CD14, CD45, and CD206. In FCM, only one CD45 antibody demonstrated strong reactivity with both Naegleria fowleri and Naegleria gruberi that was not expressed in similarly tested Acanthamoeba species. Additional testing of N. gruberi by IHC demonstrated reactivity to a different CD45 antibody. Our results suggest a possible utility of using anti-human leukocyte antibodies to screen amebic cells for similarly expressed protein epitopes. In doing so, several important items must be considered when selecting potential mAbs for testing to increase the probability of a positive result.

Mesenchymal stem cell like (MSCl) cells generated from human embryonic stem cells support pluripotent cell growth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Varga, Nora; Vereb, Zoltan; Rajnavoelgyi, Eva

2011-10-28

Highlights: Black-Right-Pointing-Pointer MSC like cells were derived from hESC by a simple and reproducible method. Black-Right-Pointing-Pointer Differentiation and immunosuppressive features of MSCl cells were similar to bmMSC. Black-Right-Pointing-Pointer MSCl cells as feeder cells support the undifferentiated growth of hESC. -- Abstract: Mesenchymal stem cell like (MSCl) cells were generated from human embryonic stem cells (hESC) through embryoid body formation, and isolated by adherence to plastic surface. MSCl cell lines could be propagated without changes in morphological or functional characteristics for more than 15 passages. These cells, as well as their fluorescent protein expressing stable derivatives, efficiently supported the growth ofmore » undifferentiated human embryonic stem cells as feeder cells. The MSCl cells did not express the embryonic (Oct4, Nanog, ABCG2, PODXL, or SSEA4), or hematopoietic (CD34, CD45, CD14, CD133, HLA-DR) stem cell markers, while were positive for the characteristic cell surface markers of MSCs (CD44, CD73, CD90, CD105). MSCl cells could be differentiated toward osteogenic, chondrogenic or adipogenic directions and exhibited significant inhibition of mitogen-activated lymphocyte proliferation, and thus presented immunosuppressive features. We suggest that cultured MSCl cells can properly model human MSCs and be applied as efficient feeders in hESC cultures.« less

Differential Expression of NK Receptors CD94 and NKG2A by T Cells in Rheumatoid Arthritis Patients in Remission Compared to Active Disease

PubMed Central

Walsh, Ceara E.; Ryan, Elizabeth J.; O’Farrelly, Cliona; Golden-Mason, Lucy; FitzGerald, Oliver; Veale, Douglas J.; Bresnihan, Barry; Fearon, Ursula

2011-01-01

Objective TNF inhibitors (TNFi) have revolutionised the treatment of rheumatoid arthritis (RA). Natural killer (NK) cells and Natural Killer Cell Receptor+ T (NKT) cells comprise important effector lymphocytes whose activity is tightly regulated through surface NK receptors (NKRs). Dysregulation of NKRs in patients with autoimmune diseases has been shown, however little is known regarding NKRs expression in patients with TNFi-induced remission and in those who maintain remission vs disease flare following TNFi withdrawal. Methods Patients with RA were recruited for this study, (i) RA patients in clinical remission following a minimum of one year of TNFi therapy (n = −15); (2) Active RA patients, not currently or ever receiving TNFi (n = 18); and healthy control volunteers (n = 15). Patients in remission were divided into two groups: those who were maintained on TNFi and those who withdrew from TNFi and maintained on DMARDS. All patients underwent full clinical assessment. Peripheral blood mononuclear cells were isolated and NKR (CD94, NKG2A, CD161, CD69, CD57, CD158a, CD158b) expression on T-(CD3+CD56−), NK-(CD3−CD56+) and NKT-(CD3+CD56+) cells was determined by flow cytometry. Results Following TNFi withdrawal, percentages and numbers of circulating T cells, NK cells or NKT cell populations were unchanged in patients in remission versus active RA or HCs. Expression of the NKRs CD161, CD57, CD94 and NKG2A was significantly increased on CD3+CD56-T cells from patients in remission compared to active RA (p<0.05). CD3+CD56-T cell expression of CD94 and NKG2A was significantly increased in patients who remained in remission compared with patients whose disease flared (p<0.05), with no differences observed for CD161 and CD57. CD3+CD56− cell expression of NKG2A was inversely related to DAS28 (r = −0.612, p<0.005). Conclusion High CD94/NKG2A expression by T cells was demonstrated in remission patients following TNFi therapy compared to active RA, while low CD94/NKG2A were associated with disease flare following withdrawal of therapy. PMID:22102879

Differential expression of NK receptors CD94 and NKG2A by T cells in rheumatoid arthritis patients in remission compared to active disease.

PubMed

Walsh, Ceara E; Ryan, Elizabeth J; O'Farrelly, Cliona; Golden-Mason, Lucy; FitzGerald, Oliver; Veale, Douglas J; Bresnihan, Barry; Fearon, Ursula

2011-01-01

TNF inhibitors (TNFi) have revolutionised the treatment of rheumatoid arthritis (RA). Natural killer (NK) cells and Natural Killer Cell Receptor+ T (NKT) cells comprise important effector lymphocytes whose activity is tightly regulated through surface NK receptors (NKRs). Dysregulation of NKRs in patients with autoimmune diseases has been shown, however little is known regarding NKRs expression in patients with TNFi-induced remission and in those who maintain remission vs disease flare following TNFi withdrawal. Patients with RA were recruited for this study, (i) RA patients in clinical remission following a minimum of one year of TNFi therapy (n = -15); (2) Active RA patients, not currently or ever receiving TNFi (n = 18); and healthy control volunteers (n = 15). Patients in remission were divided into two groups: those who were maintained on TNFi and those who withdrew from TNFi and maintained on DMARDS. All patients underwent full clinical assessment. Peripheral blood mononuclear cells were isolated and NKR (CD94, NKG2A, CD161, CD69, CD57, CD158a, CD158b) expression on T-(CD3+CD56-), NK-(CD3-CD56+) and NKT-(CD3+CD56+) cells was determined by flow cytometry. Following TNFi withdrawal, percentages and numbers of circulating T cells, NK cells or NKT cell populations were unchanged in patients in remission versus active RA or HCs. Expression of the NKRs CD161, CD57, CD94 and NKG2A was significantly increased on CD3+CD56-T cells from patients in remission compared to active RA (p<0.05). CD3+CD56-T cell expression of CD94 and NKG2A was significantly increased in patients who remained in remission compared with patients whose disease flared (p<0.05), with no differences observed for CD161 and CD57. CD3+CD56- cell expression of NKG2A was inversely related to DAS28 (r = -0.612, p<0.005). High CD94/NKG2A expression by T cells was demonstrated in remission patients following TNFi therapy compared to active RA, while low CD94/NKG2A were associated with disease flare following withdrawal of therapy.

Subclinical herpesvirus shedding among HIV-1-infected men on antiretroviral therapy.

PubMed

Agudelo-Hernandez, Arcadio; Chen, Yue; Bullotta, Arlene; Buchanan, William G; Klamar-Blain, Cynthia R; Borowski, Luann; Riddler, Sharon A; Rinaldo, Charles R; Macatangay, Bernard J C

2017-09-24

We evaluated the subclinical shedding of six different herpesviruses in antiretroviral drug-treated HIV-positive [HIV(+)] MSM, and determined how this is associated with markers of inflammation and immune activation. We obtained blood, semen, throat washing, urine, and stool from 15 antiretroviral-treated HIV-1-infected MSM with CD4 T-cell reconstitution, and 12 age-matched HIV-negative [HIV (-)] MSM from the Multicenter AIDS Cohort Study at four timepoints over 24 weeks to measure DNA levels of cytomegalovirus (CMV), Epstein-Barr virus (EBV), herpes simplex virus 1 and 2, human herpesvirus 6 (HHV6), and HHV8. T-cell activation and plasma levels of soluble markers of inflammation and activation were also measured at the corresponding timepoints. HIV(+) participants had a trend for higher total herpesvirus shedding rate. HIV(+) participants also had a significantly higher rate of shedding EBV and CMV compared with the HIV(-) group. Herpesvirus shedding was mostly seen in throat washings. In the HIV(+) group, herpesvirus shedding rate inversely correlated with plasma levels of interferon γ-induced protein 10 and soluble CD163. CMV DNA levels negatively correlated with levels of T-cell activation. There was a trend for a positive correlation between EBV shedding rate and plasma soluble CD14. HHV6 shedding rate negatively correlated with plasma levels of interleukin-6, soluble CD163, and interferon gamma-induced protein 10. Correlations were not observed among HIV(-) individuals. Among treated HIV-infected MSM, there are higher subclinical shedding rates of some herpesviruses that occur in different body compartments and negatively correlate with levels of inflammation and immune activation.

Rebound of residual plasma viremia after initial decrease following addition of intravenous immunoglobulin to effective antiretroviral treatment of HIV.

PubMed

Mellberg, Tomas; Gonzalez, Veronica D; Lindkvist, Annica; Edén, Arvid; Sönnerborg, Anders; Sandberg, Johan K; Svennerholm, Bo; Gisslén, Magnus

2011-06-28

High dosage of intravenous immunoglobulin (IVIG) has been observed as a possible activator of HIV gene expression in latently infected resting CD4+ T-cells, leading to a substantial decrease in both the reservoir and the residual plasma viremia when added to effective ART. IVIG treatment has also been reported to expand T regulatory cells (Tregs). The aim of this study was to evaluate possible long-term effect of IVIG treatment on residual viremia and T-lymphocyte activation. Nine HIV-infected subjects on effective ART included in a previously reported study on IVIG treatment were evaluated 48-104 weeks after therapy. In addition, 14 HIV-infected controls on suppressive ART were included. HIV-1 RNA was analyzed in cell-free plasma by using an ultrasensitive PCR-method with a detection limit of 2 copies/mL. T-lymphocyte activation markers and serum interleukins were measured. Plasma residual viremia rebounded to pre-treatment levels, 48-104 weeks after the initial decrease that was observed following treatment with high-dosage IVIG. No long-term effect was observed regarding T-lymphocyte activation markers, T-regulatory cells or serum interleukins. In a post-hoc analysis, a correlation between plasma HIV-1-RNA and CD4+ T-cell count was found in both IVIG-treated patients and controls. These results indicate that the decrease in the latent HIV-1 pool observed during IVIG treatment is transient. Although not our primary objective, we found a correlation between HIV-1 RNA and CD4+ T-cell count suggesting the possibility that patients with a higher CD4+ T-cell count might harbor a larger residual pool of latently infected CD4+ T-cells.