Selective anti-proliferative activities of Carica papaya leaf juice extracts against prostate cancer.

PubMed

Pandey, Saurabh; Walpole, Carina; Cabot, Peter J; Shaw, Paul N; Batra, Jyotsna; Hewavitharana, Amitha K

2017-05-01

Prostate cancer (PCa) is the leading cause of cancer related deaths in men. Carica papaya is a popular tropical plant that has been traditionally used for its nutritional and medicinal properties. We investigated the anti-proliferative responses of papaya leaf juice (LJP) and its various extracts ("biological"- in vitro digested, "physical"- size exclusion, and "chemical"-solvent extraction) on a range of cell lines representing benign hyperplasia, tumorigenic and normal cells of prostate origin. Time course analysis (by 24h, 48h and 72h) of LJP (1-0.1mg/mL) before and after in vitro digestion, and of molecular weight based fractions of LJP showed anti-proliferative responses. The medium polarity fraction of LJP (0.03-0.003mg/mL) after 72h exposure showed potent growth inhibitory (IC 50 =0.02-0.07mg/mL) and cytotoxic activities on all prostate cells, with the exception of the normal (RWPE-1 and WPMY-1) cells. Flow cytometry analysis showed S phase cell cycle arrest and apoptosis as a possible mechanism for these activities. Medium polar fraction of LJP also inhibited migration and adhesion of metastatic PC-3 cells. This is the first report suggesting selective anti-proliferative and anti-metastatic attributes of LJP extract against prostatic diseases, including PCa. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Preliminary investigations on the effects of a Strongylus vulgaris larval extract, mononuclear factors and platelet factors on equine smooth muscle cells in vitro.

PubMed

Morgan, S J; Storts, R W; Stromberg, P C; Sowa, B A; Lay, J C

1989-01-01

Factors involved in the proliferation of equine vascular smooth muscle cells were studied in vitro. The most prominent proliferative responses in cultured vascular smooth muscle cells were induced by Strongylus vulgaris larval antigen extract (LAE) and platelet-derived factors. Less significant proliferative responses were obtained with conditioned media from S. vulgaris LAE stimulated and from unstimulated equine mononuclear leukocytes. Additionally, vascular smooth muscle cells exposed to S. vulgaris LAE developed numerous perinuclear vacuoles and were more spindle-shaped than control or smooth muscle cells exposed to other factors. Equine mononuclear leukocytes exposed to LAE developed prominent morphological changes, including enlargement, clumping and increased numbers of mitotic figures.

CD4+ T-cell engagement by both wild-type and variant HCV peptides modulates the conversion of viral clearing helper T cells to Tregs

PubMed Central

Cusick, Matthew F; Libbey, Jane E; Cox Gill, Joan; Fujinami, Robert S; Eckels, David D

2013-01-01

Aim To determine whether modulation of T-cell responses by naturally occurring viral variants caused an increase in numbers of Tregs in HCV-infected patients. Patients, materials & methods Human peripheral blood mononuclear cells, having proliferative responses to a wild-type HCV-specific CD4+ T-cell epitope, were used to quantify, via proliferative assays, flow cytometry and class II tetramers, the effects of naturally occurring viral variants arising in the immunodominant epitope. Results In combination, the wild-type and variant peptides led to enhanced suppression of an anti-HCV T-cell response. The variant had a lower avidity for the wild-type-specific CD4+ T cell. Variant-stimulated CD4+ T cells had increased Foxp3, compared with wild-type-stimulated cells. Conclusion A stable viral variant from a chronic HCV subject was able to induce Tregs in multiple individuals that responded to the wild-type HCV-specific CD4+ T-cell epitope. PMID:24421862

ESR1 Mutations Affect Anti-proliferative Responses to Tamoxifen through Enhanced Cross-Talk with IGF Signaling

PubMed Central

Gelsomino, Luca; Gu, Guowei; Rechoum, Yassine; Beyer, Amanda R; Pejerrey, Sasha M; Tsimelzon, Anna; Wang, Tao; Huffman, Kenneth; Ludlow, Andrew; Ando’, Sebastiano; Fuqua, Suzanne AW

2017-01-01

It is now generally accepted that estrogen receptor (ESR1) mutations occur frequently in metastatic breast cancers, however we do not yet know how to best treat these patients. We have modeled the three most frequent hormone binding ESR1 (HBD-ESR1) mutations (Y537N, Y537S, and D538G) using stable lentiviral transduction in human breast cancer cell lines. Effects on growth were examined in response to hormonal and targeted agents, and mutation-specific changes were studied using microarray and western blot analysis. We determined that the HBD-ESR1 mutations alter anti-proliferative effects to tamoxifen (Tam), due to cell-intrinsic changes in activation of the insulin-like growth factor receptor (IGF1R) signaling pathway and levels of PIK3R1/PIK3R3. The selective estrogen receptor degrader, fulvestrant, significantly reduced the anchorage-independent growth of ESR1 mutant-expressing cells, while combination treatments with the mTOR inhibitor everolimus, or an inhibitor blocking IGF1R and the insulin receptor significantly enhanced anti-proliferative responses. Using digital drop (dd) PCR we identified mutations at high frequencies ranging from 12% for Y537N, 5% for Y537S, and 2% for D538G in archived primary breast tumors from women treated with adjuvant mono-tamoxifen therapy. The HBD-ESR1 mutations were not associated with recurrence-free or overall survival in response in this patient cohort, and suggest that knowledge of other cell-intrinsic factors in combination with ESR1 mutation status will be needed determine anti-proliferative responses to Tam. PMID:27178332

T-cell proliferative responses following sepsis in neonatal rats.

PubMed

Dallal, Ousama; Ravindranath, Thyyar M; Choudhry, Mashkoor A; Kohn, Annamarie; Muraskas, Jonathan K; Namak, Shahla Y; Alattar, Mohammad H; Sayeed, Mohammed M

2003-01-01

Both experimental and clinical evidence suggest a suppression of T-cell function in burn and sepsis. The objective of the present study was to evaluate splenocyte and purified T-cell proliferative response and IL-2 production in septic neonatal rats. We also examined if alterations in T-cell proliferation and IL-2 production in neonatal sepsis is due to elevation in PGE2. PGE2 is known to play a significant role in T-cell suppression during sepsis in adults. Sepsis was induced in 15-day-old neonatal Sprague-Dawley rats by implanting 0.1 cm3 of fecal pellet impregnated with Escherichia coli (50 CFU) and Bacteroides fragilis (10(3) CFU). Animals receiving fecal pellets without the bacteria were designated as sterile. A group of septic and sterile rats were treated with PGE2 synthesis inhibitors, NS398 and resveratrol. These treatments of animals allowed us to evaluate the role of PGE2 in T-cell suppression during neonatal sepsis. Splenocytes as well as purified T cells were prepared and then proliferative response and IL-2 productive capacities were measured. A significant suppression of splenocyte proliferation and IL-2 production was noticed in both sterile and septic animals compared to the T cells from unoperated control rats. In contrast, the proliferation and IL-2 production by nylon wool purified T cells in sterile rats was not significantly different from control rats, whereas, a significant suppression in Con A-mediated T-cell proliferation and IL-2 production noticed in septic rat T cells compared to the sterile and control rat T cells. Such decrease in T-cell proliferation and IL-2 production was accompanied with 20-25% deaths in neonates implanted with septic pellets. No mortality was noted in sterile-implanted neonates. Treatment of animals with COX-1 inhibitor had no effect on T-cell proliferation response in both septic and sterile groups, whereas COX-2 inhibitor abrogated the decrease in T-cell proliferative response in the septic group. The treatment of animals with COX-2 inhibitor also significantly prevented the sepsis-associated mortality in neonates. In conclusion, the present study demonstrated T-cell suppression during neonatal sepsis is accompanied by a decrease in IL-2 production. Such suppressions were ameliorated with COX-2 inhibitor suggesting a role for PGE2 in the suppressed T-cell-mediated immune function in neonatal sepsis. Copyright 2003 S. Karger AG, Basel

Bovine central memory T cells are highly proliferative in response to bovine tuberculosis infection

USDA-ARS?s Scientific Manuscript database

Long-term (i.e., 14 days) cultured IFN-gamma responses of peripheral blood mononuclear cells are used as a correlate of T cell central memory (Tcm) responses in both humans and cattle. With bovine tuberculosis, vaccine-elicited long-term IFN-gamma ELISPOT assays are a correlate of protection. Recent...

Phenytoin promotes Th2 type immune response in mice

PubMed Central

Okada, K; Sugiura, T; Kuroda, E; Tsuji, S; Yamashita, U

2001-01-01

The effects of chronic administration of phenytoin, a common anticonvulsive drug, on immune responses were studied in mice. Anti-keyhole limpet haemocyanin (KLH) IgE antibody response after KLH-immunization was enhanced in phenytoin-treated mice. Proliferative responses of spleen cells induced with KLH, concanavalin A (ConA), lipopolysaccharide and anti-CD3 antibody were reduced in phenytoin-treated mice. Accessory function of spleen adherent cells on ConA-induced T cell proliferative response was reduced in phenytoin-treated mice. KLH-induced IL-4 production of spleen cells was enhanced, while IFN-γ production was reduced in phenytoin-treated mice. In addition, production of IL-1α, but not IL-6 and IL-12 by spleen adherent cells from phenytoin-treated mice was reduced. Natural killer cell activity was reduced in phenytoin-treated mice. These results suggest that phenytoin treatment preferentially induces a Th2 type response. We also observed that plasma ACTH and corticosterone levels were increased in phenytoin-treated mice, and speculated that phenytoin might act directly and indirectly, through HPA axis activation, on the immune system to modulate Th1/Th2 balance. PMID:11472401

IL-3 induces apoptosis in a ras-transformed myeloid cell line.

PubMed

Ahmed, N; Anderson, S M; Berridge, M V

1999-04-01

Growth factors promote cell survival and proliferation. Homeostasis is maintained by programmed cell death which occurs when the growth stimulus is withdrawn, in response to negative growth regulators such as interferons, TNF-alpha and CD95 ligand, or following differentiation. Although acutely-transforming oncogenes often overcome the need for growth factors, growth regulatory cytokines can influence proliferative responses of transformed cells. In this study we investigated the effects of IL-3 on the proliferative responses of parental bone marrow-derived 32D cells and cells transformed with ras and abl oncogenes. We show that treatment of ras-transformed 32D cells with IL-3 reduced proliferative responses and decreased colony-forming ability. These effects were exacerbated in the absence of serum and associated with inhibition of tyrosine kinase activity, down-regulation of RAS and MYC expression, and induction of apoptosis as indicated by DNA fragmentation. In contrast, treatment of parental 32D cells with IL-3, which is obligatory for cell survival and proliferation, increased tyrosine kinase activity, upregulated MYC and RAS expression and maintained DNA integrity. With abl-transformed cells, proliferation and colony-forming ability were also inhibited by IL-3. Tyrosine kinase activity and MYC expression were reduced, but early apoptosis was not evident. Calcium uptake however, was stimulated by IL-3 in both parental and oncogene-transformed cells. These results suggest that threshold levels of tyrosine kinase activity are necessary for cell survival and proliferation and that with ras-transformed cells, IL-3 treatment may result in this threshold being breached. We conclude that in some situations, growth-promoting cytokines can inhibit proliferation of transformed cells and induce cell death by apoptosis.

Genetic control of murine T cell proliferative responses to Mycobacterium leprae. V. Evidence for cross-reactivity between host antigens and Mycobacterium leprae

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

Harris, D.P.; Jones, A.G.; Wade, S.

1988-09-01

T cell proliferative responses to Mycobacterium leprae were measured by immunization of mice at the base of the tail with Ag and challenging lymphocytes from draining lymph nodes in culture with M. leprae. C57BL/10J and B10.BR mice were identified as low responder mice and the congenic strains B10.M, B10.Q, and B10.AKM as high responders whereas F1 (high x low) hybrid mice were found to be low responders. The cellular basis of low responsiveness did not appear to result from a defect in Ag-presenting cells or the activation of suppressor T cells by M. leprae. The influence of the environment inmore » which T cells developed on responsiveness to M. leprae was analyzed in chimeric mice prepared by irradiating F1(C57BL/10J x B10.M) mice and reconstituting with bone marrow from C57BL/10J, B10.M, or F1 donors. Six weeks later, chimeric mice were immunized with M. leprae, lymph node cells were subsequently prepared, and H-2 phenotyped and challenged in culture with M. leprae Ag. T cell proliferative responses were found to be low in all cases, similar to those observed using lymph node cells from F1 hybrid mice. These results suggested that high responder B10.M lymphocytes developing in the irradiated F1 mice became tolerized to antigenic determinants found on M. leprae. This implied cross-reactive epitopes existed between some mouse strains and M. leprae. Low responsiveness to M. leprae in low responder and F1 hybrid mice may result from tolerance to H-2-encoded Ag that show cross-reactivity with M. leprae.« less

Donor cell reaction to OKT3 as predictor of chronic graft-vs-host disease in hematopoietic stem cell recipients.

PubMed

Lindemann, Monika; Ottinger, Hellmut D; Elmaagacli, Ahmet H; Trenschel, Rudolf; Rebmann, Vera; Beelen, Dietrich W; Grosse-Wilde, Hans

2006-12-01

In the hematopoietic stem cell transplantation setting, granulocyte colony-stimulating factor (G-CSF) administration can reduce donor cell reactivity in vitro, but the clinical significance of this phenomenon was only sparsely defined. We performed lymphocyte transformation tests in 28 related stem cell donors pre and 5 days post G-CSF treatment, respectively, and correlated proliferative responses of donor peripheral blood mononuclear cells with clinical parameters in the corresponding recipients. In vitro reactions towards 4 mitogens and 12 recall antigens at day 5 post G-CSF administration were predictive for the occurrence of chronic graft-vs-host disease (cGVHD). Here, proliferative responses towards the mitogen anti-CD3 monoclonal antibody (OKT3) above median were most informative; this threshold could be determined by discrimination and receiver operating curve (ROC) analyses. In the whole cohort (18 human leukocyte antigen [HLA]-identical and 10 partially mismatched donor-recipient pairs), OKT3 responses predicted cGVHD with an odds ratio of 33.0, a sensitivity of 79%, and a specificity of 90%. A subgroup analysis of HLA-identical pairs even yielded an odds ratio of 85.0. Furthermore, bivariate analysis defined HLA compatibility and responses towards OKT3 as independent risk factors for cGVHD (p = 0.02 and p = 0.0007, respectively). The proliferative capacity of G-CSF-mobilized donor cells appears as a graft factor that determines the future incidence of cGVHD in the corresponding recipient.

Dissecting the T Cell Response: Proliferation Assays vs. Cytokine Signatures by ELISPOT

PubMed Central

Anthony, Donald D.; Milkovich, Kimberly A.; Zhang, Wenji; Rodriguez, Benigno; Yonkers, Nicole L.; Tary-Lehmann, Magdalena; Lehmann, Paul V.

2012-01-01

Chronic allograft rejection is in part mediated by host T cells that recognize allogeneic antigens on transplanted tissue. One factor that determines the outcome of a T cell response is clonal size, while another is the effector quality. Studies of alloimmune predictors of transplant graft survival have most commonly focused on only one measure of the alloimmune response. Because differing qualities and frequencies of the allospecific T cell response may provide distinctly different information we analyzed the relationship between frequency of soluble antigen and allo-antigen specific memory IFN-γ secreting CD4 and CD8 T cells, their ability to secrete IL-2, and their proliferative capacity, while accounting for cognate and bystander proliferation. The results show proliferative responses primarily reflect on IL-2 production by antigen-specific T cells, and that proliferating cells in such assays entail a considerable fraction of bystander cells. On the other hand, proliferation (and IL-2 production) did not reflect on the frequency of IFN-γ producing memory cells, a finding particularly accentuated in the CD8 T cell compartment. These data provide rationale for considering both frequency and effector function of pre-transplant T cell reactivity when analyzing immune predictors of graft rejection. PMID:24710419

Screening NK-, B- and T-cell phenotype and function in patients suffering from Chronic Fatigue Syndrome.

PubMed

Curriu, Marta; Carrillo, Jorge; Massanella, Marta; Rigau, Josepa; Alegre, José; Puig, Jordi; Garcia-Quintana, Ana M; Castro-Marrero, Jesus; Negredo, Eugènia; Clotet, Bonaventura; Cabrera, Cecilia; Blanco, Julià

2013-03-20

Chronic Fatigue Syndrome (CFS) is a debilitating neuro-immune disorder of unknown etiology diagnosed by an array of clinical manifestations. Although several immunological abnormalities have been described in CFS, their heterogeneity has limited diagnostic applicability. Immunological features of CFS were screened in 22 CFS diagnosed individuals fulfilling Fukuda criteria and 30 control healthy individuals. Peripheral blood T, B and NK cell function and phenotype were analyzed by flow cytometry in both groups. CFS diagnosed individuals showed similar absolute numbers of T, B and NK cells, with minor differences in the percentage of CD4+ and CD8+ T cells. B cells showed similar subset frequencies and proliferative responses between groups. Conversely, significant differences were observed in T cell subsets. CFS individuals showed increased levels of T regulatory cells (CD25+/FOXP3+) CD4 T cells, and lower proliferative responses in vitro and in vivo. Moreover, CD8 T cells from the CFS group showed significantly lower activation and frequency of effector memory cells. No clear signs of T-cell immunosenescence were observed. NK cells from CFS individuals displayed higher expression of NKp46 and CD69 but lower expression of CD25 in all NK subsets defined. Overall, T cell and NK cell features clearly clustered CFS individuals. Our findings suggest that alterations in T-cell phenotype and proliferative response along with the specific signature of NK cell phenotype may be useful to identify CFS individuals. The striking down modulation of T cell mediated immunity may help to understand intercurrent viral infections in CFS.

Screening NK-, B- and T-cell phenotype and function in patients suffering from Chronic Fatigue Syndrome

PubMed Central

2013-01-01

Background Chronic Fatigue Syndrome (CFS) is a debilitating neuro-immune disorder of unknown etiology diagnosed by an array of clinical manifestations. Although several immunological abnormalities have been described in CFS, their heterogeneity has limited diagnostic applicability. Methods Immunological features of CFS were screened in 22 CFS diagnosed individuals fulfilling Fukuda criteria and 30 control healthy individuals. Peripheral blood T, B and NK cell function and phenotype were analyzed by flow cytometry in both groups. Results CFS diagnosed individuals showed similar absolute numbers of T, B and NK cells, with minor differences in the percentage of CD4+ and CD8+ T cells. B cells showed similar subset frequencies and proliferative responses between groups. Conversely, significant differences were observed in T cell subsets. CFS individuals showed increased levels of T regulatory cells (CD25+/FOXP3+) CD4 T cells, and lower proliferative responses in vitro and in vivo. Moreover, CD8 T cells from the CFS group showed significantly lower activation and frequency of effector memory cells. No clear signs of T-cell immunosenescence were observed. NK cells from CFS individuals displayed higher expression of NKp46 and CD69 but lower expression of CD25 in all NK subsets defined. Overall, T cell and NK cell features clearly clustered CFS individuals. Conclusions Our findings suggest that alterations in T-cell phenotype and proliferative response along with the specific signature of NK cell phenotype may be useful to identify CFS individuals. The striking down modulation of T cell mediated immunity may help to understand intercurrent viral infections in CFS. PMID:23514202

Micronutrient supplementation and T cell-mediated immune responses in patients with tuberculosis in Tanzania

PubMed Central

KAWAI, K.; MEYDANI, S. N.; URASSA, W.; WU, D.; MUGUSI, F. M.; SAATHOFF, E.; BOSCH, R. J.; VILLAMOR, E.; SPIEGELMAN, D.; FAWZI, W. W.

2017-01-01

SUMMARY Limited studies exist regarding whether incorporating micronutrient supplements during tuberculosis (TB) treatment may improve cell-mediated immune response. We examined the effect of micronutrient supplementation on lymphocyte proliferation response to mycobacteria or T-cell mitogens in a randomized trial conducted on 423 patients with pulmonary TB. Eligible participants were randomly assigned to receive a daily dose of micronutrients (vitamins A, B-complex, C, E, and selenium) or placebo at the time of initiation of TB treatment. We found no overall effect of micronutrient supplements on lymphocyte proliferative responses to phytohaemagglutinin or purified protein derivatives in HIV-negative and HIV-positive TB patients. Of HIV-negative TB patients, the micronutrient group tended to show higher proliferative responses to concanavalin A than the placebo group, although the clinical relevance of this finding is not readily notable. The role of nutritional intervention in this vulnerable population remains an important area of future research. PMID:24093552

PDC-E3BP is not a dominant T-cell autoantigen in primary biliary cirrhosis.

PubMed

McHugh, Anna; Robe, Amanda J; Palmer, Jeremy M; Jones, David E J

2006-05-01

Autoantibody responses reactive with the E2 and E3BP components of pyruvate dehydrogenase complex (PDC), which characterise primary biliary cirrhosis (PBC) crossreact, precluding the identification, from serological studies, of the antigen to which the principal breakdown of tolerance occurs. Although autoreactive T-cell responses to PDC-E2 have been well characterised it is, at present, unclear whether T-cell tolerance breakdown also occurs to PDC-E3BP. The aims of this study were to characterise autoreactive T-cell responses to PDC-E3BP in PBC and potential factors regulating their expression. Peripheral blood T-cell proliferative responses to purified recombinant human PDC-E2 and PDC-E3BP at a range of concentrations were characterised in PBC patients and control subjects. T-cell proliferative responses to both E2 and E3BP were absent from control subjects (median peak stimulation index (SI) to PDC-E2 1.2 [range 0.3-1.9], 0/10 positive (SI>2.32), median peak SI to PDC-E3BP 1.1 [0.7-2.1

Antioxidant activity, anti-proliferative activity, and amino acid profiles of ethanolic extracts of edible mushrooms.

PubMed

Panthong, S; Boonsathorn, N; Chuchawankul, S

2016-10-17

Biological activities of various mushrooms have recently been discovered, particularly, immunomodulatory and antitumor activities. Herein, three edible mushrooms, Auricularia auricula-judae (AA), Pleurotus abalonus (PA) and Pleurotus sajor-caju (PS) extracted using Soxhlet ethanol extraction were evaluated for their antioxidative, anti-proliferative effects on leukemia cells. Using the Folin-Ciocalteau method and Trolox equivalent antioxidant capacity assay, phenolics and antioxidant activity were found in all sample mushrooms. Additionally, anti-proliferative activity of mushroom extracts against U937 leukemia cells was determined using a viability assay based on mitochondrial activity. PA (0.5 mg/mL) and AA (0.25-0.5 mg/mL) significantly reduced cell viability. Interestingly, PS caused a hormetic-like biphasic dose-response. Low doses (0-0.25 mg/L) of PS promoted cell proliferation up to 140% relative to control, whereas higher doses (0.50 mg/mL) inhibited cell proliferation. Against U937 cells, AA IC 50 was 0.28 ± 0.04 mg/mL, which was lower than PS or PA IC 50 (0.45 ± 0.01 and 0.49 ± 0.001 mg/mL, respectively). Furthermore, lactate dehydrogenase (LDH) leakage conferred cytotoxicity. PS and PA were not toxic to U937 cells at any tested concentration; AA (0.50 mg/mL) showed high LDH levels and caused 50% cytotoxicity. Additionally, UPLC-HRMS data indicated several phytochemicals known to support functional activities as either antioxidant or anti-proliferative. Glutamic acid was uniquely found in ethanolic extracts of AA, and was considered an anti-cancer amino acid with potent anti-proliferative effects on U937 cells. Collectively, all mushroom extracts exhibited antioxidant effects, but their anti-proliferative effects were dose-dependent. Nevertheless, the AA extract, with highest potency, is a promising candidate for future applications.

Human immunodeficiency virus infection of helper T cell clones. Early proliferative defects despite intact antigen-specific recognition and interleukin 4 secretion.

PubMed Central

Laurence, J; Friedman, S M; Chartash, E K; Crow, M K; Posnett, D N

1989-01-01

HIV selectively inhibited the proliferative response of clonal CD4+ T lymphocytes to alloantigen while other alloantigen-dependent responses were unperturbed. Specifically, impaired blastogenesis could be dissociated from alloantigen-specific induction of the B cell activation molecule CD23, IL-4 release, and inositol lipid hydrolysis. In addition, membrane expression of pertinent T cell receptor molecules, including CD2, CD3, and T cell antigen receptor (Ti), remained intact. Using two MHC class II-specific human CD4+ helper T cell clones, the proliferative defect was shown to be an early consequence of HIV infection, occurring within 4 d of viral inoculation and preceding increases in mature virion production. It was generalizable to three distinct methods of T cell activation, all independent of antigen-presenting cells: anti-CD3 mediated cross-linking of the CD3/Ti complex; anti-CD2 and phorbol 12-myristic 13-acetate (PMA); and anti-CD28 plus PMA. These abnormalities were not mitigated by addition of exogenous IL-2, even though expression of the IL-2 receptor (CD25) was unaltered. These studies define a selective blockade in T cell function early after HIV exposure that could serve as a model for certain in vivo manifestations of AIDS. PMID:2470786

SDF-1 is both necessary and sufficient to promote proliferative retinopathy

PubMed Central

Butler, Jason M.; Guthrie, Steven M.; Koc, Mehmet; Afzal, Aqeela; Caballero, Sergio; Brooks, H. Logan; Mames, Robert N.; Segal, Mark S.; Grant, Maria B.; Scott, Edward W.

2005-01-01

Diabetic retinopathy is the leading cause of blindness in working-age adults. It is caused by oxygen starvation in the retina inducing aberrant formation of blood vessels that destroy retinal architecture. In humans, vitreal stromal cell–derived factor–1 (SDF-1) concentration increases as proliferative diabetic retinopathy progresses. Treatment of patients with triamcinolone decreases SDF-1 levels in the vitreous, with marked disease improvement. SDF-1 induces human retinal endothelial cells to increase expression of VCAM-1, a receptor for very late antigen–4 found on many hematopoietic progenitors, and reduce tight cellular junctions by reducing occludin expression. Both changes would serve to recruit hematopoietic and endothelial progenitor cells along an SDF-1 gradient. We have shown, using a murine model of proliferative adult retinopathy, that the majority of new vessels formed in response to oxygen starvation originate from hematopoietic stem cell–derived endothelial progenitor cells. We now show that the levels of SDF-1 found in patients with proliferative retinopathy induce retinopathy in our murine model. Intravitreal injection of blocking antibodies to SDF-1 prevented retinal neovascularization in our murine model, even in the presence of exogenous VEGF. Together, these data demonstrate that SDF-1 plays a major role in proliferative retinopathy and may be an ideal target for the prevention of proliferative retinopathy. PMID:15630447

In vitro study of platelets and circulating mononuclear cells of subjects presenting an intolerance to aspirin.

PubMed

Guez, S; Gualde, N; Bezian, J H; Cabanieu, G

1992-01-01

Aspirine-sensitive asthma (ASA) is a disease defined only by clinical criteria. It is an intrinsic asthma related to a hypersensitivity to aspirin. The illness is linked to abnormalities in platelet and macrophage arachidonic acid metabolism. We assessed in vitro the platelet chemoluminescence (CL) and lymphocyte proliferative response of ASA patients. We observed that platelets from patients and control do not generate any CL in the presence of aspirin. Concerning the proliferative response of lymphocytes, the in vitro effect of aspirin depends upon the origin of the lymphocytes tested. Thus aspirin clearly enhances the proliferative response of lymphocytes from normal subjects but diminishes the thymidine uptake by lymphocytes from ASA patients. This discrepancy in the in vitro response of lymphocytes from normal subjects and patients might be useful for in vitro diagnosis of ASA.

Enhanced clonal burst size corrects an otherwise defective memory response by CD8+ recent thymic emigrants

PubMed Central

Deets, Katherine A.; Berkley, Amy M.; Bergsbaken, Tessa; Fink, Pamela J.

2016-01-01

The youngest peripheral T cells (recent thymic emigrants or RTEs) are functionally distinct from naïve T cells that have completed post-thymic maturation. We now assess the RTE memory response, and find that RTEs produced less granzyme B than their mature counterparts during infection, but proliferated more and therefore generated equivalent target killing in vivo. After infection, RTE numbers contracted less dramatically than those of mature T cells, but RTEs were delayed in their transition to central memory, displaying impaired expression of CD62L, IL-2, Eomesodermin, and CXCR4, which resulted in impaired bone marrow localization. RTE-derived and mature memory cells expanded equivalently during rechallenge, indicating the robust proliferative capacity of RTEs was maintained independently of central memory phenotype. Thus, the diminished effector function and delayed central memory differentiation of RTE-derived memory cells are counterbalanced by their increased proliferative capacity, driving the efficacy of the RTE response to that of mature T cells. PMID:26873989

Cutting Edge: Enhanced Clonal Burst Size Corrects an Otherwise Defective Memory Response by CD8+ Recent Thymic Emigrants.

PubMed

Deets, Katherine A; Berkley, Amy M; Bergsbaken, Tessa; Fink, Pamela J

2016-03-15

The youngest peripheral T cells (recent thymic emigrants [RTEs]) are functionally distinct from naive T cells that have completed postthymic maturation. We assessed the RTE memory response and found that RTEs produced less granzyme B than their mature counterparts during infection but proliferated more and, therefore, generated equivalent target killing in vivo. Postinfection, RTE numbers contracted less dramatically than those of mature T cells, but RTEs were delayed in their transition to central memory, displaying impaired expression of CD62L, IL-2, Eomesodermin, and CXCR4, which resulted in impaired bone marrow localization. RTE-derived and mature memory cells expanded equivalently during rechallenge, indicating that the robust proliferative capacity of RTEs was maintained independently of central memory phenotype. Thus, the diminished effector function and delayed central memory differentiation of RTE-derived memory cells are counterbalanced by their increased proliferative capacity, driving the efficacy of the RTE response to that of mature T cells. Copyright © 2016 by The American Association of Immunologists, Inc.

Retinol induces morphological alterations and proliferative focus formation through free radical-mediated activation of multiple signaling pathways.

PubMed

Gelain, Daniel Pens; Pasquali, Matheus Augusto de Bittencourt; Caregnato, Fernanda Freitas; Castro, Mauro Antonio Alves; Moreira, José Claudio Fonseca

2012-04-01

Toxicity of retinol (vitamin A) has been previously associated with apoptosis and/or cell malignant transformation. Thus, we investigated the pathways involved in the induction of proliferation, deformation and proliferative focus formation by retinol in cultured Sertoli cells of rats. Sertoli cells were isolated from immature rats and cultured. The cells were subjected to a 24-h treatment with different concentrations of retinol. Parameters of oxidative stress and cytotoxicity were analyzed. The effects of the p38 inhibitor SB203580 (10 μmol/L), the JNK inhibitor SP600125 (10 μmol/L), the Akt inhibitor LY294002 (10 μmol/L), the ERK inhibitor U0126 (10 μmol/L) the pan-PKC inhibitor Gö6983 (10 μmol/L) and the PKA inhibitor H89 (1 μmol/L) on morphological and proliferative/transformation-associated modifications were studied. Retinol (7 and 14 μmol/L) significantly increases the reactive species production in Sertoli cells. Inhibition of p38, JNK, ERK1/2, Akt, and PKA suppressed retinol-induced [(3)H]dT incorporation into the cells, while PKC inhibition had no effect. ERK1/2 and p38 inhibition also blocked retinol-induced proliferative focus formation in the cells, while Akt and JNK inhibition partially decreased focus formation. ERK1/2 and p38 inhibition hindered transformation-associated deformation in retinol-treated cells, while other treatments had no effect. Our results suggest that activation of multiple kinases is responsible for morphological and proliferative changes associated to malignancy development in Sertoli cells by retinol at the concentrations higher than physiological level.

Bovine central memory T cells are highly proliferative in response to bovine tuberculosis infection

USDA-ARS?s Scientific Manuscript database

Long-term (i.e., 14 days) cultured IFN-gamma ELISPOT assays measure central memory T cell (Tcm) responses in both humans and cattle. With bovine tuberculosis, vaccine-elicited long-term IFN-gamma ELISPOT responses correlate with protection. In other species, Tcm’s pose low activation threshold and a...

T-dependence of human B lymphocyte proliferative response to mitogens.

PubMed

Brochier, J; Samarut, C; Gueho, J P; Revillard, J P

1976-01-01

Human peripheral blood and tonsil lymphocytes were fractionated on anti-Ig-coated Sephadex columns or by centrifugation after rosetting with native sheep erythrocytes. Both methods allowed the recovery of B and T-enriched populations the purity of which was checked by fluorescein-labelled anti-Ig serum, E and EAC rosette formation, and heterologous antisera specific for B or T lymphocytes. The proliferative response of T cells to PHA, Con A, PWM, and ALS was not found different from that of unfractionated cells, whereas no response of the B cells could be observed to these mitogens providing that no contaminating T cells were present. Addition of T lymphocytes to these unresponsive B cells allowed them to respond to phytomitogens, but not to ALS. X-irradiated T cells could, to some extent, replace the diving T lymphocytes; no T-replacing factor could be found in cell-free supernatants from T cells, whether or not they had been activated by mitrogens. This model of B-T cooperation appears useful for studying the differentiation and maturation of human B lymphocytes.

ESR1 mutations affect anti-proliferative responses to tamoxifen through enhanced cross-talk with IGF signaling.

PubMed

Gelsomino, Luca; Gu, Guowei; Rechoum, Yassine; Beyer, Amanda R; Pejerrey, Sasha M; Tsimelzon, Anna; Wang, Tao; Huffman, Kenneth; Ludlow, Andrew; Andò, Sebastiano; Fuqua, Suzanne A W

2016-06-01

The purpose of this study was to address the role of ESR1 hormone-binding mutations in breast cancer. Soft agar anchorage-independent growth assay, Western blot, ERE reporter transactivation assay, proximity ligation assay (PLA), coimmunoprecipitation assay, silencing assay, digital droplet PCR (ddPCR), Kaplan-Meier analysis, and statistical analysis. It is now generally accepted that estrogen receptor (ESR1) mutations occur frequently in metastatic breast cancers; however, we do not yet know how to best treat these patients. We have modeled the three most frequent hormone-binding ESR1 (HBD-ESR1) mutations (Y537N, Y537S, and D538G) using stable lentiviral transduction in human breast cancer cell lines. Effects on growth were examined in response to hormonal and targeted agents, and mutation-specific changes were studied using microarray and Western blot analysis. We determined that the HBD-ESR1 mutations alter anti-proliferative effects to tamoxifen (Tam), due to cell-intrinsic changes in activation of the insulin-like growth factor receptor (IGF1R) signaling pathway and levels of PIK3R1/PIK3R3. The selective estrogen receptor degrader, fulvestrant, significantly reduced the anchorage-independent growth of ESR1 mutant-expressing cells, while combination treatments with the mTOR inhibitor everolimus, or an inhibitor blocking IGF1R, and the insulin receptor significantly enhanced anti-proliferative responses. Using digital drop (dd) PCR, we identified mutations at high frequencies ranging from 12 % for Y537N, 5 % for Y537S, and 2 % for D538G in archived primary breast tumors from women treated with adjuvant mono-tamoxifen therapy. The HBD-ESR1 mutations were not associated with recurrence-free or overall survival in response in this patient cohort and suggest that knowledge of other cell-intrinsic factors in combination with ESR1 mutation status will be needed determine anti-proliferative responses to Tam.

Role of P-glycoprotein on CD69+CD4+ cells in the pathogenesis of proliferative lupus nephritis and non-responsiveness to immunosuppressive therapy

PubMed Central

Tsujimura, Shizuyo; Adachi, Tomoko; Saito, Kazuyoshi; Tanaka, Yoshiya

2017-01-01

Introduction P-glycoprotein (P-gp) expression on activated lymphocytes in systemic lupus erythematosus (SLE) plays a role in active efflux of intracellular drugs, resulting in drug resistance. The role of P-gp-expressing lymphocytes in the pathogenesis of SLE remains unclear. The aim of this study was to determine the importance of P-gp+CD4+ cells in organ manifestations in refractory SLE. Methods The proportion of P-gp+CD4+ cells was determined by flow cytometry in peripheral blood of patients with SLE (n=116) and healthy adults (n=10). Renal biopsy specimens were examined by immunohistochemistry for P-gp expression. Results CD69 is a marker of CD4 cell activation. The proportion of both P-gp-expressing CD4+ cells and CD69-expressing CD4+ cells in peripheral blood was higher in SLE than control. The proportion of P-gp+CD69+CD4+ cells correlated with Systemic Lupus Erythematosus Disease Activity Index and was higher in poor responders to corticosteroids. Furthermore, the proportion of P-gp+CD69+CD4+ cells was significantly higher in proliferative lupus nephritis (LN) with poor response to corticosteroids. The efficacy of immunosuppressive therapy depended on the regulation of the proportion of P-gp+CD69+CD4+ cells. Marked accumulation of P-gp+CD4+ cells in renal interstitial tissue and high proportion of peripheral P-gp+CD69+CD4+ cells were noted in patients with proliferative LN. Conclusions The results showed high proportion of P-gp+CD69+CD4+ cells in peripheral blood and their accumulation in renal tissue in patients with proliferative LN refractory to CS therapy, suggesting that P-gp expression on activated CD4+ T cells is a potentially useful marker for refractoriness to treatment and a novel target for treatment. PMID:29225917

Role of P-glycoprotein on CD69+CD4+ cells in the pathogenesis of proliferative lupus nephritis and non-responsiveness to immunosuppressive therapy.

PubMed

Tsujimura, Shizuyo; Adachi, Tomoko; Saito, Kazuyoshi; Tanaka, Yoshiya

2017-01-01

P-glycoprotein (P-gp) expression on activated lymphocytes in systemic lupus erythematosus (SLE) plays a role in active efflux of intracellular drugs, resulting in drug resistance. The role of P-gp-expressing lymphocytes in the pathogenesis of SLE remains unclear. The aim of this study was to determine the importance of P-gp + CD4 + cells in organ manifestations in refractory SLE. The proportion of P-gp + CD4 + cells was determined by flow cytometry in peripheral blood of patients with SLE (n=116) and healthy adults (n=10). Renal biopsy specimens were examined by immunohistochemistry for P-gp expression. CD69 is a marker of CD4 cell activation. The proportion of both P-gp-expressing CD4 + cells and CD69-expressing CD4 + cells in peripheral blood was higher in SLE than control. The proportion of P-gp + CD69 + CD4 + cells correlated with Systemic Lupus Erythematosus Disease Activity Index and was higher in poor responders to corticosteroids. Furthermore, the proportion of P-gp + CD69 + CD4 + cells was significantly higher in proliferative lupus nephritis (LN) with poor response to corticosteroids. The efficacy of immunosuppressive therapy depended on the regulation of the proportion of P-gp + CD69 + CD4 + cells. Marked accumulation of P-gp + CD4 + cells in renal interstitial tissue and high proportion of peripheral P-gp + CD69 + CD4 + cells were noted in patients with proliferative LN. The results showed high proportion of P-gp + CD69 + CD4 + cells in peripheral blood and their accumulation in renal tissue in patients with proliferative LN refractory to CS therapy, suggesting that P-gp expression on activated CD4 + T cells is a potentially useful marker for refractoriness to treatment and a novel target for treatment.

Power Frequency Magnetic Fields Affect the p38 MAPK-Mediated Regulation of NB69 Cell Proliferation Implication of Free Radicals.

PubMed

Martínez, María Antonia; Úbeda, Alejandro; Moreno, Jorge; Trillo, María Ángeles

2016-04-06

The proliferative response of the neuroblastoma line NB69 to a 100 µT, 50 Hz magnetic field (MF) has been shown mediated by activation of the MAPK-ERK1/2 pathway. This work investigates the MF effect on the cell cycle of NB69, the participation of p38 and c-Jun N-terminal (JNK) kinases in the field-induced proliferative response and the potential involvement of reactive oxygen species (ROS) in the activation of the MAPK-ERK1/2 and -p38 signaling pathways. NB69 cultures were exposed to the 100 µT MF, either intermittently for 24, 42 or 63 h, or continuously for periods of 15 to 120 min, in the presence or absence of p38 or JNK inhibitors: SB203580 and SP600125, respectively. Antioxidant N-acetylcysteine (NAC) was used as ROS scavenger. Field exposure induced transient activation of p38, JNK and ERK1/2. The MF proliferative effect, which was mediated by changes in the cell cycle, was blocked by the p38 inhibitor, but not by the JNK inhibitor. NAC blocked the field effects on cell proliferation and p38 activation, but not those on ERK1/2 activation. The MF-induced proliferative effects are exerted through sequential upregulation of MAPK-p38 and -ERK1/2 activation, and they are likely mediated by a ROS-dependent activation of p38.

Power Frequency Magnetic Fields Affect the p38 MAPK-Mediated Regulation of NB69 Cell Proliferation Implication of Free Radicals

PubMed Central

Martínez, María Antonia; Úbeda, Alejandro; Moreno, Jorge; Trillo, María Ángeles

2016-01-01

The proliferative response of the neuroblastoma line NB69 to a 100 µT, 50 Hz magnetic field (MF) has been shown mediated by activation of the MAPK-ERK1/2 pathway. This work investigates the MF effect on the cell cycle of NB69, the participation of p38 and c-Jun N-terminal (JNK) kinases in the field-induced proliferative response and the potential involvement of reactive oxygen species (ROS) in the activation of the MAPK-ERK1/2 and -p38 signaling pathways. NB69 cultures were exposed to the 100 µT MF, either intermittently for 24, 42 or 63 h, or continuously for periods of 15 to 120 min, in the presence or absence of p38 or JNK inhibitors: SB203580 and SP600125, respectively. Antioxidant N-acetylcysteine (NAC) was used as ROS scavenger. Field exposure induced transient activation of p38, JNK and ERK1/2. The MF proliferative effect, which was mediated by changes in the cell cycle, was blocked by the p38 inhibitor, but not by the JNK inhibitor. NAC blocked the field effects on cell proliferation and p38 activation, but not those on ERK1/2 activation. The MF-induced proliferative effects are exerted through sequential upregulation of MAPK-p38 and -ERK1/2 activation, and they are likely mediated by a ROS-dependent activation of p38. PMID:27058530

DNA damage signaling regulates age-dependent proliferative capacity of quiescent inner ear supporting cells

PubMed Central

Laos, Maarja; Anttonen, Tommi; Kirjavainen, Anna; Hällström, Taija af; Laiho, Marikki; Pirvola, Ulla

2014-01-01

Supporting cells (SCs) of the cochlear (auditory) and vestibular (balance) organs hold promise as a platform for therapeutic regeneration of the sensory hair cells. Prior data have shown proliferative restrictions of adult SCs forced to re-enter the cell cycle. By comparing juvenile and adult SCs in explant cultures, we have here studied how proliferative restrictions are linked with DNA damage signaling. Cyclin D1 overexpression, used to stimulate cell cycle re-entry, triggered higher proliferative activity of juvenile SCs. Phosphorylated form of histone H2AX (γH2AX) and p53 binding protein 1 (53BP1) were induced in a foci-like pattern in SCs of both ages as an indication of DNA double-strand break formation and activated DNA damage response. Compared to juvenile SCs, γH2AX and the repair protein Rad51 were resolved with slower kinetics in adult SCs, accompanied by increased apoptosis. Consistent with the in vitro data, in a Rb mutant mouse model in vivo, cell cycle re-entry of SCs was associated with γH2AX foci induction. In contrast to cell cycle reactivation, pharmacological stimulation of SC-to-hair-cell transdifferentiation in vitro did not trigger γH2AX. Thus, DNA damage and its prolonged resolution are critical barriers in the efforts to stimulate proliferation of the adult inner ear SCs. PMID:25063730

A Hydrogel-Endothelial Cell implant Mimics Infantile Hemangioma: Modulation by Survivin and the Hippo pathway*

PubMed Central

Tsuneki, Masayuki; Hardee, Steven; Michaud, Michael; Morotti, Raffaella; Lavik, Erin; Madri, Joseph A.

2015-01-01

Microvascular endothelial cells cultured in three-dimensional hydrogel scaffolds form a network of microvessel structures when implanted subcutaneously in mice, inosculate with host vessels and over time remodel into large ectatic vascular structures resembling hemangiomas. When compared to infantile hemaniomas similarities were noted including a temporal progression from a morphological appearance of a proliferative phase to the appearance of an involuted phase mimicking the proliferative and involutional phases of infantile hemangioma. Consistent with the progression of a proliferative phase to an involuted phase, both the murine implants and human biopsy tissue exhibit reduced expression of Ajuba, YAP and Survivin labeling as they progressed over time. Significant numbers of CD45+, CD11b+, Mac3+ mononuclear cells were found at the 2 week time point in our implant model which correlated with the presence of CD45+, CD68+ mononuclear cells observed in biopsies of human proliferative phase hemangiomas. At the 4 week time point in our implant model only small numbers of CD45+ cells were detected, which again correlated with our findings of significantly diminished CD45+, CD68+ mononuclear cells in human involutional phase hemangiomas. The demonstration of mononuclear cell infiltration transiently in the proliferative phase of these lesions suggests that the vascular proliferation and/or regression may be driven in part by an immune response. Gross and microscopic morphological appearances of human proliferative and involutional hemangiomas and our implant model correlate well with each other as do the expression levels of Hippo pathway components (Ajuba and YAP) and Survivin and correlate with proliferation in these entities. Inhibitors of Survivin and Ajuba (which we have demonstrated to inhibit proliferation and increase apoptosis in murine hemangioma cell tissue culture) may have potential as other beneficial treatments for proliferating infantile hemangiomas. This implant model may have potential as a modest through-put screen for testing and development of therapeutics targeted at the proliferative phase of infantile hemangiomas, reducing the subsequent post-involutional scarring sometimes associated with these lesions. PMID:25961170

The Aryl Hydrocarbon Receptor Mediates Leflunomide-Induced Growth Inhibition of Melanoma Cells

PubMed Central

O’Donnell, Edmond F.; Kopparapu, Prasad Rao; Koch, Daniel C.; Jang, Hyo Sang; Phillips, Jessica Lynne; Tanguay, Robert L.; Kerkvliet, Nancy I.; Kolluri, Siva Kumar

2012-01-01

A novel role of the dihydroorotatedehydrogenase (DHODH) inhibitor leflunomide as a potential anti-melanoma therapy was recently reported (Nature 471∶518-22, 2011). We previously reported that leflunomide strongly activates the transcriptional activity of the Aryl Hydrocarbon Receptor (AhR). We therefore tested whether the AhR regulates the anti-proliferative effects of leflunomide in melanoma. We first evaluated the expression of AhR in melanoma cells and found that AhR is highly expressed in A375 melanoma as well as in several other cancer cell types. To evaluate whether AhR plays a role in regulating the growth inhibitory effects of leflunomide in A375 cells, we generated a stable cell line from parental A375 cells expressing a doxycycline (DOX) inducible AhR shRNA. Using these cells in the absence or presence of DOX (normal AhR levels or AhR-knockdown, respectively) we found that the anti-proliferative effects of leflunomide, but not its metabolite A771726, were strongly dependent upon AhR expression. It has been well established that supplementation of cells with exogenous uridine completely rescues the anti-proliferative effects due to DHODH inhibition. Thus, we performed uridine rescue experiments in A375 cells to determine whether the anti-proliferative effects of leflunomide are solely due to DHODH inhibition as previously reported. Interestingly, saturating levels of uridine only modestly rescued A375 cells from the anti-proliferative effects of both leflunomide and A771726, indicating additional mechanism(s), apart from DHODH inhibition are responsible for the anti-proliferative effects of leflunomide in melanoma cells. Uridine also did not rescue MDA-MB-435S melanoma cell proliferation after leflunomide treatment. Our results reveal that the AhR is a molecular target of leflunomide and support the feasibility of the clinical application of leflunomide for treating melanoma. Furthermore, analysis of expression data from 967 cancer cell lines revealed that AhR is expressed in multiple different cancer types supporting the intriguing possibility of targeting the AhR for therapy in a number of cancers. PMID:22815870

Insulin-sensitizing and Anti-proliferative Effects of Argania spinosa Seed Extracts

PubMed Central

Samane, Samira; Noël, Josette; Charrouf, Zoubida; Amarouch, Hamid; Haddad, Pierre Selim

2006-01-01

Argania spinosa is an evergreen tree endemic of southwestern Morocco. Many preparations have been used in traditional Moroccan medicine for centuries to treat several illnesses including diabetes. However, scientific evidence supporting these actions is lacking. Therefore, we prepared various extracts of the argan fruit, namely keel, cake and argan oil extracts, which we tested in the HTC hepatoma cell line for their potential to affect cellular insulin responses. Cell viability was measured by Trypan Blue exclusion and the response to insulin evaluated by the activation of the extracellular regulated kinase (ERK1/2), ERK kinase (MEK1/2) and protein kinase B (PKB/Akt) signaling components. None of the extracts demonstrated significant cytotoxic activity. Certain extracts demonstrated a bi-phasic effect on ERK1/2 activation; low doses of the extract slightly increased ERK1/2 activation in response to insulin, whereas higher doses completely abolished the response. In contrast, none of the extracts had any significant effect on MEK whereas only a cake saponin subfraction enhanced insulin-induced PKB/Akt activation. The specific action of argan oil extracts on ERK1/2 activation made us consider an anti-proliferative action. We have thus tested other transformed cell lines (HT-1080 and MSV-MDCK-INV cells) and found similar results. Inhibition of ERK1/2 activation was also associated with decreased DNA synthesis as evidenced by [3H]thymidine incorporation experiments. These results suggest that the products of Argania spinosa may provide a new therapeutic avenue against proliferative diseases. PMID:16951716

Multi-scale modeling of APC and [Formula: see text]-catenin regulation in the human colonic crypt.

PubMed

Emerick, Brooks; Schleiniger, Gilberto; Boman, Bruce M

2018-06-01

Stem cell renewal and differentiation in the human colonic crypt are linked to the [Formula: see text]-catenin pathway. The spatial balance of Wnt factors in proliferative cells within the crypt maintain an appropriate level of cellular reproduction needed for normal crypt homeostasis. Mutational events at the gene level are responsible for deregulating the balance of Wnt factors along the crypt, causing an overpopulation of proliferative cells, a loss of structure of the crypt domain, and the initiation of colorectal carcinomas. We formulate a PDE model describing cell movement and reproduction in a static crypt domain. We consider a single cell population whose proliferative capabilities are determined by stemness, a quantity defined by intracellular levels of adenomatous polyposis coli (APC) scaffold protein and [Formula: see text]-catenin. We fit APC regulation parameters to biological data that describe normal protein gradients in the crypt. We also fit cell movement and protein flux parameters to normal crypt characteristics such as renewal time, total cell count, and proportion of proliferating cells. The model is used to investigate abnormal crypt dynamics when subjected to a diminished APC gradient, a scenario synonymous to mutations in the APC gene. We find that a 25% decrease in APC synthesis leads to a fraction of 0.88 proliferative, which is reflective of normal-appearing FAP crypts. A 50% drop in APC activity yields a fully proliferative crypt showing a doubling of the level of stemness, which characterizes the initial stages of colorectal cancer development. A sensitivity analysis of APC regulation parameters shows the perturbation of factors that is required to restore crypt dynamics to normal in the case of APC mutations.

Lactobacillus GG has in vitro effects on enhanced interleukin-10 and interferon-gamma release of mononuclear cells but no in vivo effects in supplemented mothers and their neonates.

PubMed

Kopp, M V; Goldstein, M; Dietschek, A; Sofke, J; Heinzmann, A; Urbanek, R

2008-04-01

The value of probiotics for primary prevention is controversial. Moreover, only little is known about the underlying immunological mechanisms of action. Therefore, we assessed the proliferative response and cytokine release in cultures of isolated mononuclear cells from pregnant women and their neonates supplemented with Lactobacillus GG (LGG) or placebo. In a double-blind, placebo-controlled prospective trial, pregnant women with at least one first-degree relative or a partner with an atopic disease were randomly assigned to receive either the probiotic LGG (ATCC 53103; 5 x 10(9) colony-forming units LGG twice daily) or placebo 4-6 weeks before expected delivery, followed by a post-natal period of 6 months. Cord blood mononuclear cells (CBMC) and peripheral blood mononuclear cells (PBMC) of the corresponding mother were isolated from cord blood and peripheral blood (n=68). The proliferative response of CBMC and PBMC was expressed as the stimulation index (SI), which was calculated according to the ratio between the mean counts per minute (c.p.m.) values measured in the wells with stimulated cells and the mean c.p.m. values measured in the wells with unstimulated cells. Additionally, the cytokines IFN-gamma, IL-10 and IL-13 in the cell culture supernatants were measured using the ELISA technique. No difference was observed between the LGG-supplemented group and the placebo group in terms of the proliferative capacity of maternal or neonatal cord blood cells in response to IL-2, beta-lactoglobulin or LGG. In vitro stimulation with LGG resulted in significantly enhanced release of IL-10 and IFN-gamma, compared with cytokine release in unstimulated controls. However, this phenomenon was observed in supernatants of maternal and neonatal MC in both groups, independent of prior supplementation with LGG. LGG has in vitro effects on enhanced IL-10 and IFN-gamma release of mononuclear cells. However, supplementation with LGG during pregnancy did not alter the proliferative capacity or cytokine pattern in their recipients.

Slc3a2 Mediates Branched-Chain Amino-Acid-Dependent Maintenance of Regulatory T Cells.

PubMed

Ikeda, Kayo; Kinoshita, Makoto; Kayama, Hisako; Nagamori, Shushi; Kongpracha, Pornparn; Umemoto, Eiji; Okumura, Ryu; Kurakawa, Takashi; Murakami, Mari; Mikami, Norihisa; Shintani, Yasunori; Ueno, Satoko; Andou, Ayatoshi; Ito, Morihiro; Tsumura, Hideki; Yasutomo, Koji; Ozono, Keiichi; Takashima, Seiji; Sakaguchi, Shimon; Kanai, Yoshikatsu; Takeda, Kiyoshi

2017-11-14

Foxp3 + regulatory T (Treg) cells, which suppress immune responses, are highly proliferative in vivo. However, it remains unclear how the active replication of Treg cells is maintained in vivo. Here, we show that branched-chain amino acids (BCAAs), including isoleucine, are required for maintenance of the proliferative state of Treg cells via the amino acid transporter Slc3a2-dependent metabolic reprogramming. Mice fed BCAA-reduced diets showed decreased numbers of Foxp3 + Treg cells with defective in vivo proliferative capacity. Mice lacking Slc3a2 specifically in Foxp3 + Treg cells showed impaired in vivo replication and decreased numbers of Treg cells. Slc3a2-deficient Treg cells showed impaired isoleucine-induced activation of the mTORC1 pathway and an altered metabolic state. Slc3a2 mutant mice did not show an isoleucine-induced increase of Treg cells in vivo and exhibited multi-organ inflammation. Taken together, these findings demonstrate that BCAA controls Treg cell maintenance via Slc3a2-dependent metabolic regulation. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Desmoglein 3-specific T regulatory 1 cells consist of two subpopulations with differential expression of the transcription factor Foxp3

PubMed Central

Veldman, Christian; Pahl, Andreas; Hertl, Michael

2009-01-01

Pemphigus vulgaris (PV) is an autoimmune bullous skin disorder associated with autoantibodies against desmoglein (Dsg) 3. An imbalance of type 1 regulatory T (Tr1) cells and T helper type 2 (Th2) cells specific for Dsg3 may be critical for the loss of tolerance against Dsg3 in PV. Within the population of Dsg3-responsive, interleukin (IL)-10-secreting Tr1 cell clones, two major subpopulations were identified and sorted by fluorescence-activated cell sorting (FACS) based on their size and granularity. Upon in vitro culture, the larger subpopulation differentiated back into the two former subpopulations of the Tr1 cell clones, while the smaller subpopulation died within 2 weeks. The smaller subpopulation of the Tr1 cell clones was characterized by the expression of Foxp3, the secretion of IL-10, transforming growth factor (TGF)-β and IL-5 upon stimulation with Dsg3, a proliferative response to IL-2 but not to Dsg3 or mitogenic stimuli, and an inhibitory effect on the proliferative response of Dsg3-responsive Th clones in a Dsg3-specific manner. In contrast, the larger subpopulation showed a Th-like phenotype, lacking Foxp3, cytotoxic T-lymphocyte antigen 4 (CTLA4) and glucocorticoid-induced tumour necrosis factor receptor (GITR) expression and IL-2 secretion, and did not mount a proliferative response to Dsg3 and mitogenic stimuli. The two Tr1 subpopulations showed expression of identical T-cell receptor (TCR) Vβ chains which varied among the PV patients studied. Upon inhibition of Foxp3, the smaller Tr1 subpopulation developed a proliferate response to Dsg3 and mitogenic stimuli, no longer suppressed Dsg3-specific Th cells, lost expression of GITR and CTLA4 and secreted IL-2. Thus, our observations suggest a distinct relationship between Dsg3-specific Tr1 and Th-like cells which may be critical for the continuous generation and survival of Dsg3-specific Tr1 cells. PMID:18800988

Lymphocyte proliferative responses of goats vaccinated with Brucella melitensis 16M or a delta purE201 strain.

PubMed Central

Olsen, S C; Cheville, N F; Stevens, M G; Houng, H H; Drazek, E S; Hadfield, T L; Warren, R L; Hoover, D L

1997-01-01

The response to a Brucella melitensis purEK deletion mutant, delta purE201 (referred to as strain 201), was compared with the response to its parental strain, 16M, in juvenile goats. Proliferative responses to gamma-irradiated bacteria were detected earlier in strain 201-infected goats. Lymphocytes from strain 16M- or 201-infected goats proliferated in response to one-dimensional polyacrylamide gel electrophoresis-separated proteins of similar mass isolated from strain 16M or Brucella abortus RB51. Data from this study suggest that some antigens stimulating cell-mediated responses are conserved among Brucella species, as 201- and 16M-infected goats recognized similar proteins expressed by RB51 and 16M. PMID:9199478

Electric stimulation at 448 kHz promotes proliferation of human mesenchymal stem cells.

PubMed

Hernández-Bule, María Luisa; Paíno, Carlos Luis; Trillo, María Ángeles; Úbeda, Alejandro

2014-01-01

Capacitive-resistive electric transfer (CRET) is a non invasive electrothermal therapy that applies electric currents within the 400 kHz - 450 kHz frequency range to the treatment of musculoskeletal lesions. Evidence exists that electric currents and electric or magnetic fields can influence proliferative and/or differentiating processes involved in tissue regeneration. This work investigates proliferative responses potentially underlying CRET effects on tissue repair. XTT assay, flow cytometry, immunofluorescence and Western Blot analyses were conducted to asses viability, proliferation and differentiation of adipose-derived stem cells (ADSC) from healthy donors, after short, repeated (5 m On/4 h Off) in vitro stimulation with a 448-kHz electric signal currently used in CRET therapy, applied at a subthermal dose of 50 μA/mm(2) RESULTS: The treatment induced PCNA and ERK1/2 upregulation, together with significant increases in the fractions of ADSC undergoing cycle phases S, G2 and M, and enhanced cell proliferation rate. This proliferative effect did not compromise the multipotential ability of ADSC for subsequent adipogenic, chondrogenic or osteogenic differentiation. These data identify cellular and molecular phenomena potentially underlying the response to CRET and indicate that CRET-induced lesion repair could be mediated by stimulation of the proliferation of stem cells present in the injured tissues. © 2014 S. Karger AG, Basel.

HIV-Infected Children Have Elevated Levels of PD-1+ Memory CD4 T Cells With Low Proliferative Capacity and High Inflammatory Cytokine Effector Functions.

PubMed

Foldi, Julia; Kozhaya, Lina; McCarty, Bret; Mwamzuka, Mussa; Marshed, Fatma; Ilmet, Tiina; Kilberg, Max; Kravietz, Adam; Ahmed, Aabid; Borkowsky, William; Unutmaz, Derya; Khaitan, Alka

2017-09-15

During human immunodeficiency virus (HIV) disease, chronic immune activation leads to T-cell exhaustion. PD-1 identifies "exhausted" CD8 T cells with impaired HIV-specific effector functions, but its role on CD4 T cells and in HIV-infected children is poorly understood. In a Kenyan cohort of vertically HIV-infected children, we measured PD-1+ CD4 T-cell frequencies and phenotype by flow cytometry and their correlation with HIV disease progression and immune activation. Second, in vitro CD4 T-cell proliferative and cytokine responses to HIV-specific and -nonspecific stimuli were assessed with and without PD-1 blockade. HIV-infected children have increased frequencies of PD-1+ memory CD4 T cells that fail to normalize with antiretroviral treatment. These cells are comprised of central and effector memory subsets and correlate with HIV disease progression, measured by viral load, CD4 percentage, CD4:CD8 T-cell ratio, and immune activation. Last, PD-1+ CD4 T cells predict impaired proliferative potential yet preferentially secrete the Th1 and Th17 cytokines interferon-γ and interleukin 17A, and are unresponsive to in vitro PD-1 blockade. This study highlights differences in PD-1+ CD4 T-cell memory phenotype and response to blockade between HIV-infected children and adults, with implications for potential immune checkpoint therapies. © 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.

Age-Dependent Netrin-1 Signaling Regulates NG2+ Glial Cell Spatial Homeostasis in Normal Adult Gray Matter

PubMed Central

Birey, Fikri

2015-01-01

Neuron–glial antigen 2-positive (NG2+) glial cells are the most proliferative glia type in the adult CNS, and their tile-like arrangement in adult gray matter is under tight regulation. However, little is known about the cues that govern this unique distribution. To this end, using a NG2+ glial cell ablation model in mice, we examined the repopulation dynamics of NG2+ glial cells in the mature and aged mice gray matter. We found that some resident NG2+ glial cells that escaped depletion rapidly enter the cell cycle to repopulate the cortex with altered spatial distribution. We reveal that netrin-1 signaling is involved in the NG2+ glial cell early proliferative, late repopulation, and distribution response after ablation in the gray matter. However, ablation of NG2+ glial cell in older animals failed to stimulate a similar repopulation response, possibly because of a decrease in the sensitivity to netrin-1. Our findings indicate that endogenous netrin-1 plays a role in NG2+ glial cell homeostasis that is distinct from its role in myelination. PMID:25926469

Cell-Autonomous Control of IL-7 Response Revealed In a Novel Stage of Precursor B Cells

PubMed Central

Sandoval, Gabriel J.; Graham, Daniel B.; Bhattacharya, Deepta; Sleckman, Barry P.; Xavier, Ramnik J.; Swat, Wojciech

2013-01-01

During early stages of B-lineage differentiation in bone marrow, signals emanating from IL-7 receptor and pre-B cell receptor (pre-BCR) are thought to synergistically induce proliferative expansion of progenitor cells. Paradoxically, loss of pre-BCR signaling components is associated with leukemia in both mice and humans. Exactly how progenitor B cells perform the task of balancing proliferative burst dependent on IL-7 with the termination of IL-7 signals and the initiation of LC gene rearrangement remains to be elucidated. In this report, we provide genetic and functional evidence that the cessation of IL-7 response of pre-B cells is controlled via a cell-autonomous mechanism that operates at a discreet developmental transition inside Fraction C’ (Large Pre-BII) marked by transient expression of c-Myc. Our data indicates that pre-BCR cooperates with IL-7R in expanding pre-B cell pool, but it is also critical to control differentiation program shutting off c-Myc gene in large pre-B cells. PMID:23420891

Toxocara canis adult worm antigen induces proliferative response of healthy human peripheral blood mononuclear cells.

PubMed

Inuo, G; Akao, N; Kohsaka, H; Saito, I; Miyasaka, N; Fujita, K

1995-02-01

The proliferative response of human peripheral blood mononuclear cells (PBMC) from healthy donors to Toxocara canis adult worm antigens (TcA) was examined. PBMC from all donors examined (n = 7) strongly responded to TcA in a dose-dependent fashion after six days of culture, irrespective of their serological reactivity. In contrast, cord blood mononuclear cells did not react to TcA. The proliferation of PBMC in response to TcA was completely inhibited by anti-HLA-DR antibody. Purified CD4+ T cells reconstituted with autologous irradiated antigen presenting cells (APC) vigorously proliferated in response to TcA, but this was abrogated by pretreatment of APC with paraformaldehyde. Significant IL-2, IL-3, IL-4, IL-5 and IFN-gamma mRNA expression was detected in PBMC stimulated with TcA, with expression peaking at 72 h after stimulation. IL-1 beta, IL-6, IL-10 and GM-CSF mRNA expression was also upregulated, peaking at 24 h after stimulation. Taken together, these results suggest that adult T. canis-derived antigens have the ability to activate human PBMC as conventional antigens, possibly due to their cross-reactivity, which may be involved in the host defence against helminth infection.

T-kininogen induces endothelial cell proliferation.

PubMed

Pérez, Viviana; Leiva-Salcedo, Elías; Acuña-Castillo, Claudio; Aravena, Mauricio; Gómez, Christian; Sabaj, Valeria; Colombo, Alicia; Nishimura, Sumiyo; Pérez, Claudio; Walter, Robin; Sierra, Felipe

2006-03-01

Basal proliferation of endothelial cells increases with age, and this might play a role in the etiology of age-related vascular diseases, as well as angiogenesis. Serum kininogen levels increase during aging in rats and humans, and T-kininogen (T-KG) can affect proliferative homeostasis in several cell models. Both kinins and kininogens have been shown previously to be angiogenic through activation of endothelial cell proliferation, and here we show that exposure of endothelial cells to T-KG results in vigorous cell proliferation, accompanied by ERK/AKT activation. In our experiments, the proliferative response requires B1 and B2 kinin receptors, even though kinins are not released from the precursor. We hypothesize that the age-related increase in T-KG could play a significant role in the age-related dysregulation of vascular physiology and function.

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

Shi, Yang, E-mail: yangshi_xz@126.com; Song, Qingwei; Hu, Dianhe

Hepatocellular carcinoma (HCC) is one of the most common cancers and can be induced by chronic HBV infection. The role of HBV-specific immune responses in mediating tumorigenesis and HCC prognosis is debated. The effect of intratumoral microenvironment on tumor-infiltrating lymphocytes (TILs) is also unclear. Here, we examined resected tumor tissue from 36 patients with HBV-induced HCC. We categorized study cohort based on ex vivo IL-10 secretion by tumor cells into high IL-10-secreting (Hi10) and low IL-10-secreting (Lo10) groups, and found that the Lo10 group was less sensitive to TLR ligand stimulation. TILs from the Lo10 group contained higher frequencies of HBV-specificmore » IFN-g-producing cells and total IFN-g-producing cells, and possessed higher proliferative capacity. Moreover, the proliferative capacity of TILs from the Hi10 group was negatively correlated with IL-10 secretion from tumor cells. Together, our data demonstrated that low IL-10-producing capacity in HBV-induced HCC tumors is associated with enhanced TIL activity. - Highlights: • We examined intratumoral IL-10 production in HBV-induced HCC. • We grouped HCC tumors into Hi10 and Lo10 groups based on their IL-10 production. • Lo10 groups had better IFN-g response by TILs. • Lo10 groups had better TIL proliferative capacity. • Lo10 group tumor cells were refractory to TLR ligand stimulation.« less

Rachycentron canadum (cobia) lectin promoted mitogenic response in mice BALB/c splenocytes.

PubMed

Coriolano, M C; de Melo, C M L; Santos, A J G; Pereira, V R A; Coelho, L C B B

2012-12-01

The mitogenic lectins are invaluable tools to study the biochemical changes associated with lymphocyte activation and proliferation of various immune cells. Rachycentron canadum lectin (RcaL) was detected and purified from serum of cobia fish. The aim of this study was to evaluate the proliferative response and cytokine production in splenocytes of mice in vitro stimulated with RcaL lectin; Canavalia ensiformis lectin (Con A) was used as positive control. A high proliferation index was induced by RcaL in relation to control cells. Furthermore, RcaL induced higher IL-2 and IL-6 production in relation to control. The cell viability was 90% in splenocytes treated with RcaL lectin, but RcaL promoted significant late apoptosis after 24 and 48 h in relation to control. RcaL induced proliferative responses suggesting that this lectin can be used as a mitogenic agent in immunostimulatory assays. © 2012 The Authors. Scandinavian Journal of Immunology © 2012 Blackwell Publishing Ltd.

Identification of helper T cell epitopes of dengue virus E-protein.

PubMed

Leclerc, C; Dériaud, E; Megret, F; Briand, J P; Van Regenmortel, M H; Deubel, V

1993-05-01

The T cell proliferative response to dengue 2 (Jamaica) E-glycoprotein (495 amino acids) was analyzed in vitro using either killed virus or E-protein fragments or synthetic peptides. Inactivated dengue virus stimulated dengue-specific lymph node (LN) CD4+T cell proliferation in BALB/c (H-2d), C3H (H-2k) and DBA/1 (H-2q) but not in C57BL/6 (H-2b) mice. Moreover, LN cells from dengue-virus primed BALB/c mice proliferated in vitro in response to three purified non-overlapping E-protein fragments expressed in E. coli as polypeptides fused to trpE (f22-205, f267-354, f366-424). To further determine T cell epitopes in the E-protein, synthetic peptides were selected using prediction algorithms for T cell epitopes. Highest proliferative responses were obtained after in vitro exposure of virus-primed LN cells to peptides p135-157, p270-298, p295-307 and p337-359. Peptide p59-78 was able to induce specific B and T cell responses in peptide-primed mice of H-2d, H-2q and H-2k haplotypes. Two peptides p59-78 corresponding to two dengue (Jamaica and Sri Lanka) isolates and differing only at position 71 cross-reacted at the B but not at the T cell level in H-2b mice. This analysis of murine T helper cell response to dengue E-protein may be of use in dengue subunit vaccine design.

Content of endoplasmic reticulum and Golgi complex membranes positively correlates with the proliferative status of brain cells.

PubMed

Silvestre, David C; Maccioni, Hugo J F; Caputto, Beatriz L

2009-03-01

Although the molecular and cellular basis of particular events that lead to the biogenesis of membranes in eukaryotic cells has been described in detail, understanding of the intrinsic complexity of the pleiotropic response by which a cell adjusts the overall activity of its endomembrane system to accomplish these requirements is limited. Here we carried out an immunocytochemical and biochemical examination of the content and quality of the endoplasmic reticulum (ER) and Golgi apparatus membranes in two in vivo situations characterized by a phase of active cell proliferation followed by a phase of declination in proliferation (rat brain tissue at early and late developmental stages) or by permanent active proliferation (gliomas and their most malignant manifestation, glioblastomas multiforme). It was found that, in highly proliferative phases of brain development (early embryo brain cells), the content of ER and Golgi apparatus membranes, measured as total lipid phosphorous content, is higher than in adult brain cells. In addition, the concentration of protein markers of ER and Golgi is also higher in early embryo brain cells and in human glioblastoma multiforme cells than in adult rat brain or in nonpathological human brain cells. Results suggest that the amount of endomembranes and the concentration of constituent functional proteins diminish as cells decline in their proliferative activity.

[Activity of non-specific T-suppressors regulating the proliferation of B- and T-lymphocytes in patients with fibroadenomatosis, breast cancer and stomach cancer].

PubMed

Grinevich, Iu A; Drannik, G N; Nikol'skiĭ, I S; Kalinina, N A; Litvishchenko, E I

1984-01-01

A decrease in proliferative rate of blood-circulating lymphocytes in response to LPS and PHA was registered in patients with fibroadenomatosis and cancer of the breast and stomach cancer. The said cells preincubated with Concanavalin A showed a weak inhibitory action on B-cells. The inhibition of T-cell proliferation by lymphocytes either remained unchanged or became less apparent in stage II breast cancer and slightly increased in stage III gastric cancer. Since no correlation was established between proliferative levels of T- and B-lymphocytes, two separate subpopulations of non-specific lymphocytes (T-T and T-B) were suggested.

A polymer-based ratiometric intracellular glucose sensor.

PubMed

Zhang, Liqiang; Su, Fengyu; Buizer, Sean; Kong, Xiangxing; Lee, Fred; Day, Kevin; Tian, Yanqing; Meldrum, Deirdre R

2014-07-04

The glucose metabolism level reflects cell proliferative status. A polymeric glucose ratiometric sensor comprising poly(N-(2-hydroxypropyl)methacrylamide) (PHPMA) and poly[2-(methacryloyloxy)ethyl]trimethylammonium chloride (PMAETMA) was synthesized. Cellular internalization and glucose response of the polymer within HeLa cells were investigated.

Effect of monoclonal antibodies (MoAb) to class I and class II HLA antigens on lectin- and MoAb OKT3-induced lymphocyte proliferation.

PubMed

Akiyama, Y; Zicht, R; Ferrone, S; Bonnard, G D; Herberman, R B

1985-04-01

We have examined the effect of several monoclonal antibodies (MoAb) to monomorphic determinants of class II HLA antigens, and MoAb to monomorphic determinants of class I HLA antigens and to beta-2-microglobulin (beta 2-mu) on lectin- and MoAb OKT3-induced proliferation of human peripheral blood mononuclear cells (PBMNC) and cultured T cells (CTC). Some, but not all, anti-class II HLA MoAb inhibited the proliferative response of PBMNC to MoAb OKT3 and pokeweed mitogen (PWM). The degree of inhibitory effect varied considerably. This effect was not limited to anti-class II HLA MoAb since anti-class I HLA MoAb and anti-beta 2-mu MoAb also inhibited MoAb OKT3- or PWM-induced proliferative responses. In contrast, the response of PBMNC to phytohemagglutinin (PHA) and concanavalin A (Con A) was not blocked by any anti-class II HLA MoAb. However, some anti-class II HLA MoAb also inhibited the proliferative response of CTC plus allogeneic peripheral blood adherent accessory cells (AC) to PHA or Con A as well as to MoAb OKT3 or PWM. This may be attributable to the substantially greater class II HLA antigen expression by CTC than by fresh lymphocytes. Pretreatment of either CTC or AC with anti-class II HLA MoAb inhibited OKT3-induced proliferation. In contrast, pretreatment of CTC, but not AC, with anti-class I HLA MoAb inhibited the proliferative response of CTC to OKT3. Pretreatment of CTC with anti-class I HLA MoAb inhibited PHA-, Con A and PWM-induced proliferation, to a greater degree than the anti-class II HLA MoAb. It appears as if lymphocyte activation by different mitogens exhibits variable requirements for the presence of cells expressing major histocompatibility determinants. Binding of Ab to membrane markers may interfere with lymphocyte-AC cooperation, perhaps by inhibiting binding of mitogens to their receptors or by interfering with lymphocyte and AC function. We also have examined the role of class II HLA antigens on CTC by depleting class II HLA-positive cells. As expected, elimination of class II HLA-positive AC with anti-class II HLA MoAb plus complement caused a decrease in proliferation of CTC in response to all the mitogens tested. In contrast, elimination of class II HLA-positive CTC was shown to clearly increase proliferation of CTC, perhaps because this may deplete class II HLA-positive suppressor cells.

Resveratrol Alters Proliferative Responses and Apoptosis in Human Activated B Lymphocytes In Vitro

USDA-ARS?s Scientific Manuscript database

We hypothesized that resveratrol, a polyphenol found in grapes, peanuts, and berries would modulate B lymphocyte proliferation, immunoglobulin synthesis, and apoptosis after activation with T-cell dependent pokeweed mitogen. Peripheral blood mononuclear cells (PBMCs) were isolated from the blood of ...

Modeling Cancer Cell Growth Dynamics In vitro in Response to Antimitotic Drug Treatment

PubMed Central

Lorz, Alexander; Botesteanu, Dana-Adriana; Levy, Doron

2017-01-01

Investigating the role of intrinsic cell heterogeneity emerging from variations in cell-cycle parameters and apoptosis is a crucial step toward better informing drug administration. Antimitotic agents, widely used in chemotherapy, target exclusively proliferative cells and commonly induce a prolonged mitotic arrest followed by cell death via apoptosis. In this paper, we developed a physiologically motivated mathematical framework for describing cancer cell growth dynamics that incorporates the intrinsic heterogeneity in the time individual cells spend in the cell-cycle and apoptosis process. More precisely, our model comprises two age-structured partial differential equations for the proliferative and apoptotic cell compartments and one ordinary differential equation for the quiescent compartment. To reflect the intrinsic cell heterogeneity that governs the growth dynamics, proliferative and apoptotic cells are structured in “age,” i.e., the amount of time remaining to be spent in each respective compartment. In our model, we considered an antimitotic drug whose effect on the cellular dynamics is to induce mitotic arrest, extending the average cell-cycle length. The prolonged mitotic arrest induced by the drug can trigger apoptosis if the time a cell will spend in the cell cycle is greater than the mitotic arrest threshold. We studied the drug’s effect on the long-term cancer cell growth dynamics using different durations of prolonged mitotic arrest induced by the drug. Our numerical simulations suggest that at confluence and in the absence of the drug, quiescence is the long-term asymptotic behavior emerging from the cancer cell growth dynamics. This pattern is maintained in the presence of small increases in the average cell-cycle length. However, intermediate increases in cell-cycle length markedly decrease the total number of cells and can drive the cancer population to extinction. Intriguingly, a large “switch-on/switch-off” increase in the average cell-cycle length maintains an active cell population in the long term, with oscillating numbers of proliferative cells and a relatively constant quiescent cell number. PMID:28913178

Impaired Upregulation of the Costimulatory Molecules, CD27 and CD28, on CD4+ T Cells from HIV Patients Receiving ART Is Associated with Poor Proliferative Responses.

PubMed

Tanaskovic, Sara; Price, Patricia; French, Martyn A; Fernandez, Sonia

2017-02-01

HIV patients beginning antiretroviral therapy (ART) with advanced immunodeficiency often retain low CD4 + T cell counts despite virological control. We examined proliferative responses and upregulation of costimulatory molecules, following anti-CD3 stimulation, in HIV patients with persistent CD4 + T cell deficiency on ART. Aviremic HIV patients with nadir CD4 + T cell counts <100 cells/μL and who had received ART for a median time of 7 (range 1-11) years were categorized into those achieving low (<350 cells/μL; n = 13) or normal (>500 cells/μL; n = 20) CD4 + T cell counts. Ten healthy controls were also recruited. CD4 + T cell proliferation (Ki67) and upregulation of costimulatory molecules (CD27 and CD28) after anti-CD3 stimulation were assessed by flow cytometry. Results were related to proportions of CD4 + T cells expressing markers of T cell senescence (CD57), activation (HLA-DR), and apoptotic potential (Fas). Expression of CD27 and/or CD28 on uncultured CD4 + T cells was similar in patients with normal CD4 + T cell counts and healthy controls, but lower in patients with low CD4 + T cell counts. Proportions of CD4 + T cells expressing CD27 and/or CD28 correlated inversely with CD4 + T cell expression of CD57, HLA-DR, and Fas. After anti-CD3 stimulation, induction of CD27 hi CD28 hi expression was independent of CD4 + T cell counts, but lower in HIV patients than in healthy controls. Induction of CD27 hi CD28 hi expression correlated with induction of Ki67 expression in total, naïve, and CD31 + naïve CD4 + T cells from patients. In HIV patients responding to ART, impaired induction of CD27 and CD28 on CD4 + T cells after stimulation with anti-CD3 is associated with poor proliferative responses as well as greater CD4 + T cell activation and immunosenescence.

Two separate defects affecting true naive or virtual memory T cell precursors combine to reduce naive T cell responses with aging.

PubMed

Renkema, Kristin R; Li, Gang; Wu, Angela; Smithey, Megan J; Nikolich-Žugich, Janko

2014-01-01

Naive T cell responses are eroded with aging. We and others have recently shown that unimmunized old mice lose ≥ 70% of Ag-specific CD8 T cell precursors and that many of the remaining precursors acquire a virtual (central) memory (VM; CD44(hi)CD62L(hi)) phenotype. In this study, we demonstrate that unimmunized TCR transgenic (TCRTg) mice also undergo massive VM conversion with age, exhibiting rapid effector function upon both TCR and cytokine triggering. Age-related VM conversion in TCRTg mice directly depended on replacement of the original TCRTg specificity by endogenous TCRα rearrangements, indicating that TCR signals must be critical in VM conversion. Importantly, we found that VM conversion had adverse functional effects in both old wild-type and old TCRTg mice; that is, old VM, but not old true naive, T cells exhibited blunted TCR-mediated, but not IL-15-mediated, proliferation. This selective proliferative senescence correlated with increased apoptosis in old VM cells in response to peptide, but decreased apoptosis in response to homeostatic cytokines IL-7 and IL-15. Our results identify TCR as the key factor in differential maintenance and function of Ag-specific precursors in unimmunized mice with aging, and they demonstrate that two separate age-related defects--drastic reduction in true naive T cell precursors and impaired proliferative capacity of their VM cousins--combine to reduce naive T cell responses with aging.

Centchroman inhibits proliferation of head and neck cancer cells through the modulation of PI3K/mTOR Pathway

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

Srivastava, Vikas Kumar; Gara, Rishi Kumar; Bhatt, M.L.B.

Research highlights: {yields} Centchroman (CC) inhibits cellular proliferation in HNSCC cells through the dual inhibition of PI3/mTOR pathway. {yields} CC treatment also inhibits STAT3 activation and alters expression of proteins involved in cell cycle regulation and DNA repair response in HNSCC cells. {yields} CC exhibits anti-proliferative activity in a variety of non-HNSCC cancer cell lines and is devoid of cytotoxicity to normal cell types of diverse origins. -- Abstract: Centchroman (CC; 67/20; INN: Ormeloxifene) is a non-steroidal antiestrogen extensively used as a female contraceptive in India. In the present study, we report the anti-proliferative effect of CC in head andmore » neck squamous cell carcinoma (HNSCC) cells. CC inhibited cell proliferation in a dose dependent manner at 24 h of treatment. Further studies showed that CC treatment induced apoptosis, inhibited Akt/mTOR and signal transducers and activators of transcription protein 3 (STAT3) signaling, altered proteins associated with cell cycle regulation and DNA damage and inhibited colony forming efficiency of HNSCC cells. In addition, CC displayed anti-proliferative activity against a variety of non-HNSCC cell lines of diverse origin. The ability of CC to serve as a dual-inhibitor of Akt/mTOR and STAT3 signaling warrants further studies into its role as a therapeutic strategy against HNSCC.« less

Molecular Effects of I3C/DIM in Prostate Cancer Cells

DTIC Science & Technology

2005-04-01

DNA fragmentation, resulting in apoptosis (40,41). To (phorbol myristate acetate, etc.), stress (pH, hypoxia, heavy further explore the mechanism of...cytosol into the ling cell proliferation, differentiation, apoptosis, inflammation, mitochondria in both cell lines treated with 13C. However, stress ...of cell prolifer- the control of cell growth, apoptosis, inflammation, ation, cell cycle, apoptosis, signal transduction, onco- stress response, and

Age-Dependent Netrin-1 Signaling Regulates NG2+ Glial Cell Spatial Homeostasis in Normal Adult Gray Matter.

PubMed

Birey, Fikri; Aguirre, Adan

2015-04-29

Neuron-glial antigen 2-positive (NG2(+)) glial cells are the most proliferative glia type in the adult CNS, and their tile-like arrangement in adult gray matter is under tight regulation. However, little is known about the cues that govern this unique distribution. To this end, using a NG2(+) glial cell ablation model in mice, we examined the repopulation dynamics of NG2(+) glial cells in the mature and aged mice gray matter. We found that some resident NG2(+) glial cells that escaped depletion rapidly enter the cell cycle to repopulate the cortex with altered spatial distribution. We reveal that netrin-1 signaling is involved in the NG2(+) glial cell early proliferative, late repopulation, and distribution response after ablation in the gray matter. However, ablation of NG2(+) glial cell in older animals failed to stimulate a similar repopulation response, possibly because of a decrease in the sensitivity to netrin-1. Our findings indicate that endogenous netrin-1 plays a role in NG2(+) glial cell homeostasis that is distinct from its role in myelination. Copyright © 2015 the authors 0270-6474/15/356946-06$15.00/0.

Human Atopic Dermatitis Skin-derived T Cells can Induce a Reaction in Mouse Keratinocytes in vivo.

PubMed

Martel, B C; Blom, L; Dyring-Andersen, B; Skov, L; Thestrup-Pedersen, K; Skov, S; Skak, K; Poulsen, L K

2015-08-01

In atopic dermatitis (AD), the inflammatory response between skin-infiltrating T cells and keratinocytes is fundamental to the development of chronic lesional eczema. The aim of this study was to investigate whether skin-derived T cells from AD patients could induce an inflammatory response in mice through keratinocyte activation and consequently cause the development of eczematous lesions. Punch biopsies of the lesional skin from AD patients were used to establish skin-derived T cell cultures, which were transferred to NOD.Cg-Prkd(scid) Il2rg(tm1Sug) /JicTac (NOG) mice. We found that the subcutaneous injection of the human AD skin-derived T cells resulted in the migration of the human T cells from subcutis to the papillary dermis followed by the development of erythema and oedema in the mouse skin. Furthermore, the human T cells induced a transient proliferative response in the mouse keratinocytes shown as increased numbers of Ki-67(+) keratinocytes and increased epidermal thickness. Out of six established AD skin-derived T cell cultures, two were superior at inducing a skin reaction in the mice, and these cultures were found to contain >10% CCR10(+) T cells compared to <2% for the other cultures. In comparison, blood-derived in vitro-differentiated Th2 cells only induced a weak response in a few of the mice. Thus, we conclude that human AD skin-derived T cells can induce a reaction in the mouse skin through the induction of a proliferative response in the mouse keratinocytes. © 2015 The Foundation for the Scandinavian Journal of Immunology.

Pathways linking major depression and immunity in ambulatory female patients.

PubMed

Miller, G E; Cohen, S; Herbert, T B

1999-01-01

The goals of this study were to investigate whether depression is associated with cellular immunity in ambulatory patients and to identify neuroendocrine and behavioral pathways that might account for this relationship. We studied 32 women who met Diagnostic and Statistical Manual of Mental Disorder, fourth edition, criteria for major depressive disorder and 32 healthy female control subjects. The groups were matched for age and ethnicity. None were taking medication, and all were free of disease involving the immune system. Depressed subjects had reduced proliferative responses to the mitogens concanavalin A and phytohemagglutinin compared with control subjects. Natural killer cell activity was reduced among older depressed subjects but enhanced among younger depressed subjects. Although depression was associated with elevated circulating levels of norepinephrine and estradiol, these hormones could not account for the immunologic differences between depressed and control subjects. Depression was also associated with greater tobacco and caffeine consumption, less physical activity, and poorer sleep quality. Mediational analyses were consistent with physical activity acting as a pathway through which depression was associated with reduced lymphocyte proliferation. Ambulatory patients with mild to moderately severe depression exhibit reduced mitogen-stimulated lymphocyte proliferative responses and altered natural killer cell cytotoxicity. The relationship between depression and proliferative responses may be mediated by physical activity.

Relationship of Blood Mercury Levels to Health Parameters in the Loggerhead Sea Turtle (Caretta caretta)

PubMed Central

Day, Rusty D.; Segars, Al L.; Arendt, Michael D.; Lee, A. Michelle; Peden-Adams, Margie M.

2007-01-01

Background Mercury is a pervasive environmental pollutant whose toxic effects have not been studied in sea turtles in spite of their threatened status and evidence of immunosuppression in diseased populations. Objectives In the present study we investigate mercury toxicity in loggerhead sea turtles (Caretta caretta) by examining trends between blood mercury concentrations and various health parameters. Methods Blood was collected from free-ranging turtles, and correlations between blood mercury concentrations and plasma chemistries, complete blood counts, lysozyme, and lymphocyte proliferation were examined. Lymphocytes were also harvested from free-ranging turtles and exposed in vitro to methylmercury to assess proliferative responses. Results Blood mercury concentrations were positively correlated with hematocrit and creatine phosphokinase activity, and negatively correlated with lymphocyte cell counts and aspartate amino-transferase. Ex vivo negative correlations between blood mercury concentrations and B-cell proliferation were observed in 2001 and 2003 under optimal assay conditions. In vitro exposure of peripheral blood leukocytes to methylmercury resulted in suppression of proliferative responses for B cells (0.1 μg/g and 0.35 μg/g) and T cells (0.7 μg/g). Conclusions The positive correlation between blood mercury concentration and hematocrit reflects the higher affinity of mercury species for erythrocytes than plasma, and demonstrates the importance of measuring hematocrit when analyzing whole blood for mercury. In vitro immunosuppression occurred at methylmercury concentrations that correspond to approximately 5% of the individuals captured in the wild. This observation and the negative correlation found ex vivo between mercury and lymphocyte numbers and mercury and B-cell proliferative responses suggests that subtle negative impacts of mercury on sea turtle immune function are possible at concentrations observed in the wild. PMID:17938730

Profound impairment of adaptive immune responses by alkylating chemotherapy

PubMed Central

Litterman, Adam J.; Zellmer, David M.; Grinnen, Karen L.; Hunt, Matthew A.; Dudek, Arkadiusz Z.; Salazar, Andres M.; Ohlfest, John R.

2013-01-01

Cancer vaccines have overall had a record of failure as an adjuvant therapy for malignancies that are treated with alkylating chemotherapy, and the contribution of standard treatment to that failure remains unclear. Vaccines aim to harness the proliferative potential of the immune system by expanding a small number of tumor-specific lymphocytes into a large number of anti-tumor effectors. Clinical trials are often conducted after treatment with alkylating chemotherapy, given either as standard therapy or for immunomodulatory effect. There is mounting evidence for synergy between chemotherapy and adoptive immunotherapy or vaccination against self-antigens; however, the impact of chemotherapy on lymphocytes primed against tumor neo-antigens remains poorly defined. We report here that clinically relevant dosages of standard alkylating chemotherapies such as temozolomide and cyclophosphamide significantly inhibit the proliferative abilities of lymphocytes in mice. This proliferative impairment was long lasting and led to quantitative and qualitative defects in B and T cell responses to neo-antigen vaccines. High affinity responder lymphocytes receiving the strongest proliferative signals from vaccines experienced the greatest DNA damage responses, skewing the response toward lower affinity responders with inferior functional characteristics. Together these defects lead to inferior efficacy and overall survival in murine tumor models treated by neo-antigen vaccines. These results suggest that clinical protocols for cancer vaccines should be designed to avoid exposing responder lymphocytes to alkylating chemotherapy. PMID:23686484

Proliferative lifespan is conserved after nuclear transfer.

PubMed

Clark, A John; Ferrier, Patricia; Aslam, Samena; Burl, Sarah; Denning, Chris; Wylie, Diana; Ross, Arlene; de Sousa, Paul; Wilmut, Ian; Cui, Wei

2003-06-01

Cultured primary cells exhibit a finite proliferative lifespan, termed the Hayflick limit. Cloning by nuclear transfer can reverse this cellular ageing process and can be accomplished with cultured cells nearing senescence. Here we describe nuclear transfer experiments in which donor cell lines at different ages and with different proliferative capacities were used to clone foetuses and animals from which new primary cell lines were generated. The rederived lines had the same proliferative capacity and rate of telomere shortening as the donor cell lines, suggesting that these are innate, genetically determined, properties that are conserved by nuclear transfer.

Regional control of Drosophila gut stem cell proliferation: EGF establishes GSSC proliferative set point & controls emergence from quiescence.

PubMed

Strand, Marie; Micchelli, Craig A

2013-01-01

Adult stem cells vary widely in their rates of proliferation. Some stem cells are constitutively active, while others divide only in response to injury. The mechanism controlling this differential proliferative set point is not well understood. The anterior-posterior (A/P) axis of the adult Drosophila midgut has a segmental organization, displaying physiological compartmentalization and region-specific epithelia. These distinct midgut regions are maintained by defined stem cell populations with unique division schedules, providing an excellent experimental model with which to investigate this question. Here, we focus on the quiescent gastric stem cells (GSSCs) of the acidic copper cell region (CCR), which exhibit the greatest period of latency between divisions of all characterized gut stem cells, to define the molecular basis of differential stem cell activity. Our molecular genetic analysis demonstrates that the mitogenic EGF signaling pathway is a limiting factor controlling GSSC proliferation. We find that under baseline conditions, when GSSCs are largely quiescent, the lowest levels of EGF ligands in the midgut are found in the CCR. However, acute epithelial injury by enteric pathogens leads to an increase in EGF ligand expression in the CCR and rapid expansion of the GSSC lineage. Thus, the unique proliferative set points for gut stem cells residing in physiologically distinct compartments are governed by regional control of niche signals along the A/P axis.

Response of animal and vegetative cells to the effect of a typical magnetic storm

NASA Astrophysics Data System (ADS)

Talikina, M. G.; Izyumov, Yu. G.; Krylov, V. V.

2013-12-01

Experimentally reproduced fluctuations of a low-frequency magnetic field in a nanotesla range (magnetic storm) affect the mitosis of animals and vegetative cells. Action of this factor during twenty four hours leads to a significant increase in the proliferative activity of embryo cells in roach ( Rutilus rutilus L.) and meristem cells of onion rootlets ( Allium cepa). The clastogenic effect statistically confirmed only in the Allium test seems to reflect the species specificity of the response and higher sensitivity of the cell association of the onion meristem to magnetic storm.

NG2 glial cells regulate neuroimmunological responses to maintain neuronal function and survival.

PubMed

Nakano, Masayuki; Tamura, Yasuhisa; Yamato, Masanori; Kume, Satoshi; Eguchi, Asami; Takata, Kumi; Watanabe, Yasuyoshi; Kataoka, Yosky

2017-02-14

NG2-expressing neural progenitor cells (i.e., NG2 glial cells) maintain their proliferative and migratory activities even in the adult mammalian central nervous system (CNS) and produce myelinating oligodendrocytes and astrocytes. Although NG2 glial cells have been observed in close proximity to neuronal cell bodies in order to receive synaptic inputs, substantive non-proliferative roles of NG2 glial cells in the adult CNS remain unclear. In the present study, we generated NG2-HSVtk transgenic rats and selectively ablated NG2 glial cells in the adult CNS. Ablation of NG2 glial cells produced defects in hippocampal neurons due to excessive neuroinflammation via activation of the interleukin-1 beta (IL-1β) pro-inflammatory pathway, resulting in hippocampal atrophy. Furthermore, we revealed that the loss of NG2 glial cell-derived hepatocyte growth factor (HGF) exacerbated these abnormalities. Our findings suggest that NG2 glial cells maintain neuronal function and survival via the control of neuroimmunological function.

Proliferative responses in the local lymph node assay associated with concomitant exposure to 1,4-phenylenediamine and methyldibromo glutaronitrile: evidence for synergy?

PubMed

Jowsey, Ian R; Basketter, David A; Irwin, Anita

2008-08-01

A key consideration when undertaking risk assessments should be the potential for synergy between contact allergens. Previously, this concept has only been investigated during elicitation in contact allergic individuals. To determine whether there exists evidence for synergy between contact allergens during the induction phase of skin sensitization using the mouse local lymph node assay (LLNA) as a model system. Proliferative responses in draining lymph nodes were assessed with increasing concentrations of 1,4-phenylenediamine (PPD), methyldibromo glutaronitrile (MDBGN), and a combination of PPD and MDBGN. Data from each of two independent experiments show that lymph node cell proliferation associated with combined exposure to PPD and MDBGN was, in general, only modestly increased relative to that predicted from a simple summation of their individual responses. Although the increase in response is very modest, it does imply a relationship between this combination of sensitizers that may not be simply additive in terms of their ability to stimulate proliferative responses in draining lymph nodes. The reproducibility of this observation should be confirmed in future studies with additional pairs of contact allergens to ascertain whether or not this represents evidence of synergy.

Achiral Mannich-Base Curcumin Analogs Induce Unfolded Protein Response and Mitochondrial Membrane Depolarization in PANC-1 Cells.

PubMed

Szebeni, Gábor J; Balázs, Árpád; Madarász, Ildikó; Pócz, Gábor; Ayaydin, Ferhan; Kanizsai, Iván; Fajka-Boja, Roberta; Alföldi, Róbert; Hackler, László; Puskás, László G

2017-10-07

Achiral Mannich-type curcumin analogs have been synthetized and assayed for their cytotoxic activity. The anti-proliferative and cytotoxic activity of curcuminoids has been tested on human non-small-cell lung carcinoma (A549), hepatocellular carcinoma (HepG2) and pancreatic cancer cell line (PANC-1). Based on the highest anti-proliferative activity nine drug candidates were further tested and proved to cause phosphatidylserine exposure as an early sign of apoptosis. Curcumin analogs with the highest apoptotic activity were selected for mechanistic studies in the most sensitive PANC-1 cells. Cytotoxic activity was accompanied by cytostatic effect since curcumin and analogs treatment led to G₀/G₁ cell cycle arrest. Moreover, cytotoxic effect could be also detected via the accumulation of curcuminoids in the endoplasmic reticulum (ER) and the up-regulation of ER stress-related unfolded protein response (UPR) genes: HSPA5 , ATF4, XBP1 , and DDIT3 . The activated UPR induced mitochondrial membrane depolarization, caspase-3 activation and subsequent DNA breakdown in PANC-1 cells. Achiral curcumin analogs, C509, C521 and C524 possessed superior, 40-times more potent cytotoxic activity compared to natural dihydroxy-dimetoxycurcumin in PANC-1 cells.

T cells which proliferate in response to concanavalin A include cells which proliferate in mixed leucocyte reactions.

PubMed

Watanabe, T; Fathman, C G; Coutinho, A

1977-09-01

Selection in long-term culture of alloreactive T cells, by successive in vitro restimulation with semi-allogeneic cells, results in primed responder cell populations which maintain full proliferative reactivity to allogeneic cells as well as to the T cell mitogens concanavalin A (Con A) and phytohemagglutinin (PHA) but are depleted of cells which can effect target cell destruction in either a specific or nonspecific manner. Con A-induced T cell blasts (selected by velocity sedimentation) can revert to small resting lymphocytes in the presence of inert "filler" cells. Con A blasts which have reverted, readily proliferate in response to Con A or allogeneic stimulator cells but are largely depleted of effector killer cells and PHA-responsive cells.

Activation of cellular death programs associated with immunosenescence-like phenotype in TPPII knockout mice

PubMed Central

Huai, Jisen; Firat, Elke; Nil, Ahmed; Million, Daniele; Gaedicke, Simone; Kanzler, Benoit; Freudenberg, Marina; van Endert, Peter; Kohler, Gabriele; Pahl, Heike L.; Aichele, Peter; Eichmann, Klaus; Niedermann, Gabriele

2008-01-01

The giant cytosolic protease tripeptidyl peptidase II (TPPII) has been implicated in the regulation of proliferation and survival of malignant cells, particularly lymphoma cells. To address its functions in normal cellular and systemic physiology we have generated TPPII-deficient mice. TPPII deficiency activates cell type-specific death programs, including proliferative apoptosis in several T lineage subsets and premature cellular senescence in fibroblasts and CD8+ T cells. This coincides with up-regulation of p53 and dysregulation of NF-κB. Prominent degenerative alterations at the organismic level were a decreased lifespan and symptoms characteristic of immunohematopoietic senescence. These symptoms include accelerated thymic involution, lymphopenia, impaired proliferative T cell responses, extramedullary hematopoiesis, and inflammation. Thus, TPPII is important for maintaining normal cellular and systemic physiology, which may be relevant for potential therapeutic applications of TPPII inhibitors. PMID:18362329

Mild myelin disruption elicits early alteration in behavior and proliferation in the subventricular zone.

PubMed

Gould, Elizabeth A; Busquet, Nicolas; Shepherd, Douglas; Dietz, Robert M; Herson, Paco S; Simoes de Souza, Fabio M; Li, Anan; George, Nicholas M; Restrepo, Diego; Macklin, Wendy B

2018-02-13

Myelin, the insulating sheath around axons, supports axon function. An important question is the impact of mild myelin disruption. In the absence of the myelin protein proteolipid protein (PLP1), myelin is generated but with age, axonal function/maintenance is disrupted. Axon disruption occurs in Plp1 -null mice as early as 2 months in cortical projection neurons. High-volume cellular quantification techniques revealed a region-specific increase in oligodendrocyte density in the olfactory bulb and rostral corpus callosum that increased during adulthood. A distinct proliferative response of progenitor cells was observed in the subventricular zone (SVZ), while the number and proliferation of parenchymal oligodendrocyte progenitor cells was unchanged. This SVZ proliferative response occurred prior to evidence of axonal disruption. Thus, a novel SVZ response contributes to the region-specific increase in oligodendrocytes in Plp1 -null mice. Young adult Plp1- null mice exhibited subtle but substantial behavioral alterations, indicative of an early impact of mild myelin disruption. © 2018, Gould et al.

Mild myelin disruption elicits early alteration in behavior and proliferation in the subventricular zone

PubMed Central

Gould, Elizabeth A; Busquet, Nicolas; Shepherd, Douglas; Dietz, Robert M; Herson, Paco S; Simoes de Souza, Fabio M; Li, Anan; George, Nicholas M

2018-01-01

Myelin, the insulating sheath around axons, supports axon function. An important question is the impact of mild myelin disruption. In the absence of the myelin protein proteolipid protein (PLP1), myelin is generated but with age, axonal function/maintenance is disrupted. Axon disruption occurs in Plp1-null mice as early as 2 months in cortical projection neurons. High-volume cellular quantification techniques revealed a region-specific increase in oligodendrocyte density in the olfactory bulb and rostral corpus callosum that increased during adulthood. A distinct proliferative response of progenitor cells was observed in the subventricular zone (SVZ), while the number and proliferation of parenchymal oligodendrocyte progenitor cells was unchanged. This SVZ proliferative response occurred prior to evidence of axonal disruption. Thus, a novel SVZ response contributes to the region-specific increase in oligodendrocytes in Plp1-null mice. Young adult Plp1-null mice exhibited subtle but substantial behavioral alterations, indicative of an early impact of mild myelin disruption. PMID:29436368

Hair cell regeneration

PubMed Central

Edge, Albert SB; Chen, Zheng-Yi

2017-01-01

The mammalian inner ear largely lacks the capacity to regenerate hair cells, the sensory cells required for hearing and balance. Recent studies in both lower vertebrates and mammals have uncovered genes and pathways important in hair cell development and have suggested ways that the sensory epithelia could be manipulated to achieve hair cell regeneration. These approaches include the use of inner ear stem cells, transdifferentiation of nonsensory cells, and induction of a proliferative response in the cells that can become hair cells. PMID:18929656

Gammadelta T lymphocytes from cystic fibrosis patients and healthy donors are high TNF-alpha and IFN-gamma-producers in response to Pseudomonas aeruginosa.

PubMed

Raga, Salvador; Julià, M Rosa; Crespí, Catalina; Figuerola, Joan; Martínez, Natalia; Milà, Joan; Matamoros, Núria

2003-01-01

Gammadelta T cells have an important immunoregulatory and effector function through cytokine release. They are involved in the responses to Gram-negative bacterium and in protection of lung epithelium integrity. On the other hand, they have been implicated in airway inflammation. The aim of the present work was to study intracytoplasmic IL-2, IL-4, IFN-gamma and TNF-alpha production by gammadelta and alphabeta T lymphocytes from cystic fibrosis patients and healthy donors in response to Pseudomonas aeruginosa (PA). Flow cytometric detection was performed after peripheral blood mononuclear cells (PBMC) culture with a cytosolic extract from PA and restimulation with phorbol ester plus ionomycine. Proliferative responses, activation markers and receptor usage of gammadelta T cells were also evaluated. The highest production of cytokine was of TNF-alpha and IFN-gamma, gammadelta being better producers than alphabeta. No differences were found between patients and controls. The Vgamma9delta2 subset of gammadelta T cells was preferentially expanded. CD25 and CD45RO expression by the alphabeta T subset and PBMC proliferative response to PA were defective in cystic fibrosis lymphocytes. Our results support the hypothesis that gammadelta T lymphocytes play an important role in the immune response to PA and in the chronic inflammatory lung reaction in cystic fibrosis patients. They do not confirm the involvement of a supressed Th1 cytokine response in the pathogenesis of this disease.

Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4+ T Cell Proliferation.

PubMed

Celada, Lindsay J; Rotsinger, Joseph E; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene; Drake, Wonder P

2017-01-01

Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4 + T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4 + T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1 + CD4 + T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P < 0.001). There was a negative correlation with PD-1 expression and proliferative capacity (r = -0.70, P < 0.001). Expression of key mediators of cell cycle progression, including PI3K and AKT, were significantly decreased. Gene and protein expression levels reverted to healthy control levels after PD-1 pathway blockade. Reduction in sarcoidosis CD4 + T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression.

Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4+ T Cell Proliferation

PubMed Central

Celada, Lindsay J.; Rotsinger, Joseph E.; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene

2017-01-01

Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4+ T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4+ T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1+ CD4+ T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P < 0.001). There was a negative correlation with PD-1 expression and proliferative capacity (r = −0.70, P < 0.001). Expression of key mediators of cell cycle progression, including PI3K and AKT, were significantly decreased. Gene and protein expression levels reverted to healthy control levels after PD-1 pathway blockade. Reduction in sarcoidosis CD4+ T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression. PMID:27564547

Increased Cellular Proliferation And Inflammatory Cytokines In Tonsils Derived From Children With Obstructive Sleep Apnea

PubMed Central

Kim, Jinkwan; Bhattacharjee, Rakesh; Dayyat, Ehab; Snow, Ayelet B.; Kheirandish-Gozal, Leila; Goldman, Julie L.; Li, Richard C.; Serpero, Laura D.; Clair, Heather B.; Gozal, David

2009-01-01

Adenotonsillar hypertrophy is the major pathophysiological mechanism underlying obstructive sleep apnea (OSA) and recurrent tonsillitis (RI) in children. The increased expression of various mediators of the inflammatory response in tonsils of OSA patients prompted our hypothesis that the enhanced local and systemic inflammation in OSA children would promote tonsillar proliferation. Mixed cell cultures from tonsils recovered during adenotonsillectomy in children with OSA and RI were established, and proliferative rates were assessed. Cells were also cultured to determine levels of pro-inflammatory cytokines and anti-oxidant protein levels and mRNA expression. Global cell proliferative rates from OSA tonsils were significantly higher than RI (P<0.01), with CD3+, CD4+, and CD8+ cell proliferation being higher in OSA (P<0.05). Moreover, pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1α were highly expressed in OSA-derived tonsils. Furthermore, thioredoxin (TRX), an anti-oxidant protein, was also highly expressed in OSA tonsils at the mRNA and protein levels (p<0.01). Thus, T-cells are in a highly proliferative state in the tonsils of children with OSA, and are associated with increased production of proinflammatory cytokines and TRX, when compared to children with RI. PMID:19581829

Proliferative reactive gliosis is compatible with glial metabolic support and neuronal function

PubMed Central

2011-01-01

Background The response of mammalian glial cells to chronic degeneration and trauma is hypothesized to be incompatible with support of neuronal function in the central nervous system (CNS) and retina. To test this hypothesis, we developed an inducible model of proliferative reactive gliosis in the absence of degenerative stimuli by genetically inactivating the cyclin-dependent kinase inhibitor p27Kip1 (p27 or Cdkn1b) in the adult mouse and determined the outcome on retinal structure and function. Results p27-deficient Müller glia reentered the cell cycle, underwent aberrant migration, and enhanced their expression of intermediate filament proteins, all of which are characteristics of Müller glia in a reactive state. Surprisingly, neuroglial interactions, retinal electrophysiology, and visual acuity were normal. Conclusion The benign outcome of proliferative reactive Müller gliosis suggests that reactive glia display context-dependent, graded and dynamic phenotypes and that reactivity in itself is not necessarily detrimental to neuronal function. PMID:21985191

SIRT1 deacetylase is overexpressed in human melanoma and its small molecule inhibition imparts anti-proliferative response via p53 activation.

PubMed

Wilking, Melissa J; Singh, Chandra; Nihal, Minakshi; Zhong, Weixiong; Ahmad, Nihal

2014-12-01

Melanoma causes more deaths than any other skin cancer, and its incidence in the US continues to rise. Current medical therapies are insufficient to control this deadly neoplasm, necessitating the development of new target-based approaches. The objective of this study was to determine the role and functional significance of the class III histone deacetylase SIRT1 in melanoma. We have found that SIRT1 is overexpressed in clinical human melanoma tissues and human melanoma cell lines (Sk-Mel-2, WM35, G361, A375, and Hs294T) compared to normal skin and normal melanocytes, respectively. In addition, treatment of melanoma cell lines A375, Hs294T, and G361 with Tenovin-1, a small molecule SIRT1 inhibitor, resulted in a significant decrease in cell growth and cell viability. Further, Tenovin-1 treatment also resulted in a marked decrease in the clonogenic survival of melanoma cells. Further experiments showed that the anti-proliferative response of Tenovin-1 was accompanied by an increase in the protein as well as activity of the tumor suppressor p53. This increase in p53 activity was substantiated by an increase in the protein level of its downstream target p21. Overall, these data suggest that small molecule inhibition of SIRT1 causes anti-proliferative effects in melanoma cells. SIRT1 appears to be acting through the activity of the tumor suppressor p53, which is not mutated in the majority of melanomas. However, future detailed studies are needed to further explore the role and mechanism of SIRT1 in melanoma development and progression and its usefulness in melanoma treatment.

TGF-{beta}-stimulated aberrant expression of class III {beta}-tubulin via the ERK signaling pathway in cultured retinal pigment epithelial cells

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

Chung, Eun Jee; Chun, Ji Na; Jung, Sun-Ah

2011-11-18

Highlights: Black-Right-Pointing-Pointer TGF-{beta} induces aberrant expression of {beta}III in RPE cells via the ERK pathway. Black-Right-Pointing-Pointer TGF-{beta} increases O-GlcNAc modification of {beta}III in RPE cells. Black-Right-Pointing-Pointer Mature RPE cells have the capacity to express a neuron-associated gene by TGF-{beta}. -- Abstract: The class III {beta}-tubulin isotype ({beta}{sub III}) is expressed exclusively by neurons within the normal human retina and is not present in normal retinal pigment epithelial (RPE) cells in situ or in the early phase of primary cultures. However, aberrant expression of class III {beta}-tubulin has been observed in passaged RPE cells and RPE cells with dedifferentiated morphology inmore » pathologic epiretinal membranes from idiopathic macular pucker, proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR). Transforming growth factor-{beta} (TGF-{beta}) has been implicated in dedifferentiation of RPE cells and has a critical role in the development of proliferative vitreoretinal diseases. Here, we investigated the potential effects of TGF-{beta} on the aberrant expression of class III {beta}-tubulin and the intracellular signaling pathway mediating these changes. TGF-{beta}-induced aberrant expression and O-linked-{beta}-N-acetylglucosamine (O-GlcNac) modification of class III {beta}-tubulin in cultured RPE cells as determined using Western blotting, RT-PCR and immunocytochemistry. TGF-{beta} also stimulated phosphorylation of ERK. TGF-{beta}-induced aberrant expression of class III {beta}-tubulin was significantly reduced by pretreatment with U0126, an inhibitor of ERK phosphorylation. Our findings indicate that TGF-{beta} stimulated aberrant expression of class III {beta}-tubulin via activation of the ERK signaling pathway. These data demonstrate that mature RPE cells have the capacity to express a neuron-associated gene in response to TGF-{beta} stimulation and provide useful information towards understanding the pathogenesis of proliferative vitreoretinal diseases.« less

Inhibition of anti-CD3 monoclonal antibody-induced T-cell proliferation and interleukin-2 secretion by physiologic combinations of dexamethasone and prostaglandin E2.

PubMed

Elliott, L; Brooks, W; Roszman, T

1993-12-01

1. The purpose of these studies was to characterize further previous observations from our laboratory indicating that physiologic concentrations of dexamethasone (DEX) and prostaglandin E2 (PGE2) added together result in synergistic inhibition of the proliferative response of T cells stimulated via the T-cell receptor CD3 signaling complex (TCR/CD3). 2. Various physiologic concentrations of DEX and PGE2 were added to T cells stimulated with immobilized anti-CD3 monoclonal antibody (mAb) and cultured at optimal and suboptimal cell densities. The results demonstrate that the proliferative response of anti-CD3 mAb-stimulated T cells cultured at a suboptimal cell density is more suppressed than that of T cells cultured at optimal concentrations. 3. The proliferative response of CD4+ T cells to immobilized anti-CD3 mAb was also determined in the presence of PGE2 and DEX. The data indicate that the CD4+ subset of T cells is more sensitive to the synergistic antiproliferative effects of DEX and PGE2 compared to whole T-cell populations. 4. Various concentrations of DEX and/or PGE2 were added to T cells stimulated with anti-CD3 mAb and the secretion of interleukin-2 (IL-2) was determined. The results demonstrate that concentrations of DEX and PGE2 which individually do not significantly suppress IL-2 synthesis act together to inhibit the synthesis of IL-2 synergistically. 5. The addition of an exogenous source of recombinant IL-2 (rIL-2) to T cells stimulated in the presence of DEX and PGE2 completely reversed the synergistic antiproliferative effect of these two compounds. This reversal was even more pronounced with T cells cultured at a suboptimal cell density. Additionally, PGE2 and DEX did not affect expression of the IL-2 receptor (IL-2R), as measured by upregulation of the alpha chain, on anti-CD3 mAb stimulated T cells. 6. Collectively these data indicate that physiologic concentrations of PGE2 and DEX, which alone have no effect on anti-CD3 mAb-induced T-cell proliferation, act synergistically to inhibit T-cell proliferation as well as IL-2 synthesis.

T-cell antigenic sites involved in myasthenia gravis: correlations with antibody titre and disease severity.

PubMed

Berrih-Aknin, S; Cohen-Kaminsky, S; Lepage, V; Neumann, D; Bach, J F; Fuchs, S

1991-02-01

We have evaluated the ability of eight synthetic peptides corresponding to selected regions of the alpha-subunit from human (H) or Torpedo (T) acetylcholine receptor (AChR) to stimulate proliferative responses of peripheral blood lymphocytes (PBL) and thymic cells from patients with Myasthenia Gravis (MG) in comparison to healthy controls. Using PBL, two of the peptides were most reactive: in the 40 myasthenic patients tested, peptide 169-181 (H) induced significant proliferative responses in 10 patients and peptide 351-368 (H) in five, while there was no response in any of the 34 healthy controls tested. Interestingly, clear associations between proliferation to peptides and clinical data were observed. Indeed, among responding patients, all presented thymic hyperplasia and most showed a high anti-AChR Ab titre and/or a severe form of the disease. In addition, responses to AChR cytoplasmic sequences were observed only in severely affected patients. Correlation with HLA-DR haplotype, sought in a subgroup of patients, indicated that response to 169-181 (H) is associated with HLA-DR5 in the patients presenting a high anti-AChR antibody titre. Using thymic lymphocytes, few responses were obtained with the human peptides, suggesting that the frequency of autoreactive cells is lower than in the blood. Similar to PBL, responses to peptides were observed only with lymphocytes isolated from hyperplastic thymuses. The correlations observed between responses to peptides and clinical parameters underline the pathophysiological relevance of our data and indicate that pathogenic and nonpathogenic T-cell antigenic sites involved in the anti-AChR response could be identified by this approach.

Lymphocyte responses to influenza and tetanus toxoid in vitro following intensive exercise and carbohydrate ingestion on consecutive days.

PubMed

Bishop, Nicolette C; Walker, Gary J; Bowley, Lee A; Evans, Kate F; Molyneux, Karen; Wallace, Fiona A; Smith, Alice C

2005-10-01

The effect of carbohydrate (CHO) ingestion on antigen- (rather than mitogen-) stimulated T-cell responses to prolonged, intensive exercise may give a more realistic insight into the effect of CHO on T-cell functional capacity and subsequent infection risk. This study investigated the effect of CHO ingestion during prolonged, intensive exercise on influenza- and tetanus toxoid-stimulated T-cell cytokine mRNA expression and proliferation. Mitogen- [phytohemagglutinin (PHA)] stimulated proliferation was assessed for comparison. Responses were assessed following exercise on consecutive mornings to determine any carryover effect. Fifteen male games players performed two exercise trials in a double-blind, randomized, crossover design. Each trial comprised 90 min of intensive, intermittent running on consecutive mornings, with either CHO (6.4% wt/vol) or placebo (PLA) beverage ingestion before, during, and after each bout of exercise. Postexercise CD3(+) cell counts were higher in PLA than CHO on both days (P < 0.05). Antigen-stimulated T-cell cytokine mRNA expression was unaffected by exercise or CHO ingestion. Before exercise on day 2, T-cell proliferative responses to PHA, influenza, and tetanus toxoid were higher in CHO than PLA by 99, 80, and 58%, respectively (P < 0.01 for PHA, P < 0.05 for influenza and tetanus toxoid). At 1 h postexercise on day 2, PHA-induced proliferation was 70% higher in CHO than PLA (P < 0.05), yet there were no differences between trials for antigen-induced proliferative responses. Therefore, mitogen-induced T-cell proliferation following strenuous exercise and CHO does not necessarily reflect responses to specific antigens and, consequently, may not provide a good model for the situation in vivo.

GAS6 Receptor Status Is Associated with Dormancy and Bone Metastatic Tumor Formation

PubMed Central

Taichman, Russell S.; Patel, Lalit R.; Bedenis, Rachel; Wang, Jingcheng; Weidner, Savannah; Schumann, Taibriana; Yumoto, Kenji; Berry, Janice E.; Shiozawa, Yusuke; Pienta, Kenneth J.

2013-01-01

Disseminated tumor cells (DTCs) are believed to lie dormant in the marrow before they can be activated to form metastases. How DTCs become dormant in the marrow and how dormant DTCs escape dormancy remains unclear. Recent work has shown that prostate cancer (PCa) cell lines express the growth-arrest specific 6 (GAS6) receptors Axl, Tyro3, and Mer, and become growth arrested in response to GAS6. We therefore hypothesized that GAS6 signaling regulates the proliferative activity of DTCs in the marrow. To explore this possibility, in vivo studies were performed where it was observed that when Tyro3 expression levels exceed Axl expression, the PCa cells exhibit rapid growth. When when Axl levels predominate, PCa cells remain largely quiescent. These findings suggest that a balance between the expression of Axl and Tyro3 is associated with a molecular switch between a dormant and a proliferative phenotype in PCa metastases. PMID:23637920

The yellow fever virus vaccine induces a broad and polyfunctional human memory CD8+ T cell response.

PubMed

Akondy, Rama S; Monson, Nathan D; Miller, Joseph D; Edupuganti, Srilatha; Teuwen, Dirk; Wu, Hong; Quyyumi, Farah; Garg, Seema; Altman, John D; Del Rio, Carlos; Keyserling, Harry L; Ploss, Alexander; Rice, Charles M; Orenstein, Walter A; Mulligan, Mark J; Ahmed, Rafi

2009-12-15

The live yellow fever vaccine (YF-17D) offers a unique opportunity to study memory CD8(+) T cell differentiation in humans following an acute viral infection. We have performed a comprehensive analysis of the virus-specific CD8(+) T cell response using overlapping peptides spanning the entire viral genome. Our results showed that the YF-17D vaccine induces a broad CD8(+) T cell response targeting several epitopes within each viral protein. We identified a dominant HLA-A2-restricted epitope in the NS4B protein and used tetramers specific for this epitope to track the CD8(+) T cell response over a 2 year period. This longitudinal analysis showed the following. 1) Memory CD8(+) T cells appear to pass through an effector phase and then gradually down-regulate expression of activation markers and effector molecules. 2) This effector phase was characterized by down-regulation of CD127, Bcl-2, CCR7, and CD45RA and was followed by a substantial contraction resulting in a pool of memory T cells that re-expressed CD127, Bcl-2, and CD45RA. 3) These memory cells were polyfunctional in terms of degranulation and production of the cytokines IFN-gamma, TNF-alpha, IL-2, and MIP-1beta. 4) The YF-17D-specific memory CD8(+) T cells had a phenotype (CCR7(-)CD45RA(+)) that is typically associated with terminally differentiated cells with limited proliferative capacity (T(EMRA)). However, these cells exhibited robust proliferative potential showing that expression of CD45RA may not always associate with terminal differentiation and, in fact, may be an indicator of highly functional memory CD8(+) T cells generated after acute viral infections.

DEC1/STRA13 is a key negative regulator of activation-induced proliferation of human B cells highly expressed in anergic cells.

PubMed

Camponeschi, Alessandro; Todi, Laura; Cristofoletti, Cristina; Lazzeri, Cristina; Carbonari, Maurizio; Mitrevski, Milica; Marrapodi, Ramona; Del Padre, Martina; Fiorilli, Massimo; Casato, Milvia; Visentini, Marcella

2018-06-01

The transcription factor DEC1/STRA13 (also known as BHLHE40 and SHARP2) is involved in a number of processes including inhibition of cell proliferation and delay of cell cycle, and is a negative regulator of B cell activation and development in mice. We show here that, unlike in mice, DEC1/STRA13 expression is induced in human naïve and memory resting B cells by activation through the B-cell receptor (BCR) or Toll-like receptor 9 (TLR9). siRNA silencing of DEC1/STRA13 increases the capacity of activated B cells to perform a high number of divisions after TLR9 ligation. This identifies DEC1/STRA13 as a critical negative regulator of clonal expansion of activated human B cells. We also show that DEC1/STRA13 is upregulated in human anergic CD21 low B cells clonally expanded in patients with HCV-associated mixed cryoglobulinemia, which fail to proliferate in response to BCR or TLR9 ligation. siRNA knockdown of DEC1/STRA13, however, fails to restore responsiveness to stimuli in these cells, although it might improve the proliferative capacity in a subset of anergic cells with less pronounced proliferative defect. Copyright © 2018 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Enhanced G2 chromatid radiosensitivity, an early stage in the neoplastic transformation of human epidermal keratinocytes in culture

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

Gantt, R.; Sanford, K.K.; Parshad, R.

1987-03-01

A deficiency in DNA repair, manifest as enhanced chromatid radiosensitivity during the G2 phase of the cell cycle, together with a proliferative stimulus such as that provided by active oncogenes may be necessary and sufficient for the malignant neoplastic transformation of human keratinocytes in culture. Normal epidermal keratinocytes established as continuous cell lines by transfection with pSV3-neo or infection with adeno 12-SV40 hybrid virus developed enhanced G2 chromatid radiosensitivity after 18 passages in culture. In contrast to cells from primary or secondary culture, these cells could be transformed to malignant neoplastic cells by infection with Kirsten murine sarcoma virus containingmore » the Ki-ras oncogene or in one line by the chemical carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine; both of these agents produced a marked proliferative response. Cytological heterogeneity and karyotypic instability characterized the cells during their progression to neoplasia. These results are interpreted in terms of a mechanism for neoplastic transformation.« less

Multiple effects of TRAIL in human carcinoma cells: Induction of apoptosis, senescence, proliferation, and cytokine production

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

Levina, Vera; Marrangoni, Adele M.; DeMarco, Richard

TRAIL is a death ligand that induces apoptosis in malignant but not normal cells. Recently the ability of TRAIL to induce proliferation in apoptosis-resistant normal and malignant cells was reported. In this study, we analyzed TRAIL effects in apoptosis sensitive MCF7, OVCAR3 and H460 human tumor cell lines. TRAIL at low concentrations preferentially induced cell proliferation. At 100 ng/ml, apoptotic death was readily observed, however surviving cells acquired higher proliferative capacity. TRAIL-stimulated production of several cytokines, IL-8, RANTES, MCP-1 and bFGF, and activation of caspases 1 and 8 was essential for this effect. Antibodies to IL-8, RANTES, and bFGF blockedmore » TRAIL-induced cell proliferation and further stimulated apoptosis. For the first time, we report that high TRAIL concentrations induced cell senescence as determined by the altered morphology and expression of several senescence markers: SA-{beta}-gal, p21{sup Waf1/Cip1}, p16{sup INK4a}, and HMGA. Caspase 9 inhibition protected TRAIL-treated cells from senescence, whereas inhibition of caspases 1 and 8 increased the yield of SLP cells. In conclusion, in cultured human carcinoma cells, TRAIL therapy results in three functional outcomes, apoptosis, proliferation and senescence. TRAIL-induced proapoptotic and prosurvival responses correlate with the strength of signaling. TRAIL-induced cytokine production is responsible for its proliferative and prosurvival effects.« less

Circulating natural killer T cells in patients with asthma.

PubMed

Ikegami, Yasuhiko; Yokoyama, Akihito; Haruta, Yoshinori; Hiyama, Keiko; Kohno, Nobuoki

2004-01-01

Recent studies suggest that therapies targeted at depletion or limiting of natural killer (NK) T cells may be a possible strategy for the treatment of asthma. In the present study, we measured the number of circulating V alpha24+ NKT cells in 32 asthmatic patients and compared these patients with 29 nonatopic healthy controls. We investigated the relationships between NKT cell number and clinical variables such as the number of eosinophils, the circulating level of IgE, and the severity of asthma. In addition, we also investigated the ability of NKT cells to proliferate in response to alpha-galactosyl ceramide (alpha-GalCer) in vitro. The V alpha24+ NKT cell counts of asthmatic patients were significantly lower than those of healthy controls. There were no significant differences observed in asthmatic patients among the subgroups in terms of atopic status and severity. There was no significant correlation between the number of NKT cells and clinical variables. The proliferative response to alpha-GalCer of the patients and controls was not significantly different, indicating no intrinsic proliferative defect of NKT cells in asthma. These results suggest that the number of circulating NKT cells was already decreased in patients with asthma. Further study, such as the evaluation of lung NKT cells, will be needed to determine the role of NKT cells in patients with asthma.

Opposing effects of TIGAR- and RAC1-derived ROS on Wnt-driven proliferation in the mouse intestine

PubMed Central

Cheung, Eric C.; Lee, Pearl; Ceteci, Fatih; Nixon, Colin; Blyth, Karen; Sansom, Owen J.; Vousden, Karen H.

2016-01-01

Reactive oxygen species (ROS) participate in numerous cell responses, including proliferation, DNA damage, and cell death. Based on these disparate activities, both promotion and inhibition of ROS have been proposed for cancer therapy. However, how the ROS response is determined is not clear. We examined the activities of ROS in a model of Apc deletion, where loss of the Wnt target gene Myc both rescues APC loss and prevents ROS accumulation. Following APC loss, Myc has been shown to up-regulate RAC1 to promote proliferative ROS through NADPH oxidase (NOX). However, APC loss also increased the expression of TIGAR, which functions to limit ROS. To explore this paradox, we used three-dimensional (3D) cultures and in vivo models to show that deletion of TIGAR increased ROS damage and inhibited proliferation. These responses were suppressed by limiting damaging ROS but enhanced by lowering proproliferative NOX-derived ROS. Despite having opposing effects on ROS levels, loss of TIGAR and RAC1 cooperated to suppress intestinal proliferation following APC loss. Our results indicate that the pro- and anti-proliferative effects of ROS can be independently modulated in the same cell, with two key targets in the Wnt pathway functioning to integrate the different ROS signals for optimal cell proliferation. PMID:26679840

Repeated episodes of ozone inhalation attenuates airway injury/repair and release of substance P, but not adaptation.

PubMed

Schelegle, Edward S; Walby, William F; Alfaro, Mario F; Wong, Viviana J; Putney, Lei; Stovall, Mary Y; Sterner-Kock, Anja; Hyde, Dallas M; Plopper, Charles G

2003-02-01

To determine the impact of repeated episodes of ozone exposure on physiologic adaptation, epithelial injury/repair, and tracheal substance P levels, adult rats were subjected to episodes of ozone (5 days, 1 ppm, 8 h/day) followed by 9 days of filtered air for four cycles. Rats were sampled on days 1 and 5 of each episode and 9 days after day 5 of episodes 1, 2, and 4. One hour before being euthanized each rat was injected with 5-bromo-2'-deoxyuridine to label proliferating cells. Each 5-day episode showed a characteristic pattern of rapid shallow breathing (days 1 and 2), epithelial injury, and interstitial and intraluminal inflammation. In contrast, the neutrophil component of inflammation, tracheal substance P release, and cell proliferation became attenuated with each consecutive episode of exposure. Concurrent with this cyclic and attenuated response there was progressive hypercellularity and hyperplasia in all airways studied and a progressive remodeling present in the terminal bronchioles. Our findings are consistent with the notion that the cumulative distal airway lesion is at least in part the result of a depressed cell proliferative response to injury in these airways. This depressed cell proliferative response may be in part the result of diminished neutrophil inflammation and/or release of mitogenic neuropeptides in response to ozone-induced injury.

Characterization of Effector and Memory T Cell Subsets in the Immune Response to Bovine Tuberculosis in Cattle

PubMed Central

Maggioli, Mayara F.; Palmer, Mitchell V.; Thacker, Tyler C.; Vordermeier, H. Martin; Waters, W. Ray

2015-01-01

Cultured IFN-γ ELISPOT assays are primarily a measure of central memory T cell (Tcm) responses with humans; however, this important subset of lymphocytes is poorly characterized in cattle. Vaccine-elicited cultured IFN-γ ELISPOT responses correlate with protection against bovine tuberculosis in cattle. However, whether this assay measures cattle Tcm responses or not is uncertain. The objective of the present study was to characterize the relative contribution of Tcm (CCR7+, CD62Lhi, CD45RO+), T effector memory (Tem, defined as: CCR7-, CD62Llow/int, CD45RO+), and T effector cells (CCR7-, CD62L-/low, CD45RO-), in the immune response to Mycobacterium bovis. Peripheral blood mononuclear cells (PBMC) from infected cattle were stimulated with a cocktail of M. bovis purified protein derivative, rTb10.4 and rAg85A for 13 days with periodic addition of fresh media and rIL-2. On day 13, cultured PBMC were re-stimulated with medium alone, rESAT-6:CFP10 or PPDb with fresh autologous adherent cells for antigen presentation. Cultured cells (13 days) or fresh PBMCs (ex vivo response) from the same calves were analyzed for IFN-γ production, proliferation, and CD4, CD45RO, CD62L, CD44, and CCR7 expression via flow cytometry after overnight stimulation. In response to mycobacterial antigens, ~75% of CD4+ IFN-γ+ cells in long-term cultures expressed a Tcm phenotype while less than 10% of the ex vivo response consisted of Tcm cells. Upon re-exposure to antigen, long-term cultured cells were highly proliferative, a distinctive characteristic of Tcm, and the predominant phenotype within the long-term cultures switched from Tcm to Tem. These findings suggest that proliferative responses of Tcm cells to some extent occurs simultaneously with reversion to effector phenotypes (mostly Tem). The present study characterizes Tcm cells of cattle and their participation in the response to M. bovis infection. PMID:25879774

The Novel PIM1 Inhibitor NMS-P645 Reverses PIM1-Dependent Effects on TMPRSS2/ERG Positive Prostate Cancer Cells And Shows Anti-Proliferative Activity in Combination with PI3K Inhibition.

PubMed

Mologni, Luca; Magistroni, Vera; Casuscelli, Francesco; Montemartini, Marisa; Gambacorti-Passerini, Carlo

2017-01-01

PIM1 is over-expressed in multiple tumors, including prostate cancer (PCa). PIM1 upregulation is mediated by direct binding of the ERG transcription factor to its promoter. About 50% of PCa cases are characterized by the presence of the TMPRSS2/ERG fusion, leading to ERG over-expression and thus to PIM1 transcriptional activation. PIM kinases are considered as weak oncogenes, but when combined with additional genetic alterations can induce strong transforming effects. Here we show anti-proliferative activity of the newly described PIM1 inhibitor NMS-P645 in combination with the PI3K inhibitor GDC-0941 in TMPRSS2/ERG positive and negative PCa cells. Treatment with NMS-P645 alone can reverse PIM1-mediated pro-survival signals in prostate cells, such as activation of STAT3 through Tyr705 phosphorylation and resistance to taxane-based treatments, but does not exert a strong anti-tumoral effect. However, the simultaneous treatment with NMS-P645 and GDC-0941 induces a significant anti-proliferative response in PCa cells. These results support the use of combination strategies with PIM and PI3K inhibitors as effective treatment for PCa cases.

A synthetic cryptochrome inhibitor induces anti-proliferative effects and increases chemosensitivity in human breast cancer cells

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

Chun, Sung Kook; Department of Biological Sciences, Seoul National University, Seoul, 151-747; Department of Brain & Cognitive Sciences, Seoul National University, Seoul, 151-747

Disruption of circadian rhythm is a major cause of breast cancer in humans. Cryptochrome (CRY), a circadian transcription factor, is a risk factor for initiation of breast cancer, and it is differentially expressed between normal and breast cancer tissues. Here, we evaluated the anti-proliferative and pro-apoptotic activity of KS15, a recently discovered small-molecule inhibitor of CRY, in human breast cancer cells. First, we investigated whether KS15 treatment could promote E-box-mediated transcription by inhibiting the activity of CRY in MCF-7 human breast cancer cells. Protein and mRNA levels of regulators of cell cycle and apoptosis, as well as core clock genes,more » were differentially modulated in response to KS15. Next, we investigated whether KS15 could inhibit proliferation and increase sensitivity to anti-tumor drugs in MCF-7 cells. We found that KS15 decreased the speed of cell growth and increased the chemosensitivity of MCF-7 cells to doxorubicin and tamoxifen, but had no effect on MCF-10A cells. These findings suggested that pharmacological inhibition of CRY by KS15 exerts an anti-proliferative effect and increases sensitivity to anti-tumor drugs in a specific type of breast cancer. - Highlights: • Cryptochrome inhibitor (KS15) has anti-tumor activity to human breast cancer cells. • KS15 induces differential changes in cell cycle regulators and pro-apoptotic genes. • KS15 inhibits MCF-7 cell growth and enhances susceptibility to anti-tumor drugs.« less

Overexpression of E2F3 promotes proliferation of functional human β cells without induction of apoptosis

PubMed Central

Rady, Brian; Chen, Yanmei; Vaca, Pilar; Wang, Qian; Wang, Yong; Salmon, Patrick; Oberholzer, José

2013-01-01

The mechanisms that control proliferation, or lack thereof, in adult human β cells are poorly understood. Controlled induction of proliferation could dramatically expand the clinical application of islet cell transplantation and represents an important component of regenerative approaches to a functional cure of diabetes. Adult human β cells are particularly resistant to common proliferative targets and often dedifferentiate during proliferation. Here we show that expression of the transcription factor E2F3 has a role in regulating β-cell quiescence and proliferation. We found human islets have virtually no expression of the pro-proliferative G1/S transcription factors E2F1–3, but an abundance of inhibitory E2Fs 4–6. In proliferative human insulinomas, inhibitory E2Fs were absent, while E2F3 is expressed. Using this pattern as a “roadmap” for proliferation, we demonstrated that ectopic expression of nuclear E2F3 induced significant expansion of insulin-positive cells in both rat and human islets. These cells did not undergo apoptosis and retained their glucose-responsive insulin secretion, showing the ability to reverse diabetes in mice. Our results suggest that E2F4–6 may help maintain quiescence in human β cells and identify E2F3 as a novel target to induce proliferation of functional β cells. Refinement of this approach may increase the islets available for cell-based therapies and research and could provide important cues for understanding in vivo proliferation of β cells. PMID:23907129

Inhibition of lymphocyte proliferative responses to Helicobacter pylori by plastic adherent cells.

PubMed

Uyub, A M; Anuar, A K

2001-03-01

A study was carried out on 49 H. pylori-positive and 11 H. pylori-negative patients to determine the reactivity of peripheral blood lymphocytes (PBL) to phytohemagglutinin (PHA) and acid glycine extract (AGE) of H. pylori, and to identify cells responsible for imunosuppression. Based on response to PHA stimulation, cell-mediated immunity of all patients were competent. In some patients, however, response to AGE of H. pylori was suppressed by plastic adherent cells. This study provided evidence of the presence of plastic adherent suppressor cells which suppressed PBL response to AGE of H. pylori but not to PHA suggesting that immunosuppression is antigen specific. There is also an indication that immunosuppression may be species-specific as PBL devoid of plastic adherent cells only responded to stimulation by AGE of H. pylori but not that to AGE of C. jejuni.

SPECIFICITIES OF ENDOMETRIAL PROLIFERATION/STEM CELL INDEX DISTRIBUTION IN ENDOMETRIOID CARCINOMA OF DIFFERENT GRADE OF MALIGNANCY.

PubMed

Kikalishvili, N; Beriashvili, R; Muzashvili, T; Burkadze, G

2018-03-01

Endometrial neoplasia is the most common malignant tumor of female genital system in developed countries. The incidence of endometrial cancer has increased in the last years and despite advances in diagnosis and treatment, the death rates have steadily been increasing over the past 20 years. Therefore aspects of endometrial cancer development, pathogenesis and effective treatment is especially urgent to this day, as much of the risk for endometrial cancer development is influenced by the environment and lifestyle. Endometrial stem cells take the special place among somatic stem cells of female reproductive system-the detection of them and identification of their location in the complex cellular hierarchy still remains challenging. Further study of endometrial stem cells will clarify their role in gynecologic pathologies associated with hyper-proliferative states of endometrium. The aim of our study was to explore the specificities of endometrial proliferative/stem cell index distribution under endometrioid carcinoma of different grade of malignancy. The study represents a retrospective research. The coded and depersonalized material data from Acad. N. Kipshidze Central University Clinic was used in the study. 3 study groups - 1st study group "Endometrioid Carcinoma Grade 1" (14 cases), 2nd study group "Endometrioid Carcinoma Grade 2" (23 cases) and 3rd study group "Endometrioid Carcinoma Grade 3" were selected from routine histopathology tissue specimens of uterus. Hematoxilyn-eosin technology and immunohistochemistry with proliferation marker ki67 and stem cell marker CD146 was performed. The proliferative/stem cell index was calculated by the ratio of Ki67-positive cell percentage value divided by CD146-positive cell percentage value. The study showed that in the 1st study group labeled as "Endometrioid Carcinoma Grade 1", the proliferative/stem cell index ranges between 21.7 and 25.5. Its mean average value in the age distribution subgroups accounts for: 1.1) reproductive age - 22.4; 1.2) menopause - 23.5; 1.3) post-menopause - 24.8. Proliferative/stem cell index reaches its maximum in the samples retrieved from post-menopause age, and decreases significantly in reproductive age individuals. In the 2nd study group labeled as "Endometrioid Carcinoma Grade 2", the proliferative/stem cell index increases and ranges within the interval 23.2-27.8. Its mean average value in the age distribution subgroups accounts for: 2.1) reproductive age -23.7; 2.2) menopause - 24.2; 2.3) post-menopause - 25.8. In the 3rd study group labeled as "Endometrioid Carcinoma Grade 3", the proliferative/stem cell index markedly increases and ranges within the interval 25.8-29.4. Its mean average value in the age distribution subgroups accounts for: 3.1) reproductive age - 28.4; 3.2) menopause - 28.5; 3.3) post-menopause - 28.5. It was found that average value of proliferative/stem cell index in the 1st and 2nd study groups (EC Grade 1/2) keeps the same tendencies of increase in age subgroups as well as at endometrial hyperplasia conditions - in particular in both study groups increase in value of the proliferative/stem cell index in age subgroups makes about 1% (1st study group-0,97%, 2nd study group-0,96%). What about 3rd study group (EC Grade 3) average value of proliferative/stem cell index in age subgroups is almost the same. It was found that average value of proliferative/stem cell index in endometrioid carcinoma most markedly differs from the norm in post-menopause period. The study showed that average value of proliferative/stem cell index in endometrioid carcinoma cases (EC Grade 1/2) tends to increase with age like endometrial hyperplasia conditions, in contrast with the norm, where it is observed to progressively decrease with aging. The attention should be given to the fact that the mean average value of proliferative/stem cell index in endometrioid carcinoma Grade 3 is almost constant.

Prophylactic fenbendazole therapy does not affect the incidence and onset of type 1 diabetes in non-obese diabetic mice.

PubMed

Franke, Deanna D H; Shirwan, Haval

2006-03-01

Fenbendazole (FBZ) is a common, highly efficacious broad-spectrum anthelmintic drug used to treat and limit rodent pinworm infections. However, the effect of its prophylactic use on the immune response of rodents is largely undefined. The non-obese diabetic (NOD) mouse is a model commonly used to study type 1 diabetes (T1D). Parasitic infections will inhibit diabetes development in NOD mice; thus, in the presence of contamination, prophylactic treatment with anthelmintics must be considered to maintain experimental research. Herein, we investigated the prophylactic use of FBZ in NOD mice to determine its effect on the incidence and onset of diabetes, lymphocyte sub-populations and T cell proliferative responses. NOD mice were separated into control and treatment groups. The treatment group received a diet containing FBZ. Animals were monitored for the incidence and onset of T1D. At matched time points, diabetic and non-diabetic mice were killed and splenic lymphocytes analyzed for various cell sub-populations and mitogen-induced proliferative responses using flow cytometry. Treated and control mice were monitored >23 weeks with no detectable effects on the incidence or onset of diabetes. Moreover, no significant differences were detected in lymphocyte sub-populations and mitogen-induced CD4(+) and CD8(+) proliferative responses between control and treatment groups. These results suggest that prophylactic FBZ treatment does not significantly alter the incidence or onset of diabetes in NOD mice. The prophylactic use of FBZ, therefore, presents a viable approach for the prevention of pinworm infection in precious experimental animals with substantial scientific and economic benefits.

Effect of L-arginine supplementation on immune responsiveness in patients with sickle cell disease.

PubMed

Scavella, Arnette; Leiva, Lily; Monjure, Hanh; Zea, Arnold H; Gardner, Renee V

2010-08-01

L-arginine (L-Arg) is deficient in sickle cell disease (SSD) during vasoocclusion. We investigated possible causal relationship between L-Arg deficiency and immune dysfunction in SSD in steady-state. Fifteen patients with SSD in steady-state and 13 controls were studied. Plasma L-Arg levels were measured using liquid chromatography. T cell subsets and CD3zeta (CD3zeta) chain expression were analyzed using flow cytometry. Lymphocyte proliferative response to phytohemagglutinin (PHA) and production of IL-6 and interferon-gamma (IFN-gamma) were evaluated with and without L-Arg. SSD patients had significantly lower L-Arg levels than controls. CD3 and CD19 cell populations were comparable for both groups, but SSD patients had above normal numbers of natural killer cells (P = 0.06). Patients and controls exhibited significantly increased lymphocyte blastogenesis to PHA after introduction of L-Arg to cultures; response of patients was significantly greater than values for control individuals. Proliferative response to candida in SSD patients was significantly lower than in controls; L-Arg supplementation did not increase this response. L-Arg had no effect on blastogenic response to PPD and candida albicans. No effect was likewise seen in production of IL-6 and IFN-gamma after addition of L-Arg. CD3zeta chain expression increased after addition of L-Arg in both groups; differences were insignificant. L-Arg levels in steady-state SSD are significantly lower than in controls. L-Arg supplementation enhanced lymphocyte blastogenesis to PHA for both controls and patients, but not in response to antigen. There were no significant differences in CD3zeta chain expression although upregulation of expression occurred after L-Arg supplementation for both groups. (c) 2010 Wiley-Liss, Inc.

COMPARISON OF PROLIFERATIVE CAPACITY OF GENETICALLY-ENGINEERED PIG AND HUMAN CORNEAL ENDOTHELIAL CELLS

PubMed Central

Fujita, Minoru; Mehra, Ruhina; Lee, Seung Eun; Roh, Danny S.; Long, Cassandra; Funderburgh, James L.; Ayares, David L.; Cooper, David K. C.; Hara, Hidetaka

2013-01-01

Purpose The possibility of providing cultured corneal endothelial cells (CECs) for clinical transplantation has gained much attention. However, the worldwide need for human (h) donor corneas far exceeds supply. The pig (p) might provide an alternative source. The aim of this study was to compare the proliferative capacity of CECs from wild-type (WT) pigs, genetically-engineered (GE) pigs, and humans. Methods The following CECs were cultured – hCECs from donors (i) ≤36 years (young), (ii) ≥49 years (old), and WT pCECs from (iii) neonatal (<5 days), (iv) young (<2 months), and (v) old (>20 months) pigs, and CECs from young (vi) GE pigs (GTKO/CD46 and GTKO/CD46/CD55). Proliferative capacity of CECs was assessed by direct cell counting over 15 days of culture and by BrdU assay. Cell viability during culture was assessed by annexin V staining. The MTT assay assessed cell metabolic activity. Results There was significantly lower proliferative capacity of old CECs than of young CECs (p<0.01) in both pigs and humans. There was no significant difference in proliferative capacity/metabolic activity between young pCECs and young hCECs. However, there was a significantly higher percentage of cell death in hCECs compared to pCECs during culture (p<0.01). Young GE pCECs showed similar proliferative capacity/cell viability/metabolic activity to young WT pCECs. Conclusions Because of the greater availability of young pigs and the excellent proliferative capacity of cultured GE pCECs, GE pigs could provide a source of CECs for clinical transplantation. PMID:23258190

Radiosensitization by PARP Inhibition in DNA Repair Proficient and Deficient Tumor Cells: Proliferative Recovery in Senescent Cells

PubMed Central

Alotaibi, Moureq; Sharma, Khushboo; Saleh, Tareq; Povirk, Lawrence F.; Hendrickson, Eric A.; Gewirtz, David A.

2016-01-01

Radiotherapy continues to be a primary modality in the treatment of cancer. DNA damage induced by radiation can promote apoptosis as well as both autophagy and senescence, where autophagy and senescence can theoretically function to prolong tumor survival. A primary aim of this work was to investigate the hypothesis that autophagy and/or senescence could be permissive for DNA repair, thereby facilitating tumor cell recovery from radiation-induced growth arrest and/or cell death. In addition, studies were designed to elucidate the involvement of autophagy and senescence in radiation sensitization by PARP inhibitors and the re-emergence of a proliferating tumor cell population. In the context of this work, the relationship between radiation-induced autophagy and senescence was also determined. Studies were performed using DNA repair proficient HCT116 colon carcinoma cells and a repair deficient Ligase IV (−/−) isogenic cell line. Irradiation promoted a parallel induction of autophagy and senescence that was strongly correlated with the extent of persistent H2AX phosphorylation in both cell lines; however inhibition of autophagy failed to suppress senescence, indicating that the two responses were dissociable. Irradiation resulted in a transient arrest in the HCT116 cells while arrest was prolonged in the Ligase IV (−/−) cells; however, both cell lines ultimately recovered proliferative function, which may reflect maintenance of DNA repair capacity. The PARP inhibitors (Olaparib) and (Niraparib) increased the extent of persistent DNA damage induced by radiation as well as the extent of both autophagy and senescence; neither cell line underwent significant apoptosis by radiation alone or in the presence of the PARP inhibitors. Inhibition of autophagy failed to attenuate radiation sensitization, indicating that autophagy was not involved in the action of the PARP inhibitors. As with radiation alone, despite sensitization by PARP inhibition, proliferative recovery was evident within a period of 10–20 days. While inhibition of DNA repair via PARP inhibition may initially sensitize tumor cells to radiation via the promotion of senescence, this strategy does not appear to interfere with proliferative recovery, which could ultimately contribute to disease recurrence. PMID:26934368

Dose-dependent responses of I3C and DIM on T-cell activation in the human T lymphocyte Jurkat cell line

USDA-ARS?s Scientific Manuscript database

Indole-3-carbinol (I3C) and its dimer diindolylmethane (DIM) are bioactive metabolites of a glucosinolate glucobrassicin found in cruciferous vegetables. Both I3C and DIM have been reported to possess anti-apoptotic, anti-proliferative and anti-carcinogenic properties via the modulation of immune p...

Ki-67 staining for determination of rhesus macaque T cell proliferative responses ex vivo1

PubMed Central

Shedlock, Devon J.; Talbott, Kendra T.; Morrow, Matthew P.; Ferraro, Bernadette; Hokey, David A.; Muthumani, Karuppiah; Weiner, David B.

2010-01-01

The capacity for robust proliferation upon re-infection is a hallmark of adaptive immunity and the basis of vaccination. A widely used animal model for the study of human disease is the rhesus macaque (RM), where capacity for proliferation can be assessed ex vivo using carboxyfluorescein succinimidyl ester (CFSE)-based dilution assays. However, we show over the course of the standard ex vivo proliferation assay that CFSE-labeling at commonly-used dye concentrations induces significant cell death, but that this phenomenon is dose-dependent. Here we describe an alternative, semi-quantitative method for estimating T cell proliferative responses that avoids the putative biases associated with chemical modification. RM peripheral blood mononuclear cells were stimulated ex vivo with cognate peptides for five days, immunostained for intracellular Ki-67, and then analyzed by flow cytometry. We describe a gating strategy using Ki-67 and side light scatter, also a marker of blastogenesis, which correlates strongly with data from CFSE dilution. We show that this method is a valid tool for measuring RM antigen-specific cellular proliferation ex vivo and can be used as an alternative to CFSE dilution assays. PMID:20104580

Emodin inhibits splenocyte proliferation and inflammation by modulating cytokine responses in a mouse model system.

PubMed

Sharma, Rahul; Tiku, Ashu Bhan

2016-01-01

Emodin, an anthraquinone derivative, was investigated for potential anti-inflammatory and anti-proliferative effects in vitro. The potential to induce these outcomes was assessed using concanavalin A (ConA)-stimulated mouse splenocytes. Dose-response studies showed that emodin at 100 µM was not cytotoxic to naive cells, and that the same dose caused proliferation to be significantly reduced in ConA-stimulated cells. In addition, emodin significantly reduced ConA-induced nitric oxide (NO) production and the formation/release of TH1 (IL-2, IFNγ, TNFα) and TH17 (IL-6 and IL-17) cell cytokines, but induced those of TH2 (IL-4) and Treg (IL-10) cells. From the results, it is concluded that earlier-reported immunomodulatory effects imparted by emodin may have been attributable, in part, to anti-proliferative effects on lymphocytes, as well as a shift within the TH1/TH2 and TH17/Treg balance (towards TH2 and Treg). These findings, while providing evidence of mechanisms of emodin immunomodulation, are also potentially important for sparking studies that ultimately may result in the potential use of this agent in preventive and/or corrective strategies against autoimmune and other inflammatory diseases.

Hypergravity Stimulates the Extracellular Matrix/Integrin-Signaling Axis and Proliferation in Primary Osteoblasts

NASA Technical Reports Server (NTRS)

Parra, M.; Vercoutere, W.; Roden, C.; Banerjee, I.; Krauser, W.; Holton, E.; Searby, N.; Globus, R.; Almeida, E.

2003-01-01

We set out to determine the molecular mechanisms involved in the proliferative response of primary rat osteoblasts to mechanical stimulation using cell culture centrifugation as a model for hypergravity. We hypothesized that this proliferative response is mediated by specific integrin/Extracellular Matrix (ECM) interactions. To investigate this question we developed a cell culture centrifuge and an automated system that performs cell fixation during hypergravity loading. We generated expression vectors for various focal adhesion and cytoskeletal proteins fused to GFP or dsRed and visualized these structures in transfected (or infected) osteoblasts. The actin cytoskeleton was also visualized using rhodamine-phalloidin staining and Focal Adhesion Kinase (FAK) levels were assessed biochemically. We observed that a 24 hour exposure to 50-g stimulated proliferation compared to the 1-g control when cells were plated on fibronectin, collagen Type I , and collagen Type IV, but not on uncoated tissue culture plastic surfaces. This proliferative response was greatest for osteoblasts grown on fibronectin (2-fold increase over 1-g control) and collagen Type I (1.4 fold increase over 1-g control), suggesting that specific matrices and integrins are involved in the signaling pathways required for proliferation. Exposing osteoblasts grown on different matrices to 10-g or 25-g showed that effects on proliferation depended on both matrix type and loading level. We found that osteoblasts exposed to a short pulse of hypergravity during adhesion spread further and had more GFP-FAK containing focal adhesions compared to their 1-g controls. While overall levels of FAK did not change, more FAK was in the active (phosphorylated) form under hypergravity than in the 1-g controls. Cytoskeletal F-actin organization into filaments was also more prominent after brief exposures to hypergravity during the first five minutes of adhesion. These results suggest that specific integrins sense hypergravity and activate distinct matrix-dependent FAK signaling pathways that can enhance proliferation. Our results also imply that brief exposures to hypergravity accelerate cell adhesion and spreading processes via the focal adhesion-signaling axis. These results support the role of the ECM/integrin-signaling axis in osteoblast response to hypergravity loading.

Ultraviolet B irradiation induces expansion of intraepithelial tumor cells in a tissue model of early cancer progression.

PubMed

Mudgil, Adarsh V; Segal, Nadav; Andriani, Frank; Wang, Youai; Fusenig, Norbert E; Garlick, Jonathan A

2003-07-01

Ultraviolet B irradiation is thought to enable skin cancer progression as clones of genetically damaged keratinocytes escape apoptosis and expand at the expense of adjacent normal cells. Mechanisms through which potentially malignant cells in human skin undergo clonal expansion, however, are not well understood. The goal of this study was to characterize the role of ultraviolet B irradiation on the intraepithelial expansion of early stage human tumor cells in organotypic skin cultures. To accomplish this, we have studied the effect of ultraviolet B irradiation on organotypic cultures that were fabricated by mixing normal human keratinocytes with beta-galactosidase-marked, intraepithelial tumor cells (HaCaT-ras, clone II-4), which bear mutations in both p53 alleles and harbor an activated H-ras oncogene. We found that when organotypic mixtures were exposed to an ultraviolet B dose of 50 mJ per cm2, intraepithelial tumor cells underwent a significant degree of proliferative expansion compared to nonirradiated cultures. To understand this response, organotypic cultures of nor-mal keratinocytes were exposed to ultraviolet B and showed a dose-dependent increase in numbers of sunburn cells and TUNEL-positive cells although their proliferation was suppressed. In contrast, neither the apoptotic nor the proliferative response of II-4 cells was altered by ultraviolet B in organotypic cultures. The differential response of these cell types suggested that II-4 cells were resistant to ultraviolet-B-induced alterations, which allowed these intraepithelial tumor cells to gain a selective growth and survival advantage relative to neighboring normal cells. These findings demonstrate that ultraviolet B exposure can induce the intraepithelial expansion of apoptosis-resistant, p53-mutant, and ras-activated keratinocytes, suggesting that this agent can act to promote the early stages of epithelial carcinogenesis.

dMyc is required in retinal progenitors to prevent JNK-mediated retinal glial activation

PubMed Central

Correia, Andreia; Santos, Marília A.; Relvas, João B.; Pereira, Paulo S.

2017-01-01

In the nervous system, glial cells provide crucial insulation and trophic support to neurons and are important for neuronal survival. In reaction to a wide variety of insults, glial cells respond with changes in cell morphology and metabolism to allow repair. Additionally, these cells can acquire migratory and proliferative potential. In particular, after axonal damage or pruning the clearance of axonal debris by glial cells is key for a healthy nervous system. Thus, bidirectional neuron-glial interactions are crucial in development, but little is known about the cellular sensors and signalling pathways involved. In here, we show that decreased cellular fitness in retinal progenitors caused by reduced Drosophila Myc expression triggers non cell-autonomous activation of retinal glia proliferation and overmigration. Glia migration occurs beyond its normal limit near the boundary between differentiated photoreceptors and precursor cells, extending into the progenitor domain. This overmigration is stimulated by JNK activation (and the function of its target Mmp1), while proliferative responses are mediated by Dpp/TGF-β signalling activation. PMID:28267791

Zinc Transporter SLC39A7/ZIP7 Promotes Intestinal Epithelial Self-Renewal by Resolving ER Stress

PubMed Central

Ohashi, Wakana; Kimura, Shunsuke; Iwanaga, Toshihiko; Furusawa, Yukihiro; Irié, Tarou; Izumi, Hironori; Watanabe, Takashi; Hara, Takafumi; Ohara, Osamu; Koseki, Haruhiko; Sato, Toshiro; Robine, Sylvie; Mori, Hisashi; Hattori, Yuichi; Mishima, Kenji; Ohno, Hiroshi; Hase, Koji; Fukada, Toshiyuki

2016-01-01

Zinc transporters play a critical role in spatiotemporal regulation of zinc homeostasis. Although disruption of zinc homeostasis has been implicated in disorders such as intestinal inflammation and aberrant epithelial morphology, it is largely unknown which zinc transporters are responsible for the intestinal epithelial homeostasis. Here, we show that Zrt-Irt-like protein (ZIP) transporter ZIP7, which is highly expressed in the intestinal crypt, is essential for intestinal epithelial proliferation. Mice lacking Zip7 in intestinal epithelium triggered endoplasmic reticulum (ER) stress in proliferative progenitor cells, leading to significant cell death of progenitor cells. Zip7 deficiency led to the loss of Olfm4+ intestinal stem cells and the degeneration of post-mitotic Paneth cells, indicating a fundamental requirement for Zip7 in homeostatic intestinal regeneration. Taken together, these findings provide evidence for the importance of ZIP7 in maintenance of intestinal epithelial homeostasis through the regulation of ER function in proliferative progenitor cells and maintenance of intestinal stem cells. Therapeutic targeting of ZIP7 could lead to effective treatment of gastrointestinal disorders. PMID:27736879

Epidermal Homeostasis and Radiation Responses in a Multiscale Tissue Modeling Framework

NASA Technical Reports Server (NTRS)

Hu, Shaowen; Cucinotta, Francis A.

2013-01-01

The surface of skin is lined with several thin layers of epithelial cells that are maintained throughout life time by a small population of stem cells. High dose radiation exposures could injure and deplete the underlying proliferative cells and induce cutaneous radiation syndrome. In this work we propose a multiscale computational model for skin epidermal dynamics that links phenomena occurring at the subcellular, cellular, and tissue levels of organization, to simulate the experimental data of the radiation response of swine epidermis, which is closely similar to human epidermis. Incorporating experimentally measured histological and cell kinetic parameters, we obtain results of population kinetics and proliferation indexes comparable to observations in unirradiated and acutely irradiated swine experiments. At the sub-cellular level, several recently published Wnt signaling controlled cell-cycle models are applied and the roles of key components and parameters are analyzed. Based on our simulation results, we demonstrate that a moderate increase of proliferation rate for the survival proliferative cells is sufficient to fully repopulate the area denuded by high dose radiation, as long as the integrity of underlying basement membrane is maintained. Our work highlights the importance of considering proliferation kinetics as well as the spatial organization of tissues when conducting in vivo investigations of radiation responses. This integrated model allow us to test the validity of several basic biological rules at the cellular level and sub-cellular mechanisms by qualitatively comparing simulation results with published research, and enhance our understanding of the pathophysiological effects of ionizing radiation on skin.

Differential effects of arsenic on intracellular free calcium levels and the proliferative response of murine mitogen-stimulated lymphocytes.

PubMed

Goytia-Acevedo, Raquel C; Cebrian, Mariano E; Calderon-Aranda, Emma S

2003-08-01

This study examined the effects of sodium arsenite treatment on free [Ca(2+)]i and cell death in mitogen-activated murine lymphocytes. The main findings of this study were that simultaneous sodium arsenite treatment inhibited PHA- but not Con A-induced T cell proliferation, induced a higher increase in free [Ca(2+)]i and an early increase in the proportion of dead cells in PHA than in Con A activated cells. Sodium arsenite pre-treatment reduced both PHA- and Con A-induced T-cell proliferation. Phorbol myristate ester (PMA) did not prevent the inhibitory effects of both sodium arsenite treatments, suggesting that sodium arsenite did not significantly decreased PKC activation or that its effects occurred on events parallel to PKC activation. Both PHA and Con A increased free [Ca(2+)]i after stimulation, yet the effect was more pronounced in mitogen-activated cells simultaneously treated with sodium arsenite and particularly in those activated with PHA. The increase in free [Ca(2+)]i was in agreement with the early cell death induced by sodium arsenite in PHA-activated cells, a finding consistent with the inhibitory effects on PHA-induced proliferation. Sodium arsenite-induced cell death occurred faster in PHA-activated cells. Further studies are needed to ascertain the relationships between the effects of sodium arsenite on free [Ca(2+)]i levels and the type of cell death induced by sodium arsenite and their relevance for the proliferative response of T cells.

Identification of a role for the nuclear receptor EAR-2 in the maintenance of clonogenic status within the leukemia cell hierarchy

PubMed Central

Ichim, CV; Atkins, HL; Iscove, NN; Wells, RA

2016-01-01

Identification of genes that regulate clonogenicity of acute myelogenous leukemia (AML) cells is hindered by the difficulty of isolating pure populations of cells with defined proliferative abilities. By analyzing the growth of clonal siblings in low passage cultures of the cell line OCI/AML4 we resolved this heterogeneous population into strata of distinct clonogenic potential, permitting analysis of the transcriptional signature of single cells with defined proliferative abilities. By microarray analysis we showed that the expression of the orphan nuclear receptor EAR-2 (NR2F6) is greater in leukemia cells with extensive proliferative capacity than in those that have lost proliferative ability. EAR-2 is expressed highly in long-term hematopoietic stem cells, relative to short-term hematopoietic stem and progenitor cells, and is downregulated in AML cells after induction of differentiation. Exogenous expression of EAR-2 increased the growth of U937 cells and prevented the proliferative arrest associated with terminal differentiation, and blocked differentiation of U937 and 32Dcl3 cells. Conversely, silencing of EAR-2 by short-hairpin RNA initiated terminal differentiation of these cell lines. These data identify EAR-2 as an important factor in the regulation of clonogenicity and differentiation, and establish that analysis of clonal siblings allows the elucidation of differences in gene expression within the AML hierarchy. PMID:21637284

A hypoallergenic variant of the major birch pollen allergen shows distinct characteristics in antigen processing and T-cell activation.

PubMed

Kitzmüller, C; Wallner, M; Deifl, S; Mutschlechner, S; Walterskirchen, C; Zlabinger, G J; Ferreira, F; Bohle, B

2012-11-01

BM4 is a novel genetically engineered variant of the major birch pollen allergen Bet v 1 that lacks the typical Bet v 1-like fold and displays negligible IgE-binding but strong T cell-activating capacity. The aim of this study was to elucidate possible differences between BM4 and Bet v 1 in internalization, antigen processing, and presentation. Proliferative responses to BM4 and Bet v 1 of peripheral blood mononuclear cells and Bet v 1-specific T-cell clones were compared. Fluorescently labeled BM4 and Bet v 1 were used to study surface binding, endocytosis, and intracellular degradation by monocyte-derived DC (mdDC). Both proteins were digested by endolysosomal extracts of mdDC. BM4- and Bet v 1-pulsed mdDC were employed to assess the kinetics of activation of Bet v 1-specific T-cell clones and the polarization of naïve T cells. BM4 displayed a significantly stronger T cell-activating capacity than Bet v 1. Furthermore, BM4 showed increased surface binding and internalization as well as faster endolysosomal degradation compared with Bet v 1. BM4-pulsed mdDC induced enhanced proliferative responses at earlier time-points in Bet v 1-specific T-cell clones and promoted less IL-5 production in T cells than Bet v 1-pulsed mdDC. The loss of the Bet v 1-fold changes the protein's interaction with the human immune system at the level of antigen-presenting cells resulting in altered T-cell responses. By combining low IgE-binding with strong and modulating T cell-activating capacity, BM4 represents a highly interesting candidate for specific immunotherapy of birch pollen allergy. © 2012 John Wiley & Sons A/S.

Fluorescence contrast-enhanced proliferative lesion imaging by enema administration of indocyanine green in a rat model of colon carcinogenesis

PubMed Central

Onda, Nobuhiko; Mizutani-Morita, Reiko; Yamashita, Susumu; Nagahara, Rei; Matsumoto, Shinya; Yoshida, Toshinori; Shibutani, Makoto

2017-01-01

The fluorescent contrast agent indocyanine green (ICG) is approved by the Food and Drug Administration for clinical applications. We previously reported that cultured human colon tumor cells preferentially take up ICG by endocytic activity in association with disruption of their tight junctions. The present study explored ICG availability in fluorescence imaging of the colon to identify proliferative lesions during colonoscopy. The cellular uptake of ICG in cultured rat colon tumor cells was examined using live-cell imaging. Colon lesions in rats administered an ICG-containing enema were further assessed in rats with azoxymethane-induced colon carcinogenesis, using in vivo endoscopy, ex vivo microscopy, and immunofluorescence microscopy. The uptake of ICG by the cultured cells was temperature-dependent. The intracellular retention of the dye in the membrane trafficking system suggested endocytosis as the uptake mechanism. ICG administered via enema accumulated in colon proliferative lesions ranging from tiny aberrant crypt foci to adenomas and localized in proliferating cells. Fluorescence endoscopy detected these ICG-positive colonic proliferative lesions in vivo. The immunoreactivity of the tight-junction molecule occludin was altered in the proliferative lesions, suggesting the disruption of the integrity of tight junctions. These results suggest that fluorescence contrast-enhanced imaging following the administration of an ICG-containing enema can enhance the detection of mucosal proliferative lesions of the colon during colonoscopy. The tissue preference of ICG in the rat model evaluated in this study can be attributed to the disruption of tight junctions, which in turn promotes endocytosis by proliferative cells and the cellular uptake of ICG. PMID:29163827

Nuclear accumulation and activation of p53 in embryonic stem cells after DNA damage.

PubMed

Solozobova, Valeriya; Rolletschek, Alexandra; Blattner, Christine

2009-06-17

P53 is a key tumor suppressor protein. In response to DNA damage, p53 accumulates to high levels in differentiated cells and activates target genes that initiate cell cycle arrest and apoptosis. Since stem cells provide the proliferative cell pool within organisms, an efficient DNA damage response is crucial. In proliferating embryonic stem cells, p53 is localized predominantly in the cytoplasm. DNA damage-induced nuclear accumulation of p53 in embryonic stem cells activates transcription of the target genes mdm2, p21, puma and noxa. We observed bi-phasic kinetics for nuclear accumulation of p53 after ionizing radiation. During the first wave of nuclear accumulation, p53 levels were increased and the p53 target genes mdm2, p21 and puma were transcribed. Transcription of noxa correlated with the second wave of nuclear accumulation. Transcriptional activation of p53 target genes resulted in an increased amount of proteins with the exception of p21. While p21 transcripts were efficiently translated in 3T3 cells, we failed to see an increase in p21 protein levels after IR in embryonal stem cells. In embryonic stem cells where (anti-proliferative) p53 activity is not necessary, or even unfavorable, p53 is retained in the cytoplasm and prevented from activating its target genes. However, if its activity is beneficial or required, p53 is allowed to accumulate in the nucleus and activates its target genes, even in embryonic stem cells.

Interleukin (IL) 15 is a novel cytokine that activates human natural killer cells via components of the IL-2 receptor

PubMed Central

1994-01-01

Interleukin 15 (IL-15) is a novel cytokine that has recently been cloned and expressed. Whereas it has no sequence homology with IL-2, IL- 15 interacts with components of the IL-2 receptor (IL-2R). In the present study we performed a functional analysis of recombinant IL-15 on phenotypically and functionally distinct populations of highly purified human natural killer (NK) cells. The CD56bright subset of human NK cells constitutively expresses the high affinity IL-2R and exhibits a brisk proliferative response after the binding of picomolar amounts of IL-2. Using a proliferation assay, IL-15 demonstrated a very steep dose-response curve that was distinct from the dose-response curve for IL-2. The proliferative effects of IL-15 could be abrogated by anti-IL-2R beta (p75), but not by anti-IL-2R alpha (p55). The proliferative effects of IL-2 on CD56bright NK cells could be inhibited by both antibodies. CD56dim NK cells express the intermediate affinity IL-2R in the absence of the high affinity IL-2R. Activation of CD56dim NK cells by IL-15 was similar to that of IL-2 as measured by enhanced NK cytotoxic activity, antibody-dependent cellular cytotoxicity, and NK cell production of interferon gamma, tumor necrosis factor alpha, and granulocyte/macrophage colony-stimulating factor. The IL-15-enhanced NK cytotoxic activity could be completely blocked by anti-IL-2R beta monoclonal antibody. The binding of radiolabeled IL-2 and IL-15 to CD56dim NK cells was inhibited in the presence of anti-IL-2R beta. Scatchard analysis of radiolabeled IL-15 and IL-2 binding to NK- enriched human lymphocytes revealed the presence of high and intermediate affinity receptors for both ligands. IL-15 is a ligand that activates human NK cells through components of the IL-2R in a pattern that is similar but not identical to that of IL-2. Unlike IL-2, IL-15 is produced by activated monocytes/macrophages. The discovery of IL-15 may increase our understanding of how monocytes/macrophages participate in the regulation of NK cell function. PMID:7523571

Tolerogenic effect of mesenchymal stromal cells on gliadin-specific T lymphocytes in celiac disease.

PubMed

Ciccocioppo, Rachele; Camarca, Alessandra; Cangemi, Giuseppina Cristina; Radano, Giorgia; Vitale, Serena; Betti, Elena; Ferrari, Davide; Visai, Livia; Strada, Elena; Badulli, Carla; Locatelli, Franco; Klersy, Catherine; Gianfrani, Carmen; Corazza, Gino Roberto

2014-08-01

Celiac disease is caused by a dysregulated immune response toward dietary gluten, whose only treatment is a lifelong gluten-free diet. We investigated the effects of mesenchymal stromal cells (MSCs) on gliadin-specific T cells, which are known to induce intestinal lesions, in view of a possible use as new therapy. Bone marrow-derived MSCs and gliadin-specific T-cell lines were obtained from allogeneic donors and mucosal specimens of celiac patients, respectively. The immunosuppressant effect of MSCs was evaluated in terms of proliferative response and interferon (IFN)-γ production upon gliadin stimulation of long-term T-cell lines; the immunomodulant effect was assessed in terms of apoptotic rate, immunophenotype and cytokine profile of short-term T-cell lines generated in the presence of MSCs. Different MSC:T-cell ratios were applied, and statistics were performed as appropriate. MSCs inhibited both proliferative response and IFN-γ production of long-term T-cell lines in a dose-dependent manner while limiting the expansion of short-term T-cell lines by increasing the apoptotic rate. Moreover, a reduction of the CD4(+) population and expansion of the regulatory FoxP3+ subset were found in T-cell lines cultured with MSCs, in which a significant decrease of interleukin (IL)-21, IFN-γ and IL-10 paralleled by an upregulation of transforming growth factor-β1, IL-6 and IL-8 were observed. Finally, an increase of the indoleamine 2,3-dioxygenase activity was found, possibly playing a key role in mediating these effects. MSCs exert potent immunomodulant effects on gliadin-specific T cells, which may be exploited for future therapeutic application in celiac disease. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

PD-1 ligand expression by human colonic myofibroblasts/fibroblasts regulates CD4+ T-cell activity.

PubMed

Pinchuk, Irina V; Saada, Jamal I; Beswick, Ellen J; Boya, Gushyalatha; Qiu, Sumin M; Mifflin, Randy C; Raju, Gottumukkala S; Reyes, Victor E; Powell, Don W

2008-10-01

A prominent role for inhibitory molecules PD-L1 and PD-L2 in peripheral tolerance has been proposed. However, the phenotype and function of PD-L-expressing cells in human gut remains unclear. Recent studies suggest that colonic myofibroblasts (CMFs) and fibroblasts are important in the switch from acute inflammation to adaptive immunity. In the normal human colon, CMFs represent a distinct population of major histocompatibility complex class II(+) cells involved in the regulation of mucosal CD4(+) T-cell responses. PD-L1 and PD-L2 expression on human CMFs was determined using Western blot, fluorescence-activated cell sorter analysis and confocal microscopy. Lymphoproliferation assays and cytokine enzyme-linked immunosorbent assays were used to evaluate the role of B7 costimulators expressed by CMFs with regard to the regulation of preactivated T-helper cell responses. We demonstrate here the expression of PD-L1/2 molecules by normal human CMF and fibroblasts in situ and in culture. Both molecules support suppressive functions of CMFs in the regulation of activated CD4(+) T-helper cell proliferative responses; blocking this interaction reverses the suppressive effect of CMFs on T-cell proliferation and leads to increased production of the major T-cell growth factor, interleukin (IL)-2. PD-L1/2-mediated CMF suppressive functions are mainly due to the inhibition of IL-2 production, because supplementation of the coculture media with exogenous IL-2 led to partial recovery of activated T-cell proliferation. Our data suggest that stromal myofibroblasts and fibroblasts may limit T-helper cell proliferative activity in the gut and, thus, might play a prominent role in mucosal intestinal tolerance.

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

Decursin inhibited proliferation and angiogenesis of endothelial cells to suppress diabetic retinopathy via VEGFR2.

PubMed

Yang, Ying; Yang, Ke; Li, Yiping; Li, Xianli; Sun, Qiangming; Meng, Hua; Zeng, Ying; Hu, Yong; Zhang, Ying

2013-09-25

Diabetes induces pathologic proliferation and angiogenesis in the retina that leads to catastrophic loss of vision. Decursin is a novel therapeutic that targets the vascular endothelial growth factor (VEGF) receptor (VEGFR) with putative anti-proliferative and anti-angiogenic activities. Thereby we utilized human retinal microvascular endothelial cells (HRMEC) and human umbilical vein endothelial cells (HUVEC) under conditions of excess glucose to explore dose-dependent responses of decursin on markers of migration, angiogenesis, and proliferation. Decursin dose-dependently inhibited tube formation, VEGFR-2 expression, along with relative metabolic activity and 5-bromo-2'-deoxy-uridine (BrdU) activity in both cell lines. We then correlated our findings to the streptozotocin-induced rat model of diabetes. Following three months of decursin treatment VEGFR-2 expression was significantly inhibited. Our data would suggest that decursin may be a potent anti-angiogenic and anti-proliferative agent targeting the VEGFR-2 signaling pathway, which significantly inhibits diabetic retinal neovascularization. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Restrictions in Cell Cycle Progression of Adult Vestibular Supporting Cells in Response to Ectopic Cyclin D1 Expression

PubMed Central

Loponen, Heidi; Ylikoski, Jukka; Albrecht, Jeffrey H.; Pirvola, Ulla

2011-01-01

Sensory hair cells and supporting cells of the mammalian inner ear are quiescent cells, which do not regenerate. In contrast, non-mammalian supporting cells have the ability to re-enter the cell cycle and produce replacement hair cells. Earlier studies have demonstrated cyclin D1 expression in the developing mouse supporting cells and its downregulation along maturation. In explant cultures of the mouse utricle, we have here focused on the cell cycle control mechanisms and proliferative potential of adult supporting cells. These cells were forced into the cell cycle through adenoviral-mediated cyclin D1 overexpression. Ectopic cyclin D1 triggered robust cell cycle re-entry of supporting cells, accompanied by changes in p27Kip1 and p21Cip1 expressions. Main part of cell cycle reactivated supporting cells were DNA damaged and arrested at the G2/M boundary. Only small numbers of mitotic supporting cells and rare cells with signs of two successive replications were found. Ectopic cyclin D1-triggered cell cycle reactivation did not lead to hyperplasia of the sensory epithelium. In addition, a part of ectopic cyclin D1 was sequestered in the cytoplasm, reflecting its ineffective nuclear import. Combined, our data reveal intrinsic barriers that limit proliferative capacity of utricular supporting cells. PMID:22073316

Restrictions in cell cycle progression of adult vestibular supporting cells in response to ectopic cyclin D1 expression.

PubMed

Loponen, Heidi; Ylikoski, Jukka; Albrecht, Jeffrey H; Pirvola, Ulla

2011-01-01

Sensory hair cells and supporting cells of the mammalian inner ear are quiescent cells, which do not regenerate. In contrast, non-mammalian supporting cells have the ability to re-enter the cell cycle and produce replacement hair cells. Earlier studies have demonstrated cyclin D1 expression in the developing mouse supporting cells and its downregulation along maturation. In explant cultures of the mouse utricle, we have here focused on the cell cycle control mechanisms and proliferative potential of adult supporting cells. These cells were forced into the cell cycle through adenoviral-mediated cyclin D1 overexpression. Ectopic cyclin D1 triggered robust cell cycle re-entry of supporting cells, accompanied by changes in p27(Kip1) and p21(Cip1) expressions. Main part of cell cycle reactivated supporting cells were DNA damaged and arrested at the G2/M boundary. Only small numbers of mitotic supporting cells and rare cells with signs of two successive replications were found. Ectopic cyclin D1-triggered cell cycle reactivation did not lead to hyperplasia of the sensory epithelium. In addition, a part of ectopic cyclin D1 was sequestered in the cytoplasm, reflecting its ineffective nuclear import. Combined, our data reveal intrinsic barriers that limit proliferative capacity of utricular supporting cells.

Rays Sting: The Acute Cellular Effects of Ionizing Radiation Exposure

PubMed Central

Franco, A; Ciccarelli, M; Sorriento, D; Napolitano, L; Fiordelisi, A; Trimarco, B; Durante, M; Iaccarino, G

2016-01-01

High-precision radiation therapy is a clinical approach that uses the targeted delivery of ionizing radiation, and the subsequent formation of reactive oxygen species (ROS) in high proliferative, radiation sensitive cancers. In particular, in thoracic cancer ratdiation treatments, can not avoid a certain amount of cardiac toxicity. Given the low proliferative rate of cardiac myocytes, research has looked at the effect of radiation on endothelial cells and consequent coronary heart disease as the mechanism of ratdiation induced cardiotoxicity. In fact, little is known concerning the direct effect of radiation on mitochondria dynamis in cardiomyocyte. The main effect of ionizing radiation is the production of ROS and recent works have uncovered that they directly participates to pivotal cell function like mitochondrial quality control. In particular ROS seems to act as check point within the cell to promote either mitochondrial biogenesis and survival or mitochondrial damage and apoptosis. Thus, it appears evident that the functional state of the cell, as well as the expression patterns of molecules involved in mitochondrial metabolism may differently modulate mitochondrial fate in response to radiation induced ROS responses. Different molecules have been described to localize to mitochondria and regulate ROS production in response to stress, in particular GRK2. In this review we will discuss the evidences on the cardiac toxicity induced by X ray radiation on cardiomyocytes with emphasis on the role played by mitochondria dynamism. PMID:27326395

Genome-wide characterization reveals complex interplay between TP53 and TP63 in response to genotoxic stress

PubMed Central

McDade, Simon S.; Patel, Daksha; Moran, Michael; Campbell, James; Fenwick, Kerry; Kozarewa, Iwanka; Orr, Nicholas J.; Lord, Christopher J.; Ashworth, Alan A.; McCance, Dennis J.

2014-01-01

In response to genotoxic stress the TP53 tumour suppressor activates target gene expression to induce cell cycle arrest or apoptosis depending on the extent of DNA damage. These canonical activities can be repressed by TP63 in normal stratifying epithelia to maintain proliferative capacity or drive proliferation of squamous cell carcinomas, where TP63 is frequently overexpressed/amplified. Here we use ChIP-sequencing, integrated with microarray analysis, to define the genome-wide interplay between TP53 and TP63 in response to genotoxic stress in normal cells. We reveal that TP53 and TP63 bind to overlapping, but distinct cistromes of sites through utilization of distinctive consensus motifs and that TP53 is constitutively bound to a number of sites. We demonstrate that cisplatin and adriamycin elicit distinct effects on TP53 and TP63 binding events, through which TP53 can induce or repress transcription of an extensive network of genes by direct binding and/or modulation of TP63 activity. Collectively, this results in a global TP53-dependent repression of cell cycle progression, mitosis and DNA damage repair concomitant with activation of anti-proliferative and pro-apoptotic canonical target genes. Further analyses reveal that in the absence of genotoxic stress TP63 plays an important role in maintaining expression of DNA repair genes, loss of which results in defective repair. PMID:24823795

VEGF-C and TGF-β reciprocally regulate mesenchymal stem cell commitment to differentiation into lymphatic endothelial or osteoblastic phenotypes.

PubMed

Igarashi, Yasuyuki; Chosa, Naoyuki; Sawada, Shunsuke; Kondo, Hisatomo; Yaegashi, Takashi; Ishisaki, Akira

2016-04-01

The direction of mesenchymal stem cell (MSC) differentiation is regulated by stimulation with various growth factors and cytokines. We recently established MSC lines, [transforming growth factor-β (TGF-β)-responsive SG‑2 cells, bone morphogenetic protein (BMP)-responsive SG‑3 cells, and TGF-β/BMP-non-responsive SG‑5 cells], derived from the bone marrow of green fluorescent protein-transgenic mice. In this study, to compare gene expression profiles in these MSC lines, we used DNA microarray analysis to characterize the specific gene expression profiles observed in the TGF-β-responsive SG‑2 cells. Among the genes that were highly expressed in the SG‑2 cells, we focused on vascular endothelial growth factor (VEGF) receptor 3 (VEGFR3), the gene product of FMS-like tyrosine kinase 4 (Flt4). We found that VEGF-C, a specific ligand of VEGFR3, significantly induced the cell proliferative activity, migratory ability (as shown by Transwell migration assay), as well as the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 in the SG‑2 cells. Additionally, VEGF-C significantly increased the expression of prospero homeobox 1 (Prox1) and lymphatic vessel endothelial hyaluronan receptor 1 (Lyve1), which are lymphatic endothelial cell markers, and decreased the expression of osteogenic differentiation marker genes in these cells. By contrast, TGF-β significantly increased the expression of early-phase osteogenic differentiation marker genes in the SG‑2 cells and markedly decreased the expression of lymphatic endothelial cell markers. The findings of our study strongly suggest the following: i) that VEGF-C promotes the proliferative activity and migratory ability of MSCs; and ii) VEGF-C and TGF-β reciprocally regulate MSC commitment to differentiation into lymphatic endothelial or osteoblastic phenotypes, respectively. Our findings provide new insight into the molecular mechanisms underlying the regenerative ability of MSCs.

[Long-term expansion of multipotent mesenchymal stromal cells under reduced oxygen tension].

PubMed

Rylova, Iu V; Buravkova, L B

2013-01-01

We have shown that the decrease in oxygen tension in the culture medium of multipotent mesenchymal stromal cells (MMSCs) results in a short-term reduction in the proportion of CD73(+)-cells in the population, without effecting the number of cells expressing other constitutive surface markers (CD90 and CD105). In this case, the heterogeneity of the cell population declined: large spread cells disappeared. The proliferative activity of MMSCs significantly increased and remained stable in conditions in which the oxygen content was close to the tissue oxygen levels (5% O2). At lower oxygen concentration, proliferative activity of the cells gradually reduced from passages 3-4. The increase in proliferative activity was not accompanied by increased expression of telomerase gene indicateding the alsance of cell transformation. However, genome-wide analysis of MMSC gene expression level revealed changes in expression of cyclins (CCND2 and PCNA), regulatory subunit cyclin-dependent kinase (CKS2) and an inhibitor of cyclin-dependent kinase (CDKN2C), regulating the cell cycle, which is obviously facilitated the increase in the proliferative capacity of cells at lower oxygen tension.

Cannabinoid receptor-dependent and -independent anti-proliferative effects of omega-3 ethanolamides in androgen receptor-positive and -negative prostate cancer cell lines

PubMed Central

Brown, Iain; Cascio, Maria G.; Wahle, Klaus W.J.; Smoum, Reem; Mechoulam, Raphael; Ross, Ruth A.; Pertwee, Roger G.; Heys, Steven D.

2010-01-01

The omega-3 fatty acid ethanolamides, docosahexaenoyl ethanolamide (DHEA) and eicosapentaenoyl ethanolamide (EPEA), displayed greater anti-proliferative potency than their parent omega-3 fatty acids, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), in LNCaP and PC3 prostate cancer cells. DHEA and EPEA activated cannabinoid CB1 and CB2 receptors in vitro with significant potency, suggesting that they are endocannabinoids. Both LNCaP and PC3 cells expressed CB1 and CB2 receptors, and the CB1- and CB2-selective antagonists, AM281 and AM630, administered separately or together, reduced the anti-proliferative potencies of EPEA and EPA but not of DHEA or DHA in PC3 cells and of EPA but not of EPEA, DHEA or DHA in LNCaP cells. Even so, EPEA and EPA may not have inhibited PC3 or LNCaP cell proliferation via cannabinoid receptors since the anti-proliferative potency of EPEA was well below the potency it displayed as a CB1 or CB2 receptor agonist. Indeed, these receptors may mediate a protective effect because the anti-proliferative potency of DHEA in LNCaP and PC3 cells was increased by separate or combined administration of AM281 and AM630. The anandamide-metabolizing enzyme, fatty acid amide hydrolase (FAAH), was highly expressed in LNCaP but not PC3 cells. Evidence was obtained that FAAH metabolizes EPEA and DHEA and that the anti-proliferative potencies of these ethanolamides in LNCaP cells can be enhanced by inhibiting this enzyme. Our findings suggest that the expression of cannabinoid receptors and of FAAH in some tumour cells could well influence the effectiveness of DHA and EPA or their ethanolamide derivatives as anticancer agents. PMID:20660502

Proliferative and morphologic changes in rat colon following bypass surgery.

PubMed

Barkla, D H; Tutton, P J

1985-06-01

In this study the proliferative and morphologic changes that occur in the colon of normal and dimethylhydrazine-treated rats following surgical bypass of the middle third of the colon are reported. Proliferative changes were measured by estimating accumulated mitotic indexes following vinblastine treatment and morphologic changes were observed with the use of light microscopy and scanning electron microscopy. Data were collected on Days 0, 7, 14, 30, and 72 after surgery. The results show that surgical bypass produces contrasting effects in the segments proximal to and distal to the suture line. In the proximal segment there was morphologic evidence of hyperplasia, although proliferative activity was unchanged except for an increase at 7 days in normal rats. In the distal segment there was a long-lived increase in the mitotic index, although morphologic changes were not seen. The results for DMH-treated rats were similar to those in normal rats. Groups of isolated dysplastic epithelial cells were often seen in the submucosa adjacent to sutures up to 72 days after surgery. Increased lymphoid infiltration was seen in segments proximal to but not distal to the suture line. It is hypothesized that the different responses of the proximal and distal segments may be related to the different embryologic origins of those segments. It is also hypothesized that the seeding of the submucosa with epithelial cells during suturing may be a factor in tumor recurrence.

Secretion of cytokines in breast cancer cells: the molecular mechanism of procathepsin D proliferative effects.

PubMed

Fusek, Martin; Vetvickova, Jana; Vetvicka, Vaclav

2007-03-01

Procathepsin D (pCD) is a major secreted protein in estrogen receptor-positive (ER+) breast cancer cell lines. Several independent studies have documented pronounced mitogenic effect of secreted pCD on cancer tissue-derived cell lines, including those from breast, lung, and prostate cancer. It has also been shown that the proliferative effect of pCD involves both autocrine and paracrine modes of action. Recent studies have suggested that pCD could act as a key paracrine communicator between cancer and stromal cells. We have shown earlier that the proliferative activity of pCD depends on the activation peptide sequence of pCD. The present study casts light on the mechanism by which pCD influences the proliferation of cancer cells expressing the ER. Results described in the current paper clearly show that pCD initiates secretion of cytokines interleukin-4 (IL-4), IL-8, IL-10, IL-13, macrophage inflammatory protein-1beta and (MIP-1beta) from such tumor cells. Secreted cytokines take part in the proliferation of the cancer cells, as proven by selective inhibition using antibodies. In addition, expression of cytokine receptors on tested cell lines corresponded to the effects of individual cytokines. An analogous pattern was also observed for fibroblasts, which, under physiologic conditions, are the cells in closest contact with the tumor tissue and play a role in tumor growth and invasion. Our observations were further supported by coculture experiments that are in agreement. Although very similar in response to addition of pCD, the invasive ER- cells do not secrete cytokines. Together with previous in vivo results, these data point to pCD as one of key molecules for therapeutic attack in breast cancer.

Olea europaea leaf extract improves the treatment response of GBM stem cells by modulating miRNA expression.

PubMed

Tezcan, Gulcin; Tunca, Berrin; Bekar, Ahmet; Budak, Ferah; Sahin, Saliha; Cecener, Gulsah; Egeli, Unal; Taskapılıoglu, Mevlut Ozgur; Kocaeli, Hasan; Tolunay, Sahsine; Malyer, Hulusi; Demir, Cevdet; Tumen, Gulendam

2014-01-01

The stem-like cells of Glioblastoma multiforme (GBM) tumors (GSCs) are one of the important determinants of recurrence and drug resistance. The aims of the current study were to evaluate the anticancer effect of Olea europaea leaf extract (OLE) on GBM cell lines, the association between OLE and TMZ responses, and the effect of OLE and the OLE-TMZ combination in GSCs and to clarify the molecular mechanism of this effect on the expression of miRNAs related to cell death. The anti-proliferative activity of OLE and the effect of the OLE-TMZ combination were tested in the T98G, U-138MG and U-87MG GBM cell lines using WST-1 assay. The mechanism of cell death was analyzed with Annexin V/FITC and TUNEL assays. The effects of OLE on the expression levels of miR-181b, miR-153, miR-145 and miR-137 and potential mRNA targets were analyzed in GSCs using RT-qPCR. OLE exhibited anti-proliferative effects via apoptosis and necrosis in the GBM cell lines. In addition, OLE significantly induced the expression of miR-153, miR-145, and miR-137 and decreased the expression of the target genes of these miRNAs in GSCs (p < 0.05). OLE causes cell death in GBM cells with different TMZ responses, and this effect is synergistically increased when the cells are treated with a combination of OLE and TMZ. This is the first study to indicate that OLE may interfere with the pluripotency of GSCs by modulating miRNA expression. Further studies are required, but we suggest that OLE may have a potential for advanced therapeutic cancer drug studies in GBM.

Participation of interleukins in synergistic effect of Bu-WSA on concanavalin A-induced DNA synthesis in mouse splenic lymphocytes.

PubMed

Nitta, T; Konno-Ejiri, H; Nemoto, K; Okumura, S; Ozawa, A; Nakano, M

1986-09-01

Butanol-extracted water-soluble adjuvant (Bu-WSA) obtained from Bacterionema matruchotii was mitogenic to murine splenic B lymphocytes, but not T lymphocytes. When murine splenic cells were cultured in the presence of Bu-WSA and concanavalin A (Con A) together, [3H]thymidine uptake of the culture cells synergically increased. The mechanism of the synergy of Con A and Bu-WSA and the participation of interleukin (IL) 1 and 2 in the synergy were studied. The proliferation cells in the synergy were Lyt-1+23- lymphocytes. Ia-positive accessory cells were required for the response. When separated cell populations and Marbrook-type culture vessels were used, a mixed cell population of T lymphocytes and B lymphocytes or macrophages (M phi) produced some active factor(s) after co-stimulation by Con A and Bu-WSA, and the factors enhanced DNA synthesis of another Con A-activated T lymphocyte population. Supernatants obtained from the spleen cell cultures or the mixed cell cultures with T lymphocytes and M phi in the presence of Con A and Bu-WSA contained greater amounts of IL-1 and IL-2 than those from cultures containing Con A or Bu-WSA alone. An addition of exogenous IL-1 or IL-2 to spleen cell cultures with Con A resulted in a proliferative response like that obtained through co-stimulation by Con A and Bu-WSA. These results suggest that the synergistic effect of Con A and Bu-WSA on the proliferative response in murine splenic cells is sustained by the enhancement of production of these T-lymphocyte growth factors.

Systematic tracking of altered haematopoiesis during sporozoite-mediated malaria development reveals multiple response points.

PubMed

Vainieri, Maria L; Blagborough, Andrew M; MacLean, Adam L; Haltalli, Myriam L R; Ruivo, Nicola; Fletcher, Helen A; Stumpf, Michael P H; Sinden, Robert E; Celso, Cristina Lo

2016-06-01

Haematopoiesis is the complex developmental process that maintains the turnover of all blood cell lineages. It critically depends on the correct functioning of rare, quiescent haematopoietic stem cells (HSCs) and more numerous, HSC-derived, highly proliferative and differentiating haematopoietic progenitor cells (HPCs). Infection is known to affect HSCs, with severe and chronic inflammatory stimuli leading to stem cell pool depletion, while acute, non-lethal infections exert transient and even potentiating effects. Both whether this paradigm applies to all infections and whether the HSC response is the dominant driver of the changes observed during stressed haematopoiesis remain open questions. We use a mouse model of malaria, based on natural, sporozoite-driven Plasmodium berghei infection, as an experimental platform to gain a global view of haematopoietic perturbations during infection progression. We observe coordinated responses by the most primitive HSCs and multiple HPCs, some starting before blood parasitaemia is detected. We show that, despite highly variable inter-host responses, primitive HSCs become highly proliferative, but mathematical modelling suggests that this alone is not sufficient to significantly impact the whole haematopoietic cascade. We observe that the dramatic expansion of Sca-1(+) progenitors results from combined proliferation of direct HSC progeny and phenotypic changes in downstream populations. We observe that the simultaneous perturbation of HSC/HPC population dynamics is coupled with early signs of anaemia onset. Our data uncover a complex relationship between Plasmodium and its host's haematopoiesis and raise the question whether the variable responses observed may affect the outcome of the infection itself and its long-term consequences on the host. © 2016 The Authors.

Systematic tracking of altered haematopoiesis during sporozoite-mediated malaria development reveals multiple response points

PubMed Central

Vainieri, Maria L.; Blagborough, Andrew M.; MacLean, Adam L.; Haltalli, Myriam L. R.; Ruivo, Nicola; Fletcher, Helen A.; Stumpf, Michael P. H.; Sinden, Robert E.; Lo Celso, Cristina

2016-01-01

Haematopoiesis is the complex developmental process that maintains the turnover of all blood cell lineages. It critically depends on the correct functioning of rare, quiescent haematopoietic stem cells (HSCs) and more numerous, HSC-derived, highly proliferative and differentiating haematopoietic progenitor cells (HPCs). Infection is known to affect HSCs, with severe and chronic inflammatory stimuli leading to stem cell pool depletion, while acute, non-lethal infections exert transient and even potentiating effects. Both whether this paradigm applies to all infections and whether the HSC response is the dominant driver of the changes observed during stressed haematopoiesis remain open questions. We use a mouse model of malaria, based on natural, sporozoite-driven Plasmodium berghei infection, as an experimental platform to gain a global view of haematopoietic perturbations during infection progression. We observe coordinated responses by the most primitive HSCs and multiple HPCs, some starting before blood parasitaemia is detected. We show that, despite highly variable inter-host responses, primitive HSCs become highly proliferative, but mathematical modelling suggests that this alone is not sufficient to significantly impact the whole haematopoietic cascade. We observe that the dramatic expansion of Sca-1+ progenitors results from combined proliferation of direct HSC progeny and phenotypic changes in downstream populations. We observe that the simultaneous perturbation of HSC/HPC population dynamics is coupled with early signs of anaemia onset. Our data uncover a complex relationship between Plasmodium and its host's haematopoiesis and raise the question whether the variable responses observed may affect the outcome of the infection itself and its long-term consequences on the host. PMID:27335321

Mouse neutrophils lacking lamin B receptor expression exhibit aberrant development and lack critical functional responses

PubMed Central

Gaines, Peter; Tien, Chiung W.; Olins, Ada L.; Olins, Donald E.; Shultz, Leonard D.; Carney, Lisa; Berliner, Nancy

2008-01-01

Objective The capacity of neutrophils to eradicate bacterial infections is dependent on normal development and the activation of functional responses, which include chemotaxis and the generation of oxygen radicals during the respiratory burst. A unique feature of the neutrophil is its highly lobulated nucleus, which is thought to facilitate chemotaxis but may also play a role in other critical neutrophil functions. Nuclear lobulation is dependent on the expression of the inner nuclear envelope protein, the lamin B receptor (LBR), mutations of which cause hypolobulated neutrophil nuclei in human Pelger-Huët anomaly (PHA) and the "ichthyosis" (ic) phenotype in mice. In this study we have investigated roles for LBR in mediating neutrophil development and the activation of multiple neutrophil functions, including chemotaxis and the respiratory burst. Materials and Methods A progenitor EML cell line was generated from an ic/ic mouse, and derived cells that lacked LBR expression were induced to mature neutrophils and then examined for abnormal morphology and functional responses. Results Neutrophils derived from EML-ic/ic cells exhibited nuclear hypolobulation identical to that observed in ichthyosis mice. The ic/ic neutrophils also displayed abnormal chemotaxis, supporting the notion that nuclear segmentation augments neutrophil extravasation. Furthermore, promyelocytic forms of ic/ic cells displayed decreased proliferative responses and produced a deficient respiratory burst upon terminal maturation. Conclusions Our studies of promyelocytes that lack LBR expression have identified roles for LBR in regulating not only the morphologic maturation of the neutrophil nucleus but also proliferative and functional responses that are critical to innate immunity. PMID:18550262

Imbalance of tumor necrosis factor receptors during progression in bovine leukemia virus infection

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

Konnai, Satoru; Usui, Tatsufumi; Ikeda, Manabu

2005-09-01

Previously, we found an up-regulation of tumor necrosis factor alpha (TNF)-{alpha} and an imbalance of TNF receptors in sheep experimentally infected with bovine leukemia virus (BLV). In order to investigate the different TNF-{alpha}-induced responses, in this study we examined the TNF-{alpha}-induced proliferative response and the expression levels of two distinct TNF receptors on peripheral blood mononuclear cells (PBMC) derived from BLV-uninfected cattle and BLV-infected cattle that were aleukemic (AL) or had persistent lymphocytosis (PL). The proliferative response of PBMC isolated from those cattle with PL in the presence of recombinant bovine TNF-{alpha} (rTNF-{alpha}) was significantly higher than those from ALmore » cattle and uninfected cattle and the cells from PL cattle expressed significantly higher mRNA levels of TNF receptor type II (TNF-RII) than those from AL and BLV-uninfected cattle. No difference was found in TNF-RI mRNA levels. Most cells expressing TNF-RII in PL cattle were CD5{sup +} or sIgM{sup +} cells and these cells showed resistance to TNF-{alpha}-induced apoptosis. Additionally, there were significant positive correlations between the changes in provirus load and TNF-RII mRNA levels, and TNF-{alpha}-induced proliferation and TNF-RII mRNA levels. These data suggest that imbalance in the expression of TNF receptors could at least in part contribute to the progression of lymphocytosis in BLV infection.« less

Transcriptome and proteome profiling of host responses to Marek's disease virus in chickens

USDA-ARS?s Scientific Manuscript database

Marek’s disease (MD) is an immunosuppressive and proliferative disease of domestic chickens caused by a highly oncogenic cell-associated alpha-herpesvirus, named Marek’s disease virus (MDV). Despite the availability of highly efficacious vaccines for control of MD and existence of lines of chickens ...

Pleural mesothelial cells promote expansion of IL-17-producing CD8+ T cells in tuberculous pleural effusion.

PubMed

Li, X; Zhou, Q; Yang, W B; Xiong, X Z; Du, R H; Zhang, J C

2013-05-01

IL-17-producing CD8(+) T lymphocytes (Tc17 cells) have recently been detected in many cancers and autoimmune diseases. However, the possible implication of Tc17 cells in tuberculous pleural effusion remains unclarified. In this study, distribution and phenotypic features of Tc17 cells in both tuberculous pleural effusion (TPE) and peripheral blood from patients with tuberculosis were determined. The effects of proinflammatory cytokines and local accessory cells (pleural mesothelial cells) on Tc17 cell expansion were also explored. We found that TPE contained more Tc17 cells than the blood. Compared with IFN-γ-producing CD8(+) T cells, Tc17 cells displayed higher expression of chemokine receptors (CCRs) and lower expression of cytotoxic molecules. In particularly, Tc17 cells in TPE exhibited high expression levels of CCR6, which could migrate in response to CCL20. Furthermore, IL-1β, IL-6, IL-23, or their various combinations could promote Tc17 cell expansion from CD8(+) T cells, whereas the proliferative response of Tc17 cells to above cytokines was lower than that of Th17 cells. Pleural mesothelial cells (PMCs) were able to stimulate Tc17 cell expansion via cell contact in an IL-1β/IL-6/IL-23 independent fashion. Thus this study demonstrates that Tc17 cells marks a subset of non-cytotoxic, CCR6(+) CD8(+) T lymphocytes with low proliferative capacity. The overrepresentation of Tc17 cells in TPE may be due to Tc17 cell expansion stimulated by pleural proinflammatory cytokines and to recruitment of Tc17 cells from peripheral blood. Additionally, PMCs may promote the production of IL-17 by CD8(+) T cells at sites of TPE via cell-cell interactions.

Tenascin-C in the extracellular matrix promotes the selection of highly proliferative and tubulogenesis-defective endothelial cells

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

Alves, Tercia Rodrigues; Universidade Federal do Rio de Janeiro; Carvalho da Fonseca, Anna Carolina

2011-09-10

The extracellular matrix (ECM) contains important cues for tissue homeostasis and morphogenesis. The matricellular protein tenascin-C (TN-C) is overexpressed in remodeling tissues and cancer. In the present work, we studied the effect of different ECM-which exhibited a significant diversity in their TN-C content-in endothelial survival, proliferation and tubulogenic differentiation: autologous (endothelial) ECM devoid of TN-C, but bearing large amounts of FN; fibroblast ECM, bearing both high TN-C and FN contents; and finally, glioma-derived matrices, usually poor in FN, but very rich in TN-C. HUVECs initially adhered to the immobilized matrix produced by U373 MG glioma cells, but significantly detached andmore » died by anoikis (50 to 80%) after 24 h, as compared with cells incubated with endothelial and fibroblast matrices. Surviving endothelial cells (20 to 50%) became up to 6-fold more proliferative and formed 74-97% less tube-like structures in vitro than cells grown on non-tumoral matrices. An antibody against the EGF-like repeats of tenascin-C (TN-C) partially rescued cells from the tubulogenic defect, indicating that this molecule is responsible for the selection of highly proliferative and tubulogenic defective endothelial cells. Interestingly, by using defined substrata, in conditions that mimic glioma and normal cell ECM composition, we observed that fibronectin (FN) modulates the TN-C-induced selection of endothelial cells. Our data show that TN-C is able to modulate endothelial branching morphogenesis in vitro and, since it is prevalent in matrices of injured and tumor tissues, also suggest a role for this protein in vascular morphogenesis, in these physiological contexts.« less

Decrease in calcitonin and parathyroid hormone mRNA levels and hormone secretion under long-term hypervitaminosis D3 in rats.

PubMed

Fernández-Santos, J M; Utrilla, J C; Conde, E; Hevia, A; Loda, M; Martín-Lacave, I

2001-04-01

In calcium homeostasis, vitamin D3 is a potent serum calcium-raising agent which in vivo regulates both calcitonin (CT) and parathyroid hormone (PTH) gene expression. Serum calcium is the major secretagogue for CT, a hormone product whose biosynthesis is the main biological activity of thyroid C-cells. Taking advantage of this regulatory mechanism, long-term vitamin D3-induced hypercalcemia has been extensively used as a model to produce hyperactivation, hyperplasia and even proliferative lesions of C-cells, supposedly to reduce the sustained high calcium serum concentrations. We have recently demonstrated that CT serum levels did not rise after long-term hypervitaminosis D3. Moreover, C-cells did not have a proliferative response, rather a decrease in CT-producing C-cell number was observed. In order to confirm the inhibitory effect of vitamin D3 on C-cells, Wistar rats were administered vitamin D3 chronically (25,000 IU/d) with or without calcium chloride (CaCl2). Under these long-term vitamin D3-hypercalcemic conditions, calcium, active metabolites of vitamin D3, CT and PTH serum concentrations were determined by RIA; CT and PTH mRNA levels were analysed by Northern blot and in situ hybridization; and, finally, the ultrastructure of calciotrophic hormone-producing cells was analysed by electron microscopy. Our results show, that, in rats, long term administration of vitamin D3 results in a decrease in hormone biosynthetic activities of both PTH and CT-producing cells, albeit at different magnitudes. Based upon these results, we conclude that hypervitaminosis D3-based methods do not stimulate C-cell activity and can not be used to induce proliferative lesions of calcitonin-producing cells.

Enhanced microglial pro-inflammatory response to lipopolysaccharide correlates with brain infiltration and blood-brain barrier dysregulation in a mouse model of telomere shortening.

PubMed

Raj, Divya D A; Moser, Jill; van der Pol, Susanne M A; van Os, Ronald P; Holtman, Inge R; Brouwer, Nieske; Oeseburg, Hisko; Schaafsma, Wandert; Wesseling, Evelyn M; den Dunnen, Wilfred; Biber, Knut P H; de Vries, Helga E; Eggen, Bart J L; Boddeke, Hendrikus W G M

2015-12-01

Microglia are a proliferative population of resident brain macrophages that under physiological conditions self-renew independent of hematopoiesis. Microglia are innate immune cells actively surveying the brain and are the earliest responders to injury. During aging, microglia elicit an enhanced innate immune response also referred to as 'priming'. To date, it remains unknown whether telomere shortening affects the proliferative capacity and induces priming of microglia. We addressed this issue using early (first-generation G1 mTerc(-/-) )- and late-generation (third-generation G3 and G4 mTerc(-/-) ) telomerase-deficient mice, which carry a homozygous deletion for the telomerase RNA component gene (mTerc). Late-generation mTerc(-/-) microglia show telomere shortening and decreased proliferation efficiency. Under physiological conditions, gene expression and functionality of G3 mTerc(-/-) microglia are comparable with microglia derived from G1 mTerc(-/-) mice despite changes in morphology. However, after intraperitoneal injection of bacterial lipopolysaccharide (LPS), G3 mTerc(-/-) microglia mice show an enhanced pro-inflammatory response. Nevertheless, this enhanced inflammatory response was not accompanied by an increased expression of genes known to be associated with age-associated microglia priming. The increased inflammatory response in microglia correlates closely with increased peripheral inflammation, a loss of blood-brain barrier integrity, and infiltration of immune cells in the brain parenchyma in this mouse model of telomere shortening. © 2015 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Effect of Neutralizing Transforming Growth Factor β1 on the Immune Response against Mycobacterium tuberculosis in Guinea Pigs

PubMed Central

Allen, Shannon Sedberry; Cassone, Lynne; Lasco, Todd M.; McMurray, David N.

2004-01-01

Transforming growth factor β (TGF-β) is a cytokine which has been shown to suppress the antimycobacterial immune responses of humans and experimental animals. In this study, the contributions of TGF-β to cytokine production in vivo were investigated by using the established guinea pig model of tuberculous pleurisy. Mycobacterium bovis BCG-vaccinated guinea pigs were injected intrapleurally with heat-killed virulent Mycobacterium tuberculosis. Eight days following induction of an antigen-specific pleural effusion, guinea pigs were injected intrapleurally with anti-TGF-β1 or isotype control antibody. The following day, pleural exudates were removed, and the fluid volume and characteristics of the infiltrating cells were determined. Pleural fluid was analyzed for total interferon (IFN) and tumor necrosis factor (TNF) protein levels by using appropriate bioassays. RNA from pleural effusion cells was examined to determine TGF-β1, TNF-α, IFN-γ, and interleukin-8 mRNA levels by using real-time PCR. Proliferative responses of pleural effusion lymphocytes were examined in response to concanavalin A and purified protein derivative (PPD) in vitro. Treatment with anti-TGF-β1 resulted in decreased pleural fluid volume and decreased cell numbers in the pleural space along with an increased percentage of lymphocytes and a decreased percentage of neutrophils. The bioactive TNF protein levels in pleural fluid were increased in guinea pigs treated with anti-TGF-β1, while the bioactive IFN protein concentrations were not altered. Expression of TGF-β1 and TNF-α mRNA was significantly increased following TGF-β1 neutralization. Finally, PPD-induced proliferative responses of pleural cells from anti-TGF-β1-treated animals were significantly enhanced. Thus, TGF-β1 may be involved in the resolution of this local, mycobacterial antigen-specific inflammatory response. PMID:14977939

Inhibition of p38/CREB phosphorylation and COX-2 expression by olive oil polyphenols underlies their anti-proliferative effects

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

Corona, Giulia; Dipartimento di Biologia Sperimentale, Sez. Patologia Sperimentale, Universita degli Studi di Cagliari, 09042 Monserrato; Deiana, Monica

2007-10-26

We investigated the anti-proliferative effects of an olive oil polyphenolic extract on human colon adenocarcinoma cells. Analysis indicated that the extract contained hydroxytyrosol, tyrosol and the various secoiridoid derivatives, including oleuropein. This extract exerted a strong inhibitory effect on cancer cell proliferation, which was linked to the induction of a G2/M phase cell cycle block. Following treatment with the extract (50 {mu}g/ml) the number of cells in the G2/M phase increased to 51.82 {+-} 2.69% relative to control cells (15.1 {+-} 2.5%). This G2/M block was mediated by the ability of olive oil polyphenols (50 {mu}g/ml) to exert rapid inhibitionmore » of p38 (38.7 {+-} 4.7%) and CREB (28.6 {+-} 5.5%) phosphorylation which led to a downstream reduction in COX-2 expression (56.9 {+-} 9.3%). Our data suggest that olive oil polyphenols may exert chemopreventative effects in the large intestine by interacting with signalling pathways responsible for colorectal cancer development.« less

Cellular responses to low-gravity: Pilot studies on suborbital rockets and orbiting spacecraft

NASA Technical Reports Server (NTRS)

Lewis, Marian L.

1993-01-01

The allocated funding supported, in part, experiments conducted on two Consort sounding rockets and five Shuttle flights. The primary parameters investigated were signal transduction in response to various mediators, cellular differentiation and metabolism in microgravity, and effect of microgravity on cytoskeletal morphology. Achievements include: demonstration of effect of spaceflight on the actin cytoskeleton in mouse osteoblasts and frog cells; confirmation that the T cell receptor-mediated signal transduction pathway in T lymphocytes is not affected by low-gravity compared to non-TCR-mediated stimulation (Con-A) which classically does not promote proliferative response; indication that microgravity may allow separation of proliferative signaling and secretory function in lymphocytes; demonstration that T lymphocytes and bone cells utilized less glucose indicating a shift in metabolism and confirming Spacelab results with WI-38 cells which used significantly less glucose, during spaceflight; confirmation that activation of human splenic B cells with a number of different mediators is not affected during spaceflight; demonstration of increased prostaglandin synthesis during reduced bone cell growth suggesting an effect of microgravity on prostaglandin-induced mitogenesis. The funding contributed significantly to the database described above and resulted in submission of six collaborative abstracts in 1993 (five to the ASGSB Annual Meeting and one to the ASCB Annual Meeting). Two abstracts were presented at the 1992 ASGSB Annual Meeting in Tucson. In addition, several peer reviewed papers are being generated and data will be included as background in preparation of future proposals, which hopefully will allow us to continue this type of extremely productive collaborative research.

A peroxisome proliferator-activated receptor ligand MCC-555 imparts anti-proliferative response in pancreatic cancer cells by PPARgamma-independent up-regulation of KLF4

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

Min, Kyung-Won; Zhang, Xiaobo; College of Animal Science and Technology, Northwest A and F University, Yangling, Shaanxi, 712100

2012-09-01

MCC-555 is a novel PPARα/γ dual ligand of the thiazolidinedione class and was recently developed as an anti-diabetic drug with unique properties. MCC-555 also has anti-proliferative activity through growth inhibition and apoptosis induction in several cancer cell types. Our group has shown that MCC-555 targets several proteins in colorectal tumorigenesis including nonsteroidal anti-inflammatory drug (NSAID)-activated gene (NAG-1) which plays an important role in chemoprevention responsible for chemopreventive compounds. NAG-1 is a member of the TGF-β superfamily and is involved in tumor progression and development; however, NAG-1's roles in pancreatic cancer have not been studied. In this report, we found thatmore » MCC-555 alters not only NAG-1 expression, but also p21 and cyclin D1 expression. NAG-1 and p21 expression was not blocked by PPARγ-specific antagonist GW9662, suggesting that MCC-555-induced NAG-1 and p21 expression is independent of PPARγ activation. However, decreasing cyclin D1 by MCC-555 seems to be affected by PPARγ activation. Further, we found that the GC box located in the NAG-1 promoter play an important role in NAG-1 transactivation by MCC-555. Subsequently, we screened several transcription factors that may bind to the GC box region in the NAG-1 promoter and found that KLF4 potentially binds to this region. Expression of KLF4 precedes NAG-1 and p21 expression in the presence of MCC-555, whereas blocking KLF4 expression using specific KLF4 siRNA showed that both NAG-1 and p21 expression by MCC-555 was blocked. In conclusion, MCC-555's actions on anti-proliferation involve both PPARγ-dependent and -independent pathways, thereby enhancing anti-tumorigenesis in pancreatic cancer cells. -- Highlights: ► PPARα/γ ligand MCC-555 exhibits anti-proliferative activity in pancreatic cancer cells. ► MCC-555 affects KLF4 expression following by NAG-1 and p21 expression in a PPARγ independent manner. ► MCC-555 also affects cyclin D1 down-regulation in a PPARγ dependent manner. ► To our knowledge, this is the first report that MCC-555 affects anti-proliferative activity in pancreatic cancer cells, along with multiple targets.« less

Nanoparticles for the delivery of zoledronic acid to prostate cancer cells: A comparative analysis through time lapse video-microscopy technique

PubMed Central

Schiraldi, Chiara; Zappavigna, Silvia; D' Agostino, Antonella; Porto, Stefania; Gaito, Ornella; Lusa, Sara; Lamberti, Monica; De Rosa, Mario; De Rosa, Giuseppe; Caraglia, Michele

2014-01-01

Time-lapse live cell imaging is a powerful tool for studying the responses of cells to drugs. Zoledronic acid (ZOL) is the most potent aminobiphosphonate able to induce cell growth inhibition at very low concentrations. The lack of clear evidence of ZOL-induced anti-cancer effects is likely due to its unfavorable pharmacokinetic profile. The use of nanotechnology-based formulations allows overcoming these limitations in ZOL pharmaco-distribution. Recently, stealth liposomes (LIPOs) and new self-assembly PEGylated nanoparticles (NPs) encapsulating ZOL were developed. Both the delivery systems showed promising anticancer activity in vitro and in vivo. In this work, we investigated the cytostatic effect of these novel formulations (LIPOs and NPs) compared with free ZOL on 2 different prostate cancer cell lines, PC 3 and DU 145 and on prostate epithelial primary cells EPN using time lapse video-microscopy (TLVM). In PC3 cells, free ZOL showed a significant anti-proliferative effect but this effect was lower than that induced by LIPOs and NPs encapsulating ZOL; moreover, LIPO-ZOL was more potent in inducing growth inhibition than NP-ZOL. On the other hand, LIPO-ZOL slightly enhanced the free ZOL activity on growth inhibition of DU 145, while the anti-proliferative effect of NP-ZOL was not statistically relevant. These novel formulations did not induce anti-proliferative effects on EPN cells. Finally, we evaluated cytotoxic effects on DU145 where, LIPO-ZOL induced the highest cytotoxicity compared with NP-ZOL and free ZOL. In conclusion, ZOL can be transformed in a powerful anticancer agent, if administered with nanotechnology-based formulations without damaging the healthy tissues. PMID:25482949

Immune response in asymptomatic smokers.

PubMed

Zeidel, A; Beilin, B; Yardeni, I; Mayburd, E; Smirnov, G; Bessler, H

2002-09-01

It has been demonstrated that cigarette smoking affects the immune system. Impairment of alveolar mononuclear cell function, described previously, may contribute to the higher rate of postoperative respiratory infections. However, increased susceptibility of smokers to infections of other origin (e.g. wound-related) implies that tobacco effect is not restricted to the respiratory immune competent cells. The present study was designed to investigate the systemic effect of tobacco smoking as it exerted on blood-derived immune cells. We measured systemic cytotoxic activity of natural killer cells, production of pro- and anti-inflammatory cytokines by blood mononuclear cells and their proliferation in response to mitogens. To minimize the immunosuppressive effect of other smoke-related factors, the smokers with chronic obstructive pulmonary disease (COPD) were excluded from this study. Peripheral blood mononuclear cells (PBMC) from 24 chronic asymptomatic smokers, and 28 controls, age and gender matched, were isolated and incubated in vitro with lipopolysaccharide (LPS) or phytohemagglutinin (PHA) to induce secretion of IL-1beta, IL-1ra, IL-6, IL-10, TNFalpha and IL-2, respectively, from mononuclear cells. The level of the cytokines in the supernatants was measured using ELISA kits. The proliferative response to the mitogens PHA and concanavalin A (ConA) was evaluated by 3H-thymidine incorporation and NK cell cytotoxicity by 51Cr release assay. Mononuclear cells from smokers showed increased production of the pro-inflammatory cytokines IL-1beta, IL-6 and TNFalpha and enhanced proliferative response to mitogens as compared to non-smoking population. The secretion of IL-2 and the anti-inflammatory cytokines IL-1ra and IL-10 was similar in both groups. NK cell cytotoxic activity was suppressed in the smokers. Cigarette smokers without chronic obstructive pulmonary disease (COPD) exhibit impaired NK cytotoxic activity in peripheral blood and unbalanced systemic production of pro- and anti-inflammatory cytokines. These changes may serve as predisposing factors for respiratory and systemic infections in the postoperative period and should alert an anesthetist during perioperative management.

Suppression of antitumour protective cytotoxic T lymphocyte responses to a human papillomavirus 16 E7 DNA vaccine by coinjection of interleukin-12 complementary DNA: involvement of nitric oxide in immune suppression

PubMed Central

Sin, Jeong-Im

2009-01-01

Interleukin-12 (IL-12) has been shown to enhance cellular immunity in vitro and in vivo. The beneficial roles of IL-12 as a DNA vaccine adjuvant have been commonly observed. Here the impact of IL-12 complementary DNA (cDNA) as an adjuvant for a human papillomavirus (HPV) type 16 E7 DNA vaccine is investigated in a mouse tumour model. Coinjection of E7 DNA vaccine with IL-12 cDNA completely suppressed antigen-specific cytotoxic T-lymphocyte (CTL) responses, leading to a complete loss of antitumour protection from a tumour cell challenge. In addition, antigen-specific antibody and T helper cell proliferative responses were also suppressed by IL-12 cDNA coinjection. This inhibition was observed over different IL-12 cDNA doses. Furthermore, separate leg injections of IL-12 and E7 cDNAs suppressed antigen-specific CTL and tumour protective responses, but not antibody and T helper cell proliferative responses, suggesting different pathways for suppression of these two separate responses. Further knockout animal studies demonstrated that interferon-γ and nitric oxide are not directly associated with suppression of antigen-specific antibody responses by IL-12 cDNA coinjection. However, nitric oxide was found to be involved in suppression of antigen-specific CTL and tumour protective responses by IL-12 cDNA coinjection. These data suggest that coinjection of IL-12 cDNA results in suppression of E7-specific CTL responses through nitric oxide, leading to a loss of antitumour resistance in this DNA vaccine model. This study further shows that the adjuvant effect of IL-12 is dependent on the antigen types tested. PMID:19740332

Suppression of antitumour protective cytotoxic T lymphocyte responses to a human papillomavirus 16 E7 DNA vaccine by coinjection of interleukin-12 complementary DNA: involvement of nitric oxide in immune suppression.

PubMed

Sin, Jeong-Im

2009-09-01

Interleukin-12 (IL-12) has been shown to enhance cellular immunity in vitro and in vivo. The beneficial roles of IL-12 as a DNA vaccine adjuvant have been commonly observed. Here the impact of IL-12 complementary DNA (cDNA) as an adjuvant for a human papillomavirus (HPV) type 16 E7 DNA vaccine is investigated in a mouse tumour model. Coinjection of E7 DNA vaccine with IL-12 cDNA completely suppressed antigen-specific cytotoxic T-lymphocyte (CTL) responses, leading to a complete loss of antitumour protection from a tumour cell challenge. In addition, antigen-specific antibody and T helper cell proliferative responses were also suppressed by IL-12 cDNA coinjection. This inhibition was observed over different IL-12 cDNA doses. Furthermore, separate leg injections of IL-12 and E7 cDNAs suppressed antigen-specific CTL and tumour protective responses, but not antibody and T helper cell proliferative responses, suggesting different pathways for suppression of these two separate responses. Further knockout animal studies demonstrated that interferon-gamma and nitric oxide are not directly associated with suppression of antigen-specific antibody responses by IL-12 cDNA coinjection. However, nitric oxide was found to be involved in suppression of antigen-specific CTL and tumour protective responses by IL-12 cDNA coinjection. These data suggest that coinjection of IL-12 cDNA results in suppression of E7-specific CTL responses through nitric oxide, leading to a loss of antitumour resistance in this DNA vaccine model. This study further shows that the adjuvant effect of IL-12 is dependent on the antigen types tested.

1-Benzyl-indole-3-carbinol is a novel indole-3-carbinol derivative with significantly enhanced potency of anti-proliferative and anti-estrogenic properties in human breast cancer cells

PubMed Central

Nguyen, Hanh H.; Lavrenov, Sergey N.; Sundar, Shyam N.; Nguyen, David H.H.; Tseng, Min; Marconett, Crystal N.; Kung, Jenny; Staub, Richard E.; Preobrazhenskaya, Maria N.; Bjeldanes, Leonard F.; Firestone, Gary L.

2012-01-01

Indole-3-carbinol (I3C), a natural autolysis product of a gluccosinolate present in Brassica vegetables such as broccoli and cabbage, has anti-proliferative and anti-estrogenic activities in human breast cancer cells. A new and significantly more potent I3C analogue, 1-benzyl-I3C was synthesized, and in comparison to I3C, this novel derivative displayed an approximate 1000-fold enhanced potency in suppressing the growth of both estrogen responsive (MCF-7) and estrogen independent (MDA-MB-231) human breast cancer cells (I3C IC50 of 52 μM, and 1-benzyl-I3C IC50 of 0.05 μM). At significantly lower concentrations, 1-benzyl-I3C induced a robust G1 cell cycle arrest and elicited the key I3C-specific effects on expression and activity of G1 acting cell cycle genes including the disruption of endogenous interactions of the Sp1 transcription factor with the CDK6 promoter. Furthermore, in estrogen responsive MCF-7 cells, with enhanced potency 1-benzyl-I3C down regulated production of estrogen receptor-alpha protein, acts with tamoxifen to arrest breast cancer cell growth more effectively than either compound alone, and inhibited the in vivo growth of human breast cancer cell-derived tumor xenografts in athymic mice. Our results implicate 1-benzyl-I3C as a novel, potent inhibitor of human breast cancer proliferation and estrogen responsiveness that could potentially be developed into a promising therapeutic agent for the treatment of indole-sensitive cancers. PMID:20570586

Generation of mesenchymal stromal cells in the presence of platelet lysate: a phenotypic and functional comparison of umbilical cord blood- and bone marrow-derived progenitors

PubMed Central

Avanzini, Maria Antonietta; Bernardo, Maria Ester; Cometa, Angela Maria; Perotti, Cesare; Zaffaroni, Nadia; Novara, Francesca; Visai, Livia; Moretta, Antonia; Del Fante, Claudia; Villa, Raffaella; Ball, Lynne M.; Fibbe, Willem E.; Maccario, Rita; Locatelli, Franco

2009-01-01

Background Mesenchymal stromal cells are employed in various different clinical settings in order to modulate immune response. However, relatively little is known about the mechanisms responsible for their immunomodulatory effects, which could be influenced by both the cell source and culture conditions. Design and Methods We tested the ability of a 5% platelet lysate-supplemented medium to support isolation and ex vivo expansion of mesenchymal stromal cells from full-term umbilical-cord blood. We also investigated the biological/functional properties of umbilical cord blood mesenchymal stromal cells, in comparison with platelet lysate-expanded bone marrow mesenchymal stromal cells. Results The success rate of isolation of mesenchymal stromal cells from umbilical cord blood was in the order of 20%. These cells exhibited typical morphology, immunophenotype and differentiation capacity. Although they have a low clonogenic efficiency, umbilical cord blood mesenchymal stromal cells may possess high proliferative potential. The genetic stability of these cells from umbilical cord blood was demonstrated by a normal molecular karyotype; in addition, these cells do not express hTERT and telomerase activity, do express p16ink4a protein and do not show anchorage-independent cell growth. Concerning alloantigen-specific immune responses, umbilical cord blood mesenchymal stromal cells were able to: (i) suppress T- and NK-lymphocyte proliferation, (ii) decrease cytotoxic activity and (iii) only slightly increase interleukin-10, while decreasing interferon-γ secretion, in mixed lymphocyte culture supernatants. While an indoleamine 2,3-dioxygenase-specific inhibitor did not reverse mesenchymal stromal cell-induced suppressive effects, a prostaglandin E2-specific inhibitor hampered the suppressive effect of both umbilical cord blood- and bone marrow-mesenchymal stromal cells on alloantigen-induced cytotoxic activity. Mesenchymal stromal cells from both sources expressed HLA-G. Conclusions Umbilical cord blood- and bone marrow-mesenchymal stromal cells may differ in terms of clonogenic efficiency, proliferative capacity and immunomodulatory properties; these differences may be relevant for clinical applications. PMID:19773264

Desquamation takes center stage at the origin of proliferative inflammatory atrophy, epithelial-mesenchymal transition, and stromal growth in benign prostate hyperplasia.

PubMed

Ferrucci, Danilo; Biancardi, Manoel F; Nishan, Umar; Rosa-Ribeiro, Rafaela; Carvalho, Hernandes F

2017-11-01

In this commentary, we propose a relationship between desquamation, initially described as the collective detachment and deletion of epithelial cell in the prostate gland after castration, and proliferative inflammatory atrophy (PIA) and stromal growth in benign prostate hyperplasia (BPH). First, in response to diverse stimuli, including inflammatory mediators, epithelial cells desquamate and leave a large surface of the luminal side of the basement membrane (BM) exposed. Basal cells are activated into intermediate-type cells, which change morphology to cover and remodel the exposed BM (simple atrophy) to a new physiological demand (such as in the hypoandrogen environment, simulated by surgical and/or chemical castration) and/or to support re-epithelialization (under normal androgen levels). In the presence of inflammation (that might be the cause of desquamation), the intermediate-type cells proliferate and characterize PIA. Second, in other circumstances, desquamation is an early step of epithelial-to-mesenchymal transition (EMT), which contributes to stromal growth, as suggested by some experimental models of BPH. The proposed associations correlate unexplored cell behaviors and reveal the remarkable plasticity of the prostate epithelium that might be at the origin of prostate diseases. © 2017 International Federation for Cell Biology.

Silicone-specific blood lymphocyte response in women with silicone breast implants.

PubMed Central

Ojo-Amaize, E A; Conte, V; Lin, H C; Brucker, R F; Agopian, M S; Peter, J B

1994-01-01

A blinded cross-sectional study was carried out with 99 women, 44 of whom had silicone breast implants. Group I consisted of 55 healthy volunteer women without breast implants; group II comprised 13 volunteer women with breast implants or explants who felt healthy; group III comprised 21 volunteer women with breast implants who had chronic fatigue, musculoskeletal symptoms, and skin disorders; and group IV comprised 10 women who had their prostheses explanted but still presented with clinical symptoms similar to those of the women in group III. Proliferative responses of peripheral blood mononuclear cells from all 99 women were measured by [3H]thymidine uptake after exposure to SiO2 silicon, or silicone gel. The levels of proliferative responses were expressed as stimulation indices, which were obtained by dividing the counts per minute of stimulated cells by the counts per minute of unstimulated cells. Abnormal responses to SiO2, silicon, or silicone gel were defined as a stimulation index of > 2.8, > 2.1, or > 2.4, respectively. Abnormal responses were observed in 0% of group I, 15% of group II, 29% of group III, and 30% of group IV (P < 0.0005 for group I versus groups II and IV). Thirty-one percent of symptomatic women with silicone gel breast implants had elevated serum silicon levels ( > 0.18 mg/liter); however, there was no significant correlation between abnormal cellular responses and silicon levels in blood serum, type of implant, time since first implantation, prosthesis explantation, number of implants, or report of implant leakage or rupture.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8556522

N-terminal tyrosine phosphorylation of caveolin-2 negates anti-proliferative effect of transforming growth factor beta in endothelial cells

PubMed Central

Abel, Britain; Willoughby, Cara; Jang, Sungchan; Cooper, Laura; Xie, Leike; Vo-Ransdell, Chi; Sowa, Grzegorz

2012-01-01

Here we show that tyrosine phosphorylation of caveolin-2 (Cav-2) negatively regulates the anti-proliferative function of transforming growth factor beta (TGF-beta) in endothelial cells. In contrast to wild-type-Cav-2, retroviral re-expression of Y19/27F-Cav-2 in Cav-2 knockout endothelial cells did not affect anti-proliferative effect of TGF-beta compared to empty vector. Conversely, although less effective than wild-type, re-expression of S23/36A-Cav-2 reduced the effect of TGF-beta compared to empty vector. This differential effect of tyrosine and serine phosphorylation mutants of Cav-2 correlated with TGF-beta-induced Smad3 phosphorylation and transcriptional activation of plasminogen activator inhibitor-1. Thus tyrosine-phosphorylated Cav-2 counteracts anti-proliferative effect of TGF-beta in endothelial cells. PMID:22819829

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

Gustafsson, Karin; Heffner, Garrett; Wenzel, Pamela L.

The widely expressed adaptor protein Shb has previously been reported to contribute to T cell function due to its association with the T cell receptor and furthermore, several of Shb's known interaction partners are established regulators of blood cell development and function. In addition, Shb deficient embryonic stem cells displayed reduced blood cell colony formation upon differentiation in vitro. The aim of the current study was therefore to explore hematopoietic stem and progenitor cell function in the Shb knockout mouse. Shb deficient bone marrow contained reduced relative numbers of long-term hematopoietic stem cells (LT-HSCs) that exhibited lower proliferation rates. Despitemore » this, Shb knockout LT-HSCs responded promptly by entering the cell cycle in response to genotoxic stress by 5-fluorouracil treatment. In competitive LT-HSC transplantations, Shb null cells initially engrafted as well as the wild-type cells but provided less myeloid expansion over time. Moreover, Shb knockout bone marrow cells exhibited elevated basal activities of focal adhesion kinase/Rac1/p21-activated kinase signaling and reduced responsiveness to Stem Cell Factor stimulation. Consequently, treatment with a focal adhesion kinase inhibitor increased Shb knockout LT-HSC proliferation. The altered signaling characteristics thus provide a plausible mechanistic explanation for the changes in LT-HSC proliferation since these signaling intermediates have all been shown to participate in LT-HSC cell cycle control. In summary, the loss of Shb dependent signaling in bone marrow cells, resulting in elevated focal adhesion kinase activity and reduced proliferative responses in LT-HSCs under steady state hematopoiesis, confers a disadvantage to the maintenance of LT-HSCs over time. -- Highlights: • Shb is an adaptor protein operating downstream of tyrosine kinase receptors. • Shb deficiency reduces hematopoietic stem cell proliferation. • The proliferative effect of Shb occurs via increased focal adhesion kinase activity. • Shb is critical for the long-term maintenance of the hematopoietic stem cell pool.« less

Impact of Helicobacter pylori on the healing process of the gastric barrier

PubMed Central

Mnich, Eliza; Kowalewicz-Kulbat, Magdalena; Sicińska, Paulina; Hinc, Krzysztof; Obuchowski, Michał; Gajewski, Adrian; Moran, Anthony P; Chmiela, Magdalena

2016-01-01

AIM To determine the impact of selected well defined Helicobacter pylori (H. pylori) antigens on gastric barrier cell turnover. METHODS In this study, using two cellular models of gastric epithelial cells and fibroblasts, we have focused on exploring the effects of well defined H. pylori soluble components such as glycine acid extract antigenic complex (GE), subunit A of urease (UreA), cytotoxin associated gene A protein (CagA) and lipopolysaccharide (LPS) on cell turnover by comparing the wound healing capacity of the cells in terms of their proliferative and metabolic activity as well as cell cycle distribution. Toxic effects of H. pylori components have been assessed in an association with damage to cell nuclei and inhibition of signal transducer and activator of transcription 3 (STAT3) phosphorylation. RESULTS We showed that H. pylori GE, CagA and UreA promoted regeneration of epithelial cells and fibroblasts, which is necessary for effective tissue healing. However, in vivo increased proliferative activity of these cells may constitute an increased risk of gastric neoplasia. In contrast, H. pylori LPS showed a dose-dependent influence on the process of wound healing. At a low concentration (1 ng/mL) H. pylori LPS accelerated of healing epithelial cells, which was linked to significantly enhanced cell proliferation and MTT reduction as well as lack of alterations in cell cycle and downregulation of epidermal growth factor (EGF) production as well as cell nuclei destruction. By comparison, H. pylori LPS at a high concentration (25 ng/mL) inhibited the process of wound repair, which was related to diminished proliferative activity of the cells, cell cycle arrest, destruction of cell nuclei and downregulation of the EGF/STAT3 signalling pathway. CONCLUSION In vivo H. pylori LPS driven effects might lead to the maintenance of chronic inflammatory response and pathological disorders on the level of the gastric mucosal barrier. PMID:27672275

Split T-cell tolerance in herpes simplex virus-infected mice and its implication for anti-viral immunity.

PubMed Central

Nash, A A; Ashford, N P

1982-01-01

Mice simultaneously injected intravenously and subcutaneously with herpes simplex virus fail to adoptively transfer delayed hypersensitivity (DH) to syngeneic recipients. The transferred lymph node cells also failed to rapidly eliminate infectious herpes from the pinna, despite the presence of cytotoxic T cells in the transferred suspension. Both primary and secondary cytotoxic cell responses in the draining lymph node were unaffected by the inhibition of DH. The lymph nodes from DH tolerized mice also contain lymphocytes capable of undergoing a proliferative response in vitro to herpes antigens. In addition, a neutralizing antibody response with IgG antibodies against herpes are also present in DH tolerized mice. These data suggest a form of split T-cell tolerance in which only DH responses are directly compromised. The implication of these findings for the pathogenesis of herpes simplex virus is discussed. PMID:6279490

Estradiol regulates the insulin-like growth factor-I (IGF-I) signalling pathway: A crucial role of phosphatidylinositol 3-kinase (PI 3-kinase) in estrogens requirement for growth of MCF-7 human breast carcinoma cells

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

Bernard, Laurence; Legay, Christine; Adriaenssens, Eric

2006-12-01

Estrogens can stimulate the proliferation of estrogen-responsive breast cancer cells by increasing their proliferative response to insulin-like growth factors. With a view to investigating the molecular mechanisms implicated, we studied the effect of estradiol on the expression of proteins implicated in the insulin-like growth factor signalling pathway. Estradiol dose- and time-dependently increased the expression of insulin receptor substrate-1 and the p85/p110 subunits of phosphatidylinositol 3-kinase but did not change those of ERK2 and Akt/PKB. ICI 182,780 did not inhibit estradiol-induced IRS-1 and p85 expression. Moreover, two distinct estradiol-BSA conjugate compounds were as effective as estradiol in inducing IRS-1 and p85/p110more » expression indicating the possible implication of an estradiol membrane receptor. Comparative analysis of steroids-depleted and steroids-treated cells showed that IGF-I only stimulates cell growth in the latter condition. Nevertheless, expression of a constitutively active form of PI 3-kinase in steroid-depleted cells triggers proliferation. These results demonstrate that estradiol positively regulates essential proteins of the IGF signalling pathway and put in evidence that phosphatidylinositol 3-kinase plays a central role in the synergistic pro-proliferative action of estradiol and IGF-I.« less

Aneurysmal bone cyst and other nonneoplastic conditions

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

Dahlin, D.C.; McLeod, R.A.

1982-08-01

Aneurysmal bone cyst is a benign proliferative tumefaction of bone. Histologic similarities indicate a kinship among classic aneurysmal bone cysts, essentially 'solid' proliferative lesions in bones; giant cell reparative granulomas of the jaws, at the base of the skull, and in the small bones of the hands and feet; skeletal lesions of hyperparathyroidism; and even pseudosarcomatous myositis ossificans, proliferative myositis, and proliferative fasciitis.

Granulocyte macrophage-colony stimulating factor and interleukin-3 increase expression of type II tumour necrosis factor receptor, increasing susceptibility to tumour necrosis factor-induced apoptosis. Control of leukaemia cell life/death switching.

PubMed

Rae, C; MacEwan, D J

2004-12-01

Tumour necrosis factor (TNF) induces apoptosis in a range of cell types via its two receptors, TNFR1 and TNFR2. Here, we demonstrate that proliferation and TNFR2 expression was increased in human leukaemic TF-1 cells by granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-3 (IL-3), with TNFR1 expression unaffected. Consequently, they switch from a proliferative to a TNF-induced apoptotic phenotype. Raised TNFR2 expression and susceptibility to TNF-induced apoptosis was not a general effect of proliferation as IL-1beta and IFN-gamma both proliferated TF-1 cells with no effect on TNFR expression or apoptosis. Although raised TNFR2 expression correlated with the apoptotic phenotype, stimulation of apoptosis in GM-CSF-pretreated cells was mediated by TNFR1, with stimulation of TNFR2 alone insufficient to initiate cell death. However, TNFR2 did play a role in apoptotic and proliferative responses as they were blocked by the presence of an antagonistic TNFR2 antibody. Additionally, coincubation with cycloheximide blocked the mitotic effects of GM-CSF or IL-3, allowing only the apoptotic responses of TNF to persist. TNF life/death was also observed in K562, but not MOLT-4 and HL-60 human leukaemic cell types. These findings show a cooperative role of TNFR2 in the TNF life/death switching phenomenon.

Analysis on pathogenesis of 50 cases of bladder proliferative lesions.

PubMed

Chen, Zhiqiang; Lan, Ruzhu; Ye, Zhangqun; Yang, Weimin

2003-01-01

In order to study the pathogenesis, clinical and pathological characteristics of proliferative lesions of the bladder, 50 cases of proliferative lesions of the bladder from 150 patients with complaints of frequency, urgency, hematuria and dysuria were subjected to cystoscopic biopsy of the suspicious foci in the bladder. In combination with the symptoms, urine and urodynamics, the relationship of proliferative lesions of the bladder to the inflammation and obstruction of the lower urinary tract was analyzed. Of the 50 cases of proliferative bladder lesions, 44 cases (88%) had lower urinary tract infection and 29 (58%) lower urinary tract obstruction. The patients with lower urinary tract obstruction were all complicated with infection. Three cases were associated with transitional cell carcinoma. Malignant cells were detected in 1 case by urinary cytologic examination. Proliferative lesions of the bladder, especially those without other obvious mucosa changes under cystoscopy, are common histological variants of urothelium in the patients with chronic inflammation and obstruction of the lower urinary tract. Chronic inflammation and obstruction of the lower urinary tract might be the causes for proliferative lesions of the bladder. It is suggested that different treatments should be applied according to the scope and histological type of the proliferative lesions.

Cytological Study of Breast Carcinoma Before and After Oncotherapy with Special Reference to Morphometry and Proliferative Activity.

PubMed

Koley, Sananda; Chakrabarti, Srabani; Pathak, Swapan; Manna, Asim Kumar; Basu, Siddhartha

2015-12-01

Our study was done to assess the cytological changes due to oncotherapy in breast carcinoma especially on morphometry and proliferative activity. Cytological aspirates were collected from a total of 32 cases of invasive ductal carcinoma both before and after oncotherapy. Morphometry was done on the stained cytological smears to assess the different morphological parameters of cell dimension by using the ocular morphometer and the software AutoCAD 2007. Staining was done with Ki-67 and proliferating cell nuclear antigen (PCNA) as proliferative markers. Different morphological parameters were compared before and after oncotherapy by unpaired Student's t test. Statistically significant differences were found in morphometric parameters, e.g., mean nuclear diameter, mean nuclear area, mean cell diameter, and mean cell area, and in the expression of proliferative markers (Ki-67 and PCNA). Statistical analysis was done by obtaining p values. There are statistically significant differences between morphological parameter of breast carcinoma cells before and after oncotherapy.

Heme oxygenase is not involved in the anti-proliferative effects of statins on pancreatic cancer cells.

PubMed

Vanova, K; Boukalova, S; Gbelcova, H; Muchova, L; Neuzil, J; Gurlich, R; Ruml, T; Vitek, L

2016-05-12

Pancreatic cancer is recognized as one of the most fatal tumors due to its aggressiveness and resistance to therapy. Statins were previously shown to inhibit the proliferation of cancer cells via various signaling pathways. In healthy tissues, statins activate the heme oxygenase pathway, nevertheless the role of heme oxygenase in pancreatic cancer is still controversial. The aim of this study was to evaluate, whether anti-proliferative effects of statins in pancreatic cancer cells are mediated via the heme oxygenase pathway. In vitro effects of various statins and hemin, a heme oxygenase inducer, on cell proliferation were evaluated in PA-TU-8902, MiaPaCa-2 and BxPC-3 human pancreatic cancer cell lines. The effect of statins on heme oxygenase activity was assessed and heme oxygenase-silenced cells were used for pancreatic cancer cell proliferation studies. Cell death rate and reactive oxygen species production were measured in PA-TU-8902 cells, followed by evaluation of the effect of cerivastatin on GFP-K-Ras trafficking and expression of markers of invasiveness, osteopontin (SPP1) and SOX2. While simvastatin and cerivastatin displayed major anti-proliferative properties in all cell lines tested, pravastatin did not affect the cell growth at all. Strong anti-proliferative effect was observed also for hemin. Co-treatment of cerivastatin and hemin increased anti-proliferative potential of these agents, via increased production of reactive oxygen species and cell death compared to individual treatment. Heme oxygenase silencing did not prevent pancreatic cancer cells from the tumor-suppressive effect of cerivastatin or hemin. Cerivastatin, but not pravastatin, protected Ras protein from trafficking to the cell membrane and significantly reduced expressions of SPP1 (p < 0.05) and SOX2 (p < 0.01). Anti-proliferative effects of statins and hemin on human pancreatic cancer cell lines do not seem to be related to the heme oxygenase pathway. While hemin triggers reactive oxygen species-induced cell death, cerivastatin targets Ras protein trafficking and affects markers of invasiveness.

Aerobic physical training does not condition against strenuous exercise-induced changes in immune function but modulates T cell proliferative responses.

PubMed

Patiño, Pablo J; Caraballo, Domingo I; Szewczyk, Katarzyna; Quintana, Juan C; Bedoya, Lady R; Ramírez, Beatriz E; Jaramillo, Andrés

2017-09-29

Exercise-induced stress induces considerable changes in the immune system. To better understand the mechanisms related to these immune changes during acute and chronic physical stress, we studied the effects of aerobic physical training (APT) on several parameters of the immune system. Previously untrained males (18-25 years of age) were divided into a group that was subjected to 6 months of APT (n=10) and a sedentary control group (n=7). The subjects performed a cardiopulmonary exercise test (CET) at 0, 3, and 6 months of the APT program. B cell (CD19+), T cell (CD4+ and CD8+), and natural killer cell (CD56+) levels, and mitogen-induced T cell proliferation and cytokine production (interleukin-1, interleukin-4, interleukin-12, and interferon-) were evaluated before and at 30 seconds and 24 hours after the CET. There was a significant increase in CD4+ T cells and natural killer cells and a significant reduction in T cell proliferation in both groups 30 seconds after the CET at 3 and 6 months of the APT program. Of note, the trained group showed significantly lower resting T cell proliferation (before and 24 hour after the CET) than the sedentary control group at 3 and 6 months of the APT program. There were no significant differences in cytokine production after the CET between both groups at any time point of the APT program. These data show that APT does not condition against strenuous exercise induced immune changes but significantly modulates T cell proliferative responses.

Human keratinocytes are efficiently immortalized by a Rho kinase inhibitor

PubMed Central

Chapman, Sandra; Liu, Xuefeng; Meyers, Craig; Schlegel, Richard; McBride, Alison A.

2010-01-01

Primary human keratinocytes are useful for studying the pathogenesis of many different diseases of the cutaneous and mucosal epithelia. In addition, they can form organotypic tissue equivalents in culture that can be used as epidermal autografts for wound repair as well as for the delivery of gene therapy. However, primary keratinocytes have a finite lifespan in culture that limits their proliferative capacity and clinical use. Here, we report that treatment of primary keratinocytes (originating from 3 different anatomical sites) with Y-27632, a Rho kinase inhibitor, greatly increased their proliferative capacity and resulted in efficient immortalization without detectable cell crisis. More importantly, the immortalized cells displayed characteristics typical of primary keratinocytes; they had a normal karyotype and an intact DNA damage response and were able to differentiate into a stratified epithelium. This is the first example to our knowledge of a defined chemical compound mediating efficient cell immortalization, and this finding could have wide-ranging and profound investigational and medical applications. PMID:20516646

Activation of double-stranded RNA-dependent protein kinase inhibits proliferation of pancreatic β-cells

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

Chen, Shan-Shan; Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing; Jiang, Teng

2014-01-17

Highlights: •PKR can be activated by glucolipitoxicity and pro-inflammatory cytokines in β-cells. •Activated PKR inhibited β-cell proliferation by arresting cell cycle at G1 phase. •Activated PKR fully abrogated the pro-proliferative effects of IGF-I on β-cells. -- Abstract: Double-stranded RNA-dependent protein kinase (PKR) is revealed to participate in the development of insulin resistance in peripheral tissues in type 2 diabetes (T2DM). Meanwhile, PKR is also characterized as a critical regulator of cell proliferation. To date, no study has focused on the impact of PKR on the proliferation of pancreatic β-cells. Here, we adopted insulinoma cell lines and mice islet β-cells tomore » investigate: (1) the effects of glucolipotoxicity and pro-inflammatory cytokines on PKR activation; (2) the effects of PKR on proliferation of pancreatic β-cells and its underlying mechanisms; (3) the actions of PKR on pro-proliferative effects of IGF-I and its underlying pathway. Our results provided the first evidence that PKR can be activated by glucolipitoxicity and pro-inflammatory cytokines in pancreatic β-cells, and activated PKR significantly inhibited cell proliferation by arresting cell cycle at G1 phase. Reductions in cyclin D1 and D2 as well as increases in p27 and p53 were associated with the anti-proliferative effects of PKR, and proteasome-dependent degradation took part in the reduction of cyclin D1 and D2. Besides, PKR activation abrogated the pro-proliferative effects of IGF-I by activating JNK and disrupting IRS1/PI3K/Akt signaling pathway. These findings indicate that the anti-proliferative actions of PKR on pancreatic β-cells may contribute to the pathogenesis of T2DM.« less

Pankiller effect of prolonged exposure to menadione on glioma cells: potentiation by vitamin C.

PubMed

Vita, Marina F; Nagachar, Nivedita; Avramidis, Dimitrios; Delwar, Zahid M; Cruz, Mabel H; Siden, Åke; Paulsson, Kajsa M; Yakisich, Juan Sebastian

2011-12-01

Menadione (Vitamin K3) has anti-tumoral effects against a wide range of cancer cells. Its potential toxicity to normal cells and narrow therapeutic range limit its use as single agent but in combination with radiation or other anti-neoplastic agents can be of therapeutic use. In this paper, we first evaluated the early (within 3 h) effect of menadione on ongoing DNA replication. In normal rat cerebral cortex mini-units menadione showed an age dependent anti-proliferative effect. In tissue mini-units prepared from newborn rats, menadione inhibited ongoing DNA replication with an IC (50) of approximately 10 μM but 50 μM had no effect on mini-units from prepared adult rat tissue. The effect of short (72 h) and prolonged exposure (1-2 weeks) to menadione alone in the DBTRG.05MG human glioma cells line and in combination with vitamin C was studied. After short period of exposure data show that menadione alone or in combination with vitamin C provided similar concentration-response curves (and IC(50) values). Prolonged exposure to these drugs was evaluated by their ability to kill 100% of glioma cells and prevent regrowth when cells are re-incubated in drug-free media. In this long-term assay, menadione:vitamin C at a ratio 1:100 showed higher anti-proliferative activity when compared to each drug alone and allowed to reduce each drug concentration between 2.5 to 5-fold. Similar anti-proliferative effect was demonstrated in 8 patient derived glioblastoma cell cultures. Our data should be able to encourage further advanced studies on animal models to evaluate the potential use of this combination therapy for glioma treatment.

Cellular immune responses to HPV-18, -31, and -53 in healthy volunteers immunized with recombinant HPV-16 L1 virus-like particles

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

Pinto, Ligia A.; Viscidi, Raphael; Harro, Clayton D.

Human papillomavirus-like particles (HPV VLP) are candidate vaccines that have shown to be efficacious in reducing infection and inducing robust antiviral immunity. Neutralizing antibodies generated by vaccination are largely type-specific, but little is known about the type-specificity of cellular immune responses to VLP vaccination. To determine whether vaccination with HPV-16 L1VLP induces cellular immunity to heterologous HPV types (HPV-18, HPV-31, and HPV-53), we examined proliferative and cytokine responses in vaccine (n = 11) and placebo (n = 5) recipients. Increased proliferative and cytokine responses to heterologous types were observed postvaccination in some individuals. The proportion of women responding to heterologousmore » types postvaccination (36%-55%) was lower than that observed in response to HPV-16 (73%). Response to HPV-16 VLP predicted response to other types. The strongest correlations in response were observed between HPV-16 and HPV-31, consistent with their phylogenetic relatedness. In summary, PBMC from HPV-16 VLP vaccine recipients can respond to L1VLP from heterologous HPV types, suggesting the presence of conserved T cell epitopes.« less

Anti-proliferative activities on HeLa cancer cell line of Thai medicinal plant recipes selected from MANOSROI II database.

PubMed

Manosroi, Jiradej; Boonpisuttinant, Korawinwich; Manosroi, Worapaka; Manosroi, Aranya

2012-07-13

The Thai/Lanna medicinal plant recipe database "MANOSROI II" contained the medicinal plant recipes of all regions in Thailand for the treatment of various diseases including anti-cancer medicinal plant recipes. To investigate anti-proliferative activity on HeLa cell lines of medicinal plant recipes selected from the Thai/Lanna medicinal plant recipe database "MANOSROI II". The forty aqueous extracts of Thai/Lanna medicinal plant recipes selected from the Thai/Lanna medicinal plant recipe database "MANOSROI II" were investigated for anti-proliferative activity on HeLa cell line by SRB assay. The apoptosis induction by caspase-3 activity and MMP-2 inhibition activity by zymography on HeLa cell line of the three selected aqueous extracts, which gave the highest anti-proliferative activity were determined. Phytochemicals and anti-oxidative activities including free radical scavenging activity, inhibition of lipid peroxidation and metal chelating inhibition activities were also investigated. Sixty percentages of the medicinal plant recipes selected from "MANOSROI II" database showed anti-proliferative activity on HeLa cell line. The recipes of N031(Albizia chinensis (Osbeck) Merr, Cassia fistula L., and Dargea volubilis Benth.ex Hook. etc.), N039 (Nymphoides indica L., Peltophorum pterocarpum (DC.), and Polyalthia debilis Finet et Gagnep etc.) and N040 (Nymphoides indica L. Kuntze, Sida rhombifolia L., and Xylinbaria minutiflora Pierre. etc.) gave higher anti-proliferative activity than the standard anti-cancer drug, cisplatin of 1.25, 1.29 and 30.18 times, respectively. The positive relationship between the anti-proliferative activity and the MMP-2 inhibition activity and metal chelating inhibition activity was observed, but no relationship between the anti-proliferative activity and apoptosis induction, free radical scavenging activity and lipid peroxidation inhibition activity. Phytochemicals found in these extracts were alkaloids, flavonoids, tannins and xanthones, but not anthraquinones and carotenoids. The recipe N040 exhibited the highest anti-proliferative and MMP-2 inhibition on HeLa cancer cell line at 30 and threefolds of cisplatin, respectively (p<0.05), while recipe N031 gave the highest caspase-3 activity (1.29-folds over the control) (p<0.05). This study has demonstrated that recipe N040 selected from MANOSROI II database appeared to be a good candidate with high potential for the further development as an anti-cancer agent. Crown Copyright © 2012. Published by Elsevier Ireland Ltd. All rights reserved.

Effect of age on immune parameters and the immune response of dogs to vaccines: a cross-sectional study.

PubMed

HogenEsch, Harm; Thompson, Steven; Dunham, Anisa; Ceddia, Michael; Hayek, Michael

2004-01-01

The evaluation of anti-aging intervention strategies in dogs would benefit from reliable quantitative biomarkers of aging. In the present study, the expression of various immune parameters was measured in young and old dogs to identify potential biomarkers of aging. The second goal of the study was to determine the effect of age on the immune response to vaccines. The immune function, including the antibody response to vaccines, was determined in 32 young adult (3.15+/-0.8 years of age) and 33 old dogs (12.1+/-1.3 years of age) of various breeds. Old dogs had a significantly lower lymphocyte proliferative response and a lower percentage of CD4+ T cells and CD45R+/CD4+ T cells, and a higher percentage of CD8+ T cells and a higher concentration of serum and salivary IgA. The most significant differences (P<0.001) occurred in the lymphocyte proliferative responses to ConA and PHA, the CD4:CD8 ratio, and the percentage of CD45R+/CD4+ T cells suggesting that these parameters are potential biomarkers of aging. There was no difference in the percentage of total T and B lymphocytes and the concentration of serum IgM and IgG. Both groups of dogs had protective titers against distemper virus, parvovirus and rabies virus before annual revaccination. The pre-vaccination titer against rabies virus was higher in the old dogs than in the young dogs, and there were no differences in post-vaccination titers against any of the viruses. This suggests that annual vaccination protocols provide adequate protection for old dogs.

Fc receptors for mouse IgG1 on human monocytes: polymorphism and role in antibody-induced T cell proliferation.

PubMed

Tax, W J; Hermes, F F; Willems, R W; Capel, P J; Koene, R A

1984-09-01

In previous studies, it was shown that there is polymorphism in the mitogenic effect of mouse IgG1 monoclonal antibodies against the T3 antigen of human T cells. This polymorphism implies that IgG1 anti-T3 antibodies are not mitogenic for T cells from 30% of healthy individuals. The present results demonstrate that this polymorphism is caused by polymorphism of an Fc receptor for mouse IgG1, present on human monocytes. The Fc receptor for murine IgG1 could be detected by a newly developed rosetting assay on monocytes from all individuals responsive to the mitogenic effect of IgG1 anti-T3 antibodies. This Fc receptor was not detectable on monocytes from those individuals exhibiting no mitogenic responses to IgG1 anti-T3 monoclonal antibodies. Cross-linking of T3 antigens appears to be essential for antibody-induced mitosis of T cells, because mononuclear cells that did not proliferate in response to WT 31 (an IgG1 antibody against T3 antigen) showed a proliferative response to Sepharose beads coated with WT 31. The Fc receptor--if functionally present--may be involved in the cross-linking of T3 antigens through anti-T3 antibodies. Further evidence for the involvement of this Fc receptor in antibody-induced T cell proliferation was provided by inhibition studies. Immune complexes containing IgG1 antibodies were able to inhibit the proliferative response to IgG1 anti-T3 antibodies. This inhibition by immune complexes appears to be mediated through the monocyte Fc receptor for mouse IgG1. These findings are important for the interpretation of previously described inhibitory effects of anti-T cell monoclonal antibodies on T cell proliferation, and show that such inhibitory effects may be monocyte-mediated (via immune complexes) rather than caused by a direct involvement of the respective T cell antigens in T cell mitosis. The Fc receptor for mouse IgG1 plays a role in antibody-induced T cell proliferation. Its polymorphism may have important implications for the therapeutic use of IgG1 monoclonal antibodies.

Restoration of the anti-proliferative and anti-migratory effects of 1,25-dihydroxyvitamin D by silibinin in vitamin D-resistant colon cancer cells.

PubMed

Bhatia, Vandanajay; Falzon, Miriam

2015-07-01

Colorectal carcinoma (CRC) is the third most common cancer in developed countries. A large fraction of cases are linked to chronic intestinal inflammation, with concomitant increased TNF-α release and elevated Snail1/Snail2 levels. These transcription factors in turn suppress vitamin D receptor (VDR) expression, resulting in loss of responsiveness to the protective anti-proliferative and anti-migratory effects of 1,25-dihydroxyvitamin D (1,25D). Experimental and epidemiologic evidence support the use of natural products to target CRC. Here we show that the flavonolignan silibinin reverses the TNF-α-induced upregulation of Snail1 and Snail2 in the 1,25D-resistant human colon carcinoma cells HT-29. These silibinin effects are accompanied by an increase in VDR levels; Snail1 overexpression reverses these silibinin effects. Silibinin also restores promoter activity from a vitamin D-response element (VDRE) reporter construct. While 1,25D had no significant effect on HT-29 and SW480-R cell proliferation and migration, co-treatment with silibinin restored 1,25D responsiveness. In addition, co-treatment with silibinin plus 1,25D decreased proliferation and migration at doses where silibinin alone had no effect. These findings demonstrate that this combination may present a novel approach to target CRC in conditions of chronic colonic inflammation. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Restoration of the anti-proliferative and anti-migratory effects of 1,25-dihydroxyvitamin D by silibinin in vitamin D-resistant colon cancer cells

PubMed Central

Bhatia, Vandanajay; Falzon, Miriam

2015-01-01

Colorectal carcinoma (CRC) is the third most common cancer in developed countries. A large fraction of cases are linked to chronic intestinal inflammation, with concomitant increased TNF-α release and elevated Snail1/Snail2 levels. These transcription factors in turn suppress vitamin D receptor (VDR) expression, resulting in loss of responsiveness to the protective anti-proliferative and anti-migratory effects of 1,25-dihydroxyvitamin D (1,25D). Experimental and epidemiologic evidence support the use of natural products to target CRC. Here we show that the flavonolignan silibinin reverses the TNF-α-induced upregulation of Snail1 and Snail2 in the 1,25D-resistant human colon carcinoma cells HT-29. These silibinin effects are accompanied by an increase in VDR levels; Snail1 overexpression reverses these silibinin effects. Silibinin also restores promoter activity from a vitamin D-response element (VDRE) reporter construct. While 1,25D had no significant effect on HT-29 and SW480-R cell proliferation and migration, co-treatment with silibinin restored 1,25D responsiveness. In addition, co-treatment with silibinin plus 1,25D decreased proliferation and migration at doses where silibinin alone had no effect. These findings demonstrate that this combination may present a novel approach to target CRC in conditions of chronic colonic inflammation. PMID:25846868

Effects of peptides on proliferative activity of retinal and pigmented epithelial cells.

PubMed

Khavinson, V Kh; Zemchikhina, V N; Trofimova, S V; Malinin, V V

2003-06-01

We studied the effects of Retinalamin (polypeptide preparation isolated from the retina) and a synthetic peptide Epithalon (Ala-Glu-Asp-Gly) on proliferative activity of retinal and pigmented epithelial cells. Experiments showed that Retinalamin and Epithalon (in certain concentrations) tissue-specifically stimulated proliferation of retinal and pigmented epithelial cell in culture.

Microvascular Remodeling and Wound Healing: A Role for Pericytes

PubMed Central

Dulmovits, Brian M.; Herman, Ira M.

2012-01-01

Physiologic wound healing is highly dependent on the coordinated functions of vascular and non-vascular cells. Resolution of tissue injury involves coagulation, inflammation, formation of granulation tissue, remodeling and scarring. Angiogenesis, the growth of microvessels the size of capillaries, is crucial for these processes, delivering blood-borne cells, nutrients and oxygen to actively remodeling areas. Central to angiogenic induction and regulation is microvascular remodeling, which is dependent upon capillary endothelial cell and pericyte interactions. Despite our growing knowledge of pericyte-endothelial cell crosstalk, it is unclear how the interplay among pericytes, inflammatory cells, glia and connective tissue elements shape microvascular injury response. Here, we consider the relationships that pericytes form with the cellular effectors of healing in normal and diabetic environments, including repair following injury and vascular complications of diabetes, such as diabetic macular edema and proliferative diabetic retinopathy. In addition, pericytes and stem cells possessing “pericyte-like” characteristics are gaining considerable attention in experimental and clinical efforts aimed at promoting healing or eradicating ocular vascular proliferative disorders. As the origin, identification and characterization of microvascular pericyte progenitor populations remains somewhat ambiguous, the molecular markers, structural and functional characteristics of pericytes will be briefly reviewed. PMID:22750474

Oral testosterone in male rats and the development of experimental autoimmune encephalomyelitis.

PubMed

Macció, Daniela R; Calfa, Gastón; Roth, German A

2005-01-01

Considering that sex steroids can influence the immune system, we studied the development of experimental autoimmune encephalomyelitis (EAE), a T-cell-mediated autoimmune disease of the central nervous system, and the concomitant cell-mediated immunity in gonadally intact and gonadectomized male Wistar rats given testosterone supplementation. Sham-operated rats and surgically castrated animals were orally self-administered with vehicle or testosterone added in the water bottle for 20 days before EAE induction. The androgenic effect of oral testosterone self-administration was evidenced by changes in body weight, and in the weights of androgen-dependent testes and seminal vesicles. Testosterone administration reduced the incidence of clinical signs of EAE in sham-operated animals and reversed the clinical symptoms of the disease associated with castrated EAE animals. The clinical signs observed in the different groups correlated with changes in delayed-type hypersensitivity and mononuclear cell-proliferative responses to the encephalitogenic myelin basic protein. Moreover, testosterone but not cholesterol supplementation in vitro suppressed the proliferative response of mononuclear cells to myelin basic protein suggesting that testosterone may affect specific immune functions through direct actions on immune cells. Finally, self-administration of testosterone induced also elevated corticosterone levels that in sham-operated rats correlated with the low incidence of the disease and in gonadectomized animals could be involved in the remission of clinical symptoms of EAE. These results suggest that orally self-administered testosterone can modulate specific cellular immune responses and serum corticosterone levels leading to changes in the development of EAE. Copyright 2005 S. Karger AG, Basel.

Identification of a subset of human natural killer cells expressing high levels of programmed death 1: A phenotypic and functional characterization.

PubMed

Pesce, Silvia; Greppi, Marco; Tabellini, Giovanna; Rampinelli, Fabio; Parolini, Silvia; Olive, Daniel; Moretta, Lorenzo; Moretta, Alessandro; Marcenaro, Emanuela

2017-01-01

Programmed death 1 (PD-1) is an immunologic checkpoint that limits immune responses by delivering potent inhibitory signals to T cells on interaction with specific ligands expressed on tumor/virus-infected cells, thus contributing to immune escape mechanisms. Therapeutic PD-1 blockade has been shown to mediate tumor eradication with impressive clinical results. Little is known about the expression/function of PD-1 on human natural killer (NK) cells. We sought to clarify whether human NK cells can express PD-1 and analyze their phenotypic/functional features. We performed multiparametric cytofluorimetric analysis of PD-1 + NK cells and their functional characterization using degranulation, cytokine production, and proliferation assays. We provide unequivocal evidence that PD-1 is highly expressed (PD-1 bright ) on an NK cell subset detectable in the peripheral blood of approximately one fourth of healthy subjects. These donors are always serologically positive for human cytomegalovirus. PD-1 is expressed by CD56 dim but not CD56 bright NK cells and is confined to fully mature NK cells characterized by the NKG2A - KIR + CD57 + phenotype. Proportions of PD-1 bright NK cells were higher in the ascites of a cohort of patients with ovarian carcinoma, suggesting their possible induction/expansion in tumor environments. Functional analysis revealed a reduced proliferative capability in response to cytokines, low degranulation, and impaired cytokine production on interaction with tumor targets. We have identified and characterized a novel subpopulation of human NK cells expressing high levels of PD-1. These cells have the phenotypic characteristics of fully mature NK cells and are increased in patients with ovarian carcinoma. They display low proliferative responses and impaired antitumor activity that can be partially restored by antibody-mediated disruption of PD-1/programmed death ligand interaction. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Heat Resistant Characteristics of Major Royal Jelly Protein 1 (MRJP1) Oligomer

PubMed Central

Moriyama, Takanori; Ito, Aimi; Omote, Sumire; Miura, Yuri; Tsumoto, Hiroki

2015-01-01

Soluble royal jelly protein is a candidate factor responsible for mammiferous cell proliferation. Major royal jelly protein 1 (MRJP1), which consists of oligomeric and monomeric forms, is an abundant proliferative protein in royal jelly. We previously reported that MRJP1 oligomer has biochemical heat resistance. Therefore, in the present study, we investigated the effects of several heat treatments (56, 65 and 96°C) on the proliferative activity of MRJP1 oligomer. Heat resistance studies showed that the oligomer molecular forms were slightly maintained until 56℃, but the molecular forms were converted to macromolecular heat-aggregated MRJP1 oligomer at 65℃ and 96℃. But, the growth activity of MRJP1 oligomer treated with 96°C was slightly attenuated when compared to unheated MRJP1 oligomer. On the other hand, the cell proliferation activity was preserved until 96℃ by the cell culture analysis of Jurkat cells. In contrast, those of IEC-6 cells were not preserved even at 56°C. The present observations suggest that the bioactive heat-resistance properties were different by the origin of the cells. The cell proliferation analysis showed that MRJP1 oligomer, but not MRJP2 and MRJP3, significantly increased cell numbers, suggesting that MRJP1 oligomer is the predominant proliferation factor for mammiferous cells. PMID:26020775

Heat Resistant Characteristics of Major Royal Jelly Protein 1 (MRJP1) Oligomer.

PubMed

Moriyama, Takanori; Ito, Aimi; Omote, Sumire; Miura, Yuri; Tsumoto, Hiroki

2015-01-01

Soluble royal jelly protein is a candidate factor responsible for mammiferous cell proliferation. Major royal jelly protein 1 (MRJP1), which consists of oligomeric and monomeric forms, is an abundant proliferative protein in royal jelly. We previously reported that MRJP1 oligomer has biochemical heat resistance. Therefore, in the present study, we investigated the effects of several heat treatments (56, 65 and 96°C) on the proliferative activity of MRJP1 oligomer. Heat resistance studies showed that the oligomer molecular forms were slightly maintained until 56℃, but the molecular forms were converted to macromolecular heat-aggregated MRJP1 oligomer at 65℃ and 96℃. But, the growth activity of MRJP1 oligomer treated with 96°C was slightly attenuated when compared to unheated MRJP1 oligomer. On the other hand, the cell proliferation activity was preserved until 96℃ by the cell culture analysis of Jurkat cells. In contrast, those of IEC-6 cells were not preserved even at 56°C. The present observations suggest that the bioactive heat-resistance properties were different by the origin of the cells. The cell proliferation analysis showed that MRJP1 oligomer, but not MRJP2 and MRJP3, significantly increased cell numbers, suggesting that MRJP1 oligomer is the predominant proliferation factor for mammiferous cells.

Nuclear calcium is required for human T cell activation

PubMed Central

Samstag, Yvonne

2016-01-01

Calcium signals in stimulated T cells are generally considered single entities that merely trigger immune responses, whereas costimulatory events specify the type of reaction. Here we show that the “T cell calcium signal” is a composite signal harboring two distinct components that antagonistically control genomic programs underlying the immune response. Using human T cells from healthy individuals, we establish nuclear calcium as a key signal in human T cell adaptogenomics that drives T cell activation and is required for signaling to cyclic adenosine monophosphate response element–binding protein and the induction of CD25, CD69, interleukin-2, and γ-interferon. In the absence of nuclear calcium signaling, cytosolic calcium activating nuclear factor of activated T cells translocation directed the genomic response toward enhanced expression of genes that negatively modulate T cell activation and are associated with a hyporesponsive state. Thus, nuclear calcium controls the T cell fate decision between a proliferative immune response and tolerance. Modulators of nuclear calcium–driven transcription may be used to develop a new type of pro-tolerance immunosuppressive therapy. PMID:27810914

In vitro maturation of immature thymocytes into immunocompetent T cells in the absence of direct thymic influence.

PubMed

Irlé, C; Piguet, P F; Vassalli, P

1978-07-01

Peanut lectin (PNL) binds to a majority of mouse thymocytes (Thc) in suspension. By using cell affinity chromatography on a column of anti-PNL antibody, Thc populations at least 96 percent pure in PNL + or - cells, as judged by immunofluorescence, were obtained. PNL(+) cells are rich in Thy 1 and poor in H(2) antigens, cortisone sensitive, unresponsive to phytohemagglutinin (PHA), and immunologically incompetent, as judged by mixed lymphocyte reaction, popliteal lymph node graft-versus-host assay, and by testing helper activity in a primary in vitro antibody response to sheep erythrocytes; the converse is true of PNL(-) cells. Thus, PNL(+) and (-) cells appear to correspond to cortical and medullary Thc, respectively, as previously suggested. In culture, PNL(+) Thc show poor viability and a weak proliferative response to concanavalin A (Con A), except when supernate (SUP) of 24 h Con A stimulated lymph node lymphocyte cultures, or irradiated lymph node cells, are added, in which cases a strong proliferative response to the mitogen is observed. A variety of control experiments showed that the proliferating cells did not result from preferential stimulation of a few contaminating PNL(-) Thc present in the PNL(+) Thc cultures. The blasts resulting from PNL(+) Thc proliferation display mitogen-induced cytotoxicity, and give rise to a population of medium-sized lymphocytes, mostly PNL(-), poor in Thy 1 and rich in H(2) antigens, PHA responsive, and immunologically competent in the above-mentioned assays. Fresh PNL(+) Thc responded in mixed lymphocyte reaction in the presence of SUP (lectin depleted) and since incubation in SUP alone did not confer reactivity on PNL(+) Thc, it appears therefore that (a) immature Thc possess alloantigen and mitogen-specific surface receptors but lack the capacity to respond by proliferation to receptor triggering without the help of extracellular factor(s) released by mature lymphoid cells stimulated by mitogens (b) cell division is associated with the acquisition of immunological responsiveness, characteristic of mature T lymphocytes. The implications of these findings for the ontogenesis of thymus-derived lymphocytes, and for the possible traffic of Thc within and from the thymus, are discussed.

Concurrent targeting of EP1/EP4 receptors and COX-2 induces synergistic apoptosis in KSHV and EBV associated non-Hodgkin lymphoma cell lines

PubMed Central

Paul, Arun George; Chandran, Bala; Sharma-Walia, Neelam

2014-01-01

The effective anti-tumorigenic potential of non-steroidal anti-inflammatory drugs (NSAIDs) and eicosonoid (EP; EP1–4) receptor antagonists prompted us to test their efficacy in Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) related lymphomas. Our study demonstrated that (1) EP1–4 receptor protein levels vary among the various non-Hodgkin’s lymphoma (NHL) cell lines tested (BCBL-1:KSHV+/EBV−;BC-3: KSHV+/EBV−; Akata/EBV+: KSHV−/EBV+; and JSC-1 cells: KSHV+/EBV+ cells); (2) 5.0 µM of EP1 antagonist (SC-51322) had a significant anti-proliferative effect on BCBL-1, BC-3, Akata/EBV+, and JSC-1 cells; (3) 50.0 µM of EP2 antagonist (AH6809) was required to induce a significant anti-proliferative effect on BCBL-1, Akata/EBV+, and JSC-1 cells; (4) 5.0 µM of EP4 antagonist (GW 627368X) had a significant anti-proliferative effect on BC-3, Akata/EBV+, and JSC-1 cells; (5) COX-2 selective inhibitor celecoxib (5.0µM) had significant anti-proliferative effects on BCBL-1, BC-3, Akata/EBV+, and JSC-1 cells; and (6) a combination of 1.0µM each of celecoxib, SC-51322 and GW 627368X could potentiate the pro-apoptotic properties of celecoxib or vice-versa. Overall, our studies identified the synergistic anti-proliferative effect of NSAIDs and EP receptor blockers on KSHV and EBV related B cell malignancies. PMID:23523954

Segregation of human peripheral blood lymphocytes according to their affinity for insolubilized histamine. Principal differences between males and females.

PubMed Central

Tartakovsky, B; Segal, S; Shani, A; Hellerstein, S; Weinstein, Y; Bentwich, Z

1979-01-01

An attempt was made to investigate the possible existence of differences in the composition of peripheral blood lymphocytes between males and females. Using affinity chromatography of human peripheral mononuclear cells on insolubilized histamine together with staining by fluoresceinated histamine-rabbit serum albumin (HRSA) we revealed that males possess a significantly higher proportion of mononuclear cells which bind to HRSA. These results are also reflected in sex-related differences in proliferative responses of the HRSA-non-adherent mononuclear cell population to T cell-dependent mitogens antigens and allogeneic mononuclear cells. PMID:160849

Immunization of Aged Pigs with Attenuated Pseudorabies Virus Vaccine Combined with CpG Oligodeoxynucleotide Restores Defective Th1 Immune Responses

PubMed Central

Chu, Pinpin; Ma, Miaopeng; Shi, Juqing; Cai, Haiming; Huang, Chaoyuan; Li, Huazhou; Jiang, Zhenggu; Wang, Houguang; Wang, Weifang; Zhang, Shuiqing; Zhang, Linghua

2013-01-01

Background and Aims Attempts to immunize aged subjects often result in the failure to elicit a protective immune response. Murine model studies have shown that oligonucleotides containing CpG motifs (CpG ODN) can stimulate immune system in aged mice as effectively as in young mice. Since many physiological and pathophysiological data of pigs can be transferred to humans, research in pigs is important to confirm murine data. Here we investigated whether immunization of aged pig model with attenuated pseudorabies virus vaccine (PRV vaccine) formulated with CpG ODN could promote a successful development of immune responses that were comparable to those induced in young pigs in a similar manner. Methodology Young and aged pigs were immunized IM with PRV vaccine alone, or in combination with CpG ODN respectively. At days 3, 7, 14 post immunization sera were assayed by ELISA for IgG titres, at day 7 for IgG1 and IgG2 subtypes titres. All blood samples collected in evacuated test tubes with K-EDTA at day 7 were analyzed for flow cytometer assay. Blood samples at day 7 collected in evacuated test tubes with heparin were analysed for antigen-specific cytokines production and peripheral blood mononuclear cells (PBMCs) proliferative responses. Results CpG ODN could enhance Th1 responses (PRV-specific IgG2/IgG1 ratio, proliferative responses, Th1 cytokines production) when used as an adjuvant for the vaccination of aged pigs, which were correlated with enhanced CD4+ T cells percentage, decreased CD4+CD8+CD45RO+ T cells percentage and improved PRV-specific CD4+ T cells activation. Conclusions Our results demonstrate a utility for CpG ODN, as a safe vaccine adjuvant for promoting effective systemic immune responses in aged pig model. This agent could have important clinical uses in overcoming some of age-associated depressions in immune function that occur in response to vaccination. PMID:23785433

Deconvoluting Post-Transplant Immunity: Cell Subset-Specific Mapping Reveals Pathways for Activation and Expansion of Memory T, Monocytes and B Cells

PubMed Central

Grigoryev, Yevgeniy A.; Kurian, Sunil M.; Avnur, Zafi; Borie, Dominic; Deng, Jun; Campbell, Daniel; Sung, Joanna; Nikolcheva, Tania; Quinn, Anthony; Schulman, Howard; Peng, Stanford L.; Schaffer, Randolph; Fisher, Jonathan; Mondala, Tony; Head, Steven; Flechner, Stuart M.; Kantor, Aaron B.; Marsh, Christopher; Salomon, Daniel R.

2010-01-01

A major challenge for the field of transplantation is the lack of understanding of genomic and molecular drivers of early post-transplant immunity. The early immune response creates a complex milieu that determines the course of ensuing immune events and the ultimate outcome of the transplant. The objective of the current study was to mechanistically deconvolute the early immune response by purifying and profiling the constituent cell subsets of the peripheral blood. We employed genome-wide profiling of whole blood and purified CD4, CD8, B cells and monocytes in tandem with high-throughput laser-scanning cytometry in 10 kidney transplants sampled serially pre-transplant, 1, 2, 4, 8 and 12 weeks. Cytometry confirmed early cell subset depletion by antibody induction and immunosuppression. Multiple markers revealed the activation and proliferative expansion of CD45RO+CD62L− effector memory CD4/CD8 T cells as well as progressive activation of monocytes and B cells. Next, we mechanistically deconvoluted early post-transplant immunity by serial monitoring of whole blood using DNA microarrays. Parallel analysis of cell subset-specific gene expression revealed a unique spectrum of time-dependent changes and functional pathways. Gene expression profiling results were validated with 157 different probesets matching all 65 antigens detected by cytometry. Thus, serial blood cell monitoring reflects the profound changes in blood cell composition and immune activation early post-transplant. Each cell subset reveals distinct pathways and functional programs. These changes illuminate a complex, early phase of immunity and inflammation that includes activation and proliferative expansion of the memory effector and regulatory cells that may determine the phenotype and outcome of the kidney transplant. PMID:20976225

Deconvoluting post-transplant immunity: cell subset-specific mapping reveals pathways for activation and expansion of memory T, monocytes and B cells.

PubMed

Grigoryev, Yevgeniy A; Kurian, Sunil M; Avnur, Zafi; Borie, Dominic; Deng, Jun; Campbell, Daniel; Sung, Joanna; Nikolcheva, Tania; Quinn, Anthony; Schulman, Howard; Peng, Stanford L; Schaffer, Randolph; Fisher, Jonathan; Mondala, Tony; Head, Steven; Flechner, Stuart M; Kantor, Aaron B; Marsh, Christopher; Salomon, Daniel R

2010-10-14

A major challenge for the field of transplantation is the lack of understanding of genomic and molecular drivers of early post-transplant immunity. The early immune response creates a complex milieu that determines the course of ensuing immune events and the ultimate outcome of the transplant. The objective of the current study was to mechanistically deconvolute the early immune response by purifying and profiling the constituent cell subsets of the peripheral blood. We employed genome-wide profiling of whole blood and purified CD4, CD8, B cells and monocytes in tandem with high-throughput laser-scanning cytometry in 10 kidney transplants sampled serially pre-transplant, 1, 2, 4, 8 and 12 weeks. Cytometry confirmed early cell subset depletion by antibody induction and immunosuppression. Multiple markers revealed the activation and proliferative expansion of CD45RO(+)CD62L(-) effector memory CD4/CD8 T cells as well as progressive activation of monocytes and B cells. Next, we mechanistically deconvoluted early post-transplant immunity by serial monitoring of whole blood using DNA microarrays. Parallel analysis of cell subset-specific gene expression revealed a unique spectrum of time-dependent changes and functional pathways. Gene expression profiling results were validated with 157 different probesets matching all 65 antigens detected by cytometry. Thus, serial blood cell monitoring reflects the profound changes in blood cell composition and immune activation early post-transplant. Each cell subset reveals distinct pathways and functional programs. These changes illuminate a complex, early phase of immunity and inflammation that includes activation and proliferative expansion of the memory effector and regulatory cells that may determine the phenotype and outcome of the kidney transplant.

In vitro stimulation of rabbit T lymphocytes by cells expressing herpes simplex antigens.

PubMed

Kapoor, A K; Ling, N R; Nash, A A; Bachan, A; Wildy, P

1982-04-01

Lymphocyte stimulation responses to herpes antigens were studied using virus-infected X-irradiated cells. Rabbits were immunized with herpes simplex virus type 1 (strain HFEM) grown in RK 13 cells. For in vitro stimulation assay BHK21 cells were X-irradiated (15 000 rad) and infected with a high m.o.i. of a temperature-sensitive (ts) mutant (N102) of HFEM strain at the non-permissive temperature (38.5 degrees C) of virus. Virus antigens were expressed on the infected cells and there was no leakage of infectious virus into the medium at 38.5 degrees C. T lymphocytes from rabbits immunized with herpes simplex virus were specifically activated by herpesvirus-infected X-irradiated cells; lymph node cells from rabbits immunized with RK13 cells and from non-immune rabbits showed no proliferative response.

Protection against murine potomac horse fever by an inactivated Ehrlichia risticii vaccine.

PubMed

Rikihisa, Y

1991-05-01

Ehrlichia risticii propagated in a murine macrophage cell line were freed from the host cell by hypotonic lysis of the infected cells. The cell-free ehrlichiae were inactivated with beta-propiolactone and combined or not combined with polymyxin-B. The vaccines were administered to mice with Quil-A (saponin) as an adjuvant twice at 2 to 3 week intervals and the mice were challenged with live E. risticii 2 to 3 weeks after the last vaccination. With or without the addition of polymyxin-B, the vaccine preparations protected mice from developing clinical signs and gross pathologic changes such as thymic atrophy, splenomegaly, and increase in whole intestinal weight. Mice vaccinated with or without polymyxin-B developed high titer IgG antibody against E. risticii before and after the challenge with live E. risticii. Spleen lymphocyte proliferative response assay at 11 days post challenge revealed that with polymyxin-B a higher lymphocyte proliferation occurred as compared with that of the mice which received polymyxin-B-free vaccine. Spleen lymphocytes of the placebo (polymyxin-B and Quil-A) pretreated/challenged mice showed no proliferative activity. Western blot analysis revealed that vaccinated mice reacted mainly with 110, 57 and 33 kDa antigen bands before and after challenge. The placebo (polymyxin-B and Quil-A)/challenged mice showed a very weak response to ehrlichial antigens at day 10 to 11 post challenge. Comparison with inactivated Renografin-purified E. risticii or 0.25% SDS-insoluble fraction of E. risticii with the inactivated host cell-free vaccine revealed no increased protection. These results indicate that inactivated host cell-free E. risticii can protect mice from murine Potomac horse fever. The presence of polymyxin-B appeared to be not harmful but rather beneficial for lymphocyte proliferation response upon challenge with live E. risticii.

Involvement of Semaphorin (Sema4D) in T-Dependent Activation of B Cells.

PubMed

Kuklina, Е М; Nekrasova, I V; Valieva, Yu V

2017-08-01

The involvement of endogenous semaphorin (Sema4D) into the key stage of T-dependent differentiation of B cells, formation of plasmoblasts, was demonstrated in vitro in T/B cell co-culture under conditions of polyclonal activation of T cells. The effect of semaphorin was not associated with activation of high-affinity Sema4D receptor plexin B1, but involves lowaffinity receptor CD72. These data indicate that Sema4D-dependent signal regulates not only the initial stage of B-cell activation, proliferative response to the antigen, but also further differentiation of B cells into plasma cells.

Dysregulation of B Cell Activity During Proliferative Kidney Disease in Rainbow Trout.

PubMed

Abos, Beatriz; Estensoro, Itziar; Perdiguero, Pedro; Faber, Marc; Hu, Yehfang; Díaz Rosales, Patricia; Granja, Aitor G; Secombes, Christopher J; Holland, Jason W; Tafalla, Carolina

2018-01-01

Proliferative kidney disease (PKD) is a widespread disease caused by the endoparasite Tetracapsuloides bryosalmonae (Myxozoa: Malacosporea). Clinical disease, provoked by the proliferation of extrasporogonic parasite stages, is characterized by a chronic kidney pathology with underlying transcriptional changes indicative of altered B cell responses and dysregulated T-helper cell-like activities. Despite the relevance of PKD to European and North American salmonid aquaculture, no studies, to date, have focused on further characterizing the B cell response during the course of this disease. Thus, in this work, we have studied the behavior of diverse B cell populations in rainbow trout ( Oncorhynchus mykiss ) naturally infected with T. bryosalmonae at different stages of preclinical and clinical disease. Our results show a clear upregulation of all trout immunoglobulins (Igs) (IgM, IgD, and IgT) demonstrated by immunohistochemistry and Western blot analysis, suggesting the alteration of diverse B cell populations that coexist in the infected kidney. Substantial changes in IgM, IgD, and IgT repertoires were also identified throughout the course of the disease further pointing to the involvement of the three Igs in PKD through what appear to be independently regulated mechanisms. Thus, our results provide strong evidence of the involvement of IgD in the humoral response to a specific pathogen for the first time in teleosts. Nevertheless, it was IgT, a fish-specific Ig isotype thought to be specialized in mucosal immunity, which seemed to play a prevailing role in the kidney response to T. bryosalmonae . We found that IgT was the main Ig coating extrasporogonic parasite stages, IgT + B cells were the main B cell subset that proliferated in the kidney with increasing kidney pathology, and IgT was the Ig for which more significant changes in repertoire were detected. Hence, although our results demonstrate a profound dysregulation of different B cell subsets during PKD, they point to a major involvement of IgT in the immune response to the parasite. These results provide further insights into the pathology of PKD that may facilitate the future development of control strategies.

Lack of immunotoxicity of saquinavir (Ro 31-8959) used alone or in double or triple combination with AZT and ddC.

PubMed

Viora, M; Di Genova, G; Quaranta, M G; Boirivant, M; Camponeschi, B

1998-09-01

Saquinavir (Ro 31-8959; SQV) has been demonstrated to be a potent inhibitor of human immunodeficiency virus (HIV) proteinases and acts synergistically with dideoxynucleoside analogues. The aim of this study was to investigate the in vitro immunomodulatory effects of SQV on normal human peripheral blood mononuclear cells (PBMC) and on lamina propria mononuclear cells (LPMC). We used the drug either alone or in double and triple combination with AZT and ddC to assess whether SQV enhances the immunomodulatory effects induced by AZT and ddC that we previously observed. We demonstrated that SQV did not induce any modulation of the proliferative response either in PBMC or in LPMC. Similarly, NK cell-mediated cytotoxic activity and cytokine production were not modified by SQV. More importantly, SQV/AZT, SQV/ddC, and SQV/AZT/ddC combinations did not strengthen neither the inhibition of PBMC and LPMC proliferative response or the modulation of cytokine production induced by AZT, ddC, and AZT/ddC. On the other hand, the increased IL-2 production induced by AZT and ddC was not observed adding SQV to the dideoxynucleoside analogues. In conclusion, we demonstrated that SQV used in combination with AZT and ddC did not add any further immunotoxicity.

Selective Roles of Normal and Mutant Huntingtin in Neural Induction and Early Neurogenesis

PubMed Central

Nguyen, Giang D.; Gokhan, Solen; Molero, Aldrin E.; Mehler, Mark F.

2013-01-01

Huntington's disease (HD) is a neurodegenerative disorder caused by abnormal polyglutamine expansion in the amino-terminal end of the huntingtin protein (Htt) and characterized by progressive striatal and cortical pathology. Previous reports have shown that Htt is essential for embryogenesis, and a recent study by our group revealed that the pathogenic form of Htt (mHtt) causes impairments in multiple stages of striatal development. In this study, we have examined whether HD-associated striatal developmental deficits are reflective of earlier maturational alterations occurring at the time of neurulation by assessing differential roles of Htt and mHtt during neural induction and early neurogenesis using an in vitro mouse embryonic stem cell (ESC) clonal assay system. We demonstrated that the loss of Htt in ESCs (KO ESCs) severely disrupts the specification of primitive and definitive neural stem cells (pNSCs, dNSCs, respectively) during the process of neural induction. In addition, clonally derived KO pNSCs and dNSCs displayed impaired proliferative potential, enhanced cell death and altered multi-lineage potential. Conversely, as observed in HD knock-in ESCs (Q111 ESCs), mHtt enhanced the number and size of pNSC clones, which exhibited enhanced proliferative potential and precocious neuronal differentiation. The transition from Q111 pNSCs to fibroblast growth factor 2 (FGF2)-responsive dNSCs was marked by potentiation in the number of dNSCs and altered proliferative potential. The multi-lineage potential of Q111 dNSCs was also enhanced with precocious neurogenesis and oligodendrocyte progenitor elaboration. The generation of Q111 epidermal growth factor (EGF)-responsive dNSCs was also compromised, whereas their multi-lineage potential was unaltered. These abnormalities in neural induction were associated with differential alterations in the expression profiles of Notch, Hes1 and Hes5. These cumulative observations indicate that Htt is required for multiple stages of neural induction, whereas mHtt enhances this process and promotes precocious neurogenesis and oligodendrocyte progenitor cell elaboration. PMID:23691206

Increased telomere length and proliferative potential in peripheral blood mononuclear cells of adults of different ages stimulated with concanavalin A

PubMed Central

2013-01-01

Background Recently, a direct correlation with telomere length, proliferative potential and telomerase activity has been found in the process of aging in peripheral blood cells. The objective of the study was to evaluate telomere length and proliferative potential in peripheral blood mononuclear cells (PBMCs) after stimulation with Concanavalin A (ConA) of young adults compared with older adults. Methods Blood samples were obtained from 20 healthy young males (20–25 years old) (group Y) and 20 males (60–65 years old) (group O). We compared PBMC proliferation before and after stimulation with ConA. DNA was isolated from cells separated before and after culture with ConA for telomeric measurement by real-time polymerase chain reaction. Results In vitro stimulation of PBMCs from young subjects induced an increase of telomere length as well as a higher replicative capacity of cell proliferation. Samples from older adults showed higher loss of telomeric DNA (p = 0.03) and higher levels of senescent (≤6.2 kb) telomeric DNA (p = 0.02) and displayed a marked decrease of proliferation capacity. Viability cell counts and CFSE tracking in 72-h-old cell cultures indicated that group O PBMCs (CD8+ and CD4+ T cells) underwent fewer mitotic cycles and had shorter telomeres than group Y (p = 0.04). Conclusions Our findings confirm that telomere length in older-age adults is shorter than in younger subjects. After stimulation with ConA, cells are not restored to the previous telomere length and undergo replicative senescence. This is in sharp contrast to the response observed in young adults after ConA stimulation where cells increase in telomere length and replicative capacity. The mechanisms involved in this phenomenon are not yet clear and merit further investigation. PMID:24063536

Local and Systemic CD4+ T Cell Exhaustion Reverses with Clinical Resolution of Pulmonary Sarcoidosis

PubMed Central

Hawkins, Charlene; Shaginurova, Guzel; Shelton, D. Auriel; Herazo-Maya, Jose D.; Oswald-Richter, Kyra A.; Young, Anjuli; Celada, Lindsay J.; Kaminski, Naftali; Sevin, Carla

2017-01-01

Investigation of the Th1 immune response in sarcoidosis CD4+ T cells has revealed reduced proliferative capacity and cytokine expression upon TCR stimulation. In other disease models, such cellular dysfunction has been associated with a step-wise, progressive loss of T cell function that results from chronic antigenic stimulation. T cell exhaustion is defined by decreased cytokine production upon TCR activation, decreased proliferation, increased expression of inhibitory cell surface receptors, and increased susceptibility to apoptosis. We characterized sarcoidosis CD4+ T cell immune function in systemic and local environments among subjects undergoing disease progression compared to those experiencing disease resolution. Spontaneous and TCR-stimulated Th1 cytokine expression and proliferation assays were performed in 53 sarcoidosis subjects and 30 healthy controls. PD-1 expression and apoptosis were assessed by flow cytometry. Compared to healthy controls, sarcoidosis CD4+ T cells demonstrated reductions in Th1 cytokine expression, proliferative capacity (p < 0.05), enhanced apoptosis (p < 0.01), and increased PD-1 expression (p < 0.001). BAL-derived CD4+ T cells also demonstrated multiple facets of T cell exhaustion (p < 0.05). Reversal of CD4+ T cell exhaustion was observed in subjects undergoing spontaneous resolution (p < 0.05). Sarcoidosis CD4+ T cells exhibit loss of cellular function during progressive disease that follows the archetype of T cell exhaustion. PMID:29234685

SLC6A4 expression and anti-proliferative responses to serotonin transporter ligands chlomipramine and fluoxetine in primary B-cell malignancies.

PubMed

Chamba, Anita; Holder, Michelle J; Jarrett, Ruth F; Shield, Lesley; Toellner, Kai M; Drayson, Mark T; Barnes, Nicholas M; Gordon, John

2010-08-01

B-cell lines of diverse neoplastic origin express the serotonin transporter (SERT/SLC6A4) and growth arrest in response to SERT-ligands, including the antidepressants chlomipramine and fluoxetine. Here we detail SLC6A4 transcript (Q-PCR) and protein (FACS) expression in primary cells from patients with: chronic lymphocytic leukaemia; mantle cell lymphoma; follicular lymphoma; Burkitt's lymphoma; and diffuse large B-cell lymphoma. The ability of the SERT-binding antidepressants to impact the growth of these cells when sustained on CD154-transfected fibroblasts was also determined. The results reveal a broad spectrum of primary B-cell malignancies expressing SLC6A4 with a proportion additionally displaying growth arrest on SERT-ligand exposure. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

In vitro anti-proliferative activity on colon cancer cell line (HT-29) of Thai medicinal plants selected from Thai/Lanna medicinal plant recipe database "MANOSROI III".

PubMed

Manosroi, Aranya; Akazawa, Hiroyuki; Akihisa, Toshihiro; Jantrawut, Pensak; Kitdamrongtham, Worapong; Manosroi, Worapaka; Manosroi, Jiradej

2015-02-23

Thai/Lanna region has its own folklore wisdoms including the traditional medicinal plant recipes. Thai/Lanna medicinal plant recipe database "MANOSROI III" has been developed by Prof. Dr. Jiradej Manosroi. It consists of over 200,000 recipes for all diseases including cancer. To investigate the anti-proliferative and apoptotic activities on human colon cancer cell line (HT-29) as well as the cancer cell selectivity of the methanolic extracts (MEs) and fractions of the 23 selected plants from the "MANOSROI III" database. The 23 selected plants were extracted with methanol under reflux and evaluated for their anti-proliferative activity by sulforhodamine B assay. The 5 plants (Gloriosa superba, Caesalpinia sappan, Fibraurea tinctoria, Ventilago denticulata and Psophocarpus tetragonolobus) with potent anti-proliferative activity were fractionated by liquid-liquid partition to give 4 fractions including each hexane (HF), methanol-water (MF), n-butanol (BF) and water (WF) fractions. They were tested for anti-proliferative activity and cancer cell selectivity. The ME and fractions of G. superba which showed potent anti-proliferative activity were further examined for morphological changes and apoptotic activities by acridine orange (AO)/ethidium bromide (EB) staining. The ME of G. superba root showed active with the highest anti-proliferative activity at 9.17 and 1.58 folds of cisplatin and doxorubicin, respectively. After liquid-liquid partition, HF of V. denticulata, MFs of F. tinctoria, V. denticulata and BF of P. tetragonolobus showed higher anti-proliferative activities than their MEs. The MF of G. superba indicated the highest anti-proliferative activity at 7.73 and 1.34 folds of cisplatin and doxorubicin, respectively, but only 0.86 fold of its ME. The ME and HF, MF and BF of G. superba and MF of F. tinctoria demonstrated high cancer cell selectivity. At 50 µg/ml, ME, HF, MF and BF of G. superba demonstrated higher apoptotic activities than the two standard drugs. This present study has not only confirmed the traditional use of the Thai/Lanna medicinal plant recipes for cancer treatments, but also the potential of the selected plant, G. superba for the further development as a modern anti-cancer drug. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Activation of innate immunity by prostate specific antigen (PSA).

PubMed

Kodak, James A; Mann, Dean L; Klyushnenkova, Elena N; Alexander, Richard B

2006-11-01

Prostate specific antigen (PSA) is a serine protease secreted by the prostatic epithelium. The only known function of the protein is to cleave seminogelin. We wished to determine if PSA activated peripheral blood mononuclear cells (PBMC). PBMC and selected sub-populations were cultured with purified PSA. Secretion of IFNgamma was measured by cytokine capture flow cytometry and enzyme-linked immunosorbent assay. We observed secretion of IFNgamma and a proliferative response in PBMC cultured with PSA. We found that NK cells were the source of the IFNgamma but NK cells were not directly stimulated by PSA. Rather, a soluble factor secreted primarily by CD14 monocytes in response to PSA stimulated NK cells to secrete IFNgamma. PSA induces a pro-inflammatory response that results in the secretion of INFgamma by NK cells. The presence of large amounts of PSA could contribute to the common finding of inflammatory infiltrates in the prostate.

A T-cell-dependent humoral immune response is preserved during the administration of the nerve agent pre-treatment pyridostigmine bromide in a murine model.

PubMed

Griffiths, Gareth D; Telford, Gary; Hooi, Doreen S W; Cook, David L; Wilkinson, Lucy J; Green, Christopher A; Pritchard, David I

2005-03-01

Immune regulation, either via the autonomic nervous system or by a proposed "non-neuronal" cholinergic system, suggests that the immune system may be susceptible to perturbation by compounds affecting cholinergic function. Here, the current UK and US nerve agent pre-treatment, pyridostigmine bromide (PB) and the related anti-acetylcholinesterase (AChE) compounds physostigmine (PHY) and BW284c51 were tested for their ability to affect mouse splenocyte function in vitro. In addition, PB, at a dose equivalent to that received during pre-treatment for nerve agent poisoning, was tested for its effect on a T-cell-dependent humoral response to antigen in vivo in the mouse. None of the anti-AChEs tested affected concanavalin A (Con A)-, anti-CD3- or lipopolysaccharide LPS-driven splenocyte proliferation, in vitro, at concentrations expected to give effective nerve agent pre-treatment. However, higher concentrations (>100 microM) particularly of PHY caused some inhibition of the proliferative responses. In vivo, PB or saline was administered via 28-day mini-osmotic pumps to give a 25-40% inhibition of whole blood AChE in the PB-treated animals. During PB or saline administration, primary and secondary doses (i.p.) of sheep red blood cells (SRBC) were given and the humoral response determined by monitoring anti-SRBC IgM and IgG levels. Splenocytes isolated from the experimental animals were also examined for their proliferative and cytokine responses to stimulation. No remarkable effects of PB were seen during the period of AChE inhibition on the humoral immune response. However, a modest elevation in IL-2 and IFN(gamma) in Con A-stimulated lymphocytes was seen in PB-treated animals following pump removal. Overall these data suggest that, in vivo, the SRBC stimulated T-cell-dependent immune response is unaffected by the administration of PB at pre-treatment doses.

CXC195 suppresses proliferation and inflammatory response in LPS-induced human hepatocellular carcinoma cells via regulating TLR4-MyD88-TAK1-mediated NF-κB and MAPK pathway

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

Wang, Yiting; Tu, Qunfei; Yan, Wei

Highlights: • CXC195 exhibited significant anti-proliferative effect and induced cell cycle arrest in LPS-induced HepG2 cells. • CXC195 suppressed the release of pro-inflammatory mediators in LPS-induced HepG2 cells. • CXC195 regulated TLR4-MyD88-TAK1-mediated NF-κB and MAPK pathway in LPS-induced HepG2 cells. - Abstract: CXC195 showed strong protective effects in neuronal apoptosis by exerting its antioxidant activity. However, the anti-cancer effects of CXC195 is still with limited acquaintance. Here, we investigated the role of CXC195 in lipopolysaccharide (LPS)-induced human hepatocellular carcinoma (HCC) cells lines (HepG2) and the possible signaling pathways. CXC195 exhibited significant anti-proliferative effect and induced cell cycle arrest in LPS-inducedmore » HepG2 cells. In addition, CXC195 suppressed the release of pro-inflammatory mediators in LPS-induced HepG2 cells, including TNF-α, iNOS, IL-1β, IL-6, CC chemokine ligand (CCL)-2, CCL-22 and epidermal growth factor receptor (EGFR). Moreover, CXC195 inhibited the expressions and interactions of TLR4, MyD88 and TAK1, NF-κB translocation to nucleus and its DNA binding activity, phosphorylation of ERK1/2, p38 and JNK. Our results suggested that treatment with CXC195 could attenuate the TLR4-mediated proliferation and inflammatory response in LPS-induced HepG2 cells, thus might be beneficial for the treatment of HCC.« less

Cell output, cell cycle duration and neuronal specification: a model of integrated mechanisms of the neocortical proliferative process

NASA Technical Reports Server (NTRS)

Caviness, V. S. Jr; Goto, T.; Tarui, T.; Takahashi, T.; Bhide, P. G.; Nowakowski, R. S.

2003-01-01

The neurons of the neocortex are generated over a 6 day neuronogenetic interval that comprises 11 cell cycles. During these 11 cell cycles, the length of cell cycle increases and the proportion of cells that exits (Q) versus re-enters (P) the cell cycle changes systematically. At the same time, the fate of the neurons produced at each of the 11 cell cycles appears to be specified at least in terms of their laminar destination. As a first step towards determining the causal interrelationships of the proliferative process with the process of laminar specification, we present a two-pronged approach. This consists of (i) a mathematical model that integrates the output of the proliferative process with the laminar fate of the output and predicts the effects of induced changes in Q and P during the neuronogenetic interval on the developing and mature cortex and (ii) an experimental system that allows the manipulation of Q and P in vivo. Here we show that the predictions of the model and the results of the experiments agree. The results indicate that events affecting the output of the proliferative population affect both the number of neurons produced and their specification with regard to their laminar fate.

Mesenchymal Stromal Cells Express GARP/LRRC32 on Their Surface: Effects on Their Biology and Immunomodulatory Capacity

PubMed Central

Carrillo-Galvez, Ana Belén; Cobo, Marién; Cuevas-Ocaña, Sara; Gutiérrez-Guerrero, Alejandra; Sánchez-Gilabert, Almudena; Bongarzone, Pierpaolo; García-Pérez, Angélica; Muñoz, Pilar; Benabdellah, Karim; Toscano, Miguel G; Martín, Francisco; Anderson, Per

2015-01-01

Mesenchymal stromal cells (MSCs) represent a promising tool for therapy in regenerative medicine, transplantation, and autoimmune disease due to their trophic and immunomodulatory activities. However, we are still far from understanding the mechanisms of action of MSCs in these processes. Transforming growth factor (TGF)-β1 is a pleiotropic cytokine involved in MSC migration, differentiation, and immunomodulation. Recently, glycoprotein A repetitions predominant (GARP) was shown to bind latency-associated peptide (LAP)/TGF-β1 to the cell surface of activated Foxp3+ regulatory T cells (Tregs) and megakaryocytes/platelets. In this manuscript, we show that human and mouse MSCs express GARP which presents LAP/TGF-β1 on their cell surface. Silencing GARP expression in MSCs increased their secretion and activation of TGF-β1 and reduced their proliferative capacity in a TGF-β1-independent manner. Importantly, we showed that GARP expression on MSCs contributed to their ability to inhibit T-cell responses in vitro. In summary, we have found that GARP is an essential molecule for MSC biology, regulating their immunomodulatory and proliferative activities. We envision GARP as a new target for improving the therapeutic efficacy of MSCs and also as a novel MSC marker. Stem Cells 2015;33:183–195 PMID:25182959

Mitochondria-Mediated Protein Regulation Mechanism of Polymorphs-Dependent Inhibition of Nanoselenium on Cancer Cells

NASA Astrophysics Data System (ADS)

Wang, Ge; Guo, Yuming; Yang, Gai; Yang, Lin; Ma, Xiaoming; Wang, Kui; Zhu, Lin; Sun, Jiaojiao; Wang, Xiaobing; Zhang, Hua

2016-08-01

The present study was (i) to prepare two types of selenium nanoparticles, namely an amorphous form of selenium quantum dots (A-SeQDs) and a crystalline form of selenium quantum dots (C-SeQDs); and (ii) to investigate the nano-bio interactions of A-SeQDs and C-SeQDs in MCF-7, HepG2, HeLa, NIH/3T3, L929 cells and BRL-3A cells. It was found that A-SeQDs could induce the mitochondria-mediated apoptosis, necrosis and death of cells, while C-SeQDs had much weaker effects. This polymorphs-dependent anti-proliferative activity of nano-selenium was scarcely reported. Further investigation demonstrated that A-SeQDs could differentially regulate 61 proteins and several pathways related to stress response, protein synthesis, cell migration and cell cycle, including “p38 MAPK Signaling”, “p53 Signaling”, “14-3-3-mediated Signaling”, “p70S6K Signaling” and “Protein Ubiquitination Pathway”. This was the first report to demonstrate the involvement of protein synthesis and post-translational modification pathways in the anti-proliferative activity associated with NMs. Compared with previously fragmentary studies, this study use a nanomics approach combining bioinformatics and proteomics to systematically investigate the nano-bio interactions of selenium nanoparticles in cancer cells.

Smart, Injury-Triggered Therapy for Ocular Trauma

DTIC Science & Technology

2016-10-01

prognosis due to retinal cell death , scar formation, and lack of functional regeneration. Proliferative vitreoretinopathy (PVR), a form of intraocular...Proteases, Metalloproteinases, Cell death , Gene Therapy 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a. NAME...vision has a poor prognosis due to retinal cell death , scar formation, and lack of functional regeneration. Proliferative vitreoretinopathy (PVR), a

Extrinsic and Intrinsic Apoptotic Responses Induced by Shiitake Culinary-Medicinal Mushroom Lentinus edodes (Agaricomycetes) Aqueous Extract against a Larynx Carcinoma Cell Line.

PubMed

Finimundy, Tiane C; Scola, Gustavo; Scariot, Fernando J; Dillon, Aldo J P; Moura, Sidnei; Echeverrigaray, Sérgio; Henriques, João Pegas; Roesch-Ely, Mariana

2018-01-01

Cumulative evidence from research studies has shown that the shiitake culinary-medicinal mushroom, Lentinus edodes, is an excellent source of natural antitumor agents and is capable of inhibiting cancer cell growth. However, the cell signaling pathway that leads tumor cells to apoptosis is not well understood because many chemical compounds may be acting. This study investigated the chemopreventive effects of an L. edodes aqueous extract on human HEp-2 epithelial larynx carcinoma cells and normal human MRC-5 lung fibroblasts by identifying proliferative and apoptotic pathways. The chemical characterization of the dry powder was assessed by high-performance liquid chromatography. Antiproliferative and proapoptotic effects induced by the extract were evaluated by assessing proliferative markers, cell sorting through flow cytometry, and expression levels of apoptotic proteins with Western blotting. The results suggest that inhibition of cell proliferation was more prominent in HEp-2 than in MRC-5 cells. Cell death analysis showed the appearance of cell populations in the sub-G1 phase, with late apoptotic signal increased in a dose-dependent manner. In addition, the aqueous extract induced depolarization of mitochondria, activating the generation of intracellular reactive oxygen species in HEp-2 cells. These observations suggest that L. edodes extract may exert a chemopreventive effect, regulating mitotic induction of apoptogenic signals. These findings highlight the mushroom's pharmacological potential in cancer treatment.

CD4+ T-cell clones obtained from cattle chronically infected with Fasciola hepatica and specific for adult worm antigen express both unrestricted and Th2 cytokine profiles.

PubMed Central

Brown, W C; Davis, W C; Dobbelaere, D A; Rice-Ficht, A C

1994-01-01

The well-established importance of helper T (Th)-cell subsets in immunity and immunoregulation of many experimental helminth infections prompted a detailed study of the cellular immune response against Fasciola hepatica in the natural bovine host. T-cell lines established from two cattle infected with F. hepatica were characterized for the expression of T-cell surface markers and proliferative responses against F. hepatica adult worm antigen. Parasite-specific T-cell lines contained a mixture of CD4+, CD8+, and gamma/delta T-cell-receptor-bearing T cells. However, cell lines containing either fewer than 10% CD8+ T cells or depleted of gamma/delta T cells proliferated vigorously against F. hepatica antigen, indicating that these T-cell subsets are not required for proliferative responses in vitro. Seventeen F. hepatica-specific CD4+ Th-cell clones were examined for cytokine expression following concanavalin A stimulation. Biological assays to measure interleukin-2 (IL-2) or IL-4, gamma interferon (IFN-gamma), and tumor necrosis factor and Northern (RNA) blot analysis to verify the expression of IL-2, IL-4, and IFN-gamma revealed that the Th-cell clones expressed a spectrum of cytokine profiles. Several Th-cell clones were identified as Th2 cells by the strong expression of IL-4 but little or no IL-2 or IFN-gamma mRNA. The majority of Th-cell clones were classified as Th0 cells by the expression of either all three cytokines or combinations of IL-2 and IL-4 or IL-4 and IFN-gamma. No Th1-cell clones were obtained. All of the Th-cell clones expressed a typical memory cell surface phenotype, characterized as CD45Rlow, and all expressed the lymph node homing receptor (L selectin). These results are the first to describe cytokine responses of F. hepatica-specific T cells obtained from infected cattle and extend our previous analysis of Th0 and Th1 cells from cattle immune to Babesia bovis (W. C. Brown, V. M. Woods, D. A. E. Dobbelaere, and K. S. Logan, Infect. Immun. 61:3273-3281, 1993) to include F. hepatica-specific Th2 cells. Images PMID:7509319

In pursuit of synergy: An investigation of the PI3K/mTOR/MEK co-targeted inhibition strategy in NSCLC

PubMed Central

Heavey, Susan; Cuffe, Sinead; Finn, Stephen; Young, Vincent; Ryan, Ronan; Nicholson, Siobhan; Leonard, Niamh; McVeigh, Niall; Barr, Martin; O'Byrne, Kenneth; Gately, Kathy

2016-01-01

Clinical PI3K inhibition has been somewhat disappointing, due to both inadequate patient stratification and compensatory cell signalling through bypass mechanisms. As such, investigation of PI3K-MEK co-targeted inhibition has been recommended. With high mortality rates and a clear need for new therapeutic intervention strategies, non-small cell lung cancer (NSCLC) is an important setting to investigate the effectiveness of this approach. Here, 174 NSCLC tumours were screened for 150 mutations by Fluidigm technology, with 15 patients being profiled for phosphoprotein expression. The effects of GDC-0941 (a pan PI3K inhibitor), GDC-0980 (a dual PI3K/mTOR inhibitor) and GDC-0973 (a MEK inhibitor) alone and in combination were assessed in 3 NSCLC cell lines. PIK3CA was mutated in 5.17% of NSCLC patients. GDC-0941 and GDC-0980 treatment induced anti-proliferative and pro-apoptotic responses across all NSCLC cell lines, while GDC-0973 treatment induced only anti-proliferative responses. GDC-0980 and GDC-0973 combined treatment led to significant increases in apoptosis and synergistic reductions in proliferation across the panel of cell lines. This study found that the PI3K/MEK co-targeted inhibition strategy is synergistic in all 3 molecular subtypes of NSCLC investigated. Consequently, we would advocate clinical trials for NSCLC patients combining GDC-0980 and GDC-0973, each of which are separately under clinical investigation currently. PMID:27765909

In pursuit of synergy: An investigation of the PI3K/mTOR/MEK co-targeted inhibition strategy in NSCLC.

PubMed

Heavey, Susan; Cuffe, Sinead; Finn, Stephen; Young, Vincent; Ryan, Ronan; Nicholson, Siobhan; Leonard, Niamh; McVeigh, Niall; Barr, Martin; O'Byrne, Kenneth; Gately, Kathy

2016-11-29

Clinical PI3K inhibition has been somewhat disappointing, due to both inadequate patient stratification and compensatory cell signalling through bypass mechanisms. As such, investigation of PI3K-MEK co-targeted inhibition has been recommended. With high mortality rates and a clear need for new therapeutic intervention strategies, non-small cell lung cancer (NSCLC) is an important setting to investigate the effectiveness of this approach.Here, 174 NSCLC tumours were screened for 150 mutations by Fluidigm technology, with 15 patients being profiled for phosphoprotein expression. The effects of GDC-0941 (a pan PI3K inhibitor), GDC-0980 (a dual PI3K/mTOR inhibitor) and GDC-0973 (a MEK inhibitor) alone and in combination were assessed in 3 NSCLC cell lines.PIK3CA was mutated in 5.17% of NSCLC patients. GDC-0941 and GDC-0980 treatment induced anti-proliferative and pro-apoptotic responses across all NSCLC cell lines, while GDC-0973 treatment induced only anti-proliferative responses. GDC-0980 and GDC-0973 combined treatment led to significant increases in apoptosis and synergistic reductions in proliferation across the panel of cell lines.This study found that the PI3K/MEK co-targeted inhibition strategy is synergistic in all 3 molecular subtypes of NSCLC investigated. Consequently, we would advocate clinical trials for NSCLC patients combining GDC-0980 and GDC-0973, each of which are separately under clinical investigation currently.

Effect of polysaccharide of dendrobium candidum on proliferation and apoptosis of human corneal epithelial cells in high glucose

PubMed Central

Li, Qiangxiang; Chen, Jing; Li, Yajia; Chen, Ting; Zou, Jing; Wang, Hua

2017-01-01

Abstract Background: The aim of the study was to observe the effect of polysaccharide of dendrobium candidum (PDC) and high glucose on proliferation, apoptosis of human corneal epithelial cells (HCEC). Methods: The MTT method was used to screen and take the optimal high-glucose concentration, treatment time, and PDC concentration using HCEC and divide it into 4 groups: control group (C), high glucose group (HG), PDC group, and HG + PDC group. We observed and compared the effect of the 4 groups on HCEC proliferation by MTT, apoptosis by Annexin V-FITC/PI double fluorescent staining and flow cytometry (FCM), and expression of bax mRNA and bcl-2 mRNA by RT-qPCR. Results: Compared with the control group, proliferative activity of HCEC cells was reduced; the cells apoptosis ratio was increased; the expression of bax mRNA was increased, and the expression of bcl-2 mRNA was reduced in the HG group. Proliferative activity of HCEC cells in the PDC group was increased, and the expression of bcl-2 mRNA was increased but that of bax mRNA was decreased. Proliferative activity of HCEC cells in the HG + PDC group was increased, but it could not restore to the normal level; the expression of bax mRNA was significantly decreased but the expression of bcl-2 mRNA was significantly increased. Conclusions: Our results demonstrate that high glucose can inhibit proliferative activity and induce apoptosis of HCEC. PDC can improve the proliferative activity of HCEC cells under the high glucose environment and reduce the apoptosis of cells by regulating the expression of bax and bcl-2. PDC play a very important role on protecting and repairing of corneal epithelial cells damage in high glucose. PMID:28796073

Differentiation expression during proliferative activity induced through different pathways: in situ hybridization study of thyroglobulin gene expression in thyroid epithelial cells

PubMed Central

1990-01-01

In canine thyrocytes in primary culture, our previous studies have identified three mitogenic agents and pathways: thyrotropin (TSH) acting through cyclic AMP (cAMP), EGF and its receptor tyrosine protein kinase, and the phorbol esters that stimulate protein kinase C. TSH enhances, while EGF and phorbol esters inhibit, the expression of differentiation. Given that growth and differentiation expression are often considered as mutually exclusive activities of the cells, it was conceivable that the differentiating action of TSH was restricted to noncycling (Go) cells, while the inhibition of the differentiation expression by EGF and phorbol esters only concerned proliferating cells. Therefore, the capacity to express the thyroglobulin (Tg) gene, the most prominent marker of differentiation in thyrocytes, was studied in proliferative cells (with insulin) and in quiescent cells (without insulin). Using cRNA in situ hybridization, we observed that TSH (and, to a lesser extent, insulin and insulin-like growth factor I) restored or maintained the expression of the Tg gene. Without these hormones, the Tg mRNA content became undetectable in most of the cells. EGF and 12-0-tetradecanoyl phorbol-13-acetate (TPA) inhibited the Tg mRNA accumulation induced by TSH (and/or insulin). Most of the cells (up to 90%) responded to both TSH and EGF. Nevertheless, the range of individual response was quite variable. The effects of TSH and EGF on differentiation expression were not dependent on insulin and can therefore be dissociated from their mitogenic effects. Cell cycling did not affect the induction of Tg gene. Indeed, the same cell distribution of Tg mRNA content was observed in quiescent cells stimulated by TSH alone, or in cells approximately 50% of which had performed one mitotic cycle in response to TSH + insulin. Moreover, after proliferation in "dedifferentiating" conditions (EGF + serum + insulin), thyrocytes had acquired a fusiform fibroblast-like morphology, and responded to TSH by regaining a characteristic epithelial shape and high Tg mRNA content. 32 h after the replacement of EGF by TSH, cells in mitosis presented the same distribution of the Tg mRNA content as the rest of the cell population. This implies that cell cycling (at least 27 h, as previously shown) did not affect the induction of the Tg gene which is clearly detectable after a time lag of at least 24 h. The data unequivocally show that the reexpression of differentiation and proliferative activity are separate but fully compatible processes when induced by cAMP in thyrocytes.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:2199463

GLD-4-Mediated Translational Activation Regulates the Size of the Proliferative Germ Cell Pool in the Adult C. elegans Germ Line

PubMed Central

Millonigg, Sophia; Eckmann, Christian R.

2014-01-01

To avoid organ dysfunction as a consequence of tissue diminution or tumorous growth, a tight balance between cell proliferation and differentiation is maintained in metazoans. However, cell-intrinsic gene expression mechanisms controlling adult tissue homeostasis remain poorly understood. By focusing on the adult Caenorhabditis elegans reproductive tissue, we show that translational activation of mRNAs is a fundamental mechanism to maintain tissue homeostasis. Our genetic experiments identified the Trf4/5-type cytoplasmic poly(A) polymerase (cytoPAP) GLD-4 and its enzymatic activator GLS-1 to perform a dual role in regulating the size of the proliferative zone. Consistent with a ubiquitous expression of GLD-4 cytoPAP in proliferative germ cells, its genetic activity is required to maintain a robust proliferative adult germ cell pool, presumably by regulating many mRNA targets encoding proliferation-promoting factors. Based on translational reporters and endogenous protein expression analyses, we found that gld-4 activity promotes GLP-1/Notch receptor expression, an essential factor of continued germ cell proliferation. RNA-protein interaction assays documented also a physical association of the GLD-4/GLS-1 cytoPAP complex with glp-1 mRNA, and ribosomal fractionation studies established that GLD-4 cytoPAP activity facilitates translational efficiency of glp-1 mRNA. Moreover, we found that in proliferative cells the differentiation-promoting factor, GLD-2 cytoPAP, is translationally repressed by the stem cell factor and PUF-type RNA-binding protein, FBF. This suggests that cytoPAP-mediated translational activation of proliferation-promoting factors, paired with PUF-mediated translational repression of differentiation factors, forms a translational control circuit that expands the proliferative germ cell pool. Our additional genetic experiments uncovered that the GLD-4/GLS-1 cytoPAP complex promotes also differentiation, forming a redundant translational circuit with GLD-2 cytoPAP and the translational repressor GLD-1 to restrict proliferation. Together with previous findings, our combined data reveals two interconnected translational activation/repression circuitries of broadly conserved RNA regulators that maintain the balance between adult germ cell proliferation and differentiation. PMID:25254367

Proliferative and morphologic changes in rat colon following bypass surgery.

PubMed Central

Barkla, D. H.; Tutton, P. J.

1985-01-01

In this study the proliferative and morphologic changes that occur in the colon of normal and dimethylhydrazine-treated rats following surgical bypass of the middle third of the colon are reported. Proliferative changes were measured by estimating accumulated mitotic indexes following vinblastine treatment and morphologic changes were observed with the use of light microscopy and scanning electron microscopy. Data were collected on Days 0, 7, 14, 30, and 72 after surgery. The results show that surgical bypass produces contrasting effects in the segments proximal to and distal to the suture line. In the proximal segment there was morphologic evidence of hyperplasia, although proliferative activity was unchanged except for an increase at 7 days in normal rats. In the distal segment there was a long-lived increase in the mitotic index, although morphologic changes were not seen. The results for DMH-treated rats were similar to those in normal rats. Groups of isolated dysplastic epithelial cells were often seen in the submucosa adjacent to sutures up to 72 days after surgery. Increased lymphoid infiltration was seen in segments proximal to but not distal to the suture line. It is hypothesized that the different responses of the proximal and distal segments may be related to the different embryologic origins of those segments. It is also hypothesized that the seeding of the submucosa with epithelial cells during suturing may be a factor in tumor recurrence. Images Figure 19 Figure 20 Figure 21 Figure 22 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Figure 16 Figure 17 Figure 18 PMID:4014432

From the Hayflick mosaic to the mosaics of ageing. Role of stress-induced premature senescence in human ageing.

PubMed

Toussaint, Olivier; Remacle, Jose; Dierick, Jean-François; Pascal, Thierry; Frippiat, Christophe; Zdanov, Stéphanie; Magalhaes, Joao Pedro; Royer, Véronique; Chainiaux, Florence

2002-11-01

The Hayflick limit-senescence of proliferative cell types-is a fundamental feature of proliferative cells in vitro. Various human proliferative cell types exposed in vitro to many types of subcytotoxic stresses undergo stress-induced premature senescence (SIPS) (also called stress-induced premature senescence-like phenotype, according to the definition of senescence). The known mechanisms of appearance the main features of SIPS are reviewed: senescent-like morphology, growth arrest, senescence-related changes in gene expression, telomere shortening. Long before telomere-shortening induces senescence, other factors such as culture conditions or lack of 'feeder cells' can trigger either SIPS or prolonged reversible G(0) phase of the cell cycle. In vivo, 'proliferative' cell types of aged individuals are likely to compose a mosaic made of cells irreversibly growth arrested or not. The higher level of stress to which these cells have been exposed throughout their life span, the higher proportion of the cells of this mosaic will be in SIPS rather than in telomere-shortening dependent senescence. All cell types undergoing SIPS in vivo, most notably the ones in stressful conditions, are likely to participate in the tissular changes observed along ageing. For instance, human diploid fibroblasts (HDFs) exposed in vivo and in vitro to pro-inflammatory cytokines display biomarkers of senescence and might participate in the degradation of the extracellular matrix observed in ageing.

Anti-proliferative and mutagenic activities of aqueous and methanol extracts of leaves from Pereskia bleo (Kunth) DC (Cactaceae).

PubMed

Er, Hui Meng; Cheng, En-Hsiang; Radhakrishnan, Ammu Kutty

2007-09-25

The anti-proliferative effects of the aqueous and methanol extracts of leaves of Pereskia bleo (Kunth) DC (Cactaceae) against a mouse mammary cancer cell line (4T1) and a normal mouse fibroblast cell line (NIH/3T3) were evaluated under an optimal (in culture medium containing 10% foetal bovine serum (FBS)) and a sub-optimal (in culture medium containing 0.5% FBS) conditions. Under the optimal condition, the aqueous extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in 4T1 cells and 300 microg/mL in NIH/3T3 cells, whereas the methanol extract did not show any notable anti-proliferative effect in these cell lines, at any of the concentrations tested. Under the sub-optimal condition, the aqueous extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in NIH/3T3 cells, whilst the methanol extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in both cell lines. An upward trend of apoptosis was observed in both 4T1 and NIH/3T3 cells treated with increasing concentrations of the aqueous extract. The level of apoptosis observed at all the concentrations of the aqueous extract tested was consistently higher than necrosis. There was a significant (p<0.05) increase in the level of necrosis observed in the 4T1 cells treated with 300 microg/mL of the methanol extract. Generally, the level of necrosis was noted to be higher than that of apoptosis in the methanol extract-treated cells. The mutagenicity assay performed showed that in the absence of S-9 liver metabolic activation, the extract was not mutagenic up to the concentration of 165 microg/mL . However, in the presence of S-9 liver metabolic activation, the aqueous extract was mutagenic at all the concentrations tested. This study shows that both the aqueous and methanol extracts of the leaves from Pereskia bleo (Kunth) DC (Cactaceae) do not have appreciable anti-proliferative effect on the 4T1 and NIH/3T3 cells as the EC(50) values obtained are greater than 50 microg/mL when tested under optimal culture condition. Moreover, the aqueous extract may form mutagenic compound(s) upon the metabolisation by liver enzymes.

Melicope ptelefolia leaf extracts exhibit antioxidant activity and exert anti-proliferative effect with apoptosis induction on four different cancer cell lines.

PubMed

Kabir, Mohammad Faujul; Mohd Ali, Johari; Abolmaesoomi, Mitra; Hashim, Onn Haji

2017-05-05

Melicope ptelefolia is a well-known herb in a number of Asian countries. It is often used as vegetable salad and traditional medicine to address various ailments. However, not many studies have been currently done to evaluate the medicinal benefits of M. ptelefolia (MP). The present study reports antioxidant, anti-proliferative, and apoptosis induction activities of MP leaf extracts. Young MP leaves were dried, powdered and extracted sequentially using hexane (HX), ethyl acetate (EA), methanol (MeOH) and water (W). Antioxidant activity was evaluated using ferric reducing antioxidant power (FRAP), 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) radicals scavenging and cellular antioxidant activity (CAA) assays. Anti-proliferative activity was evaluated through cell viability assay, using the following four human cancer cell lines: breast (HCC1937, MDA-MB-231), colorectal (HCT116) and liver (HepG2). The anti-proliferative activity was further confirmed through cell cycle and apoptosis assays, including annexin-V/7-aminoactinomycin D staining and measurements of caspase enzymes activation and inhibition. Overall, MP-HX extract exhibited the highest antioxidant potential, with IC 50 values of 267.73 ± 5.58 and 327.40 ± 3.80 μg/mL for ABTS and DPPH radical-scavenging assays, respectively. MP-HX demonstrated the highest CAA activity in Hs27 cells, with EC 50 of 11.30 ± 0.68 μg/mL, while MP-EA showed EC 50 value of 37.32 ± 0.68 μg/mL. MP-HX and MP-EA showed promising anti-proliferative activity towards the four cancer cell lines, with IC 50 values that were mostly below 100 μg/mL. MP-HX showed the most notable anti-proliferative activity against MDA-MB-231 (IC 50  = 57.81 ± 3.49 μg/mL) and HCT116 (IC 50  = 58.04 ± 0.96 μg/mL) while MP-EA showed strongest anti-proliferative activity in HCT116 (IC 50  = 64.69 ± 0.72 μg/mL). The anticancer potential of MP-HX and MP-EA were also demonstrated by their ability to induce caspase-dependent apoptotic cell death in all of the cancer cell lines tested. Cell cycle analysis suggested that both the MP-HX and MP-EA extracts were able to disrupt the cell cycle in most of the cancer cell lines. MP-HX and MP-EA extracts demonstrated notable antioxidant, anti-proliferative, apoptosis induction and cancer cell cycle inhibition activities. These findings reflect the promising potentials of MP to be a source of novel phytochemical(s) with health promoting benefits that are also valuable for nutraceutical industry and cancer therapy.

High-molecular-weight tropomyosins localize to the contractile rings of dividing CNS cells but are absent from malignant pediatric and adult CNS tumors.

PubMed

Hughes, Julie A I; Cooke-Yarborough, Claire M; Chadwick, Nigel C; Schevzov, Galina; Arbuckle, Susan M; Gunning, Peter; Weinberger, Ron P

2003-04-01

Tropomyosin has been implicated in the control of actin filament dynamics during cell migration, morphogenesis, and cytokinesis. In order to gain insight into the role of tropomyosins in cell division, we examined their expression in developing and neoplastic brain tissue. We found that the high-molecular-weight tropomyosins are downregulated at birth, which correlates with glial cell differentiation and withdrawal of most cells from the cell cycle. Expression of these isoforms was restricted to proliferative areas in the embryonic brain and was absent from the adult, where the majority of cells are quiescent. However, they were induced under conditions where glial cells became proliferative in response to injury. During cytokinesis, these tropomyosin isoforms were associated with the contractile ring. We also investigated tropomyosin expression in neoplastic CNS tissues. Low-grade astrocytic tumors expressed high-molecular-weight tropomyosins, while highly malignant CNS tumors of diverse origin did not (P

Regulation of gene expression in autoimmune disease loci and the genetic basis of proliferation in CD4+ effector memory T cells.

PubMed

Hu, Xinli; Kim, Hyun; Raj, Towfique; Brennan, Patrick J; Trynka, Gosia; Teslovich, Nikola; Slowikowski, Kamil; Chen, Wei-Min; Onengut, Suna; Baecher-Allan, Clare; De Jager, Philip L; Rich, Stephen S; Stranger, Barbara E; Brenner, Michael B; Raychaudhuri, Soumya

2014-06-01

Genome-wide association studies (GWAS) and subsequent dense-genotyping of associated loci identified over a hundred single-nucleotide polymorphism (SNP) variants associated with the risk of rheumatoid arthritis (RA), type 1 diabetes (T1D), and celiac disease (CeD). Immunological and genetic studies suggest a role for CD4-positive effector memory T (CD+ TEM) cells in the pathogenesis of these diseases. To elucidate mechanisms of autoimmune disease alleles, we investigated molecular phenotypes in CD4+ effector memory T cells potentially affected by these variants. In a cohort of genotyped healthy individuals, we isolated high purity CD4+ TEM cells from peripheral blood, then assayed relative abundance, proliferation upon T cell receptor (TCR) stimulation, and the transcription of 215 genes within disease loci before and after stimulation. We identified 46 genes regulated by cis-acting expression quantitative trait loci (eQTL), the majority of which we detected in stimulated cells. Eleven of the 46 genes with eQTLs were previously undetected in peripheral blood mononuclear cells. Of 96 risk alleles of RA, T1D, and/or CeD in densely genotyped loci, eleven overlapped cis-eQTLs, of which five alleles completely explained the respective signals. A non-coding variant, rs389862A, increased proliferative response (p=4.75 × 10-8). In addition, baseline expression of seventeen genes in resting cells reliably predicted proliferative response after TCR stimulation. Strikingly, however, there was no evidence that risk alleles modulated CD4+ TEM abundance or proliferation. Our study underscores the power of examining molecular phenotypes in relevant cells and conditions for understanding pathogenic mechanisms of disease variants.

Identification of molecular subtypes of gastric cancer with different responses to PI3-kinase inhibitors and 5-fluorouracil.

PubMed

Lei, Zhengdeng; Tan, Iain Beehuat; Das, Kakoli; Deng, Niantao; Zouridis, Hermioni; Pattison, Sharon; Chua, Clarinda; Feng, Zhu; Guan, Yeoh Khay; Ooi, Chia Huey; Ivanova, Tatiana; Zhang, Shenli; Lee, Minghui; Wu, Jeanie; Ngo, Anna; Manesh, Sravanthy; Tan, Elisabeth; Teh, Bin Tean; So, Jimmy Bok Yan; Goh, Liang Kee; Boussioutas, Alex; Lim, Tony Kiat Hon; Flotow, Horst; Tan, Patrick; Rozen, Steven G

2013-09-01

Almost all gastric cancers are adenocarcinomas, which have considerable heterogeneity among patients. We sought to identify subtypes of gastric adenocarcinomas with particular biological properties and responses to chemotherapy and targeted agents. We compared gene expression patterns among 248 gastric tumors; using a robust method of unsupervised clustering, consensus hierarchical clustering with iterative feature selection, we identified 3 major subtypes. We developed a classifier for these subtypes and validated it in 70 tumors from a different population. We identified distinct genomic and epigenomic properties of the subtypes. We determined drug sensitivities of the subtypes in primary tumors using clinical survival data, and in cell lines through high-throughput drug screening. We identified 3 subtypes of gastric adenocarcinoma: proliferative, metabolic, and mesenchymal. Tumors of the proliferative subtype had high levels of genomic instability, TP53 mutations, and DNA hypomethylation. Cancer cells of the metabolic subtype were more sensitive to 5-fluorouracil than the other subtypes. Furthermore, in 2 independent groups of patients, those with tumors of the metabolic subtype appeared to have greater benefits with 5-fluorouracil treatment. Tumors of the mesenchymal subtype contain cells with features of cancer stem cells, and cell lines of this subtype are particularly sensitive to phosphatidylinositol 3-kinase-AKT-mTOR inhibitors in vitro. Based on gene expression patterns, we classified gastric cancers into 3 subtypes, and validated these in an independent set of tumors. The subgroups have differences in molecular and genetic features and response to therapy; this information might be used to select specific treatment approaches for patients with gastric cancer. Copyright © 2013 AGA Institute. Published by Elsevier Inc. All rights reserved.

Transcript and protein profiling identifies signaling, growth arrest, apoptosis, and NF-κB survival signatures following GNRH receptor activation

PubMed Central

Meyer, Colette; Sims, Andrew H; Morgan, Kevin; Harrison, Beth; Muir, Morwenna; Bai, Jianing; Faratian, Dana; Millar, Robert P; Langdon, Simon P

2013-01-01

GNRH significantly inhibits proliferation of a proportion of cancer cell lines by activating GNRH receptor (GNRHR)-G protein signaling. Therefore, manipulation of GNRHR signaling may have an under-utilized role in treating certain breast and ovarian cancers. However, the precise signaling pathways necessary for the effect and the features of cellular responses remain poorly defined. We used transcriptomic and proteomic profiling approaches to characterize the effects of GNRHR activation in sensitive cells (HEK293-GNRHR, SCL60) in vitro and in vivo, compared to unresponsive HEK293. Analyses of gene expression demonstrated a dynamic response to the GNRH superagonist Triptorelin. Early and mid-phase changes (0.5–1.0 h) comprised mainly transcription factors. Later changes (8–24 h) included a GNRH target gene, CGA, and up- or downregulation of transcripts encoding signaling and cell division machinery. Pathway analysis identified altered MAPK and cell cycle pathways, consistent with occurrence of G2/M arrest and apoptosis. Nuclear factor kappa B (NF-κB) pathway gene transcripts were differentially expressed between control and Triptorelin-treated SCL60 cultures. Reverse-phase protein and phospho-proteomic array analyses profiled responses in cultured cells and SCL60 xenografts in vivo during Triptorelin anti-proliferation. Increased phosphorylated NF-κB (p65) occurred in SCL60 in vitro, and p-NF-κB and IκBϵ were higher in treated xenografts than controls after 4 days Triptorelin. NF-κB inhibition enhanced the anti-proliferative effect of Triptorelin in SCL60 cultures. This study reveals details of pathways interacting with intense GNRHR signaling, identifies potential anti-proliferative target genes, and implicates the NF-κB survival pathway as a node for enhancing GNRH agonist-induced anti-proliferation. PMID:23202794

Differences in immune cell function between tuberculosis positive and negative Asian elephants.

PubMed

Landolfi, Jennifer A; Miller, Michele; Maddox, Carol; Zuckermann, Federico; Langan, Jennifer N; Terio, Karen A

2014-07-01

Tuberculosis is an important health concern for Asian elephant (Elephas maximus) populations worldwide, however, mechanisms underlying susceptibility to Mycobacterium tuberculosis are unknown. Proliferative responses assessed via brominated uridine incorporation and cytokine expression measured by real-time RT-PCR were evaluated in peripheral blood mononuclear cell (PBMC) cultures from 8 tuberculosis negative and 8 positive Asian elephants. Cultures were stimulated with Mycobacterium bovis purified protein derivative (PPD-B), M. tuberculosis culture filtrate protein (CFP)-10, and Mycobacterium avium PPD (PPD-A). Following stimulation with PPD-B, proliferation was higher (α = 0.005) in positive samples; no significant differences were detected following CFP-10 or PPD-A stimulation. Tumor necrosis factor (TNF)-α, interleukin (IL)-12, and interferon (IFN)-γ expression was greater in samples from positive elephants following stimulation with PPD-B (α = 0.025) and CFP-10 (α = 0.025 TNF-α and IL-12; α = 0.005 IFN-γ). Stimulation with PPD-A also produced enhanced IL-12 expression in positive samples (α = 0.025). Findings suggested that differences in immune cell function exist between tuberculosis positive and negative elephants. Proliferative responses and expression of TNF-α, IL-12, and IFN-γ in response to stimulation with PPD-B and CFP-10 differ between tuberculosis positive and negative elephants, suggesting these parameters may be important to tuberculosis immunopathogenesis in this species. Copyright © 2014 Elsevier Ltd. All rights reserved.

Hydrodynamics of stratified epithelium: Steady state and linearized dynamics

NASA Astrophysics Data System (ADS)

Yeh, Wei-Ting; Chen, Hsuan-Yi

2016-05-01

A theoretical model for stratified epithelium is presented. The viscoelastic properties of the tissue are assumed to be dependent on the spatial distribution of proliferative and differentiated cells. Based on this assumption, a hydrodynamic description of tissue dynamics at the long-wavelength, long-time limit is developed, and the analysis reveals important insights into the dynamics of an epithelium close to its steady state. When the proliferative cells occupy a thin region close to the basal membrane, the relaxation rate towards the steady state is enhanced by cell division and cell apoptosis. On the other hand, when the region where proliferative cells reside becomes sufficiently thick, a flow induced by cell apoptosis close to the apical surface enhances small perturbations. This destabilizing mechanism is general for continuous self-renewal multilayered tissues; it could be related to the origin of certain tissue morphology, tumor growth, and the development pattern.

Telomerase Reverse Transcriptase Deficiency Prevents Neointima Formation Through Chromatin Silencing of E2F1 Target Genes.

PubMed

Endorf, Elizabeth B; Qing, Hua; Aono, Jun; Terami, Naoto; Doyon, Geneviève; Hyzny, Eric; Jones, Karrie L; Findeisen, Hannes M; Bruemmer, Dennis

2017-02-01

Aberrant proliferation of smooth muscle cells (SMC) in response to injury induces pathological vascular remodeling during atherosclerosis and neointima formation. Telomerase is rate limiting for tissue renewal and cell replication; however, the physiological role of telomerase in vascular diseases remains to be determined. The goal of the present study was to determine whether telomerase reverse transcriptase (TERT) affects proliferative vascular remodeling and to define the molecular mechanism by which TERT supports SMC proliferation. We first demonstrate high levels of TERT expression in replicating SMC of atherosclerotic and neointimal lesions. Using a model of guidewire-induced arterial injury, we demonstrate decreased neointima formation in TERT-deficient mice. Studies in SMC isolated from TERT-deficient and TERT overexpressing mice with normal telomere length established that TERT is necessary and sufficient for cell proliferation. TERT deficiency did not induce a senescent phenotype but resulted in G1 arrest albeit hyperphosphorylation of the retinoblastoma protein. This proliferative arrest was associated with stable silencing of the E2F1-dependent S-phase gene expression program and not reversed by ectopic overexpression of E2F1. Finally, chromatin immunoprecipitation and accessibility assays revealed that TERT is recruited to E2F1 target sites and promotes chromatin accessibility for E2F1 by facilitating the acquisition of permissive histone modifications. These data indicate a previously unrecognized role for TERT in neointima formation through epigenetic regulation of proliferative gene expression in SMC. © 2016 American Heart Association, Inc.

Effects of silibinin on growth and invasive properties of human ovarian carcinoma cells through suppression of heregulin/HER3 pathway.

PubMed

Momeny, Majid; Ghasemi, Reza; Valenti, Giovanni; Miranda, Mariska; Zekri, Ali; Zarrinrad, Ghazaleh; Javadikooshesh, Sepehr; Yaghmaie, Marjan; Alimoghaddam, Kamran; Ghavamzadeh, Ardeshir; Ghaffari, Seyed H

2016-03-01

Epithelial ovarian cancer (EOC) is the most fatal gynecological malignancy due to its high proliferative and invasive capacities. A heregulin (HRG)/HER3 autocrine loop increases proliferative and metastatic properties of EOC cells, suggesting that modulators of this signaling pathway may prove effective to trammel growth and motility of these cells. This study aimed to evaluate the effects of multi-tyrosine kinase inhibitor silibinin on proliferative and invasive characteristics of EOC cell lines OVCAR8 and SKOV3 through suppression of the HRG/HER3 pathway. To achieve this, the effects of silibinin on proliferation, DNA synthesis, clonogenicity, cell cycle progression, cathepsin B enzymatic activity, and migration and invasion were explored in vitro. Silibinin suppressed proliferation, DNA synthesis, and clonogenic abilities of OVCAR8 and SKOV3 cells through inhibition of the autocrine HRG/HER3 circuit. Silibinin-mediated attenuation of the HER3 signaling disabled the HER3/AKT/survivin axis and thereby, induced G1/S cell cycle arrest. Furthermore, silibinin reduced invasive potentials of the EOC cells through quelling the HRG/HER3 pathway and suppression of cathepsin B activity. Altogether, these results suggest that silibinin is a potential anti-cancer drug to inhibit proliferative and invasive characteristics of the EOC cells that exhibit an autocrine HRG/HER3 pathway.

Elucidation of proliferative capability of mononuclear tetraploid cells, emerging spontaneously from diploid cells, using image cytometry and fluorescence in situ hybridization.

PubMed

Ito, Hideaki; Oga, Atsunori; Furuya, Tomoko; Ikemoto, Kenzo; Amakawa, Genta; Chochi, Yasuyo; Kawauchi, Shigeto; Sasaki, Kohsuke

2013-06-01

Proliferation of tetraploid cells (TCs) emerging from diploid cells is considered to be a critical event toward tumourigenesis, or cancer progression. Recently, several studies have reported that binuclear TCs emerging from normal cells are capable of mitosis, however, it has not been confirmed directly whether mononuclear TCs emerging from normal cells could proliferate, even cancer cells. The aim of this study is to detect mononuclear TCs in vitro, spontaneously emerging from diploid cells and to elucidate their proliferative capability directly. For this purpose, we have developed a novel method. In this study, two completely disomic cell lines were used, TIG-7, a fibroblast cell line and CAL-51, a breast cancer cell line. Cells were cultured on microscope slides and their DNA content was determined using an image cytometer. On the same slides, chromosome numbers were scored using centromere fluorescence in situ hybridization (FISH). For evaluating proliferative capability of TCs, bromodeoxyuridine (BrdUrd) incorporation and colony-forming ability were examined. Using our method, spontaneous emergence of mononuclear TCs was detected in both TIG-7 and CAL-51. Colonies of TIG-7 TCs were not observed, but were observed of CAL-51 TCs. Our method enables detection of mononuclear TCs and elucidation of their proliferative capability, directly; this evidence reveals that mononuclear TIG-7 TCs do not proliferate but that mononuclear CAL-51 TCs are able to. © 2013 Blackwell Publishing Ltd.

Low dose/low fluence ionizing radiation-induced biological effects: The role of intercellular communication and oxidative metabolism

NASA Astrophysics Data System (ADS)

Azzam, Edouard

Mechanistic investigations have been considered critical to understanding the health risks of exposure to ionizing radiation. To gain greater insight in the biological effects of exposure to low dose/low fluence space radiations with different linear energy transfer (LET) properties, we examined short and long-term biological responses to energetic protons and high charge (Z) and high energy (E) ions (HZE particles) in human cells maintained in culture and in targeted and non-targeted tissues of irradiated rodents. Particular focus of the studies has been on mod-ulation of gene expression, proliferative capacity, induction of DNA damage and perturbations in oxidative metabolism. Exposure to mean doses of 1000 MeV/nucleon iron ions, by which a small to moderate proportion of cells in an exposed population is targeted through the nucleus by an HZE particle, induced stressful effects in the irradiated and non-irradiated cells in the population. Direct intercellular communication via gap-junctions was a primary mediator of the propagation of stressful effects from irradiated to non-irradiated cells. Compromised prolif-erative capacity, elevated level of DNA damage and oxidative stress evaluated by measurements of protein carbonylation, lipid peroxidation and activity of metabolic enzymes persisted in the progeny of irradiated and non-irradiated cells. In contrast, progeny of cells exposed to high or low doses from 150-1000 MeV protons retained the ability to form colonies and harbored similar levels of micronuclei, a surrogate form of DNA damage, as control, which correlated with normal reactive oxygen species (ROS) levels. Importantly, a significant increase in the spontaneous neoplastic transformation frequency was observed in progeny of bystander mouse embryo fibroblasts (MEFs) co-cultured with MEFs irradiated with energetic iron ions but not protons. Of particular significance, stressful effects were detected in non-targeted tissues of rats that received partial body irradiation, 20 months earlier, from low mean doses of HZE particles. These effects were associated with disruption of mitochondrial function in the non-irradiated tissues and in modulation of immune cell populations. Collectively, our data support the concept that the response of the organism to high LET radiations involves irradiated and non-irradiated cells/tissues and is associated with changes in several physiological functions. Supported by the US National Aeronautics and Space Administration

Mesenchymal stromal cells express GARP/LRRC32 on their surface: effects on their biology and immunomodulatory capacity.

PubMed

Carrillo-Galvez, Ana Belén; Cobo, Marién; Cuevas-Ocaña, Sara; Gutiérrez-Guerrero, Alejandra; Sánchez-Gilabert, Almudena; Bongarzone, Pierpaolo; García-Pérez, Angélica; Muñoz, Pilar; Benabdellah, Karim; Toscano, Miguel G; Martín, Francisco; Anderson, Per

2015-01-01

Mesenchymal stromal cells (MSCs) represent a promising tool for therapy in regenerative medicine, transplantation, and autoimmune disease due to their trophic and immunomodulatory activities. However, we are still far from understanding the mechanisms of action of MSCs in these processes. Transforming growth factor (TGF)-β1 is a pleiotropic cytokine involved in MSC migration, differentiation, and immunomodulation. Recently, glycoprotein A repetitions predominant (GARP) was shown to bind latency-associated peptide (LAP)/TGF-β1 to the cell surface of activated Foxp3(+) regulatory T cells (Tregs) and megakaryocytes/platelets. In this manuscript, we show that human and mouse MSCs express GARP which presents LAP/TGF-β1 on their cell surface. Silencing GARP expression in MSCs increased their secretion and activation of TGF-β1 and reduced their proliferative capacity in a TGF-β1-independent manner. Importantly, we showed that GARP expression on MSCs contributed to their ability to inhibit T-cell responses in vitro. In summary, we have found that GARP is an essential molecule for MSC biology, regulating their immunomodulatory and proliferative activities. We envision GARP as a new target for improving the therapeutic efficacy of MSCs and also as a novel MSC marker. © 2014 AlphaMed Press.

MYC activation is a hallmark of cancer initiation and maintenance.

PubMed

Gabay, Meital; Li, Yulin; Felsher, Dean W

2014-06-02

The MYC proto-oncogene has been implicated in the pathogenesis of most types of human tumors. MYC activation alone in many normal cells is restrained from causing tumorigenesis through multiple genetic and epigenetically controlled checkpoint mechanisms, including proliferative arrest, apoptosis, and cellular senescence. When pathologically activated in a permissive epigenetic and/or genetic context, MYC bypasses these mechanisms, enforcing many of the "hallmark" features of cancer, including relentless tumor growth associated with DNA replication and transcription, cellular proliferation and growth, protein synthesis, and altered cellular metabolism. MYC mandates tumor cell fate, by inducing stemness and blocking cellular senescence and differentiation. Additionally, MYC orchestrates changes in the tumor microenvironment, including the activation of angiogenesis and suppression of the host immune response. Provocatively, brief or even partial suppression of MYC back to its physiological levels of activation can result in the restoration of intrinsic checkpoint mechanisms, resulting in acute and sustained tumor regression, associated with tumor cells undergoing proliferative arrest, differentiation, senescence, and apoptosis, as well as remodeling of the tumor microenvironment, recruitment of an immune response, and shutdown of angiogenesis. Hence, tumors appear to be "addicted" to MYC because of both tumor cell-intrinsic, cell-autonomous and host-dependent, immune cell-dependent mechanisms. Both the trajectory and persistence of many human cancers require sustained MYC activation. Multiscale mathematical modeling may be useful to predict when tumors will be addicted to MYC. MYC is a hallmark molecular feature of both the initiation and maintenance of tumorigenesis. Copyright © 2014 Cold Spring Harbor Laboratory Press; all rights reserved.

Host Immune Response to Bacterial Cyclic Diguanylic Acid (c-di-GMP)

DTIC Science & Technology

2009-07-01

at 4°C. Sera were harvested and kept at 20°C until tested. The institutional ethics committee on animal experimentation of the Faculté des Sciences...On day 5, nonadherent DCs were harvested by gentle pipetting, counted, and plated in fresh medium containing GM-CSF and IL-4 (50 ng/ml each). On day...incubation, cells were pulsed with 1 Ci of [3H]thymidine/well for 18 h and were harvested on filter paper. Proliferative responses were measured as [3H

Tolerogenic dendritic cells inhibit antiphospholipid syndrome derived effector/memory CD4⁺ T cell response to β2GPI.

PubMed

Torres-Aguilar, Honorio; Blank, Miri; Kivity, Shaye; Misgav, Mudi; Luboshitz, Jacob; Pierangeli, Silvia S; Shoenfeld, Yehuda

2012-01-01

The importance of β(2)-glycoprotein I (β(2)GPI)-specific CD4(+) T cells in the development of pathogenic processes in patients with antiphospholipid syndrome (APS) and APS mouse models is well established. Therefore, our objective is to manipulate the β2GPI specific CD4(+) T cells using tolerogenic dendritic cells (tDCs) to induce tolerance. We aim to evaluate the capability of tDCs to induce antigen-specific tolerance in effector/memory T cells from patients with APS and to elucidate the involved mechanism. DCs and tDCs were produced from patients with APS peripheral-blood-monocytes, using specific cytokines. β(2)GPI-specific tolerance induction was investigated by coculturing control DC (cDC) or tDC, β(2)GPI-loaded, with autologous effector/memory T cells, evaluating the proliferative response, phenotype, cytokines secretion, viability and regulatory T cells. Human monocyte-derived DCs treated with interleukin (IL)-10 and transforming growth factor β-1 (10/TGF-DC) induced β(2)GPI-specific-unresponsiveness in effector/memory CD4(+) T cells (46.5% ± 26.0 less proliferation) in 16 of 20 analysed patients with APS, without affecting the proliferative response to an unrelated candidin. In five analysed patients, 10/TGF-DC-stimulated T cells acquired an IL-2(low)interferon γ(low)IL-10(high) cytokine profile, with just a propensity to express higher numbers of Foxp3(+)CTLA-4(+) cells, but with an evident suppressive ability. In four of 10 analysed patients, 10/TGF-DC-stimulated T cell hyporesponsiveness could not be reverted and showed higher percentages of late apoptosis, p<0.02. The inherent tolerance induction resistance of activated T cells present during the development of autoimmune diseases has delayed the application of tDC as an alternative therapy. This study highlights the 10/TGF-DC feasibility to induce antigen-specific unresponsiveness in autoreactive T cells generated in patients with APS by inducing apoptosis or T cells with regulatory abilities.

Suppression of Proinflammatory Cytokines in Functionalized Fullerene-Exposed Dermal Keratinocytes

DOE PAGES

Gao, Jun; Wang, Hsing-Lin; Iyer, Rashi

2010-01-01

Initial experiments using differentially functionalized fullerenes, CD-, hexa-, and tris-, suggested a properties dependent effect on cytotoxic and proliferative responses in human skin keratinocytes. In the present study we investigated the cytokine secretion profile of dermal epithelial cells exposed to functionalized fullerenes. Keratinocyte-derived cytokines affect homing and trafficking of normal and malignant epidermal immune as well as nonimmune cells in vivo. These cytokines are critical for regulating activation, proliferation, and differentiation of epidermal cells. Our results indicate that tris- (size range <100 nm) significantly reduces inflammatory cytokine release in a dose- and time-dependent manner. In contrast CD- demonstrated a relatively pro-inflammatorymore » cytokine response, while hexa- did not significantly perturb cytokine responses. Physical and chemical characterizations of these engineered nanomaterials suggest that the disparate biological responses observed may potentially be a function of the aggregation properties of these fullerenes.« less

Actual Proliferating Index and p53 protein expression as prognostic marker in odontogenic cysts.

PubMed

Gadbail, A R; Chaudhary, M; Patil, S; Gawande, M

2009-10-01

The purpose of this study was to evaluate the biological aggressiveness of odontogenic keratocyst/keratocystic odontogenic tumour (KCOT), radicular cyst (RC) and dentigerous cyst (DC) by observing the actual proliferative activity of epithelium, and p53 protein expression. The actual proliferative activity was measured by Ki-67 Labelling Index and argyrophilic nucleolar organizing regions (AgNOR) count per nucleus. The p53 protein expression was also evaluated. Ki-67 positive cells were observed higher in suprabasal cell layers of KCOT with uniform distribution, a few of them were predominantly observed in basal cell layer in RC and DC. The AgNOR count was significantly higher in suprabasal cell layers of KCOT. The actual proliferative activity was noted to be higher in suprabasal cell layers of KCOT. The p53 immunolabelling was dense and scattered in basal and suprabasal cell layers in KCOT. The weakly stained p53 positive cells were observed diffusely distributed in KCOT, whereas they were mainly seen in basal cell layer of RC and DC. The quantitative and qualitative differences of the proliferative activity and the p53 protein expression in sporadic KCOT may be associated with intrinsic growth potential that could play a role in its development and explain locally aggressive biological behaviour. AgNOR count and p53 protein detection in odontogenic lesions can be of great consequence to predict the biological behaviour and prognosis.

Cellular basis for neonatally induced T-suppressor activity. Primary B cell maturation is blocked by suppressor-helper interactions restricted by loci on chromosome 12

PubMed Central

1985-01-01

The cellular mechanism and genetic restriction of neonatally induced HA- specific suppressor T (Ts) cells have been examined. The in vivo effect of these Ts cells on antibody production, primary B cell proliferation, B cell surface marker changes, and helper T (Th) cell priming during primary responses to HA have been determined. The results indicate that, although antigen-induced B cell proliferative responses and surface marker changes occur in the presence of Ts cells, differentiation to Ig secretion, and long-lived memory B cell production are prevented. Further, antigen-specific Th cell priming is completely ablated by Ts cells, suggesting that Ts act by preventing the delivery of Th signals required for both the later stages of primary B cell maturation, and the formation of memory B cell populations. Finally, in vivo cell mixing experiments using congenic mice indicate that this Ts-Th interaction is restricted by loci on mouse chromosome 12. PMID:2580040

Chronic toxic and carcinogenic effects of oral cadmium in the Noble (NBL/Cr) rat: induction of neoplastic and proliferative lesions of the adrenal, kidney, prostate, and testes.

PubMed

Waalkes, M P; Anver, M R; Diwan, B A

1999-10-29

Based on the occurrence of pulmonary cancers in exposed populations, cadmium is classified as a human carcinogen. More controversial target sites for cadmium in humans include the prostate and kidney, where some studies have shown a link between cadmium and cancer. In Wistar rats cadmium induces tumors in the ventral prostate. The relevance of such lesions to humans is debated since the rat ventral lobe, unlike the dorsolateral lobe, has no embryological homolog in the human prostate. Cadmium has not been linked with renal tumors in rodents but is a potent nephrotoxin. In this work we studied the effects of oral cadmium in the Noble (NBL/Cr) rat with particular attention to proliferative lesions of the prostate and kidneys. Cadmium (as CdCl2) was given ad libitum throughout the study in the drinking water at doses of 0, 25, 50, 100, and 200 ppm Cd to groups (initial n = 30) of male rats, which were observed for up to 102 wk. At the lower doses of cadmium (< or =50 ppm) a clear dose-related increase in total proliferative lesions of the prostate (ventral and dorsolateral lesions combined) occurred (0 ppm = 21% incidence, 25 ppm = 46%, 50 ppm = 50%; trend p < .03). These lesions were described as intraepithelial hyperplasia with occasional areas of atypical epithelial cells without stromal invasion. The lesions occurred primarily in the dorsolateral prostate with cadmium exposure and most frequently showed three or more foci within each specimen. At higher doses, prostatic proliferative lesions declined to control levels. The loss of prostatic response at the higher doses was likely due to diminished testicular function secondary to cadmium treatment. This was reflected in lesions indicative of testicular hypofunction at the highest cadmium dose, namely, interstitial cell hyperplasia, and a strong correlation between cadmium dose and total proliferative lesions of the testes (hyperplasias and tumors combined). Renal tumors (mainly mesenchymal and pelvic transitional cell), although few in number, showed a positive correlation with cadmium dose, as did pelvic transitional epithelial hyperplasia. Renal lesions were not associated with any cadmium-induced changes in age-related chronic nephropathy. The incidence of pheochromocytomas of the adrenal was increased by cadmium but only at the 50 ppm dose. Inflammatory lesions of the liver and spleen were common at higher doses and showed strong trends based on dose. These results indicate that oral cadmium can induce proliferative lesions in the prostate and kidney of the Noble rat. The finding of proliferative lesions of dorsolateral prostate in rats has presumed relevance to human prostate cancers.

Effect of indinavir used alone or in double or triple combination with AZT and ddC on human immune functions.

PubMed

Mattioli, Benedetta; Giordani, Luciana; Quaranta, Maria Giovanna; Viora, Marina

2004-03-19

Indinavir (IDV) is a potent and selective human immunodeficiency virus type 1 (HIV-1) protease inhibitor (PI) widely used in antiretroviral therapy, but its effects on the immune system are relatively unknown. In this study we have investigated the in vitro effect of IDV on normal human peripheral blood mononuclear cells (PBMC). We used the drug alone or in double and triple combination with AZT and ddC to assess whether IDV interferes with the previously observed immunomodulatory effects induced by AZT and ddC. We found that proliferative response, induction of immunoglobulins (Ig) production and cytokine production was not modulated by IDV. More importantly, IDV used in double or triple combination with AZT and ddC, does not further strenghten the inhibition of proliferative response induced by AZT and is able to abrogate the inhibitory effect induced by ddC on proliferative response. Similarly, IDV/AZT, IDV/ddC and IDV/AZT/ddC combinations does not strenghten the modulation of TNF-alpha, IFN-gamma and IL-4 induced by AZT, ddC and AZT/ddC. On the other hand, IDV neutralizes the up-regulating effects of AZT on IL-2 production while the up-regulating effects of ddC on IL-2 production is not affected. These data suggest that IDV used in combination with AZT and ddC did not add any further immunotoxicity.

Immunity to herpes simplex virus type 2. Suppression of virus-induced immune responses in ultraviolet B-irradiated mice

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

Yasumoto, S.; Hayashi, Y.; Aurelian, L.

1987-10-15

Ultraviolet B irradiation (280 to 320 nm) of mice at the site of intradermal infection with herpes simplex virus type 2 increased the severity of the herpes simplex virus type 2 disease and decreased delayed-type hypersensitivity (DTH) responses to viral antigen. Decrease in DTH resulted from the induction of suppressor T cells, as evidenced by the ability of spleen cells from UV-irradiated mice to inhibit DTH and proliferative responses after adoptive transfer. Lymph node cells from UV-irradiated animals did not transfer suppression. DTH was suppressed at the induction but not the expression phase. Suppressor T cells were Lyt-1+, L3T4+, andmore » their activity was antigen-specific. However, after in vitro culture of spleen cells from UV-irradiated mice with herpes simplex virus type 2 antigen, suppressor activity was mediated by Lyt-2+ cells. Culture supernatants contained soluble nonantigen-specific suppressive factors.« less

The apoptotic and anti-proliferative activity of Origanum majorana extracts on human leukemic cell line.

PubMed

Abdel-Massih, Roula M; Fares, Rida; Bazzi, Samer; El-Chami, Nisrine; Baydoun, Elias

2010-08-01

Scientists are constantly searching for phytochemicals and compounds with anti-cancer and antioxidant activity. In this study, the anti-proliferative activity of plant extracts from Origanum majorana (marjoram) was tested on human lymphoblastic leukemia cell line Jurkat. Cytotoxicity was examined using non-radioactive cytotoxicity assay and the IC(50) was calculated. At non-cytotoxic concentrations, the viability of cells decreased with increase of concentration of plant extract. The anti-proliferative effect was also found to be dose-dependent. Analysis via flow cytometry shows that marjoram extracts stimulated apoptosis. Induction of apoptosis was caused by an up-regulation of p53 protein levels and down-regulation of Bcl-2alpha. Marjoram exhibited a strong scavenging activity (SC(50)=0.03mg dry weight). The conclusions from this study suggest that marjoram extracts exhibit anti-proliferative effect and high antioxidant activity. For that it merits further investigation as a potential therapeutic agent. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Lycopene Protects Keratinocytes Against UVB Radiation-Induced Carcinogenesis via Negative Regulation of FOXO3a Through the mTORC2/AKT Signaling Pathway.

PubMed

Chen, Ping; Xu, Shina; Qu, Jinlong

2018-01-01

Lycopene, one of the most potent anti-oxidants, has been reported to exhibit potent anti-proliferative properties in a wide range of cancer cells through modulation of the cell cycle and apoptosis. Forkhead box O3 (FOXO3a) plays a pivotal role in modulating the expression of genes involved in cell death. Herein, we investigated the role of FOXO3a signaling in the anti-cancer effects of lycopene. Results showed that lycopene pretreatment attenuated UVB-induced cell hyper-proliferation and promoted apoptosis, accompanied by decreased cyclin-dependent kinase 2 (CDK2) and CDK4 complex in both human keratinocytes and SKH-1 hairless mice. FOXO3a is phosphorylated in response to UVB irradiation and sequestered in the cytoplasm, while lycopene pretreatment rescued this sensitization. Gene ablation of FOXO3a attenuated lycopene-induced decrease in cell hyper-proliferation, CDK2, and CDK4 complex, indicating a critical role of FOXO3a in the lycopene-induced anti-proliferative effect of keratinocytes during UVB irradiation. Transfection with FOXO3a siRNA inhibited the lycopene-induced increase in cell apoptosis, BAX and cleaved PARP expression. Moreover, loss of AKT induced further accelerated lycopene-induced FOXO3a dephosphorylation, while loss of mechanistic target of rapamycin complex 2 (mTORC2) by transfection with RICTOR siRNA induced levels of AKT phosphorylation comparable to those obtained with lycopene. In contrast, overexpression of AKT or mTORC2 decreased the effects of lycopene on the expression of FOXO3a as well as AKT phosphorylation, suggesting that lycopene depends on the negative modulation of mTORC2/AKT signaling. Taken together, our findings demonstrate that the mTORC2/AKT/FOXO3a axis plays a critical role in the anti-proliferative and pro-apoptotic effects of lycopene in UVB-induced photocarcinogenesis. J. Cell. Biochem. 119: 366-377, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Role of muscarinic receptors in the regulation of immune and inflammatory responses

PubMed Central

Razani-Boroujerdi, Seddigheh; Behl, Muskaan; Hahn, Fletcher F.; Pena-Philippides, Juan Carlos; Hutt, Julie; Sopori, Mohan L.

2008-01-01

Leukocytes contain both nicotinic and muscarinic receptors, and while activation of nicotinic receptors suppresses immune/inflammatory responses, the role of muscarinic receptors in immunity is unclear. We examined the effects of a muscarinic receptor antagonist (atropine) and agonist (oxotremorine), administered chronically through miniosmotic pumps, on immune/inflammatory responses in the rat. Results show that while oxotremorine stimulated, atropine inhibited the antibody and T-cell proliferative responses. Moreover, atropine also suppressed the turpentine-induced leukocytic infiltration and tissue injury, and inhibited chemotaxis of leukocytes toward neutrophil and monocyte/lymphocyte chemoattractants. Thus, activation of nicotinic and muscarinic receptors has opposite effects on the immune/inflammatory responses. PMID:18190972

Strategies for immortalization of primary hepatocytes

PubMed Central

Eva, Ramboer; Bram, De Craene; Joery, De Kock; Tamara, Vanhaecke; Geert, Berx; Vera, Rogiers; Mathieu, Vinken

2014-01-01

The liver has the unique capacity to regenerate in response to a damaging event. Liver regeneration is hereby largely driven by hepatocyte proliferation, which in turn relies on cell cycling. The hepatocyte cell cycle is a complex process that is tightly regulated by several well-established mechanisms. In vitro, isolated hepatocytes do not longer retain this proliferative capacity. However, in vitro cell growth can be boosted by immortalization of hepatocytes. Well-defined immortalization genes can be artificially overexpressed in hepatocytes or the cells can be conditionally immortalized leading to controlled cell proliferation. This paper discusses the current immortalization techniques and provides a state-of-the-art overview of the actually available immortalized hepatocyte-derived cell lines and their applications. PMID:24911463

The initiation of lateral roots in the primary roots of maize (Zea mays L.) implies a reactivation of cell proliferation in a group of founder pericycle cells.

PubMed

Alarcón, M Victoria; Lloret, Pedro G; Martín-Partido, Gervasio; Salguero, Julio

2016-03-15

The initiation of lateral roots (LRs) has generally been viewed as a reactivation of proliferative activity in pericycle cells that are committed to initiate primordia. However, it is also possible that pericycle founder cells that initiate LRs never cease proliferative activity but rather are displaced to the most distal root zones while undertaking successive stages of LR initiation. In this study, we tested these two alternative hypotheses by examining the incorporation of 5-bromo-2'-deoxyuridine (BrdU) into the DNA of meristematic root cells of Zea mays. According to the values for the length of the cell cycle and values for cell displacement along the maize root, our results strongly suggest that pericycle cells that initiate LR primordia ceased proliferative activity upon exiting the meristematic zone. This finding is supported by the existence of a root zone between 4 and 20mm from the root cap junction, in which neither mitotic cells nor labelled nuclei were observed in phloem pericycle cells. Copyright © 2016 Elsevier GmbH. All rights reserved.

Genetic control of T cell responsiveness to the Friend murine leukemia virus envelope antigen. Identification of class II loci of the H-2 as immune response genes

PubMed Central

1988-01-01

T cells primed specifically for the envelope glycoprotein of Friend murine leukemia helper virus (F-MuLV) were prepared by immunizing mice with a recombinant vaccinia virus that expressed the entire env gene of F-MuLV. Significant proliferative responses of F-MuLV envelope- specific, H-2a/b T cells were observed when the T cells were stimulated with antigen-pulsed peritoneal exudate cells (PEC) having the b allele at the K, A beta, A alpha, and E beta loci of the H-2. On the other hand, PEC having only the kappa allele at these loci did not induce the envelope-specific T cell proliferation, even when the PEC had the b allele at the E alpha, S, or D loci. F-MuLV envelope-specific proliferation of H-2a/b T cells under the stimulation of antigen- pulsed, H-2a/b PEC was specifically blocked with anti-I-Ab and anti-I- Ek mAbs but not with anti-Kb, anti-Kk, or anti-I-Ak mAbs. Moreover, (B10.MBR x A/WySn)F1 mice that have the b allele only at the K locus but not in I-A subregion were nonresponders to the envelope glycoprotein, and the bm12 mutation at the A beta locus completely abolished the T cell responsiveness to this antigen. These results indicate that proliferative T cells recognize a limited number of epitopes on F-MuLV envelope protein in the context of I-Ab, hybrid I- Ak/b, and/or hybrid I-Ek/b class II MHC molecules but fail to recognize the same envelope protein in the context of I-Ak or I-Ek molecules. This influence of the H-2I region on T cell recognition of the envelope glycoprotein appeared to control in vivo induction of protective immunity against Friend virus complex after immunization with the vaccinia-F-MuLV env vaccine. Thus, these results provide, for the first time, direct evidence for Ir gene-controlled responder/nonresponder phenotypes influencing the immune response to a pathogenic virus of mice. PMID:3141552

[Complex estimation of proliferative activity of epithelial cells of the large intestine damaged by polyps and cancer].

PubMed

Nalieskina, L A; Zabarko, L B; Polishchuk, L Z; Oliĭnichenko, G P; Zakhartseva, L M; Koshel', K V

2001-01-01

Peculiarities of mitotic regime and expression of proliferating cell nuclear antigen were investigated in 18 polyps and 35 cases of colorectal cancer. Direct relationship between spectrum and degree of manifestation of proliferative activity, level of morphological malignant tumors and accumulation of oncopathology in the patient pedigrees was established.

L-Leucyl-L-Leucine Methyl Ester Treatment of Canine Marrow and Peripheral Blood Cells: Inhibition of Proliferative Responses with Maintenance of the Capacity for Autologous Marrow Engraftment

DTIC Science & Technology

1988-11-01

Copyright 0 198 by The Winiams & Wilkins Co. Printed in U.S.A. L-LEUCYL-L-LEUCINE METHYL ESTER TREATMENT OF CANINE MARROW AND PERIPHERAL BLOOD CELLS...Reearch CeThs eatetle, Washington 9%104 tInaiyuba on o canine UMrrowt and peripher hi Recently, Thiele and Lipsky have described adipeptide nionon clear...that marrow iincubation with Leu-Leu. Leu-Leu-OMe is a feasible method to deplete canine marrows of aloreactive and cytotoxic T cells prior to OMe

Antagonizing Effects of Aspartic Acid against Ultraviolet A-Induced Downregulation of the Stemness of Human Adipose Tissue-Derived Mesenchymal Stem Cells.

PubMed

Jung, Kwangseon; Cho, Jae Youl; Soh, Young-Jin; Lee, Jienny; Shin, Seoung Woo; Jang, Sunghee; Jung, Eunsun; Kim, Min Hee; Lee, Jongsung

2015-01-01

Ultraviolet A (UVA) irradiation is responsible for a variety of changes in cell biology. The purpose of this study was to investigate effects of aspartic acid on UVA irradiation-induced damages in the stemness properties of human adipose tissue-derived mesenchymal stem cells (hAMSCs). Furthermore, we elucidated the UVA-antagonizing mechanisms of aspartic acid. The results of this study showed that aspartic acid attenuated the UVA-induced reduction of the proliferative potential and stemness of hAMSCs, as evidenced by increased proliferative activity in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and upregulation of stemness-related genes OCT4, NANOG, and SOX2 in response to the aspartic acid treatment. UVA-induced reduction in the mRNA level of hypoxia-inducible factor (HIF)-1α was also significantly recovered by aspartic acid. In addition, the antagonizing effects of aspartic acid against the UVA effects were found to be mediated by reduced production of PGE2 through the inhibition of JNK and p42/44 MAPK. Taken together, these findings show that aspartic acid improves reduced stemness of hAMSCs induced by UVA and its effects are mediated by upregulation of HIF-1α via the inhibition of PGE2-cAMP signaling. In addition, aspartic acid may be used as an antagonizing agent to mitigate the effects of UVA.

Proliferation kinetics and cyclic AMP as prognostic factors in adult acute leukemia.

PubMed

Paietta, E; Mittermayer, K; Schwarzmeier, J

1980-07-01

In 41 adult patients with acute leukemia (myeloblastic, lymphoblastic, and undifferentiated), proliferation kinetics (as determined by double-label autoradiography) and cyclic adenosine 3',5'-monophosphate (cAMP) concentration were studied for their significance in the prediction of responsiveness to cytostatic therapy. Patients with good clinical response had significantly shorter turnover times and higher labeling indices in the bone marrow than did those who failed to respond to treatment. Cases for which cell kinetics did not correlate with clinical response were explained by variance in the distribution of leukemic blasts between the proliferative cell cycle and the resting pool. Good clinical response was also found to be associated with low levels of cAMP in leukemic cells prior to therapy, whereas high cAMP contents predicted failure. Low cAMP concentrations, however, did not necessarily correlate with short turnover times and vice versa. This might be due to fluctuations of the cAMP concentrations during the cell cycle.

Differential p53 engagement in response to oxidative and oncogenic stresses in Fanconi anemia mice.

PubMed

Rani, Reena; Li, Jie; Pang, Qishen

2008-12-01

Members of the Fanconi anemia (FA) protein family are involved in repair of genetic damage caused by DNA cross-linkers. It is not clear whether the FA proteins function in oxidative DNA damage and oncogenic stress response. Here, we report that deficiency in the Fanca gene in mice elicits a p53-dependent growth arrest and DNA damage response to oxidative DNA damage and oncogenic stress. Using a Fanca-/-Trp53-/- double knockout model and a functionally switchable p53 retrovirus, we define the kinetics, dependence, and persistence of p53-mediated response to oxidative and oncogenic stresses in Fanca-/- cells. Notably, oxidative stress induces persistent p53 response in Fanca-/- cells, likely due to accumulation of unrepaired DNA damage. On the other hand, whereas wild-type cells exhibit prolonged response to oncogene activation, the p53-activating signals induced by oncogenic ras are short-lived in Fanca-/- cells, suggesting that Fanca may be required for the cell to engage p53 during constitutive ras activation. We propose that the FA proteins protect cells from stress-induced proliferative arrest and tumor evolution by acting as a modulator of the signaling pathways that link FA to p53.

Differential p53 engagement in response to oxidative and oncogenic stresses in Fanconi anemia mice

PubMed Central

Rani, Reena; Li, Jie; Pang, Qishen

2008-01-01

Members of the Fanconi anemia (FA) protein family are involved in repair of genetic damage caused by DNA cross-linkers. It is not clear whether the FA proteins function in oxidative DNA damage and oncogenic stress response. Here we report that deficiency in the Fanca gene in mice elicits a p53-dependent growth arrest and DNA damage response to oxidative DNA damage and oncogenic stress. Using a Fanca-/- Trp53-/- double knockout model and a functionally switchable p53 retrovirus, we define the kinetics, dependence, and persistence of p53-mediated response to oxidative and oncogenic stresses in Fanca-/- cells. Notably, oxidative stress induces persistent p53 response in Fanca-/- cells, likely due to accumulation of unrepaired DNA damage. On the other hand, whereas WT cells exhibit prolonged response to oncogene activation, the p53-activating signals induced by oncogenic ras are short-lived in Fanca-/- cells, suggesting that Fanca may be required for the cell to engage p53 during constitutive ras activation. We propose that the FA proteins protect cells from stress-induced proliferative arrest and tumor evolution by acting as a modulator of the signaling pathways that link FA to p53. PMID:19047147

The state of T cells before cryopreservation: Effects on post-thaw proliferation and function.

PubMed

Luo, Ying; Wang, Peng; Liu, Hui; Zhu, Zhengyan; Li, Chenglong; Gao, Yingtang

2017-12-01

We aim to assess the effect of the state of T cells before cryopreservation on the post-thaw proliferative capacity, phenotype and functional response. Peripheral blood mononuclear cells (PBMCs) were isolated from a hepatocellular carcinoma (HCC) patient, and the T cells were frozen during cell culture according to our experimental design. After a period of re-culture, the proliferative capacity of the cryopreserved cells, the expression of T cell surface markers and the secretion of IFN-γ and IL-10 were assayed. There was >90% cell viability after thaw in every group. Lymphocytes cryopreserved at day 4, 8 or 12 during the cell culture were allowed to recover for 24 h, whereas lymphocytes cryopreserved while freshly isolated were allowed to recover for 72 h. After the period of re-culture, cryopreservation at day 4, 8 or 12 during T cell culture was not found to alter the T cell subpopulation. The proportions of NKT and Treg cells were unchanged when cells were cryopreserved at day 12 during T cell culture. IFN-γ secretion was not impacted by cryopreservation, and IL-10 secretion was significantly decreased when cells were cryopreserved at day 8 or 12 during T cell culture. The state of T cells before cryopreservation has effects on the post-thaw proliferation capacity, the phenotype and the secretion of IFN-γ and IL-10. Cryopreservation of lymphocytes at day 8 or 12 during the cell culture may be the best choice for T cell immunotherapy. Copyright © 2017 Elsevier Inc. All rights reserved.

Cancer metabolism, stemness and tumor recurrence: MCT1 and MCT4 are functional biomarkers of metabolic symbiosis in head and neck cancer.

PubMed

Curry, Joseph M; Tuluc, Madalina; Whitaker-Menezes, Diana; Ames, Julie A; Anantharaman, Archana; Butera, Aileen; Leiby, Benjamin; Cognetti, David M; Sotgia, Federica; Lisanti, Michael P; Martinez-Outschoorn, Ubaldo E

2013-05-01

Here, we interrogated head and neck cancer (HNSCC) specimens (n = 12) to examine if different metabolic compartments (oxidative vs. glycolytic) co-exist in human tumors. A large panel of well-established biomarkers was employed to determine the metabolic state of proliferative cancer cells. Interestingly, cell proliferation in cancer cells, as marked by Ki-67 immunostaining, was strictly correlated with oxidative mitochondrial metabolism (OXPHOS) and the uptake of mitochondrial fuels, as detected via MCT1 expression (p < 0.001). More specifically, three metabolic tumor compartments were delineated: (1) proliferative and mitochondrial-rich cancer cells (Ki-67+/TOMM20+/COX+/MCT1+); (2) non-proliferative and mitochondrial-poor cancer cells (Ki-67-/TOMM20-/COX-/MCT1-); and (3) non-proliferative and mitochondrial-poor stromal cells (Ki-67-/TOMM20-/COX-/MCT1-). In addition, high oxidative stress (MCT4+) was very specific for cancer tissues. Thus, we next evaluated the prognostic value of MCT4 in a second independent patient cohort (n = 40). Most importantly, oxidative stress (MCT4+) in non-proliferating epithelial cancer cells predicted poor clinical outcome (tumor recurrence; p < 0.0001; log-rank test), and was functionally associated with FDG-PET avidity (p < 0.04). Similarly, oxidative stress (MCT4+) in tumor stromal cells was specifically associated with higher tumor stage (p < 0.03), and was a highly specific marker for cancer-associated fibroblasts (p < 0.001). We propose that oxidative stress is a key hallmark of tumor tissues that drives high-energy metabolism in adjacent proliferating mitochondrial-rich cancer cells, via the paracrine transfer of mitochondrial fuels (such as L-lactate and ketone bodies). New antioxidants and MCT4 inhibitors should be developed to metabolically target "three-compartment tumor metabolism" in head and neck cancers. It is remarkable that two "non-proliferating" populations of cells (Ki-67-/MCT4+) within the tumor can actually determine clinical outcome, likely by providing high-energy mitochondrial "fuels" for proliferative cancer cells to burn. Finally, we also show that in normal mucosal tissue, the basal epithelial "stem cell" layer is hyper-proliferative (Ki-67+), mitochondrial-rich (TOMM20+/COX+) and is metabolically programmed to use mitochondrial fuels (MCT1+), such as ketone bodies and L-lactate. Thus, oxidative mitochondrial metabolism (OXPHOS) is a common feature of both (1) normal stem cells and (2) proliferating cancer cells. As such, we should consider metabolically treating cancer patients with mitochondrial inhibitors (such as Metformin), and/or with a combination of MCT1 and MCT4 inhibitors, to target "metabolic symbiosis."

Analysis of in situ proliferative activity in oral gingival epithelium in patients with xerostomia.

PubMed

Celenligil-Nazliel, Haviye; Palali, Ali; Ayhan, Ayşe; Ruacan, Sevket

2003-02-01

Sjögren's syndrome is an autoimmune disease characterized by xerostomia and keratoconjunctivitis sicca. The relationship between xero-stomia and proliferative activity in human gingival epithelium is not known. Proliferating cell nuclear antigen (PCNA) is a nuclear protein associated with the cell cycle. Nuclear PCNA immunoreactivity is found in the proliferative compartment of normal tissues. The aims of this study were to evaluate PCNA expression in oral gingival epithelium of healthy and inflamed gingiva obtained from patients with Sjögren's syndrome, and to compare the results to age- and gender-matched subjects with normal salivary function. Eighteen Sjögren's syndrome patients and 28 controls (14 with chronic periodontitis and 14 with no clinical evidence of periodontal disease) were included in the study. Biopsies were obtained from both inflamed and healthy gingiva. The expression of PCNA was evaluated in formalin-fixed, paraffin-embedded gingival samples using an immunoperoxidase technique and PC10 monoclonal antibody to PCNA. PCNA expression was observed both in the basal and suprabasal layers, and was found to be more prominent in the suprabasal layers. Proliferative index (PI) in inflamed gingiva was significantly lower in the Sjögren's syndrome group. However, no significant difference was observed between the study and control groups with respect to PI in healthy gingiva. In both groups, PI was found to be increased due to inflammation. Our data indicate that proliferative activity is observed in the suprabasal layers and, less frequently, in the basal layer. Inflammation caused increased proliferative activity. However, this positive effect of inflammation on epithelial cell proliferation decreased significantly with a lack of saliva. Therefore, it appears that saliva-derived biological mediators may also contribute to increased proliferative activity observed during inflammation.

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens

PubMed Central

Kleiveland, Charlotte R.; Minic, Rajna; Moen, Lars F.; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Eijsink, Vincent G. H.

2016-01-01

ABSTRACT Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum. The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo. The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. IMPORTANCE This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this antigen to the bacterial cell wall or to the cell membrane. The recombinant strains elicited proliferative antigen-specific T-cell responses in white blood cells from tuberculosis-positive humans and induced specific immune responses after nasal and oral administrations in mice. PMID:27815271

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens.

PubMed

Kuczkowska, Katarzyna; Kleiveland, Charlotte R; Minic, Rajna; Moen, Lars F; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Mathiesen, Geir; Eijsink, Vincent G H

2017-01-15

Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this antigen to the bacterial cell wall or to the cell membrane. The recombinant strains elicited proliferative antigen-specific T-cell responses in white blood cells from tuberculosis-positive humans and induced specific immune responses after nasal and oral administrations in mice. Copyright © 2016 American Society for Microbiology.

Chemical Constituents from Cimicifuga dahurica and Their Anti-Proliferative Effects on MCF-7 Breast Cancer Cells.

PubMed

Huyen, Chu Thi Thanh; Luyen, Bui Thi Thuy; Khan, Ghulam Jilany; Oanh, Ha Van; Hung, Ta Manh; Li, Hui-Jun; Li, Ping

2018-05-04

This study was designed to search for novel anti-cancer compounds from natural plants. The 70% ethanolic extract from the rizhomes of Cimicifuga dahurica (Turcz.) Maxim. (Ranunculaceae) was found to possess significant in vitro anti-proliferative effects on MCF-7 breast cancer cells. A phytochemical investigation using assay-guided fractionation of the ethanolic extract of C. dahurica resulted in the isolation of one new phenolic amide glycoside 3 , two new lignan glycosides 4 and 7 , one new 9,19-cycloartane triterpenoid glycoside 6 , and thirteen known constituents 1 , 2 , 5 , and 8 ⁻ 17 . The structures of 3 , 4 , 6 , and 7 were established using contemporary NMR methods and from their HRESIMS data. The anti-proliferative effects of isolated compounds were evaluated using the BrdU-proliferation kit. Five among the 17 isolated compounds showed significant anti-proliferative effects ( p ≤ 0.05), wherein compound 7 showed the most significant anti-proliferative and cell cycle arresting effect ( p ≤ 0.05) which followed a dose dependent manner. Western blot protein expression analysis showed a down expression of c-Myc and cyclin D1 which further elucidated the anti-proliferation mechanism of compound 7 while apoptotic effects were found in association with Bcl-2 family protein expression variations. Conclusively this study reports the isolation and identification of 17 compounds from C. dahurica , including four novel molecules, in addition to the fact that compound 7 possesses significant anti-proliferative and apoptotic effects in vitro that may require further exploration.

Non-specific factor enhancement of human in vitro antigen-dependent antibody synthesis: role of B cell activation and T cell help.

PubMed Central

Brenner, M K; North, M E; Chadda, H R; Farrant, J

1984-01-01

Lectin-free supernatants obtained from PWM-stimulated lymphocytes, enable B cells to proliferate and secrete immunoglobulin. Both functions are augmented by the addition of irradiated T cells. In the presence of antigen, these supernatants also enhance specific anti-tetanus toxoid antibody production. The components of the supernatant responsible for these activities have a molecular weight between 30,000 and 60,000, and have the characteristics of non-specific factors: they are genetically unrestricted, and do not bind to either antigen or anti-DR affinity columns. There is no evidence that the partial T dependency of these factors is an indication that their target is a T cell. Instead, T cells appear necessary to move the B cell into a state of activation in which it becomes responsive to the factor. Alternative activation signals such as Staph. A. Cowan can substitute for T cell help in the proliferative response, but not for immunoglobulin or antibody synthesis. The implications of these results for the approaches used to detect and classify B cell growth factors are discussed. PMID:6608488

The mechanism of retrovirus suppression of human T cell proliferation in vitro.

PubMed

Copelan, E A; Rinehart, J J; Lewis, M; Mathes, L; Olsen, R; Sagone, A

1983-10-01

Immunosuppression is commonly associated with retrovirus-induced animal tumors. Studies in the murine and feline retrovirus systems suggest that the 15,000-dalton envelope protein (p15E) of the virion may contribute to immunosuppression by interfering with normal lymphocyte function. We examined the effect of inactivated feline leukemia virus (UV-FeLV) and p15E derived from this virus on concanavalin A (Con A) driven human T cell proliferation. Virus and p15E markedly suppressed mononuclear cell proliferative response to Con A. Suppression was not due to inhibition of monocyte accessory cell function, or interleukin 1 (IL 1) secretion. In fact, the presence of monocytes partially protected T cells from UV-FeLV suppression. UV-FeLV, however, suppressed T cell secretion of and response to interleukin 2 (IL 2). We conclude that UV-FeLV and derived p15E inhibit T cell proliferation by direct inhibition of T cell function. These findings, extended to the in vivo situations, suggest that retrovirus-associated suppression of the immune response involves the induction of T cell but not monocyte dysfunction.

TNFα-senescence initiates a STAT-dependent positive feedback loop, leading to a sustained interferon signature, DNA damage, and cytokine secretion

PubMed Central

Kandhaya-Pillai, Renuka; Miro-Mur, Francesc; Alijotas-Reig, Jaume; Tchkonia, Tamara; Kirkland, James L.; Schwartz, Simo

2017-01-01

Cellular senescence is a cell fate program that entails essentially irreversible proliferative arrest in response to damage signals. Tumor necrosis factor-alpha (TNFα), an important pro-inflammatory cytokine secreted by some types of senescent cells, can induce senescence in mouse and human cells. However, downstream signaling pathways linking TNFα-related inflammation to senescence are not fully characterized. Using human umbilical vein endothelial cells (HUVECs) as a model, we show that TNFα induces permanent growth arrest and increases p21CIP1, p16INK4A, and SA-β-gal, accompanied by persistent DNA damage and ROS production. By gene expression profiling, we identified the crucial involvement of inflammatory and JAK/STAT pathways in TNFα-mediated senescence. We found that TNFα activates a STAT-dependent autocrine loop that sustains cytokine secretion and an interferon signature to lock cells into senescence. Furthermore, we show STAT1/3 activation is necessary for cytokine and ROS production during TNFα-induced senescence. However, inhibition of STAT1/3 did not rescue cells from proliferative arrest, but rather suppressed cell cycle regulatory genes and altered TNFα-induced senescence. Our findings suggest a positive feedback mechanism via the STAT pathway that sustains cytokine production and reveal a reciprocal regulatory role of JAK/STAT in TNFα-mediated senescence. PMID:29176033

Epigenetic regulation of vascular smooth muscle cell proliferation and neointima formation by histone deacetylase inhibition.

PubMed

Findeisen, Hannes M; Gizard, Florence; Zhao, Yue; Qing, Hua; Heywood, Elizabeth B; Jones, Karrie L; Cohn, Dianne; Bruemmer, Dennis

2011-04-01

Proliferation of smooth muscle cells (SMC) in response to vascular injury is central to neointimal vascular remodeling. There is accumulating evidence that histone acetylation constitutes a major epigenetic modification for the transcriptional control of proliferative gene expression; however, the physiological role of histone acetylation for proliferative vascular disease remains elusive. In the present study, we investigated the role of histone deacetylase (HDAC) inhibition in SMC proliferation and neointimal remodeling. We demonstrate that mitogens induce transcription of HDAC 1, 2, and 3 in SMC. Short interfering RNA-mediated knockdown of either HDAC 1, 2, or 3 and pharmacological inhibition of HDAC prevented mitogen-induced SMC proliferation. The mechanisms underlying this reduction of SMC proliferation by HDAC inhibition involve a growth arrest in the G(1) phase of the cell cycle that is due to an inhibition of retinoblastoma protein phosphorylation. HDAC inhibition resulted in a transcriptional and posttranscriptional regulation of the cyclin-dependent kinase inhibitors p21(Cip1) and p27(Kip). Furthermore, HDAC inhibition repressed mitogen-induced cyclin D1 mRNA expression and cyclin D1 promoter activity. As a result of this differential cell cycle-regulatory gene expression by HDAC inhibition, the retinoblastoma protein retains a transcriptional repression of its downstream target genes required for S phase entry. Finally, we provide evidence that these observations are applicable in vivo by demonstrating that HDAC inhibition decreased neointima formation and expression of cyclin D1 in a murine model of vascular injury. These findings identify HDAC as a critical component of a transcriptional cascade regulating SMC proliferation and suggest that HDAC might play a pivotal role in the development of proliferative vascular diseases, including atherosclerosis and in-stent restenosis.

Epigenetic Regulation of Vascular Smooth Muscle Cell Proliferation and Neointima Formation by Histone Deacetylase Inhibition

PubMed Central

Findeisen, Hannes M.; Gizard, Florence; Zhao, Yue; Qing, Hua; Heywood, Elizabeth B.; Jones, Karrie L.; Cohn, Dianne; Bruemmer, Dennis

2011-01-01

Objective Proliferation of smooth muscle cells (SMC) in response to vascular injury is central to neointimal vascular remodeling. There is accumulating evidence that histone acetylation constitutes a major epigenetic modification for the transcriptional control of proliferative gene expression; however, the physiological role of histone acetylation for proliferative vascular disease remains elusive. Methods and Results In the present study, we investigated the role of histone deacetylase (HDAC) inhibition in SMC proliferation and neointimal remodeling. We demonstrate that mitogens induce transcription of HDAC 1, 2 and 3 in SMC. siRNA-mediated knock-down of either HDAC 1, 2 or 3 and pharmacologic inhibition of HDAC prevented mitogen-induced SMC proliferation. The mechanisms underlying this reduction of SMC proliferation by HDAC inhibition involve a growth arrest in the G1-phase of the cell cycle due to an inhibition of retinoblastoma protein phosphorylation. HDAC inhibition resulted in a transcriptional and posttranscriptional regulation of the cyclin-dependent kinase inhibitors p21Cip1 and p27Kip. Furthermore, HDAC inhibition repressed mitogen-induced cyclin D1 mRNA expression and cyclin D1 promoter activity. As a result of this differential cell cycle-regulatory gene expression by HDAC inhibition, the retinoblastoma protein retains a transcriptional repression of its downstream target genes required for S phase entry. Finally, we provide evidence that these observations are applicable in vivo by demonstrating that HDAC inhibition decreased neointima formation and expression of cyclin D1 in a murine model of vascular injury. Conclusion These findings identify HDAC as a critical component of a transcriptional cascade regulating SMC proliferation and suggest that HDAC might play a pivotal role in the development of proliferative vascular diseases, including atherosclerosis and in-stent restenosis. PMID:21233448

Cytokine-mediated induction of endothelial adhesion molecule and histocompatibility leukocyte antigen expression by cytomegalovirus-activated T cells.

PubMed Central

Waldman, W. J.; Knight, D. A.

1996-01-01

Cytomegalovirus (CMV) has been associated with allograft rejection and transplantation-associated arteriosclerosis. CMV infects endothelium, the interface between allograft tissue and the host immune system; however, mechanisms by which such interaction might exacerbate the rejection process remain unresolved. Here we test the hypothesis that host immune activity, triggered by CMV-infected graft endothelial cells (ECs), can result in the production of cytokines capable of enhancing the alloimmunogenicity of nearby uninfected endothelia. To model these phenomena in vitro, confluent monolayers of ECs derived from human umbilical vein or adult gonadal vein were incubated 5 days beneath trans-well culture inserts containing CMV-seropositive or CMV-seronegative donor-derived CD3+ or CD4+ T cells alone or in combination with CMV-infected or uninfected allogeneic ECs. The extent of T cell proliferation was determined by [3H]thymidine labeling of trans-well contents after transfer to microtiter plates. Endothelial responses to soluble factors elaborated by CMV-activated T cells were determined by immunohistochemical staining and immunofluorescence flow cytometric analysis of underlying EC monolayers. Results of experiments with CMV-seropositive donor-derived CD4+ T cells demonstrated enhancement of ICAM-1 and histocompatibility leukocyte antigen class I, as well as induction of histocompatibility leukocyte antigen DR on ECs incubated beneath T cell/EC/CMV trans-well co-cultures. Total (CD3+) T cells co-cultured with EC/CMV induced VCAM-1 as well. Furthermore, [3H]thymidine incorporation by these T cells indicated a strong proliferative response. Endothelial responses to T cells alone or in combination with uninfected ECs were minimal, and T cells cultured under these conditions showed little proliferative activity. Similarly, little or no endothelial responses were apparent in monolayers beneath trans-wells containing T cells isolated from CMV-seronegative individuals regardless of the CMV status of stimulator ECs. Finally, experiments employing blocking antibodies identified interferon-gamma and tumor necrosis factor-alpha as inducing agents in this co-culture system. These findings suggest that allograft endothelium harboring CMV has the potential to activate host T cells and that the consequent release of cytokines shows potential to raise surrounding endothelia to a fully activated, highly immunogenic state. Results of these studies thus provide insight into mechanisms that help elucidate the association between CMV and transplantation-associated arteriosclerosis and/or allograft rejection. Images Figure 1 Figure 5 PMID:8546198

Uric acid stimulates proliferative pathways in vascular smooth muscle cells through the activation of p38 MAPK, p44/42 MAPK and PDGFRβ.

PubMed

Kırça, M; Oğuz, N; Çetin, A; Uzuner, F; Yeşilkaya, A

2017-04-01

Hyperuricemia and angiotensin II (Ang II) may have a pathogenetic role in the development of hypertension and atherosclerosis as well as cardiovascular disease (CVD) and its prognosis. The purpose of this study was to investigate whether uric acid can induce proliferative pathways of vascular smooth muscle cell (VSMC) that are thought to be responsible for the development of CVD. The phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), p44/42 mitogen-activated protein kinase (p44/42 MAPK) and platelet-derived growth factor receptor β (PDGFRβ) was measured by Elisa and Western blot techniques to determine the activation of proliferative pathways in primary cultured VSMCs from rat aorta. Results demonstrated that uric acid can stimulate p38 MAPK, p44/42 MAPK and PDGFRβ phosphorylation in a time- and concentration-dependent manner. Furthermore, treatment of VSMCs with the angiotensin II type I receptor (AT1R) inhibitor losartan suppressed p38 MAPK and p44/42 MAPK induction by uric acid. The stimulatory effect of uric acid on p38 MAPK was higher compared to that of Ang II. The results of this study show for the first time that uric acid-induced PDGFRβ phosphorylation plays a crucial role in the development of CVDs and that elevated uric acid levels could be a potential therapeutical target in CVD patients.

Skeletal muscle satellite cells

NASA Technical Reports Server (NTRS)

Schultz, E.; McCormick, K. M.

1994-01-01

Evidence now suggests that satellite cells constitute a class of myogenic cells that differ distinctly from other embryonic myoblasts. Satellite cells arise from somites and first appear as a distinct myoblast type well before birth. Satellite cells from different muscles cannot be functionally distinguished from one another and are able to provide nuclei to all fibers without regard to phenotype. Thus, it is difficult to ascribe any significant function to establishing or stabilizing fiber type, even during regeneration. Within a muscle, satellite cells exhibit marked heterogeneity with respect to their proliferative behavior. The satellite cell population on a fiber can be partitioned into those that function as stem cells and those which are readily available for fusion. Recent studies have shown that the cells are not simply spindle shaped, but are very diverse in their morphology and have multiple branches emanating from the poles of the cells. This finding is consistent with other studies indicating that the cells have the capacity for extensive migration within, and perhaps between, muscles. Complexity of cell shape usually reflects increased cytoplasmic volume and organelles including a well developed Golgi, and is usually associated with growing postnatal muscle or muscles undergoing some form of induced adaptive change or repair. The appearance of activated satellite cells suggests some function of the cells in the adaptive process through elaboration and secretion of a product. Significant advances have been made in determining the potential secretion products that satellite cells make. The manner in which satellite cell proliferative and fusion behavior is controlled has also been studied. There seems to be little doubt that cellcell coupling is not how satellite cells and myofibers communicate. Rather satellite cell regulation is through a number of potential growth factors that arise from a number of sources. Critical to the understanding of this form of control is to determine which of the many growth factors that can alter satellite cell behavior in vitro are at work in vivo. Little work has been done to determine what controls are at work after a regeneration response has been initiated. It seems likely that, after injury, growth factors are liberated through proteolytic activity and initiate an activation process whereby cells enter into a proliferative phase. After myofibers are formed, it also seems likely that satellite cell behavior is regulated through diffusible factors arising from the fibers rather than continuous control by circulating factors.(ABSTRACT TRUNCATED AT 400 WORDS).

Aqueous exposure to Aroclor 1254 modulates the mitogenic response of Atlantic salmon anterior kidney T-cells: indications of short- and long-term immunomodulation.

PubMed

Iwanowicz, Luke R; Lerner, Darren T; Blazer, Vicki S; McCormick, Stephen D

2005-05-15

Polychlorinated biphenyls (PCBs) exist as persistent organic pollutants in numerous river systems in the United States. Unfortunately, some of these rivers are sites of active Atlantic salmon restoration programs, and polychlorinated biphenyls have been implicated as ancillary factors contributing to failed salmon restoration. Here, we investigate the immediate and chronic effects of intermediate duration aqueous PCB exposure (1 or 10 microgL-1 Aroclor 1254) on the mitogen-stimulated lymphoproliferative response of Atlantic salmon anterior kidney leukocytes (AKLs). A short-term study was designed to examine immunomodulation in Atlantic salmon smolts immediately following 21 days of aqueous exposure, while a long-term study evaluated chronic impacts in the mitogen response in parr 15 months post-exposure as larvae. The proliferative response of AKLs to the mitogens concanavalin A (CON A), phytohemaglutinnin-P (PHA-P), pokeweed mitogen (PWM), and lipopolysaccharide were used as an indice of immunomodulation. The proliferative response to the T-cell mitogens CON A and PHA-P was significantly increased in the 10 microgL-1 group (n=10; P=0.043 and 0.002, respectively) immediately following exposure of smolts. Additionally, The PHA-P response was significantly increased in the 1 microgL-1 exposure group (n=10, P=0.036). In fish treated as larvae and tested 15 months later, the PHA-P sensitive populations exhibited elevated proliferation in the 1 and 10 microgL-1 groups (n=12, P<0.04) relative to the vehicle control while the PWM response was significantly increased (n=12, P=0.036) only in the 10 microgL-1 treated groups. These results demonstrate an immunomodulatory effect of PCBs on T-cell mitogen sensitive populations of lymphocytes in Atlantic salmon as well as long-term immunomodulation in PHA-P and PWM sensitive populations.

Aqueous exposure to Aroclor 1254 modulates the mitogenic response of Atlantic salmon anterior kidney T-cells: Indications of short- and long-term immunomodulation

USGS Publications Warehouse

Iwanowicz, L.R.; Lerner, D.T.; Blazer, V.S.; McCormick, S.D.

2005-01-01

Polychlorinated biphenyls (PCBs) exist as persistent organic pollutants in numerous river systems in the United States. Unfortunately, some of these rivers are sites of active Atlantic salmon restoration programs, and polychlorinated biphenyls have been implicated as ancillary factors contributing to failed salmon restoration. Here, we investigate the immediate and chronic effects of intermediate duration aqueous PCB exposure (1 or 10 ??g L-1 Aroclor 1254) on the mitogen-stimulated lymphoproliferative response of Atlantic salmon anterior kidney leukocytes (AKLs). A short-term study was designed to examine immunomodulation in Atlantic salmon smolts immediately following 21 days of aqueous exposure, while a long-term study evaluated chronic impacts in the mitogen response in parr 15 months post-exposure as larvae. The proliferative response of AKLs to the mitogens concanavalin A (CON A), phytohemaglutinnin-P (PHA-P), pokeweed mitogen (PWM), and lipopolysaccharide were used as an indice of immunomodulation. The proliferative response to the T-cell mitogens CON A and PHA-P was significantly increased in the 10 ??g L-1 group (n = 10; P = 0.043 and 0.002, respectively) immediately following exposure of smolts. Additionally, The PHA-P response was significantly increased in the 1 ??g L-1 exposure group (n = 10, P = 0.036). In fish treated as larvae and tested 15 months later, the PHA-P sensitive populations exhibited elevated proliferation in the 1 and 10 ??g L-1 groups (n = 12, P < 0.04) relative to the vehicle control while the PWM response was significantly increased (n = 12, P = 0.036) only in the 10 ??g L-1 treated groups. These results demonstrate an immunomodulatory effect of PCBs on T-cell mitogen sensitive populations of lymphocytes in Atlantic salmon as well as long-term immunomodulation in PHA-P and PWM sensitive populations. ?? 2005 Elsevier B.V. All rights reserved.

Cancer metabolism, stemness and tumor recurrence

PubMed Central

Curry, Joseph M.; Tuluc, Madalina; Whitaker-Menezes, Diana; Ames, Julie A.; Anantharaman, Archana; Butera, Aileen; Leiby, Benjamin; Cognetti, David M.; Sotgia, Federica; Lisanti, Michael P.; Martinez-Outschoorn, Ubaldo E.

2013-01-01

Here, we interrogated head and neck cancer (HNSCC) specimens (n = 12) to examine if different metabolic compartments (oxidative vs. glycolytic) co-exist in human tumors. A large panel of well-established biomarkers was employed to determine the metabolic state of proliferative cancer cells. Interestingly, cell proliferation in cancer cells, as marked by Ki-67 immunostaining, was strictly correlated with oxidative mitochondrial metabolism (OXPHOS) and the uptake of mitochondrial fuels, as detected via MCT1 expression (p < 0.001). More specifically, three metabolic tumor compartments were delineated: (1) proliferative and mitochondrial-rich cancer cells (Ki-67+/TOMM20+/COX+/MCT1+); (2) non-proliferative and mitochondrial-poor cancer cells (Ki-67−/TOMM20−/COX−/MCT1−); and (3) non-proliferative and mitochondrial-poor stromal cells (Ki-67−/TOMM20−/COX−/MCT1−). In addition, high oxidative stress (MCT4+) was very specific for cancer tissues. Thus, we next evaluated the prognostic value of MCT4 in a second independent patient cohort (n = 40). Most importantly, oxidative stress (MCT4+) in non-proliferating epithelial cancer cells predicted poor clinical outcome (tumor recurrence; p < 0.0001; log-rank test), and was functionally associated with FDG-PET avidity (p < 0.04). Similarly, oxidative stress (MCT4+) in tumor stromal cells was specifically associated with higher tumor stage (p < 0.03), and was a highly specific marker for cancer-associated fibroblasts (p < 0.001). We propose that oxidative stress is a key hallmark of tumor tissues that drives high-energy metabolism in adjacent proliferating mitochondrial-rich cancer cells, via the paracrine transfer of mitochondrial fuels (such as L-lactate and ketone bodies). New antioxidants and MCT4 inhibitors should be developed to metabolically target “three-compartment tumor metabolism” in head and neck cancers. It is remarkable that two “non-proliferating” populations of cells (Ki-67−/MCT4+) within the tumor can actually determine clinical outcome, likely by providing high-energy mitochondrial “fuels” for proliferative cancer cells to burn. Finally, we also show that in normal mucosal tissue, the basal epithelial “stem cell” layer is hyper-proliferative (Ki-67+), mitochondrial-rich (TOMM20+/COX+) and is metabolically programmed to use mitochondrial fuels (MCT1+), such as ketone bodies and L-lactate. Thus, oxidative mitochondrial metabolism (OXPHOS) is a common feature of both (1) normal stem cells and (2) proliferating cancer cells. As such, we should consider metabolically treating cancer patients with mitochondrial inhibitors (such as Metformin), and/or with a combination of MCT1 and MCT4 inhibitors, to target “metabolic symbiosis.” PMID:23574725

Transcriptional Regulation of S Phase Kinase-associated Protein 2 by NR4A Orphan Nuclear Receptor NOR1 in Vascular Smooth Muscle Cells*

PubMed Central

Gizard, Florence; Zhao, Yue; Findeisen, Hannes M.; Qing, Hua; Cohn, Dianne; Heywood, Elizabeth B.; Jones, Karrie L.; Nomiyama, Takashi; Bruemmer, Dennis

2011-01-01

Members of the NR4A subgroup of the nuclear hormone receptor superfamily have emerged as key transcriptional regulators of proliferation and inflammation. NOR1 constitutes a ligand-independent transcription factor of this subgroup and induces cell proliferation; however, the transcriptional mechanisms underlying this mitogenic role remain to be defined. Here, we demonstrate that the F-box protein SKP2 (S phase kinase-associated protein 2), the substrate-specific receptor of the ubiquitin ligase responsible for the degradation of p27KIP1 through the proteasome pathway, constitutes a direct transcriptional target for NOR1. Mitogen-induced Skp2 expression is silenced in vascular smooth muscle cells (VSMC) isolated from Nor1-deficient mice or transfected with Nor1 siRNA. Conversely, adenovirus-mediated overexpression of NOR1 induces Skp2 expression in VSMC and decreases protein abundance of its target p27. Transient transfection experiments establish that NOR1 transactivates the Skp2 promoter through a nerve growth factor-induced clone B response element (NBRE). Electrophoretic mobility shift and chromatin immunoprecipitation assays further revealed that NOR1 is recruited to this NBRE site in the Skp2 promoter in response to mitogenic stimulation. In vivo Skp2 expression is increased during the proliferative response underlying neointima formation, and this transcriptional induction depends on the expression of NOR1. Finally, we demonstrate that overexpression of Skp2 rescues the proliferative arrest of Nor1-deficient VSMC. Collectively, these results characterize Skp2 as a novel NOR1-regulated target gene and detail a previously unrecognized transcriptional cascade regulating mitogen-induced VSMC proliferation. PMID:21868379

Mechanical Tension Increases CCN2/CTGF Expression and Proliferation in Gingival Fibroblasts via a TGFβ-Dependent Mechanism

PubMed Central

Guo, Fen; Carter, David E.; Leask, Andrew

2011-01-01

Unlike skin, oral gingival do not scar in response to tissue injury. Fibroblasts, the cell type responsible for connective tissue repair and scarring, are exposed to mechanical tension during normal and pathological conditions including wound healing and fibrogenesis. Understanding how human gingival fibroblasts respond to mechanical tension is likely to yield valuable insights not only into gingival function but also into the molecular basis of scarless repair. CCN2/connective tissue growth factor is potently induced in fibroblasts during tissue repair and fibrogenesis. We subjected gingival fibroblasts to cyclical strain (up to 72 hours) using the Flexercell system and showed that CCN2 mRNA and protein was induced by strain. Strain caused the rapid activation of latent TGFβ, in a fashion that was reduced by blebbistatin and FAK/src inhibition, and the induction of endothelin (ET-1) mRNA and protein expression. Strain did not cause induction of α-smooth muscle actin or collagen type I mRNAs (proteins promoting scarring); but induced a cohort of pro-proliferative mRNAs and cell proliferation. Compared to dermal fibroblasts, gingival fibroblasts showed reduced ability to respond to TGFβ by inducing fibrogenic mRNAs; addition of ET-1 rescued this phenotype. Pharmacological inhibition of the TGFβ type I (ALK5) receptor, the endothelin A/B receptors and FAK/src significantly reduced the induction of CCN2 and pro-proliferative mRNAs and cell proliferation. Controlling TGFβ, ET-1 and FAK/src activity may be useful in controlling responses to mechanical strain in the gingiva and may be of value in controlling fibroproliferative conditions such as gingival hyperplasia; controlling ET-1 may be of benefit in controlling scarring in response to injury in the skin. PMID:21611193

An APC:WNT Counter-Current-Like Mechanism Regulates Cell Division Along the Human Colonic Crypt Axis: A Mechanism That Explains How APC Mutations Induce Proliferative Abnormalities That Drive Colon Cancer Development

PubMed Central

Boman, Bruce M.; Fields, Jeremy Z.

2013-01-01

APC normally down-regulates WNT signaling in human colon, and APC mutations cause proliferative abnormalities in premalignant crypts leading to colon cancer, but the mechanisms are unclear at the level of spatial and functional organization of the crypt. Accordingly, we postulated a counter-current-like mechanism based on gradients of factors (APC;WNT) that regulate colonocyte proliferation along the crypt axis. During crypt renewal, stem cells (SCs) at the crypt bottom generate non-SC daughter cells that proliferate and differentiate while migrating upwards. The APC concentration is low at the crypt bottom and high at the top (where differentiated cells reside). WNT signaling, in contrast, is high at the bottom (where SCs reside) and low at the top. Given that WNT and APC gradients are counter to one another, we hypothesized that a counter-current-like mechanism exists. Since both APC and WNT signaling components (e.g., survivin) are required for mitosis, this mechanism establishes a zone in the lower crypt where conditions are optimal for maximal cell division and mitosis orientation (symmetric versus asymmetric). APC haploinsufficiency diminishes the APC gradient, shifts the proliferative zone upwards, and increases symmetric division, which causes SC overpopulation. In homozygote mutant crypts, these changes are exacerbated. Thus, APC-mutation-induced changes in the counter-current-like mechanism cause expansion of proliferative populations (SCs, rapidly proliferating cells) during tumorigenesis. We propose this mechanism also drives crypt fission, functions in the crypt cycle, and underlies adenoma development. Novel chemoprevention approaches designed to normalize the two gradients and readjust the proliferative zone downwards, might thwart progression of these premalignant changes. PMID:24224156

The novel HDAC inhibitor AR-42-induced anti-colon cancer cell activity is associated with ceramide production

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

Xu, Weihong; Xu, Bin; Yao, Yiting

In the current study, we investigated the potential activity of AR-42, a novel histone deacetylase (HDAC) inhibitor, against colon cancer cells. Our in vitro results showed that AR-42 induced ceramide production, exerted potent anti-proliferative and pro-apoptotic activities in established (SW-620 and HCT-116 lines) and primary human colon cancer cells. Exogenously-added sphingosine 1-phosphate (S1P) suppressed AR-42-induced activity, yet a cell-permeable ceramide (C4) facilitated AR-42-induced cytotoxicity against colon cancer cells. In addition, AR-42-induced ceramide production and anti-colon cancer cell activity were inhibited by the ceramide synthase inhibitor fumonisin B1, but were exacerbated by PDMP, which is a ceramide glucosylation inhibitor. In vivo, oral administrationmore » of a single dose of AR-42 dramatically inhibited SW-620 xenograft growth in severe combined immunodeficient (SCID) mice, without inducing overt toxicities. Together, these results show that AR-42 dramatically inhibits colon cancer cell proliferation in vitro and in vivo, and ceramide production might be the key mechanism responsible for its actions. - Highlights: • AR-42 is anti-proliferative against primary/established colon cancer cells. • AR-42 induces significant apoptotic death in primary/established colon cancer cells. • Ceramide production mediates AR-42-induced cytotoxicity in colon cancer cells. • AR-42 oral administration potently inhibits SW-620 xenograft growth in SCID mice.« less

Design, synthesis, and biological evaluation of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetate derivatives as anti-proliferative agents through ROS-induced cell apoptosis.

PubMed

Song, Zhuang; Chen, Cai-Ping; Liu, Jun; Wen, Xiaoan; Sun, Hongbin; Yuan, Haoliang

2016-11-29

A novel class of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetate derivatives were designed and synthesized as potent anti-proliferative agents. Most of these compounds showed potent anti-proliferative activity against some tumor cell lines, including SK-BR-3, MDA-MB-231, HCT-116, SW480, Ovcar-3, HL-60, Saos-2 and HepG2. Compounds 8c and 11h were identified as the most potent ones, while HL-60, HCT116 and MDA-MB-231 were the most sensitive cell lines. Mechanistic study revealed that compound 8c enhanced reactive oxygen species level by inhibiting TrxR and then induced apoptosis by activating apoptosis proteins, bax and cleaved-caspase 3 in HCT116 cells. Preliminary SAR analysis indicated that modifications of the double bond and ester group made great effects on the anti-proliferative activity. Our findings suggested that it was worth further studies on the antitumor potency of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetates. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Proliferative human cell sources applied as biocomponent in bioartificial livers: a review.

PubMed

Nibourg, Geert A A; Chamuleau, Robert A F M; van Gulik, Thomas M; Hoekstra, Ruurdtje

2012-07-01

Bioartificial livers (BALs) are urgently needed to bridge severe liver failure patients to liver transplantation or liver regeneration. When based on primary hepatocytes, their efficacy has been shown in animal experiments and their safety was confirmed in clinical trials. However, a proliferative human cell source with therapeutic functionality is needed to secure availability and move BAL application forward. This review compares the performance of BALs based on proliferative human biocomponents and primary hepatocytes. This review evaluates relevant studies identified by searching the MEDLINE database until July 2011 and some of our own unpublished data. All the discussed hepatocyte-like biocomponents show deficiencies in their hepatic functionality compared with primary hepatocytes, particularly functions occurring late in liver development. Nonetheless, the HepaRG, HepG2-GS-CYP3A4, and mesenchymal stem cells show efficacy in a statistically well-powered animal model of acute liver failure, when applied in a BAL device. Various methods to gain higher functionality of BALs, including genetic modification, the usage of combinatory cell sources, and improvement of culture methods, have scarcely been applied, but may further pave the path for BAL application. Clinical implementation of a BAL based on a human proliferative biocomponent is still several years away.

Lung Inflammation, Injury, and Proliferative Response after Repetitive Particulate Hexavalent Chromium Exposure

PubMed Central

Beaver, Laura M.; Stemmy, Erik J.; Schwartz, Arnold M.; Damsker, Jesse M.; Constant, Stephanie L.; Ceryak, Susan M.; Patierno, Steven R.

2009-01-01

Background Chronic inflammation is implicated in the development of several human cancers, including lung cancer. Certain particulate hexavalent chromium [Cr(VI)] compounds are well-documented human respiratory carcinogens that release genotoxic soluble chromate and are associated with fibrosis, fibrosarcomas, adenocarcinomas, and squamous cell carcinomas of the lung. Despite this, little is known about the pathologic injury and immune responses after repetitive exposure to particulate chromates. Objectives In this study we investigated the lung injury, inflammation, proliferation, and survival signaling responses after repetitive exposure to particulate chromate. Methods BALB/c mice were repetitively treated with particulate basic zinc chromate or saline using an intranasal exposure regimen. We assessed lungs for Cr(VI)-induced changes by bronchoalveolar lavage, histologic examination, and immunohistochemistry. Results Single exposure to Cr(VI) resulted in inflammation of lung tissue that persists for up to 21 days. Repetitive Cr(VI) exposure induced a neutrophilic inflammatory airway response 24 hr after each treatment. Neutrophils were subsequently replaced by increasing numbers of macrophages by 5 days after treatment. Repetitive Cr(VI) exposure induced chronic peribronchial inflammation with alveolar and interstitial pneumonitis dominated by lymphocytes and macrophages. Moreover, chronic toxic mucosal injury was observed and accompanied by increased airway pro-matrix metalloprotease-9. Injury and inflammation correlated with airways becoming immunoreactive for phosphorylation of the survival signaling protein Akt and the proliferation marker Ki-67. We observed a reactive proliferative response in epithelial cells lining airways of chromate-exposed animals. Conclusions These data illustrate that repetitive exposure to particulate chromate induces chronic injury and an inflammatory microenvironment that may promote Cr(VI) carcinogenesis. PMID:20049209

Developmental control of transcriptional and proliferative potency during the evolutionary emergence of animals

PubMed Central

Arenas-Mena, Cesar; Coffman, James A.

2016-01-01

Summary It is proposed that the evolution of complex animals required repressive genetic mechanisms for controlling the transcriptional and proliferative potency of cells. Unicellular organisms are transcriptionally potent, able to express their full genetic complement as the need arises through their life cycle, whereas differentiated cells of multicellular organisms can only express a fraction of their genomic potential. Likewise, whereas cell proliferation in unicellular organisms is primarily limited by nutrient availability, cell proliferation in multicellular organisms is developmentally regulated. Repressive genetic controls limiting the potency of cells at the end of ontogeny would have stabilized the gene expression states of differentiated cells and prevented disruptive proliferation, allowing the emergence of diverse cell types and functional shapes. We propose that distal cis-regulatory elements represent the primary innovations that set the stage for the evolution of developmental gene regulatory networks and the repressive control of key multipotency and cell-cycle control genes. The testable prediction of this model is that the genomes of extant animals, unlike those of our unicellular relatives, encode gene regulatory circuits dedicated to the developmental control of transcriptional and proliferative potency. PMID:26173445

H3K4 demethylase activities repress proliferative and postmitotic aging

PubMed Central

Alvares, Stacy M; Mayberry, Gaea A; Joyner, Ebony Y; Lakowski, Bernard; Ahmed, Shawn

2014-01-01

Homeostasis of postmitotic and proliferating cells is maintained by pathways that repress stress. We found that the Caenorhabditis elegans histone 3 lysine 4 (H3K4) demethylases RBR-2 and SPR-5 promoted postmitotic longevity of stress-resistant daf-2 adults, altered pools of methylated H3K4, and promoted silencing of some daf-2 target genes. In addition, RBR-2 and SPR-5 were required for germ cell immortality at a high temperature. Transgenerational proliferative aging was enhanced for spr-5; rbr-2 double mutants, suggesting that these histone demethylases may function sequentially to promote germ cell immortality by targeting distinct H3K4 methyl marks. RBR-2 did not play a comparable role in the maintenance of quiescent germ cells in dauer larvae, implying that it represses stress that occurs as a consequence of germ cell proliferation, rather than stress that accumulates in nondividing cells. We propose that H3K4 demethylase activities promote the maintenance of chromatin states during stressful growth conditions, thereby repressing postmitotic aging of somatic cells as well as proliferative aging of germ cells. PMID:24134677

Triterpenoid Saponins from Anemone rivularis var. Flore-Minore and Their Anti-Proliferative Activity on HSC-T6 Cells.

PubMed

Wang, Xiao-Yang; Gao, Hui; Xie, Xiao-Jie; Jurhiin, Jirimubatu; Zhang, Mu-Zi-He; Zhou, Yan-Ping; Liu, Rui; Ning, Meng; Han, Jin; Tang, Hai-Feng

2018-02-23

Five previously undescribed triterpenoid saponins ( 1 - 5 ), along with eight known ones ( 6 - 13 ), were isolated from the whole plants of Anemone rivularis var. flore-minore . Their structures were clarified by extensive spectroscopic data and chemical evidence. For the first time, the lupane-type saponins ( 3 and 12 ) were reported from the Anemone genus. The anti-proliferative activity of all isolated saponins was evaluated on hepatic stellate cells (HSC-T6). Saponins 12 and 13 , which possess more monosaccharides than the others, displayed potent anti-proliferative activity, with IC 50 values of 18.21 and 15.56 μM, respectively.

Immunoregulation by Taenia crassiceps and its antigens.

PubMed

Peón, Alberto N; Espinoza-Jiménez, Arlett; Terrazas, Luis I

2013-01-01

Taenia crassiceps is a cestode parasite of rodents (in its larval stage) and canids (in its adult stage) that can also parasitize immunocompromised humans. We have studied the immune response elicited by this helminth and its antigens in mice and human cells, and have discovered that they have a strong capacity to induce chronic Th2-type responses that are primarily characterized by high levels of Th2 cytokines, low proliferative responses in lymphocytes, an immature and LPS-tolerogenic profile in dendritic cells, the recruitment of myeloid-derived suppressor cells and, specially, alternatively activated macrophages. We also have utilized the immunoregulatory capabilities of this helminth to successfully modulate autoimmune responses and the outcome of other infectious diseases. In the present paper, we review the work of others and ourselves with regard to the immune response induced by T. crassiceps and its antigens, and we compare the advances in our understanding of this parasitic infection model with the knowledge that has been obtained from other selected models.

Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT, NFκB and ERK

PubMed Central

Badr, Gamal; Sayed, Ayat; Abdel-Maksoud, Mostafa A.; Mohamed, Amany O.; El-Amir, Azza; Abdel-Ghaffar, Fathy A.; Al-Quraishy, Saleh; Mahmoud, Mohamed H.

2015-01-01

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity. PMID:25909640

Possible mechanisms for arsenic-induced proliferative diseases

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

Wetterhahn, K.E.; Dudek, E.J.; Shumilla, J.A.

1996-12-31

Possible mechanisms for cardiovascular diseases and cancers which have been observed on chronic exposure to arsenic have been investigated. We tested the hypothesis that nonlethal levels of arsenic are mitogenic, cause oxidative stress, increase nuclear translocation of trans-acting factors, and increase expression of genes involved in proliferation. Cultured porcine vascular (from aorta) endothelial cells were used as a model cell system to study the effects of arsenic on the target cells for cardiovascular diseases. Treatment of postconfluent cell cultures with nonovertly toxic concentrations of arsenite increased DNA synthesis, similar to the mitogenic response observed with hydrogen peroxide. Within 1 hourmore » of adding noncytotoxic concentrations of arsenite, cellular levels of oxidants increased relative to control levels, indicating that arsenite promotes cellular oxidations. Arsenite treatment increased nuclear translocation of NF-{kappa}B, an oxidative stress-responsive transcription factor, in a manner similar to that observed with hydrogen peroxide. Pretreatment of intact cells with the antioxidants N-acetylcysteine and dimethylfumarate prevented the arsenite-induced increases in cellular oxidant formation and NF-KB translocation. Arsenite had little or no effect on binding of NF-KB to its DNA recognition sequence in vitro, indicating that it is unlikely that arsenite directly affects NF-KB. The steady-state mRNA levels of intracellular adhesion molecule and urokinase-like plasminogen activator, genes associated with the active endothelial phenotype in arteriosclerosis and cancer metastasis, were increased by nontoxic concentrations of arsenite. These data suggest that arsenite promotes proliferative diseases like heart disease and cancer by activating oxidant-sensitive endothelial cell signaling and gene expression. It is possible that antioxidant therapy would be useful in preventing arsenic-induced cardiovascular disease and cancer.« less

Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT, NFκB and ERK.

PubMed

Badr, Gamal; Sayed, Ayat; Abdel-Maksoud, Mostafa A; Mohamed, Amany O; El-Amir, Azza; Abdel-Ghaffar, Fathy A; Al-Quraishy, Saleh; Mahmoud, Mohamed H

2015-01-01

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity.

Substance P prevents development of proliferative vitreoretinopathy in mice by modulating TNF-α

PubMed Central

Yoo, Kyungsang; Son, Bo Kwon; Kim, Suna; Son, Youngsook; Yu, Seung-Young

2017-01-01

Purpose Proliferative vitreoretinopathy (PVR) is an inflammatory fibrotic disease resulting from the inflammatory milieu after retinal detachment, which can prevent retinal healing. This study aimed to elucidate the effect of substance P (SP) on retinal degeneration caused by retinal detachment in vivo and to examine the role of SP in the tumor necrosis factor-alpha (TNF-α)-induced epithelial-mesenchymal transition (EMT) of human RPE cells in vitro. Methods PVR-like retinal damage was induced by intravitreally injecting dispase into mice, and SP was systemically injected twice a week for 3 weeks. Histological analysis and cytokine profile with enzyme-linked immunosorbent assay (ELISA) were performed. The direct effect of SP on induction of EMT in vitro was studied by adding SP to TNF-α-treated ARPE-19 cells and then evaluating the change in the characteristics of the epithelial and mesenchymal cells. Results Dispase injection led to a PVR-like retinal condition, demonstrating an inflammatory response with disruption of RPE interaction within 1 week and severe destruction with enfolding within 3 weeks after the dispase injection. The inflammatory environment promoted apoptosis and migration of fibroblast-like cells in the retinal layer, which can cause fibrotic disease, such as PVR. However, SP treatment suppressed early inflammatory responses by reducing TNF-α and elevating interleukin-10 (IL-10), with cell death and the appearance of fibroblastic cells inhibited and the progression of retinal degeneration obviously delayed. Moreover, SP ameliorated TNF-α-induced EMT of the RPE and directly prevented fibrotic change in the RPE. Conclusions This study revealed that SP can block apoptosis and EMT due to retinal inflammation and inhibit the development of PVR. This effect most likely occurred by modulating the secretion and action of TNF-α.. PMID:29296073

Regulatory function of cytomegalovirus-specific CD4{sup +}CD27{sup -}CD28{sup -} T cells

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

Tovar-Salazar, Adriana; Patterson-Bartlett, Julie; Jesser, Renee

2010-03-15

CMV infection is characterized by high of frequencies of CD27{sup -}CD28{sup -} T cells. Here we demonstrate that CMV-specific CD4{sup +}CD27{sup -}CD28{sup -} cells are regulatory T cells (T{sub R}). CD4{sup +}CD27{sup -}CD28{sup -} cells sorted from CMV-stimulated PBMC of CMV-seropositive donors inhibited de novo CMV-specific proliferation of autologous PBMC in a dose-dependent fashion. Compared with the entire CMV-stimulated CD4{sup +} T-cell population, higher proportions of CD4{sup +}CD27{sup -}CD28{sup -} T{sub R} expressed FoxP3, TGFbeta, granzyme B, perforin, GITR and PD-1, lower proportions expressed CD127 and PD1-L and similar proportions expressed CD25, CTLA4, Fas-L and GITR-L. CMV-CD4{sup +}CD27{sup -}CD28{sup -}more » T{sub R} expanded in response to IL-2, but not to CMV antigenic restimulation. The anti-proliferative effect of CMV-CD4{sup +}CD27{sup -}CD28{sup -} T{sub R} significantly decreased after granzyme B or TGFbeta inhibition. The CMV-CD4{sup +}CD27{sup -}CD28{sup -} T{sub R} of HIV-infected and uninfected donors had similar phenotypes and anti-proliferative potency, but HIV-infected individuals had higher proportions of CMV-CD4{sup +}CD27{sup -}CD28{sup -} T{sub R}. The CMV-CD4{sup +}CD27{sup -}CD28{sup -} T{sub R} may contribute to the downregulation of CMV-specific and nonspecific immune responses of CMV-infected individuals.« less

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

Perkins, Timothy N.; Dentener, Mieke A.

Growth and development of the mature lung is a complex process orchestrated by a number of intricate developmental signaling pathways. Wingless-type MMTV-integration site (WNT) signaling plays critical roles in controlling branching morphogenesis cell differentiation, and formation of the conducting and respiratory airways. In addition, WNT pathways are often re-activated in mature lungs during repair and regeneration. WNT- signaling has been elucidated as a crucial contributor to the development of idiopathic pulmonary fibrosis as well as other hyper-proliferative lung diseases. Silicosis, a detrimental occupational lung disease caused by excessive inhalation of crystalline silica dust, is hallmarked by repeated cycles of damagingmore » inflammation, epithelial hyperplasia, and formation of dense, hyalinized nodules of whorled collagen. However, mechanisms of epithelial cell hyperplasia and matrix deposition are not well understood, as most research efforts have focused on the pronounced inflammatory response. Microarray data from our previous studies has revealed a number of WNT-signaling and WNT-target genes altered by crystalline silica in human lung epithelial cells. In the present study, we utilize pathway analysis to designate connections between genes altered by silica in WNT-signaling networks. Furthermore, we confirm microarray findings by QRT-PCR and demonstrate both activation of canonical (β-catenin) and down-regulation of non-canonical (WNT5A) signaling in immortalized (BEAS-2B) and primary (PBEC) human bronchial epithelial cells. These findings suggest that WNT-signaling and cross-talk with other pathways (e.g. Notch), may contribute to proliferative, fibrogenic and inflammatory responses to silica in lung epithelial cells. - Highlights: • Pathway analysis reveals silica-induced WNT-signaling in lung epithelial cells. • Silica-induced canonical WNT-signaling is mediated by autocrine/paracrine signals. • Crystalline silica decreases non-canonical WNT5A signaling. • Microarray reveals WNT as a novel complex signaling network in silica-mediated injury.« less

5-(Furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f), a new synthetic compound, causes human fibrosarcoma HT-1080 cell apoptosis by disrupting tubulin polymerisation and inducing G2/M arrest.

PubMed

Zuo, Daiying; Pang, Lili; Shen, Jiwei; Guan, Qi; Bai, Zhaoshi; Zhang, Huijuan; Li, Yao; Lu, Guodong; Zhang, Weige; Wu, Yingliang

2017-06-01

In the current study, we synthesized a series of new compounds targeting tubulin and tested their anti-proliferative activities. Among these new synthetic com-pounds, 5-(furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f) exhibited significant anti-proliferative activity against different human cancer cell lines including human gastric adenocarcinoma SGC-7901, human non-small cell lung cancer A549, and human fibrosarcoma HT-1080. As a result, 6f was selected to further test the sensitivity to different cancer cell lines including human cervical cancer cell line HeLa, human breast cancer cell line MCF-7, non-small cell lung cancer cell line A549, human liver carcinoma cell line HepG-2, human oral squamous cell carcinoma cell lines KB, SGC-7901 and HT-1080. Among these cell lines, HT-1080 and HeLa are the most sensitive. Therefore, HT-1080 was selected to further explore the properties of anti-proliferative activity and the underlying mechanisms. Our data proved that 6f exhibited strong anti-proliferative effects against HT-1080 cells in a time- and dose-dependent manner. We showed that the growth inhibitory effect of 6f in HT-1080 cells was related with microtubule depolymerisation. Molecular docking studies revealed that 6f interacted and bound efficiently with the colchicine-binding site of tubulin. In addition, 6f treatment induced G2/M cell cycle arrest dose-dependently and subsequently induced cell apoptosis. Western blot study indicated that upregulation of cyclin B1 and p-cdc2 was related with G2/M arrest. 6f-induced cell apoptosis was associated with both mitochondrial and death receptor pathway. In conclusion, our data showed that 6f, among the newly synthetic compounds, exhibited highest anti-proliferative activity by disrupting the microtubule polymerisation, causing G2/M arrest and subsequently inducing cell apoptosis in HT-1080 cells. Hence, 6f is a promising microtubule depolymerising agent for the treatment of various cancers especially human fibrosarcoma.

A Convenient and Efficient Method to Enrich and Maintain Highly Proliferative Human Fetal Liver Stem Cells

PubMed Central

Guo, Xuan; Wang, Shu; Dou, Ya-ling; Guo, Xiang-fei; Chen, Zhao-li; Wang, Xin-wei; Shen, Zhi-qiang; Qiu, Zhi-gang

2015-01-01

Abstract Pluripotent human hepatic stem cells have broad research and clinical applications, which are, however, restricted by both limited resources and technical difficulties with respect to isolation of stem cells from the adult or fetal liver. In this study, we developed a convenient and efficient method involving a two-step in situ collagenase perfusion, gravity sedimentation, and Percoll density gradient centrifugation to enrich and maintain highly proliferative human fetal liver stem cells (hFLSCs). Using this method, the isolated hFLSCs entered into the exponential growth phase within 10 days and maintained sufficient proliferative activity to permit subculture for at least 20 passages without differentiation. Immunocytochemistry, immunofluorescence, and flow cytometry results showed that these cells expressed stem cell markers, such as c-kit, CD44, epithelial cell adhesion molecule (EpCAM), oval cell marker-6 (OV-6), epithelial marker cytokeratin 18 (CK18), biliary ductal marker CK19, and alpha-fetoprotein (AFP). Gene expression analysis showed that these cells had stable mRNA expression of c-Kit, EpCAM, neural cell adhesion molecule (NCAM), CK19, CK18, AFP, and claudin 3 (CLDN-3) throughout each passage while maintaining low levels of ALB, but with complete absence of cytochrome P450 3A4 (C3A4), phosphoenolpyruvate carboxykinase (PEPCK), telomeric repeat binding factor (TRF), and connexin 26 (CX26) expression. When grown in appropriate medium, these isolated liver stem cells could differentiate into hepatocytes, cholangiocytes, osteoblasts, adipocytes, or endothelial cells. Thus, we have demonstrated a more economical and efficient method to isolate hFLSCs than magnetic-activated cell sorting (MACS). This novel approach may provide an excellent tool to isolate highly proliferative hFLSCs for tissue engineering and regenerative therapies. PMID:25556695

Stochastic cellular automata model of neurosphere growth: Roles of proliferative potential, contact inhibition, cell death, and phagocytosis.

PubMed

Sipahi, Rifat; Zupanc, Günther K H

2018-05-14

Neural stem and progenitor cells isolated from the central nervous system form, under specific culture conditions, clonal cell clusters known as neurospheres. The neurosphere assay has proven to be a powerful in vitro system to study the behavior of such cells and the development of their progeny. However, the theory of neurosphere growth has remained poorly understood. To overcome this limitation, we have, in the present paper, developed a cellular automata model, with which we examined the effects of proliferative potential, contact inhibition, cell death, and clearance of dead cells on growth rate, final size, and composition of neurospheres. Simulations based on this model indicated that the proliferative potential of the founder cell and its progenitors has a major influence on neurosphere size. On the other hand, contact inhibition of proliferation limits the final size, and reduces the growth rate, of neurospheres. The effect of this inhibition is particularly dramatic when a stem cell becomes encapsulated by differentiated or other non-proliferating cells, thereby suppressing any further mitotic division - despite the existing proliferative potential of the stem cell. Conversely, clearance of dead cells through phagocytosis is predicted to accelerate growth by reducing contact inhibition. A surprising prediction derived from our model is that cell death, while resulting in a decrease in growth rate and final size of neurospheres, increases the degree of differentiation of neurosphere cells. It is likely that the cellular automata model developed as part of the present investigation is applicable to the study of tissue growth in a wide range of systems. Copyright © 2018 Elsevier Ltd. All rights reserved.

A Convenient and Efficient Method to Enrich and Maintain Highly Proliferative Human Fetal Liver Stem Cells.

PubMed

Guo, Xuan; Wang, Shu; Dou, Ya-ling; Guo, Xiang-fei; Chen, Zhao-li; Wang, Xin-wei; Shen, Zhi-qiang; Qiu, Zhi-gang; Jin, Min; Li, Jun-wen

2015-06-01

Pluripotent human hepatic stem cells have broad research and clinical applications, which are, however, restricted by both limited resources and technical difficulties with respect to isolation of stem cells from the adult or fetal liver. In this study, we developed a convenient and efficient method involving a two-step in situ collagenase perfusion, gravity sedimentation, and Percoll density gradient centrifugation to enrich and maintain highly proliferative human fetal liver stem cells (hFLSCs). Using this method, the isolated hFLSCs entered into the exponential growth phase within 10 days and maintained sufficient proliferative activity to permit subculture for at least 20 passages without differentiation. Immunocytochemistry, immunofluorescence, and flow cytometry results showed that these cells expressed stem cell markers, such as c-kit, CD44, epithelial cell adhesion molecule (EpCAM), oval cell marker-6 (OV-6), epithelial marker cytokeratin 18 (CK18), biliary ductal marker CK19, and alpha-fetoprotein (AFP). Gene expression analysis showed that these cells had stable mRNA expression of c-Kit, EpCAM, neural cell adhesion molecule (NCAM), CK19, CK18, AFP, and claudin 3 (CLDN-3) throughout each passage while maintaining low levels of ALB, but with complete absence of cytochrome P450 3A4 (C3A4), phosphoenolpyruvate carboxykinase (PEPCK), telomeric repeat binding factor (TRF), and connexin 26 (CX26) expression. When grown in appropriate medium, these isolated liver stem cells could differentiate into hepatocytes, cholangiocytes, osteoblasts, adipocytes, or endothelial cells. Thus, we have demonstrated a more economical and efficient method to isolate hFLSCs than magnetic-activated cell sorting (MACS). This novel approach may provide an excellent tool to isolate highly proliferative hFLSCs for tissue engineering and regenerative therapies.

Clonal analysis of T-cell responses to herpes simplex virus: isolation, characterization and antiviral properties of an antigen-specific helper T-cell clone.

PubMed

Leung, K N; Nash, A A; Sia, D Y; Wildy, P

1984-12-01

A herpes simplex virus (HSV)-specific long-term T-cell clone has been established from the draining lymph node cells of BALB/c mice; the cells required repeated in vitro restimulation with UV-irradiated virus. The established T-cell clone expresses the Thy-1 and Lyt-1+2,3- surface antigens. For optimal proliferation of the cloned cells, both the presence of specific antigen and an exogenous source of T-cell growth factor are required. The proliferative response of the cloned T cells was found to be virus-specific but it did not distinguish between HSV-1 and HSV-2. Adoptive cell transfer of the cloned T cells helped primed B cells to produce anti-herpes antibodies: the response was antigen-specific and cell dose-dependent. The clone failed to produce a significant DTH reaction in vivo, but did produce high levels of macrophage-activating factor. Furthermore, the T-cell clone could protect from HSV infection, as measured by a reduction in local virus growth, and by enhanced survival following the challenge of mice with a lethal dose of virus. The mechanism(s) whereby this clone protects in vivo is discussed.

Fever-range hyperthermia inhibits cells immune response to protein-bound polysaccharides derived from Coriolus versicolor extract.

PubMed

Pawlikowska, Małgorzata; Jędrzejewski, Tomasz; Piotrowski, Jakub; Kozak, Wiesław

2016-12-01

The aim of the study was to explore whether fever-range hyperthermia (FRH) might enhance the anticancer and immunoregulatory activities of protein-bound polysaccharides (PBP), a class of fungus derived immunomodifiers used in the cancer adjuvant therapy. Blood lymphocytes and breast cancer cells (MCF-7) were cultured at 39.5°C in humidified atmosphere containing 5% CO 2 for 2h. After rested at 37°C for 6h, the cells were treated with PBP extract at 100- and 300μg/ml concentration. After indicated time, the proliferative response was analyzed and cytokine mRNA expression assessment was performed by qRT-PCR. In animal model, the FRH was induced by placing rats in the Homeothermic Controller with heating blanket. Animals were heated until Tb reached 39.5°C (±0.2°C) and were maintained at this temperature for 30min. The protein-bound polysaccharides solution was injected i.p. at a dose of 100 mg/kg 6h post FRH. Twenty four hours after treatment, the blood was collected and cytokines expression analysis were performed. The results have shown that fever-range hyperthermia has an inhibitory effect on PBP extract-induced proliferative response of blood lymphocytes, as well as IL-1β and IL-6 mRNA expression. Moreover, the temperature of 39.5°C blocks PBP-induced cytotoxicity against MCF-7 cells, which correlates with significant reduction in TNF-α level. Combined treatment of rats (FRH+PBP) results in decrease of IL-1β, IL-6 and TNF-α mRNA expression in peripheral blood mononuclear cells compared to cells derived from rats treated with protein-bound polysaccharides extract alone. This study demonstrates that fever-range temperature inhibits immunostimulatory as well as anticancer effects mediated by protein-bound polysaccharides. Copyright © 2016. Published by Elsevier Ltd.

Development of novobiocin analogues that manifest anti-proliferative activity against several cancer cell lines.

PubMed

Burlison, Joseph A; Avila, Christopher; Vielhauer, George; Lubbers, Donna J; Holzbeierlein, Jeffrey; Blagg, Brian S J

2008-03-21

Recent studies have shown that the DNA gyrase inhibitor, novobiocin, binds to a previously unrecognized ATP-binding site located at the C-terminus of Hsp90 and induces degradation of Hsp90-dependent client proteins at approximately 700 microM. As a result of these studies, several analogues of the coumarin family of antibiotics have been reported and shown to exhibit increased Hsp90 inhibitory activity; however, the monomeric species lacked the ability to manifest anti-proliferative activity against cancer cell lines at concentrations tested. In an effort to develop more efficacious compounds that produce growth inhibitory activity against cancer cell lines, structure-activity relationships were investigated surrounding the prenylated benzamide side chain of the natural product. Results obtained from these studies have produced the first novobiocin analogues that manifest anti-proliferative activity against several cancer cell lines.

The effects of erythropoietin signaling on telomerase regulation in non-erythroid malignant and non-malignant cells

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

Uziel, Orit, E-mail: Oritu@clalit.org.il; Kanfer, Gil; Dep. of Human Molecular Genetics and Biochemistry, Sackler School of Medicine, Tel-Aviv University, Ramat-Aviv

Highlights: • We assumed that some of erythropoietin adverse effects may be mediated by telomerase activity. • EPO administration increased telomerase activity, cells proliferation and migration. • The inhibition of telomerase modestly repressed the proliferative effect of erythropoietin. • Telomere shortening caused by long term inhibition of the enzyme totally abolished that effect. • This effect was mediated via the Lyn–AKT axis and not by the canonical JAK2–STAT pathway. - Abstract: Treatment with erythropoietin (EPO) in several cancers is associated with decreased survival due to cancer progression. Due to the major importance of telomerase in cancer biology we hypothesized thatmore » some of these effects may be mediated through EPO effect on telomerase. For this aim we explored the possible effects of EPO on telomerase regulation, cell migration and chemosensitivity in non-erythroid malignant and non-malignant cells. Cell proliferation, telomerase activity (TA) and cell migration increased in response to EPO. EPO had no effect on cancer cells sensitivity to cisplatinum and on the cell cycle status. The inhibition of telomerase modestly repressed the proliferative effect of EPO. Telomere shortening caused by long term inhibition of the enzyme abolished the effect of EPO, suggesting that EPO effects on cancer cells are related to telomere dynamics. TA was correlated with the levels of Epo-R. The increase in TA was mediated post-translationally through the Lyn-Src and not the canonical JAK2 pathway.« less

Zoledronic Acid-Induced Expansion of γδ T Cells from Early-Stage Breast Cancer Patients: Effect of IL-18 on Helper NK Cells

PubMed Central

Sugie, Tomoharu; Murata-Hirai, Kaoru; Iwasaki, Masashi; Morita, Craig T.; Li, Wen; Okamura, Haruki; Minato, Nagahiro; Toi, Masakazu; Tanaka, Yoshimasa

2013-01-01

Human γδ T cells display potent cytotoxicity against various tumor cells pretreated with zoledronic acid (Zol). Zol has shown benefits when added to adjuvant endocrine therapy for patients with early-stage breast cancer or to standard chemotherapy for patients with multiple myeloma. Although γδ T cells may contribute to this additive effect, the responsiveness of γδ T cells from early-stage breast cancer patients has not been fully investigated. In this study, we determined the number, frequency, and responsiveness of Vγ2Vδ2 T cells from early- and late-stage breast cancer patients and examined the effect of IL-18 on their ex vivo expansion. The responsiveness of Vγ2Vδ2 T cells from patients with low frequencies of Vγ2Vδ2 T cells was significantly diminished. IL-18, however, enhanced ex vivo proliferative responses of Vγ2Vδ2 T cells and helper NK cells from patients with either low or high frequencies of Vγ2Vδ2 T cells. Treatment of breast cancer patients with Zol alone decreased the number of Vγ2Vδ2 T cells and reduced their ex vivo responsiveness. These results demonstrate that Zol can elicit immunological responses by γδ T cells from early-stage breast cancer patients but that frequent in vivo treatment reduces Vγ2Vδ2 T cell numbers and their responsiveness to stimulation. PMID:23151944

Chronic hyperglicemia and nitric oxide bioavailability play a pivotal role in pro-atherogenic vascular modifications

PubMed Central

De Filippis, Elena Anna

2007-01-01

Diabetes is associated with accelerated atherosclerosis and macrovascular complications are a major cause of morbidity and mortality in this disease. Although our understanding of vascular pathology has lately greatly improved, the mechanism(s) underlying enhanced atherosclerosis in diabetes remain unclear. Endothelial cell dysfunction is emerging as a key component in the pathophysiology of cardiovascular abnormalities associated with diabetes. Although it has been established that endothelium plays a critical role in overall homeostasis of the vessels, vascular smooth muscle cells (vSMC) in the arterial intima have a relevant part in the development of atherosclerosis in diabetes. However, high glucose induced alterations in vSMC behaviour are not fully characterized. Several studies have reported that impaired nitric oxide (NO) synthesis and/or actions are often present in diabetes and endothelial dysfunction. Furthermore, although endothelial cells are by far the main site of vascular NO synthesis, vSMC do express nitric oxyde synthases (NOSs) and NO synthesis in vSMC might be important in vessel’s function. Although it is known that vSMC contribute to vascular pathology in diabetes by their change from a quiescent state to an activated proliferative and migratory phenotype (termed phenotypic modulation), whether this altered phenotypic modulation might also involve alterations in the nitrergic systems is still controversial. Our recent data indicate that, in vivo, chronic hyperglycemia might induce an increased number of vSMC proliferative clones which persist in culture and are associated with increased eNOS expression and activity. However, upregulation of eNOS and increased NO synthesis occur in the presence of a marked concomitant increase of O2− production. Since NO bioavailabilty might not be increased in high glucose stimulated vSMC, it is tempting to hypothesize that the proliferative phenotype observed in cells from diabetic rats is associated with a redox imbalance responsible quenching and/or trapping of NO, with the consequent loss of its biological activity. This might provide new insight on the mechanisms responsible for accelerated atherosclerosis in diabetes. PMID:18850175

AZD1480 delays tumor growth in a melanoma model while enhancing the suppressive activity of myeloid-derived suppressor cells

PubMed Central

Maenhout, Sarah K.; Four, Stephanie Du; Corthals, Jurgen; Neyns, Bart; Thielemans, Kris; Aerts, Joeri L.

2014-01-01

AZD1480 is a potent, competitive small-molecule inhibitor of JAK1/2 kinase which inhibits STAT3 phosphorylation and tumor growth. Here we investigated the effects of AZD1480 on the function of different immune cell populations in a melanoma model. When MO4 tumor-bearing mice were treated with AZD1480 we observed a strong inhibition of tumor growth as well as a prolonged survival. Moreover, a significant decrease in the percentage of myeloid-derived suppressor cells (MDSCs) was observed after treatment with AZD1480. However, AZD1480 enhanced the suppressive capacity of murine MDSCs while at the same time impairing the proliferative as well as the IFN-γ secretion capacity of murine T cells. The addition of AZD1480 to co-cultures of human MDSCs and T cells does not affect the suppressive activity of MDSCs but it does reduce the IFN-γ secretion and the proliferative capacity of T cells. We showed that although AZD1480 has the ability to delay the tumor growth of MO4 tumor-bearing mice, this drug has detrimental effects on several aspects of the immune system. These data indicate that systemic targeting of the JAK/STAT pathway by JAK1/2 inhibition can have divergent effects on tumor growth and anti-tumor immune responses. PMID:25149535

A Combination Histochemical and Autoradiographic Method for Analysis of Enzymatic and Proliferative Responses to Immunoreactive Osteoinducer

DTIC Science & Technology

1988-01-15

DNA replication and alkaline phosphate activity in the responding cells was shown to result in effective differential labeling of these features in mildly fixed tissue sections. Application of this method with monoclonal antibodies specific for induction-associated determinants and with modifications to permit ultrastructural analyses may provide important information relevant to the mechanism of matrix-induced bone formation. Keywords: Osteogenic implants; Immunohistochemistry;

Dietary strawberries increase proliferative response of CD3/CD28-activated CD8+ T cells and production of TNF-alpha in lipopolysaccharide-stimulated monocytes from obese human subjects

USDA-ARS?s Scientific Manuscript database

Obesity increases the risk of developing bacterial and viral infections compared to normal weight. In a 7 wk double-blind, randomized, crossover trial, twenty obese volunteers (20-50 y old, BMI between 30-40 kg/m2) were fed freeze-dried strawberry powder or strawberry-flavored placebo preparations ...

Substance P promotes expansion of human mesenteric preadipocytes through proliferative and antiapoptotic pathways.

PubMed

Gross, Kara; Karagiannides, Iordanes; Thomou, Thomas; Koon, Hon Wai; Bowe, Collin; Kim, Ho; Giorgadze, Nino; Tchkonia, Tamara; Pirtskhalava, Tamara; Kirkland, James L; Pothoulakis, Charalabos

2009-05-01

White adipose tissue is intimately involved in the regulation of immunity and inflammation. We reported that human mesenteric preadipocytes express the substance P (SP)-mediated neurokinin-1 receptor (NK-1R), which signals proinflammatory responses. Here we tested the hypothesis that SP promotes proliferation and survival of human mesenteric preadipocytes and investigated responsible mechanism(s). Preadipocytes were isolated from mesenteric fat biopsies during gastric bypass surgery. Proliferative and antiapoptotic responses were delineated in 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS), bromodeoxyuridine (BrdU), caspase-3, and TUNEL assays, as well as Western immunoanalysis. SP (10(-7) M) increased MTS and proliferation (BrdU) and time dependently (15-30 min) induced Akt, EGF receptor, IGF receptor, integrin alphaVbeta3, phosphatidylinositol 3-kinase, and PKC-theta phosphorylation. Furthermore, pharmacological antagonism of Akt and PKC-theta activation significantly attenuated SP-induced preadipocyte proliferation. Exposure of preadipocytes to the proapoptotic Fas ligand (FasL, 100 microM) resulted in nuclear DNA fragmentation (TUNEL assay), as well as increased cleaved poly (ADP-ribose) polymerase, cleaved caspase-7, and caspase-3 expression. Cotreatment with SP almost completely abolished these responses in a NK-1R-dependent fashion. SP (10(-7) M) also time dependently stimulated expression 4E binding protein 1 and phosphorylation of p70 S6 kinase, which increased protein translation efficiency. SP increases preadipocyte viability, reduces apoptosis, and stimulates proliferation, possibly via cell cycle upregulation and increased protein translation efficiency. SP-induced proliferative and antiapoptotic pathways in fat depots may contribute to development of the creeping fat and inflammation characteristic of Crohn's disease.

Eco-friendly synthesis, in vitro anti-proliferative evaluation, and 3D-QSAR analysis of a novel series of monocationic 2-aryl/heteroaryl-substituted 6-(2-imidazolinyl)benzothiazole mesylates.

PubMed

Racané, Livio; Ptiček, Lucija; Sedić, Mirela; Grbčić, Petra; Kraljević Pavelić, Sandra; Bertoša, Branimir; Sović, Irena; Karminski-Zamola, Grace

2018-04-17

Herein, we describe the synthesis of twenty-one novel water-soluble monocationic 2-aryl/heteroaryl-substituted 6-(2-imidazolinyl)benzothiazole mesylates 3a-3u and present the results of their anti-proliferative assays. Efficient syntheses were achieved by three complementary simple two-step synthetic protocols based on the condensation reaction of aryl/heteroaryl carbaldehydes or carboxylic acid. We developed an eco-friendly synthetic protocol using glycerol as green solvent, particularly appropriate for the condensation of thermally and acid-sensitive heterocycles such as furan, benzofuran, pyrrole, and indole. Screening of anti-proliferative activity was performed on four human tumour cell lines in vitro including pancreatic cancer (CFPAC-1), metastatic colon cancer (SW620), hepatocellular carcinoma (HepG2), and cervical cancer (HeLa), as well as in normal human fibroblast cell lines. All tested compounds showed strong to moderate anti-proliferative activity on tested cell lines depending on the structure containing aryl/heteroaryl moiety coupled to 6-(2-imidazolinyl)benzothiazole moiety. The most potent cytostatic effects on all tested cell lines with [Formula: see text] values ranging from 0.1 to 3.70 [Formula: see text] were observed for benzothiazoles substituted with naphthalene-2-yl 3c, benzofuran-2-yl 3e, indole-3-yl 3j, indole-2-yl 3k, quinoline-2-yl 3s, and quinoline-3-yl 3t and derivatives substituted with phenyl 3a, naphthalene-1-yl 3b, benzothiazole-2-yl 3g, benzothiazole-6-yl 3h, N-methylindole-3-yl 3l, benzimidazole-2-yl 3n, benzimidazole-5(6)-yl 3o, and quinolone-4-yl 3u with [Formula: see text] values ranging from 1.1 to 29.1 [Formula: see text]. Based on obtained anti-proliferative activities, 3D-QSAR models for five cell lines were derived. Molecular volume, molecular surface, the sum of hydrophobic surface areas, molecular mass, and possibility of making dispersion forces were identified by QSAR analyses as molecular properties that are positively correlated with anti-proliferative activity, while compound's capability to accept H-bond was identified as a negatively correlated property. Comparison of molecular properties identified for different cell lines enabled assumptions about similarity of mode of action through which anti-proliferative activities against different cell lines are accomplished. Novel compounds that are predicted to have enhanced activities in comparison with herein presented ones were designed using 3D-QSAR analysis as guideline.

Phytochemical properties and anti-proliferative activity of Olea europaea L. leaf extracts against pancreatic cancer cells.

PubMed

Goldsmith, Chloe D; Vuong, Quan V; Sadeqzadeh, Elham; Stathopoulos, Costas E; Roach, Paul D; Scarlett, Christopher J

2015-07-17

Olea europaea L. leaves are an agricultural waste product with a high concentration of phenolic compounds; especially oleuropein. Oleuropein has been shown to exhibit anti-proliferative activity against a number of cancer types. However, they have not been tested against pancreatic cancer, the fifth leading cause of cancer related death in Western countries. Therefore, water, 50% ethanol and 50% methanol extracts of Corregiola and Frantoio variety Olea europaea L. leaves were investigated for their total phenolic compounds, total flavonoids and oleuropein content, antioxidant capacity and anti-proliferative activity against MiaPaCa-2 pancreatic cancer cells. The extracts only had slight differences in their phytochemical properties, and at 100 and 200 μg/mL, all decreased the viability of the pancreatic cancer cells relative to controls. At 50 μg/mL, the water extract from the Corregiola leaves exhibited the highest anti-proliferative activity with the effect possibly due to early eluting HPLC peaks. For this reason, olive leaf extracts warrant further investigation into their potential anti-pancreatic cancer benefits.

Role of Mesenchymal-Derived Stem Cells in Stimulating Dormant Tumor Cells to Proliferate and Form Clinical Metastases

DTIC Science & Technology

2016-07-01

have determined the break in dormancy is dependent on collagen and other fibrotic extracellular matrix components for the induction of a proliferative...state in these dormant D2.0R breast cancer cell lines. Performing gene expression array on these dormant D2.0R cells exposed to collagen to induce a...D2.0R”) and proliferate when cultured in matrigel supplemented with collagen type-1 (“proliferative D2.0R”) (Barkan, Cancer Research, 2008, 68(15

Suppressive effects of chlorphenesin on lymphocyte function in mice and humans.

PubMed

Stites, D P; Brecher, G; Schmidt, L; Berger, F M

1979-12-01

The immunosuppressive action of chlorphenesin was investigated in a wide variety of in vitro assays for cellular immunity in humans and mice. Chlorphenesin, at doses of 20-50 micrograms/ml, inhibited mitogenic responses of both mouse and human B and T cells. These doses did not kill cells exposed to the drug for 72 hr. Mixed lymphocyte reactions in inbred strains of mice and in unrelated humans were also inhibited at concentrations of about 50 micrograms/ml. However, the generation of cytotoxic T cells in cell-mediated lympholysis assays was not inhibited to the same degree as proliferation in mixed lymphocyte reaction and the cytotoxic potential of presensitized mouse T cells for allogeneic targets was totally unaffected. These studies suggest that chlorphenesin may have a broad spectrum of suppressive effects both on T and B cells and that the predominant inhibition of proliferative responses in these cells may reduce the expansion of clones of immunocompetent cells in vivo.

The inhibition of Caco-2 proliferation by astaxanthin from Xanthophyllomyces dendrorhous.

PubMed

Wayakanon, Kornchanok; Rueangyotchanthana, Kanjana; Wayakanon, Praween; Suwannachart, Chatrudee

2018-04-01

To investigate the efficiency of natural astaxanthin that has been extracted from Xanthophyllomyces dendrorhous in inhibiting the proliferation and viability of colorectal adenocarcinoma cell line (Caco-2; colon cancer cells). Caco-2 cells and normal human oralkeratinocytes (NOKs) were treated with different concentrations of extracted astaxanthin, ranging from 0.075 to 10 mg ml -1 , for 24, 48 and 72 h. The number of cells was determined via MTS assay and the proliferating cells were investigated by bromodeoxyuridine (BrdU) assay.Results/Key findings. Of the Caco-2 cells, 30-50 % remained viable, while the NOKs showed 110-120 % survival when treated with 5 mg ml -1 astaxanthin. The Caco-2 cells showed distinct structural shrinkage when treated with the same concentration of astaxanthin. Fluorescent labelling of the DNA of the proliferative cells with BrdU showed a significant decrease in the number of the proliferative Caco-2 cells when the concentration of astaxanthin was increased to 5 mg ml -1 . The natural astaxanthin from X. dendrorhous, at an appropriate concentration, is effective in terminating the viability of, or retarding the proliferative activity of, Caco-2 cells, without harmful effects on NOKs.

Investigating the anti-proliferative activity of styrylazanaphthalenes and azanaphthalenediones.

PubMed

Mrozek-Wilczkiewicz, Anna; Kalinowski, Danuta S; Musiol, Robert; Finster, Jacek; Szurko, Agnieszka; Serafin, Katarzyna; Knas, Magdalena; Kamalapuram, Sishir K; Kovacevic, Zaklina; Jampilek, Josef; Ratuszna, Alicja; Rzeszowska-Wolny, Joanna; Richardson, Des R; Polanski, Jaroslaw

2010-04-01

A group of styrylazanaphthalenes and azanaphthalenediones were synthesized and tested for their anti-proliferative activity. Most of the compounds were obtained with the use of microwave-assisted synthesis. The lipophilicity of the compounds was measured by RP-HPLC and their anti-proliferative activity was assayed against the human SK-N-MC neuroepithelioma and HCT116 human colon carcinoma cell lines. Active compounds were also tested in clonogenity and comet assays. Several quinazolinone and styrylquinazoline analogues were found to have markedly greater anti-proliferative activity than desferoxamine and cis-platin. Copyright 2010 Elsevier Ltd. All rights reserved.

Slow-cycling stem cells in hydra contribute to head regeneration

PubMed Central

Govindasamy, Niraimathi; Murthy, Supriya; Ghanekar, Yashoda

2014-01-01

ABSTRACT Adult stem cells face the challenge of maintaining tissue homeostasis by self-renewal while maintaining their proliferation potential over the lifetime of an organism. Continuous proliferation can cause genotoxic/metabolic stress that can compromise the genomic integrity of stem cells. To prevent stem cell exhaustion, highly proliferative adult tissues maintain a pool of quiescent stem cells that divide only in response to injury and thus remain protected from genotoxic stress. Hydra is a remarkable organism with highly proliferative stem cells and ability to regenerate at whole animal level. Intriguingly, hydra does not display consequences of high proliferation, such as senescence or tumour formation. In this study, we investigate if hydra harbours a pool of slow-cycling stem cells that could help prevent undesirable consequences of continuous proliferation. Hydra were pulsed with the thymidine analogue 5-ethynyl-2′-deoxyuridine (EdU) and then chased in the absence of EdU to monitor the presence of EdU-retaining cells. A significant number of undifferentiated cells of all three lineages in hydra retained EdU for about 8–10 cell cycles, indicating that these cells did not enter cell cycle. These label-retaining cells were resistant to hydroxyurea treatment and were predominantly in the G2 phase of cell cycle. Most significantly, similar to mammalian quiescent stem cells, these cells rapidly entered cell division during head regeneration. This study shows for the first time that, contrary to current beliefs, cells in hydra display heterogeneity in their cell cycle potential and the slow-cycling cells in this population enter cell cycle during head regeneration. These results suggest an early evolution of slow-cycling stem cells in multicellular animals. PMID:25432513

Oxidative stress triggers cytokinesis failure in hepatocytes upon isolation.

PubMed

Tormos, A M; Taléns-Visconti, R; Bonora-Centelles, A; Pérez, S; Sastre, J

2015-01-01

Primary hepatocytes are highly differentiated cells and proliferatively quiescent. However, the stress produced during liver digestion seems to activate cell cycle entry by proliferative/dedifferentiation programs that still remain unclear. The aim of this work was to assess whether the oxidative stress associated with hepatocyte isolation affects cell cycle and particularly cytokinesis, the final step of mitosis. Hepatocytes were isolated from C57BL/6 mice by collagenase perfusion in the absence and presence of N-acetyl cysteine (NAC). Polyploidy, cell cycle, and reactive oxygen species (ROS) were studied by flow cytometry (DNA, phospho-histone 3, and CellROX(®) Deep Red) and Western blotting (cyclins B1 and D1, and proliferating cell nuclear antigen). mRNA expression of cyclins A1, B1, B2, D1, and F by reverse transcription (RT)-PCR was also assessed. Glutathione levels were measured by mass spectrometry. Here we show that hepatocyte isolation enhanced cell cycle entry, increased hepatocyte binucleation, and caused marked glutathione oxidation. Addition of 5 mM NAC to the hepatocyte isolation media prevented glutathione depletion, partially blocked ROS production and cell cycle entry of hepatocytes, and avoided the blockade of mitosis progression, abrogating defective cytokinesis and diminishing the formation of binucleated hepatocytes during isolation. Therefore, addition of NAC to the isolation media decreased the generation of polyploid hepatocytes confirming that oxidative stress occurs during hepatocyte isolation and it is responsible, at least in part, for cytokinesis failure and hepatocyte binucleation.

The long non-coding RNA HOTAIR promotes the proliferation of serous ovarian cancer cells through the regulation of cell cycle arrest and apoptosis

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

Qiu, Jun-jun; Department of Obstetrics and Gynecology of Shanghai Medical College, Fudan University, 138 Yixueyuan Road, Shanghai 200032; Shanghai Key Laboratory of Female Reproductive Endocrine-Related Diseases, 413 Zhaozhou Road, Shanghai 200011

HOX transcript antisense RNA (HOTAIR) is a well-known long non-coding RNA (lncRNA) whose dysregulation correlates with poor prognosis and malignant progression in many forms of cancer. Here, we investigate the expression pattern, clinical significance, and biological function of HOTAIR in serous ovarian cancer (SOC). Clinically, we found that HOTAIR levels were overexpressed in SOC tissues compared with normal controls and that HOTAIR overexpression was correlated with an advanced FIGO stage and a high histological grade. Multivariate analysis revealed that HOTAIR is an independent prognostic factor for predicting overall survival in SOC patients. We demonstrated that HOTAIR silencing inhibited A2780 andmore » OVCA429 SOC cell proliferation in vitro and that the anti-proliferative effects of HOTAIR silencing also occurred in vivo. Further investigation into the mechanisms responsible for the growth inhibitory effects by HOTAIR silencing revealed that its knockdown resulted in the induction of cell cycle arrest and apoptosis through certain cell cycle-related and apoptosis-related proteins. Together, these results highlight a critical role of HOTAIR in SOC cell proliferation and contribute to a better understanding of the importance of dysregulated lncRNAs in SOC progression. - Highlights: • HOTAIR overexpression correlates with an aggressive tumour phenotype and a poor prognosis in SOC. • HOTAIR promotes SOC cell proliferation both in vitro and in vivo. • The proliferative role of HOTAIR is associated with regulation of the cell cycle and apoptosis.« less

Extracellular NAMPT/Visfatin induces proliferation through ERK1/2 and AKT and inhibits apoptosis in breast cancer cells.

PubMed

Gholinejad, Zafar; Kheiripour, Nejat; Nourbakhsh, Mitra; Ilbeigi, Davod; Behroozfar, Kiarash; Hesari, Zahra; Golestani, Abolfazl; Shabani, Mohammad; Einollahi, Nahid

2017-06-01

Visfatin is a novel adipokine and proinflammatory cytokine which is implicated in breast cancer progression. The exact proliferative and anti-apoptotic mechanisms of visfatin are still under debate. In this study, the effect of extracellular visfatin on proliferation and apoptosis of breast cancer cells were investigated considering key regulatory molecules in these procedures. BrdU (Bromodeoxyuridine) experiment was used to assess cell proliferation in response to visfatin treatment. Cell viability and apoptosis were assessed using MTT assay and flowcytometry, respectively. Phosphorylation levels of AKT and ERK1/2 as well as survivin levels and Poly ADP ribose polymerase (PARP) cleavage were investigated by western blot analysis. Visfatin induced proliferation of MCF-7 and MDA-MB-231 cells, an effect that was repressed by using AKT and ERK1/2 inhibitors, indicating involvement of these two signaling pathways in the proliferative effect of visfatin. Similarly, phosphorylation of AKT and ERK1/2 were elevated by visfatin treatment. On the other hand, visfatin improved cell viability and prevented TNF-α-induced apoptosis as well as PARP cleavage. Visfatin also exerted a protective effect on survivin. The results of this study suggest that visfatin induces breast cancer cell proliferation through AKT/PI3K and ERK/MAPK activation and protects against apoptosis in these cells. Thus increased visfatin levels may augment breast cancer development and attenuate treatment efficiency in breast cancer patients. Copyright © 2017 Elsevier Inc. All rights reserved.

The Monocyte Chemoattractant Protein-1/Cognate CC Chemokine Receptor 2 System Affects Cell Motility in Cultured Human Podocytes

PubMed Central

Burt, Davina; Salvidio, Gennaro; Tarabra, Elena; Barutta, Federica; Pinach, Silvia; Dentelli, Patrizia; Camussi, Giovanni; Perin, Paolo Cavallo; Gruden, Gabriella

2007-01-01

In crescentic glomerulonephritis (GN), monocyte chemoattractant protein-1 (MCP-1) is overexpressed within the glomeruli, and MCP-1 blockade has renoprotective effects. Adult podocytes are in a quiescent state, but acquisition of a migratory/proliferative phenotype has been described in crescentic GN and implicated in crescent formation. The cognate CC chemokine receptor 2 (CCR2), the MCP-1 receptor, is expressed by other cell types besides monocytes and has been implicated in both cell proliferation and migration. We investigated whether MCP-1 binding to CCR2 can induce a migratory/proliferative response in cultured podocytes. MCP-1 binding to CCR2 enhanced podocyte chemotaxis/haptotaxis in a concentration-dependent manner and had a modest effect on cell proliferation. Closure of a wounded podocyte monolayer was delayed by CCR2 blockade, and CCR2 was overexpressed at the wound edge, suggesting a role for CCR2 in driving podocyte migration. Immunohistochemical analysis of kidney biopsies from patients with crescentic GN demonstrated CCR2 expression in both podocytes and cellular crescents, confirming the clinical relevance of our in vitro findings. In conclusion, the MCP-1/CCR2 system is functionally active in podocytes and may be implicated in the migratory events triggered by podocyte injury in crescentic GN and other glomerular diseases. PMID:18055544

Mitochondria, calcium, and tumor suppressor Fus1: At the crossroad of cancer, inflammation, and autoimmunity

PubMed Central

Uzhachenko, Roman; Shanker, Anil; Yarbrough, Wendell G.; Ivanova, Alla V.

2015-01-01

Mitochondria present a unique set of key intracellular functions such as ATP synthesis, production of reactive oxygen species (ROS) and Ca2+ buffering. Mitochondria both encode and decode Ca2+ signals and these interrelated functions have a direct impact on cell signaling and metabolism. High proliferative potential is a key energy-demanding feature shared by cancer cells and activated T lymphocytes. Switch of a metabolic state mediated by alterations in mitochondrial homeostasis plays a fundamental role in maintenance of the proliferative state. Recent studies show that tumor suppressors have the ability to affect mitochondrial homeostasis controlling both cancer and autoimmunity. Herein, we discuss established and putative mechanisms of calcium–dependent regulation of both T cell and tumor cell activities. We use the mitochondrial protein Fus1 as a case of tumor suppressor that controls immune response and tumor growth via maintenance of mitochondrial homeostasis. We focus on the regulation of mitochondrial Ca2+ handling as a key function of Fus1 and highlight the mechanisms of a crosstalk between Ca2+ accumulation and mitochondrial homeostasis. Given the important role of Ca2+ signaling, mitochondrial Ca2+ transport and ROS production in the activation of NFAT and NF-κB transcription factors, we outline the importance of Fus1 activities in this context. PMID:26246474

Combined changes in Wnt signaling response and contact inhibition induce altered proliferation in radiation-treated intestinal crypts

PubMed Central

Dunn, S.-J.; Osborne, J. M.; Appleton, P. L.; Näthke, I.

2016-01-01

Curative intervention is possible if colorectal cancer is identified early, underscoring the need to detect the earliest stages of malignant transformation. A candidate biomarker is the expanded proliferative zone observed in crypts before adenoma formation, also found in irradiated crypts. However, the underlying driving mechanism for this is not known. Wnt signaling is a key regulator of proliferation, and elevated Wnt signaling is implicated in cancer. Nonetheless, how cells differentiate Wnt signals of varying strengths is not understood. We use computational modeling to compare alternative hypotheses about how Wnt signaling and contact inhibition affect proliferation. Direct comparison of simulations with published experimental data revealed that the model that best reproduces proliferation patterns in normal crypts stipulates that proliferative fate and cell cycle duration are set by the Wnt stimulus experienced at birth. The model also showed that the broadened proliferation zone induced by tumorigenic radiation can be attributed to cells responding to lower Wnt concentrations and dividing at smaller volumes. Application of the model to data from irradiated crypts after an extended recovery period permitted deductions about the extent of the initial insult. Application of computational modeling to experimental data revealed how mechanisms that control cell dynamics are altered at the earliest stages of carcinogenesis. PMID:27053661

Steroid hormones as regulators of the proliferative activity of normal and neoplastic intestinal epithelial cells (review).

PubMed

Tutton, P J; Barkla, D H

1988-01-01

Glucocorticoid and mineralocorticoid receptors are present in normal epithelial cells of both the small and large intestine and there have also been contentious reports of androgen, oestrogen and progesterone receptors in the epithelium of the normal large intestine. The majority of reports suggest that stimulation of the intestinal glucocorticoid receptors results in increased proliferation of epithelial cells in the small bowel, as does stimulation of androgen receptors and possibly mineralocorticoid receptors. The proliferative response of the normal intestine to oestrogens is difficult to evaluate and that to progestigens appears not to have been reported. Epidemiological studies reveal a higher incidence of bowel cancer in premenopausal women than in men of the same age and yet there is a lower incidence of these tumors in women of higher parity. These findings have been atributted to a variety of non-epithelial gender characteristic such as differences in bile metabolism, colonic bacterial and fecal transit times. In experimental animals, androgens have also been shown to influence carcinogenesis and this could well be attributed to changes in food intake etc. However, many studies have now revealed steroid hormone receptors on colorectal tumor cells and thus a direct effect of the steroid hormones on the epithelium during and after malignant transformation must now be considered.

Acquisition of Pneumococci Specific Effector and Regulatory Cd4+ T Cells Localising within Human Upper Respiratory-Tract Mucosal Lymphoid Tissue

PubMed Central

Pido-Lopez, Jeffrey; Kwok, William W.; Mitchell, Timothy J.; Heyderman, Robert S.; Williams, Neil A.

2011-01-01

The upper respiratory tract mucosa is the location for commensal Streptococcus (S.) pneumoniae colonization and therefore represents a major site of contact between host and bacteria. The CD4+ T cell response to pneumococcus is increasingly recognised as an important mediator of immunity that protects against invasive disease, with data suggesting a critical role for Th17 cells in mucosal clearance. By assessing CD4 T cell proliferative responses we demonstrate age-related sequestration of Th1 and Th17 CD4+ T cells reactive to pneumococcal protein antigens within mucosal lymphoid tissue. CD25hi T cell depletion and utilisation of pneumococcal specific MHCII tetramers revealed the presence of antigen specific Tregs that utilised CTLA-4 and PDL-1 surface molecules to suppress these responses. The balance between mucosal effector and regulatory CD4+ T cell immunity is likely to be critical to pneumococcal commensalism and the prevention of unwanted pathology associated with carriage. However, if dysregulated, such responses may render the host more susceptible to invasive pneumococcal infection and adversely affect the successful implementation of both polysaccharide-conjugate and novel protein-based pneumococcal vaccines. PMID:22144893

Isolation of osteoprogenitors from human jaw periosteal cells: a comparison of two magnetic separation methods.

PubMed

Olbrich, Marcus; Rieger, Melanie; Reinert, Siegmar; Alexander, Dorothea

2012-01-01

Human jaw periosteum tissue contains osteoprogenitors that have potential for tissue engineering applications in oral and maxillofacial surgeries. To isolate osteoprogenitor cells from heterogeneous cell populations, we used the specific mesenchymal stem cell antigen-1 (MSCA-1) antibody and compared two magnetic separation methods. We analyzed the obtained MSCA-1(+) and MSCA-1(-) fractions in terms of purity, yield of positive/negative cells and proliferative and mineralization potentials. The analysis of cell viability after separation revealed that the EasySep method yielded higher viability rates, whereas the flow cytometry results showed a higher purity for the MACS-separated cell fractions. The mineralization capacity of the osteogenic induced MSCA-1(+) cells compared with the MSCA-1(-) controls using MACS was 5-fold higher, whereas the same comparison after EasySep showed no significant differences between both fractions. By analyzing cell proliferation, we detected a significant difference between the proliferative potential of the osteogenic cells versus untreated cells after the MACS and EasySep separations. The differentiated cells after MACS separation adjusted their proliferative capacity, whereas the EasySep-separated cells failed to do so. The protein expression analysis showed small differences between the two separation methods. Our findings suggest that MACS is a more suitable separation method to isolate osteoprogenitors from the entire jaw periosteal cell population.

Phylogeny of lymphocyte heterogeneity: the cellular requirements for the mixed leucocyte reaction with channel catfish.

PubMed Central

Miller, N W; Deuter, A; Clem, L W

1986-01-01

Vigorous mixed leucocyte reactions (MLR) were obtained using channel catfish peripheral blood leucocytes (PBL) when equal numbers of responder and stimulator cells (5 X 10(5) cells each) were cocultured. The use of 2000 rads of X-irradiation was sufficient to block subsequent proliferative responses of the stimulator cells. The cellular requirements for channel catfish MLR responses were assessed by using three functionally distinct leucocyte subpopulations isolated from the PBL. B cells (sIg+ lymphocytes) and T cells (sIg- lymphocytes) were isolated by an indirect panning procedure employing a monoclonal antibody specific for channel catfish Ig. A third population, monocytes, was isolated or depleted by adherence to baby hamster kidney cell microexudate-coated surfaces or adherence to Sephadex G-10, respectively. The results indicated that only the T cells were able to respond in the fish MLR, with monocytes being required as accessory cells. In contrast, all three cell types could function as stimulator cells. In addition, it was observed that low in vitro culture temperatures inhibited the generation of channel catfish MLRs, thereby supporting the contention that low temperature immunosuppression in fish results from a preferential inhibition of the generation of primary T-cell responses. PMID:2944817

Green synthesis of bacterial mediated anti-proliferative gold nanoparticles: inducing mitotic arrest (G2/M phase) and apoptosis (intrinsic pathway)

NASA Astrophysics Data System (ADS)

Ganesh Kumar, C.; Poornachandra, Y.; Chandrasekhar, Cheemalamarri

2015-11-01

The physiochemical and biological properties of microbial derived gold nanoparticles have potential applications in various biomedical domains as well as in cancer therapy. We have fabricated anti-proliferative bacterial mediated gold nanoparticles (b-Au NPs) using a culture supernatant of Streptomyces clavuligerus and later characterized them by UV-visible, TEM, DLS, XRD and FT-IR spectroscopic techniques. The capping agent responsible for the nanoparticle formation was characterized based on SDS-PAGE and MALDI-TOF-MS analyses. They were tested for anticancer activity in A549, HeLa and DU145 cell lines. The biocompatibility and non-toxic nature of the nanoparticles were tested on normal human lung cell line (MRC-5). The b-Au NPs induced the cell cycle arrest in G2/M phase and also inhibited the microtubule assembly in DU145 cells. Mechanistic studies, such as ROS, MMP, Cyt-c, GSH, caspases 9, 8 and 3 activation and the Annexin V-FITC staining, along with the above parameters tested provided sufficient evidence that the b-Au NPs induced apoptosis through the intrinsic pathway. The results supported the use of b-Au NPs for future therapeutic application in cancer therapy and other biomedical applications.The physiochemical and biological properties of microbial derived gold nanoparticles have potential applications in various biomedical domains as well as in cancer therapy. We have fabricated anti-proliferative bacterial mediated gold nanoparticles (b-Au NPs) using a culture supernatant of Streptomyces clavuligerus and later characterized them by UV-visible, TEM, DLS, XRD and FT-IR spectroscopic techniques. The capping agent responsible for the nanoparticle formation was characterized based on SDS-PAGE and MALDI-TOF-MS analyses. They were tested for anticancer activity in A549, HeLa and DU145 cell lines. The biocompatibility and non-toxic nature of the nanoparticles were tested on normal human lung cell line (MRC-5). The b-Au NPs induced the cell cycle arrest in G2/M phase and also inhibited the microtubule assembly in DU145 cells. Mechanistic studies, such as ROS, MMP, Cyt-c, GSH, caspases 9, 8 and 3 activation and the Annexin V-FITC staining, along with the above parameters tested provided sufficient evidence that the b-Au NPs induced apoptosis through the intrinsic pathway. The results supported the use of b-Au NPs for future therapeutic application in cancer therapy and other biomedical applications. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr04577k

[Quantitative variations of lymphocyte subsets in various kinds of cancer patients in the terminal stage].

PubMed

Nakajima, Y; Akimoto, M; Iwasaki, H; Matano, S; Hirakawa, H; Kimura, M

1986-12-01

Immunological studies of the peripheral blood were made in terminal breast cancer and terminal abdominal cancer patients. Two immunological parameters were studied: (1) lymphocyte subsets and (2) proliferative response to PHA. A decrease in the number of OKT-3(+) cells and an increase in that of OKT-8(+) cells were observed in abdominal cancer. It was suggested that the immunological status in abdominal cancer is more suppressive than in breast cancer. Increases in the number of OK-M1(+) cells and Leu-7(+) cells were observed in breast cancer. It is suggested that cytotoxic lymphocytes increase in number in breast cancer more than in abdominal cancer.

Lymphocyte-mediated regulation of platelet activation during desensitization in patients with hymenoptera venom hypersensitivity.

PubMed Central

Ledru, E; Pestel, J; Tsicopoulos, A; Joseph, M; Wallaert, B; Tonnel, A B; Capron, A

1988-01-01

T cells from peripheral blood of hymenoptera sensitive patients were studied before and after venom desensitization. Before treatment, T cells showed a variable but higher proliferative response to allergen than T cells of treated patients or controls. While before desensitization, T cell products, specifically released after in vitro allergen stimulation, were able to amplify the IgE-dependent platelet activity, we showed that after treatment of the same patients, T cell products strongly reduced platelet activation. Considering the modifications in platelet activation previously observed in patients treated by specific immunotherapy, the present results suggest that, through a modification of T cell reactivity to allergen, T cell functions are modulated by desensitization, and emphasize the involvement of T cell products in the desensitization mechanisms. PMID:3263227

Proliferative verrucous leukoplakia: diagnosis, management and current advances.

PubMed

Capella, Diogo Lenzi; Gonçalves, Jussara Maria; Abrantes, Adelino António Artur; Grando, Liliane Janete; Daniel, Filipe Ivan

Proliferative verrucous leukoplakia is a multifocal and progressive lesion of the oral mucosa, with unknown etiology, and commonly resistant to all therapy attempts with frequent recurrences. It is characterized by a high rate of oral squamous cell carcinoma and verrucou carcinoma transformations. To analyze the studies about Proliferative verrucous leukoplakia and develop a concise update. A Pubmed search identifying studies (laboratory research, case series and reviews of literature) that examined patients with Proliferative verrucous leukoplakia was realized. There are not enough studies about Proliferative verrucous leukoplakia in the literature. The few found studies not present a consensus about its etiology and diagnosis criteria. Although several treatment strategies have been proposed, most of them still show a high recurrence rate. More research about Proliferative verrucous leukoplakia is necessary to understand and treat this disease. Copyright © 2017 Associação Brasileira de Otorrinolaringologia e Cirurgia Cérvico-Facial. Published by Elsevier Editora Ltda. All rights reserved.

Long-term memory cellular immune response to dengue virus after a natural primary infection.

PubMed

Sierra, Beatríz; García, Gissel; Pérez, Ana B; Morier, Luis; Rodríguez, Rayner; Alvarez, Mayling; Guzmán, María G

2002-06-01

This study was conducted to examine the memory T-cell response to dengue virus 20 years after a primary infection. We took advantage of the exceptional epidemiologic situation in Cuba, where the population initially suffered two large successive epidemics due to dengue virus 1 and 2 respectively over a 4-year period. Thereafter, no dengue virus circulation was subsequently observed, except for the Santiago de Cuba municipality. T-cell response was evaluated in peripheral blood mononuclear cells (PBMCs) from 20 individuals with history of a primary infection by dengue virus 1 or 2. Methods previously shown to induce lymphoproliferation of CD4+ memory T-cell subpopulations were used. We evaluated the proliferative responses generated in those PBMCs after stimulation with dengue virus 1, 2, 3 and 4 antigens in a serotype-specific and serotype-crossreactive way. Serotype-specific and serotype-crossreactive lymphoproliferative responses in all PBMCs donated by dengue immune donors were observed. The serotype-crossreactive response for dengue 2 was stronger than for the rest of the serotypes. This is the first report of cellular memory lymphocyte response specific for dengue virus detected 20 years after a primary infection by dengue.

Synthesis and anti-proliferative activity of fluoro-substituted chalcones.

PubMed

Burmaoglu, Serdar; Algul, Oztekin; Anıl, Derya Aktas; Gobek, Arzu; Duran, Gulay Gulbol; Ersan, Ronak Haj; Duran, Nizami

2016-07-01

A series of novel fluoro-substituted chalcone derivatives have been synthesized. All synthesized compounds were characterized by (1)H nuclear magnetic resonance (NMR), (13)C NMR, and elemental analysis. Their anti-proliferative activities were evaluated against five cancer cells lines, namely, A549, A498, HeLa, A375, and HepG2 using the MTT method. Most of the compounds showed moderate to high activity with IC50 values in the range of 0.029-0.729μM. Of all the synthesized compounds, 10 and 19 exhibited the most potent anti-proliferative activities against cancer cells, and 10 was identified as the most promising compound. Copyright © 2016 Elsevier Ltd. All rights reserved.

Macrophage Migration Inhibitory Factor Mediates Proliferative GN via CD74

PubMed Central

Djudjaj, Sonja; Lue, Hongqi; Rong, Song; Papasotiriou, Marios; Klinkhammer, Barbara M.; Zok, Stephanie; Klaener, Ole; Braun, Gerald S.; Lindenmeyer, Maja T.; Cohen, Clemens D.; Bucala, Richard; Tittel, Andre P.; Kurts, Christian; Moeller, Marcus J.; Floege, Juergen; Ostendorf, Tammo

2016-01-01

Pathologic proliferation of mesangial and parietal epithelial cells (PECs) is a hallmark of various glomerulonephritides. Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that mediates inflammation by engagement of a receptor complex involving the components CD74, CD44, CXCR2, and CXCR4. The proliferative effects of MIF may involve CD74 together with the coreceptor and PEC activation marker CD44. Herein, we analyzed the effects of local glomerular MIF/CD74/CD44 signaling in proliferative glomerulonephritides. MIF, CD74, and CD44 were upregulated in the glomeruli of patients and mice with proliferative glomerulonephritides. During disease, CD74 and CD44 were expressed de novo in PECs and colocalized in both PECs and mesangial cells. Stress stimuli induced MIF secretion from glomerular cells in vitro and in vivo, in particular from podocytes, and MIF stimulation induced proliferation of PECs and mesangial cells via CD74. In murine crescentic GN, Mif-deficient mice were almost completely protected from glomerular injury, the development of cellular crescents, and the activation and proliferation of PECs and mesangial cells, whereas wild-type mice were not. Bone marrow reconstitution studies showed that deficiency of both nonmyeloid and bone marrow–derived Mif reduced glomerular cell proliferation and injury. In contrast to wild-type mice, Cd74-deficient mice also were protected from glomerular injury and ensuing activation and proliferation of PECs and mesangial cells. Our data suggest a novel molecular mechanism and glomerular cell crosstalk by which local upregulation of MIF and its receptor complex CD74/CD44 mediate glomerular injury and pathologic proliferation in GN. PMID:26453615

Nuclear Migration During Retinal Development

PubMed Central

Baye, Lisa M.; Link, Brian A.

2009-01-01

In this review we focus on the mechanisms, regulation, and cellular consequences of nuclear migration in the developing retina. In the nervous system, nuclear migration is prominent during both proliferative and post-mitotic phases of development. Interkinetic nuclear migration is the process where the nucleus oscillates from the apical to basal surfaces in proliferative neuroepithelia. Proliferative nuclear movement occurs in step with the cell cycle, with M-phase being confined to the apical surface and G1-, S-, and G2-phases occurring at more basal locations. Later, following cell cycle exit, some neuron precursors migrate by nuclear translocation. In this mode of cellular migration, nuclear movement is the driving force for motility. Following discussion of the key components and important regulators for each of these processes, we present an emerging model where interkinetic nuclear migration functions to distinguish cell fates among retinal neuroepithelia. PMID:17560964

Anti‑inflammatory and anti‑proliferative effects of Rhus verniciflua Stokes in RAW264.7 cells.

PubMed

Choi, Han-Seok; Seo, Hye Sook; Kim, Soon Re; Choi, Youn Kyung; Jang, Bo-Hyoung; Shin, Yong-Cheol; Ko, Seong-Gyu

2014-01-01

Inflammatory response is a major defense mechanism against pathogens and chemical or mechanical injury. Rhus verniciflua Stokes (RVS) has traditionally been used as an ingredient in East Asian medicine for the treatment of gastritis, stomach cancer and atherosclerosis. The aim of the current study was to analyze the effect of RVS on LPS‑induced inflammatory responses in the RAW264.7 mouse macrophage cell line. RAW264.7 cells were treated with various concentrations of RVS and LPS at specific time points. WST assay, trypan blue assay and quantification of activated cells revealed that RVS suppressed cell proliferation in a dose‑dependent manner. RVS induced G1 cell cycle arrest, suppressed iNOS and COX‑2 mRNA expression induced by LPS and decreased intracellular ROS levels induced by LPS. In addition, RVS induced PARP and caspase‑3 cleavage suggesting that RVS causes cell death. Results of the present study indicate that RVS may be advantageous in treating inflammatory disease.

Inhibition of T cell-mediated functions by MVM(i), a parvovirus closely related to minute virus of mice.

PubMed

Engers, H D; Louis, J A; Zubler, R H; Hirt, B

1981-12-01

A purified preparation of MVM(i), a murine parvovirus closely related to minute virus of mice (MVM), was found to inhibit various functions mediated by murine T cells in vitro. Addition of MVM(i) virus to secondary allogeneic mixed leukocyte cultures resulted in the inhibition of both lymphocyte proliferation (3H-thymidine incorporation) and the generation of cytolytic T lymphocyte activity but not interferon production. MVM(i) virus also inhibited the growth and cytolytic activity of several cloned, long-term Lyt-2+ cytolytic T cell lines. Furthermore, the antigen-induced proliferative responses of parasite- (Leishmania) specific Lyt-1+ T cells in vitro was abrogated by the addition of MVM(i) virus to the culture. Finally, the suppression of an in vitro antibody response to SRBC by MVM(i) virus was the result of the inhibition of T helper cells required for the B cell response. These suppressive effects were specific for MVM(i); parallel studies in which the prototype MVM parvovirus was used showed no significant inhibition in the various systems tested.

RNA sequencing-based cell proliferation analysis across 19 cancers identifies a subset of proliferation-informative cancers with a common survival signature.

PubMed

Ramaker, Ryne C; Lasseigne, Brittany N; Hardigan, Andrew A; Palacio, Laura; Gunther, David S; Myers, Richard M; Cooper, Sara J

2017-06-13

Despite advances in cancer diagnosis and treatment strategies, robust prognostic signatures remain elusive in most cancers. Cell proliferation has long been recognized as a prognostic marker in cancer, but the generation of comprehensive, publicly available datasets allows examination of the links between cell proliferation and cancer characteristics such as mutation rate, stage, and patient outcomes. Here we explore the role of cell proliferation across 19 cancers (n = 6,581 patients) by using tissue-based RNA sequencing data from The Cancer Genome Atlas Project and calculating a 'proliferative index' derived from gene expression associated with Proliferating Cell Nuclear Antigen (PCNA) levels. This proliferative index is significantly associated with patient survival (Cox, p-value < 0.05) in 7 of 19 cancers, which we have defined as "proliferation-informative cancers" (PICs). In PICs, the proliferative index is strongly correlated with tumor stage and nodal invasion. PICs demonstrate reduced baseline expression of proliferation machinery relative to non-PICs. Additionally, we find the proliferative index is significantly associated with gross somatic mutation burden (Spearman, p = 1.76 x 10-23) as well as with mutations in individual driver genes. This analysis provides a comprehensive characterization of tumor proliferation indices and their association with disease progression and prognosis in multiple cancer types and highlights specific cancers that may be particularly susceptible to improved targeting of this classic cancer hallmark.

Epithelial atrophy in oral submucous fibrosis is mediated by copper (II) and arecoline of areca nut

PubMed Central

Khan, Imran; Pant, Ila; Narra, Sivakrishna; Radhesh, Rekha; Ranganathan, Kannan; Rao, Somanahalli Girish; Kondaiah, Paturu

2015-01-01

Exposure of oral cavity to areca nut is associated with several pathological conditions including oral submucous fibrosis (OSF). Histopathologically OSF is characterized by epithelial atrophy, chronic inflammation, juxtaepithelial hyalinization, leading to fibrosis of submucosal tissue and affects 0.5% of the population in the Indian subcontinent. As the molecular mechanisms leading to atrophied epithelium and fibrosis are poorly understood, we studied areca nut actions on human keratinocyte and gingival fibroblast cells. Areca nut water extract (ANW) was cytotoxic to epithelial cells and had a pro-proliferative effect on fibroblasts. This opposite effect of ANW on epithelial and fibroblast cells was intriguing but reflects the OSF histopathology such as epithelial atrophy and proliferation of fibroblasts. We demonstrate that the pro-proliferative effects of ANW on fibroblasts are dependent on insulin-like growth factor signalling while the cytotoxic effects on keratinocytes are dependent on the generation of reactive oxygen species. Treatment of keratinocytes with arecoline which is a component of ANW along with copper resulted in enhanced cytotoxicity which becomes comparable to IC50 of ANW. Furthermore, studies using cyclic voltammetry, mass spectrometry and plasmid cleavage assay suggested that the presence of arecoline increases oxidation reduction potential of copper leading to enhanced cleavage of DNA which could generate an apoptotic response. Terminal deoxynucleotidyl transferase dUTP Nick End Labeling assay and Ki-67 index of OSF tissue sections suggested epithelial apoptosis, which could be responsible for the atrophy of OSF epithelium. PMID:26248978

Effect of glutamine supplementation on changes in the immune system induced by repeated exercise.

PubMed

Rohde, T; MacLean, D A; Pedersen, B K

1998-06-01

The ability of lymphocytes to proliferate and generate lymphokine activated killer (LAK) cell activity in vitro is dependent on glutamine. In relation to intense exercise the lymphocyte concentration, the proliferative response, the natural killer and LAK cell activity, and the plasma glutamine concentration decline. It has been hypothesized that in relation to physical activity a lack of glutamine may temporarily affect the function of the immune system. The purpose of this study was to examine the influence of glutamine supplementation on exercise-induced immune changes. In a randomized cross-over placebo-controlled study, eight healthy male subjects performed three bouts of ergometer bicycle exercise lasting 60, 45, and 30 min at 75% of their VO2max separated by 2 h of rest. The arterial plasma glutamine concentration declined from 508 +/- 35 (pre-exercise) to 402 +/- 38 microM (2 h after the last exercise bout) in the placebo trial and was maintained above pre-exercise levels in the glutamine supplementation trial. The numbers of circulating lymphocytes and the phytohemagglutinin-stimulated lymphocyte proliferative response declined 2 h after, respectively, during each bout of exercise, whereas the LAK cell activity declined 2 h after the third bout. Glutamine supplementation in vivo, given in the described doses at the specific times, did not influence these changes. The present study does not appear to support the hypothesis that those aspects of postexercise immune changes studied are caused by decreased plasma glutamine concentrations.

Antagonizing Effects of Aspartic Acid against Ultraviolet A-Induced Downregulation of the Stemness of Human Adipose Tissue-Derived Mesenchymal Stem Cells

PubMed Central

Lee, Jienny; Shin, Seoung Woo; Jang, Sunghee; Jung, Eunsun; Kim, Min Hee; Lee, Jongsung

2015-01-01

Ultraviolet A (UVA) irradiation is responsible for a variety of changes in cell biology. The purpose of this study was to investigate effects of aspartic acid on UVA irradiation-induced damages in the stemness properties of human adipose tissue-derived mesenchymal stem cells (hAMSCs). Furthermore, we elucidated the UVA-antagonizing mechanisms of aspartic acid. The results of this study showed that aspartic acid attenuated the UVA-induced reduction of the proliferative potential and stemness of hAMSCs, as evidenced by increased proliferative activity in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and upregulation of stemness-related genes OCT4, NANOG, and SOX2 in response to the aspartic acid treatment. UVA-induced reduction in the mRNA level of hypoxia-inducible factor (HIF)-1α was also significantly recovered by aspartic acid. In addition, the antagonizing effects of aspartic acid against the UVA effects were found to be mediated by reduced production of PGE2 through the inhibition of JNK and p42/44 MAPK. Taken together, these findings show that aspartic acid improves reduced stemness of hAMSCs induced by UVA and its effects are mediated by upregulation of HIF-1α via the inhibition of PGE2-cAMP signaling. In addition, aspartic acid may be used as an antagonizing agent to mitigate the effects of UVA. PMID:25909857

Apicidin sensitizes pancreatic cancer cells to gemcitabine by epigenetically regulating MUC4 expression.

PubMed

Ansari, Daniel; Urey, Carlos; Hilmersson, Katarzyna Said; Bauden, Monika P; Ek, Fredrik; Olsson, Roger; Andersson, Roland

2014-10-01

Mucin 4 (MUC4) has been linked to resistance to gemcitabine in pancreatic cancer cells. The aim of the present study was to assess whether epigenetic control of MUC4 expression can sensitize pancreatic cancer cells to gemcitabine treatment. A 76-member combined epigenetics and phosphatase small-molecule inhibitor library was screened for anti-proliferative activity against the MUC4(+) gemcitabine-resistant pancreatic cancer cell line Capan-1, followed by high-content screening of protein expression. Apicidin, a histone deacetylase inhibitor, showed the greatest anti-proliferative activity with a lethal dose 50 (LD50) value of 5.17 μM. Apicidin significantly reduced the expression of MUC4 and its transcription factor hepatocyte nuclear factor 4α. Combined treatment with a sub-therapeutic concentration of apicidin and gemcitabine synergistically inhibited growth of Capan-1 cells. Apicidin appears to be a novel anti-proliferative agent against pancreatic cancer cells that may reverse chemoresistance by epigenetically regulating MUC4 expression. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Presence of natural killer-cell clones with variable proliferative capacity in chronic active Epstein-Barr virus infection.

PubMed

Nagata, H; Numata, T; Konno, A; Mikata, I; Kurasawa, K; Hara, S; Nishimura, M; Yamamoto, K; Shimizu, N

2001-10-01

Chronic active Epstein-Barr virus infection (CAEBV) is a syndrome that takes diverse clinical courses and is often associated with lymphoproliferative disorders of T/natural killer (NK)-cell lineage. We describe a patient with CAEBV associated with persistent pharyngeal ulcer, and with subsequent nasal T/NK-cell lymphoma in her neck lymph nodes and nasopharynx. Immunophenotyping of lymphoid cells showed that the lineage of Epstein-Barr virus (EBV)-positive cells in the patient was of NK-cell origin. By means of high-dose recombinant interleukin-2, we established an EBV-positive cell line of NK-cell lineage from her peripheral blood. Southern blot analysis for the number of terminal repeat sequences of EBV detected three NK-cell clones in the patient's lymph node. One of these clones was identical to the established cell line but was not observed in the pharyngeal ulcer, while the other two clones were present in the pharyngeal ulcer. These results suggest that the patient had expansion of the three NK-cell clones, one of which had proliferative capacity in vitro and was involved in the formation of the lymphoma. Moreover, the results suggest that the proliferative capacity of EBV-positive cells can be variable even in a single patient, and this variability may explain the clinical diversity in CAEBV.

Polish Natural Bee Honeys Are Anti-Proliferative and Anti-Metastatic Agents in Human Glioblastoma multiforme U87MG Cell Line

PubMed Central

Moskwa, Justyna; Borawska, Maria H.; Markiewicz-Zukowska, Renata; Puscion-Jakubik, Anna; Naliwajko, Sylwia K.; Socha, Katarzyna; Soroczynska, Jolanta

2014-01-01

Honey has been used as food and a traditional medicament since ancient times. However, recently many scientists have been concentrating on the anti-oxidant, anti-proliferative, anti-inflammatory and other properties of honey. In this study, we investigated for the first time an anticancer effect of different honeys from Poland on tumor cell line - glioblastoma multiforme U87MG. Anti-proliferative activity of honeys and its interferences with temozolomide were determined by a cytotoxicity test and DNA binding by [H3]-thymidine incorporation. A gelatin zymography was used to conduct an evaluation of metalloproteinases (MMP-2 and MMP-9) expression in U87MG treatment with honey samples. The honeys were previously tested qualitatively (diastase activity, total phenolic content, lead and cadmium content). The data demonstrated that the examined honeys have a potent anti-proliferative effect on U87MG cell line in a time- and dose-dependent manner, being effective at concentrations as low as 0.5% (multifloral light honey - viability 53% after 72 h of incubation). We observed that after 48 h, combining honey with temozolomide showed a significantly higher inhibitory effect than the samples of honey alone. We observed a strong inhibition of MMP-2 and MMP-9 for the tested honeys (from 20 to 56% and from 5 to 58% compared to control, respectively). Our results suggest that Polish honeys have an anti-proliferative and anti-metastatic effect on U87MG cell line. Therefore, natural bee honey can be considered as a promising adjuvant treatment for brain tumors. PMID:24594866

Quinazolinones-Phenylquinoxaline hybrids with unsaturation/saturation linkers as novel anti-proliferative agents.

PubMed

Palem, Jyothsna Devi; Alugubelli, Gopi Reddy; Bantu, Rajashaker; Nagarapu, Lingaiah; Polepalli, Sowjanya; Jain, S Nishanth; Bathini, Raju; Manga, Vijjulatha

2016-07-01

A new series of novel quinazolinones with allylphenyl quinoxaline hybrids 9a-n were efficiently synthesized in good yields by the reaction of 3-allyl-2-methylquinazolin-4(3H)-one (5a-n) with bromophenyl)quinoxaline (8) utilizing Pd catalyzed Heck-cross coupling and evaluated for anti-proliferative activity against four cancer cell lines such as HeLa (cervical), MIAPACA (pancreatic), MDA-MB-231 (breast) and IMR32 (neuroblastoma). Compounds 9a, 9e, 9g and 9h exhibited promising anti-proliferative activity with GI50 values ranging from 0.06 to 0.2μM against four cell lines, while compounds 9e and 9k showed significant activity against HeLa and MIAPACA cell lines and compounds 9b, 9d, 9h and 9j showed selective potency against IMR32 and MDA-MB-231 cell lines. This is the first report on the synthesis and in vitro anti-proliferative evaluation of E-2-(4-substituted)-3-(3-(4-(quinoxalin-2-yl)phenyl)allyl)quinazolin-4(3H)-ones (9a-n). Docking results indicate a sign of good correlation between experimental activity and calculated binding affinity (dock score), suggesting that these compounds could act as promising DNA intercalates. Copyright © 2016 Elsevier Ltd. All rights reserved.

In vitro antigen-specific induction of IL-22 in human subjects that resolved HCV infection

PubMed Central

Cusick, Matthew F; Libbey, Jane E; Fujinami, Robert S; Eckels, David D

2012-01-01

Aims To determine if in vitro production of IL-22 and IL-17 correlated with resolution of HCV infection. Materials & methods Human peripheral blood cells isolated from a well-defined cohort of resolved and chronic HCV-infected subjects were used to measure HCV-, influenza- and mitogen-activated T-cell proliferation. In addition, IL-22 and IL-17 production was measured via ELISAs and flow cytometry. Results Resolved HCV subjects had a significantly higher T-cell proliferative response to recombinant NS3 protein compared with chronic HCV subjects. Resolved subjects had a dose-dependent IL-22 response to recombinant NS3 compared with chronic HCV subjects. Conclusion IL-22 production is associated with antigen-specific induction of CD4 + T cells in individuals that resolved HCV infection, suggesting a potential role for IL-22 in HCV clearance. PMID:23185211

Phytochemical regulation of the tumor suppressive microRNA, miR-34a, by p53-dependent and independent responses in human breast cancer cells

PubMed Central

Hargraves, Kris G.; He, Lin; Firestone, Gary L.

2016-01-01

The tumor suppressive microRNA miR-34a is transcriptionally regulated by p53 and shown to inhibit breast cancer cell proliferation as well as being a marker of increased disease free survival. Indole-3-carbinol (I3C) derived from cruciferous vegetables, artemisinin, extracted from the sweet wormwood plant, and artesunate, a semi-synthetic derivative of artemisinin, are phytochemicals with anti-tumorigenic properties however, little is known about the role of microRNAs in their mechanism of action. Human breast cancer cells expressing wild-type (MCF-7) or mutant p53 (T47D) were treated with a concentration range and time course of each phytochemical under conditions of cell cycle arrest as detected by flow cytometry to examine the potential connection between miR-34a expression and their anti-proliferative responses. Real-time PCR and western blot analysis of extracted RNA and total protein revealed artemsinin and artesunate increased miR-34a expression in a dose-dependent manner correlating with down-regulation of the miR-34a target gene, CDK4. I3C stimulation of miR-34a expression required functional p53, whereas, both artemisinin and artesunate up-regulated miR-34a expression regardless of p53 mutational status or in the presence of dominant negative p53. Phytochemical treatments inhibited the luciferase activity of a construct containing the wild-type 3′UTR of CDK4, but not those with a mutated miR-34a binding site, whereas, transfection of miR-34a inhibitors ablated the phytochemical mediated down-regulation of CDK4 and induction of cell cycle arrest. Our results suggest that miR-34a is an essential component of the anti-proliferative activities of I3C, artemisinin and artesunate and demonstrate that both wild-type p53 dependent and independent pathways are responsible for miR-34a induction. PMID:25789847

Does dietary fibre stimulate intestinal epithelial cell proliferation in germ free rats?

PubMed Central

Goodlad, R A; Ratcliffe, B; Fordham, J P; Wright, N A

1989-01-01

The aim of the present experiment was to investigate the role of hind gut fermentation in the proliferative response of the intestinal epithelium to dietary fibre. We have previously shown that refeeding starved rats with an elemental diet supplemented with fermentable dietary fibre (but not inert bulk) is capable of stimulating intestinal epithelial cell proliferation throughout the gastrointestinal tract. Three groups of 10 germ free (GF) rats and three groups of 10 conventional (CV) rats, were used. All groups were starved for three days and then refed for two days with either an elemental diet (Flexical); Flexical plus 30% kaolin; or Flexical plus 30% of a fibre mixture. Cell production was determined by the accumulation of vincristine arrested metaphases in microdissected crypts. There was no significant difference between refeeding the rats with an elemental diet alone or with kaolin supplementation, however, the addition of fibre in CV rats was associated with a significant increase in intestinal crypt cell production rate in both the small intestine (p less than 0.01) and the colon (p less than 0.001). This marked proliferative effects of fibre was abolished in the GF rats. It can be concluded that it is the products of hind gut fermentation, not fibre per se that stimulate intestinal epithelial cell proliferation in the colon and small intestine. PMID:2546871

Loss of embryonic MET signaling alters profiles of hippocampal interneurons.

PubMed

Martins, Gabriela J; Plachez, Céline; Powell, Elizabeth M

2007-01-01

Hippocampal interneurons arise in the ventral forebrain and migrate dorsally in response to cues, including hepatocyte growth factor/scatter factor which signals via its receptor MET. Examination of the hippocampus in adult mice in which MET had been inactivated in the embryonic proliferative zones showed an increase in parvalbumin-expressing cells in the dentate gyrus, but a loss of these cells in the CA3 region. An overall loss of calretinin-expressing cells was seen throughout the hippocampus. A similar CA3 deficit of parvalbumin and calretinin cells was observed when MET was eliminated only in postmitotic cells. These data suggest that MET is required for the proper hippocampal development, and embryonic perturbations lead to long-term anatomical defects with possible learning and memory dysfunction.

RB mutation and RAS overexpression induce resistance to NK cell-mediated cytotoxicity in glioma cells.

PubMed

Orozco-Morales, Mario; Sánchez-García, Francisco Javier; Golán-Cancela, Irene; Hernández-Pedro, Norma; Costoya, Jose A; de la Cruz, Verónica Pérez; Moreno-Jiménez, Sergio; Sotelo, Julio; Pineda, Benjamín

2015-01-01

Several theories aim to explain the malignant transformation of cells, including the mutation of tumor suppressors and proto-oncogenes. Deletion of Rb (a tumor suppressor), overexpression of mutated Ras (a proto-oncogene), or both, are sufficient for in vitro gliomagenesis, and these genetic traits are associated with their proliferative capacity. An emerging hallmark of cancer is the ability of tumor cells to evade the immune system. Whether specific mutations are related with this, remains to be analyzed. To address this issue, three transformed glioma cell lines were obtained (Rb(-/-), Ras(V12), and Rb(-/-)/Ras(V12)) by in vitro retroviral transformation of astrocytes, as previously reported. In addition, Ras(V12) and Rb(-/-)/Ras(V12) transformed cells were injected into SCID mice and after tumor growth two stable glioma cell lines were derived. All these cells were characterized in terms of Rb and Ras gene expression, morphology, proliferative capacity, expression of MHC I, Rae1δ, and Rae1αβγδε, mult1, H60a, H60b, H60c, as ligands for NK cell receptors, and their susceptibility to NK cell-mediated cytotoxicity. Our results show that transformation of astrocytes (Rb loss, Ras overexpression, or both) induced phenotypical and functional changes associated with resistance to NK cell-mediated cytotoxicity. Moreover, the transfer of cell lines of transformed astrocytes into SCID mice increased resistance to NK cell-mediated cytotoxicity, thus suggesting that specific changes in a tumor suppressor (Rb) and a proto-oncogene (Ras) are enough to confer resistance to NK cell-mediated cytotoxicity in glioma cells and therefore provide some insight into the ability of tumor cells to evade immune responses.

Proliferative capability of parietal and zymogen cells.

PubMed Central

Chen, K Y; Withers, H R

1975-01-01

Six to eight week old male mice of C3Hf/Bu strain were killed and studied at various times from 30 minutes to 30 days after tritiated thymidine (3H-TdR) injection. Labelling of parietal and zymogen cells was observed in autoradiographic histological specimens. There were 590 +/- 22 gastric glands per circumference at the body of the stomach. Parietal cells were counted as 16-2 (14-0-18-4) per gland or 9600 per circumference. There were 11-4 (9-4-13-7) zymogen cells per gland or 6700 per circumference. Two labelled immature parietal cells per circumference were seen 30 minutes after 3H-TdR injection, equivalent to a ratio of 1:5000; more mature labelled parietal cells were seen at later times up to 30 days. There was evidence for proliferation during maturation and downward migration of cells toward the body and the lower part of the gastric gland. Our data support earlier evidence that immature parietal cells constantly supply mature parietal cells through migration from a proliferative zone. Whether the parietal cells are derived from the same stem cell compartment as surface epithelium cells is unclear at the present time. An average of 2-6 zymogen cells per circumference, or 1:2500, was found to be labelled. While most zymogen cells were not proliferating, cells entered the proliferating cycle at random. Based on the findings reported in this paper and the radiation responses of both parietal and zymogen cells, it is postulated that they form part of a slow renewal system. Images Fig. 1 (cont.) Fig. 1 (cont.) Fig. 1 (cont.) Fig. 1 Fig. 2 (cont.) Fig. 2 (cont.) Fig. 2 PMID:1213947

Establishment of proliferative tetraploid cells from telomerase-immortalized normal human fibroblasts.

PubMed

Ohshima, Susumu; Seyama, Atsushi

2016-06-01

Aneuploidy is observed in the majority of human cancers and is considered to be causally related to carcinogenesis. Although malignant aneuploid cells are suggested to develop from polyploid cells formed in precancerous lesions, the mechanisms of this process remain elusive. This is partly because no experimental model is available where nontransformed polyploid human cells propagate in vitro. We previously showed that proliferative tetraploid cells can be established from normal human fibroblasts by treatment with the spindle poison demecolcine (DC). However, the limited lifespan of these cells hampered detailed analysis of a link between chromosomal instability and the oncogenic transformation of polyploid cells. Here, we report the establishment of proliferative tetraploid cells from the telomerase-immortalized normal human fibroblast cell line TIG-1. Treatment of immortalized diploid cells with DC for 4 days resulted in proliferation of cells with tetraploid DNA content and near-tetraploid/tetraploid chromosome counts. Established tetraploid cells had functional TP53 despite growing at almost the same rate as diploid cells. The frequency of clonal and sporadic chromosome aberrations in tetraploid cells was higher than in diploid cells and in one experiment, gradually increased with repeated subculture. This study suggests that tetraploid cells established from telomerase-immortalized normal human fibroblasts can be a valuable model for studying chromosomal instability and the oncogenic potential of polyploid cells. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Potent anti-proliferative effects against oral and cervical cancers of Thai medicinal plants selected from the Thai/Lanna medicinal plant recipe database "MANOSROI III".

PubMed

Manosroi, Aranya; Akazawa, Hiroyuki; Pattamapun, Kassara; Kitdamrongtham, Worapong; Akihisa, Toshihiro; Manosroi, Worapaka; Manosroi, Jiradej

2015-07-01

Thai/Lanna medicinal plant recipes have been used for the treatment of several diseases including oral and cervical cancers. To investigate anti-proliferative activity on human cervical (HeLa) and oral (KB) cancer cell lines of medicinal plants selected from Thai/Lanna medicinal plant recipe database "MANOSROI III". Twenty-three methanolic plant crude extracts were tested for phytochemicals and anti-proliferative activity on HeLa and KB cell lines for 24 h by the sulforhodamine B (SRB) assay at the doses of 1 × 10(1)-1 × 10(-6 )mg/ml. The nine extracts with the concentrations giving 50% growth inhibition (GI50) lower than 100 µg/ml were further semi-purified by liquid/liquid partition in order to evaluate and enhance the anti-proliferative potency. All extracts contained steroids/triterpenoids, but not xanthones. The methanolic extracts of Gloriosa superba L. (Colchinaceae) root and Albizia chinensis (Osbeck) Merr. (Leguminosae-Mimosoideae) wood gave the highest anti-proliferative activity on HeLa and KB cell lines with the GI50 values of 0.91 (6.0- and 0.31-fold of cisplatin and doxorubicin) and 0.16 µg/ml (28.78- and 82.29-fold of cisplatin and doxorubicin), respectively. Hexane and methanol-water fractions of G. superba exhibited the highest anti-proliferative activity on HeLa and KB cell lines with the GI50 values of 0.15 (37- and 1.9-fold of cisplatin and doxorubicin) and 0.058 µg/ml (77.45- and 221.46-fold of cisplatin and doxorubicin), respectively. This study has demonstrated the potential of plants selected from MANOSROI III database especially G. superba and A. chinensis for further development as anti-oral and cervical cancer agents.

Pupillary responses in non-proliferative diabetic retinopathy.

PubMed

Park, Jason C; Chen, Yi-Fan; Blair, Norman P; Chau, Felix Y; Lim, Jennifer I; Leiderman, Yannek I; Shahidi, Mahnaz; McAnany, J Jason

2017-03-23

The goal of this study was to determine the extent of rod-, cone-, and melanopsin-mediated pupillary light reflex (PLR) abnormalities in diabetic patients who have non-proliferative diabetic retinopathy (NPDR). Fifty diabetic subjects who have different stages of NPDR and 25 age-equivalent, non-diabetic controls participated. PLRs were measured in response to full-field, brief-flash stimuli under conditions that target the rod, cone, and intrinsically-photosensitive (melanopsin) retinal ganglion cell pathways. Pupil responses were compared among the subjects groups using age-corrected linear mixed models. Compared to control, the mean baseline pupil diameters were significantly smaller for all patient groups in the dark (all p < 0.001) and for the moderate-severe NPDR group in the light (p = 0.003). Pairwise comparisons indicated: (1) the mean melanopsin-mediated PLR was significantly reduced in the mild and moderate-severe groups (both p < 0.001); (2) the mean cone-mediated PLR was reduced significantly in the moderate-severe group (p = 0.008); (3) no significant differences in the mean rod-mediated responses. The data indicate abnormalities in NPDR patients under conditions that separately assess pupil function driven by different photoreceptor classes. The results provide evidence for compromised neural function in these patients and provide a promising approach for quantifying their neural abnormalities.

Chondron curvature mapping in growth plate cartilage under compressive loading.

PubMed

Vendra, Bhavya B; Roan, Esra; Williams, John L

2018-05-18

The physis, or growth plate, is a layer of cartilage responsible for long bone growth. It is organized into reserve, proliferative and hypertrophic zones. Unlike the reserve zone where chondrocytes are randomly arranged, either singly or in pairs, the proliferative and hypertrophic chondrocytes are arranged within tubular structures called chondrons. In previous studies, the strain patterns within the compressed growth plate have been reported to be nonuniform and inhomogeneous, with an apparent random pattern in compressive strains and a localized appearance of tensile strains. In this study we measured structural deformations along the entire lengths of chondrons when the physis was subjected to physiological (20%) and hyper-physiological (30% and 40%) levels of compression. This provided a means to interpret the apparent random strain patterns seen in texture correlation maps in terms of bending deformations of chondron structures and provided a physical explanation for the inhomogeneous and nonuniform strain patterns reported in previous studies. We observed relatively large bending deformations (kinking) of the chondron structures at the interface of the reserve and proliferative zones during compression. Bending in this region may induce dividing cells to align longitudinally to maintain column formation and drive longitudinal growth. Copyright © 2018 Elsevier Ltd. All rights reserved.

Essential role of Stat6 in IL-4 signalling.

PubMed

Takeda, K; Tanaka, T; Shi, W; Matsumoto, M; Minami, M; Kashiwamura, S; Nakanishi, K; Yoshida, N; Kishimoto, T; Akira, S

1996-04-18

Interleukin-4 (IL-4) is a pleiotropic lymphokine which plays an important role in the immune system. IL-4 activates two distinct signalling pathways through tyrosine phosphorylation of Stat6, a signal transducer and activator of transcription, and of a 170K protein called 4PS. To investigate the functional role of Stat6 in IL-4 signalling, we generated mice deficient in Stat6 by gene targeting. We report here that in the mutant mice, expression of CD23 and major histocompatibility complex (MHC) class II in resting B cells was not enhanced in response to IL-4. IL-4 induced B-cell proliferation costimulated by anti-IgM antibody was abolished. The T-cell proliferative response was also notably reduced. Furthermore, production of Th2 cytokines from T cells as well as IgE and IgG1 responses after nematode infection were profoundly reduced. These findings agreed with those obtained in IL-4 deficient mice or using antibodies to IL-4 and the IL-4 receptor. We conclude that Stat6 plays a central role in exerting IL-4 mediated biological responses.

Study of the quantitative, functional, cytogenetic, and immunoregulatory properties of bone marrow mesenchymal stem cells in patients with B-cell chronic lymphocytic leukemia.

PubMed

Pontikoglou, Charalampos; Kastrinaki, Maria-Christina; Klaus, Mirjam; Kalpadakis, Christina; Katonis, Pavlos; Alpantaki, Kalliopi; Pangalis, Gerassimos A; Papadaki, Helen A

2013-05-01

The bone marrow (BM) microenvironment has clearly been implicated in the pathogenesis of B-cell chronic lymphocytic leukemia (B-CLL). However, the potential involvement of BM stromal progenitors, the mesenchymal stem cells (MSCs), in the pathophysiology of the disease has not been extensively investigated. We expanded in vitro BM-MSCs from B-CLL patients (n=11) and healthy individuals (n=16) and comparatively assessed their reserves, proliferative potential, differentiation capacity, and immunoregulatory effects on T- and B-cells. We also evaluated the anti-apoptotic effect of patient-derived MSCs on leukemic cells and studied their cytogenetic characteristics in comparison to BM hematopoietic cells. B-CLL-derived BM MSCs exhibit a similar phenotype, differentiation potential, and ability to suppress T-cell proliferative responses as compared with MSCs from normal controls. Furthermore, they do not carry the cytogenetic abnormalities of the leukemic clone, and they exert a similar anti-apoptotic effect on leukemic cells and healthy donor-derived B-cells, as their normal counterparts. On the other hand, MSCs from B-CLL patients significantly promote normal B-cell proliferation and IgG production, in contrast to healthy-donor-derived MSCs. Furthermore, they have impaired reserves, defective cellular growth due to increased apoptotic cell death and exhibit aberrant production of stromal cell-derived factor 1, B-cell activating factor, a proliferation inducing ligand, and transforming growth factor β1, cytokines that are crucial for the survival/nourishing of the leukemic cells. We conclude that ex vivo expanded B-CLL-derived MSCs harbor intrinsic qualitative and quantitative abnormalities that may be implicated in disease development and/or progression.

Direct inhibition of interleukin-2 receptor alpha-mediated signaling pathway induces G1 arrest and apoptosis in human head-and-neck cancer cells.

PubMed

Kuhn, Deborah J; Dou, Q Ping

2005-05-15

Overexpression of the interleukin-2 receptor (IL-2R) alpha chain in tumor cells is associated with tumor progression and a poor patient prognosis. IL-2Ralpha is responsible for the high affinity binding of the receptor to IL-2, leading to activation of several proliferative and anti-apoptotic intracellular signaling pathways. We have previously shown that human squamous cell carcinoma of a head-and-neck line (PCI-13) genetically engineered to overexpress IL-2Ralpha exhibit increased transforming activity, proliferation, and drug resistance, compared to the vector control cells (J Cell Biochem 2003;89:824-836). In this study, we report that IL-2Ralpha(+) cells express high levels of total and phosphorylated Jak3 protein and are more resistant to apoptosis induced by a Jak3 inhibitor than the control LacZ cells. Furthermore, we used daclizumab, a monoclonal antibody specific to IL-2Ralpha, and determined the effects of IL-2Ralpha inhibition on cell cycle and apoptosis as well as the involvement of potential cell cycle and apoptosis regulatory proteins. We found that daclizumab induces G(1) arrest, associated with down-regulation of cyclin A protein, preferentially in IL-2Ralpha(+) cells, but not in LacZ cells. In addition, daclizumab activates apoptotic death program via Bcl-2 down-regulation preferentially in IL-2Ralpha(+) cells. Finally, daclizumab also sensitizes IL-2Ralpha(+) cells to other apoptotic stimuli, although the effect is moderate. These results indicate that daclizumab inhibits the proliferative potential of IL-2Ralpha(+) cells via inhibition of cell cycle progression and induction of apoptosis.

Submolecular recognition of the C-terminal domain of the heavy chain of botulinum neurotoxin type A by T cells from toxin-treated cervical dystonia patients.

PubMed

Oshima, Minako; Deitiker, Philip; Jankovic, Joseph; Aoki, K Roger; Atassi, M Zouhair

2016-04-01

We determined the T-cell proliferative responses of the peripheral blood lymphocytes (PBL) from 25 botulinum neurotoxin (BoNT)-treated patients to 31 overlapping synthetic peptides encompassing the C-terminal half (residues 855-1296) of BoNT/A heavy chain. Responses of PBL to HC peptides varied among patients. Samples from 14 patients treated solely with BoNT/A recognized 2-13 (average 6.4) peptides/sample at Z>3.0 level. Six peptide regions representing residues 855-873, 1023-1041, 1051-1069, 1093-1111, 1135-1153 and 1247-1265 were frequently recognized by 36-57% of these PBLs. Influence of treatment parameters on T-cell recognition of the peptides was also investigated. Copyright © 2015 Elsevier GmbH. All rights reserved.

Long-Term Outcome of Administration of c-kitPOS Cardiac Progenitor Cells After Acute Myocardial Infarction: Transplanted Cells Do Not Become Cardiomyocytes, Structural and Functional Improvement and Proliferation of Endogenous Cells Persist for at Least One Year

PubMed Central

Tang, Xian-Liang; Li, Qianhong; Rokosh, Gregg; Sanganalmath, Santosh K.; Chen, Ning; Ou, Qinghui; Stowers, Heather; Hunt, Greg; Bolli, Roberto

2016-01-01

RATIONALE Cardiac progenitor cells (CPCs) improve left ventricular (LV) remodeling and function after acute or chronic myocardial infarction (MI). However, the long-term (>5 weeks) effects, potential tumorigenicity, and fate of transplanted CPCs are unknown. OBJECTIVE To assess the outcome of CPC therapy at 1 year. METHODS AND RESULTS Female rats underwent a 90-min coronary occlusion; 4 h after reperfusion, they received intracoronarily vehicle or 1 million male, syngeneic CPCs. One year later, CPC-treated rats exhibited smaller scars and more viable myocardium in the risk region, along with improved LV remodeling and regional and global LV function. No tumors were observed. Some transplanted (Y-chromosomePOS) CPCs (or their progeny) persisted and continued to proliferate, but they failed to acquire a mature cardiomyocyte phenotype and were too few (4-8% of nuclei) to account for the benefits of CPC therapy. Surprisingly, CPC transplantation triggered a prolonged proliferative response of endogenous cells, resulting in increased formation of endothelial cells and Y-chromosomeNEG CPCs for 12 months and increased formation, for at least 7 months, of small cells that expressed cardiomyocytic proteins (α-sarcomeric actin) but did not have a mature cardiomyocyte phenotype. CONCLUSIONS The beneficial effects of CPCs on LV remodeling and dysfunction are sustained for at least 1 year, and thus are likely to be permanent. Since transplanted CPCs do not differentiate into mature myocytes, their major mechanism of action must involve paracrine actions. These paracrine mechanisms could be very prolonged because some CPCs engraft, proliferate, and persist at 1 year. This is the first report that transplantation of any cell type in the heart induces a proliferative response that lasts at least 1 year. The results strongly support the safety and clinical utility of CPC therapy. PMID:26838790

Quantitative Receptor-Based Imaging of Tumor Proliferation with the Sigma-2 Ligand [18F]ISO-1

PubMed Central

Shoghi, Kooresh I.; Xu, Jinbin; Su, Yi; He, June; Rowland, Douglas; Yan, Ying; Garbow, Joel R.; Tu, Zhude; Jones, Lynne A.; Higashikubo, Ryuji; Wheeler, Kenneth T.; Lubet, Ronald A.; Mach, Robert H.; You, Ming

2013-01-01

The sigma-2 receptor is expressed in higher density in proliferating (P) tumor cells versus quiescent (Q) tumor cells, thus providing an attractive target for imaging the proliferative status (i.e., P:Q ratio) of solid tumors. Here we evaluate the utility of the sigma-2 receptor ligand 2-(2-[18F]fluoroethoxy)-N-(4-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)butyl)-5-methyl-benzamide, [18F]ISO-1, in two different rodent models of breast cancer. In the first study, small animal Positron Emission Tomography (PET) imaging studies were conducted with [18F]ISO-1 and 18FDG in xenografts of mouse mammary tumor 66 and tracer uptake was correlated with the in vivo P:Q ratio determined by flow cytometric measures of BrdU-labeled tumor cells. The second model utilized a chemically-induced (N-methyl-N-nitrosourea [MNU]) model of rat mammary carcinoma to correlate measures of [18F]ISO-1 and FDG uptake with MR-based volumetric measures of tumor growth. In addition, [18F]ISO-1 and FDG were used to assess the response of MNU-induced tumors to bexarotene and Vorozole therapy. In the mouse mammary 66 tumors, a strong linear correlation was observed between the [18F]ISO-1 tumor: background ratio and the proliferative status (P:Q ratio) of the tumor (R = 0.87). Similarly, measures of [18F]ISO-1 uptake in MNU-induced tumors significantly correlated (R = 0.68, P<0.003) with changes in tumor volume between consecutive MR imaging sessions. Our data suggest that PET studies of [18F]ISO-1 provide a measure of both the proliferative status and tumor growth rate, which would be valuable in designing an appropriate treatment strategy. PMID:24073202

Biodegradable composite scaffolds: a strategy to modulate stem cell behaviour.

PubMed

Armentano, Ilaria; Fortunati, Elena; Mattioli, Samantha; Rescignano, Nicolatta; Kenny, José M

2013-04-01

The application of new biomaterial technologies offers the potential to direct the stem cell fate, targeting the delivery of cells and reducing immune rejection, thereby supporting the development of regenerative medicine. Cells respond to their surrounding structure and with nanostructures exhibit unique proliferative and differentiation properties. This review presents the relevance, the promising perspectives and challenges of current biodegradable composite scaffolds in terms of material properties, processing technology and surface modification, focusing on significant recent patents in these fields. It has been reported how biodegradable porous composite scaffolds can be engineered with initial properties that reproduce the anisotropy, viscoelasticity, tension-compression non-linearity of different tissues by introducing specific nanostructures. Moreover the modulation of electrical, morphological, surface and topographic scaffold properties enables specific stem cell response. Recent advances in nanotechnology have allowed to engineer novel biomaterials with these complexity levels. Understanding the specific biological response triggered by various aspects of the fibrous environment is important in guiding the design and engineering of novel substrates that mimic the native cell matrix interactions in vivo.

Evaluation of an ex vivo murine local lymph node assay: multiple endpoint comparison.

PubMed

Piccotti, Joseph R; Knight, Stephanie A; Gillhouse, Kimberly; Lagattuta, Mark S; Bleavins, Michael R

2006-01-01

The local lymph node assay (LLNA) is used to assess the skin sensitization potential of chemicals. In the standard assay, mice are treated topically on the dorsum of both ears with test substance for 3 days. Following 2 days of rest, the initiation of the hypersensitivity response is evaluated by injecting (3)H-thymidine into a tail vein, and then measuring the levels of radioisotope incorporated into the DNA of lymph node cells draining the ears. In the current study, BALB/c mice were treated with the contact sensitizers hexylcinnamic aldehyde (HCA) and oxazolone, and the nonsensitizer methyl salicylate. The proliferative response of lymph node cells was evaluated in an ex vivo assay, in which isolated cells were cultured in vitro with (3)H-thymidine. Treatment of mice with HCA at 5-50% resulted in concentration-related increases in (3)H-thymidine incorporation, with stimulation indices ranging from 3 to 14. Low animal-to-animal variability was seen in three replicate assays testing HCA at 25%. As anticipated, the proliferative response induced by the potent sensitizer oxazolone at 0.25% was greater than HCA at all concentrations tested. Stimulation indices of 1.5 and 3 were seen in two independent experiments with methyl salicylate. These equivocal findings were likely due to the irritancy properties of the compound. Importantly, measuring ex vivo (3)H-thymidine incorporation was more sensitive than evaluating lymph node weight and cellularity, and in vitro bromodeoxyuridine incorporation. Furthermore, the results of the ex vivo LLNA were comparable to the standard assay. This study provided evidence that supports the use of an ex vivo LLNA for hazard assessment of contact hypersensitivity. Copyright 2006 John Wiley & Sons, Ltd.

Recombinant adenovirus-p21 attenuates proliferative responses associated with excessive scarring.

PubMed

Gu, Danling; Atencio, Isabella; Kang, David W; Looper, L David; Ahmed, C M I; Levy, Alina; Maneval, Dan; Zepeda, Monica L

2005-01-01

Excessive cutaneous scarring is an important clinical disorder resulting in adverse tissue growth and function as well as undesirable cosmetic appearance. p21WAF-1/Cip-1 is a cyclin-dependent kinase inhibitor that blocks cell cycle progression and inhibits cell proliferation. We used a recombinant adenovirus containing the human p21WAF-1/Cip-1 cDNA (rAd-p21) to evaluate proliferative responses in skin models. In vitro dose-response studies using primary human dermal fibroblasts resulted in a dose-dependent expression of p21WAF-1/Cip-1 protein and a 3- to 80-fold reduction in cell proliferation as measured by 5-bromodeoxyuridine incorporation. Further, rAd-p21 reduced type I procollagen production when compared to control virus. A rat polyvinyl alcohol sponge model was used to determine rAd-p21 effects on granulation tissue formation in vivo. Sponges pretreated with a granulation tissue stimulator, rAd-PDGF-B and subsequently rAd-p21 on a second injection, showed a p21WAF-1/Cip-1 specific dose-dependent decrease in percent granulation fill as the rAd-p21 dose increased (p < 0.001). Immunohistochemistry identified human p21WAF-1/Cip-1 expression in sponges treated with rAd-p21 5 days postinjection. Additionally, 5-bromodeoxyuridine and Ki67 staining in sponges treated with rAd-p21 showed a significant decrease in proliferation when compared to rAd-platelet-derived growth factor-B alone or vehicle control groups (p < 0.01). These data support the utility of p21WAF-1/Cip-1 in targeting hyperproliferative disorders of the skin.

Effects of vitamin K3 and K5 on proliferation, cytokine production, and regulatory T cell-frequency in human peripheral-blood mononuclear cells.

PubMed

Hatanaka, Hiroshige; Ishizawa, Hitomi; Nakamura, Yurie; Tadokoro, Hiroko; Tanaka, Sachiko; Onda, Kenji; Sugiyama, Kentaro; Hirano, Toshihiko

2014-03-18

The effects of vitamin K (VK) derivatives VK3 and VK5 on human immune cells have not been extensively investigated. We examined the effects of VK3 and VK5 on proliferation, apoptosis, cytokine production, and CD4+CD25+Foxp3+ regulatory T (Treg) cell-frequency in human peripheral blood mononuclear cells (PBMCs) activated by T cell mitogen in vitro. Anti-proliferative effects of VK3 and VK5 on T-cell mitogen activated PBMCs were assessed by WST assay procedures. Apoptotic cells were determined as Annexin V positive/propidium iodide (PI) negative cells. Cytokine concentrations in the supernatant of the culture medium were measured with bead-array procedures followed by analysis with flow cytometry. The CD4+CD25+Foxp3+Treg cells in mitogen-activated PBMCs were stained with fluorescence-labeled specific antibodies followed by flow cytometry. VK3 and VK5 suppressed the mitogen-activated proliferation of PBMCs significantly at 10-100μM (p<0.05). The data also suggest that VK3 and VK5 promote apoptosis in the mitogen-activated T cells. VK3 and VK5 significantly inhibited the production of tumor necrosis factor (TNF) α, interleukin (IL)-4, -6, and -10 from the activated PBMCs at 10-100μM (p<0.05). In contrast, VK3 and VK5 significantly increased Treg cell-frequency in the activated PBMCs at concentrations more than 10μM (p<0.001). Our data suggest that VK3 and VK5 attenuate T cell mediated immunity by inhibiting the proliferative response and inducing apoptosis in activated T cells. Copyright © 2014 Elsevier Inc. All rights reserved.

p21 in cancer: intricate networks and multiple activities.

PubMed

Abbas, Tarek; Dutta, Anindya

2009-06-01

One of the main engines that drives cellular transformation is the loss of proper control of the mammalian cell cycle. The cyclin-dependent kinase inhibitor p21 (also known as p21WAF1/Cip1) promotes cell cycle arrest in response to many stimuli. It is well positioned to function as both a sensor and an effector of multiple anti-proliferative signals. This Review focuses on recent advances in our understanding of the regulation of p21 and its biological functions with emphasis on its p53-independent tumour suppressor activities and paradoxical tumour-promoting activities, and their implications in cancer.

Curcumin conjugated with PLGA potentiates sustainability, anti-proliferative activity and apoptosis in human colon carcinoma cells.

PubMed

Waghela, Bhargav N; Sharma, Anupama; Dhumale, Suhashini; Pandey, Shashibahl M; Pathak, Chandramani

2015-01-01

Curcumin, an ingredient of turmeric, exhibits a variety of biological activities such as anti-inflammatory, anti-atherosclerotic, anti-proliferative, anti-oxidant, anti-cancer and anti-metastatic. It is a highly pleiotropic molecule that inhibits cell proliferation and induces apoptosis in cancer cells. Despite its imperative biological activities, chemical instability, photo-instability and poor bioavailability limits its utilization as an effective therapeutic agent. Therefore, enhancing the bioavailability of curcumin may improve its therapeutic index for clinical setting. In the present study, we have conjugated curcumin with a biodegradable polymer Poly (D, L-lactic-co-glycolic acid) and evaluated its apoptotic potential in human colon carcinoma cells (HCT 116). The results show that curcumin-PLGA conjugate efficiently inhibits cell proliferation and cell survival in human colon carcinoma cells as compared to native curcumin. Additionally, curcumin conjugated with PLGA shows improved cellular uptake and exhibits controlled release at physiological pH as compared to native curcumin. The curcumin-PLGA conjugate efficiently activates the cascade of caspases and promotes intrinsic apoptotic signaling. Thus, the results suggest that conjugation potentiates the sustainability, anti-proliferative and apoptotic activity of curcumin. This approach could be a promising strategy to improve the therapeutic index of cancer therapy.

T cell function in tuatara (Sphenodon punctatus).

PubMed

Burnham, D Kim; Keall, Susan N; Nelson, Nicola J; Daugherty, Charles H

2005-05-01

Tuatara are the sole survivors of an entire order of reptiles that thrived during the age of the dinosaurs. Therefore, knowledge of their physiology is critical to understanding the phylogeny of reptiles. Previous studies of the immune system of the tuatara did not assess T cell function. We analyzed T cell function among six captive tuatara by assessing concanavalin A (Con A), phytohemagglutinin (PHA) and mixed lymphocyte reaction (MLR) induced T cell proliferation. Peripheral blood mononuclear cells from six out of six and four out of four tuatara tested exhibited significant proliferative responses to Con A and PHA, respectively, as measured by an MTT reduction assay. A lower level of proliferation was detected in an MLR. However, Con A activated lymphocytes were not cytotoxic for a xenogeneic murine mastocytoma cell line (P815).

Clonal analysis of T-cell responses to herpes simplex virus: isolation, characterization and antiviral properties of an antigen-specific helper T-cell clone.

PubMed Central

Leung, K N; Nash, A A; Sia, D Y; Wildy, P

1984-01-01

A herpes simplex virus (HSV)-specific long-term T-cell clone has been established from the draining lymph node cells of BALB/c mice; the cells required repeated in vitro restimulation with UV-irradiated virus. The established T-cell clone expresses the Thy-1 and Lyt-1+2,3- surface antigens. For optimal proliferation of the cloned cells, both the presence of specific antigen and an exogenous source of T-cell growth factor are required. The proliferative response of the cloned T cells was found to be virus-specific but it did not distinguish between HSV-1 and HSV-2. Adoptive cell transfer of the cloned T cells helped primed B cells to produce anti-herpes antibodies: the response was antigen-specific and cell dose-dependent. The clone failed to produce a significant DTH reaction in vivo, but did produce high levels of macrophage-activating factor. Furthermore, the T-cell clone could protect from HSV infection, as measured by a reduction in local virus growth, and by enhanced survival following the challenge of mice with a lethal dose of virus. The mechanism(s) whereby this clone protects in vivo is discussed. PMID:6209206

A p53-dependent response limits the viability of mammalian haploid cells

PubMed Central

Olbrich, Teresa; Mayor-Ruiz, Cristina; Vega-Sendino, Maria; Gomez, Carmen; Ortega, Sagrario; Ruiz, Sergio; Fernandez-Capetillo, Oscar

2017-01-01

The recent development of haploid cell lines has facilitated forward genetic screenings in mammalian cells. These lines include near-haploid human cell lines isolated from a patient with chronic myelogenous leukemia (KBM7 and HAP1), as well as haploid embryonic stem cells derived from several organisms. In all cases, haploidy was shown to be an unstable state, so that cultures of mammalian haploid cells rapidly become enriched in diploids. Here we show that the observed diploidization is due to a proliferative disadvantage of haploid cells compared with diploid cells. Accordingly, single-cell–sorted haploid mammalian cells maintain the haploid state for prolonged periods, owing to the absence of competing diploids. Although the duration of interphase is similar in haploid and diploid cells, haploid cells spend longer in mitosis, indicative of problems in chromosome segregation. In agreement with this, a substantial proportion of the haploids die at or shortly after the last mitosis through activation of a p53-dependent cytotoxic response. Finally, we show that p53 deletion stabilizes haploidy in human HAP1 cells and haploid mouse embryonic stem cells. We propose that, similar to aneuploidy or tetraploidy, haploidy triggers a p53-dependent response that limits the fitness of mammalian cells. PMID:28808015

T-helper cell receptors from long-term survivors after telomerase cancer vaccination for use in adoptive cell therapy.

PubMed

Kyte, Jon Amund; Gaudernack, Gustav; Faane, Anne; Lislerud, Kari; Inderberg, Else Marit; Brunsvig, Paal; Aamdal, Steinar; Kvalheim, Gunnar; Wälchli, Sébastien; Pule, Martin

2016-01-01

We herein report retargeting of T-helper (Th) cells against the universal cancer antigen telomerase for use in adoptive cell therapy. The redirected Th cells may counter tumor tolerance, transform the inflammatory milieu, and induce epitope spreading and cancer senescence. We have previously conducted a series of trials evaluating vaccination with telomerase peptides. From long-term survivors, we isolated >100 CD4 + Th-cell clones recognizing telomerase epitopes. The clones were characterized with regard to HLA restriction, functional avidity, fine specificity, proliferative capacity, cytokine profile, and recognition of naturally processed epitopes. DP4 is the most prevalent HLA molecule worldwide. Two DP4-restricted T-cell clones with different functional avidity, C13 and D71, were selected for molecular T-cell receptor (TCR) cloning. Both clones showed a high proliferative capacity, recognition of naturally processed telomerase epitopes, and a polyfunctional and Th1-weighted cytokine profile. TCR C13 and D71 were cloned into the retroviral vector MP71 together with the compact and GMP-applicable marker/suicide gene RQR8. Both TCRs were expressed well in recipient T cells after PBMC transduction. The transduced T cells co-expressed RQR8 and acquired the desired telomerase specificity, with a polyfunctional response including production of TNFa, IFNγ, and CD107a. Interestingly, the DP4-restricted TCRs were expressed and functional both in CD4 + and CD8 + T cells. The findings demonstrate that the cloned TCRs confer recipient T cells with the desired hTERT-specificity and functionality. We hypothesize that adoptive therapy with Th cells may offer a powerful novel approach for overcoming tumor tolerance and synergize with other forms of immunotherapy.

Phylogeny of Immune Recognition: Processing and Presentation of Structurally Defined Proteins in Channel Catfish Immune Responses

PubMed Central

Vallejo, Abbe N.; Miller, Norman W.

1991-01-01

This work was undertaken to investigate whether or not antigen processing and presentation are important in channel catfish in vitro secondary immune responses elicited with structurally defined proteins, namely, pigeon heart cytochrome C (pCytC), hen egg lysozyme, and horse myoglobin. The use of in vitro antigen-pulsed and fixed B cells or monocytes as antigen presenting cells (APC) resulted in autologous peripheral blood leukocytes (PBL) responding with vigorous proliferation and antibody production in vitro. In addition, several long-term catfish monocyte lines have been found to function as efficient APC with autologous but not allogeneic responders. Subsequent separation of the responding PBL into sIg- (T-cell-enriched) and B (sIg+) cell subsets showed that both underwent proliferative responses to antigen-pulsed and fixed APC. Moreover, allogeneic cells used as APC were found to induce only strong mixed leukocyte reactions without specific in vitro antibody production. Initial attempts at identifying the immunogenic region(s) of the protein antigens for catfish indicated there are two such regions for pCytC, namely, peptides 66-80 and 81-104. PMID:1668258

SAP modulates B cell functions in a genetic background-dependent manner.

PubMed

Detre, Cynthia; Yigit, Burcu; Keszei, Marton; Castro, Wilson; Magelky, Erica M; Terhorst, Cox

2013-06-01

Mutations affecting the SLAM-associated protein (SAP) are responsible for the X-linked lympho-proliferative syndrome (XLP), a severe primary immunodeficiency syndrome with disease manifestations that include fatal mononucleosis, B cell lymphoma and dysgammaglobulinemia. It is well accepted that insufficient help by SAP-/- CD4+ T cells, in particular during the germinal center reaction, is a component of dysgammaglobulinemia in XLP patients and SAP-/- animals. It is however not well understood whether in XLP patients and SAP-/- mice B cell functions are affected, even though B cells themselves do not express SAP. Here we report that B cell intrinsic responses to haptenated protein antigens are impaired in SAP-/- mice and in Rag-/- mice into which B cells derived from SAP-/- mice together with wt CD4+ T cells had been transferred. This impaired B cells functions are in part depending on the genetic background of the SAP-/- mouse, which affects B cell homeostasis. Surprisingly, stimulation with an agonistic anti-CD40 causes strong in vivo and in vitro B cell responses in SAP-/- mice. Taken together, the data demonstrate that genetic factors play an important role in the SAP-related B cell functions. The finding that anti-CD40 can in part restore impaired B cell responses in SAP-/- mice, suggests potentially novel therapeutic interventions in subsets of XLP patients. Copyright © 2013 Elsevier B.V. All rights reserved.

Synthesis, anti-proliferative activity, SAR study, and preliminary in vivo toxicity study of substituted N,N'-bis(arylmethyl)benzimidazolium salts against a panel of non-small cell lung cancer cell lines.

PubMed

Shelton, Kerri L; DeBord, Michael A; Wagers, Patrick O; Southerland, Marie R; Williams, Travis M; Robishaw, Nikki K; Shriver, Leah P; Tessier, Claire A; Panzner, Matthew J; Youngs, Wiley J

2017-01-01

A series of N,N'-bis(arylmethyl)benzimidazolium salts have been synthesized and evaluated for their in vitro anti-cancer activity against select non-small cell lung cancer cell lines to create a structure activity relationship profile. The results indicate that hydrophobic substituents on the salts increase the overall anti-proliferative activity. Our data confirms that naphthylmethyl substituents at the nitrogen atoms (N 1 (N 3 )) and highly lipophilic substituents at the carbon atoms (C 2 and C 5 (C 6 )) can generate benzimidazolium salts with anti-proliferative activity that is comparable to that of cisplatin. The National Cancer Institute's Developmental Therapeutics Program tested 1, 3-5, 10, 11, 13-18, 20-25, and 28-30 in their 60 human tumor cell line screen. Results were supportive of data observed in our lab. Compounds with hydrophobic substituents have higher anti-cancer activity than compounds with hydrophilic substituents. Copyright © 2016 Elsevier Ltd. All rights reserved.

Proliferative effects of apical, but not basal, matrix metalloproteinase-7 activity in polarized MDCK cells

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

Harrell, Permila C.; McCawley, Lisa J.; Fingleton, Barbara

2005-02-15

Matrix metalloproteinase-7 (MMP-7) is primarily expressed in glandular epithelium. Therefore, its mechanism of action may be influenced by its regulated vectorial release to either the apical and/or basolateral compartments, where it would act on its various substrates. To gain a better understanding of where MMP-7 is released in polarized epithelium, we have analyzed its pattern of secretion in polarized MDCK cells expressing stably transfected human MMP-7 (MDCK-MMP-7), and HCA-7 and Caco2 human colon cancer cell lines. In all cell lines, latent MMP-7 was secreted to both cellular compartments, but was 1.5- to 3-fold more abundant in the basolateral compartment asmore » compared to the apical. However, studies in the MDCK system demonstrated that MMP-7 activity was 2-fold greater in the apical compartment of MDCK-MMP-7{sup HIGH}-polarized monolayers, which suggests the apical co-release of an MMP-7 activator. In functional assays, MMP-7 over-expression increased cell saturation density as a result of increased cell proliferation with no effect on apoptosis. Apical MMP-7 activity was shown to be responsible for the proliferative effect, which occurred, as demonstrated by media transfer experiments, through cleavage of an apical substrate and not through the generation of a soluble factor. Taken together, our findings demonstrate the importance of MMP-7 secretion in relation to its mechanism of action when expressed in a polarized epithelium.« less

Glycogen synthase kinase 3 has a limited role in cell cycle regulation of cyclin D1 levels.

PubMed

Yang, Ke; Guo, Yang; Stacey, William C; Harwalkar, Jyoti; Fretthold, Jonathan; Hitomi, Masahiro; Stacey, Dennis W

2006-08-30

The expression level of cyclin D1 plays a vital role in the control of proliferation. This protein is reported to be degraded following phosphorylation by glycogen synthase kinase 3 (GSK3) on Thr-286. We recently showed that phosphorylation of Thr-286 is responsible for a decline in cyclin D1 levels during S phase, an event required for efficient DNA synthesis. These studies were undertaken to test the possibility that phosphorylation by GSK3 is responsible for the S phase specific decline in cyclin D1 levels, and that this event is regulated by the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway which controls GSK3. We found, however, that neither PI3K, AKT, GSK3, nor proliferative signaling activity in general is responsible for the S phase decline in cyclin D1 levels. In fact, the activity of these signaling kinases does not vary through the cell cycle of proliferating cells. Moreover, we found that GSK3 activity has little influence over cyclin D1 expression levels during any cell cycle phase. Inhibition of GSK3 activity by siRNA, LiCl, or other chemical inhibitors failed to influence cyclin D1 phosphorylation on Thr-286, even though LiCl efficiently blocked phosphorylation of beta-catenin, a known substrate of GSK3. Likewise, the expression of a constitutively active GSK3 mutant protein failed to influence cyclin D1 phosphorylation or total protein expression level. Because we were unable to identify any proliferative signaling molecule or pathway which is regulated through the cell cycle, or which is able to influence cyclin D1 levels, we conclude that the suppression of cyclin D1 levels during S phase is regulated by cell cycle position rather than signaling activity. We propose that this mechanism guarantees the decline in cyclin D1 levels during each S phase; and that in so doing it reduces the likelihood that simple over expression of cyclin D1 can lead to uncontrolled cell growth.

Zoster Vaccination Increases the Breadth of CD4+ T Cells Responsive to Varicella Zoster Virus

PubMed Central

Laing, Kerry J.; Russell, Ronnie M.; Dong, Lichun; Schmid, D. Scott; Stern, Michael; Magaret, Amalia; Haas, Jürgen G.; Johnston, Christine; Wald, Anna; Koelle, David M.

2015-01-01

Background. The live, attenuated varicella vaccine strain (vOka) is the only licensed therapeutic vaccine. Boost of varicella zoster virus (VZV)–specific cellular immunity is a likely mechanism of action. We examined memory CD4+ T-cell responses to each VZV protein at baseline and after zoster vaccination. Methods. Serial blood samples were collected from 12 subjects vaccinated with Zostavax and immunogenicity confirmed by ex vivo VZV-specific T-cell and antibody assays. CD4+ T-cell lines enriched for VZV specificity were generated and probed for proliferative responses to every VZV protein and selected peptide sets. Results. Zoster vaccination increased the median magnitude (2.3-fold) and breadth (4.2-fold) of VZV-specific CD4+ T cells one month post-vaccination. Both measures declined by 6 months. The most prevalent responses at baseline included VZV open reading frames (ORFs) 68, 4, 37, and 63. After vaccination, responses to ORFs 40, 67, 9, 59, 12, 62, and 18 were also prevalent. The immunogenicity of ORF9 and ORF18 were confirmed using peptides, defining a large number of discrete CD4 T-cell epitopes. Conclusions. The breadth and magnitude of the VZV-specific CD4+ T-cell response increase after zoster vaccination. In addition to glycoprotein E (ORF68), we identified antigenic ORFs that may be useful components of subunit vaccines. PMID:25784732

Neural stem cells in the immature, but not the mature, subventricular zone respond robustly to traumatic brain injury.

PubMed

Goodus, Matthew T; Guzman, Alanna M; Calderon, Frances; Jiang, Yuhui; Levison, Steven W

2015-01-01

Pediatric traumatic brain injury is a significant problem that affects many children each year. Progress is being made in developing neuroprotective strategies to combat these injuries. However, investigators are a long way from therapies to fully preserve injured neurons and glia. To restore neurological function, regenerative strategies will be required. Given the importance of stem cells in repairing damaged tissues and the known persistence of neural precursors in the subventricular zone (SVZ), we evaluated regenerative responses of the SVZ to a focal brain lesion. As tissues repair more slowly with aging, injury responses of male Sprague Dawley rats at 6, 11, 17, and 60 days of age and C57Bl/6 mice at 14 days of age were compared. In the injured immature animals, cell proliferation in the dorsolateral SVZ more than doubled by 48 h. By contrast, the proliferative response was almost undetectable in the adult brain. Three approaches were used to assess the relative numbers of bona fide neural stem cells, as follows: the neurosphere assay (on rats injured at postnatal day 11, P11), flow cytometry using a novel 4-marker panel (on mice injured at P14) and staining for stem/progenitor cell markers in the niche (on rats injured at P17). Precursors from the injured immature SVZ formed almost twice as many spheres as precursors from uninjured age-matched brains. Furthermore, spheres formed from the injured brain were larger, indicating that the neural precursors that formed these spheres divided more rapidly. Flow cytometry revealed a 2-fold increase in the percentage of stem cells, a 4-fold increase in multipotential progenitor-3 cells and a 2.5-fold increase in glial-restricted progenitor-2/multipotential-3 cells. Analogously, there was a 2-fold increase in the mitotic index of nestin+/Mash1- immunoreactive cells within the immediately subependymal region. As the early postnatal SVZ is predominantly generating glial cells, an expansion of precursors might not necessarily lead to the production of many new neurons. On the contrary, many BrdU+/doublecortin+ cells were observed streaming out of the SVZ into the neocortex 2 weeks after injuries to P11 rats. However, very few new mature neurons were seen adjacent to the lesion 28 days after injury. Altogether, these data indicate that immature SVZ cells mount a more robust proliferative response to a focal brain injury than adult cells, which includes an expansion of stem cells, primitive progenitors and neuroblasts. Nonetheless, this regenerative response does not result in significant neuronal replacement, indicating that new strategies need to be implemented to retain the regenerated neurons and glia that are being produced. © 2014 S. Karger AG, Basel.

Rotation is the primary motion of paired human epidermal keratinocytes.

PubMed

Tate, Sota; Imai, Matome; Matsushita, Natsuki; Nishimura, Emi K; Higashiyama, Shigeki; Nanba, Daisuke

2015-09-01

Collective motion of keratinocytes is involved in morphogenesis, homeostasis, and wound healing of the epidermis. Yet how the collective motion of keratinocytes emerges from the behavior of individual cells is still largely unknown. The aim of this study was to find the cellular behavior that links single and collective motion of keratinocytes. We investigated the behavior of two-cell colonies of HaCaT keratinocytes by a combination of time-lapse imaging and image processing. The two-cell colonies of HaCaT cells were formed as a contacted pair of keratinocyte clones. Image analysis and cell culture experiments revealed that the rotational speed of two-cell colonies was positively associated with their proliferative capacity. α6 integrin was required for the rotational motion of two-cell keratinocyte colonies. We also confirmed that two-cell colonies of keratinocytes predominantly exhibited the rotational, but not translational, motion, two modes of motion in a contact pair of rotating objects. The rotational motion is the primary motion of two-cell keratinocyte colonies and its speed is positively associated with their proliferative capacity. This study suggests that the assembly of rotating keratinocytes generates the collective motion of proliferative keratinocytes during morphogenesis and wound healing of the epidermis. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Characterizing invading glioma cells based on IDH1-R132H and Ki-67 immunofluorescence.

PubMed

Sabit, Hemragul; Nakada, Mitsutoshi; Furuta, Takuya; Watanabe, Takuya; Hayashi, Yutaka; Sato, Hiroshi; Kato, Yukinari; Hamada, Jun-ichiro

2014-10-01

Glioma, the most common primary brain tumor, is characterized by proliferative-invasive growth. However, the detailed biological characteristics of invading glioma cells remain to be elucidated. A monoclonal antibody (clone HMab-1) that specifically and sensitively recognizes the isocitrate dehydrogenase-1 (IDH1) protein carrying the R132H mutation can identify invading glioma cells by immunostaining. To investigate the degree of invasion in gliomas of distinct grades and the proliferative capacity of the invading cells, immunofluorescent staining was conducted using antibodies against IDH1-R132H and Ki-67 on 11 surgical and autopsy specimens of the tumor core and the invading area. Higher numbers of IDH1-R132H-positive cells in the invading area correlated with a higher tumor grade. Double staining for IDH1-R132H and Ki-67 demonstrated that most invading cells that expressed IDH1-R132H were not stained by the Ki-67 antibody, and the ratio of Ki-67-positive cells among IDH1-R132H-positive cells was significantly lower in the invasion area than in the tumor core in all grades of glioma. These data suggest that higher grade gliomas have a greater invasive potential and that invading cells possess low proliferative capacity.

Gingival wound healing: an essential response disturbed by aging?

PubMed

Smith, P C; Cáceres, M; Martínez, C; Oyarzún, A; Martínez, J

2015-03-01

Gingival wound healing comprises a series of sequential responses that allow the closure of breaches in the masticatory mucosa. This process is of critical importance to prevent the invasion of microbes or other agents into tissues, avoiding the establishment of a chronic infection. Wound healing may also play an important role during cell and tissue reaction to long-term injury, as it may occur during inflammatory responses and cancer. Recent experimental data have shown that gingival wound healing is severely affected by the aging process. These defects may alter distinct phases of the wound-healing process, including epithelial migration, granulation tissue formation, and tissue remodeling. The cellular and molecular defects that may explain these deficiencies include several biological responses such as an increased inflammatory response, altered integrin signaling, reduced growth factor activity, decreased cell proliferation, diminished angiogenesis, reduced collagen synthesis, augmented collagen remodeling, and deterioration of the proliferative and differentiation potential of stem cells. In this review, we explore the cellular and molecular basis of these defects and their possible clinical implications. © International & American Associations for Dental Research 2014.

Breast cancer cell-associated endopeptidase EC 24.11 modulates proliferative response to bombesin.

PubMed

Burns, D M; Walker, B; Gray, J; Nelson, J

1999-01-01

We have investigated the production, growth and inactivation of gastrin-releasing peptide (GRP)-like peptides in human breast cancer cell lines. Radioimmunoassay detected GRP-like immunoreactivity (GRP-LI) in T47D breast cancer cells but not in the conditioned medium, indicating rapid clearance. No GRP-LI was found in the ZR-75-1 or MDA-MB-436 cells or their conditioned medium. High-performance liquid chromatography (HPLC) analysis of the GRP-LI in the T47D cells revealed a major peak, which co-eluted with GRP(18-27), and a minor more hydrophilic peak. In vitro stimulation of T47D cell growth by bombesin (BN) was enhanced to 138% of control levels (bombesin alone) by the addition of the selective endopeptidase EC 3.4.24.11 inhibitor phosphoramidon (0.1 ng ml(-1)). Fluorogenic analysis using whole cells confirmed low levels of this phosphoramidon-sensitive enzyme on the T47D cells. This enzyme, previously unreported in human breast cancer cells, significantly modulates both T47D growth and its response to BN-induced growth.

Breast cancer cell-associated endopeptidase EC 24.11 modulates proliferative response to bombesin

PubMed Central

Burns, D M; Walker, B; Gray, J; Nelson, J

1999-01-01

We have investigated the production, growth and inactivation of gastrin-releasing peptide (GRP)-like peptides in human breast cancer cell lines. Radioimmunoassay detected GRP-like immunoreactivity (GRP-LI) in T47D breast cancer cells but not in the conditioned medium, indicating rapid clearance. No GRP-LI was found in the ZR-75-1 or MDA-MB-436 cells or their conditioned medium. High-performance liquid chromatography (HPLC) analysis of the GRP-LI in the T47D cells revealed a major peak, which co-eluted with GRP18–27, and a minor more hydrophilic peak. In vitro stimulation of T47D cell growth by bombesin (BN) was enhanced to 138% of control levels (bombesin alone) by the addition of the selective endopeptidase EC 3.4.24.11 inhibitor phosphoramidon (0.1 ng ml−;1). Fluorogenic analysis using whole cells confirmed low levels of this phosphoramidon-sensitive enzyme on the T47D cells. This enzyme, previously unreported in human breast cancer cells, significantly modulates both T47D growth and its response to BN-induced growth. © 1999 Cancer Research Campaign PMID:9888460

Acquired resistance to rechallenge injury in rats recovered from subclinical renal damage with uranyl acetate-Importance of proliferative activity of tubular cells

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

Sun, Yuan; Fujigaki, Yoshihide, E-mail: yf0516@hama-med.ac.j; Sakakima, Masanori

Animals recovered from acute renal failure are resistant to subsequent insult. We investigated whether rats recovered from mild proximal tubule (PT) injury without renal dysfunction (subclinical renal damage) acquire the same resistance. Rats 14 days after recovering from subclinical renal damage, which was induced by 0.2 mg/kg of uranyl acetate (UA) (sub-toxic dose), were rechallenged with 4 mg/kg of UA (nephrotoxic dose). Fate of PT cells and renal function were examined in response to nephrotoxic dose of UA. All divided cells after sub-toxic dose of UA insult were labeled with bromodeoxyuridine (BrdU) for 14 days then the number of PTmore » cells with or without BrdU-labeling was counted following nephrotoxic dose of UA insult. Rats recovered from subclinical renal damage gained resistance to nephrotoxic dose of UA with reduced renal dysfunction, less severity of peak damage (necrotic and TUNEL+ apoptotic cells) and accelerated PT cell proliferation, but with earlier peak of PT damage. The decrease in number of PT cells in the early phase of rechallenge injury with nephrotoxic UA was more in rats pretreated with sub-toxic dose of UA than vehicle pretreated rats. The exaggerated loss of PT cells was mainly caused by the exaggerated loss of BrdU+ divided cells. In contrast, accelerated cell proliferation in rats recovered from sub-toxic dose of UA was observed mainly in BrdU- non-divided cells. The findings suggest that rats recovered from subclinical renal damage showed partial acquired resistance to nephrotoxic insult. Accelerated recovery with increased proliferative activity of non-divided PT cells after subclinical renal damage may mainly contribute to acquired resistance.« less

Proteomics of the human endometrial glandular epithelium and stroma from the proliferative and secretory phases of the menstrual cycle.

PubMed

Hood, Brian L; Liu, Baoquan; Alkhas, Addie; Shoji, Yutaka; Challa, Rusheeswar; Wang, Guisong; Ferguson, Susan; Oliver, Julie; Mitchell, Dave; Bateman, Nicholas W; Zahn, Christopher M; Hamilton, Chad A; Payson, Mark; Lessey, Bruce; Fazleabas, Asgerally T; Maxwell, G Larry; Conrads, Thomas P; Risinger, John I

2015-04-01

Despite its importance in reproductive biology and women's health, a detailed molecular-level understanding of the human endometrium is lacking. Indeed, no comprehensive studies have been undertaken to elucidate the important protein expression differences between the endometrial glandular epithelium and surrounding stroma during the proliferative and midsecretory phases of the menstrual cycle. We utilized laser microdissection to harvest epithelial cells and stromal compartments from proliferative and secretory premenopausal endometrial tissue and performed a global, quantitative mass spectrometry-based proteomics analysis. This analysis identified 1224 total proteins from epithelial cells, among which 318 were differentially abundant between the proliferative and secretory phases (q < 0.05), and 1005 proteins from the stromal compartments, 19 of which were differentially abundant between the phases (q < 0.05). Several proteins were chosen for validation by immunohistochemistry in an independent set of uterine tissues, including carboxypeptidase M, tenascin C, neprilysin, and ectonucleotide pyrophosphatase/phosphodiesterase family member 3 (ENPP3). ENPP3, which was elevated in epithelial glandular cells in the secretory phase, was confirmed to be elevated in midsecretory-phase baboon uterine lavage samples and also observed to have an N-linked glycosylated form that was not observed in the proliferative phase. This study provides a detailed view into the global proteomic alterations of the epithelial cells and stromal compartments of the cycling premenopausal endometrium. These proteomic alterations during endometrial remodeling provide a basis for numerous follow-up investigations on the function of these differentially regulated proteins and their role in reproductive biology and endometrial pathologies. © 2015 by the Society for the Study of Reproduction, Inc.

N-terminal acylation of somatostatin analog with long chain fatty acids enhances its stability and anti-proliferative activity in human breast adenocarcinoma cells.

PubMed

Dasgupta, Piyali; Singh, Anu; Mukherjee, Rama

2002-01-01

The anti-proliferative activity of the somatostatin analog RC-160 is limited by its short serum half life. To circumvent this limitation, fatty acids of chain lengths ranging from 4 to 18 were individually conjugated to the N-terminal residue of RC-160. The lipophilized derivatives of RC-160 were synthesized, purified and characterized. The anti-proliferative activity of lipophilized-RC-160 on the human breast carcinoma cell line MCF-7, was evaluated in vitro. The long chain lipopeptides like pamitoyl-RC-160 exhibited significantly higher anti-proliferative activity on MCF-7 cells (p<0.001), relative to RC-160. The affinity of RC-160 towards somatostatin receptors remained unaltered by pamitoylation. However, the observed increase in bioactivity was manifested within an optimum range of chain length of the lipoppetide. Increasing the peptide hydrophobicity beyond this range reduced the bioactivity of lipophilized-RC-160. Accordingly, stearoyl-RC-160, manifested lower anti-neoplastic activity and receptor affinity relative to pamitoyl-RC-160 and RC-160 itself. The signaling pathways underlying the antineoplastic activity of these lipopeptides were found to be similar to RC-160. Pamitoyl-RC-160 displayed enhanced inhibition of protein tyrosine kinase activity and intracellular cAMP levels in MCF-7 cells, relative to butanoyl-RC-160 or RC-160 itself. Pamitoyl-RC-160 also displayed greater resistance towards trypsin and serum degradation than RC-160. Lipophilization of RC-160 with long chain fatty acids like pamitic acid improves its stability and anti-proliferative activity, thereby improving the scope of enhancing its therapeutic index. However, the optimization of peptide hydrophobicity seems to be a crucial factor governing the efficacy of bioactive lipopeptides.

Krüppel-like factor 4 is induced by rapamycin and mediates the anti-proliferative effect of rapamycin in rat carotid arteries after balloon injury.

PubMed

Wang, Ying; Zhao, Beilei; Zhang, Yi; Tang, Zhihui; Shen, Qiang; Zhang, Youyi; Zhang, Weizhen; Du, Jie; Chien, Shu; Wang, Nanping

2012-04-01

The transcription factor, Krüppel-like factor 4 (KLF4), plays an important role in regulating the proliferation of vascular smooth muscle cells. This study aimed to examine the effect of rapamycin on the expression of KLF4 and the role of KLF4 in arterial neointimal formation. Expression of KLF4 was monitored using real-time PCR and immunoblotting in cultured vascular smooth muscle cells. and in rat carotid arteries in vivo after balloon injury. Adenovirus-mediated overexpression and siRNA-mediated knockdown of KLF4 were used to examine the role of KLF4 in mediating the anti-proliferative role of rapamycin . KLF4-regulated genes were identified using cDNA microarray. Rapamycin induced the expression of KLF4 in vitro and in vivo. Overexpression of KLF4 inhibited cell proliferation and the activity of mammalian target of rapamycin (mTOR) and its downstream pathways, including 4EBP-1 and p70S6K in vascular smooth muscle cells and prevented the neointimal formation in the balloon-injured arteries. KLF4 up-regulated the expression of GADD45β, p57(kip2) and p27(kip1) . Furthermore, knockdown of KLF4 attenuated the anti-proliferative effect of rapamycin both in vitro and in vivo. KLF4 plays an important role in mediating the anti-proliferative effect of rapamycin in VSMCs and balloon-injured arteries. Thus, it is a potential target for the treatment of proliferative vascular disorders such as restenosis after angioplasty. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Protein oxidation and degradation during proliferative senescence of human MRC-5 fibroblasts.

PubMed

Sitte, N; Merker, K; von Zglinicki, T; Grune, T

2000-03-01

One of the highlights of age-related changes of cellular metabolism is the accumulation of oxidized proteins. The aging process on a cellular level can be treated either as the ongoing proliferation until a certain number of cell divisions is reached (the Hayflick limit) or as the aging of nondividing cells, that is, the age-related changes in cells without proliferation. The present investigation was undertaken to reveal the changes in protein turnover, proteasome activity, and protein oxidation status during proliferative senescence. We were able to demonstrate that the activity of the cytosolic proteasomal system declines dramatically during the proliferative senescence of human MRC-5 fibroblasts. Regardless of the loss in activity, it could be demonstrated that there are no changes in the transcription and translation of proteasomal subunits. This decline in proteasome activity was accompanied by an increased concentration of oxidized proteins. Cells at higher proliferation stages were no longer able to respond with increased degradation of endogenous [(35)S]-Met-radiolabeled proteins after hydrogen peroxide- or quinone-induced oxidative stress. It could be demonstrated that oxidized proteins in senescent human MRC-5 fibroblasts are not as quickly removed as they are in young cells. Therefore, our study demonstrates that the accumulation of oxidized proteins and decline in protein turnover and activity of the proteasomal system are not only a process of postmitotic aging but also occur during proliferative senescence and result in an increased half-life of oxidized proteins.

Immunomodulation by memantine in therapy of Alzheimer's disease is mediated through inhibition of Kv1.3 channels and T cell responsiveness

PubMed Central

Lowinus, Theresa; Bose, Tanima; Busse, Stefan; Busse, Mandy; Reinhold, Dirk; Schraven, Burkhart; Bommhardt, Ursula H.H.

2016-01-01

Memantine is approved for the treatment of advanced Alzheimer's disease (AD) and reduces glutamate-mediated neuronal excitotoxicity by antagonism of N-methyl-D-aspartate receptors. In the pathophysiology of AD immune responses deviate and infectious side effects are observed during memantine therapy. However, the particular effects of memantine on human T lymphocytes are unresolved. Here, we provide evidence that memantine blocks Kv1.3 potassium channels, inhibits CD3-antibody- and alloantigen-induced proliferation and suppresses chemokine-induced migration of peripheral blood T cells of healthy donors. Concurrent with the in vitro data, CD4+ T cells from AD patients receiving therapeutic doses of memantine show a transient decline of Kv1.3 channel activity and a long-lasting reduced proliferative response to alloantigens in mixed lymphocyte reactions. Furthermore, memantine treatment provokes a profound depletion of peripheral blood memory CD45RO+ CD4+ T cells. Thus, standard doses of memantine profoundly reduce T cell responses in treated patients through blockade of Kv1.3 channels. This may normalize deviant immunopathology in AD and contribute to the beneficial effects of memantine, but may also account for the enhanced infection rate. PMID:27462773

Immunomodulation by memantine in therapy of Alzheimer's disease is mediated through inhibition of Kv1.3 channels and T cell responsiveness.

PubMed

Lowinus, Theresa; Bose, Tanima; Busse, Stefan; Busse, Mandy; Reinhold, Dirk; Schraven, Burkhart; Bommhardt, Ursula H H

2016-08-16

Memantine is approved for the treatment of advanced Alzheimer´s disease (AD) and reduces glutamate-mediated neuronal excitotoxicity by antagonism of N-methyl-D-aspartate receptors. In the pathophysiology of AD immune responses deviate and infectious side effects are observed during memantine therapy. However, the particular effects of memantine on human T lymphocytes are unresolved. Here, we provide evidence that memantine blocks Kv1.3 potassium channels, inhibits CD3-antibody- and alloantigen-induced proliferation and suppresses chemokine-induced migration of peripheral blood T cells of healthy donors. Concurrent with the in vitro data, CD4+ T cells from AD patients receiving therapeutic doses of memantine show a transient decline of Kv1.3 channel activity and a long-lasting reduced proliferative response to alloantigens in mixed lymphocyte reactions. Furthermore, memantine treatment provokes a profound depletion of peripheral blood memory CD45RO+ CD4+ T cells. Thus, standard doses of memantine profoundly reduce T cell responses in treated patients through blockade of Kv1.3 channels. This may normalize deviant immunopathology in AD and contribute to the beneficial effects of memantine, but may also account for the enhanced infection rate.

The Immunology of Wild Rodents: Current Status and Future Prospects

PubMed Central

Viney, Mark; Riley, Eleanor M.

2017-01-01

Wild animals’ immune responses contribute to their evolutionary fitness. These responses are moulded by selection to be appropriate to the actual antigenic environment in which the animals live, but without imposing an excessive energetic demand which compromises other component of fitness. But, exactly what these responses are, and how they compare with those of laboratory animals, has been little studied. Here, we review the very small number of published studies of immune responses of wild rodents, finding general agreement that their humoral (antibody) responses are highly elevated when compared with those of laboratory animals, and that wild rodents’ cellular immune system reveals extensive antigenic exposure. In contrast, proliferative and cytokine responses of ex vivo-stimulated immune cells of wild rodents are typically depressed compared with those of laboratory animals. Collectively, these responses are appropriate to wild animals’ lives, because the elevated responses reflect the cumulative exposure to infection, while the depressed proliferative and cytokine responses are indicative of effective immune homeostasis that minimizes immunopathology. A more comprehensive understanding of the immune ecology of wild animals requires (i) understanding the antigenic load to which wild animals are exposed, and identification of any key antigens that mould the immune repertoire, (ii) identifying immunoregulatory processes of wild animals and the events that induce them, and (iii) understanding the actual resource state of wild animals, and the immunological consequences that flow from this. Together, by extending studies of wild rodents, particularly addressing these questions (while drawing on our immunological understanding of laboratory animals), we will be better able to understand how rodents’ immune responses contribute to their fitness in the wild. PMID:29184549

The Immunology of Wild Rodents: Current Status and Future Prospects.

PubMed

Viney, Mark; Riley, Eleanor M

2017-01-01

Wild animals' immune responses contribute to their evolutionary fitness. These responses are moulded by selection to be appropriate to the actual antigenic environment in which the animals live, but without imposing an excessive energetic demand which compromises other component of fitness. But, exactly what these responses are, and how they compare with those of laboratory animals, has been little studied. Here, we review the very small number of published studies of immune responses of wild rodents, finding general agreement that their humoral (antibody) responses are highly elevated when compared with those of laboratory animals, and that wild rodents' cellular immune system reveals extensive antigenic exposure. In contrast, proliferative and cytokine responses of ex vivo -stimulated immune cells of wild rodents are typically depressed compared with those of laboratory animals. Collectively, these responses are appropriate to wild animals' lives, because the elevated responses reflect the cumulative exposure to infection, while the depressed proliferative and cytokine responses are indicative of effective immune homeostasis that minimizes immunopathology. A more comprehensive understanding of the immune ecology of wild animals requires (i) understanding the antigenic load to which wild animals are exposed, and identification of any key antigens that mould the immune repertoire, (ii) identifying immunoregulatory processes of wild animals and the events that induce them, and (iii) understanding the actual resource state of wild animals, and the immunological consequences that flow from this. Together, by extending studies of wild rodents, particularly addressing these questions (while drawing on our immunological understanding of laboratory animals), we will be better able to understand how rodents' immune responses contribute to their fitness in the wild.

Benzene-induced myelotoxicity: application of flow cytofluorometry for the evaluation of early proliferative change in bone marrow.

PubMed Central

Irons, R D

1981-01-01

A detailed description of flow cytofluorometric DNA cell cycle analysis is presented. A number of studies by the author and other investigators are reviewed in which a method is developed for the analysis of cell cycle phase in bone marrow of experimental animals. Bone marrow cell cycle analysis is a sensitive indicator of changes in bone marrow proliferative activity occurring early in chemically-induced myelotoxicity. Cell cycle analysis, used together with other hematologic methods, has revealed benzene-induced toxicity in proliferating bone marrow cells to be cycle specific, appearing to affect a population in late S phase which then accumulate in G2/M. PMID:7016521

Intracrine prostaglandin E2 pro-tumoral actions in prostate epithelial cells originate from non-canonical pathways.

PubMed

Madrigal-Martínez, Antonio; Fernández-Martínez, Ana B; Lucio Cazaña, Francisco J

2018-04-01

Prostaglandin E 2 (PGE 2 ) increases cell proliferation and stimulates migratory and angiogenic abilities in prostate cancer cells. However, the effects of PGE 2 on non-transformed prostate epithelial cells are unknown, despite the fact that PGE 2 overproduction has been found in benign hyperplastic prostates. In the present work we studied the effects of PGE 2 in immortalized, non-malignant prostate epithelial RWPE-1 cells and found that PGE 2 increased cell proliferation, cell migration, and production of vascular endothelial growth factor-A, and activated in vitro angiogenesis. These actions involved a non-canonic intracrine mechanism in which the actual effector was intracellular PGE 2 (iPGE 2 ) instead of extracellular PGE 2 : inhibition of the prostaglandin uptake transporter (PGT) or antagonism of EP receptors prevented the effects of PGE 2 , which indicated that PGE 2 activity depended on its carrier-mediated translocation from the outside to the inside of cells and that EP receptors located intracellularly (iEP) mediated the effects of PGE 2 . iPGE 2 acted through transactivation of epidermal growth factor-receptor (EGFR) by iEP, leading to increased expression and activity of hypoxia-inducible factor-1α (HIF-1α). Interestingly, iPGE 2 also mediates the effects of PGE 2 on prostate cancer PC3 cells through the axis iPGE 2 -iEP receptors-EGFR-HIF-1α. Thus, this axis might be responsible for the growth-stimulating effects of PGE 2 on prostate epithelial cells, thereby contributing to prostate proliferative diseases associated with chronic inflammation. Since this PGT-dependent non-canonic intracrine mechanism of PGE 2 action operates in both benign and malignant prostate epithelial cells, PGT inhibitors should be tested as a novel therapeutic modality to treat prostate proliferative disease. © 2017 Wiley Periodicals, Inc.

Oligomeric proanthocyanidin complexes (OPC) exert anti-proliferative and pro-apoptotic effects on prostate cancer cells.

PubMed

Neuwirt, Hannes; Arias, Mercedes Cedeno; Puhr, Martin; Hobisch, Alfred; Culig, Zoran

2008-11-01

Oligomeric proanthocyanidin complexes (OPC) are extracted from grape seeds or maritime pine bark and have been used for enhancement of capillary stability and lymphatic drainage. Since a role for OPC in cancer prevention was postulated, we asked whether they have an effect on prostate cancer cells. Cell proliferation was determined by (3)H-thymidine assay and cell cycle status was analyzed on a flow cytometer. Expression of regulators of proliferation and apoptosis was determined by Western blot. We found that androgen-responsive cells LNCaP are more sensitive to OPC in terms of inhibition of proliferation in comparison to androgen receptor-negative PC3 or DU145 cells. Treatment with OPC resulted in a decrease in the percentage of LNCaP cells in the S phase and an increase in the percentage of cells in sub G1 phase. The anti-proliferative and pro-apoptotic effect of OPC in the LNCaP cell line was associated with down-regulation of expression of the androgen receptor. Interestingly, similar regulatory effects of OPC, such as inhibition of expression of cyclin-dependent kinases and cyclins and stimulation of tumor suppressors p21 and p27, were seen in LNCaP and PC3 cells. Favorable changes in the Bcl-2/Bax ratio were observed in LNCaP and PC3 cells after the treatment with OPC. OPC caused an increase of phosphorylated mitogen-activated protein kinase p44 and p42, thus suggesting induction of cellular differentiation. We conclude that OPC is a candidate that fulfills criteria for chemopreventive strategies in prostate cancer that might be established in following in vivo studies. (c) 2008 Wiley-Liss, Inc.

Proliferative effects of 'fibre' on the intestinal epithelium: relationship to gastrin, enteroglucagon and PYY.

PubMed Central

Goodlad, R A; Lenton, W; Ghatei, M A; Adrian, T E; Bloom, S R; Wright, N A

1987-01-01

Refeeding starved rats with a fibre free 'elemental' diet increased crypt cell production rate (CCPR) in the proximal small intestine but not in the distal regions of the gut. Little effect on CCPR was seen when inert bulk (kaolin) was added to the 'elemental' diet. Addition of a poorly fermentable dietary 'fibre' (purified wood cellulose) had little effect on intestinal epithelial cell proliferation except in the distal colon where it significantly increased CCPR. A more readily fermentable 'fibre' (purified wheat bran) caused a large proliferative response in the proximal, mid and distal colon and in the distal small intestine. A gel forming 'fibre' also stimulated proliferation in the distal colon. There was no significant correlation between CCPR and plasma gastrin concentrations, but plasma enteroglucagon concentrations were significantly correlated with CCPR in almost all the sites studied. Plasma PYY concentrations also showed some correlation with CCPR, especially in the colon. Thus, whilst inert bulk cannot stimulate colonic epithelial cell proliferation, fermentable 'fibre' is capable of stimulating proliferation in the colon, and especially in the distal colon: it can also stimulate proliferation in the distal small intestine and it is likely that plasma enteroglucagon may have a role to play in this process. PMID:2826311

ERK1/2 and p38 MAP kinases control prion protein fragment 90-231-induced astrocyte proliferation and microglia activation.

PubMed

Thellung, Stefano; Villa, Valentina; Corsaro, Alessandro; Pellistri, Francesca; Venezia, Valentina; Russo, Claudio; Aceto, Antonio; Robello, Mauro; Florio, Tullio

2007-11-01

Astrogliosis and microglial activation are a common feature during prion diseases, causing the release of chemoattractant and proinflammatory factors as well as reactive free radicals, involved in neuronal degeneration. The recombinant protease-resistant domain of the prion protein (PrP90-231) displays in vitro neurotoxic properties when refolded in a beta-sheet-rich conformer. Here, we report that PrP90-231 induces the secretion of several cytokines, chemokines, and nitric oxide (NO) release, in both type I astrocytes and microglial cells. PrP90-231 elicited in both cell types the activation of ERK1/2 MAP kinase that displays, in astrocytes, a rapid kinetics and a proliferative response. Conversely, in microglia, PrP90-231-dependent MAP kinase activation was delayed and long lasting, inducing functional activation and growth arrest. In microglial cells, NO release, dependent on the expression of the inducible NO synthase (iNOS), and the secretion of the chemokine CCL5 were Ca(2+) dependent and under the control of the MAP kinases ERK1/2 and p38: ERK1/2 inhibition, using PD98059, reduced iNOS expression, while p38 blockade by PD169316 inhibited CCL5 release. In summary, we demonstrate that glial cells are activated by extracellular misfolded PrP90-231 resulting in a proliferative/secretive response of astrocytes and functional activation of microglia, both dependent on MAP kinase activation. In particular, in microglia, PrP90-231 activated a complex signalling cascade involved in the regulation of NO and chemokine release. These data argue in favor of a causal role for misfolded prion protein in sustaining glial activation and, possibly, glia-mediated neuronal death.

Phospholipase PLA2G7, associated with aggressive prostate cancer, promotes prostate cancer cell migration and invasion and is inhibited by statins

PubMed Central

Vainio, Paula; Lehtinen, Laura; Mirtti, Tuomas; Hilvo, Mika; Seppänen-Laakso, Tuulikki; Virtanen, Johannes; Sankila, Anna; Nordling, Stig; Lundin, Johan; Rannikko, Antti; Orešič, Matej; Kallioniemi, Olli; Iljin, Kristiina

2011-01-01

Prostate cancer is the second leading cause of cancer mortality in men in developed countries. Due to the heterogeneous nature of the disease, design of novel personalized treatments is required to achieve efficient therapeutic responses. We have recently identified phospholipase 2 group VII (PLA2G7) as a potential drug target especially in ERG oncogene positive prostate cancers. Here, the expression profile of PLA2G7 was studied in 1137 prostate cancer and 409 adjacent non-malignant prostate tissues using immunohistochemistry to validate its biomarker potential and putative association with disease progression. In order to reveal the molecular alterations induced by PLA2G7 impairment, lipidomic and gene expression profiling was performed in response to PLA2G7 silencing in cultured prostate cancer cells. Moreover, the antineoplastic effect of statins combined with PLA2G7 impairment was studied in prostate cancer cells to evaluate the potential of repositioning of in vivo compatible drugs developed for other indications towards anti-cancer purposes. The results indicated that PLA2G7 is a cancer-selective biomarker in 50% of prostate cancers and associates with aggressive disease. The alterations induced by PLA2G7 silencing highlighted the potential of PLA2G7 inhibition as an anti-proliferative, pro-apoptotic and anti-migratorial therapeutic approach in prostate cancer. Moreover, the anti-proliferative effect of PLA2G7 silencing was potentiated by lipid-lowering statins in prostate cancer cells. Taken together, our results support the potential of PLA2G7 as a biomarker and a drug target in prostate cancer and present a rationale for combining PLA2G7 inhibition with the use of statins in prostate cancer management. PMID:22202492

Distinct cellular pathways select germline-encoded and somatically mutated antibodies into immunological memory

PubMed Central

Kaji, Tomohiro; Ishige, Akiko; Hikida, Masaki; Taka, Junko; Hijikata, Atsushi; Kubo, Masato; Nagashima, Takeshi; Takahashi, Yoshimasa; Kurosaki, Tomohiro; Okada, Mariko; Ohara, Osamu

2012-01-01

One component of memory in the antibody system is long-lived memory B cells selected for the expression of somatically mutated, high-affinity antibodies in the T cell–dependent germinal center (GC) reaction. A puzzling observation has been that the memory B cell compartment also contains cells expressing unmutated, low-affinity antibodies. Using conditional Bcl6 ablation, we demonstrate that these cells are generated through proliferative expansion early after immunization in a T cell–dependent but GC-independent manner. They soon become resting and long-lived and display a novel distinct gene expression signature which distinguishes memory B cells from other classes of B cells. GC-independent memory B cells are later joined by somatically mutated GC descendants at roughly equal proportions and these two types of memory cells efficiently generate adoptive secondary antibody responses. Deletion of T follicular helper (Tfh) cells significantly reduces the generation of mutated, but not unmutated, memory cells early on in the response. Thus, B cell memory is generated along two fundamentally distinct cellular differentiation pathways. One pathway is dedicated to the generation of high-affinity somatic antibody mutants, whereas the other preserves germ line antibody specificities and may prepare the organism for rapid responses to antigenic variants of the invading pathogen. PMID:23027924

Ex vivo immunosuppressive effects of mesenchymal stem cells on Crohn's disease mucosal T cells are largely dependent on indoleamine 2,3-dioxygenase activity and cell-cell contact.

PubMed

Ciccocioppo, Rachele; Cangemi, Giuseppina C; Kruzliak, Peter; Gallia, Alessandra; Betti, Elena; Badulli, Carla; Martinetti, Miryam; Cervio, Marila; Pecci, Alessandro; Bozzi, Valeria; Dionigi, Paolo; Visai, Livia; Gurrado, Antonella; Alvisi, Costanza; Picone, Cristina; Monti, Manuela; Bernardo, Maria E; Gobbi, Paolo; Corazza, Gino R

2015-07-24

Crohn's disease (CD) is a disabling chronic enteropathy sustained by a harmful T-cell response toward antigens of the gut microbiota in genetically susceptible subjects. Growing evidence highlights the safety and possible efficacy of mesenchymal stem cells (MSCs) as a new therapeutic tool for this condition. Therefore, we aimed to investigate the effects of bone marrow-derived MSCs on pathogenic T cells with a view to clinical application. T-cell lines from both inflamed and non-inflamed colonic mucosal specimens of CD patients and from healthy mucosa of control subjects were grown with the antigen muramyl-dipeptide in the absence or presence of donors' MSCs. The MSC effects were evaluated in terms of T-cell viability, apoptotic rate, proliferative response, immunophenotype, and cytokine profile. The role of the indoleamine 2,3-dioxygenase (IDO) was established by adding a specific inhibitor, the 1-methyl-DL-tryptophan, and by using MSCs transfected with the small interfering RNA (siRNA) targeting IDO. The relevance of cell-cell contact was evaluated by applying transwell membranes. A significant reduction in both cell viability and proliferative response to muramyl-dipeptide, with simultaneous increase in the apoptotic rate, was found in T cells from both inflamed and non-inflamed CD mucosa when co-cultured with MSCs and was reverted by inhibiting IDO activity and expression. A reduction of the activated CD4(+)CD25(+) subset and increase of the CD3(+)CD69(+) population were also observed when T-cell lines from CD mucosa were co-cultured with MSCs. In parallel, an inhibitory effect was evident on the expression of the pro-inflammatory cytokines tumor necrosis factor-α, interferon-γ, interleukin-17A and -21, whereas that of the transforming growth factor-β and interleukin-6 were increased, and production of the tolerogenic molecule soluble HLA-G was high. These latter effects were almost completely eliminated by blocking the IDO, whose activity was upregulated in MSCs co-cultured with CD T cells. The use of a semipermeable membrane partially inhibited the MSC immunosuppressive effects. Finally, hardly any effects of MSCs were observed when T cells obtained from control subjects were used. MSCs exert potent immunomodulant effects on antigen-specific T cells in CD through a complex paracrine and cell-cell contact-mediated action, which may be exploited for widespread therapeutic use.

Glyphosate induces human breast cancer cells growth via estrogen receptors.

PubMed

Thongprakaisang, Siriporn; Thiantanawat, Apinya; Rangkadilok, Nuchanart; Suriyo, Tawit; Satayavivad, Jutamaad

2013-09-01

Glyphosate is an active ingredient of the most widely used herbicide and it is believed to be less toxic than other pesticides. However, several recent studies showed its potential adverse health effects to humans as it may be an endocrine disruptor. This study focuses on the effects of pure glyphosate on estrogen receptors (ERs) mediated transcriptional activity and their expressions. Glyphosate exerted proliferative effects only in human hormone-dependent breast cancer, T47D cells, but not in hormone-independent breast cancer, MDA-MB231 cells, at 10⁻¹² to 10⁻⁶M in estrogen withdrawal condition. The proliferative concentrations of glyphosate that induced the activation of estrogen response element (ERE) transcription activity were 5-13 fold of control in T47D-KBluc cells and this activation was inhibited by an estrogen antagonist, ICI 182780, indicating that the estrogenic activity of glyphosate was mediated via ERs. Furthermore, glyphosate also altered both ERα and β expression. These results indicated that low and environmentally relevant concentrations of glyphosate possessed estrogenic activity. Glyphosate-based herbicides are widely used for soybean cultivation, and our results also found that there was an additive estrogenic effect between glyphosate and genistein, a phytoestrogen in soybeans. However, these additive effects of glyphosate contamination in soybeans need further animal study. Copyright © 2013 Elsevier Ltd. All rights reserved.

The in vitro mitogenic response to intact bacteria by murine B cells does not predict in vivo susceptibility to Salmonella typhimurium.

PubMed

Elkins, K; Metcalf, E S

1986-05-01

We are interested in developing in vitro culture systems that will permit immune responses to intact Salmonella typhimurium, since these systems would have certain advantages over in vivo infection models for the characterization of the host's responding cell types. In this report, the in vitro proliferative response of nonimmune murine spleen cells to four different killed preparations of S. typhimurium, strain TML (TML), are examined. These studies show that UV-killed TML, heat-killed TML, glutaraldehyde-killed TML, and acetone-killed and dried TML, all elicit a nonspecific mitogenic spleen cell response in vitro, as does a live, avirulent, temperature-sensitive mutant of TML, TS27. This response reaches a maximum on day 2 after initiation of culture, which is similar to the time course of a conventional lipopolysaccharide (LPS) response. Unlike the LPS response, little 3H-thymidine incorporation is observed in low-density cultures (2 X 10(5) cells/well), which suggests a critical role for accessory cells. The responding cell types include, but are not necessarily limited to, the B-cell population. The response cannot be readily inhibited by polymyxin B, which binds specifically to the lipid A portion of LPS. Thus, the bacterial components required for mitogenicity are not yet definitively identified. A survey of the mitogenic responses of lymphocytes from various inbred mouse strains, including the C3H/HeJ LPS hyporesponsive strain, indicates that all B cells tested are capable of proliferating vigorously in response to intact TML, regardless of the in vivo susceptibility to virulent infection. These results also emphasize the importance of assessing the nonspecific components of the immune response when studying the specific immune response to intact S. typhimurium.

Blockade of the Programmed Death-1 Pathway Restores Sarcoidosis CD4+ T-Cell Proliferative Capacity

PubMed Central

Braun, Nicole A.; Celada, Lindsay J.; Herazo-Maya, Jose D.; Abraham, Susamma; Shaginurova, Guzel; Sevin, Carla M.; Grutters, Jan; Culver, Daniel A.; Dworski, Ryszard; Sheller, James; Massion, Pierre P.; Polosukhin, Vasiliy V.; Johnson, Joyce E.; Kaminski, Naftali; Wilkes, David S.; Oswald-Richter, Kyra A.

2014-01-01

Rationale: Effective therapeutic interventions for chronic, idiopathic lung diseases remain elusive. Normalized T-cell function is an important contributor to spontaneous resolution of pulmonary sarcoidosis. Up-regulation of inhibitor receptors, such as programmed death-1 (PD-1) and its ligand, PD-L1, are important inhibitors of T-cell function. Objectives: To determine the effects of PD-1 pathway blockade on sarcoidosis CD4+ T-cell proliferative capacity. Methods: Gene expression profiles of sarcoidosis and healthy control peripheral blood mononuclear cells were analyzed at baseline and follow-up. Flow cytometry was used to measure ex vivo expression of PD-1 and PD-L1 on systemic and bronchoalveolar lavage–derived cells of subjects with sarcoidosis and control subjects, as well as the effects of PD-1 pathway blockade on cellular proliferation after T-cell receptor stimulation. Immunohistochemistry analysis for PD-1/PD-L1 expression was conducted on sarcoidosis, malignant, and healthy control lung specimens. Measurements and Main Results: Microarray analysis demonstrates longitudinal increase in PDCD1 gene expression in sarcoidosis peripheral blood mononuclear cells. Immunohistochemistry analysis revealed increased PD-L1 expression within sarcoidosis granulomas and lung malignancy, but this was absent in healthy lungs. Increased numbers of sarcoidosis PD-1+ CD4+ T cells are present systemically, compared with healthy control subjects (P < 0.0001). Lymphocytes with reduced proliferative capacity exhibited increased proliferation with PD-1 pathway blockade. Longitudinal analysis of subjects with sarcoidosis revealed reduced PD-1+ CD4+ T cells with spontaneous clinical resolution but not with disease progression. Conclusions: Analogous to the effects in other chronic lung diseases, these findings demonstrate that the PD-1 pathway is an important contributor to sarcoidosis CD4+ T-cell proliferative capacity and clinical outcome. Blockade of the PD-1 pathway may be a viable therapeutic target to optimize clinical outcomes. PMID:25073001

CCR4+T cell recruitment to the skin in mycosis fungoides: potential contributions by thymic stromal lymphopoietin and interleukin-16.

PubMed

Tuzova, Marina; Richmond, Jillian; Wolpowitz, Deon; Curiel-Lewandrowski, Clara; Chaney, Keri; Kupper, Thomas; Cruikshank, William

2015-02-01

Mycosis fungoides (MF) is characterized by skin accumulation of CCR4+CCR7- effector memory T cells; however the mechanism for their recruitment is not clearly identified. Thymic Stromal Lymphopoietin (TSLP) is a keratinocyte-derived cytokine that triggers Th2 immunity and is associated with T cell recruitment to the skin in atopic dermatitis. Interleukin-16 (IL-16) is a chemoattractant and growth factor for CD4+T cells. We hypothesized that TSLP and IL-16 could contribute to recruitment of malignant T cells in MF. We found elevated TSLP and IL-16 in very early stage patients' plasma and skin biopsies, prior to elevation in CCL22. Both TSLP and IL-16 induced migratory responses of CCR4+TSLPR+CD4+CCR7-CD31+cells, characteristic of malignant T cells in the skin. Co-stimulation also resulted in significant proliferative responses. We conclude that TSLP and IL-16, expressed at early stages of disease, function to recruit malignant T cells to the skin and contribute to their enhanced proliferation.

Carcinogenic effects of cadmium in the noble (NBL/Cr) rat: induction of pituitary, testicular, and injection site tumors and intraepithelial proliferative lesions of the dorsolateral prostate.

PubMed

Waalkes, M P; Anver, M; Diwan, B A

1999-12-01

Cadmium is a known human carcinogen based on findings of lung cancer in exposed populations. A more controversial target site for cadmium is the human prostate gland, for which some studies indicate a link between cadmium exposure and cancer. Our work in various strains of Wistar rats has shown that cadmium can induce tumors in the ventral lobe of the prostate. The relevance of this type of lesion to human prostate cancer has been questioned because the ventral lobe of the rat prostate, unlike the dorsolateral lobe, has no embryological homolog in the human gland. In this study we investigated the chronic toxic and carcinogenic effects of cadmium in the Noble (NBL/Cr) rat, with particular attention to lesions of the prostate. Cadmium chloride (CdCl2) was given as a single sc injection (0, 1, 2, 4, 8, 16, or 32 micromol/kg) to groups (initially n = 30) of 10-week-old rats. Rats were observed for up to 72 weeks following exposure. In rats that were injected with the lower doses of cadmium (< or =4 micromol/kg), a clear dose-related increase in proliferative lesions of the dorsolateral prostate occurred (0 micromol/kg = 36% incidence, 1 micromol/kg = 62%, 2 micromol/kg = 65%; 4 micromol/kg = 79%; trend p < 0.003). Lesions were described as intraepithelial hyperplasia with occasional areas of atypical epithelial cells, without stromal invasion. At higher doses (> or =8 micromol/kg) the proliferative-lesion response in the dorsolateral prostate gradually declined to near control levels (8 micromol/kg = 63%; 16 micromol/kg = 60%; 32 micromol/kg = 52%). The loss of prostatic response at the higher doses of cadmium was probably due to loss of testicular function secondary to cadmium treatment. This was reflected in a very high incidence (>90%) of lesions, indicative of testicular hypofunction, including tubular degeneration, mineralization, and interstitial (Leydig) cell tumors, at doses in excess of 16 micromol/kg. Malignant injection-site sarcomas occurred at the two highest doses of cadmium, while pituitary adenomas were elevated by cadmium exposure at the highest dose. These results show that cadmium induces proliferative lesions in the dorsolateral prostate of the Noble rat, a model having a presumed relevance to human prostate cancers.

Maternal allergic disease does not affect the phenotype of T and B cells or the immune response to allergens in neonates.

PubMed

Rindsjö, E; Joerink, M; Johansson, C; Bremme, K; Malmström, V; Scheynius, A

2010-07-01

It is hypothesized that the in utero environment in allergic mothers can affect the neonatal immune responses. The aim of this study was to analyse the effect of maternal allergic disease on cord blood mononuclear cell (CBMC) phenotype and proliferative responses upon allergen stimulation. Peripheral blood mononuclear cells (PBMC) from 12 allergic and 14 nonallergic mothers and CBMC from their children were analysed. In the mothers, we determined cell proliferation, production of IL-4 and expression of FOXP3 in response to allergen stimulation. In the children, we evaluated cell proliferation and FOXP3 expression following allergen stimulation. Furthermore, expression of different homing markers on T cells and regulatory T cells and maturity of the T cells and B cell subsets were evaluated directly ex vivo. The timothy- and birch-allergic mothers responded with increased proliferation and/or IL-4 production towards timothy and birch extract, respectively, when compared to nonallergic mothers. This could not be explained by impairment of FOXP3(+) regulatory T cells in the allergic mothers. CBMC proliferation and FOXP3 expression in response to allergens were not affected by the allergic status of the mother. Also, phenotype of T cells, FOXP3(+) regulatory T cells and B cells was not affected by the allergic status of the mother. Our results suggest that maternal allergic disease has no effect on the neonatal response to allergens or the phenotype of neonatal lymphocytes. The factors studied here could, however, still affect later development of allergy.

Biogenic amines as regulators of the proliferative activity of normal and neoplastic intestinal epithelial cells (review).

PubMed

Tutton, P J; Barkla, D H

1987-01-01

The role of extracellular amines such as noradrenaline and serotonin and their interaction with cyclic nucleotides and intracellular polyamines in the regulation of intestinal epithelial cell proliferation is reviewed with particular reference to the differences between normal and neoplastic cells. In respect to the normal epithelium of the small intestine there is a strong case to support the notion that cell proliferation is controlled by, amongst other things, sympathetic nerves. In colonic carcinomas, antagonists for certain serotonin receptors, for histamine H2 receptors and for dopamine D2 receptors inhibit both cell division and tumour growth. Because of the reproducible variations between tumour lines in the response to these antagonists, this inhibition appears to be due to a direct effect on the tumour cells rather than an indirect effect via the tumour host or stroma. This conclusion is supported by the cytocidal effects of toxic congeners of serotonin on the tumour cells. The most salient difference between the amine responses of normal and neoplastic cells relates to the issue of amine uptake. Proliferation of crypt cells is promoted by amine uptake inhibitors, presumably because they block amine re-uptake by the amine secreting cells--sympathetic neurones and enteroendocrine cells. However, tumour cell proliferation is strongly inhibited by amine uptake inhibitors, suggesting that neoplastic cells can, and need to take up the amine before being stimulated by it. Recent revelations in the field of oncogenes also support an important association between amines, cyclic nucleotides and cell division. The ras oncogenes code for a protein that is a member of a family of molecules which relay information from extracellular regulators, such as biogenic amines, to the intracellular regulators, including cyclic nucleotides. Evidence is presented suggesting that enteroendocrine cells, enterocytes, carcinoid tumour cells and adenocarcinoma cells all have the same embryonic origin and that cells exhibiting an admixture of endocrine and proliferative properties exist in colonic tumours, but not in the normal intestinal epithelium. Thus, it appears that in the normal intestine a clear structural and functional distinction exists between the regulating cells (i.e. the sympathetic neurones and enteroendocrine cells) and the regulated cells (i.e. the undifferentiated crypt cells): cells that have acquired a regulating role are no longer able to divide and cells which are able to divide do not take up or store amines.(ABSTRACT TRUNCATED AT 400 WORDS)

Anatomical gradients in proliferation and differentiation of embryonic rat CNS accessed by buoyant density fractionation: alpha 3, beta 3 and gamma 2 GABAA receptor subunit co-expression by post-mitotic neocortical neurons correlates directly with cell buoyancy.

PubMed

Maric, D; Maric, I; Ma, W; Lahojuji, F; Somogyi, R; Wen, X; Sieghart, W; Fritschy, J M; Barker, J L

1997-03-01

Development of the CNS occurs as a complex cascade of pre-programmed events involving distinct phases of cell proliferation and differentiation. Here we show these phases correlate with cells of specific buoyant densities which can be readily accessed by density gradient fractionation. Sprague-Dawley dams were pulse-labelled with bromodeoxyuridine (BrdU) and selected regions of embryonic (E) CNS tissues at E11-22 dissociated with papain into single-cell suspensions. Proliferative cell populations were assessed by anti-BrdU and propidium iodide staining using flow cytometry. Cell differentiation was evaluated using molecular and immunocytochemical probes against mRNAs and antigens differentiating the neuroepithelial, neuronal and glial cell lineages. The results show the emergence of distinctive spatiotemporal changes in BrdU+ populations throughout the CNS during embryonic development, which were followed by corresponding changes in the cellular distributions of antigens distinguishing specific cell types. Fractionation of neocortical cells using discontinuous Percoll gradients revealed that an increasing number of cells increase their buoyancy during corticogenesis. Immunocytochemical and molecular characterization showed that the proliferative and progenitor cell populations are for the most part associated with lower buoyancy or higher specific buoyant densities (> 1.056 g/ml) whereas the post-mitotic, differentiated neurons generally separated into fractions of higher buoyancy or lower specific buoyant densities (< 1.043 g/ml). Immunostaining with antibodies against several GABAA receptor subunits (alpha 3, beta 3, gamma 2) revealed that the highest percent (70-90%) of immunopositive cells could be identified in the most buoyant, differentiating neurons found in the cortical plate/subplate regions, with the lowest percent of the immunopositive cells found in the least buoyant, proliferative and progenitor cell populations originating from the ventricular/subventricular zones. Taken together, these results indicate that buoyant density is a distinguishing characteristic of embryonic CNS cells transforming from primarily proliferative to mainly differentiating, and that fractionation of these cells according to their buoyant densities provides rapid access to the properties of specific cell lineages during the prenatal period of CNS development.

Influence of parturition and diets enriched in n-3 or n-6 polyunsaturated fatty acids on immune response of dairy cows during the transition period.

PubMed

Lessard, M; Gagnon, N; Godson, D L; Petit, H V

2004-07-01

The objectives of this study were to evaluate the functional properties of immunocompetent cells in dairy cows fed diets enriched in n-3 or n-6 polyunsaturated fatty acids during the transition period. Six weeks before calving, 21 primiparous and 27 multiparous pregnant Holstein dairy cows were randomly allotted to 1 of 3 dietary fat treatments: calcium salts of palm oil (Megalac), micronized soybeans, or whole flaxseed, which are, respectively, rich in saturated, n-6, or n-3 fatty acids. On wk 6 and 3 before parturition, cows received a subcutaneous injection of ovalbumin to measure the antibody response in colostrum and serum. Colostrum samples were collected at the first milking after calving, and blood samples were taken 6, 3, and 1 wk before the expected calving date and 1, 3, and 6 wk after calving. Blood mononuclear cells were cultured to evaluate the proliferative response to concanavalin A and the in vitro productions of interferon-gamma, tumor necrosis factor-alpha, nitric oxide, and prostaglandin E2. The serum antibody response to ovalbumin was unaffected by dietary fatty acids, but the response was lower in primiparous cows than in multiparous cows. A significant diet x parity interaction indicated that colostral antibody level against ovalbumin was significantly higher in multiparous cows fed soybeans than in those fed flaxseed or Megalac; there was no difference among treatments for primiparous cows. The lymphocyte response to concanavalin A was lower in cows fed soybeans than in those receiving flaxseed or Megalac when the cells were incubated with autologous serum. The proliferative response of mononuclear cells incubated with autologous serum was suppressed in the 1st wk after calving in both primiparous and multiparous cows, and multiparous cows showed a higher response than primiparous cows throughout the experiment. There was a significant interaction between parity and diet as a result of a greater production of interferon-gamma by mononuclear cells incubated with autologous serum in multiparous cows than in primiparous cows fed flaxseed; there was no difference among cows fed the other diets. Interferon-gamma production was reduced around calving while the inverse was observed for productions of nitric oxide and tumor necrosis factor-alpha. Productions of nitric oxide, prostaglandin E2, and tumor necrosis factor-gamma were greater in primiparous cows than in multiparous cows. In conclusion, functional properties of lymphocytes and monocyte/macrophage lineage of dairy cows during the transition period are modulated by parturition and the composition of polyunsaturated fatty acids in the diet.

Quantitative and qualitative iNKT repertoire associations with disease susceptibility and outcome in macaque tuberculosis infection.

PubMed

Chancellor, Andrew; White, Andrew; Tocheva, Anna S; Fenn, Joe R; Dennis, Mike; Tezera, Liku; Singhania, Akul; Elliott, Tim; Tebruegge, Marc; Elkington, Paul; Gadola, Stephan; Sharpe, Sally; Mansour, Salah

2017-07-01

Correlates of immune protection that reliably predict vaccine efficacy against Mycobacterium tuberculosis (Mtb) infection are urgently needed. Invariant NKT cells (iNKTs) are CD1d-dependent innate T cells that augment host antimicrobial immunity through production of cytokines, including interferon (IFN)-γ and tumour necrosis factor (TNF)-α. We determined peripheral blood iNKT numbers, their proliferative responses and iNKT subset proportions after in vitro antigen expansion by α-galactosylceramide (αGC) in a large cohort of mycobacteria-naïve non-human primates, and macaques from Bacillus Calmette-Guerin (BCG) vaccine and Mtb challenge studies. Animals studied included four genetically distinct groups of macaques within cynomolgus and rhesus species that differ in their susceptibility to Mtb infection. We demonstrate significant differences in ex vivo iNKT frequency between groups, which trends towards an association with susceptibility to Mtb, but no significant difference in overall iNKT proliferative responses. Susceptible animals exhibited a skewed CD4 + /CD8 + iNKT subset ratio in comparison to more Mtb-resistant groups. Correlation of iNKT subsets post BCG vaccination with clinical disease manifestations following Mtb challenge in the Chinese cynomolgus and Indian rhesus macaques identified a consistent trend linking increased CD8 + iNKTs with favourable disease outcome. Finally, a similar iNKT profile was conferred by BCG vaccination in rhesus macaques. Our study provides the first detailed characterisation of iNKT cells in macaque tuberculosis infection, suggesting that iNKT repertoire differences may impact on disease outcome, which warrants further investigation. Copyright © 2017 Elsevier Ltd. All rights reserved.

GPER and ERα expression in abnormal endometrial proliferations.

PubMed

Tica, Andrei Adrian; Tica, Oana Sorina; Georgescu, Claudia Valentina; Pirici, Daniel; Bogdan, Maria; Ciurea, Tudorel; Mogoantă, Stelian ŞtefăniŢă; Georgescu, Corneliu Cristian; Comănescu, Alexandru Cristian; Bălşeanu, Tudor Adrian; Ciurea, Raluca Niculina; Osiac, Eugen; Buga, Ana Maria; Ciurea, Marius Eugen

2016-01-01

G-protein coupled estrogen receptor 1 (GPER), a particular extranuclear estrogen receptor (ER), seems not to be significantly involved in normal female phenotype development but especially associated with severe genital malignancies. This study investigated the GPER expression in different types of normal and abnormal proliferative endometrium, and the correlation with the presence of ERα. GPER was much highly expressed in cytoplasm (than onto cell membrane), contrary to ERα, which was almost exclusively located in the nucleus. Both ERs' densities were higher in columnar epithelial then in stromal cells, according with higher estrogen-sensitivity of epithelial cells. GPER and ERα density decreased as follows: complex endometrial hyperplasia (CEH) > simple endometrial hyperplasia (SHE) > normal proliferative endometrium (NPE) > atypical endometrial hyperplasia (AEH), ERα' density being constantly higher. In endometrial adenocarcinomas, both ERs were significant lower expressed, and widely varied, but GPER÷ERα ratio was significantly increased in high-grade lesions. The nuclear ERα is responsible for the genomic (the most important) mechanism of action of estrogens, involved in cell growth and multiplication. In normal and benign proliferations, ERα expression is increased as an evidence of its effects on cells with conserved architecture, in atypical and especially in malignant cells ERα's (and GPER's) density being much lower. Cytoplasmic GPER probably interfere with different tyrosine÷protein kinases signaling pathways, also involved in cell growth and proliferation. In benign endometrial lesions, GPER's presence is, at least partially, the result of an inductor effect of ERα on GPER gene transcription. In high-grade lesions, GPER÷ERα ratio was increased, demonstrating that GPER is involved per se in malignant endometrial proliferations.

Suppressor cell hyperactivity relative to allogeneic lymphocyte proliferation as a manifestation of defective T-T-cell interactions in systemic lupus erythematosus

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

Stenina, M.A.; Potapova, A.A.; Biryukov, A.V.

1987-01-01

The authors study the state of immunoregulatory process in patients with systemic lupus erythematosus at the T-T-cell interaction level and seek to test the possibility of the pharmacological modulation of this process. The proliferative activity of mononuclear lymphocytes, extracted from the blood of ten lupus patients, was assessed by measuring the incorporation of tritiated thymidine into cultures stimulated by phytohemagglutinin, concanavalin, and theophylline. The comparative effects of each of these agents on the immunoregulatory and proliferative activity of the lymphocytes are reported.

Cell cycle gene expression networks discovered using systems biology: Significance in carcinogenesis

PubMed Central

Scott, RE; Ghule, PN; Stein, JL; Stein, GS

2015-01-01

The early stages of carcinogenesis are linked to defects in the cell cycle. A series of cell cycle checkpoints are involved in this process. The G1/S checkpoint that serves to integrate the control of cell proliferation and differentiation is linked to carcinogenesis and the mitotic spindle checkpoint with the development of chromosomal instability. This paper presents the outcome of systems biology studies designed to evaluate if networks of covariate cell cycle gene transcripts exist in proliferative mammalian tissues including mice, rats and humans. The GeneNetwork website that contains numerous gene expression datasets from different species, sexes and tissues represents the foundational resource for these studies (www.genenetwork.org). In addition, WebGestalt, a gene ontology tool, facilitated the identification of expression networks of genes that co-vary with key cell cycle targets, especially Cdc20 and Plk1 (www.bioinfo.vanderbilt.edu/webgestalt). Cell cycle expression networks of such covariate mRNAs exist in multiple proliferative tissues including liver, lung, pituitary, adipose and lymphoid tissues among others but not in brain or retina that have low proliferative potential. Sixty-three covariate cell cycle gene transcripts (mRNAs) compose the average cell cycle network with p = e−13 to e−36. Cell cycle expression networks show species, sex and tissue variability and they are enriched in mRNA transcripts associated with mitosis many of which are associated with chromosomal instability. PMID:25808367

Testicular Development in Male Rats Is Sensitive to a Soy-Based Diet in the Neonatal Period1

PubMed Central

Napier, India D.; Simon, Liz; Perry, Devin; Cooke, Paul S.; Stocco, Douglas M.; Sepehr, Estatira; Doerge, Daniel R.; Kemppainen, Barbara W.; Morrison, Edward E.; Akingbemi, Benson T.

2014-01-01

ABSTRACT Approximately 30% of infants in the United States are exposed to high doses of isoflavones resulting from soy infant formula consumption. Soybeans contain the isoflavones genistin and daidzin, which are hydrolyzed in the gastrointestinal tract to their genistein and daidzein aglycones. Both aglycones possess hormonal activity and may interfere with male reproductive development. Testosterone, which supports male fertility, is mainly produced by testicular Leydig cells. Our previous studies indicated that perinatal exposure of male rats to isoflavones induced proliferative activity in Leydig cells and increased testosterone concentrations into adulthood. However, the relevance of the neonatal period as part of the perinatal window of isoflavone exposure remains to be established. The present study examined the effects of exposure to isoflavones on male offspring of dams maintained on a casein-based control or whole soybean diet in the neonatal period, that is, Days 2 to 21 postpartum. The results showed that the soybean diet stimulated proliferative activity in developing Leydig cells while suppressing their steroidogenic capacity in adulthood. In addition, isoflavone exposure decreased production of anti-Müllerian hormone by Sertoli cells. Similar to our previous in vitro studies of genistein action in Leydig cells, daidzein induced proliferation and interfered with signaling pathways to suppress steroidogenic activity. Overall, the data showed that the neonatal period is a sensitive window of exposure to isoflavones and support the view that both genistein and daidzein are responsible for biological effects associated with soy-based diets. PMID:24451983

Biologic consequences of Stat1-independent IFN signaling

PubMed Central

Gil, M. Pilar; Bohn, Erwin; O'Guin, Andrew K.; Ramana, Chilakamarti V.; Levine, Beth; Stark, George R.; Virgin, Herbert W.; Schreiber, Robert D.

2001-01-01

Although Stat1 is required for many IFN-dependent responses, recent work has shown that IFNγ functions independently of Stat1 to affect the growth of tumor cells or immortalized fibroblasts. We now demonstrate that both IFNγ and IFNα/β regulate proliferative responses in cells of the mononuclear phagocyte lineage derived from Stat1-null mice. Using both representational difference analysis and gene arrays, we show that IFNγ exerts its Stat1-independent actions on mononuclear phagocytes by regulating the expression of many genes. This result was confirmed by monitoring changes in expression and function of the corresponding gene products. Regulation of the expression of these genes requires the IFNγ receptor and Jak1. The physiologic relevance of IFN-dependent, Stat1-independent signaling was demonstrated by monitoring antiviral responses in Stat1-null mice. Thus, the IFN receptors engage alternative Stat1-independent signaling pathways that have important physiological consequences. PMID:11390995

Reciprocal Activation of Transcription Factors Underlies the Dichotomy between Proliferation and Invasion of Glioma Cells

PubMed Central

Dhruv, Harshil D.; McDonough Winslow, Wendy S.; Armstrong, Brock; Tuncali, Serdar; Eschbacher, Jenny; Kislin, Kerri; Loftus, Joseph C.; Tran, Nhan L.; Berens, Michael E.

2013-01-01

Histology of malignant glioma depicts dense proliferative areas rich in angiogenesis as well as dissemination of neoplastic cells into adjacent brain tissue. Although the mechanisms that trigger transition from proliferative to invasive phenotypes are complex, the dichotomy of cell proliferation and migration, the “Go or Grow” hypothesis, argues for specific and coordinated regulation of these phenotypes. We investigated transcriptional elements that accompany the phenotypes of migration and proliferation, and consider the therapeutic significance of the “Go or Grow” hypothesis. Interrogation of matched core and rim regions from human glioblastoma biopsy specimens in situ (n = 44) revealed higher proliferation (Ki67 labeling index) in cells residing at the core compared to the rim. Profiling activated transcription factors in a panel of migration-activated versus migration-restricted GBM cells portrayed strong NF-κB activity in the migratory cell population. In contrast, increased c-Myc activity was found in migration-restricted proliferative cells. Validation of transcriptional activity by NF-κB- or c-Myc-driven GFP or RFP, respectively, showed an increased NF-κB activity in the active migrating cells, whereas the proliferative, migration restricted cells displayed increased c-Myc activity. Immunohistochemistry on clinical specimens validated a robust phosphorylated c-Myc staining in tumor cells at the core, whereas increased phosphorylated NF-κB staining was detected in the invasive tumor cells at the rim. Functional genomics revealed that depletion of c-Myc expression by siRNA oligonucleotides reduced cell proliferation in vitro, but surprisingly, cell migration was enhanced significantly. Conversely, inhibition of NF-κB by pharmacological inhibitors, SN50 or BAY-11, decreased both cell migration in vitro and invasion ex vivo. Notably, inhibition of NF-κB was found to have no effect on the proliferation rate of glioma cells. These findings suggest that the reciprocal and coordinated suppression/activation of transcription factors, such as c-Myc and NF-κB may underlie the shift of glioma cells from a “growing-to-going” phenotype. PMID:23967279

Cloning and characterization of a DNA polymerase beta gene from Trypanosoma cruzi.

PubMed

Venegas, Juan A; Aslund, Lena; Solari, Aldo

2009-06-01

A gene coding for a DNA polymerase beta from the Trypanosoma cruzi Miranda clone, belonging to the TcI lineage, was cloned (Miranda Tcpol beta), using the information from eight peptides of the T. cruzi beta-like DNA polymerase purified previously. The gene encodes for a protein of 403 amino acids which is very similar to the two T. cruzi CL Brener (TcIIe lineage) sequences published, but has three different residues in highly conserved segments. At the amino acid level, the identity of TcI-pol beta with mitochondrial pol beta and pol beta-PAK from other trypanosomatids was between 68-80% and 22-30%, respectively. Miranda Tc-pol beta protein has an N-terminal sequence similar to that described in the mitochondrial Crithidia fasciculata pol beta, which suggests that the TcI-pol beta plays a role in the organelle. Northern and Western analyses showed that this T. cruzi gene is highly expressed both in proliferative and non-proliferative developmental forms. These results suggest that, in addition to replication of kDNA in proliferative cells, this enzyme may have another function in non-proliferative cells, such as DNA repair role similar to that which has extensively been described in a vast spectrum of eukaryotic cells.

Relationship between DNA ploidy and proliferative cell nuclear antigen index in canine hemangiopericytoma.

PubMed

Kang, Seong-Kwi; Park, Nam-Yong; Cho, Ho-Sung; Shin, Sung-Shik; Kang, Mun-Il; Kim, Sang-Ki; Hyun, Changbaig; Park, In-Chul; Kim, Jong-Tack; Jeong, Cheol; Park, Sung-Hee; Park, Su-Jin; Jeong, Jae-Ho; Kim, You-Jung; Ochiai, Kenji; Umemura, Takashi; Cho, Kyoung-Oh

2006-03-01

The mitotic index is reported to be correlated with recurrence, mean patient survival, and metastasis of canine hemangiopericytoma (CHP). However, to the authors' knowledge, studies investigating the parameters that can predict recurrence or metastasis of CHP with low mitotic index have not been done. To evaluate growth kinetics of CHP with low mitotic index, a retrospective analysis of the proliferative activity by antiproliferative cell nuclear antigen monoclonal antibody and DNA contents by flow cytometry (FCM) was performed with 21 formalin-fixed and paraffin-embedded CHP samples. Of the 21 tumors evaluated by FCM, 6 (26.6%) were aneuploid tumors, and 15 (71.4%) were diploid tumors. There was significant correlation between the PCNA index and ploidy pattern. The diploid group had 39.1 +/- 9.2 PCNA index, whereas the aneuploid group's proliferative cell nuclear antigen (PCNA) index was 63.1 +/- 8.2. The diploid group had mean mitotic index value of 1.140 +/- 0.855, and the aneuploid group had a mean value of 1.067 +/- 0.767. From these results, the CHP samples with low mitotic index were classified into either the aneuploid group with higher PCNA index or the diploid group with lower PCNA index, suggesting that DNA ploidy and proliferative activity may give an indication about malignancy of CHPs with a low mitotic index.

Canine corneal epithelial cells possess a sustained proliferative capacity and generate a spontaneously derived cell line.

PubMed

Morita, Maresuke; Fujita, Naoki; Abe, Momoko; Hayashimoto, Koji; Nakagawa, Takayuki; Nishimura, Ryohei; Tsuzuki, Keiko

2018-06-01

We have previously reported characteristics of canine corneal epithelial cells in vitro and found that canine corneal epithelial cells could maintain their proliferative capacity even after continuous culture without the use of feeder cells and growth promoting additives. The objective of this study was to elucidate proliferative characteristics of canine corneal epithelial cells independent of feeder cells and growth promoting additives, with the aim of developing a spontaneously derived corneal epithelial cell line. Canine and rabbit corneal epithelial cells were harvested from the limbus and cultured with, or without, feeder cells and growth promoting additives, and both were passaged continuously until growth arrest. Canine corneal epithelial cells could proliferate independently, and could be passaged more times than rabbit cells. A canine corneal epithelial cell line, cCEpi, which could be passaged more than 100 times without using feeder cells and growth promoting additives, was established. cCEpi cells maintained a cell morphology close to the primary culture and expressed p63, cytokeratin 15 (K15), and K3. Although changes in colony morphology, shortening of the population doubling time and a heteroploid karyotype were observed, cCEpi was not tumorigenic. Stratified cell sheets cultured from cCEpi were morphologically and immunohistologically similar to sheets cultivated from early passage cells. In conclusion, canine corneal epithelial cells can proliferate independent of feeder cells and growth promoting additives. cCEpi maintains properties similar to normal corneal epithelial cells and could be a useful source for studies in cellular biology and for developing novel therapies. Copyright © 2018 Elsevier Ltd. All rights reserved.

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

Becerikli, Mustafa; Jacobsen, Frank; Rittig, Andrea

Soft tissue sarcomas (STS) are characterized by co-participation of several epigenetic and genetic events during tumorigenesis. Having bypassed cellular senescence barriers during oncogenic transformation, the factors further affecting growth rate of STS cells remain poorly understood. Therefore, we investigated the role of gene silencing (DNA promoter methylation of LINE-1, PTEN), genetic aberrations (karyotype, KRAS and BRAF mutations) as well as their contribution to the proliferation rate and migratory potential that underlies “initial” and “final” passage sarcoma cells. Three different cell lines were used, SW982 (synovial sarcoma), U2197 (malignant fibrous histiocytoma (MFH)) and HT1080 (fibrosarcoma). Increased proliferative potential of final passagemore » STS cells was not associated with significant differences in methylation (LINE-1, PTEN) and mutation status (KRAS, BRAF), but it was dependent on the amount of chromosomal aberrations. Collectively, our data demonstrate that these fairly differentiated/advanced cancer cell lines have still the potential to gain an additional spontaneous growth benefit without external influences and that maintenance of increased proliferative potential towards longevity of STS cells (having crossed senescence barriers) may be independent of overt epigenetic alterations. -- Highlights: Increased proliferative potential of late passage STS cells was: • Not associated with epigenetic changes (methylation changes at LINE-1, PTEN). • Not associated with mutation status of KRAS, BRAF. • Dependent on presence/absence of chromosomal aberrations.« less

CLEFMA- An Anti-Proliferative Curcuminoid from Structure Activity Relationship Studies on 3,5-bis(benzylidene)-4-piperidones

PubMed Central

Lagisetty, Pallavi; Vilekar, Prachi; Sahoo, Kaustuv; Anant, Shrikant; Awasthi, Vibhudutta

2010-01-01

3,5-bis(benzylidene)-4-piperidones are being advanced as synthetic analogs of curcumin for anticancer and anti-inflammatory properties. We performed structure-activity relationship studies, by testing several synthesized 3,5-bis(benzylidene)-4-piperidones for anti-proliferative activity in lung adenocarcinoma H441 cells. Compared to the lead compound 1, or 3,5-bis(2-fluorobenzylidene)-4-piperidone, five compounds were found to be more potent (IC50 < 30 μM), and sixteen compounds possessed reduced cell-killing efficacy (IC50 > 50 μM). Based on the observations, we synthesized 4-[3,5-bis(2-chlorobenzylidene-4-oxo-piperidine-1-yl)-4-oxo-2-butenoic acid] (29 or CLEFMA) as a novel analog of 1. CLEFMA was evaluated for anti-proliferative activity in H441 cells, and was found to be several folds more potent than compound 1. We did not find apoptotic cell population in flow cytometry, and the absence of apoptosis was confirmed by the lack of caspase cleavage. The electron microscopy of H441cells indicated that CLEFMA and compound 1 induce autophagic cell death that was inhibited by specific autophagy inhibitor 3-methyladenine. The results suggest that the potent and novel curcuminoid, CLEFMA, offers an alternative mode of cell death in apoptosis-resistant cancers. PMID:20638855

Curcumin Conjugated with PLGA Potentiates Sustainability, Anti-Proliferative Activity and Apoptosis in Human Colon Carcinoma Cells

PubMed Central

Waghela, Bhargav N.; Sharma, Anupama; Dhumale, Suhashini; Pandey, Shashibahl M.; Pathak, Chandramani

2015-01-01

Curcumin, an ingredient of turmeric, exhibits a variety of biological activities such as anti-inflammatory, anti-atherosclerotic, anti-proliferative, anti-oxidant, anti-cancer and anti-metastatic. It is a highly pleiotropic molecule that inhibits cell proliferation and induces apoptosis in cancer cells. Despite its imperative biological activities, chemical instability, photo-instability and poor bioavailability limits its utilization as an effective therapeutic agent. Therefore, enhancing the bioavailability of curcumin may improve its therapeutic index for clinical setting. In the present study, we have conjugated curcumin with a biodegradable polymer Poly (D, L-lactic-co-glycolic acid) and evaluated its apoptotic potential in human colon carcinoma cells (HCT 116). The results show that curcumin-PLGA conjugate efficiently inhibits cell proliferation and cell survival in human colon carcinoma cells as compared to native curcumin. Additionally, curcumin conjugated with PLGA shows improved cellular uptake and exhibits controlled release at physiological pH as compared to native curcumin. The curcumin-PLGA conjugate efficiently activates the cascade of caspases and promotes intrinsic apoptotic signaling. Thus, the results suggest that conjugation potentiates the sustainability, anti-proliferative and apoptotic activity of curcumin. This approach could be a promising strategy to improve the therapeutic index of cancer therapy. PMID:25692854

Development and characterization of a novel C-terminal inhibitor of Hsp90 in androgen dependent and independent prostate cancer cells

PubMed Central

2011-01-01

Background The molecular chaperone, heat shock protein 90 (Hsp90) has been shown to be overexpressed in a number of cancers, including prostate cancer, making it an important target for drug discovery. Unfortunately, results with N-terminal inhibitors from initial clinical trials have been disappointing, as toxicity and resistance resulting from induction of the heat shock response (HSR) has led to both scheduling and administration concerns. Therefore, Hsp90 inhibitors that do not induce the heat shock response represent a promising new direction for the treatment of prostate cancer. Herein, the development of a C-terminal Hsp90 inhibitor, KU174, is described, which demonstrates anti-cancer activity in prostate cancer cells in the absence of a HSR and describe a novel approach to characterize Hsp90 inhibition in cancer cells. Methods PC3-MM2 and LNCaP-LN3 cells were used in both direct and indirect in vitro Hsp90 inhibition assays (DARTS, Surface Plasmon Resonance, co-immunoprecipitation, luciferase, Western blot, anti-proliferative, cytotoxicity and size exclusion chromatography) to characterize the effects of KU174 in prostate cancer cells. Pilot in vivo efficacy studies were also conducted with KU174 in PC3-MM2 xenograft studies. Results KU174 exhibits robust anti-proliferative and cytotoxic activity along with client protein degradation and disruption of Hsp90 native complexes without induction of a HSR. Furthermore, KU174 demonstrates direct binding to the Hsp90 protein and Hsp90 complexes in cancer cells. In addition, in pilot in-vivo proof-of-concept studies KU174 demonstrates efficacy at 75 mg/kg in a PC3-MM2 rat tumor model. Conclusions Overall, these findings suggest C-terminal Hsp90 inhibitors have potential as therapeutic agents for the treatment of prostate cancer. PMID:22039910

Regional variations in growth plate chondrocyte deformation as predicted by three-dimensional multi-scale simulations.

PubMed

Gao, Jie; Roan, Esra; Williams, John L

2015-01-01

The physis, or growth plate, is a complex disc-shaped cartilage structure that is responsible for longitudinal bone growth. In this study, a multi-scale computational approach was undertaken to better understand how physiological loads are experienced by chondrocytes embedded inside chondrons when subjected to moderate strain under instantaneous compressive loading of the growth plate. Models of representative samples of compressed bone/growth-plate/bone from a 0.67 mm thick 4-month old bovine proximal tibial physis were subjected to a prescribed displacement equal to 20% of the growth plate thickness. At the macroscale level, the applied compressive deformation resulted in an overall compressive strain across the proliferative-hypertrophic zone of 17%. The microscale model predicted that chondrocytes sustained compressive height strains of 12% and 6% in the proliferative and hypertrophic zones, respectively, in the interior regions of the plate. This pattern was reversed within the outer 300 μm region at the free surface where cells were compressed by 10% in the proliferative and 26% in the hypertrophic zones, in agreement with experimental observations. This work provides a new approach to study growth plate behavior under compression and illustrates the need for combining computational and experimental methods to better understand the chondrocyte mechanics in the growth plate cartilage. While the current model is relevant to fast dynamic events, such as heel strike in walking, we believe this approach provides new insight into the mechanical factors that regulate bone growth at the cell level and provides a basis for developing models to help interpret experimental results at varying time scales.

The microenvironment of proliferative diabetic retinopathy supports lymphatic neovascularization.

PubMed

Gucciardo, Erika; Loukovaara, Sirpa; Korhonen, Ani; Repo, Pauliina; Martins, Beatriz; Vihinen, Helena; Jokitalo, Eija; Lehti, Kaisa

2018-06-01

Proliferative diabetic retinopathy (PDR) is a major diabetic microvascular complication characterized by pathological angiogenesis. Several retinopathy animal models have been developed to study the disease mechanisms and putative targets. However, knowledge on the human proliferative disease remains incomplete, relying on steady-state results from thin histological neovascular tissue sections and vitreous samples. New translational models are thus required to comprehensively understand the disease pathophysiology and develop improved therapeutic interventions. We describe here a clinically relevant model, whereby the native multicellular PDR landscape and neo(fibro)vascular processes can be analysed ex vivo and related to clinical data. As characterized by three-dimensional whole-mount immunofluorescence and electron microscopy, heterogeneity in patient-derived PDR neovascular tissues included discontinuous capillaries coupled with aberrantly differentiated, lymphatic-like and tortuous endothelia. Spatially confined apoptosis and proliferation coexisted with inflammatory cell infiltration and unique vascular islet formation. Ex vivo-cultured explants retained multicellularity, islet patterning and capillary or fibrotic outgrowth in response to vitreoretinal factors. Strikingly, PDR neovascular tissues, whose matched vitreous samples enhanced lymphatic endothelial cell sprouting, contained lymphatic-like capillaries in vivo and developed Prox1 + capillaries and sprouts with lymphatic endothelial ultrastructures ex vivo. Among multiple vitreal components, vascular endothelial growth factor C was one factor found at lymphatic endothelium-activating concentrations. These results indicate that the ischaemia-induced and inflammation-induced human PDR microenvironment supports pathological neolymphovascularization, providing a new concept regarding PDR mechanisms and targeting options. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Regional Variations in Growth Plate Chondrocyte Deformation as Predicted By Three-Dimensional Multi-Scale Simulations

PubMed Central

Gao, Jie; Roan, Esra; Williams, John L.

2015-01-01

The physis, or growth plate, is a complex disc-shaped cartilage structure that is responsible for longitudinal bone growth. In this study, a multi-scale computational approach was undertaken to better understand how physiological loads are experienced by chondrocytes embedded inside chondrons when subjected to moderate strain under instantaneous compressive loading of the growth plate. Models of representative samples of compressed bone/growth-plate/bone from a 0.67 mm thick 4-month old bovine proximal tibial physis were subjected to a prescribed displacement equal to 20% of the growth plate thickness. At the macroscale level, the applied compressive deformation resulted in an overall compressive strain across the proliferative-hypertrophic zone of 17%. The microscale model predicted that chondrocytes sustained compressive height strains of 12% and 6% in the proliferative and hypertrophic zones, respectively, in the interior regions of the plate. This pattern was reversed within the outer 300 μm region at the free surface where cells were compressed by 10% in the proliferative and 26% in the hypertrophic zones, in agreement with experimental observations. This work provides a new approach to study growth plate behavior under compression and illustrates the need for combining computational and experimental methods to better understand the chondrocyte mechanics in the growth plate cartilage. While the current model is relevant to fast dynamic events, such as heel strike in walking, we believe this approach provides new insight into the mechanical factors that regulate bone growth at the cell level and provides a basis for developing models to help interpret experimental results at varying time scales. PMID:25885547

Evaluation of the anti-proliferative and cytostatic effect of Citrus sinensis (orange) fruit juice.

PubMed

Chinedu, Enegide; Arome, David; Ameh, Solomon F; Ameh, Gift E

2014-09-01

This work has been designed to evaluate the anti-proliferative and cytostatic effects of Citrus sinensis (orange) fruit juice on rapidly proliferating cells. The study was carried out on the seeds of Sorghum bicolor for 72 h. The mean radicle length (mm) of the seeds was taken at 48 and 72 h. The result showed that when compared with the control, methotrexate, the standard drug showed a significant (P < 0.001) anti-proliferative effect throughout the experiment. The inhibition of the radicle growth was more after 72 h (87.42%). At a dose of 5% (v/v), the juice showed a slightly significant (P < 0.05) effect affect after 72 h; however, there was no significant effect at 48 h. The juice at doses of 10% and 20% (v/v) showed a highly significant (P < 0.001) anti-proliferative effect throughout the experiment; however, the percentage inhibitions were higher at 72 h. At 72 h, the percentage inhibition for juice at 10% (v/v) was 72.37% and at 20% (v/v) was 91.96%. The concentrations of 40% and 60% (v/v) showed cytostatic effects as no appreciable growth of the radicles of the seeds was observed throughout the experiment. The percentage inhibition for 40% (v/v) was 100% and 99.72% for 48 and 72 h, respectively, while that for the juice concentration of 60% (v/v) was 100% throughout the study. The experiment has shown that C. sinensis fruit juice has a potential for causing both anti-proliferative and cytostatic effects on fast proliferating cells and hence cancerous cells.

Anti-proliferative Effects of Androctonus amoreuxi Scorpion and Cerastes cerastes Snake Venoms on Human Prostate Cancer Cells

PubMed Central

Akef, Hassan; Kotb, Nahla; Abo-Elmatty, Dina; Salem, Sayed

2017-01-01

The present study evaluated the effects of Androctonus amoreuxi scorpion venom, Cerastes cerastes snake venom and their mixture on prostate cancer cells (PC3). An MTT assay was used to determine the anti-proliferative effect of the venoms, while quantitative real time PCR was used to evaluate the expression of apoptosis-related genes (Bax and Bcl-2). Furthermore, colorimetric assays were used to measure the levels of malondialdehyde (MDA) and antioxidant enzymes. Our results show that the venoms significantly reduced PC3 cell viability in a dose-dependent manner. On the other hand, these venoms significantly decreased Bcl-2 gene expression. Additionally, C. cerastes venom significantly reduced Bax gene expression, while A. amoreuxi venom and a mixture of A. amoreuxi & C. cerastes venoms did not alter Bax expression. Consequently, these venoms significantly increased the Bax/Bcl-2 ratio and the oxidative stress biomarker MDA. Furthermore, these venoms also increased the activity levels of the antioxidant enzymes, catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase, and glutathione-S-transferase. Overall, the venoms have cytotoxic and anti-proliferative effects on PC3 cells. PMID:28382285

Multiple bidirectional alterations of phenotype and changes in proliferative potential during the in vitro and in vivo passage of clonal mast cell populations derived from mouse peritoneal mast cells

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

Kanakura, Y.; Thompson, H.; Nakano, T.

1988-09-01

Mouse peritoneal mast cells (PMC) express a connective tissue-type mast cell (CTMC) phenotype, including reactivity with the heparin-binding fluorescent dye berberine sulfate and incorporation of (35S) sulfate predominantly into heparin proteoglycans. When PMC purified to greater than 99% purity were cultured in methylcellulose with IL-3 and IL-4, approximately 25% of the PMC formed colonies, all of which contained both berberine sulfate-positive and berberine sulfate-negative mast cells. When these mast cells were transferred to suspension culture, they generated populations that were 100% berberine sulfate-negative, a characteristic similar to that of mucosal mast cells (MMC), and that synthesized predominantly chondroitin sulfate (35S)more » proteoglycans. When ''MMC-like'' cultured mast cells derived from WBB6F1-+/+ PMC were injected into the peritoneal cavities of mast cell-deficient WBB6F1-W/Wv mice, the adoptively transferred mast cell population became 100% berberine sulfate-positive. In methylcellulose culture, these ''second generation PMC'' formed clonal colonies containing both berberine sulfate-positive and berberine sulfate-negative cells, but exhibited significantly less proliferative ability than did normal +/+ PMC. Thus, clonal mast cell populations initially derived from single PMC exhibited multiple and bidirectional alterations between CTMC-like and MMC-like phenotypes. However, this process was associated with a progressive diminution of the mast cells' proliferative ability.« less

Autophagic Cell Death, Polyploidy and Senescence Induced in Breast Tumor Cells by the Substituted Pyrrole JG-03-14, a Novel Microtubule Poison

PubMed Central

Arthur, Christopher R.; Gupton, John T.; Kellogg, Glen E.; Yeudall, W. Andrew; Cabot, Myles C.; Newsham, Irene; Gewirtz, David A.

2007-01-01

JG-03-14, a substituted pyrrole that inhibits microtubule polymerization, was screened against MCF-7 (p53 wild type), MDA-MB 231 (p53 mutant), MCF-7/caspase 3 and MCF-7/ADR (multidrug resistant) breast tumor cell lines. Cell viability and growth inhibition were assessed by the crystal violet dye assay. Apoptosis was evaluated by the TUNEL assay, cell cycle distribution by flow cytometry, autophagy by acridine orange staining of vesicle formation, and senescence based on β-galactosidase staining and cell morphology. Our studies indicate that exposure to JG-03-14, at a concentration of 500 nM, induces time dependent cell death in the MCF-7 and MDA-MB 231 cell lines. In MCF-7 cells, a residual surviving cell population was found to be senescent; in contrast, there was no surviving senescent population in treated MDA-MB 231 cells. No proliferative recovery was detected over a period of 15 days post-treatment in either cell line. Both the TUNEL assay and FLOW cytometry indicated a relatively limited degree of apoptosis (< 10%) in response to drug treatment in MCF-7 cells with more extensive apoptosis (but < 20%) in MDA-MB231 cells; acidic vacuole formation indicative of autophagic cell death was relatively extensive in both MCF-7 and MDA-MB231 cells. In addition, JG-03-14 induced the formation of a large hyperdiploid cell population in MDA-MB231 cells. JG-03-14 also demonstrated pronounced anti-proliferative activity in MCF-7/caspase 3 cells and in the MCF-7/ADR cell line. The observation that JG-03-14 promotes autophagic cell death and also retains activity in tumor cells expressing the multidrug resistance pump indicates that novel microtubule poisons of the substituted pyrroles class may hold promise in the treatment of breast cancer. PMID:17692290

B-cell activation with CD40L or CpG measures the function of B-cell subsets and identifies specific defects in immunodeficient patients.

PubMed

Marasco, Emiliano; Farroni, Chiara; Cascioli, Simona; Marcellini, Valentina; Scarsella, Marco; Giorda, Ezio; Piano Mortari, Eva; Leonardi, Lucia; Scarselli, Alessia; Valentini, Diletta; Cancrini, Caterina; Duse, Marzia; Grimsholm, Ola; Carsetti, Rita

2017-01-01

Around 65% of primary immunodeficiencies are antibody deficiencies. Functional tests are useful tools to study B-cell functions in vitro. However, no accepted guidelines for performing and evaluating functional tests have been issued yet. Here, we report our experience on the study of B-cell functions in infancy and throughout childhood. We show that T-independent stimulation with CpG measures proliferation and differentiation potential of memory B cells. Switched memory B cells respond better than IgM memory B cells. On the other hand, CD40L, a T-dependent stimulus, does not induce plasma cell differentiation, but causes proliferation of naïve and memory B cells. During childhood, the production of plasmablasts in response to CpG increases with age mirroring the development of memory B cells. The response to CD40L does not change with age. In patients with selective IgA deficiency (SIgAD), we observed that switched memory B cells are reduced due to the absence of IgA memory B cells. In agreement, IgA plasma cells are not generated in response to CpG. Unexpectedly, B cells from SIgAD patients show a reduced proliferative response to CD40L. Our results demonstrate that functional tests are an important tool to assess the functions of the humoral immune system. © 2016 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mitogenic activity of a water-soluble adjuvant (Bu-WSA) obtained from Bacterionema matruchotii. IV. Synergistic effects of Bu-WSA on Concanavalin A-induced proliferative response of human peripheral blood lymphocytes.

PubMed

Nitta, T; Okumura, S; Tsushi, M; Nakano, M

1982-01-01

Butanol-extracted water-soluble adjuvant (Bu-WSA) obtained from Bacterionema matruchotii was cultured with peripheral blood mononuclear cells (PBM) in the presence of sub- and/or supra-optimal mitogenic concentrations of concanavalin A (Con A). The addition of Bu-WSA resulted in increased tritiated thymidine incorporation above that produced by Con A alone. Bu-WSA by itself is not mitogenic for PBM and in fact produced a decrease in thymidine uptake compared to the control. We investigated the response of subpopulation(s) of PBM to Bu-WSA, Con A and a mixture of Bu-WSA and Con A. Separation of PBM into purified T cells, B cells and macrophages showed that cell-cell cooperation of T cells with B cells or macrophages is necessary for the observed synergistic effect of Bu-WSA with Con A. A marked increase in thymidine incorporation by the mixture of T and B cell populations occurred, while only a small amount of thymidine was incorporated when the B cell population was absent. Mitomycin treatment revealed that the response could be ascribed to the T-cell response with a B-cell helper effect. Moreover, Con A and Bu-WSA appeared to act on the same T cell population. This model may provide unique information about the activation of human peripheral blood T cells compared with the activation of these cells by other mitogens.

Antiproliferative and Antiangiogenic Effects of Punica granatum Juice (PGJ) in Multiple Myeloma (MM)

PubMed Central

Tibullo, Daniele; Caporarello, Nunzia; Giallongo, Cesarina; Anfuso, Carmelina Daniela; Genovese, Claudia; Arlotta, Carmen; Puglisi, Fabrizio; Parrinello, Nunziatina L.; Bramanti, Vincenzo; Romano, Alessandra; Lupo, Gabriella; Toscano, Valeria; Avola, Roberto; Brundo, Maria Violetta; Di Raimondo, Francesco; Raccuia, Salvatore Antonio

2016-01-01

Multiple myeloma (MM) is a clonal B-cell malignancy characterized by an accumulation of clonal plasma cells (PC) in the bone marrow (BM) leading to bone destruction and BM failure. Despite recent advances in pharmacological therapy, MM remains a largely incurable pathology. Therefore, novel effective and less toxic agents are urgently necessary. In the last few years, pomegranate has been studied for its potential therapeutic properties including treatment and prevention of cancer. Pomegranate juice (PGJ) contains a number of potential active compounds including organic acids, vitamins, sugars, and phenolic components that are all responsible of the pro-apoptotic effects observed in tumor cell line. The aim of present investigation is to assess the antiproliferative and antiangiogenic potential of the PGJ in human multiple myeloma cell lines. Our data demonstrate the anti-proliferative potential of PGJ in MM cells; its ability to induce G0/G1 cell cycle block and its anti-angiogenic effects. Interestingly, sequential combination of bortezomib/PGJ improved the cytotoxic effect of the proteosome inhibitor. We investigated the effect of PGJ on angiogenesis and cell migration/invasion. Interestingly, we observed an inhibitory effect on the tube formation, microvessel outgrowth aorting ring and decreased cell migration and invasion as showed by wound-healing and transwell assays, respectively. Analysis of angiogenic genes expression in endothelial cells confirmed the anti-angiogenic properties of pomegranate. Therefore, PGJ administration could represent a good tool in order to identify novel therapeutic strategies for MM treatment, exploiting its anti-proliferative and anti-angiogenic effects. Finally, the present research supports the evidence that PGJ could play a key role of a future therapeutic approach for treatment of MM in order to optimize the pharmacological effect of bortezomib, especially as adjuvant after treatment. PMID:27706074

Cultured Human Fibroblast Biostimulation Using a 940 nm Diode Laser

PubMed Central

Illescas-Montes, Rebeca; Melguizo-Rodríguez, Lucía; Manzano-Moreno, Francisco Javier; García-Martínez, Olga; Ruiz, Concepción

2017-01-01

Background: Fibroblasts are the main cells involved in regeneration during wound healing. The objective was to determine the effect of 940 nm diode laser on cultured human fibroblasts using different irradiation regimens. Methods: The CCD-1064Sk human epithelial fibroblast cell line was treated with a 940 nm diode laser at different energy doses (power: 0.2–1 W and energy density: 1–7 J/cm2) using different transmission modes (continuous or pulsed). The effect on cell growth at 24 and 72 h post-treatment was examined by measuring the proliferative capacity, the impact on the cell cycle, and the effect on cell differentiation. Results: fibroblast proliferative capacity was increased at 24 and 72 h post-treatment as a function of the energy dose. The greatest increase was observed with a power of 0.2 or 0.5 W and energy density between 1 and 4 J/cm2; no difference was observed between continuous and pulsed modes. There were no significant differences in cell cycle between treated groups and controls. α-actin expression was increased by treatment, indicating enhanced cell differentiation. Conclusion: The 940 nm diode laser has biostimulating effects on fibroblasts, stimulating proliferative capacity and cell differentiation without altering the cell cycle. Further researches are necessary to explore its potential clinical usefulness in wound healing. PMID:28773152

Arctigenin in combination with quercetin synergistically enhances the anti-proliferative effect in prostate cancer cells

PubMed Central

Wang, Piwen; Phan, Tien; Gordon, David; Chung, Seyung; Henning, Susanne M.; Vadgama, Jaydutt V.

2014-01-01

Scope We investigated whether a combination of two promising chemopreventive agents arctigenin and quercetin increases the anti-carcinogenic potency at lower concentrations than necessary when used individually in prostate cancer. Methods and results Androgen-dependent LAPC-4 and LNCaP prostate cancer cells were treated with low doses of arctigenin and quercetin alone or in combination for 48h. The anti-proliferative activity of arctigenin was 10-20 fold stronger than quercetin in both cell lines. Their combination synergistically enhanced the anti-proliferative effect, with a stronger effect in androgen receptor (AR) wild-type LAPC-4 cells than in AR mutated LNCaP cells. Arctigenin demonstrated a strong ability to inhibit AR protein expression in LAPC-4 cells. The combination treatment significantly inhibited both AR and PI3K/Akt pathways compared to control. A protein array analysis revealed that the mixture targets multiple pathways particularly in LAPC-4 cells including Stat3 pathway. The mixture significantly inhibited the expression of several oncogenic microRNAs including miR-21, miR-19b, and miR-148a compared to control. The mixture also enhanced the inhibition of cell migration in both cell lines compared to individual compounds tested. Conclusion The combination of arctigenin and quercetin, that target similar pathways, at low physiological doses, provides a novel regimen with enhanced chemoprevention in prostate cancer. PMID:25380086

Interleukin-7 Availability Is Maintained by a Hematopoietic Cytokine Sink Comprising Innate Lymphoid Cells and T Cells.

PubMed

Martin, Christopher E; Spasova, Darina S; Frimpong-Boateng, Kwesi; Kim, Hee-Ok; Lee, Minji; Kim, Kwang Soon; Surh, Charles D

2017-07-18

Interleukin-7 (IL-7) availability determines the size and proliferative state of the resting T cell pool. However, the mechanisms that regulate steady-state IL-7 amounts are unclear. Using experimental lymphopenic mouse models and IL-7-induced homeostatic proliferation to measure IL-7 availability in vivo, we found that radioresistant cells were the source of IL-7 for both CD4 + and CD8 + T cells. Hematopoietic lineage cells, although irrelevant as a source of IL-7, were primarily responsible for limiting IL-7 availability via their expression of IL-7R. Unexpectedly, innate lymphoid cells were found to have a potent influence on IL-7 amounts in the primary and secondary lymphoid tissues. These results demonstrate that IL-7 homeostasis is achieved through consumption by multiple subsets of innate and adaptive immune cells. Copyright © 2017 Elsevier Inc. All rights reserved.

Polyclonal cell activity of a repeat peptide derived from the sequence of an 85-kilodalton surface protein of Trypanosoma cruzi trypomastigotes.

PubMed Central

Pestel, J; Defoort, J P; Gras-Masse, H; Afchain, D; Capron, A; Tartar, A; Ouaissi, A

1992-01-01

Some in vitro and in vivo biological activities of an octadecapeptide derived from an 85-kDa surface protein of Trypanosoma cruzi trypomastigote were studied. The peptide coupled to a carrier protein induced the proliferative response of lymph node cells from mice immunized with various antigens. Moreover, sera from mice immunized with the coupled peptide were found to contain antibodies against a number of self and nonself antigens: fibronectin, bovine serum albumin, myosin, tetanus toxoid, ovalbumin, keyhole limpet hemocyanin, and DNA. These results are discussed in the context of Chagas' disease immunopathology. PMID:1730508

TGF-beta-induced apoptosis in human thyrocytes is mediated by p27kip1 reduction and is overridden in neoplastic thyrocytes by NF-kappaB activation.

PubMed

Bravo, Susana B; Pampín, Sandra; Cameselle-Teijeiro, José; Carneiro, Carmen; Domínguez, Fernando; Barreiro, Francisco; Alvarez, Clara V

2003-10-30

Millions of people worldwide suffer goiter, a proliferative disease of the follicular cells of the thyroid that may become neoplastic. Thyroid neoplasms have low proliferative index, low apoptotic index and a high incidence of metastasis. TGF-beta is overexpressed in thyroid follicular tumor cells. To investigate the role of TGF-beta in thyroid tumor progression, we established cultures of human thyrocytes from different proliferative pathologies (Grave's disease, multinodular goiter, follicular adenoma, papillary carcinoma), lymph node metastasis, and a normal thyroid sample. All cultures maintained the thyrocyte phenotype. TGF-beta induced cell-cycle arrest in all cultures, in contrast with results reported for other epithelial tumors. In deprived medium, TGF-beta induced apoptosis in normal thyrocyte cultures and all neoplastic cultures except the metastatic cultures. This apoptosis was mediated by a reduction in p27kip1 levels, inducing cell-cycle initiation. Antisense p27 expression induced apoptosis in the absence of TGF-beta. By contrast, in cells in which p27 was overexpressed, TGF-beta had a survival effect. In growth medium, a net survival effect occurs in neoplastic thyrocytes only, not normal thyrocytes, due to activation of the NF-kappaB survival program. Together, these findings suggest that (a) thyroid neoplasms are due to reduced apoptosis, not increased division, in line with the low proliferative index of these pathologies, and (b) TGF-beta induces apoptosis in normal thyrocytes via p27 reduction, but that in neoplastic thyrocytes this effect is overridden by activation of the NF-kappaB program.

GSK3 as a Sensor Determining Cell Fate in the Brain.

PubMed

Cole, Adam R

2012-01-01

Glycogen synthase kinase 3 (GSK3) is an unusual serine/threonine kinase that controls many neuronal functions, including neurite outgrowth, synapse formation, neurotransmission, and neurogenesis. It mediates these functions by phosphorylating a wide range of substrates involved in gene transcription, metabolism, apoptosis, cytoskeletal dynamics, signal transduction, lipid membrane dynamics, and trafficking, amongst others. This complicated list of diverse substrates generally follow a more simple pattern: substrates negatively regulated by GSK3-mediated phosphorylation favor a proliferative/survival state, while substrates positively regulated by GSK3 favor a more differentiated/functional state. Accordingly, GSK3 activity is higher in differentiated cells than undifferentiated cells and physiological (Wnt, growth factors) and pharmacological inhibitors of GSK3 promote the proliferative capacity of embryonic stem cells. In the brain, the level of GSK3 activity influences neural progenitor cell proliferation/differentiation in neuroplasticity and repair, as well as efficient neurotransmission in differentiated adult neurons. While defects in GSK3 activity are unlikely to be the primary cause of neurodegenerative diseases, therapeutic regulation of its activity to promote a proliferative/survival versus differentiated/mature functional environment in the brain could be a powerful strategy for treatment of neurodegenerative and other mental disorders.

GSK3 as a Sensor Determining Cell Fate in the Brain

PubMed Central

Cole, Adam R.

2012-01-01

Glycogen synthase kinase 3 (GSK3) is an unusual serine/threonine kinase that controls many neuronal functions, including neurite outgrowth, synapse formation, neurotransmission, and neurogenesis. It mediates these functions by phosphorylating a wide range of substrates involved in gene transcription, metabolism, apoptosis, cytoskeletal dynamics, signal transduction, lipid membrane dynamics, and trafficking, amongst others. This complicated list of diverse substrates generally follow a more simple pattern: substrates negatively regulated by GSK3-mediated phosphorylation favor a proliferative/survival state, while substrates positively regulated by GSK3 favor a more differentiated/functional state. Accordingly, GSK3 activity is higher in differentiated cells than undifferentiated cells and physiological (Wnt, growth factors) and pharmacological inhibitors of GSK3 promote the proliferative capacity of embryonic stem cells. In the brain, the level of GSK3 activity influences neural progenitor cell proliferation/differentiation in neuroplasticity and repair, as well as efficient neurotransmission in differentiated adult neurons. While defects in GSK3 activity are unlikely to be the primary cause of neurodegenerative diseases, therapeutic regulation of its activity to promote a proliferative/survival versus differentiated/mature functional environment in the brain could be a powerful strategy for treatment of neurodegenerative and other mental disorders. PMID:22363258

Mdm4 loss in the intestinal epithelium leads to compartmentalized cell death but no tissue abnormalities

PubMed Central

Valentin-Vega, Yasmine A.; Box, Neil; Terzian, Tamara; Lozano, Guillermina

2014-01-01

Mdm4 is a critical inhibitor of the p53 tumor suppressor. Mdm4 null mice die early during embryogenesis due to increased p53 activity. In this study, we explore the role that Mdm4 plays in the intestinal epithelium by crossing mice carrying the Mdm4 floxed allele to mice with the Villin Cre transgene. Our data show that loss of Mdm4 (Mdm4intΔ) in this tissue resulted in viable animals with no obvious morphological abnormalities. However, these mutants displayed increased p53 levels and apoptosis exclusively in the proliferative compartment of the intestinal epithelium. This phenotype was completely rescued in a p53 null background. Notably, the observed compartmentalized apoptosis in proliferative intestinal epithelial cells was not due to restricted Mdm4 expression in this region. Thus, in this specific cellular context, p53 is negatively regulated by Mdm4 exclusively in highly proliferative cells. PMID:19371999

[Effect of low-intensity electromagnetic fields of industrial frequency on the ultrastructure and proliferative activity of rat's thymus cells].

PubMed

Zhitkevich, T I; Bokut', T B; Netukova, N I

2001-01-01

Effects of two types of low-intensity electromagnetic fields (EMF) of industrial frequency (50 Hz) on the fine structure and proliferative activity of thymic cells in white rats were studied. It was found that a weak EMF with a prevailing electrical component (380-480 V/m, 120-140 nT1) did not affect the DNA synthesis intensity. An EMF with a stronger magnetic induction (10-15 V/m, 800-1500 nT1) diminished the glucose-6-phosphate dehydrogenase activity and proliferative processes in cultured stimulated lymphocytes. Electron microscopic investigation of the thymus after both types of exposure revealed an accumulation of lymphocytes with pyknotic nuclei and electron-dense cytoplasm, as well as hypoplasia of the vascular endothelium. At the same time, EMF with a prevailing magnetic component produced a more marked negative effect on the ultrastructure of thymic cells, which indicated a lowered secretory activity of epitheliocytes.

Expression of retinoblastoma gene product (pRb) in mantle cell lymphomas. Correlation with cyclin D1 (PRAD1/CCND1) mRNA levels and proliferative activity.

PubMed Central

Jares, P.; Campo, E.; Pinyol, M.; Bosch, F.; Miquel, R.; Fernandez, P. L.; Sanchez-Beato, M.; Soler, F.; Perez-Losada, A.; Nayach, I.; Mallofré, C.; Piris, M. A.; Montserrat, E.; Cardesa, A.

1996-01-01

Mantle cell lymphomas (MCLs) are molecularly characterized by bcl-1 rearrangement and constant cyclin D1 (PRAD-1/CCND1) gene overexpression. Cyclin D1 is a G1 cyclin that participates in the control of the cell cycle progression by interacting with the retinoblastoma gene product (pRb). Inactivation of the Rb tumor suppressor gene has been implicated in the development of different types of human tumors including some high grade non-Hodgkin's lymphomas. To determine the role of the retinoblastoma gene in the pathogenesis of MCLs and its possible interaction with cyclin D1, pRb expression was examined in 23 MCLs including 17 typical and 6 blastic variants by immunohistochemistry and Western blot. Rb gene structure was studied in 13 cases by Southern blot. Cytogenetic analysis was performed in 5 cases. The results were compared with the cyclin D1 mRNA levels examined by Northern analysis, and the proliferative activity of the tumors was measured by Ki-67 growth fraction and flow cytometry. pRb was expressed in all MCLs. The expression varied from case to case (mean, 14.1% of positive cells; range, 1.3 to 42%) with a significant correlation with the proliferative activity of the tumors (mitotic index r = 0.85; Ki-67 r = 0.7; S phase = 0.73). Blastic variants showed higher numbers of pRb-positive cells (mean, 29%) than the typical cases (10%; P < 0.005) by immunohistochemistry and, concordantly, higher levels of expression by Western blot. In addition, the blastic cases also had an increased expression of the phosphorylated protein. No alterations in Rb gene structure were observed by Southern blot analysis. Cyclin D1 mRNA levels were independent of pRb expression and the proliferative activity of the tumors. These findings suggest that pRb in MCLs is normally regulated in relation to the proliferative activity of the tumors. Cyclin D1 overexpression may play a role in the maintenance of cell proliferation by overcoming the suppressive growth control of pRb. Images Figure 1 Figure 2 Figure 4 PMID:8623927

In vitro anti-proliferative and anti-angiogenic activities of thalidomide dithiocarbamate analogs.

PubMed

El-Aarag, Bishoy Y A; Kasai, Tomonari; Zahran, Magdy A H; Zakhary, Nadia I; Shigehiro, Tsukasa; Sekhar, Sreeja C; Agwa, Hussein S; Mizutani, Akifumi; Murakami, Hiroshi; Kakuta, Hiroki; Seno, Masaharu

2014-08-01

Inhibition of angiogenesis is currently perceived as a promising strategy in the treatment of cancer. The anti-angiogenicity of thalidomide has inspired a second wave of research on this teratogenic drug. The present study aimed to investigate the anti-proliferative and anti-angiogenic activities of two thalidomide dithiocarbamate analogs by studying their anti-proliferative effects on human umbilical vein endothelial cells (HUVECs) and MDA-MB-231 human breast cancer cell lines. Their action on the expression levels of IL-6, IL-8, TNF-α, VEGF165, and MMP-2 was also assessed. Furthermore, their effect on angiogenesis was evaluated through wound healing, migration, tube formation, and nitric oxide (NO) assays. Results illustrated that the proliferation of HUVECs and MDA-MB-231 cells was not significantly affected by thalidomide at 6.25-100μM. Thalidomide failed to block angiogenesis at similar concentrations. By contrast, thalidomide dithiocarbamate analogs exhibited significant anti-proliferative action on HUVECs and MDA-MB-231 cells without causing cytotoxicity and also showed powerful anti-angiogenicity in wound healing, migration, tube formation, and NO assays. Thalidomide analogs 1 and 2 demonstrated more potent activity to suppress expression levels of IL-6, IL-8, TNF-α, VEGF165, and MMP-2 than thalidomide. Analog 1 consistently, showed the highest potency and efficacy in all the assays. Taken together, our results support further development and evaluation of novel thalidomide analogs as anti-tumor and anti-angiogenic agents. Copyright © 2014. Published by Elsevier B.V.

Activation of the PI3K/AKT pathway by microRNA-22 results in CLL B-cell proliferation.

PubMed

Palacios, F; Abreu, C; Prieto, D; Morande, P; Ruiz, S; Fernández-Calero, T; Naya, H; Libisch, G; Robello, C; Landoni, A I; Gabus, R; Dighiero, G; Oppezzo, P

2015-01-01

Chronic lymphocytic leukemia (CLL) is characterized by accumulation of clonal B cells arrested in G0/G1 stages that coexist, in different proportions, with proliferative B cells. Understanding the crosstalk between the proliferative subsets and their milieu could provide clues on CLL biology. We previously identified one of these subpopulations in the peripheral blood from unmutated patients that appears to be a hallmark of a progressive disease. Aiming to characterize the molecular mechanism underlying this proliferative behavior, we performed gene expression analysis comparing the global mRNA and microRNA expression of this leukemic subpopulation, and compared it with their quiescent counterparts. Our results suggest that proliferation of this fraction depend on microRNA-22 overexpression that induces phosphatase and tensin homolog downregulation and phosphoinositide 3-kinase (PI3K)/AKT pathway activation. Transfection experiments demonstrated that miR-22 overexpression in CLL B cells switches on PI3K/AKT, leading to downregulation of p27(-Kip1) and overexpression of Survivin and Ki-67 proteins. We also demonstrated that this pathway could be triggered by microenvironment signals like CD40 ligand/interleukin-4 and, more importantly, that this regulatory loop is also present in lymph nodes from progressive unmutated patients. Altogether, these results underline the key role of PI3K/AKT pathway in the generation of the CLL proliferative pool and provide additional rationale for the usage of PI3K inhibitors.

The downregulation of Wnt/β-catenin signaling pathway is associated with zinc deficiency-induced proliferative deficit of C17.2 neural stem cells.

PubMed

Zhao, Jianya; Han, Jingling; Jiang, Junkang; Shi, Shangshi; Ma, Xia; Liu, Xinhang; Wang, Cheng; Nie, Xiaoke; He, Yunhua; Jiang, Shengyang; Wan, Chunhua

2015-07-30

Zinc is an essential nutrient that is important for normal brain development. Zinc deficiency has been linked to aberrant neurological development and functioning. However, the molecular mechanisms underlying Zinc deficiency-induced neurological disorders remain largely elusive. In the present study, we showed that the proliferation of C17.2 neural stem cells (NSCs) was evidently impaired after exposed to low levels of Zinc chelator, N,N,N',N'-tetrakis-(2-pyridylmethy) ethylenediamine (TPEN). In addition, we found that TPEN-induced proliferative deficit of NSCs was related with significant downregulation of Wnt/β-catenin signaling. Zinc deficiency impaired the proliferation of neural stem cells in dose- and time-dependent manners. Western blot revealed that the levels of p-Ser9-glycogensynthase kinase-3β (p-GSK-3β) and β-catenin were remarkably downregulated during TPEN-induced C17.2 proliferative impairment. Moreover, immunofluorescent analysis indicated that the level of nuclear β-catenin was apparently decreased following TPEN exposure. Furthermore, application with GSK-3β inhibitor lithium chloride (LiCl) reversed TPEN-induced downregulation of β-catenin and impairment of cell proliferation. Flow cytometry analysis also showed that TPEN-induced impairment of NSC proliferation could be reversed by LiCl. Taken together, these findings suggested that the disturbance of canonical Wnt/β-catenin signaling pathway partially accounted for Zinc deficiency-induced proliferative impairment of NSCs. Copyright © 2015 Elsevier B.V. All rights reserved.

Lipophilization of somatostatin analog RC-160 with long chain fatty acid improves its anti-proliferative activity on human oral carcinoma cells in vitro.

PubMed

Dasgupta, P; Singh, A T; Mukherjee, R

2000-03-01

Oral cancer which comprises about 40% of total cancers in India, has one of the lowest relative survival rates of all cancers. Epidermal growth factor (EGF) has been known to play a role in the proliferation/malignant transformation of oral neoplasms. Since, the somatostatin analog RC-160 is reported to be a potent inhibitor of EGF stimulated cell proliferation, its anti-proliferative activity in the human oral carcinoma cell line KB was investigated, in this study. RC-160 was found to potently inhibit EGF-induced proliferation in KB cells in vitro, suggesting a therapeutic potential of the same in oral carcinoma. However, the therapeutic potential of RC-160 is limited by its short serum half life. To overcome this limitation, fatty acids namely butanoic acid and myristic acid individually were coupled to RC-160. The lipophilized derivatives of RC-160 were synthesized, purified and characterized. The anti-proliferative activity of lipophilized derivatives of RC-160 on KB cells was evaluated in vitro. Myristoyl-RC-160 (0.75 nM) inhibited the growth of KB cells at a 10-fold lower concentration relative to RC-160 (8.8 nM) and at a 100-fold lower concentration relative to butanoyl-RC-160 (0.83 microM) (p<0.001). The affinity of RC-160 towards somatostatin receptors remains unaltered by lipophilization. The signaling pathways underlying the antineoplastic activity of these lipopeptides are similar to RC-160, and do not involve the stimulation of a protein tyrosine phosphatase or a serine threonine phosphatase 1A and 2A. The anti-proliferative activity of the lipopeptides was found to be mediated by somatostatin receptors and correlates with the inhibition of protein tyrosine kinase activity and decrease in intracellular cAMP levels. Myristoyl-RC-160 displayed significantly greater resistance towards trypsin and serum degradation than RC-160 (p<0.01). These findings demonstrate that RC-160 can inhibit the growth of oral cancer cells in vitro. Lipophilization of RC-160 with long chain fatty acids like myristic acid improves its stability and anti-proliferative activity, in human oral carcinoma cells in vitro, thereby enhancing the scope of improving its therapeutic index.

The potential for chemical mixtures from the environment to enable the cancer hallmark of sustained proliferative signalling

PubMed Central

Engström, Wilhelm; Darbre, Philippa; Eriksson, Staffan; Gulliver, Linda; Hultman, Tove; Karamouzis, Michalis V.; Klaunig, James E.; Mehta, Rekha; Moorwood, Kim; Sanderson, Thomas; Sone, Hideko; Vadgama, Pankaj; Wagemaker, Gerard; Ward, Andrew; Singh, Neetu; Al-Mulla, Fahd; Al-Temaimi, Rabeah; Amedei, Amedeo; Colacci, Anna Maria; Vaccari, Monica; Mondello, Chiara; Scovassi, A. Ivana; Raju, Jayadev; Hamid, Roslida A.; Memeo, Lorenzo; Forte, Stefano; Roy, Rabindra; Woodrick, Jordan; Salem, Hosni K.; Ryan, Elizabeth; Brown, Dustin G.; Bisson, William H.

2015-01-01

The aim of this work is to review current knowledge relating the established cancer hallmark, sustained cell proliferation to the existence of chemicals present as low dose mixtures in the environment. Normal cell proliferation is under tight control, i.e. cells respond to a signal to proliferate, and although most cells continue to proliferate into adult life, the multiplication ceases once the stimulatory signal disappears or if the cells are exposed to growth inhibitory signals. Under such circumstances, normal cells remain quiescent until they are stimulated to resume further proliferation. In contrast, tumour cells are unable to halt proliferation, either when subjected to growth inhibitory signals or in the absence of growth stimulatory signals. Environmental chemicals with carcinogenic potential may cause sustained cell proliferation by interfering with some cell proliferation control mechanisms committing cells to an indefinite proliferative span. PMID:26106143

Fibrin/fibrinogen degradation products in children with renal disease

PubMed Central

Uttley, W. S.; Maxwell, Heather; Cash, J. D.

1974-01-01

Fibrin/fibrinogen degradation products (FDP) were measured in the serum and urine of children with various forms of renal disease. Serum FDP was raised both with nephrosis and with active proliferative nephritis. Urine FDP was rarely present in nephrosis but was significantly increased during the active phase of proliferative nephritis and also in urinary tract infection with frank haematuria. Urinary FDP correlated with total urinary protein in proliferative nephritis but not in nephrosis, nor did it correlate with serum FDP in either condition. The major application of urinary FDP determination in clinical practice is as an indicator of activity and possible response to treatment in the management of active proliferative nephritis. PMID:4817446

Scanning and transmission electron microscopy of the damage to small intestinal mucosa following X irradiation or hyperthermia

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

Carr, K.E.; Hume, S.P.; Marigold, J.C.

Scanning and transmission electron microscopy (S.E.M. and T.E.M.) and resin histology have been used to investigate the effects on mouse small intestinal villi of heating at 43 degrees C for 20 minutes and of irradiation with 10 Gy X-rays. Damage after irradiation included conical villi and giant cells. Damage after heating included the production of conical and rudimentary villi and the stacking of enterocytes. Individual cells showed signs of abnormalities in their cell membranes, nuclei and cytoplasmic components. The differences in the response after irradiation and hyperthermia are linked to the fact that heating has a primary effect on villousmore » structure, whereas irradiation mainly affects the proliferative pool of crypt cells.« less

The PPARα/p16INK4a Pathway inhibits Vascular Smooth Muscle Cell Proliferation by repressing Cell Cycle-dependent Telomerase Activation

PubMed Central

Gizard, Florence; Nomiyama, Takashi; Zhao, Yue; Findeisen, Hannes M.; Heywood, Elizabeth B.; Jones, Karrie L.; Staels, Bart; Bruemmer, Dennis

2009-01-01

Peroxisome Proliferator-Activated Receptor (PPAR) α, the molecular target for fibrates used to treat dyslipidemia, exerts pleiotropic effects on vascular cells. In vascular smooth muscle cells (VSMCs), we have previously demonstrated that PPARα activation suppresses G1→S cell cycle progression by targeting the cyclin-dependent kinase inhibitor p16INK4a (p16). In the present study, we demonstrate that this inhibition of VSMC proliferation by PPARα is mediated through a p16-dependent suppression of telomerase activity, which has been implicated in key cellular functions including proliferation. PPARα activation inhibited mitogen-induced telomerase activity by repressing the catalytic subunit telomerase reverse transcriptase (TERT) through negative cross-talk with an E2F-1-dependent trans-activation of the TERT promoter. This trans-repression involved the recruitment of the retinoblastoma (RB) family proteins p107 and p130 to the TERT promoter resulting in impaired E2F-1 binding, an effect which was dependent on p16. The inhibition of cell proliferation by PPARα activation was lost in VSMC following TERT overexpression or knock-down, pointing to a key role of telomerase as a target for the antiproliferative effects of PPARα. Finally, we demonstrate that PPARα agonists suppress telomerase activation during the proliferative response following vascular injury indicating that these findings are applicable in vivo. In concert, these results demonstrate that the anti-proliferative effects of PPARα in VSMCs depend on the suppression of telomerase activity by targeting the p16/RB/E2F transcriptional cascade. PMID:18818403

Branched-chain amino acid supplementation and the immune response of long-distance athletes.

PubMed

Bassit, Reinaldo A; Sawada, Letícia A; Bacurau, Reury F P; Navarro, Franciso; Martins, Eivor; Santos, Ronaldo V T; Caperuto, Erico C; Rogeri, Patrícia; Costa Rosa, Luís F B P

2002-05-01

Intense long-duration exercise has been associated with immunosuppression, which affects natural killer cells, lymphokine-activated killer cells, and lymphocytes. The mechanisms involved, however, are not fully determined and seem to be multifactorial, including endocrine changes and alteration of plasma glutamine concentration. Therefore, we evaluated the effect of branched-chain amino acid supplementation on the immune response of triathletes and long-distance runners. Peripheral blood was collected prior to and immediately after an Olympic Triathlon or a 30k run. Lymphocyte proliferation, cytokine production by cultured cells, and plasma glutamine were measured. After the exercise bout, athletes from the placebo group presented a decreased plasma glutamine concentration that was abolished by branched-chain amino acid supplementation and an increased proliferative response in their peripheral blood mononuclear cells. Those cells also produced, after exercise, less tumor necrosis factor, interleukins-1 and -4, and interferon and 48% more interleukin-2. Supplementation stimulated the production of interleukin-2 and interferon after exercise and a more pronounced decrease in the production of interleukin-4, indicating a diversion toward a Th1 type immune response. Our results indicate that branched-chain amino acid (BCAA) supplementation recovers the ability of peripheral blood mononuclear cells proliferate in response to mitogens after a long distance intense exercise, as well as plasma glutamine concentration. The amino acids also modify the pattern of cytokine production leading to a diversion of the immune response toward a Th1 type of immune response.

Type I IFNs Are Required to Promote Central Nervous System Immune Surveillance through the Recruitment of Inflammatory Monocytes upon Systemic Inflammation

PubMed Central

Peralta Ramos, Javier María; Bussi, Claudio; Gaviglio, Emilia Andrea; Arroyo, Daniela Soledad; Baez, Natalia Soledad; Rodriguez-Galan, Maria Cecilia; Iribarren, Pablo

2017-01-01

Brain-resident microglia and peripheral migratory leukocytes play essential roles in shaping the immune response in the central nervous system. These cells activate and migrate in response to chemokines produced during active immune responses and may contribute to the progression of neuroinflammation. Herein, we addressed the participation of type I–II interferons in the response displayed by microglia and inflammatory monocytes to comprehend the contribution of these cytokines in the establishment and development of a neuroinflammatory process. Following systemic lipopolysaccharide (LPS) challenge, we found glial reactivity and an active recruitment of CD45hi leukocytes close to CD31+ vascular endothelial cells in circumventricular organs. Isolated CD11b+ CD45hi Ly6Chi Ly6G−-primed inflammatory monocytes were able to induce T cell proliferation, unlike CD11b+ CD45lo microglia. Moreover, ex vivo re-stimulation with LPS exhibited an enhancement of T cell proliferative response promoted by inflammatory monocytes. These myeloid cells also proved to be recruited in a type I interferon-dependent fashion as opposed to neutrophils, unveiling a role of these cytokines in their trafficking. Together, our results compares the phenotypic and functional features between tissue-resident vs peripheral recruited cells in an inflamed microenvironment, identifying inflammatory monocytes as key sentinels in a LPS-induced murine model of neuroinflammation. PMID:29255461

Different Regions of the Malaria Merozoite Surface Protein 1 of Plasmodium chabaudi Elicit Distinct T-Cell and Antibody Isotype Responses

PubMed Central

Quin, Stuart J.; Langhorne, Jean

2001-01-01

In this study we have investigated the antibody and CD4 T-cell responses to the well-characterized malaria vaccine candidate MSP-1 during the course of a primary Plasmodium chabaudi chabaudi (AS) infection. Specific antibody responses can be detected within the first week of infection, and CD4 T cells can be detected after 3 weeks of infection. The magnitude of the CD4 T-cell response elicited during a primary infection depended upon the region of MSP-1. In general, the highest precursor frequencies were obtained when a recombinant MSP-1 fragment corresponding to amino acids 900 to 1507 was used as the antigen in vitro. By contrast, proliferative and cytokine responses against amino acids 1508 to 1766 containing the C-terminal 21-kDa region of the molecule were low. The characteristic interleukin 4 (IL-4) switch that occurs in the CD4 T-cell population after an acute blood stage P. c. chabaudi infection was only consistently observed in the response to the amino acid 900 to 1507 MSP1 fragment. A lower frequency of IL-4-producing cells was seen in response to other regions. Although the magnitudes of the immunoglobulin G antibody responses to the different regions of MSP-1 were similar, the isotype composition of each response was distinct, and there was no obvious relationship with the type of T helper cells generated. Interestingly, a relatively high antibody response to the C-terminal region of MSP-1 was observed, suggesting that T-cell epitopes outside of this region may provide the necessary cognate help for specific antibody production. PMID:11254580

Inulin based glutathione-responsive delivery system for colon cancer treatment.

PubMed

Wang, Dongdong; Sun, Feifei; Lu, Chunbo; Chen, Peng; Wang, Zhaojie; Qiu, Yuanhao; Mu, Haibo; Miao, Zehong; Duan, Jinyou

2018-05-01

Colorectal cancer is one of the most common types of tumor in the world. Here we developed a lipoic acid esterified polysaccharide (inulin) delivery system for tanshinone IIA to treat colorectal cancer in vitro. The release of tanshinone IIA in the system was highly responsive to glutathione, which is commonly abundant in cancer cells. In addition, this drug delivery system was proliferative to Bifidobacterium longum, the common inhabitant of human intestine. Thus, this strategy might be useful to improve colon cancer therapy efficacy of anticancer drugs and meanwhile promote the growth of beneficial commensal flora in the gut. Copyright © 2018 Elsevier B.V. All rights reserved.

Immunization of Cattle by Infection with Cowdria ruminantium Elicits T Lymphocytes That Recognize Autologous, Infected Endothelial Cells and Monocytes†

PubMed Central

Mwangi, Duncan M.; Mahan, Suman M.; Nyanjui, John K.; Taracha, Evans L. N.; McKeever, Declan J.

1998-01-01

Peripheral blood mononuclear cells (PBMC) from immune cattle proliferate in the presence of autologous Cowdria ruminantium-infected endothelial cells and monocytes. Endothelial cells required treatment with T-cell growth factors to induce class II major histocompatibility complex expression prior to infection and use as stimulators. Proliferative responses to both infected autologous endothelial cells and monocytes were characterized by expansion of a mixture of CD4+, CD8+, and γδ T cells. However, γδ T cells dominated following several restimulations. Reverse transcription-PCR analysis of cytokine expression by C. ruminantium-specific T-cell lines and immune PBMC revealed weak interleukin-2 (IL-2), IL-4, and gamma interferon (IFN-γ) transcripts at 3 to 24 h after stimulation. Strong expression of IFN-γ, tumor necrosis factor alpha (TNF-α), TNF-β, and IL-2 receptor α-chain mRNA was detected in T-cell lines 48 h after antigen stimulation. Supernatants from these T-cell cultures contained IFN-γ protein. Our findings suggest that in immune cattle a C. ruminantium-specific T-cell response is induced and that infected endothelial cells and monocytes may present C. ruminantium antigens to specific T lymphocytes in vivo during infection and thereby play a role in induction of protective immune responses to the pathogen. PMID:9573061

Early Increases in Superantigen-Specific Foxp3+ Regulatory T Cells during Mouse Mammary Tumor Virus Infection▿ †

PubMed Central

Cabrera, Gabriel; Burzyn, Dalia; Mundiñano, Juliana; Courreges, M. Cecilia; Camicia, Gabriela; Lorenzo, Daniela; Costa, Héctor; Ross, Susan R.; Nepomnaschy, Irene; Piazzon, Isabel

2008-01-01

Mouse mammary tumor virus (MMTV) is a milk-borne betaretrovirus that has developed strategies to exploit and subvert the host immune system. Here, we show in a natural model of MMTV infection that the virus causes early and progressive increases in superantigen (SAg)-specific Foxp3+ regulatory T cells (Treg) in Peyer's patches (PP). These increases were shown to be dependent on the presence of dendritic cells. CD4+ CD25+ T cells from the PP of infected mice preferentially suppress the proliferative response of T cells to SAg-expressing antigen-presenting cells ex vivo. We investigated the influence of the depletion of CD25+ cells at different stages of the infection. When CD25+ cells were depleted before MMTV infection, an increase in the number of PP SAg-cognate Foxp3− T cells was found at day 6 of infection. Since the SAg response is associated with viral amplification, the possibility exists that Treg cells attenuate the increase in viral load at the beginning of the infection. In contrast, depletion of CD25+ cells once the initial SAg response has developed caused a lower viral load, suggesting that at later stages Treg cells may favor viral persistence. Thus, our results indicated that Treg cells play an important and complex role during MMTV infection. PMID:18495774

Innate signals overcome acquired TCR signaling pathway regulation and govern the fate of human CD161(hi) CD8α⁺ semi-invariant T cells.

PubMed

Turtle, Cameron J; Delrow, Jeff; Joslyn, Rochelle C; Swanson, Hillary M; Basom, Ryan; Tabellini, Laura; Delaney, Colleen; Heimfeld, Shelly; Hansen, John A; Riddell, Stanley R

2011-09-08

Type 17 programmed CD161(hi)CD8α(+) T cells contribute to mucosal immunity to bacteria and yeast. In early life, microbial colonization induces proliferation of CD161(hi) cells that is dependent on their expression of a semi-invariant Vα7.2(+) TCR. Although prevalent in adults, CD161(hi)CD8α(+) cells exhibit weak proliferative and cytokine responses to TCR ligation. The mechanisms responsible for the dichotomous response of neonatal and adult CD161(hi) cells, and the signals that enable their effector function, have not been established. We describe acquired regulation of TCR signaling in adult memory CD161(hi)CD8α(+) T cells that is absent in cord CD161(hi) cells and adult CD161(lo) cells. Regulated TCR signaling in CD161(hi) cells was due to profound alterations in TCR signaling pathway gene expression and could be overcome by costimulation through CD28 or innate cytokine receptors, which dictated the fate of their progeny. Costimulation with IL-1β during TCR ligation markedly increased proinflammatory IL-17 production, while IL-12-induced Tc1-like function and restored the response to TCR ligation without costimulation. CD161(hi) cells from umbilical cord blood and granulocyte colony stimulating factor-mobilized leukaphereses differed in frequency and function, suggesting future evaluation of the contribution of CD161(hi) cells in hematopoietic stem cell grafts to transplant outcomes is warranted.

Characterisation of cell cycle arrest and terminal differentiation in a maximally proliferative human epithelial tissue: Lessons from the human hair follicle matrix.

PubMed

Purba, Talveen S; Brunken, Lars; Peake, Michael; Shahmalak, Asim; Chaves, Asuncion; Poblet, Enrique; Ceballos, Laura; Gandarillas, Alberto; Paus, Ralf

2017-09-01

Human hair follicle (HF) growth and hair shaft formation require terminal differentiation-associated cell cycle arrest of highly proliferative matrix keratinocytes. However, the regulation of this complex event remains unknown. CIP/KIP family member proteins (p21 CIP1 , p27 KIP1 and p57 KIP2 ) regulate cell cycle progression/arrest, endoreplication, differentiation and apoptosis. Since they have not yet been adequately characterized in the human HF, we asked whether and where CIP/KIP proteins localise in the human hair matrix and pre-cortex in relation to cell cycle activity and HF-specific epithelial cell differentiation that is marked by keratin 85 (K85) protein expression. K85 expression coincided with loss or reduction in cell cycle activity markers, including in situ DNA synthesis (EdU incorporation), Ki-67, phospho-histone H3 and cyclins A and B1, affirming a post-mitotic state of pre-cortical HF keratinocytes. Expression of CIP/KIP proteins was found abundantly within the proliferative hair matrix, concomitant with a role in cell cycle checkpoint control. p21 CIP1 , p27 KIP1 and cyclin E persisted within post-mitotic keratinocytes of the pre-cortex, whereas p57 KIP2 protein decreased but became nuclear. These data imply a supportive role for CIP/KIP proteins in maintaining proliferative arrest, differentiation and anti-apoptotic pathways, promoting continuous hair bulb growth and hair shaft formation in anagen VI. Moreover, post-mitotic hair matrix regions contained cells with enlarged nuclei, and DNA in situ hybridisation showed cells that were >2N in the pre-cortex. This suggests that CIP/KIP proteins might counterbalance cyclin E to control further rounds of DNA replication in a cell population that has a propensity to become tetraploid. These data shed new light on the in situ-biography of human hair matrix keratinocytes on their path of active cell cycling, arrest and terminal differentiation, and showcase the human HF as an excellent, clinically relevant model system for cell cycle physiology research of human epithelial cells within their natural tissue habitat. Crown Copyright © 2017. Published by Elsevier GmbH. All rights reserved.

Size distribution of retrovirally marked lineages matches prediction from population measurements of cell cycle behavior

NASA Technical Reports Server (NTRS)

Cai, Li; Hayes, Nancy L.; Takahashi, Takao; Caviness, Verne S Jr; Nowakowski, Richard S.

2002-01-01

Mechanisms that regulate neuron production in the developing mouse neocortex were examined by using a retroviral lineage marking method to determine the sizes of the lineages remaining in the proliferating population of the ventricular zone during the period of neuron production. The distribution of clade sizes obtained experimentally in four different injection-survival paradigms (E11-E13, E11-E14, E11-E15, and E12-E15) from a total of over 500 labeled lineages was compared with that obtained from three models in which the average behavior of the proliferating population [i.e., the proportion of cells remaining in the proliferative population (P) vs. that exiting the proliferative population (Q)] was quantitatively related to lineage size distribution. In model 1, different proportions of asymmetric, symmetric terminal, and symmetric nonterminal cell divisions coexisted during the entire developmental period. In model 2, the developmental period was divided into two epochs: During the first, asymmetric and symmetric nonterminal cell divisions occurred, but, during the second, asymmetric and symmetric terminal cell divisions occurred. In model 3, the shifts in P and Q are accounted for by changes in the proportions of the two types of symmetric cell divisions without the inclusion of any asymmetric cell divisions. The results obtained from the retroviral experiments were well accounted for by model 1 but not by model 2 or 3. These findings demonstrate that: 1) asymmetric and both types of symmetric cell divisions coexist during the entire period of neurogenesis in the mouse, 2) neuron production is regulated in the proliferative population by the independent decisions of the two daughter cells to reenter S phase, and 3) neurons are produced by both asymmetric and symmetric terminal cell divisions. In addition, the findings mean that cell death and/or tangential movements of cells in the proliferative population occur at only a low rate and that there are no proliferating lineages "reserved" to make particular laminae or cell types. Copyright 2002 Wiley-Liss, Inc.

Toxicological and pharmacological assessment of AGEN1884, a novel human IgG1 anti-CTLA-4 antibody

PubMed Central

Gonzalez, Ana; Manrique, Mariana; Chand, Dhan; Savitsky, David; Morin, Benjamin; Breous-Nystrom, Ekaterina; Dupont, Christopher; Ward, Rebecca A.; Mundt, Cornelia; Duckless, Benjamin; Tang, Hao; Findeis, Mark A.; Schuster, Andrea; Waight, Jeremy D.; Underwood, Dennis; Clarke, Christopher; Ritter, Gerd; Merghoub, Taha; Schaer, David; Wolchok, Jedd D.; van Dijk, Marc; Buell, Jennifer S.; Cuillerot, Jean-Marie; Stein, Robert; Drouin, Elise E.

2018-01-01

CTLA-4 and CD28 exemplify a co-inhibitory and co-stimulatory signaling axis that dynamically sculpts the interaction of antigen-specific T cells with antigen-presenting cells. Anti-CTLA-4 antibodies enhance tumor-specific immunity through a variety of mechanisms including: blockade of CD80 or CD86 binding to CTLA-4, repressing regulatory T cell function and selective elimination of intratumoral regulatory T cells via an Fcγ receptor-dependent mechanism. AGEN1884 is a novel IgG1 antibody targeting CTLA-4. It potently enhanced antigen-specific T cell responsiveness that could be potentiated in combination with other immunomodulatory antibodies. AGEN1884 was well-tolerated in non-human primates and enhanced vaccine-mediated antigen-specific immunity. AGEN1884 combined effectively with PD-1 blockade to elicit a T cell proliferative response in the periphery. Interestingly, an IgG2 variant of AGEN1884 revealed distinct functional differences that may have implications for optimal dosing regimens in patients. Taken together, the pharmacological properties of AGEN1884 support its clinical investigation as a single therapeutic and combination agent. PMID:29617360

The Effects of Orbital Spaceflight on Human Osteoblastic Cell Physiology and Gene Expression

NASA Technical Reports Server (NTRS)

Turner, R. T.

1999-01-01

The purpose of the proposed study is to establish whether changes in gravitational loading have a direct effect on osteoblasts to regulate TGF-6 expression. The effects of spaceflight and reloading on TGF-B MRNA and peptide levels will be studied in a newly developed line of immortalized human fetal osteoblasts (HFOB) transfected with an SV-40 temperature dependent mutant to generate proliferating, undifferentiated hFOB cells at 33-34 C and a non-proliferating, differentiated HFOB cells at 37-39'C. Unlike previous cell culture models, HFOB cells have unlimited proliferative capacity yet can be precisely regulated to differentiate into mature cells which express mature osteoblast function. If isolated osteoblasts respond to changes in mechanical loading in a manner similar to their response in animals, the cell system could provide a powerful model to investigate the signal transduction pathway for gravitational loading.

Functional and molecular alterations in T Cells induced by CCL5.

PubMed

Cridge, T J; Horowitz, K M; Marinucci, M N; Rose, K M; Wells, M; Werner, M T; Kurt, Robert A

2006-01-01

To delineate whether, and the extent to which, CCL5 could impact T cell function we examined cytokine production and proliferative ability following CCL5 treatment in vitro. We report a decreased ability of splenic T cells to produce IFN-? and TNF-a as well as proliferate in response to crosslinking with antibody to CD3 after 72, but not 24 hours of CCL5 exposure. To identify a mechanism by which CCL5 modulated T cell function, we examined T cell receptor translocation and lipid raft clustering. After exposure to CCL5, T cells were less efficient at translocating the TCR and clustering lipid rafts. Since TCR translocation and lipid raft clustering are required for creation of an immunological synapse, these data suggest that extended exposure to CCL5 may impact T cell effector function by modulating the ability to create a functional immunological synapse.

Long-Term Hematopoietic Engraftment of Congenic Amniotic Fluid Stem Cells After in Utero Intraperitoneal Transplantation to Immune Competent Mice

PubMed Central

Shangaris, Panicos; Loukogeorgakis, Stavros P.; Blundell, Michael P.; Petra, Eleni; Shaw, Steven W.; Ramachandra, Durrgah L.; Maghsoudlou, Panagiotis; Urbani, Luca; Thrasher, Adrian J.

2018-01-01

Clinical success of in utero transplantation (IUT) using allogeneic hematopoietic stem cells (HSCs) has been limited to fetuses that lack an immune response to allogeneic cells due to severe immunological defects, and where transplanted genetically normal cells have a proliferative or survival advantage. Amniotic fluid (AF) is an autologous source of stem cells with hematopoietic potential that could be used to treat congenital blood disorders. We compared the ability of congenic and allogeneic mouse AF stem cells (AFSC) to engraft the hematopoietic system of time-mated C57BL/6J mice (E13.5). At 4 and 16 weeks of age, multilineage donor engraftment was higher in congenic versus allogeneic animals. In vitro mixed lymphocyte reaction confirmed an immune response in the allogeneic group with higher CD4 and CD8 cell counts and increased proliferation of stimulated lymphocytes. IUT with congenic cells resulted in 100% of donor animals having chimerism of around 8% and successful hematopoietic long-term engraftment in immune-competent mice when compared with IUT with allogeneic cells. AFSCs may be useful for autologous cell/gene therapy approaches in fetuses diagnosed with congenital hematopoietic disorders. PMID:29482456

Ethanol extracts from the branch of Taxillus yadoriki parasitic to Neolitsea sericea induces cyclin D1 proteasomal degradation through cyclin D1 nuclear export.

PubMed

Park, Su Bin; Park, Gwang Hun; Kim, Ha Na; Song, Hun Min; Son, Ho-Jun; Park, Ji Ae; Kim, Hyun-Seok; Jeong, Jin Boo

2018-06-20

Although the inhibitory effect of mistletoe on cancer cell growth has been reported, the underlying mechanisms to explain its anti-proliferative activity are not fully studied. Thus, we elucidated the potential molecular mechanism of the branch from Taxillus yadoriki (TY) parasitic to Neolitsea sericea (NS) (TY-NS-B) for the anti-proliferative effect. Anti-cell proliferative effect was evaluated by MTT assay. The change of cyclin D1 protein or mRNA level was evaluated by Western blot and RT-RCR, respectively. In comparison of anti-proliferative effect of TY from the host trees such as Cryptomeria japonica (CJ), Neolitsea sericea (NS), Prunus serrulata (PS), Cinnamomum camphora (CC) and Quercus acutissima (QA), TY-NS showed higher anti-cell proliferative effect than TY-CJ, TY-PS, TY-CC or TY-QA. In addition, the anti-proliferative effect of branch from TY from all host trees was better than leaves. Thus, we selected the branch from Taxillus yadoriki parasitic to Neolitsea sericea (TY-NS-B) for the further study. TY-NS-B inhibited the cell proliferation in the various cancer cells and downregulated cyclin D1 protein level. MG132 treatment attenuated cyclin D1 downregulation of cyclin D1 protein level by TY-NS-B. In addition, TY-NS-B increased threonine-286 (T286) phosphorylation of cyclin D1, and the mutation of T286 to alanine (T286A) blocked cyclin D1 proteasomal degradation by TY-NS-B. But the upstream factors related to cyclin D1 degradation such as ERK1/2, p38, JNK, GSK3β, PI3K, IκK or ROS did not affect cyclin D1 degradation by TY-NS-B. However, LMB treatment was observed to inhibit cyclin D1 degradation by TY-NS-B, and T286A blocked cyclin D1 degradation through suppressing cyclin D1 redistribution from nucleus to cytoplasm by TY-NS-B. In addition, TY-NS-B activated CRM1 expression. Our results suggest that TY-NS-B may suppress cell proliferation by downregulating cyclin D1 protein level through proteasomal degradation via T286 phosphorylation-dependent cyclin D1 nuclear export. These findings will provide the evidence that TY-NS-B has potential to be a candidate for the development of chemoprevention or therapeutic agents for human cancer.

WITHAFERIN A INDUCES APOPTOSIS IN RAT C6 GLIOMA CELLS THROUGH REGULATING NF-KB NUCLEAR TRANSLOCATION AND ACTIVATION OF CASPASE CASCADE.

PubMed

Hou, Wei-Chen; Miao, Xiao-Hui; Ma, Lian-Jun; Bai, Xiao-Xue; Liu, Qun; Song, Lei

2017-01-01

The demand for the chemopreventive drug from the plant source is increasing in recent times, owing to its various biological activities without any adverse effect. The intention of this current study was to examine the anti-glioma effect of Withaferin A (WFA) on C6 glioma cell line model. C6 glioma cells were administrated with different concentration of WFA (50, 100, 200 and 500 μg/mL) and DMSO (control) group to examine its anti-proliferative, anti-inflammatory and pro-apoptotic activities. Treatment with WFA showed a significant decline in the glioma cell count in a dose-dependent manner and thus proving its anti-proliferative effect. Similarly, inflammatory markers were also substantially lowered upon treatment with different concentration of WFA. However, DNA fragmentation and apoptotic markers like Caspase-3 and 9 were concomitantly enhanced after co-cultured with different concentration of WFA and thus exhibiting its cytotoxicity efficacy. Furthermore, the protein expression of Bcl2 and Bax were markedly downregulated and upregulated respectively; upon treatment with WFA on C6 glioma cells. The outcome of this study evidently demonstrates that C6 glioma cells co-cultured with increased concentration of WFA, showed an anti-proliferative, anti-inflammatory and pro-apoptotic effect in a dose-dependent fashion.

Interkinetic and migratory behavior of a cohort of neocortical neurons arising in the early embryonic murine cerebral wall

NASA Technical Reports Server (NTRS)

Takahashi, T.; Nowakowski, R. S.; Caviness, V. S. Jr

1996-01-01

Neocortical neuronogenesis occurs in the pseudostratified ventricular epithelium (PVE) where nuclei of proliferative cells undergo interkinetic nuclear movement. A fraction of daughter cells exits the cell cycle as neurons (the quiescent, or Q, fraction), whereas a complementary fraction remains in the cell cycle (the proliferative, or P, fraction). By means of sequential thymidine and bromodeoxyuridine injections in mouse on embryonic day 14, we have monitored the proliferative and post-mitotic migratory behaviors of 1 and 2 hr cohorts of PVE cells defined by the injection protocols. Soon after mitosis, the Q fraction partitions into a rapidly exiting (up to 50 microns/hr) subpopulation (Qr) and a more slowly exiting (6 microns/hr) subpopulation (Qs). Qr and Qs are separated as two distributions on exit from the ventricular zone with an interpeak distance of approximately 40 microns. Cells in Qr and Qs migrate through the intermediate zone with no significant change in the interpeak distance, suggesting that they migrate at approximately the same velocities. The rate of migration increases with ascent through the intermediate zone (average 2-6.4 microns/hr) slowing only transiently on entry into the developing cortex. Within the cortex, Qr and Qs merge to form a single distribution most concentrated over layer V.

Mesenchymal stem cells induce epithelial proliferation within the inflamed stomach.

PubMed

Donnelly, Jessica M; Engevik, Amy; Feng, Rui; Xiao, Chang; Boivin, Gregory P; Li, Jing; Houghton, JeanMarie; Zavros, Yana

2014-06-15

Bone marrow-derived mesenchymal stem cells (MSCs) sustain cancer cells by creating a microenvironment favorable for tumor growth. In particular, MSCs have been implicated in gastric cancer development. There is extensive evidence suggesting that Hedgehog signaling regulates tumor growth. However, very little is known regarding the precise roles of Hedgehog signaling and MSCs in tumor development within the stomach. The current study tests that hypothesis that Sonic Hedgehog (Shh), secreted from MSCs, provides a proliferative stimulus for the gastric epithelium in the presence of inflammation. Red fluorescent protein-expressing MSCs transformed in vitro (stMSCs) were transduced with lentiviral constructs containing a vector control (stMSC(vect)) or short hairpin RNA (shRNA) targeting the Shh gene (stMSC(ShhKO)). Gastric submucosal transplantation of wild-type MSCs (wtMSCs), wild-type MSCs overexpressing Shh (wtMSC(Shh)), stMSC(vect), or stMSC(ShhKO) cells in C57BL/6 control (BL/6) or gastrin-deficient (GKO) mice was performed and mice analyzed 30 and 60 days posttransplantation. Compared with BL/6 mice transplanted with wtMSC(Shh) and stMSC(vect) cells, inflamed GKO mice developed aggressive gastric tumors. Tumor development was not observed in mouse stomachs transplanted with wtMSC or stMSC(ShhKO) cells. Compared with stMSC(ShhKO)-transplanted mice, within the inflamed GKO mouse stomach, Shh-expressing stMSC(vect)- and wtMSC(Shh)-induced proliferation of CD44-positive cells. CD44-positive cells clustered in gland-like structures within the tumor stroma and were positive for Patched (Ptch) expression. We conclude that Shh, secreted from MSCs, provides a proliferative stimulus for the gastric epithelium that is associated with tumor development, a response that is sustained by chronic inflammation. Copyright © 2014 the American Physiological Society.

Mesenchymal stem cells induce epithelial proliferation within the inflamed stomach

PubMed Central

Donnelly, Jessica M.; Engevik, Amy; Feng, Rui; Xiao, Chang; Boivin, Gregory P.; Li, Jing; Houghton, JeanMarie

2014-01-01

Bone marrow-derived mesenchymal stem cells (MSCs) sustain cancer cells by creating a microenvironment favorable for tumor growth. In particular, MSCs have been implicated in gastric cancer development. There is extensive evidence suggesting that Hedgehog signaling regulates tumor growth. However, very little is known regarding the precise roles of Hedgehog signaling and MSCs in tumor development within the stomach. The current study tests that hypothesis that Sonic Hedgehog (Shh), secreted from MSCs, provides a proliferative stimulus for the gastric epithelium in the presence of inflammation. Red fluorescent protein-expressing MSCs transformed in vitro (stMSCs) were transduced with lentiviral constructs containing a vector control (stMSCvect) or short hairpin RNA (shRNA) targeting the Shh gene (stMSCShhKO). Gastric submucosal transplantation of wild-type MSCs (wtMSCs), wild-type MSCs overexpressing Shh (wtMSCShh), stMSCvect, or stMSCShhKO cells in C57BL/6 control (BL/6) or gastrin-deficient (GKO) mice was performed and mice analyzed 30 and 60 days posttransplantation. Compared with BL/6 mice transplanted with wtMSCShh and stMSCvect cells, inflamed GKO mice developed aggressive gastric tumors. Tumor development was not observed in mouse stomachs transplanted with wtMSC or stMSCShhKO cells. Compared with stMSCShhKO-transplanted mice, within the inflamed GKO mouse stomach, Shh-expressing stMSCvect- and wtMSCShh-induced proliferation of CD44-positive cells. CD44-positive cells clustered in gland-like structures within the tumor stroma and were positive for Patched (Ptch) expression. We conclude that Shh, secreted from MSCs, provides a proliferative stimulus for the gastric epithelium that is associated with tumor development, a response that is sustained by chronic inflammation. PMID:24789207

Memory T cell proliferative responses and IFN-γ productivity sustain long-lasting efficacy of a Cap-based PCV2 vaccine upon PCV2 natural infection and associated disease.

PubMed

Ferrari, Luca; Borghetti, Paolo; De Angelis, Elena; Martelli, Paolo

2014-04-16

Porcine circovirus type 2 (PCV2) vaccination represents an important measure to cope with PCV2 infection; however, data regarding the modulation of the immune cell compartment are still limited, especially under field conditions. This study is aimed at investigating the features of the cellular immune response in conventional piglets induced by vaccination using a capsid (Cap) protein-based PCV2 vaccine compared to unvaccinated animals when exposed to PCV2 natural infection. Immune reactivity was evaluated by quantifying peripheral cell subsets involved in the anti-viral response and characterizing the interferon-gamma (IFN-γ) secreting cell (SC) responsiveness both in vivo and upon in vitro whole PCV2 recall. The vaccination triggered an early and intense IFN-γ secreting cell response and induced the activation of peripheral lymphocytes. The early increase of IFN-γ SC frequencies resulted in a remarkable and transient tendency to increased IFN-γ productivity in vaccinated pigs. In vaccinated animals, soon before the onset of infection occurred 15-16 weeks post-vaccination, the recalled PCV2-specific immune response was characterized by moderate PCV2-specific IFN-γ secreting cell frequencies and augmented productivity together with reactive CD4+CD8+ memory T cells. Conversely, upon infection, unvaccinated animals showed very high frequencies of IFN-γ secreting cells and a tendency to lower productivity, which paralleled with effector CD4-CD8+ cytotoxic cell responsiveness. The study shows that PCV2 vaccination induces a long-lasting immunity sustained by memory T cells and IFN-γ secreting cells that potentially played a role in preventing the onset of infection; the extent and duration of this reactivity can be an important feature for evaluating the protective immunity induced by vaccination.

The pharmacology of second-generation chimeric antigen receptors.

PubMed

van der Stegen, Sjoukje J C; Hamieh, Mohamad; Sadelain, Michel

2015-07-01

Second-generation chimeric antigen receptors (CARs) retarget and reprogramme T cells to augment their antitumour efficacy. The combined activating and co-stimulatory domains incorporated in these CARs critically determine the function, differentiation, metabolism and persistence of engineered T cells. CD19-targeted CARs that incorporate CD28 or 4-1BB signalling domains are the best known to date. Both have shown remarkable complete remission rates in patients with refractory B cell malignancies. Recent data indicate that CD28-based CARs direct a brisk proliferative response and boost effector functions, whereas 4-1BB-based CARs induce a more progressive T cell accumulation that may compensate for less immediate potency. These distinct kinetic features can be exploited to further develop CAR-based T cell therapies for a variety of cancers. A new field of immunopharmacology is emerging.

Treatment of both native and deamidated gluten peptides with an endo-peptidase from Aspergillus niger prevents stimulation of gut-derived gluten-reactive T cells from either children or adults with celiac disease.

PubMed

Toft-Hansen, Henrik; Rasmussen, Karina S; Staal, Anne; Roggen, Erwin L; Sollid, Ludvig M; Lillevang, Søren T; Barington, Torben; Husby, Steffen

2014-08-01

Celiac disease (CD) is characterized by an inappropriate immunological reaction against gluten driven by gluten-specific CD4+ T cells. We screened 25 proteases and tested 10 for their potential to degrade gluten in vitro. Five proteases were further tested for their ability to prevent the proliferative response by a gluten-specific CD4+ T cell clone and seven gluten-reactive T cell lines to protease-digested gluten peptides. A proline-specific endo-peptidase from Aspergillus niger (AnP2) was particularly efficient at diminishing proliferation after stimulation with cleaved antigen, and could completely block the response against both native and deamidated gluten peptides. We found that AnP2 was efficient down to a 1:64 protease:substrate ratio (w:w). When AnP2 was tested in assays using seven gluten-reactive T cell lines from individual CD patients (three adults and four children), the response to gluten was diminished in all cases. Our study indicates a therapeutic benefit of AnP2 to CD patients. Copyright © 2014 Elsevier Inc. All rights reserved.

Antigen presenting cells (APCs) from thermally injured and/or septic rats modulate CD4+ T cell responses of naive rat.

PubMed

Fazal, Nadeem; Raziuddin, Syed; Khan, Mehdi; Al-Ghoul, Walid M

2006-01-01

Regulation of immune response is marked by complex interactions among the cells that recognize and present antigens. Antigen presenting cells (APCs), the antigen presenting cell component of the innate immune response plays an important role in effector CD4+ T cell response. Thermal injury and/or superimposed sepsis in rats' leads to suppressed CD4+ T cell functions. We investigated modulations of CD4+ T cell function by APCs (purified non-T cells) from thermally injured and/or septic rats. Rats were subjected to 30% total body surface area scald burn or exposed to 37 degrees C water (Sham burn) and sepsis was induced by cecal-ligation and puncture (CLP) method. At day 3 post-injury animals were sacrificed and CD4+ T cells and APCs from mesenteric lymph nodes (MLN) were obtained using magnetic microbead isolation procedure. APCs from injured rats were co-cultured with sham rat MLN CD4+ T cells and proliferative responses (thymidine incorporation), phenotypic changes (Flow cytometry), IL-2 production (ELISA) and CTLA-4 mRNA (RT-PCR) were determined in naive rat CD4+ T cells. The data indicate that APCs from thermally injured and/or septic rats when co-cultured with CD4+ T cells suppressed CD4+ T cell effector functions. This lack of CD4+ T cell activation was accompanied with altered co-stimulatory molecules, i.e., CD28 and/or CTLA-4 (CD152). In conclusion, our studies indicated that defective APCs from thermally injured and/or septic rats modulate CD4+ T cell functions via changes in co-stimulatory molecules expressed on naive CD4+ T cells. This altered APC: CD4+ T cell interaction leads to suppressed CD4+ T cell activation of healthy animals.

Triple selectin knockout (ELP-/-) mice fail to develop OVA-induced acute asthma phenotype

PubMed Central

2011-01-01

Objective The recruitment of leukocytes from circulation to sites of inflammation requires several families of adhesion molecules among which are selectins expressed on a variety of cells. In addition, they have also been shown to play key roles in the activation of cells in inflammation. Methods To explore the collective role of E-, L-, and P- selectins in OVA-induced Th2 mediated response in acute asthma pathophysiology, ELP-/- mice were used and compared with age-matched wildtype (WT). Results Asthma phenotype was assessed by measuring pulmonary function, inflammation and OVA-specific serum IgE, which were completely abrogated in ELP-/- mice. Adoptive transfer of sensitized L selectin+CD4+ T cells into naïve ELP-/- mice which post-OVA challenge, developed asthma, suggesting that L-selectin may be critically involved in the onset of Th2 response in asthma. Tissue resident ELP-deficient cells were otherwise functionally competent as proved by normal proliferative response. Conclusions: Comparative studies between ELP-/- and WT mice uncovered functional roles of these three integrins in inflammatory response in allergic asthma. All three selectins seem to impede inflammatory migration while only L-selectin also possibly regulates activation of specific T cell subsets in lung and airways. PMID:21835035

Gastrin and the growth of the gastrointestinal tract.

PubMed Central

Ekundayo, A A; Lee, C Y; Goodlad, R A

1995-01-01

While the proliferative effects of gastrin in the gastric fundus are well established, there is a considerable degree of confusion regarding the role of gastrin on the growth of the small intestine and colon. The hypothesis that gastrin is trophic throughout the gut was tested by giving three doses of pentagastrin and one of gastrin 17 to rats maintained by total parenteral nutrition (TPN). The rats were fed intravenously for one week, with the various peptides added to the TPN diet. The number of vincristine arrested metaphases per gland or crypt was then scored to determine the proliferative state. Both gastrin 17 and pentagastrin were found to be trophic in the gastric fundus, but not to the gastric antrum. A proliferative response was also seen in the duodenum, but with little evidence of a dose response element. No effect on small bowel weight was seen, and no proliferative effect was noted in the mid small bowel, thus the duodenal effect could be attributed to a local action of increased acid output on the duodenum, not a general role throughout the small intestine. No proliferative effects of pentagastrin or gastrin were seen in the colon. It is therefore concluded that the trophic role of gastrin is restricted to the gastric fundus and the proximal duodenum. PMID:7883218

BCR ligation induced by IgM stimulation results in gene expression and functional changes only in IgV H unmutated chronic lymphocytic leukemia (CLL) cells.

PubMed

Guarini, Anna; Chiaretti, Sabina; Tavolaro, Simona; Maggio, Roberta; Peragine, Nadia; Citarella, Franca; Ricciardi, Maria Rosaria; Santangelo, Simona; Marinelli, Marilisa; De Propris, Maria Stefania; Messina, Monica; Mauro, Francesca Romana; Del Giudice, Ilaria; Foà, Robert

2008-08-01

Chronic lymphocytic leukemia (CLL) patients exhibit a variable clinical course. To investigate the association between clinicobiologic features and responsiveness of CLL cells to anti-IgM stimulation, we evaluated gene expression changes and modifications in cell-cycle distribution, proliferation, and apoptosis of IgV(H) mutated (M) and unmutated (UM) samples upon BCR cross-linking. Unsupervised analysis highlighted a different response profile to BCR stimulation between UM and M samples. Supervised analysis identified several genes modulated exclusively in the UM cases upon BCR cross-linking. Functional gene groups, including signal transduction, transcription, cell-cycle regulation, and cytoskeleton organization, were up-regulated upon stimulation in UM cases. Cell-cycle and proliferation analyses confirmed that IgM cross-linking induced a significant progression into the G(1) phase and a moderate increase of proliferative activity exclusively in UM patients. Moreover, we observed only a small reduction in the percentage of subG(0/1) cells, without changes in apoptosis, in UM cases; contrariwise, a significant increase of apoptotic levels was observed in stimulated cells from M cases. These results document that a differential genotypic and functional response to BCR ligation between IgV(H) M and UM cases is operational in CLL, indicating that response to antigenic stimulation plays a pivotal role in disease progression.

Characterization of outer membrane vesicles from a neonatal meningitic strain of Cronobacter sakazakii.

PubMed

Alzahrani, Hayat; Winter, Jody; Boocock, David; De Girolamo, Luigi; Forsythe, Stephen J

2015-06-01

Cronobacter sakazakii is associated with severe and often fatal cases of infant meningitis and necrotizing enterocolitis. The form of meningitis differs from that due to Neisseria meningitidis and Streptococcus spp., in that it is highly invasive and destructive towards human brain cells. However, there is relatively little understanding of the cytopathogenic interaction of C. sakazakii with host cells which results in stimulation of an inflammatory immune response. The production of Cronobacter outer membrane vesicles (OMV) and their potential pathogenic functions have not yet been elucidated. This study is the first to show that C. sakazakii produce OMV, which may play a role in the activation of cytopathogenic and host cell responses on human intestinal epithelial cells. Cronobacter sakazakii strain 767 was used which had been isolated from a fatal outbreak of neonatal meningitis and necrotizing enterocolitis. Cronobacter sakazakii OMV were internalized by Caco-2 cells, increased cell proliferation and stimulated the host's innate proinflammatory response without inducing overt toxicity. A total of 18 OMV-associated proteins were identified by mass spectrometry and their potential pathogenicity roles were evaluated. Collectively, these data indicate that C. sakazakii OMV could play a role in pathogenesis by delivering bacterial toxins into host epithelial cells, driving proliferative and proinflammatory responses. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

The effect of burn injury on CD8+ and CD4+ T cells in an irradiation model of homeostatic proliferation.

PubMed

Buchanan, Ian B; Maile, Robert; Frelinger, Jeffrey A; Fair, Jeffrey H; Meyer, Anthony A; Cairns, Bruce A

2006-11-01

Homeostatic proliferation of T cells has recently been shown to be an important mechanism in the host response to infection. However, its role in the T cell response to burn injury is unknown. In this study, we examine the effect of burn injury on CD4+ and CD8+ T cell homeostatic proliferation after irradiation. Wild-type C57BL/6 female mice were irradiated with six grays ionizing radiation and 48 hours later, syngeneic whole splenocytes or purified CD4+ or CD8+ T cells labeled with carboxy-fluorescein diacetate, succinimidyl ester were adoptively transferred. Two days later, mice underwent a 20% burn injury, followed by splenocyte harvest 3 and 10 days after injury. Burn mice demonstrate increased splenic cellularity and CD8+ T cell proliferation after adoptive transfer of either purified CD8+ cells or whole spleen populations compared with unburned (sham) mice. In contrast, CD4+ T cell proliferation after burn injury is unchanged after adoptive transfer of whole spleen cells and drastically decreased after adoptive transfer of a purified CD4+ population compared with sham mice. Ten days after burn injury CD8+ T cells continue to demonstrate greater proliferation than CD4+ T cells. CD8+ T cells are more robust than CD4+ T cells in their proliferative response after burn injury. In addition, CD8+ T cell proliferation appears less reliant on other immune cells than purified CD4+ T cell proliferation. These data reiterate the importance of CD8+ T cells in the initial immune response to burn injury.

Postnatal ablation of osteoblast Smad4 enhances proliferative responses to canonical Wnt signaling through interactions with β-catenin

PubMed Central

Salazar, Valerie S.; Zarkadis, Nicholas; Huang, Lisa; Watkins, Marcus; Kading, Jacqueline; Bonar, Sheri; Norris, Jin; Mbalaviele, Gabriel; Civitelli, Roberto

2013-01-01

Summary Canonical Wnt (cWnt) signaling through β-catenin regulates osteoblast proliferation and differentiation to enhance bone formation. We previously reported that osteogenic action of β-catenin is dependent on BMP signaling. Here, we further examined interactions between cWnt and BMP in bone. In osteoprogenitors stimulated with BMP2, β-catenin localizes to the nucleus, physically interacts with Smad4, and is recruited to DNA-binding transcription complexes containing Smad4, R-Smad1/5 and TCF4. Furthermore, Tcf/Lef-dependent transcription, Ccnd1 expression and proliferation all increase when Smad4, 1 or 5 levels are low, whereas TCF/Lef activities decrease when Smad4 expression is high. The ability of Smad4 to antagonize transcription of Ccnd1 is dependent on DNA-binding activity but Smad4-dependent transcription is not required. In mice, conditional deletion of Smad4 in osterix+ cells increases mitosis of cells on trabecular bone surfaces as well as in primary osteoblast cultures from adult bone marrow and neonatal calvaria. By contrast, ablation of Smad4 delays differentiation and matrix mineralization by primary osteoblasts in response to Wnt3a, indicating that loss of Smad4 perturbs the balance between proliferation and differentiation in osteoprogenitors. We propose that Smad4 and Tcf/Lef transcription complexes compete for β-catenin, thus restraining cWnt-dependent proliferative signals while favoring the matrix synthesizing activity of osteoblasts. PMID:24101723

Cellular stress induced by resazurin leads to autophagy and cell death via production of reactive oxygen species and mitochondrial impairment.

PubMed

Erikstein, Bjarte S; Hagland, Hanne R; Nikolaisen, Julie; Kulawiec, Mariola; Singh, Keshav K; Gjertsen, Bjørn T; Tronstad, Karl J

2010-10-15

Mitochondrial bioenergetics and reactive oxygen species (ROS) often play important roles in cellular stress mechanisms. In this study we investigated how these factors are involved in the stress response triggered by resazurin (Alamar Blue) in cultured cancer cells. Resazurin is a redox reactive compound widely used as reporter agent in assays of cell biology (e.g. cell viability and metabolic activity) due to its colorimetric and fluorimetric properties. In order to investigate resazurin-induced stress mechanisms we employed cells affording different metabolic and regulatory phenotypes. In HL-60 and Jurkat leukemia cells resazurin caused mitochondrial disintegration, respiratory dysfunction, reduced proliferation, and cell death. These effects were preceded by a burst of ROS, especially in HL-60 cells which were also more sensitive and contained autophagic vesicles. Studies in Rho(0) cells (devoid of mitochondrial DNA) indicated that the stress response does not depend on the rates of mitochondrial respiration. The anti-proliferative effect of resazurin was confirmed in native acute myelogenous leukemia (AML) blasts. In conclusion, the data suggest that resazurin triggers cellular ROS production and thereby initiates a stress response leading to mitochondrial dysfunction, reduced proliferation, autophagy, and cell degradation. The ability of cells to tolerate this type of stress may be important in toxicity and chemoresistance. © 2010 Wiley-Liss, Inc.

Investigation of functional activity human dental pulp stem cells at acute and chronic pulpitis.

PubMed

Ustiashvili, M; Kordzaia, D; Mamaladze, M; Jangavadze, M; Sanodze, L

2014-09-01

It is already recognized that together with the other connective tissues organ-specific progenic stem cells are also found in postnatal dental pulp. This group of undifferentiated cells is only 1% of total cell population of the pulp. The aim of the study was the identification of stem cells in human dental pulp, detection of their localization and assessment of functional activity during inflammation process and/or at norm. The obtained results showed that at acute pulpitis the pulp stroma is hypocellular in comparison with the norm but cells proliferative activity is low. CD 133 and NCAM (CD 56) positive stem cells were found in perivascularl space of the pulp stroma and in Hohle layer. At process prolongation and transition to the chronic phase pulp stroma is hypercellular, the cells with large, rounded or oval-shaped nuclei with clear chromatin appear together with fibroblasts. They are distributed as about entire thickness of the stroma as especially Hohle layer. In such cells higher proliferative activity (Ki67 expression) was observed. The cells in the mentioned proliferation phase are intensively marked by CD133, the rate of which is high in Hohle layer and along it. A large number of NCAM (CD 56) positive cells appear in pulp stroma. During pulpitis an involvement of stem cells into the process of reparative dentinogenesis should be conducted stepwise. In acute cases of the disease, stem cell perivascularl mobilization and proliferation and its migration to Hohle layer occur in response to irritation /stimulation. Chronification of the process leads not only to the migration of stem cells to the periphery of the pulp but also s their В«maturationВ» (increase of NCAM expression in the stem cells), which causes an increase the number of dentin producing active odontoblasts and initiation of reparative dentinogenesis.

Maternal embryonic leucine zipper kinase is a novel target for proliferation-associated high-risk myeloma

PubMed Central

Bolomsky, Arnold; Heusschen, Roy; Schlangen, Karin; Stangelberger, Kathrin; Muller, Joséphine; Schreiner, Wolfgang; Zojer, Niklas; Caers, Jo; Ludwig, Heinz

2018-01-01

Treatment of high-risk patients is a major challenge in multiple myeloma. This is especially true for patients assigned to the gene expression profiling-defined proliferation subgroup. Although recent efforts have identified some key players of proliferative myeloma, genetic interactions and players that can be targeted with clinically effective drugs have to be identified in order to overcome the poor prognosis of these patients. We therefore examined maternal embryonic leucine zipper kinase (MELK) for its implications in hyper-proliferative myeloma and analyzed the activity of the MELK inhibitor OTSSP167 both in vitro and in vivo. MELK was found to be significantly overexpressed in the proliferative subgroup of myeloma. This finding translated into poor overall survival in patients with high vs. low MELK expression. Enrichment analysis of upregulated genes in myeloma cells of MELKhigh patients confirmed the strong implications in myeloma cell proliferation. Targeting MELK with OTSSP167 impaired the growth and survival of myeloma cells, thereby affecting central survival factors such as MCL-1 and IRF4. This activity was also observed in the 5TGM.1 murine model of myeloma. OTSSP167 reduced bone marrow infiltration and serum paraprotein levels in a dose-dependent manner. In addition, we revealed a strong link between MELK and other proliferation-associated high-risk genes (PLK-1, EZH2, FOXM1, DEPDC1) and MELK inhibition also impaired the expression of those genes. We therefore conclude that MELK is an essential component of a proliferative gene signature and that pharmacological inhibition of MELK represents an attractive novel approach to overcome the poor prognosis of high-risk patients with a proliferative expression pattern. PMID:29122991

Evaluation of the anti-proliferative and cytostatic effect of Citrus sinensis (orange) fruit juice

PubMed Central

Chinedu, Enegide; Arome, David; Ameh, Solomon F; Ameh, Gift E

2014-01-01

Aim: This work has been designed to evaluate the anti-proliferative and cytostatic effects of Citrus sinensis (orange) fruit juice on rapidly proliferating cells. Materials and Methods: The study was carried out on the seeds of Sorghum bicolor for 72 h. The mean radicle length (mm) of the seeds was taken at 48 and 72 h. Result: The result showed that when compared with the control, methotrexate, the standard drug showed a significant (P < 0.001) anti-proliferative effect throughout the experiment. The inhibition of the radicle growth was more after 72 h (87.42%). At a dose of 5% (v/v), the juice showed a slightly significant (P < 0.05) effect affect after 72 h; however, there was no significant effect at 48 h. The juice at doses of 10% and 20% (v/v) showed a highly significant (P < 0.001) anti-proliferative effect throughout the experiment; however, the percentage inhibitions were higher at 72 h. At 72 h, the percentage inhibition for juice at 10% (v/v) was 72.37% and at 20% (v/v) was 91.96%. The concentrations of 40% and 60% (v/v) showed cytostatic effects as no appreciable growth of the radicles of the seeds was observed throughout the experiment. The percentage inhibition for 40% (v/v) was 100% and 99.72% for 48 and 72 h, respectively, while that for the juice concentration of 60% (v/v) was 100% throughout the study. Conclusion: The experiment has shown that C. sinensis fruit juice has a potential for causing both anti-proliferative and cytostatic effects on fast proliferating cells and hence cancerous cells. PMID:25298937

Replacement of Lost Lgr5-Positive Stem Cells through Plasticity of Their Enterocyte-Lineage Daughters.

PubMed

Tetteh, Paul W; Basak, Onur; Farin, Henner F; Wiebrands, Kay; Kretzschmar, Kai; Begthel, Harry; van den Born, Maaike; Korving, Jeroen; de Sauvage, Frederic; van Es, Johan H; van Oudenaarden, Alexander; Clevers, Hans

2016-02-04

Intestinal crypts display robust regeneration upon injury. The relatively rare secretory precursors can replace lost stem cells, but it is unknown if the abundant enterocyte progenitors that express the Alkaline phosphate intestinal (Alpi) gene also have this capacity. We created an Alpi-IRES-CreERT2 (Alpi(CreER)) knockin allele for lineage tracing. Marked clones consist entirely of enterocytes and are all lost from villus tips within days. Genetic fate-mapping of Alpi(+) cells before or during targeted ablation of Lgr5-expressing stem cells generated numerous long-lived crypt-villus "ribbons," indicative of dedifferentiation of enterocyte precursors into Lgr5(+) stems. By single-cell analysis of dedifferentiating enterocytes, we observed the generation of Paneth-like cells and proliferative stem cells. We conclude that the highly proliferative, short-lived enterocyte precursors serve as a large reservoir of potential stem cells during crypt regeneration. Copyright © 2016 Elsevier Inc. All rights reserved.

Comparative study of allogenic and xenogeneic mesenchymal stem cells on cisplatin-induced acute kidney injury in Sprague-Dawley rats.

PubMed

Ashour, Rehab H; Saad, Mohamed-Ahdy; Sobh, Mohamed-Ahmed; Al-Husseiny, Fatma; Abouelkheir, Mohamed; Awad, Amal; Elghannam, Doaa; Abdel-Ghaffar, Hassan; Sobh, Mohamed

2016-09-01

The paracrine and regenerative activities of mesenchymal stem cells (MSCs) may vary with different stem cell sources. The aim of the present study is to compare the effects of MSCs from different sources on acute kidney injury (AKI) induced by cisplatin and their influence on renal regeneration. A single intraperitoneal injection of cisplatin (5 mg/kg) was used to induce AKI in 120 Sprague-Dawley rats. Rats were treated with either rat bone marrow stem cells (rBMSCs), human adipose tissue-derived stem cells (hADSCs), or human amniotic fluid-derived stem cells (hAFSCs). 5 × 10(6) MSCs of different sources were administered through rat tail vein in a single dose, 24 hours after cisplatin injection. Within each group, rats were sacrificed at the 4th, 7th, 11th, and 30th day after cisplatin injection. Serum creatinine, BUN, and renal tissue oxidative stress parameters were measured. Renal tissue was scored histopathologically for evidence of injury, regeneration, and chronicity. Immunohistochemistry was also done using Ki67 for renal proliferative activity evaluation. MSCs of the three sources were able to ameliorate cisplatin-induced renal function deterioration and tissue damage. The rat BMSCs-treated group had the lowest serum creatinine by day 30 (0.52 ± 0.06) compared to hADSCs and hAFSCs. All MSC-treated groups had nearly equal antioxidant activity as indicated by the decreased renal tissue malondialdehyde (MDA) and increased reduced glutathione (GSH) level and superoxide dismutase (SOD) activity at different time intervals. Additionally, all MSCs improved injury and regenerative scores. Rat BMSCs had the highest count and earliest proliferative activity in the renal cortex by day 7 as identified by Ki67; while, hAFSCs seem to have the greatest improvement in the regenerative and proliferative activities with a higher count of renal cortex Ki67-positive cells at day 11 and with the least necrotic lesions. Rat BMSCs, hADSCs, and hAFSCs, in early single IV dose, had a renoprotective effect against cisplatin-induced AKI, and were able to reduce oxidative stress markers. Rat BMSCs had the earliest proliferative activity by day 7; however, hAFSCs seemed to have the greatest improvement in the regenerative activities. Human ADSCs were the least effective in the terms of proliferative and regenerative activities.

Abnormalities in the basement membrane structure promote basal keratinocytes in the epidermis of hypertrophic scars to adopt a proliferative phenotype.

PubMed

Yang, Shaowei; Sun, Yexiao; Geng, Zhijun; Ma, Kui; Sun, Xiaoyan; Fu, Xiaobing

2016-05-01

The majority of studies on scar formation have mainly focused on the dermis and little is known of the involvement of the epidermis. Previous research has demonstrated that the scar tissue-derived keratinocytes are different from normal cells at both the genetic and cell biological levels; however, the mechanisms responsible for the fundamental abnormalities in keratinocytes during scar development remain elusive. For this purpose, in this study, we used normal, wound edge and hypertrophic scar tissue to examine the morphological changes which occur during epidermal regeneration as part of the wound healing process and found that the histological structure of hypertrophic scar tissues differed from that of normal skin, with a significant increase in epidermal thickness. Notably, staining of the basement membrane (BM) appeared to be absent in the scar tissues. Moreover, immunofluorescence staining for cytokeratin (CK)10, CK14, CK5, CK19 and integrin-β1 indicated the differential expression of cell markers in the epidermal keratinocytes among the normal, wound edge and hypertrophic scar tissues, which corresponded with the altered BM structures. By using a panel of proteins associated with BM components, we validated our hypothesis that the BM plays a significant role in regulating the cell fate decision of epidermal keratinocytes during skin wound healing. Alterations in the structure of the BM promote basal keratinocytes to adopt a proliferative phenotype both in vivo and in vitro.

Abnormalities in the basement membrane structure promote basal keratinocytes in the epidermis of hypertrophic scars to adopt a proliferative phenotype

PubMed Central

YANG, SHAOWEI; SUN, YEXIAO; GENG, ZHIJUN; MA, KUI; SUN, XIAOYAN; FU, XIAOBING

2016-01-01

The majority of studies on scar formation have mainly focused on the dermis and little is known of the involvement of the epidermis. Previous research has demonstrated that the scar tissue-derived keratinocytes are different from normal cells at both the genetic and cell biological levels; however, the mechanisms responsible for the fundamental abnormalities in keratinocytes during scar development remain elusive. For this purpose, in this study, we used normal, wound edge and hypertrophic scar tissue to examine the morphological changes which occur during epidermal regeneration as part of the wound healing process and found that the histological structure of hypertrophic scar tissues differed from that of normal skin, with a significant increase in epidermal thickness. Notably, staining of the basement membrane (BM) appeared to be absent in the scar tissues. Moreover, immunofluorescence staining for cytokeratin (CK)10, CK14, CK5, CK19 and integrin-β1 indicated the differential expression of cell markers in the epidermal keratinocytes among the normal, wound edge and hypertrophic scar tissues, which corresponded with the altered BM structures. By using a panel of proteins associated with BM components, we validated our hypothesis that the BM plays a significant role in regulating the cell fate decision of epidermal keratinocytes during skin wound healing. Alterations in the structure of the BM promote basal keratinocytes to adopt a proliferative phenotype both in vivo and in vitro. PMID:26986690

Regulation of Telomere Homeostasis during Epstein-Barr virus Infection and Immortalization.

PubMed

Kamranvar, Siamak A; Masucci, Maria G

2017-08-09

The acquisition of unlimited proliferative potential is dependent on the activation of mechanisms for telomere maintenance, which counteracts telomere shortening and the consequent triggering of the DNA damage response, cell cycle arrest, and apoptosis. The capacity of Epstein Barr virus (EBV) to infect B-lymphocytes in vitro and transform the infected cells into autonomously proliferating immortal cell lines underlies the association of this human gamma-herpesvirus with a broad variety of lymphoid and epithelial cell malignancies. Current evidence suggests that both telomerase-dependent and -independent pathways of telomere elongation are activated in the infected cells during the early and late phases of virus-induced immortalization. Here we review the interaction of EBV with different components of the telomere maintenance machinery and the mechanisms by which the virus regulates telomere homeostasis in proliferating cells. We also discuss how these viral strategies may contribute to malignant transformation.

Real-time tracking of cell cycle progression during CD8+ effector and memory T-cell differentiation

PubMed Central

Kinjyo, Ichiko; Qin, Jim; Tan, Sioh-Yang; Wellard, Cameron J.; Mrass, Paulus; Ritchie, William; Doi, Atsushi; Cavanagh, Lois L.; Tomura, Michio; Sakaue-Sawano, Asako; Kanagawa, Osami; Miyawaki, Atsushi; Hodgkin, Philip D.; Weninger, Wolfgang

2015-01-01

The precise pathways of memory T-cell differentiation are incompletely understood. Here we exploit transgenic mice expressing fluorescent cell cycle indicators to longitudinally track the division dynamics of individual CD8+ T cells. During influenza virus infection in vivo, naive T cells enter a CD62Lintermediate state of fast proliferation, which continues for at least nine generations. At the peak of the anti-viral immune response, a subpopulation of these cells markedly reduces their cycling speed and acquires a CD62Lhi central memory cell phenotype. Construction of T-cell family division trees in vitro reveals two patterns of proliferation dynamics. While cells initially divide rapidly with moderate stochastic variations of cycling times after each generation, a slow-cycling subpopulation displaying a CD62Lhi memory phenotype appears after eight divisions. Phenotype and cell cycle duration are inherited by the progeny of slow cyclers. We propose that memory precursors cell-intrinsically modulate their proliferative activity to diversify differentiation pathways. PMID:25709008

Real-time tracking of cell cycle progression during CD8+ effector and memory T-cell differentiation.

PubMed

Kinjyo, Ichiko; Qin, Jim; Tan, Sioh-Yang; Wellard, Cameron J; Mrass, Paulus; Ritchie, William; Doi, Atsushi; Cavanagh, Lois L; Tomura, Michio; Sakaue-Sawano, Asako; Kanagawa, Osami; Miyawaki, Atsushi; Hodgkin, Philip D; Weninger, Wolfgang

2015-02-24

The precise pathways of memory T-cell differentiation are incompletely understood. Here we exploit transgenic mice expressing fluorescent cell cycle indicators to longitudinally track the division dynamics of individual CD8(+) T cells. During influenza virus infection in vivo, naive T cells enter a CD62L(intermediate) state of fast proliferation, which continues for at least nine generations. At the peak of the anti-viral immune response, a subpopulation of these cells markedly reduces their cycling speed and acquires a CD62L(hi) central memory cell phenotype. Construction of T-cell family division trees in vitro reveals two patterns of proliferation dynamics. While cells initially divide rapidly with moderate stochastic variations of cycling times after each generation, a slow-cycling subpopulation displaying a CD62L(hi) memory phenotype appears after eight divisions. Phenotype and cell cycle duration are inherited by the progeny of slow cyclers. We propose that memory precursors cell-intrinsically modulate their proliferative activity to diversify differentiation pathways.

The touch dome defines an epidermal niche specialized for mechanosensory signaling

PubMed Central

Doucet, Yanne S.; Woo, Seung-Hyun; Ruiz, Marlon E.; Owens, David M.

2013-01-01

Summary In mammalian skin, Merkel cells are mechanoreceptor cells that are required for the perception of gentle touch. Recent evidence indicates that mature Merkel cells descend from the proliferative layer of skin epidermis; however, the stem cell niche for Merkel cell homeostasis has not been reported. Here, we provide the first genetic evidence for maintenance of mature Merkel cells during homeostasis by Krt17+ stem cells located in epidermal touch domes of hairy skin and in the tips of the rete ridges of glabrous skin. Lineage tracing analysis indicated that the entire pool of mature Merkel cells is turned over every 7–8 weeks in adult epidermis and that Krt17+ stem cells also maintain squamous differentiation in the touch dome and in glabrous skin. Finally, selective genetic ablation of Krt17+ touch dome keratinocytes indicates that these cells, and not mature Merkel cells, are primarily responsible for maintaining innervation of the Merkel cell-neurite complex. PMID:23727240

Quiescent and Proliferative Fibroblasts Exhibit Differential p300 HAT Activation through Control of 5-Methoxytryptophan Production

PubMed Central

Chu, Ling-yun; Chang, Tzu-Ching; Kuo, Cheng-Chin; Wu, Kenneth K.

2014-01-01

Quiescent fibroblasts possess unique genetic program and exhibit high metabolic activity distinct from proliferative fibroblasts. In response to inflammatory stimulation, quiescent fibroblasts are more active in expressing cyclooxygenase-2 and other proinflammatory genes than proliferative fibroblasts. The underlying transcriptional mechanism is unclear. Here we show that phorbol 12-myristate 13-acetate (PMA) and cytokines increased p300 histone acetyltransferase activity to a higher magnitude (> 2 fold) in quiescent fibroblasts than in proliferative fibroblasts. Binding of p300 to cyclooxygenase-2 promoter was reduced in proliferative fibroblasts. By ultrahigh-performance liquid chromatography coupled with a quadrupole time of flight mass spectrometer and enzyme-immunoassay, we found that production of 5-methoxytryptophan was 2–3 folds higher in proliferative fibroblasts than that in quiescent fibroblasts. Addition of 5-methoxytryptophan and its metabolic precursor, 5-hydroxytryptophan, to quiescent fibroblasts suppressed PMA-induced p300 histone acetyltransferase activity and cyclooxygenase-2 expression to the level of proliferative fibroblasts. Silencing of tryptophan hydroxylase-1 or hydroxyindole O-methyltransferase in proliferative fibroblasts with siRNA resulted in elevation of PMA-induced p300 histone acetyltransferase activity to the level of that in quiescent fibroblasts, which was rescued by addition of 5-hydroxytryptophan or 5-methoxytryptophan. Our findings indicate that robust inflammatory gene expression in quiescent fibroblasts vs. proliferative fibroblasts is attributed to uncontrolled p300 histone acetyltransferase activation due to deficiency of 5-methoxytryptophan production. 5-methoxytryptophan thus is a potential valuable lead compound for new anti-inflammatory drug development. PMID:24523905

[Immunohistochemical description of proliferative activity and apoptosis of lung squamous cell carcinoma (literature review)].

PubMed

Филенко, Борис Н; Ройко, Наталия В; Степанчук, Алла П; Проскурня, Сергей А

2016-01-01

The analysis of the publications are describe immunohistochemical study of proliferative activity and apoptosis of lung squamous cell carcinoma. Established that the imbalance between proliferation and cell death is a key process in the development of tumors. However, the value of tumor markers in histogenesis and morfogenesis of tumors and forecast their occurrence is not studied enough. Despite the significant amount of scientific literature devoted to this issue, has not yet established a clear link expression of immunohistochemical markers of proliferation and apoptosis with the degree of differentiation of squamous cell lung cancer. Analysis of the literature shows that the morphology of this histogenetics type lung cancer at the cellular, subcellular structural and functional levels are controversial and require detailed investigation.

Induction of sister chromatid exchanges and inhibition of cellular proliferation in vitro. I. Caffeine

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

Guglielmi, G.E.; Vogt, T.F.; Tice, R.R.

1982-01-01

While many agents have been examined for their ability to induce SCE's, complete dose-response information has often been lacking. We have reexamined the ability of one such compound - caffeine - to induce SCEs and also to inhibit cellular proliferation in human peripheral lymphocytes in vitro. An acute exposure to caffeine prior to the DNA synthetic period did not affect either SCE frequency or the rate of cellular proliferation. Chronic exposure to caffeine throughout the culture period lead to both a dose-dependent increase in SCEs (SCE/sub d/ or doubling dose = 2.4 mM; SCE/sub 10/ or the dose capable ofmore » inducing 10 SCE = 1.4 mM) and a dose-dependent inhibition of cellular proliferation (IC/sub 50/ or the 50% inhibition concentration = 2.6 mM). The relative proportion of first generation metaphase cells, an assessment of proliferative inhibiton, increased linearly with increasing caffeine concentrations. However, SCE frequency increased nonlinearly over the same range of caffeine concentrations. Examination of the ratio of nonsymmetrical to symmetrical SCEs in third generation metaphase cells indicated that caffeine induced SCEs in equal frequency in each of three successive generations. The dependency of SCE induction and cellular proliferative inhibition on caffeine's presence during the DNA synthetic period suggests that caffeine may act as an antimetabolite in normal human cells.« less

Premature chromosome condensation studies in human leukemia. I. Pretreatment characteristics.

PubMed

Hittelman, W N; Broussard, L C; McCredie, K

1979-11-01

The phenomenon of premature chromosome condensation (PCC) was used to compare the bone marrow proliferation characteristics of 163 patients with various forms of leukemia prior to the initiation of new therapy. The proliferative potential index (PPI, or fraction of G1 cells in late G1 phase) and the fraction of cells in S phase was determined and compared to the type of disease and the bone marrow blast infiltrate for each patient. Previously untreated patients with acute leukemia exhibited an average PPI value three times that of normal bone marrow (37.5% for acute myeloblastic leukemia [AML], acute monomyeloblastic leukemia [AMML], or acute promyelocytic leukemia [APML] and 42% for acute lymphocytic leukemia [ALL] or acute undifferentiated leukemia [AUL]). Untreated chronic myelogenous leukemia (CML) patients showed intermediate PPI values (25.2%), whereas CML patients with controlled disease exhibited nearly normal PPI values (14.6%). On the other hand, blastic-phase CML patients exhibited PPI values closer to that observed in patients with acute leukemia (35.4%). Seven patients with chronic lymphocytic leukemia (CLL) exhibited even higher PPI values. No correlations were observed between PPI values, fraction of cells in S phase, and marrow blast infiltrate. For untreated acute disease patients, PPI values were prognostic for response only at low and high PPI values. These results suggest that the PCC-determined proliferative potential is a biologic reflection of the degree of malignancy within the bone marrow.

Stem Cell-Containing Hyaluronic Acid-Based Spongy Hydrogels for Integrated Diabetic Wound Healing.

PubMed

da Silva, Lucília Pereira; Santos, Tírcia Carlos; Rodrigues, Daniel Barreira; Pirraco, Rogério Pedro; Cerqueira, Mariana Teixeira; Reis, Rui Luís; Correlo, Vitor Manuel; Marques, Alexandra Pinto

2017-07-01

The detailed pathophysiology of diabetic foot ulcers is yet to be established and improved treatments are still required. We propose a strategy that directs inflammation, neovascularization, and neoinnervation of diabetic wounds. Aiming to potentiate a relevant secretome for nerve regeneration, stem cells were precultured in hyaluronic acid-based spongy hydrogels under neurogenic/standard media before transplantation into diabetic mice full-thickness wounds. Acellular spongy hydrogels and empty wounds were used as controls. Re-epithelialization was attained 4 weeks after transplantation independently of the test groups, whereas a thicker and more differentiated epidermis was observed for the cellular spongy hydrogels. A switch from the inflammatory to the proliferative phase of wound healing was revealed for all the experimental groups 2 weeks after injury, but a significantly higher M2(CD163 + )/M1(CD86 + ) subtype ratio was observed in the neurogenic preconditioned group that also failed to promote neoinnervation. A higher number of intraepidermal nerve fibers were observed for the unconditioned group probably due to a more controlled transition from the inflammatory to the proliferative phase. Overall, stem cell-containing spongy hydrogels represent a promising approach to enhance diabetic wound healing by positively impacting re-epithelialization and by modulating the inflammatory response to promote a successful neoinnervation. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Nonlinear ghost waves accelerate the progression of high-grade brain tumors

NASA Astrophysics Data System (ADS)

Pardo, Rosa; Martínez-González, Alicia; Pérez-García, Víctor M.

2016-10-01

We study a reduced continuous model describing the evolution of high grade gliomas in response to hypoxic events through the interplay of different cellular phenotypes. We show that hypoxic events, even when sporadic and/or limited in space, may have a crucial role on the acceleration of high grade gliomas growth. Our modeling approach is based on two cellular phenotypes. One of them is more migratory and a second one is more proliferative. Transitions between both phenotypes are driven by the local oxygen values, assumed in this simple model to be uniform. Surprisingly, even very localized in time hypoxia events leading to transient migratory populations have the potential to accelerate the tumor's invasion speed up to speeds close to those of the migratory phenotype. The high invasion speed persists for times much longer than the lifetime of the hypoxic event. Moreover, the phenomenon is observed both when the migratory cells form a persistent wave of cells located on the invasion front and when they form a evanescent "ghost" wave disappearing after a short time by decay to the more proliferative phenotype. Our findings are obtained through numerical simulations of the model equations both in 1D and higher dimensional scenarios. We also provide a deeper mathematical analysis of some aspects of the problem such as the conditions for the existence of persistent waves of cells with a more migratory phenotype.

Murine c-mpl: a member of the hematopoietic growth factor receptor superfamily that transduces a proliferative signal.

PubMed Central

Skoda, R C; Seldin, D C; Chiang, M K; Peichel, C L; Vogt, T F; Leder, P

1993-01-01

The murine myeloproliferative leukemia virus has previously been shown to contain a fragment of the coding region of the c-mpl gene, a member of the cytokine receptor superfamily. We have isolated cDNA and genomic clones encoding murine c-mpl and localized the c-mpl gene to mouse chromosome 4. Since some members of this superfamily function by transducing a proliferative signal and since the putative ligand of mpl is unknown, we have generated a chimeric receptor to test the functional potential of mpl. The chimera consists of the extracellular domain of the human interleukin-4 receptor and the cytoplasmic domain of mpl. A mouse hematopoietic cell line transfected with this construct proliferates in response to human interleukin-4, thereby demonstrating that the cytoplasmic domain of mpl contains all elements necessary to transmit a growth stimulatory signal. In addition, we show that 25-40% of mpl mRNA found in the spleen corresponds to a novel truncated and potentially soluble isoform of mpl and that both full-length and truncated forms of mpl protein can be immunoprecipitated from lysates of transfected COS cells. Interestingly, however, although the truncated form of the receptor possesses a functional signal sequence and lacks a transmembrane domain, it is not detected in the culture media of transfected cells. Images PMID:8334987