Klotho gene expression decreases in peripheral blood mononuclear cells (PBMCs) of patients with relapsing-remitting multiple sclerosis.

PubMed

Karami, Masoumeh; Mehrabi, Farzad; Allameh, Abdolamir; Pahlevan Kakhki, Majid; Amiri, Mehdi; Emami Aleagha, Mohammad Sajad

2017-10-15

we recently showed that a hypothesized anti-aging and anti-inflammatory protein, namely Klotho, may contribute to the etiology and/or pathogenesis of multiple sclerosis (MS). In addition, Klotho function and its gene expression are dependent on inflammatory pathways. Accordingly, the aim of this study was to investigate the Klotho gene expression within peripheral blood mononuclear cells (PBMCs) of patients with MS. Altogether, 30 patients with relapsing-remitting MS (RRMS) along with 30 age and sex-matched healthy individuals were enrolled in this study. Blood samples were obtained from all participants and then PBMCs were isolated. The quantitative Real-Time PCR was carried out for Klotho mRNA derived from PBMCs. The results showed that klotho gene expression in the PBMCs of patients with RRMS is nearly 2.5-fold less than healthy individuals (P=0.0006). This is the first study demonstrating a possible role of Klotho in the PBMCs of MS patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Desipramine decreases expression of human and murine indoleamine-2,3-dioxygenases

PubMed Central

Brooks, Alexandra K.; Janda, Tiffany M.; Lawson, Marcus A.; Rytych, Jennifer L.; Smith, Robin A.; Ocampo-Solis, Cecilia; McCusker, Robert H.

2017-01-01

Abundant evidence connects depression symptomology with immune system activation, stress and subsequently elevated levels of kynurenine. Anti-depressants, such as the tricyclic norepinephrine/serotonin reuptake inhibitor desipramine (Desip), were developed under the premise that increasing extracellular neurotransmitter level was the sole mechanism by which they alleviate depressive symptomologies. However, evidence suggests that anti-depressants have additional actions that contribute to their therapeutic potential. The Kynurenine Pathway produces tryptophan metabolites that modulate neurotransmitter activity. This recognition identified another putative pathway for anti-depressant targeting. Considering a recognized role of the Kynurenine Pathway in depression, we investigated the potential for Desip to alter expression of rate-limiting enzymes of this pathway: indoleamine-2,3-dioxygenases (Ido1 and Ido2). Mice were administered lipopolysaccharide (LPS) or synthetic glucocorticoid dexamethasone (Dex) with Desip to determine if Desip alters indoleamine-dioxygenase (DO) expression in vivo following a modeled immune and stress response. This work was followed by treating murine and human peripheral blood mononuclear cells (PBMCs) with interferon-gamma (IFNγ) and Desip. In vivo: Desip blocked LPS-induced Ido1 expression in hippocampi, astrocytes, microglia and PBMCs and Ido2 expression by PBMCs. Ex vivo: Desip decreased IFNγ-induced Ido1 and Ido2 expression in murine PBMCs. This effect was directly translatable to the human system as Desip decreased IDO1 and IDO2 expression by human PBMCs. These data demonstrate for the first time that an anti-depressant alters expression of Ido1 and Ido2, identifying a possible new mechanism behind anti-depressant activity. Furthermore, we propose the assessment of PBMCs for anti-depressant responsiveness using IDO expression as a biomarker. PMID:28212884

Dental Calculus Stimulates Interleukin-1β Secretion by Activating NLRP3 Inflammasome in Human and Mouse Phagocytes

PubMed Central

Montenegro Raudales, Jorge Luis; Yoshimura, Atsutoshi; SM, Ziauddin; Kaneko, Takashi; Ozaki, Yukio; Ukai, Takashi; Miyazaki, Toshihiro; Latz, Eicke; Hara, Yoshitaka

2016-01-01

Dental calculus is a mineralized deposit associated with periodontitis. The bacterial components contained in dental calculus can be recognized by host immune sensors, such as Toll-like receptors (TLRs), and induce transcription of proinflammatory cytokines, such as IL-1β. Studies have shown that cellular uptake of crystalline particles may trigger NLRP3 inflammasome activation, leading to the cleavage of the IL-1β precursor to its mature form. Phagocytosis of dental calculus in the periodontal pocket may therefore lead to the secretion of IL-1β, promoting inflammatory responses in periodontal tissues. However, the capacity of dental calculus to induce IL-1β secretion in human phagocytes has not been explored. To study this, we stimulated human polymorphonuclear leukocytes (PMNs) and peripheral blood mononuclear cells (PBMCs) with dental calculus collected from periodontitis patients, and measured IL-1β secretion by ELISA. We found that calculus induced IL-1β secretion in both human PMNs and PBMCs. Calculus also induced IL-1β in macrophages from wild-type mice, but not in macrophages from NLRP3- and ASC-deficient mice, indicating the involvement of NLRP3 and ASC. IL-1β induction was inhibited by polymyxin B, suggesting that LPS is one of the components of calculus that induces pro-IL-1β transcription. To analyze the effect of the inorganic structure, we baked calculus at 250°C for 1 h. This baked calculus failed to induce pro-IL-1β transcription. However, it did induce IL-1β secretion in lipid A-primed cells, indicating that the crystalline structure of calculus induces inflammasome activation. Furthermore, hydroxyapatite crystals, a component of dental calculus, induced IL-1β in mouse macrophages, and baked calculus induced IL-1β in lipid A-primed human PMNs and PBMCs. These results indicate that dental calculus stimulates IL-1β secretion via NLRP3 inflammasome in human and mouse phagocytes, and that the crystalline structure has a partial role in

Dental Calculus Stimulates Interleukin-1β Secretion by Activating NLRP3 Inflammasome in Human and Mouse Phagocytes.

PubMed

Montenegro Raudales, Jorge Luis; Yoshimura, Atsutoshi; Sm, Ziauddin; Kaneko, Takashi; Ozaki, Yukio; Ukai, Takashi; Miyazaki, Toshihiro; Latz, Eicke; Hara, Yoshitaka

2016-01-01

Dental calculus is a mineralized deposit associated with periodontitis. The bacterial components contained in dental calculus can be recognized by host immune sensors, such as Toll-like receptors (TLRs), and induce transcription of proinflammatory cytokines, such as IL-1β. Studies have shown that cellular uptake of crystalline particles may trigger NLRP3 inflammasome activation, leading to the cleavage of the IL-1β precursor to its mature form. Phagocytosis of dental calculus in the periodontal pocket may therefore lead to the secretion of IL-1β, promoting inflammatory responses in periodontal tissues. However, the capacity of dental calculus to induce IL-1β secretion in human phagocytes has not been explored. To study this, we stimulated human polymorphonuclear leukocytes (PMNs) and peripheral blood mononuclear cells (PBMCs) with dental calculus collected from periodontitis patients, and measured IL-1β secretion by ELISA. We found that calculus induced IL-1β secretion in both human PMNs and PBMCs. Calculus also induced IL-1β in macrophages from wild-type mice, but not in macrophages from NLRP3- and ASC-deficient mice, indicating the involvement of NLRP3 and ASC. IL-1β induction was inhibited by polymyxin B, suggesting that LPS is one of the components of calculus that induces pro-IL-1β transcription. To analyze the effect of the inorganic structure, we baked calculus at 250°C for 1 h. This baked calculus failed to induce pro-IL-1β transcription. However, it did induce IL-1β secretion in lipid A-primed cells, indicating that the crystalline structure of calculus induces inflammasome activation. Furthermore, hydroxyapatite crystals, a component of dental calculus, induced IL-1β in mouse macrophages, and baked calculus induced IL-1β in lipid A-primed human PMNs and PBMCs. These results indicate that dental calculus stimulates IL-1β secretion via NLRP3 inflammasome in human and mouse phagocytes, and that the crystalline structure has a partial role in

Stress-related methylation of the catechol-O-methyltransferase Val 158 allele predicts human prefrontal cognition and activity.

PubMed

Ursini, Gianluca; Bollati, Valentina; Fazio, Leonardo; Porcelli, Annamaria; Iacovelli, Luisa; Catalani, Assia; Sinibaldi, Lorenzo; Gelao, Barbara; Romano, Raffaella; Rampino, Antonio; Taurisano, Paolo; Mancini, Marina; Di Giorgio, Annabella; Popolizio, Teresa; Baccarelli, Andrea; De Blasi, Antonio; Blasi, Giuseppe; Bertolino, Alessandro

2011-05-04

DNA methylation at CpG dinucleotides is associated with gene silencing, stress, and memory. The catechol-O-methyltransferase (COMT) Val(158) allele in rs4680 is associated with differential enzyme activity, stress responsivity, and prefrontal activity during working memory (WM), and it creates a CpG dinucleotide. We report that methylation of the Val(158) allele measured from peripheral blood mononuclear cells (PBMCs) of Val/Val humans is associated negatively with lifetime stress and positively with WM performance; it interacts with stress to modulate prefrontal activity during WM, such that greater stress and lower methylation are related to reduced cortical efficiency; and it is inversely related to mRNA expression and protein levels, potentially explaining the in vivo effects. Finally, methylation of COMT in prefrontal cortex and that in PBMCs of rats are correlated. The relationship of methylation of the COMT Val(158) allele with stress, gene expression, WM performance, and related brain activity suggests that stress-related methylation is associated with silencing of the gene, which partially compensates the physiological role of the high-activity Val allele in prefrontal cognition and activity. Moreover, these results demonstrate how stress-related DNA methylation of specific functional alleles impacts directly on human brain physiology beyond sequence variation.

Phospholipase A2 Inhibitor from Crotalus durissus terrificus rattlesnake: Effects on human peripheral blood mononuclear cells and human neutrophils cells.

PubMed

Xavier, Caroline V; da S Setúbal, Sulamita; Lacouth-Silva, Fabianne; Pontes, Adriana S; Nery, Neriane M; de Castro, Onassis Boeri; Fernandes, Carla F C; Soares, Andreimar M; Fortes-Dias, Consuelo L; Zuliani, Juliana P

2017-12-01

Crotalus Neutralizing Factor (CNF) is an inhibitor of phospholipase A 2 (PLA 2 ), present in the blood plasma of Crotalus durissus terrificus snake. This inhibitor neutralizes the lethal and enzymatic activity of crotoxin, the main neurotoxin from this venom. In this study, we investigated the effects of CNF on the functionality of human peripheral blood mononuclear cells (PBMCs) and human neutrophils. The following parameters were evaluated: viability and proliferation, chemotaxis, cytokines and LTB 4 production, cytosolic PLA 2 s activity, myeloperoxidase (MPO) and superoxide anion (O 2 - ) production. CNF showed no toxicity on PBMCs or neutrophils, and acts by stimulating the release of TNF-α and LTB 4 , but neither stimulates IL-10 and IL-2 nor affects PBMCs proliferation and O 2 - release. In neutrophils, CNF induces chemotaxis but does not induce the release of both MPO and O 2 - . However, it induces LTB 4 and IL-8 production. These data show the influence of CNF on PBMCs' function by inducing TNF-α and LTB 4 production, and on neutrophils, by stimulating chemotaxis and LTB 4 production, via cytosolic PLA 2 activity, and IL-8 release. The inflammatory profile produced by CNF is shown for the first time. Our present results suggest that CNF has a role in activation of leukocytes and exert proinflammatory effects on these cell. Copyright © 2017 Elsevier B.V. All rights reserved.

Perspectives on Systems Modeling of Human Peripheral Blood Mononuclear Cells

PubMed Central

Sen, Partho; Kemppainen, Esko; Orešič, Matej

2018-01-01

Human peripheral blood mononuclear cells (PBMCs) are the key drivers of the immune responses. These cells undergo activation, proliferation and differentiation into various subsets. During these processes they initiate metabolic reprogramming, which is coordinated by specific gene and protein activities. PBMCs as a model system have been widely used to study metabolic and autoimmune diseases. Herein we review various omics and systems-based approaches such as transcriptomics, epigenomics, proteomics, and metabolomics as applied to PBMCs, particularly T helper subsets, that unveiled disease markers and the underlying mechanisms. We also discuss and emphasize several aspects of T cell metabolic modeling in healthy and disease states using genome-scale metabolic models. PMID:29376056

Recombinant methionyl human leptin administration activates signal transducer and activator of transcription 3 signaling in peripheral blood mononuclear cells in vivo and regulates soluble tumor necrosis factor-alpha receptor levels in humans with relative leptin deficiency.

PubMed

Chan, Jean L; Moschos, Stergios J; Bullen, John; Heist, Kathleen; Li, Xian; Kim, Young-Bum; Kahn, Barbara B; Mantzoros, Christos S

2005-03-01

Studies of congenital complete leptin deficiency in animals and humans support a role for leptin in regulating immune function. Whether acquired relative leptin deficiency affects immunological parameters in healthy humans remains unknown. We thus used experimental models of relative leptin deficiency and recombinant methionyl human leptin (r-metHuLeptin) administration in humans to investigate whether r-metHuLeptin would activate signaling pathways in peripheral blood mononuclear cells (PBMCs) and whether acquired relative leptin deficiency and/or increasing circulating leptin levels into the physiologic range would change PBMC subpopulations and cytokines important in the T-helper cell and systemic immune responses. We found that r-metHuLeptin administration to healthy humans activates signal transducer and activator of transcription-3 signaling in PBMCs in vivo. Neither short-term leptin deficiency, induced by 3-d complete fasting, nor physiologic r-metHuLeptin replacement for the same period of time had a major effect on PBMC subpopulations or serum cytokines in healthy men. In contrast, normalizing serum leptin levels over 8 wk in lean women with relative leptin deficiency for 5.1 +/- 1.4 yr (mean +/- se) due to chronic energy deficit increased soluble TNFalpha receptor levels, indicating activation of the TNFalpha system. These findings suggest that relative leptin deficiency due to more long-term energy deprivation is associated with defects in immunological parameters that may be corrected with exogenous r-metHuLeptin administration. Further studies are warranted to assess the implications of acquired relative hypoleptinemia and/or r-metHuLeptin administration on the immunosuppression associated with energy- and leptin-deficient states in humans.

Replacing with whole grains and legumes reduces Lp-PLA2 activities in plasma and PBMCs in patients with prediabetes or T2D1

PubMed Central

Kim, Minjoo; Jeung, Se Ri; Jeong, Tae-Sook; Lee, Sang-Hyun; Lee, Jong Ho

2014-01-01

To determine dietary effects on circulating lipoprotein-associated phospholipase A2 (Lp-PLA2) activity and enzyme activity in peripheral blood mononuclear cells (PBMCs), 99 patients with impaired fasting glucose, impaired glucose tolerance, or newly-diagnosed T2D were randomly assigned to either a control group (usual diet with refined rice) or the whole grain and legume group. Substitution of whole grains and legumes for refined rice was associated with the replacement of 7% of energy from carbohydrates with energy from protein (about 4%) and fat. After 12 weeks, the whole grain and legume group showed a significant decrease in fasting glucose, insulin, homeostasis model assessment-insulin resistance, hemoglobin A1c, malondialdehyde, plasma Lp-PLA2 activity, and oxidized LDL (ox-LDL), and an increase in LDL particle size. The changes (Δs) in these variables in the whole grain and legume group were significantly different from those in controls after adjustment for the baseline levels. When all subjects were considered, Δ plasma Lp-PLA2 positively correlated with Δ glucose, Δ PBMC Lp-PLA2, Δ ox-LDL, and Δ urinary 8-epi-prostaglandin F2α after being adjusted for confounding factors. The Δ PBMC Lp-PLA2 correlated positively with Δ glucose and Δ ox-LDL, and negatively with Δ LDL particle size and baseline PBMC Lp-PLA2. The substitution of whole grains and legumes for refined rice resulted in a reduction in Lp-PLA2 activities in plasma and PBMCs partly through improved glycemic control, increased consumption of protein relative to carbohydrate, and reduced lipid peroxides. PMID:24904022

Microgramma vacciniifolia (Polypodiaceae) fronds contain a multifunctional lectin with immunomodulatory properties on human cells.

PubMed

de Siqueira Patriota, Leydianne Leite; Procópio, Thamara Figueiredo; de Santana Brito, Jéssica; Sebag, Virginie; de Oliveira, Ana Patrícia Silva; de Araújo Soares, Ana Karine; Moreira, Leyllane Rafael; de Albuquerque Lima, Thâmarah; Soares, Tatiana; da Silva, Túlio Diego; Paiva, Patrícia Maria Guedes; de Lorena, Virgínia Maria Barros; de Melo, Cristiane Moutinho Lagos; de Albuquerque, Lidiane Pereira; Napoleão, Thiago Henrique

2017-10-01

In this study, we report the purification and characterization of a multifunctional lectin (MvFL) from Microgramma vacciniifolia fronds as well as its immunomodulatory properties on human peripheral blood mononuclear cells (PBMCs). MvFL (pI 4.51; 54kDa) is a glycoprotein able to inhibit trypsin activity and that has sequence similarities (32% coverage) with a plant RNA-binding protein. Hemagglutinating activity of MvFL was not altered by heating at 100°C for 30min, but was reduced in alkaline pH (8.0 and 9.0). Fluorimetric analyses showed that this lectin did not undergo marked conformational changes when heated. However, the MvFL conformation changed depending on the pH. MvFL at 6.25-25μg/mL was not cytotoxic to lymphocytes present among PBMCs. The PBMCs incubated for 24h with the lectin (12.5μg/mL) showed increased TNF-α, IFN-γ, IL-6, IL-10, and nitric oxide production. MvFL also stimulated T lymphocytes from PBMCs to differentiate into CD8 + cells. The activation (indicated by CD28 expression) of these cells was also stimulated. In conclusion, MvFL is a heat-stable and multifunctional protein, with both lectin and trypsin inhibitor activities, and capable of inducing predominantly a Th1 response in human PBMCs as well as activation and differentiation of T lymphocytes. Copyright © 2017 Elsevier B.V. All rights reserved.

Antimony resistant Leishmania donovani but not sensitive ones drives greater frequency of potent T-regulatory cells upon interaction with human PBMCs: role of IL-10 and TGF-β in early immune response.

PubMed

Guha, Rajan; Das, Shantanabha; Ghosh, June; Sundar, Shyam; Dujardin, Jean Claude; Roy, Syamal

2014-07-01

In India the sand fly, Phlebotomus argentipes, transmitted parasitic disease termed kala-azar is caused by Leishmania donovani (LD) in humans. These immune-evading parasites have increasingly developed resistance to the drug sodium antimony gluconate in endemic regions. Lack of early diagnosis methods for the disease limits the information available regarding the early interactions of this parasite with either human tissues or cell lineages. We reasoned that peripheral blood mononuclear cells (PBMCs) from healthy human beings could help compare some of their immune signatures once they were exposed for up to 8 days, to either pentavalent antimony sensitive (Sb(S)-LD) or resistant (Sb(R)-LD) Leishmania donovani isolates. At day 2, PBMC cultures exposed to Sb(S)-LD and Sb(R)-LD stationary phase promastigotes had four and seven fold higher frequency of IL-10 secreting monocyte-macrophage respectively, compared to cultures unexposed to parasites. Contrasting with the CD4(+)CD25-CD127- type-1 T-regulatory (Tr1) cell population that displayed similar features whatever the culture conditions, there was a pronounced increase in the IL-10 producing CD4(+)CD25(+)CD127low/- inducible T-regulatory cells (iTregs) in the PBMC cultures sampled at day 8 post addition of Sb(R)-LD. Sorted iTregs from different cultures on day 8 were added to anti-CD3/CD28 induced naïve PBMCs to assess their suppressive ability. We observed that iTregs from Sb(R)-LD exposed PBMCs had more pronounced suppressive ability compared to Sb(S)-LD counterpart on a per cell basis and is dependent on both IL-10 and TGF-β, whereas IL-10 being the major factor contributing to the suppressive ability of iTregs sorted from PBMC cultures exposed to Sb(S)-LD. Of note, iTreg population frequency value remained at the basal level after addition of genetically modified Sb(R)-LD lacking unique terminal sugar in surface glycan. Even with limitations of this artificial in vitro model of L. donovani-human PBMC

Human umbilical cord mesenchymal stem cells hUC-MSCs exert immunosuppressive activities through a PGE2-dependent mechanism.

PubMed

Chen, Ke; Wang, Ding; Du, Wei Ting; Han, Zhi-Bo; Ren, He; Chi, Ying; Yang, Shao Guang; Zhu, Delin; Bayard, Francis; Han, Zhong Chao

2010-06-01

Human umbilical-cord-derived mesenchymal stem cells (hUC-MSCs) constitute an attractive alternative to bone-marrow-derived MSCs for potential clinical applications because of easy preparation and lower risk of viral contamination. In this study, both proliferation of human peripheral blood mononuclear cells (hPBMCs) and their IFN-gamma production in response to mitogenic or allogeneic stimulus were effectively inhibited by hUC-MSCs. Co-culture experiments in transwell systems indicated that the suppression was largely mediated by soluble factor(s). Blocking experiments identified prostaglandin E(2) (PGE(2)) as the major factor, because inhibition of PGE(2) synthesis almost completely mitigated the immunosuppressive effects, whereas neutralization of TGF-beta, IDO, and NO activities had little effects. Moreover, the inflammatory cytokines, IFN-gamma and IL-1beta, produced by hPBMCs upon activation notably upregulated the expression of cyclooxygenase-2 (COX-2) and the production of PGE(2) by hUC-MSCs. In conclusion, our data have demonstrated for the first time the PGE(2)-mediated mechanism by which hUC-MSCs exert their immunomodulatory effects. Copyright 2010 Elsevier Inc. All rights reserved.

Transcriptome analysis of the human T lymphocyte cell line Jurkat and human peripheral blood mononuclear cells exposed to deoxynivalenol (DON): New mechanistic insights

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

Katika, Madhumohan R.; Department of Health Risk Analysis and Toxicology, Maastricht University; Netherlands Toxicogenomics Centre

Deoxynivalenol (DON) or vomitoxin is a commonly encountered type-B trichothecene mycotoxin, produced by Fusarium species predominantly found in cereals and grains. DON is known to exert toxic effects on the gastrointestinal, reproductive and neuroendocrine systems, and particularly on the immune system. Depending on dose and exposure time, it can either stimulate or suppress immune function. The main objective of this study was to obtain a deeper insight into DON-induced effects on lymphoid cells. For this, we exposed the human T-lymphocyte cell line Jurkat and human peripheral blood mononuclear cells (PBMCs) to various concentrations of DON for various times and examinedmore » gene expression changes by DNA microarray analysis. Jurkat cells were exposed to 0.25 and 0.5 μM DON for 3, 6 and 24 h. Biological interpretation of the microarray data indicated that DON affects various processes in these cells: It upregulates genes involved in ribosome structure and function, RNA/protein synthesis and processing, endoplasmic reticulum (ER) stress, calcium-mediated signaling, mitochondrial function, oxidative stress, the NFAT and NF-κB/TNF-α pathways, T cell activation and apoptosis. The effects of DON on the expression of genes involved in ER stress, NFAT activation and apoptosis were confirmed by qRT-PCR. Other biochemical experiments confirmed that DON activates calcium-dependent proteins such as calcineurin and M-calpain that are known to be involved in T cell activation and apoptosis. Induction of T cell activation was also confirmed by demonstrating that DON activates NFATC1 and induces its translocation from the cytoplasm to the nucleus. For the gene expression profiling of PBMCs, cells were exposed to 2 and 4 μM DON for 6 and 24 h. Comparison of the Jurkat microarray data with those obtained with PBMCs showed that most of the processes affected by DON in the Jurkat cell line were also affected in the PBMCs. -- Highlights: ► The human T cell line Jurkat and

Enrofloxacin in therapeutic doses alters cytokine production by porcine PBMCs induced by lipopolysaccharide.

PubMed

Pomorska-Mól, Małgorzata; Czyżewska-Dors, Ewelina; Kwit, Krzysztof; Pejsak, Zygmunt

2017-07-01

The effect of enrofloxacin on cytokine secretion by porcine peripheral blood mononuclear cells (PBMCs) was studied. Twenty 8-20-week-old pigs were randomly divided into two groups: control (C, n = 10) and experimental (E, n = 10) were used. Pigs from group E received enrofloxacin at therapeutic dose for 5 consecutive days. Blood samples were collected at 0 (before antibiotic administration), 2, 4 (during antibiotic therapy) 6, 9, 14 21, 35, 49, and 63 d of study (after treatment). PBMCs of pigs from both groups were incubated with or without lipopolysaccharide (LPS). Ex vivo production on interleukin (IL)-4, IL-6, IL-10, INF-γ, and TNF-α were analyzed using ELISA assay. Intramuscular administration of enrofloxacin to healthy pigs for 5 consecutive days induced a transitory reduction of the ex vivo response of PBMCs to LPS in terms of IL-6 and TNF-α secretion. The level of IL-6 returned to day 0 level shortly after end of treatment, while the TNF-α production remained reduced 10 d after the end of treatment. Our results indicate that enrofloxacin given in vivo in therapeutic doses has an immunomodulatory effect through its capacity to inhibit ex vivo secretion of IL-6 and TNF-α by porcine PBMC after LPS stimulation.

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.

In vitro immunomodulatory effects of cuphiin D1 on human mononuclear cells.

PubMed

Wang, Ching-Chiung; Chen, Lih-Geeng; Yang, Ling-Ling

2002-01-01

Cuphiin D1 (CD1), a macrocyclic hydrolyzable tannin isolated from Cuphea hyssopifolia, has been shown to exert antitumor activity both in vitro and in vivo. Moreover, the antitumor effects of CD1 are not only related to its cytotoxicity to carcinoma cell lines, but also depend on host-mediated mechanisms. In the present study, CD1 was investigated for its effects on the proliferation and cytokine secretion of human peripheral blood mononuclear cells (PBMCs). At concentrations of from 6.25 to 50 micrograms/ml, it enhanced the 3H-thymidine incorporation of concanavalin A (Con A)-stimulated PBMCs in a dose-dependent manner. Excretion of IL-1 beta, IL-2 and TNF-alpha by CD1-stimulated PBMCs was markedly increased in a dose-dependent manner. The results show that CD1 could stimulate PBMCs release of IL-1 beta, IL-2 and TNF-alpha and then activate T cells. Therefore, CD1-activated T cells via IL-1 beta in vitro might account for the host-mediated CD1 mechanism of action.

Differential MicroRNA Analyses of Burkholderia pseudomallei- and Francisella tularensis-Exposed hPBMCs Reveal Potential Biomarkers.

PubMed

Cer, Regina Z; Herrera-Galeano, J Enrique; Frey, Kenneth G; Schully, Kevin L; Luu, Truong V; Pesce, John; Mokashi, Vishwesh P; Keane-Myers, Andrea M; Bishop-Lilly, Kimberly A

2017-01-01

Increasing evidence that microRNAs (miRNAs) play important roles in the immune response against infectious agents suggests that miRNA might be exploitable as signatures of exposure to specific infectious agents. In order to identify potential early miRNA biomarkers of bacterial infections, human peripheral blood mononuclear cells (hPBMCs) were exposed to two select agents, Burkholderia pseudomallei K96243 and Francisella tularensis SHU S4, as well as to the nonpathogenic control Escherichia coli DH5 α . RNA samples were harvested at three early time points, 30, 60, and 120 minutes postexposure, then sequenced. RNAseq analyses identified 87 miRNAs to be differentially expressed (DE) in a linear fashion. Of these, 31 miRNAs were tested using the miScript miRNA qPCR assay. Through RNAseq identification and qPCR validation, we identified differentially expressed miRNA species that may be involved in the early response to bacterial infections. Based upon its upregulation at early time points postexposure in two different individuals, hsa-mir-30c-5p is a miRNA species that could be studied further as a potential biomarker for exposure to these gram-negative intracellular pathogens. Gene ontology functional analyses demonstrated that programmed cell death is the first ranking biological process associated with miRNAs that are upregulated in F. tularensis -exposed hPBMCs.

Differential MicroRNA Analyses of Burkholderia pseudomallei- and Francisella tularensis-Exposed hPBMCs Reveal Potential Biomarkers

PubMed Central

Herrera-Galeano, J. Enrique; Frey, Kenneth G.; Schully, Kevin L.; Luu, Truong V.; Pesce, John; Mokashi, Vishwesh P.; Keane-Myers, Andrea M.; Bishop-Lilly, Kimberly A.

2017-01-01

Increasing evidence that microRNAs (miRNAs) play important roles in the immune response against infectious agents suggests that miRNA might be exploitable as signatures of exposure to specific infectious agents. In order to identify potential early miRNA biomarkers of bacterial infections, human peripheral blood mononuclear cells (hPBMCs) were exposed to two select agents, Burkholderia pseudomallei K96243 and Francisella tularensis SHU S4, as well as to the nonpathogenic control Escherichia coli DH5α. RNA samples were harvested at three early time points, 30, 60, and 120 minutes postexposure, then sequenced. RNAseq analyses identified 87 miRNAs to be differentially expressed (DE) in a linear fashion. Of these, 31 miRNAs were tested using the miScript miRNA qPCR assay. Through RNAseq identification and qPCR validation, we identified differentially expressed miRNA species that may be involved in the early response to bacterial infections. Based upon its upregulation at early time points postexposure in two different individuals, hsa-mir-30c-5p is a miRNA species that could be studied further as a potential biomarker for exposure to these gram-negative intracellular pathogens. Gene ontology functional analyses demonstrated that programmed cell death is the first ranking biological process associated with miRNAs that are upregulated in F. tularensis-exposed hPBMCs. PMID:28791299

Associations of Epstein-Barr Virus DNA in PBMCs and the Subtypes with Breast Cancer Risk

PubMed Central

Zhang, Wei; Wang, Min-Yi; Wei, Xue-Ling; Lin, Ying; Su, Feng-Xi; Xie, Xiao-Ming; Tang, Lu-Ying; Ren, Ze-Fang

2017-01-01

Objective: Epstein-Barr virus (EBV) has been found to be implicated in the development of breast cancer. The purpose of the present study was to identify the associations of EBV DNA and the subtypes in peripheral blood mononuclear cells (PBMCs) with the risk of breast cancer. Material and Methods: A case-control study with 671 breast cancer cases and 859 age-matched controls was conducted in Guangzhou, China. Face-to-face interviews were performed and blood samples were collected immediately after admission to the hospital for patients or after the interview for controls. EBV DNA in PBMCs and the subtypes were detected using Polymerase Chain Reaction (PCR) and restricted fragment length polymorphisms (RFLP). IgA antibodies against EBV VCA-p18 and EBNA-1 were examined using commercial enzyme-linked immunosorbent assay kits. Unconditional logistic regression analysis was applied to evaluate the associations of the DNA positivity and subtypes of EBV with the risk of breast cancer. Results: Among the 1530 subjects, 164 cases (24.4 %) and 206 controls (24.0 %) were positive for EBV DNA in PBMCs and no significant difference occurred between cases and controls. The presence of EBV DNA was related to the positivity of EBV IgA antibodies. Of the DNA positive samples, 71 cases and 109 controls for F/f subtype and 58 cases and 112 controls for C/D subtype were successfully obtained. The D subtype was associated with an increased breast cancer risk compared with the C subtype [OR (95% CI): 2.86 (1.25~6.53)]. We did not find an association of the F/f polymorphism with breast cancer risk. Conclusions: The present study suggested that the presence of EBV DNA in PBMCs may not be an appropriate biomarker for breast cancer risk. The subtype D of EBV was likely to be related to breast tumorigenesis. PMID:28928885

Immunomodulatory activity of a novel, synthetic beta-glucan (β-glu6) in murine macrophages and human peripheral blood mononuclear cells.

PubMed

Li, Xiaofei; Wang, Jing; Wang, Wei; Liu, Chunhong; Sun, Shuhui; Gu, Jianxin; Wang, Xun; Boraschi, Diana; Huang, Yuxian; Qu, Di

2013-01-01

Natural β-glucans extracted from plants and fungi have been used in clinical therapies since the late 20th century. However, the heterogeneity of natural β-glucans limits their clinical applicability. We have synthesized β-glu6, which is an analog of the lentinan basic unit, β-(1→6)-branched β-(1→3) glucohexaose, that contains an α-(1→3)-linked bond. We have demonstrated the stimulatory effect of this molecule on the immune response, but the mechanisms by which β-glu6 activates innate immunity have not been elucidated. In this study, murine macrophages and human PBMCs were used to evaluate the immunomodulatory effects of β-glu6. We showed that β-glu6 activated ERK and c-Raf phosphorylation but suppressed the AKT signaling pathway in murine macrophages. Additionally, β-glu6 enhanced the secretion of large levels of cytokines and chemokines, including CD54, IL-1α, IL-1β, IL-16, IL-17, IL-23, IFN-γ, CCL1, CCL3, CCL4, CCL12, CXCL10, tissue inhibitor of metalloproteinase-1 (TIMP-1) and G-CSF in murine macrophages as well as IL-6, CCL2, CCL3, CCL5, CXCL1 and macrophage migration inhibitory factor (MIF) in human PBMCs. In summary, it demonstrates the immunomodulatory activity of β-glu6 in innate immunity.

Immunomodulatory activity of a plant extract containing human papillomavirus 16-E7 protein in human monocyte-derived dendritic cells.

PubMed

Di Bonito, P; Grasso, F; Mangino, G; Massa, S; Illiano, E; Franconi, R; Fanales-Belasio, E; Falchi, M; Affabris, E; Giorgi, C

2009-01-01

This study reports the immunomodulatory activity on human monocyte derived dendritic cells (MDDCs) of a vaccine preparation shown to be effective against an HPV16-related tumour in an animal model. The vaccine is composed of extract from Nicotiana benthamiana leaves containing HPV16 E7 protein expressed by a potato virus X-derived vector (NbPVX-E7). The effect of the extract was evaluated on MDDC differentiation and maturation by monitoring the phenotypic expression of specific markers. The results show that NbPVX-E7 does not induce monocyte differentiation to dendritic cells, but does induce MDDC maturation. Plant extract does not influence MDDC-uptake of E7-FITC while it significantly improves the Ovalbumin-FITC uptake, considered as a model antigen. Importantly, NbPVX-E7-pulsed MDDCs/PBMCs are able to prime human blood-derived lymphocytes from healthy individuals to induce HPV16 E7-specific cytotoxic activity. This is a propaedeutic study for a possible use of E7-containing plant extract in human immunotherapy of HPV-related lesions.

Human Peripheral Blood Mononuclear Cell Function and Dendritic Cell Differentiation Are Affected by Bisphenol-A Exposure

PubMed Central

Ariemma, Fabiana; Cimmino, Ilaria; Bruzzese, Dario; Scerbo, Roberta; Picascia, Stefania; D’Esposito, Vittoria; Beguinot, Francesco; Formisano, Pietro

2016-01-01

Environmental pollutants, including endocrine disruptor chemicals (EDCs), interfere on human health, leading to hormonal, immune and metabolic perturbations. Bisphenol-A (BPA), a main component of polycarbonate plastics, has been receiving increased attention due to its worldwide distribution with a large exposure. In humans, BPA, for its estrogenic activity, may have a role in autoimmunity, inflammatory and allergic diseases. To this aim, we assessed the effect of low BPA doses on functionality of human peripheral blood mononuclear cells (PBMCs), and on in vitro differentiation of dendritic cells from monocytes (mDCs). Fresh peripheral blood samples were obtained from 12 healthy adult volunteers. PBMCs were left unstimulated or were activated with the mitogen phytohemagglutinin (PHA) or the anti-CD3 and anti-CD28 antibodies and incubated in presence or absence of BPA at 0.1 and 1nM concentrations. The immune-modulatory effect of BPA was assessed by evaluating the cell proliferation and the levels of interferon-γ (IFN-γ), interleukin-4 (IL-4), interleukin-10 (IL-10) and interleukin-13 (IL-13) secreted by PBMCs. mDCs were differentiated with IL-4 and GC-CSF with or without BPA and the expression of differentiation/maturation markers (CD11c, CD1a, CD86, HLA-DR) was evaluated by flow cytometry; furthermore, a panel of 27 different cytokines, growth factors and chemokines were assayed in the mDC culture supernatants. PBMCs proliferation significantly increased upon BPA exposure compared to BPA untreated cells. In addition, a significant decrease in IL-10 secretion was observed in PBMCs incubated with BPA, either in unstimulated or mitogen-stimulated cells, and at both 0.1 and 1nM BPA concentrations. Similarly, IL-13 was reduced, mainly in cells activated by antiCD3/CD28. By contrast, no significant changes in IFN-γ and IL-4 production were found in any condition assayed. Finally, BPA at 1nM increased the density of dendritic cells expressing CD1a and concomitantly

TRIBUTYLTIN ALTERS SECRETION OF INTERLEUKIN 1 BETA FROM HUMAN IMMUNE CELLS

PubMed Central

Brown, Shyretha; Whalen, Margaret

2014-01-01

Tributyltin (TBT) has been used as a biocide in industrial applications such as wood preservation, antifouling paint, and antifungal agents. Due to its many uses, it contaminates the environment and has been found in human blood samples. Interleukin 1 beta (IL-1β) is a pro-inflammatory cytokine that promotes cell growth, tissue repair, and immune response regulation. Produced predominately by both monocytes and macrophages, IL-1β appears to increase the invasiveness of certain tumors. This study shows that TBT modifies the secretion of IL-1β from increasingly reconstituted preparations of human immune cells. IL-1β secretion was examined after 24h, 48h, or 6 day exposures to TBT in highly enriched human NK cells, monocyte-depleted (MD) peripheral blood mononuclear cells (MD-PBMCs), PBMCs, granulocytes, and a preparation combining both PBMCs and granulocytes (PBMCs+granulocytes). TBT altered IL-1β secretion from all of the cells preparations. The 200 nM concentration of TBT normally blocked the secretion of IL-1β, while lower concentrations (usually 5-50 nM) elevated secretion of IL-1β. Examination of the signaling pathway(s) responsible for the elevated secretion of IL-1β were carried out in MD-PBMCs. Pathways examined were IL-1β processing (Caspase-1), mitogen-activated protein kinases (MAPKs), and nuclear factor kappa B (NFκB). Results indicated that MAPK pathways (p44/42 and p38) appear to be the targets of TBT that lead to increased IL-1β secretion from immune cells. These results from human immune cells show IL-1β dysregulation by TBT is occurring ex vivo. Thus, potential for in vivo effects on pro-inflammatory cytokine levels may possibly be a consequence of TBT exposures. PMID:25382723

Tributyltin alters secretion of interleukin 1 beta from human immune cells.

PubMed

Brown, Shyretha; Whalen, Margaret

2015-08-01

Tributyltin (TBT) has been used as a biocide in industrial applications such as wood preservation, antifouling paint and antifungal agents. Owing to its many uses, it contaminates the environment and has been found in human blood samples. Interleukin-1 beta (IL-1β) is a pro-inflammatory cytokine that promotes cell growth, tissue repair and immune response regulation. Produced predominately by both monocytes and macrophages, IL-1β appears to increase the invasiveness of certain tumors. This study shows that TBT modifies the secretion of IL-1β from increasingly reconstituted preparations of human immune cells. IL-1β secretion was examined after 24-, 48-h or 6-day exposures to TBT in highly enriched human natural killer (NK) cells, monocyte-depleted peripheral blood mononuclear cells (MD-PBMCs), PBMCs, granulocytes and a preparation combining both PBMCs and granulocytes (PBMCs+granulocytes). TBT altered IL-1β secretion from all of the cell preparations. The 200 nM concentration of TBT normally blocked the secretion of IL-1β, whereas lower concentrations (usually 5-50 nM) elevated secretion of IL-1β. Examination of the signaling pathway(s) responsible for the elevated secretion of IL-1β was carried out in MD-PBMCs. Pathways examined were IL-1β processing (Caspase-1), mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NFκB). Results indicated that MAPK pathways (p44/42 and p38) appear to be the targets of TBT that lead to increased IL-1β secretion from immune cells. These results from human immune cells show IL-1β dysregulation by TBT is occurring ex vivo. Thus, the potential for in vivo effects on pro-inflammatory cytokine levels may possibly be a consequence of TBT exposures. Copyright © 2014 John Wiley & Sons, Ltd.

Elimination of human T cell leukemia virus type-1-infected cells by neutralizing and antibody-dependent cellular cytotoxicity-inducing antibodies against human t cell leukemia virus type-1 envelope gp46.

PubMed

Tanaka, Yuetsu; Takahashi, Yoshiaki; Tanaka, Reiko; Kodama, Akira; Fujii, Hideki; Hasegawa, Atsuhiko; Kannagi, Mari; Ansari, Aftab A; Saito, Mineki

2014-06-01

Human T cell leukemia virus type-1 (HTLV-1) is prevalent worldwide with foci of high prevalence. However, to date no effective vaccine or drug against HTLV-1 infection has been developed. In efforts to define the role of antibodies in the control of HTLV-1 infection, we capitalized on the use of our previously defined anti-gp46 neutralizing monoclonal antibody (mAb) (clone LAT-27) and high titers of human anti-HTLV-1 IgG purified from HAM/TSP patients (HAM-IgG). LAT-27 and HAM-IgG completely blocked syncytium formation and T cell immortalization mediated by HTLV-1 in vitro. The addition of these antibodies to cultures of CD8(+) T cell-depleted peripheral blood mononuclear cells (PBMCs) from HAM/TSP patients at the initiation of culture not only decreased the numbers of Tax-expressing cells and the production of HTLV-1 p24 but also inhibited the spontaneous immortalization of T cells. Coculture of in vitro-HTLV-1-immortalized T cell lines with autologous PBMCs in the presence of LAT-27 or HAM-IgG, but not an F(ab')2 fragment of LAT-27 or nonneutralizing anti-gp46 mAbs, resulted in depletion of HTLV-1-infected cells. A 24-h (51)Cr release assay showed the presence of significant antibody-dependent cellular cytotoxicity (ADCC) activity in LAT-27 and HAM-IgG, but not F(ab')2 of LAT-27, resulting in the depletion of HTLV-1-infected T cells by autologous PBMCs. The depletion of natural killer (NK) cells from the effector PBMCs reduced this ADCC activity. Altogether, the present data demonstrate that the neutralizing and ADCC-inducing activities of anti-HTLV-1 antibodies are capable of reducing infection and eliminating HTLV-1-infected cells in the presence of autologous PBMCs.

Irradiated and activated autologous PBMCs induce expansion of highly cytotoxic human NK cells in vitro.

PubMed

Ahn, Yong-Oon; Kim, Saerom; Kim, Tae Min; Song, Eun Young; Park, Myoung Hee; Heo, Dae Seog

2013-09-01

Adoptive cell transfer of ex vivo-activated natural killer (NK) cells is a promising therapy for cancer treatment. Because of inhibitory signaling through killer immunoglobulin-like receptor (KIR)-KIR ligands, KIR-mismatched allogeneic NK cell transfer is considered to be a more effective strategy than is autologous transfer. However, purified NK cells do not expand well enough in vitro with good manufacturing practice-compliant components for clinical use. Some investigators have developed selective expansion of NK cells from peripheral blood mononuclear cells, but these cells have the risk of graft-versus-host disease in allogeneic settings because of T cells contamination. In this study, we developed a novel method for NK cell activation and expansion. Using only good manufacturing practice-compliant components and autologous feeder cells, once purified NK cells were effectively expanded (2500-fold at day 17). The expanded cells were highly purified NK cells, and the use of these cells is suitable for allogeneic transfer without the risk of graft-versus-host disease induction. Importantly, the expanded NK cells also showed enhanced cytotoxicity compared with NK cells conventionally expanded by recombinant human interleukin 2. Finally, induction of NKG2D ligand expression on feeder cells implies that the NKG2D-NKG2DL interaction may play a role in NK cell expansion. In conclusion, this method can be used to obtain NK cells for more successful allogeneic NK cell adoptive transfer for use in antitumor immune therapy.

Aspergillus Cell Wall Chitin Induces Anti- and Proinflammatory Cytokines in Human PBMCs via the Fc-γ Receptor/Syk/PI3K Pathway.

PubMed

Becker, K L; Aimanianda, V; Wang, X; Gresnigt, M S; Ammerdorffer, A; Jacobs, C W; Gazendam, R P; Joosten, L A B; Netea, M G; Latgé, J P; van de Veerdonk, F L

2016-05-31

Chitin is an important cell wall component of Aspergillus fumigatus conidia, of which hundreds are inhaled on a daily basis. Previous studies have shown that chitin has both anti- and proinflammatory properties; however the exact mechanisms determining the inflammatory signature of chitin are poorly understood, especially in human immune cells. Human peripheral blood mononuclear cells were isolated from healthy volunteers and stimulated with chitin from Aspergillus fumigatus Transcription and production of the proinflammatory cytokine interleukin-1β (IL-1β) and the anti-inflammatory cytokine IL-1 receptor antagonist (IL-1Ra) were measured from the cell culture supernatant by quantitative PCR (qPCR) or enzyme-linked immunosorbent assay (ELISA), respectively. Chitin induced an anti-inflammatory signature characterized by the production of IL-1Ra in the presence of human serum, which was abrogated in immunoglobulin-depleted serum. Fc-γ-receptor-dependent recognition and phagocytosis of IgG-opsonized chitin was identified as a novel IL-1Ra-inducing mechanism by chitin. IL-1Ra production induced by chitin was dependent on Syk kinase and phosphatidylinositol 3-kinase (PI3K) activation. In contrast, costimulation of chitin with the pattern recognition receptor (PRR) ligands lipopolysaccharide, Pam3Cys, or muramyl dipeptide, but not β-glucan, had synergistic effects on the induction of proinflammatory cytokines by human peripheral blood mononuclear cells (PBMCs). In conclusion, chitin can have both pro- and anti-inflammatory properties, depending on the presence of pathogen-associated molecular patterns and immunoglobulins, thus explaining the various inflammatory signatures reported for chitin. Invasive aspergillosis and allergic aspergillosis are increasing health care problems. Patients get infected by inhalation of the airborne spores of Aspergillus fumigatus A profound knowledge of how Aspergillus and its cell wall components are recognized by the host cell and

Nitric oxide affects IL-6 expression in human peripheral blood mononuclear cells involving cGMP-dependent modulation of NF-κB activity.

PubMed

Siednienko, Jakub; Nowak, Joanna; Moynagh, Paul N; Gorczyca, Wojciech A

2011-06-01

Interleukin 6 (IL-6) and nitric oxide (NO) are important mediators of the inflammatory response. We report that in human peripheral blood mononuclear cells (PBMCs), NO exerts a biphasic effect on the expression of IL-6. Using sodium nitroprusside (SNP) and S-nitrosoglutathione (GSNO) as NO-donating compounds, we observed that both mRNA and protein levels of IL-6 increased at lower (≤10μM) and decreased at higher (>100μM) concentrations of NO donors. Changes in the expression of IL-6 correlated with changes in the activity of NF-κB, which increased at lower and decreased at higher concentrations of both NO donors as shown by the electrophoretic mobility shift assay (EMSA). The effects of NO on NF-κB activity were cGMP-dependent because they were reversed in the presence of ODQ, the inhibitor of soluble guanylyl cyclase (sGC), and KT5823, the inhibitor of cGMP-dependent protein kinase (PKG). Moreover, the membrane permeable analog of cGMP (8-Br-cGMP) mimicked the effect of the NO donors. These observations show that NO, depending on its concentration, may act in human PBMCs as a stimulator of IL-6 expression involving the sGC/cGMP/PKG pathway. Copyright © 2011 Elsevier Ltd. All rights reserved.

Human umbilical cord-derived mesenchymal stem cells suppress proliferation of PHA-activated lymphocytes in vitro by inducing CD4(+)CD25(high)CD45RA(+) regulatory T cell production and modulating cytokine secretion.

PubMed

Yang, Hongna; Sun, Jinhua; Li, Yan; Duan, Wei-Ming; Bi, Jianzhong; Qu, Tingyu

2016-04-01

Bone marrow-derived mesenchymal stem cells (MSCs) are promising candidate cells for therapeutic application in autoimmune diseases due to their immunomodulatory properties. Unused human umbilical cords (UC) offer an abundant and noninvasive source of MSCs without ethical issues and are emerging as a valuable alternative to bone marrow tissue for producing MSCs. We thus investigated the immunomodulation effect of umbilical cord-derived MSCs (UC-MSCs) on human peripheral blood mononuclear cells (PBMCs), T cells in particular, in a co-culture system. We found that UC-MSCs efficiently suppressed the proliferation of phytohaemagglutinin (PHA)-stimulated PBMCs (p<0.01). Kinetic analysis revealed that UC-MSCs primarily inhibited the division of generation 3 (G3) and 4 (G4) of PBMCs. In addition, UC-MSCs augmented the expression of CD127(+) and CD45RA(+) but reduced the expression of CD25(+) in PBMCs stimulated by PHA (p<0.05). Furthermore, UC-MSCs inhibited PHA-resulted increase in the frequency of CD4(+)CD25(+)CD127(low/-) Tregs significantly (p<0.01) but augmented PHA-resulted increase in the frequency of CD4(+)CD25(high)CD45RA(+) Tregs to about three times in PBMCs. The levels of anti-inflammatory cytokines, PEG2, TGF-β, and IL-10 were greatly up-regulated, accompanied by a significant down-regulation of pro-inflammatory IFN-γ in the co-culture (p<0.01). Our results showed that UC-MSCs are able to suppress mitogen-induced PBMC activation and proliferation in vitro by altering T lymphocyte phenotypes, increasing the frequency of CD4(+)CD25(high)CD45RA(+) Tregs, and modulating the associated cytokine production. Further studies are warranted to investigate the therapeutic potential of UC-MSCs in immunologically-diseased conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

AMPA, NMDA and kainate glutamate receptor subunits are expressed in human peripheral blood mononuclear cells (PBMCs) where the expression of GluK4 is altered by pregnancy and GluN2D by depression in pregnant women.

PubMed

Bhandage, Amol K; Jin, Zhe; Hellgren, Charlotte; Korol, Sergiy V; Nowak, Krzysztof; Williamsson, Louise; Sundström-Poromaa, Inger; Birnir, Bryndis

2017-04-15

The amino acid glutamate opens cation permeable ion channels, the iGlu receptors. These ion channels are abundantly expressed in the mammalian brain where glutamate is the main excitatory neurotransmitter. The neurotransmitters and their receptors are being increasingly detected in the cells of immune system. Here we examined the expression of the 18 known subunits of the iGlu receptors families; α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), kainate, N-methyl-d-aspartate (NMDA) and delta in human peripheral blood mononuclear cells (PBMCs). We compared the expression of the subunits between four groups: men, non-pregnant women, healthy pregnant women and depressed pregnant women. Out of 18 subunits of the iGlu receptors, mRNAs for 11 subunits were detected in PBMCs from men and non-pregnant women; AMPA: GluA3, GluA4, kainate: GluK2, GluK4, GluK5, NMDA: GluN1, GluN2C, GluN2D, GluN3A, GluN3B, and delta: GluD1. In the healthy and the depressed pregnant women, in addition, the delta GluD2 subunit was identified. The mRNAs for GluK4, GluK5, GluN2C and GluN2D were expressed at a higher level than other subunits. Gender, pregnancy or depression during pregnancy altered the expression of GluA3, GluK4, GluN2D, GluN3B and GluD1 iGlu subunit mRNAs. The greatest changes recorded were the lower GluA3 and GluK4 mRNA levels in pregnant women and the higher GluN2D mRNA level in healthy but not in depressed pregnant women as compared to non-pregnant individuals. Using subunit specific antibodies, the GluK4, GluK5, GluN1, GluN2C and GluN2D subunit proteins were identified in the PBMCs. The results show expression of specific iGlu receptor subunit in the PBMCs and support the idea of physiology-driven changes of iGlu receptors subtypes in the immune cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Gc protein-derived macrophage-activating factor (GcMAF) stimulates cAMP formation in human mononuclear cells and inhibits angiogenesis in chick embryo chorionallantoic membrane assay.

PubMed

Pacini, Stefania; Morucci, Gabriele; Punzi, Tiziana; Gulisano, Massimo; Ruggiero, Marco

2011-04-01

The effects of Gc protein-derived macrophage-activating factor (GcMAF) have been studied in cancer and other conditions where angiogenesis is deregulated. In this study, we demonstrate for the first time that the mitogenic response of human peripheral blood mononuclear cells (PBMCs) to GcMAF was associated with 3'-5'-cyclic adenosine monophosphate (cAMP) formation. The effect was dose dependent, and maximal stimulation was achieved using 0.1 ng/ml. Heparin inhibited the stimulatory effect of GcMAF on PBMCs. In addition, we demonstrate that GcMAF (1 ng/ml) inhibited prostaglandin E(1)- and human breast cancer cell-stimulated angiogenesis in chick embryo chorionallantoic membrane (CAM) assay. Finally, we tested different GcMAF preparations on CAM, and the assay proved to be a reliable, reproducible and inexpensive method to determine the relative potencies of different preparations and their stability; we observed that storage at room temperature for 15 days decreased GcMAF potency by about 50%. These data could prove useful for upcoming clinical trials on GcMAF.

Changes in gene expression in PBMCs profiles of PPARα target genes in obese and non-obese individuals during fasting.

PubMed

Felicidade, Ingrid; Marcarini, Juliana Cristina; Carreira, Clísia Mara; Amarante, Marla Karine; Afman, Lydia A; Mantovani, Mário Sérgio; Ribeiro, Lúcia Regina

2015-01-01

The prevalence of obesity has risen dramatically and the World Health Organization estimates that 700 million people will be obese worldwide by 2015. Approximately, 50% of the Brazilian population above 20 years of age is overweight, and 16% is obese. This study aimed to evaluate the differences in the expression of PPARα target genes in human peripheral blood mononuclear cells (PBMCs) and free fatty acids (FFA) in obese and non-obese individuals after 24 h of fasting. We first presented evidence that Brazilian people exhibit expression changes in PPARα target genes in PBMCs under fasting conditions. Q-PCR was utilized to assess the mRNA expression levels of target genes. In both groups, the FFA concentrations increased significantly after 24 h of fasting. The basal FFA mean concentration was two-fold higher in the obese group compared with the non-obese group. After fasting, all genes evaluated in this study showed increased expression levels compared with basal expression in both groups. However, our results reveal no differences in gene expression between the obese and non-obese, more studies are necessary to precisely delineate the associated mechanisms, particularly those that include groups with different degrees of obesity and patients with diabetes mellitus type 2 because the expression of the main genes that are involved in β-oxidation and glucose level maintenance are affected by these factors. © 2014 S. Karger AG, Basel.

Evaluation of HTLV-1 activity in HAM/TSP patients using proviral load and Tax mRNA expression after In Vitro lymphocyte activation.

PubMed

Yari, Atefeh; Rezaee, Seyyed Abdolrahim; Valizadeh, Narges; Rajaee, Taraneh; Jazayeri, Seyyed Mohammad; Soltani, Mojdeh; Norouzi, Mehdi

2014-07-01

HTLV-1 is the first human retrovirus that has been recognized and is associated with HAM/TSP and ATLL. Studies have shown that less than five percent of HTLV-1 infected carriers develop HAM/TSP or ATLL and about ninety-five percent remain asymptomatic. Therefore, the proviral load with Tax may affect cellular genes such as cytokines and oncogenes, as well as involve in pathogenicity. Thirty HAM/TSP patients, thirty HTLV-1 healthy carriers, and MT-2 cell line were evaluated for HTLV-1 activity. PBMCs were isolated and activated using PMA and ionomycine. Real-time PCR and TaqMan methods were performed using specific primers and fluorescence probes for Tax expression and proviral load assessment. B2microglobulin (β2m) and albumin were used as controls in Tax expression and in proviral load, respectively. An insignificant increase in Tax expression was observed in rest PBMCs of HAM/TSP patients compared to healthy carriers. However, after lymphocyte activation there was a significant increase in Tax expression in HAM/TSP patients (P=0.042). The Proviral load in patients was significantly higher than in carriers. Moreover, there was a significant correlation between Tax mRNA expression in activated PBMCs and proviral load (R=0.37, P=0.012). Although proviral load had been addressed as a valuable index for monitoring HTLV-1 infected subjects, the results of this study demonstrated that Tax expression in activated PBMCs along with proviral load assessment in HAM/TSP patients are a more reliable factor for determining the prognosis and monitoring healthy carriers and HAM/TSP patients.

Decreased epigenetic age of PBMCs from Italian semi-supercentenarians and their offspring.

PubMed

Horvath, Steve; Pirazzini, Chiara; Bacalini, Maria Giulia; Gentilini, Davide; Di Blasio, Anna Maria; Delledonne, Massimo; Mari, Daniela; Arosio, Beatrice; Monti, Daniela; Passarino, Giuseppe; De Rango, Francesco; D'Aquila, Patrizia; Giuliani, Cristina; Marasco, Elena; Collino, Sebastiano; Descombes, Patrick; Garagnani, Paolo; Franceschi, Claudio

2015-12-01

Given the dramatic increase in ageing populations, it is of great importance to understand the genetic and molecular determinants of healthy ageing and longevity. Semi-supercentenarians (subjects who reached an age of 105-109 years) arguably represent the gold standard of successful human ageing because they managed to avoid or postpone the onset of major age-related diseases. Relatively few studies have looked at epigenetic determinants of extreme longevity in humans. Here we test whether families with extreme longevity are epigenetically distinct from controls according to an epigenetic biomarker of ageing which is known as "epigenetic clock". We analyze the DNA methylation levels of peripheral blood mononuclear cells (PBMCs) from Italian families constituted of 82 semi-supercentenarians (mean age: 105.6 ± 1.6 years), 63 semi-supercentenarians' offspring (mean age: 71.8 ± 7.8 years), and 47 age-matched controls (mean age: 69.8 ± 7.2 years). We demonstrate that the offspring of semi-supercentenarians have a lower epigenetic age than age-matched controls (age difference=5.1 years, p=0.00043) and that centenarians are younger (8.6 years) than expected based on their chronological age. By contrast, no significant difference could be observed for estimated blood cell counts (such as naïve or exhausted cytotoxic T cells or helper T cells). Future studies will be needed to replicate these findings in different populations and to extend them to other tissues. Overall, our results suggest that epigenetic processes might play a role in extreme longevity and healthy human ageing.

Decreased epigenetic age of PBMCs from Italian semi-supercentenarians and their offspring

PubMed Central

Horvath, Steve; Pirazzini, Chiara; Bacalini, Maria Giulia; Gentilini, Davide; Di Blasio, Anna Maria; Delledonne, Massimo; Mari, Daniela; Arosio, Beatrice; Monti, Daniela; Passarino, Giuseppe; De Rango, Francesco; D'Aquila, Patrizia; Giuliani, Cristina; Marasco, Elena; Collino, Sebastiano; Descombes, Patrick; Garagnani, Paolo; Franceschi, Claudio

2015-01-01

Given the dramatic increase in ageing populations, it is of great importance to understand the genetic and molecular determinants of healthy ageing and longevity. Semi-supercentenarians (subjects who reached an age of 105-109 years) arguably represent the gold standard of successful human ageing because they managed to avoid or postpone the onset of major age-related diseases. Relatively few studies have looked at epigenetic determinants of extreme longevity in humans. Here we test whether families with extreme longevity are epigenetically distinct from controls according to an epigenetic biomarker of ageing which is known as “epigenetic clock”. We analyze the DNA methylation levels of peripheral blood mononuclear cells (PBMCs) from Italian families constituted of 82 semi-supercentenarians (mean age: 105.6 ± 1.6 years), 63 semi-supercentenarians' offspring (mean age: 71.8 ± 7.8 years), and 47 age-matched controls (mean age: 69.8 ± 7.2 years). We demonstrate that the offspring of semi-supercentenarians have a lower epigenetic age than age-matched controls (age difference=5.1 years, p=0.00043) and that centenarians are younger (8.6 years) than expected based on their chronological age. By contrast, no significant difference could be observed for estimated blood cell counts (such as naïve or exhausted cytotoxic T cells or helper T cells). Future studies will be needed to replicate these findings in different populations and to extend them to other tissues. Overall, our results suggest that epigenetic processes might play a role in extreme longevity and healthy human ageing. PMID:26678252

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 ...

Human mesenchymal stromal cells transiently increase cytokine production by activated T cells before suppressing T-cell proliferation: effect of interferon-γ and tumor necrosis factor-α stimulation.

PubMed

Cuerquis, Jessica; Romieu-Mourez, Raphaëlle; François, Moïra; Routy, Jean-Pierre; Young, Yoon Kow; Zhao, Jing; Eliopoulos, Nicoletta

2014-02-01

Mesenchymal stromal cells (MSCs) suppress T-cell proliferation, especially after activation with inflammatory cytokines. We compared the dynamic action of unprimed and interferon (IFN)-γ plus tumor necrosis factor (TNF)-α-pretreated human bone marrow-derived MSCs on resting or activated T cells. MSCs were co-cultured with allogeneic peripheral blood mononuclear cells (PBMCs) at high MSC-to-PBMC ratios in the absence or presence of concomitant CD3/CD28-induced T-cell activation. The kinetic effects of MSCs on cytokine production and T-cell proliferation, cell cycle and apoptosis were assessed. Unprimed MSCs increased the early production of IFN-γ and interleukin (IL)-2 by CD3/CD28-activated PBMCs before suppressing T-cell proliferation. In non-activated PBMC co-cultures, low levels of IL-2 and IL-10 synthesis were observed with MSCs in addition to low levels of CD69 expression by T cells and no T-cell proliferation. MSCs also decreased apoptosis in resting and activated T cells and inhibited the transition of these cells into the sub-G0/G1 and the S phases. With inhibition of indoleamine 2,3 dioxygenase, MSCs increased CD3/CD28-induced T-cell proliferation. After priming with IFN-γ plus TNF-α, MSCs were less potent at increasing cytokine production by CD3/CD28-activated PBMCs and more effective at inhibiting T-cell proliferation but had preserved anti-apoptotic functions. Unprimed MSCs induce a transient increase in IFN-γ and IL-2 synthesis by activated T cells. Pre-treatment of MSCs with IFN-γ plus TNF-α may increase their effectiveness and safety in vivo. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

50 Hz sinusoidal magnetic fields do not affect human lymphocyte activation and proliferation in vitro

NASA Astrophysics Data System (ADS)

Capri, Miriam; Mesirca, Pietro; Remondini, Daniel; Carosella, Simona; Pasi, Sara; Castellani, Gastone; Franceschi, Claudio; Bersani, Ferdinando

2004-12-01

In the last 30 years, an increasing public concern about the possible harmful effects of electromagnetic fields generated by power lines and domestic appliances has pushed the scientific community to search for a correct and comprehensive answer to this problem. In this work the effects of exposure to 50 Hz sinusoidal magnetic fields, with a magnetic flux density of 0.05 mT and 2.5 mT (peak values), were studied on human peripheral blood mononuclear cells (PBMCs) collected from healthy young and elderly donors. Cell activation and proliferation were investigated by using flow cytometry techniques and 3H-TdR incorporation assays, respectively. The results obtained indicated that exposure to the fields altered neither DNA synthesis nor the capacity of lymphocytes to enter the activation phase and progress into the cell cycle. Thus, the conclusions are that two important functional phases of human lymphocytes, such as activation and proliferation, are not affected by exposures to 50 Hz magnetic fields similar to those found under power lines.

Aptamer-conjugated live human immune cell based biosensors for the accurate detection of C-reactive protein

NASA Astrophysics Data System (ADS)

Hwang, Jangsun; Seo, Youngmin; Jo, Yeonho; Son, Jaewoo; Choi, Jonghoon

2016-10-01

C-reactive protein (CRP) is a pentameric protein that is present in the bloodstream during inflammatory events, e.g., liver failure, leukemia, and/or bacterial infection. The level of CRP indicates the progress and prognosis of certain diseases; it is therefore necessary to measure CRP levels in the blood accurately. The normal concentration of CRP is reported to be 1-3 mg/L. Inflammatory events increase the level of CRP by up to 500 times; accordingly, CRP is a biomarker of acute inflammatory disease. In this study, we demonstrated the preparation of DNA aptamer-conjugated peripheral blood mononuclear cells (Apt-PBMCs) that specifically capture human CRP. Live PBMCs functionalized with aptamers could detect different levels of human CRP by producing immune complexes with reporter antibody. The binding behavior of Apt-PBMCs toward highly concentrated CRP sites was also investigated. The immune responses of Apt-PBMCs were evaluated by measuring TNF-alpha secretion after stimulating the PBMCs with lipopolysaccharides. In summary, engineered Apt-PBMCs have potential applications as live cell based biosensors and for in vitro tracing of CRP secretion sites.

High concentration of branched-chain amino acids promotes oxidative stress, inflammation and migration of human peripheral blood mononuclear cells via mTORC1 activation.

PubMed

Zhenyukh, Olha; Civantos, Esther; Ruiz-Ortega, Marta; Sánchez, Maria Soledad; Vázquez, Clotilde; Peiró, Concepción; Egido, Jesús; Mas, Sebastián

2017-03-01

Leucine, isoleucine and valine are essential aminoacids termed branched-chain amino acids (BCAA) due to its aliphatic side-chain. In several pathological and physiological conditions increased BCAA plasma concentrations have been described. Elevated BCAA levels predict insulin resistance development. Moreover, BCAA levels higher than 2mmol/L are neurotoxic by inducing microglial activation in maple syrup urine disease. However, there are no studies about the direct effects of BCAA in circulating cells. We have explored whether BCAA could promote oxidative stress and pro-inflammatory status in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors. In cultured PBMCs, 10mmol/L BCAA increased the production of reactive oxygen species (ROS) via both NADPH oxidase and the mitochondria, and activated Akt-mTOR signalling. By using several inhibitors and activators of these molecular pathways we have described that mTOR activation by BCAA is linked to ROS production and mitochondrial dysfunction. BCAA stimulated the activation of the redox-sensitive transcription factor NF-κB, which resulted in the release of pro-inflammatory molecules, such as interleukin-6, tumor necrosis factor-α, intracellular adhesion molecule-1 or CD40L, and the migration of PBMCs. In conclusion, elevated BCAA blood levels can promote the activation of circulating PBMCs, by a mechanism that involving ROS production and NF-κB pathway activation. These data suggest that high concentrations of BCAA could exert deleterious effects on circulating blood cells and therefore contribute to the pro-inflammatory and oxidative status observed in several pathophysiological conditions. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

A novel recombinantly produced banana lectin isoform is a valuable tool for glycoproteomics and a potent modulator of the proliferation response in CD3+, CD4+, and CD8+ populations of human PBMCs.

PubMed

Gavrovic-Jankulovic, Marija; Poulsen, Knud; Brckalo, Tamara; Bobic, Sonja; Lindner, Buko; Petersen, Arnd

2008-01-01

Lectins as carbohydrate-binding proteins have been employed in various biological assays for the detection and characterization of glycan structures on glycoproteins, including clinical biomarkers in disease states. A mannose-specific banana lectin (BanLec) is unique in its specificity for internal alpha1,3 linkages as well as beta1,3 linkages at the reducing termini. The immunomodulatory potential of natural BanLec was recognized by a strong immunoglobulin G4 antibody response and T cell mitogen activity in humans. To explore its applicability in glycoproteomics and its modulatory potential, the gene of banana lectin was cloned, sequenced and a recombinant protein was produced in Escherichia coli. The obtained cDNA revealed a novel banana lectin isoform, with an open reading frame of 426 nucleotides, encoding a cytoplasmatic protein of 141 amino acids. The molecular mass of rBanLec determined by ESI FT-MS and N-terminal sequencing confirmed the cDNA at the protein level. The specificity of rBanLec for detection glycan structures was the same as for natural BanLec as examined with five protein extracts rich in glycoprotein content, as well as with horseradish peroxidase glycoprotein. Besides, the immunomodulatory potential of rBanLec and nBanLec were comparable as assessed by an inhibition assay and a human T cell proliferation assay where they induced a strong proliferation response in CD3+, CD4+, and CD8+ populations of human PBMCs. This recombinant BanLec is a useful reagent for glycoproteomics and lectin microarrays, with a potential for modulation of the immune response.

Human innate responses and adjuvant activity of TLR ligands in vivo in mice reconstituted with a human immune system.

PubMed

Cheng, Liang; Zhang, Zheng; Li, Guangming; Li, Feng; Wang, Li; Zhang, Liguo; Zurawski, Sandra M; Zurawski, Gerard; Levy, Yves; Su, Lishan

2017-10-27

TLR ligands (TLR-Ls) represent a class of novel vaccine adjuvants. However, their immunologic effects in humans remain poorly defined in vivo. Using a humanized mouse model with a functional human immune system, we investigated how different TLR-Ls stimulated human innate immune response in vivo and their applications as vaccine adjuvants for enhancing human cellular immune response. We found that splenocytes from humanized mice showed identical responses to various TLR-Ls as human PBMCs in vitro. To our surprise, various TLR-Ls stimulated human cytokines and chemokines differently in vivo compared to that in vitro. For example, CpG-A was most efficient to induce IFN-α production in vitro. In contrast, CpG-B, R848 and Poly I:C stimulated much more IFN-α than CpG-A in vivo. Importantly, the human innate immune response to specific TLR-Ls in humanized mice was different from that reported in C57BL/6 mice, but similar to that reported in nonhuman primates. Furthermore, we found that different TLR-Ls distinctively activated and mobilized human plasmacytoid dendritic cells (pDCs), myeloid DCs (mDCs) and monocytes in different organs. Finally, we showed that, as adjuvants, CpG-B, R848 and Poly I:C can all enhance antigen specific CD4 + T cell response, while only R848 and Poly I:C induced CD8 + cytotoxic T cells response to a CD40-targeting HIV vaccine in humanized mice, correlated with their ability to activate human mDCs but not pDCs. We conclude that humanized mice serve as a highly relevant model to evaluate and rank the human immunologic effects of novel adjuvants in vivo prior to testing in humans. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Metabolic Biofuel Cell: Conversion of Human Leukocyte Metabolic Activity to Electrical Currents

PubMed Central

2011-01-01

An investigation of the electrochemical activity of human white blood cells (WBC) for biofuel cell (BFC) applications is described. WBCs isolated from whole human blood were suspended in PBS and introduced into the anode compartment of a proton exchange membrane (PEM) fuel cell. The cathode compartment contained a 50 mM potassium ferricyanide solution. Average current densities between 0.9 and 1.6 μA cm-2 and open circuit potentials (Voc) between 83 and 102 mV were obtained, which were both higher than control values. Cyclic voltammetry was used to investigate the electrochemical activity of the activated WBCs in an attempt to elucidate the mechanism of electron transfer between the cells and electrode. Voltammograms were obtained for the WBCs, including peripheral blood mononuclear cells (PBMCs - a lymphocyte-monocyte mixture isolated on a Ficoll gradient), a B lymphoblastoid cell line (BLCL), and two leukemia cell lines, namely K562 and Jurkat. An oxidation peak at about 363 mV vs. SCE for the PMA (phorbol ester) activated primary cells, with a notable absence of a reduction peak was observed. Oxidation peaks were not observed for the BLCL, K562 or Jurkat cell lines. HPLC confirmed the release of serotonin (5-HT) from the PMA activated primary cells. It is believed that serotonin, among other biochemical species released by the activated cells, contributes to the observed BFC currents. PMID:21569243

Activation of lysosomal cathepsins in pregnant bovine leukocytes.

PubMed

Talukder, Md Abdus Shabur; Balboula, Ahmed Zaky; Shirozu, Takahiro; Kim, Sung Woo; Kunii, Hiroki; Suzuki, Toshiyuki; Ito, Tsukino; Kimura, Koji; Takahashi, Masashi

2018-06-01

In ruminants, interferon-tau (IFNT) - mediated expression of interferon-stimulated genes in peripheral blood leukocytes (PBLs) can indicate pregnancy. Recently, type 1 IFN-mediated activation of lysosomes and lysosomal cathepsins (CTSs) was observed in immune cells. This study investigated the status of lysosomal CTSs and lysosomes in PBLs collected from pregnant (P) and non-pregnant (NP) dairy cows, and conducted in vitro IFNT stimulation of NP blood leukocytes. Blood samples were collected 0, 7, 14 and 18 days post-artificial insemination, and the peripheral blood mononuclear cells (PBMCs) and polymorphonuclear granulocytes (PMNs) separated. The fluorescent activity of CTSB and CTSK in PMNs significantly increased with the progress of pregnancy, especially on day 18. In vitro supplementation of IFNT significantly increased the activities of CTSB and CTSK in NP PBMCs and PMNs. CTSB expression was significantly higher in PBMCs and PMNs collected from P day-18 cows than from NP cows, whereas there was no difference in CTSK expression. IFNT increased CTSB expression but did not affect CTSK expression. Immunodetection showed an increase of CTSB in P day-18 PBMCs and PMNs. In vitro stimulation of IFNT increased CTSB in NP PBMCs and PMNs. Lysosomal acidification showed a significant increase in P day-18 PBMCs and PMNs. IFNT also stimulated lysosomal acidification. Expressions of lysosome-associated membrane protein (LAMP) 1 and LAMP2 were significantly higher in P day-18 PBMCs and PMNs. The results suggest that pregnancy-specific activation of lysosomal functions by CTS activation in blood leukocytes is highly associated with IFNT during maternal and fetal recognition of pregnancy. © 2018 Society for Reproduction and Fertility.

Time-Course Study of the Transcriptome of Peripheral Blood Mononuclear Cells (PBMCs) from Sheep Infected with Fasciola hepatica

PubMed Central

Scheerlinck, Jean-Pierre; Ansell, Brendan R. E.; Hall, Ross S.; Gasser, Robin B.; Jex, Aaron R.

2016-01-01

Fasciola hepatica is a parasitic trematode that infects a wide range of mammalian hosts, including livestock and humans, in temperate and tropical regions globally. This trematode causes the disease fascioliasis, which consists of an acute phase (≤ 12 weeks) during which juvenile parasites migrate through the host liver tissues, and a chronic phase (> 12 weeks) following the establishment of adult parasites in the liver bile ducts. Few studies have explored the progression of the host response over the course of Fasciola infection in the same animals. In this study, we characterized transcriptomic changes in peripheral blood mononuclear cells (PBMCs) collected from sheep at three time points over the first eight weeks of infection relative to uninfected controls. In total, 183 and 76 genes were found to be differentially transcribed at two and eight weeks post-infection respectively. Functional and pathway analysis of differentially transcribed genes revealed changes related to T-cell activation that may underpin a Th2-biased immune response against this parasite. This first insight into the dynamics of host responses during the early stages of infection improves the understanding of the pathogenesis of acute fascioliasis, informs vaccine development and presents a set of PBMC markers with diagnostic potential. PMID:27438474

Co-culture of primary human tumor hepatocytes from patients with hepatocellular carcinoma with autologous peripheral blood mononuclear cells: study of their in vitro immunological interactions

PubMed Central

2013-01-01

Background Many studies have suggested that the immune response may play a crucial role in the progression of hepatocellular carcinoma (HCC). Therefore, our aim was to establish a (i) functional culture of primary human tumor hepatocytes and non-tumor from patients with hepatocellular carcinoma (HCC) and (ii) a co-culture system of HCC and non-HCC hepatocytes with autologous peripheral blood mononuclear cells (PBMCs) in order to study in vitro cell-to-cell interactions. Methods Tumor (HCC) and non-tumor (non-HCC) hepatocytes were isolated from the liver resection specimens of 11 patients operated for HCC, while PBMCs were retrieved immediately prior to surgery. Four biopsies were obtained from patients with no liver disease who had surgery for non malignant tumor (normal hepatocytes). Hepatocytes were either cultured alone (monoculture) or co-cultured with PBMCs. Flow cytometry measurements for MHC class II expression, apoptosis, necrosis and viability (7AAD) were performed 24 h, 48 h and 72 h in co-culture and monocultures. Results HCC and non-HCC hepatocytes exhibited increased MHC-II expression at 48h and 72h in co-culture with PBMCs as compared to monoculture, with MHC II-expressing HCC hepatocytes showing increased viability at 72 h. PBMCs showed increased MHC-II expression (activation) in co-culture with HCC as compared to non-HCC hepatocytes at all time points. Moreover, CD8+ T cells had significantly increased apoptosis and necrosis at 48h in co-culture with HCC hepatocytes as compared to monocultures. Interestingly, MHC-II expression on both HCC and non-HCC hepatocytes in co-culture was positively correlated with the respective activated CD8+ T cells. Conclusions We have established an in vitro co-culture model to study interactions between autologous PBMCs and primary HCC and non-HCC hepatocytes. This direct interaction leads to increased antigen presenting ability of HCC hepatocytes, activation of PBMCs with a concomitant apoptosis of activated CD8+ T

Association Between Schizophrenia and DNA Demethylase Activity in Human Peripheral Blood Mononuclear Cells.

PubMed

Zhang, Lili; Pang, Bo; Zhang, Wenbin; Bai, Wei; Yu, Weiying; Li, Yuanyuan; Hua, Wanqing; Li, Wenjun; Kou, Changgui

2018-06-01

DNA demethylase is a crucial enzyme in the epigenetic modification and regulation mechanisms of gene transcription. Based on previous assertions that the pathophysiology of schizophrenia is associated with epigenetics, we aimed to explore whether DNA demethylase activity might be related to schizophrenia in northeast China. We recruited 25 patients with first-episode schizophrenia and 29 normal controls from a northeast Chinese Han population. The diagnostic criteria of schizophrenia were determined according to diseases and related health problems, the tenth revision (ICD-10), and criteria of mental disorders, the third revised edition (CCMD3). DNA demethylase activity in human peripheral blood mononuclear cells (PBMCs) was measured using a DNA demethylase activity colorimetric assay ultra kit. Using Student's t-test, activation of DNA demethylase and its activity were higher in schizophrenia patients compared to healthy individuals (p < 0.001). Furthermore, the level of DNA demethylase activity in male and female subjects with schizophrenia significantly increased (all p < 0.05). Our data showed that DNA demethylase might play a role in the pathophysiology of schizophrenia, and individuals with higher DNA demethylase activity were susceptible to schizophrenia in a northeast Chinese Han population. To the best of our knowledge, this is the first time directly measured human blood samples to examine the association between first-episode schizophrenia patients and DNA demethylase activity, which will provide new insight to explore the effect on the mechanism of schizophrenia.

Comparative study of the effect of chloro-, dichloro-, bromo-, and dibromoacetic acid on necrotic, apoptotic and morphological changes in human peripheral blood mononuclear cells (in vitro study).

PubMed

Michałowicz, Jaromir; Wróblewski, Wojciech; Mokra, Katarzyna; Maćczak, Aneta; Kwiatkowska, Marta

2015-10-01

In this study, the effect of monochloroacetic acid (MCAA), dichloroacetic acid (DCAA), monobromoacetic acid (MBAA) and dibromoacetic acid (DBAA) on human peripheral blood mononuclear cells (PBMCs) was assessed. HAAs studied induced at millimolar concentrations necrotic alterations in PBMCs with the strongest effect noted for MBAA and DBAA. Chloro- and bromoacetic acids also provoked changes in PBMCs morphology because they caused a strong decrease in cell size (particularly DCAA and DBAA) and increase in cell granulation (mainly MBAA and DBAA). All HAAs studied, and DCAA and DBAA in particular (at lower concentrations than those, which caused necrosis) induced apoptotic changes, which was confirmed by analysis of alterations in cell membrane permeability and caspase 8, 9 and 3 activation. Moreover, HAAs examined (mainly dihalogenated acids) strongly increased transmembrane mitochondrial potential and enhanced ROS (mainly hydroxyl radical) formation, which was possibly associated with apoptotic changes provoked by those substances. The results showed that DBAA exhibited the strongest effects on PBMCs. Copyright © 2015 Elsevier Ltd. All rights reserved.

The Impact of Glyphosate, Its Metabolites and Impurities on Viability, ATP Level and Morphological changes in Human Peripheral Blood Mononuclear Cells

PubMed Central

Kwiatkowska, Marta; Jarosiewicz, Paweł; Michałowicz, Jaromir; Koter-Michalak, Maria; Huras, Bogumiła; Bukowska, Bożena

2016-01-01

The toxicity of herbicides to animals and human is an issue of worldwide concern. The present study has been undertaken to assess toxic effect of widely used pesticide—glyphosate, its metabolites: aminomethylphosphonic acid (AMPA) and methylphosphonic acid and its impurities: N-(phosphonomethyl)iminodiacetic acid (PMIDA), N-methylglyphosate, hydroxymethylphosphonic acid and bis-(phosphonomethyl)amine on human peripheral blood mononuclear cells (PBMCs). We have evaluated the effect of those compounds on viability, ATP level, size (FSC-A parameter) and granulation (SSC-A parameter) of the cells studied. Human peripheral blood mononuclear cells were exposed to different concentrations of glyphosate, its metabolites and impurities (0.01–10 mM) for 4 and 24 h. It was found that investigated compounds caused statistically significant decrease in viability and ATP level of PBMCs. The strongest changes in cell viability and ATP level were observed after 24 h incubation of PBMCs with bis-(phosphonomethyl)amine, and particularly PMIDA. Moreover, all studied compounds changed cell granularity, while PMIDA and bis-(phosphonomethyl)amine altered PBMCs size. It may be concluded that bis-(phosphonomethyl)amine, and PMIDA caused a slightly stronger damage to PBMCs than did glyphosate. Changes in the parameters studied in PBMCs were observed only at high concentrations of the compounds examined, which clearly shows that they may occur in this cell type only as a result of acute poisoning of human organism with these substances. PMID:27280764

In vitro transdifferentiation of human peripheral blood mononuclear cells to photoreceptor-like cells

PubMed Central

Komuta, Yukari; Ishii, Toshiyuki; Kaneda, Makoto; Ueda, Yasuji; Miyamoto, Kiyoko; Toyoda, Masashi; Umezawa, Akihiro

2016-01-01

ABSTRACT Direct reprogramming is a promising, simple and low-cost approach to generate target cells from somatic cells without using induced pluripotent stem cells. Recently, peripheral blood mononuclear cells (PBMCs) have attracted considerable attention as a somatic cell source for reprogramming. As a cell source, PBMCs have an advantage over dermal fibroblasts with respect to the ease of collecting tissues. Based on our studies involving generation of photosensitive photoreceptor cells from human iris cells and human dermal fibroblasts by transduction of photoreceptor-related transcription factors via retrovirus vectors, we transduced these transcription factors into PBMCs via Sendai virus vectors. We found that retinal disease-related genes were efficiently detected in CRX-transduced cells, most of which are crucial to photoreceptor functions. In functional studies, a light-induced inward current was detected in some CRX-transduced cells. Moreover, by modification of the culture conditions including additional transduction of RAX1 and NEUROD1, we found a greater variety of retinal disease-related genes than that observed in CRX-transduced PBMCs. These data suggest that CRX acts as a master control gene for reprogramming PBMCs into photoreceptor-like cells and that our induced photoreceptor-like cells might contribute to individualized drug screening and disease modeling of inherited retinal degeneration. PMID:27170256

Amino acids exhibit anti-inflammatory effects in human monocytic leukemia cell line, THP-1 cells.

PubMed

Hasegawa, Shunji; Ichiyama, Takashi; Sonaka, Ichiro; Ohsaki, Ayami; Hirano, Reiji; Haneda, Yasuhiro; Fukano, Reiji; Hara, Masami; Furukawa, Susumu

2011-11-01

The elemental diet is one of the effective therapies for inflammatory bowel disease. However, the mechanism remains unclear, and there have never been reports about the inhibitory effects of amino acids in human monocytes/macrophages. We investigated the inhibitory effects of amino acids on cytokine production or expression of adhesion molecules that are involved in inflammatory diseases, in human monocytes/macrophages. We examined the inhibitory effects of cysteine, histidine or glycine on the induction of nuclear factor-κB (NF-κB) activation, expression of intracellular adhesion molecule-1 (ICAM-1, CD54) and production of interleukin-8 (IL-8) in THP-1 cells, a human monocytic leukemia cell line, and peripheral blood mononuclear cells (PBMCs) stimulated with tumor necrosis factor-α (TNF-α). Cysteine, histidine and glycine significantly reduced the activation of NF-κB in THP-1 cells stimulated with TNF-α. In addition, cysteine and histidine significantly inhibited the expression of ICAM-1 and production of IL-8 in THP-1 cells and PBMCs. Our results suggest that cysteine and histidine exhibit anti-inflammatory effects in THP-1 cells, and may be responsible for the efficacy of treatment in inflammatory bowel diseases.

Excretory-secretory product of third-stage Gnathostoma spinigerum larvae induces apoptosis in human peripheral blood mononuclear cells.

PubMed

Viseshakul, Nareerat; Dechkhajorn, Wilanee; Benjathummarak, Surachet; Nuamtanong, Supaporn; Maneerat, Yaowapa

2017-10-01

Human gnathostomiasis caused by third-stage Gnathostoma spinigerum larvae (G. spinigerum L3) is an important zoonotic disease in tropical areas of the world. The excretory-secretory products (ES) that are excreted by infective larva play a significant role in host immune evasion and tissue destruction. To investigate the poorly understood mechanisms of G. spinigerum L3 pathogenesis, we focused on the potential effect of ES on inducing apoptosis in human immune cells by using human peripheral blood mononuclear cells (PBMCs) as a model. Early and late apoptosis of PBMCs were assessed following the exposure of these cells to G. spinigerum L3 ES (0.1, 0.5, and 1.0 μg/ml) for 6-48 h. The apoptotic cells were identified by flow cytometric staining of PBMC with FITC-annexin V and propidium iodide. The expression of regulatory genes related to apoptosis mechanisms in ES-treated PBMCs was investigated using a Human Apoptosis RT 2 Profiler™ PCR Array. The results showed significant levels of early phase apoptosis at 18 h and of late phase apoptosis at 24 h. We speculate that this apoptosis in PBMCs occurs via the extrinsic pathway. Apoptosis in the ES-induced PBMCs was observed as quickly as 90 min after exposure, and the highest effect was observed at 18-24 h. Furthermore, ES can trigger apoptosis lasting for 48 h. Our findings expand the understanding of one of the mechanisms involved, immune-evasive strategy mechanism used by G. spinigerum larvae during human gnathostomiasis.

High viral load in lymph nodes and latent human immunodeficiency virus (HIV) in peripheral blood cells of HIV-1-infected chimpanzees.

PubMed Central

Saksela, K; Muchmore, E; Girard, M; Fultz, P; Baltimore, D

1993-01-01

We have examined human immunodeficiency virus type 1 (HIV-1) infection in chimpanzees by analyzing HIV-1 DNA and RNA in lymph nodes and peripheral mononuclear cells (PBMCs). Like certain asymptomatic HIV-infected persons, these chimpanzees had no detectable viral replication in their PBMCs. However, viral replication and a high viral load were observed in the lymphatic tissue. Despite the absence of viral replication in PBMCs, 1/1,000 to 1/10,000 of the PBMCs contained HIV-1 proviral DNA, and HIV transcription could be rapidly induced in these cells in vitro. These results provide direct evidence of cellular latency of HIV in vivo and suggest that HIV infection in chimpanzees may be a useful model for clinical latency of HIV infection in humans. Images PMID:8230463

Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment

PubMed Central

Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

2015-01-01

The aim of this study was to quantify the proportion of regulatory T cells (Treg) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)+ Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4+ T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3+ Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3+ Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3+ Treg cells. PMID:25354724

Transcriptomic Response of Porcine PBMCs to Vaccination with Tetanus Toxoid as a Model Antigen

PubMed Central

Adler, Marcel; Murani, Eduard; Brunner, Ronald; Ponsuksili, Siriluck; Wimmers, Klaus

2013-01-01

The aim of the present study was to characterize in vivo genome-wide transcriptional responses to immune stimulation in order to get insight into the resulting changes of allocation of resources. Vaccination with tetanus toxoid was used as a model for a mixed Th1 and Th2 immune response in pig. Expression profiles of PBMCs (peripheral blood mononuclear cells) before and at 12 time points over a period of four weeks after initial and booster vaccination at day 14 were studied by use of Affymetrix GeneChip microarrays and Ingenuity Pathway Analysis (IPA). The transcriptome data in total comprised more than 5000 genes with different transcript abundances (DE-genes). Within the single time stages the numbers of DE-genes were between several hundred and more than 1000. Ingenuity Pathway Analysis mainly revealed canonical pathways of cellular immune response and cytokine signaling as well as a broad range of processes in cellular and organismal growth, proliferation and development, cell signaling, biosynthesis and metabolism. Significant changes in the expression profiles of PBMCs already occurred very early after immune stimulation. At two hours after the first vaccination 679 DE-genes corresponding to 110 canonical pathways of cytokine signaling, cellular immune response and other multiple cellular functions were found. Immune competence and global disease resistance are heritable but difficult to measure and to address by breeding. Besides QTL mapping of immune traits gene expression profiling facilitates the detection of functional gene networks and thus functional candidate genes. PMID:23536793

Transcriptomic response of porcine PBMCs to vaccination with tetanus toxoid as a model antigen.

PubMed

Adler, Marcel; Murani, Eduard; Brunner, Ronald; Ponsuksili, Siriluck; Wimmers, Klaus

2013-01-01

The aim of the present study was to characterize in vivo genome-wide transcriptional responses to immune stimulation in order to get insight into the resulting changes of allocation of resources. Vaccination with tetanus toxoid was used as a model for a mixed Th1 and Th2 immune response in pig. Expression profiles of PBMCs (peripheral blood mononuclear cells) before and at 12 time points over a period of four weeks after initial and booster vaccination at day 14 were studied by use of Affymetrix GeneChip microarrays and Ingenuity Pathway Analysis (IPA). The transcriptome data in total comprised more than 5000 genes with different transcript abundances (DE-genes). Within the single time stages the numbers of DE-genes were between several hundred and more than 1000. Ingenuity Pathway Analysis mainly revealed canonical pathways of cellular immune response and cytokine signaling as well as a broad range of processes in cellular and organismal growth, proliferation and development, cell signaling, biosynthesis and metabolism. Significant changes in the expression profiles of PBMCs already occurred very early after immune stimulation. At two hours after the first vaccination 679 DE-genes corresponding to 110 canonical pathways of cytokine signaling, cellular immune response and other multiple cellular functions were found. Immune competence and global disease resistance are heritable but difficult to measure and to address by breeding. Besides QTL mapping of immune traits gene expression profiling facilitates the detection of functional gene networks and thus functional candidate genes.

Septic Shock Alters Mitochondrial Respiration of Lymphoid Cell-Lines and Human Peripheral Blood Mononuclear Cells: The Role of Plasma.

PubMed

Clere-Jehl, Raphael; Helms, Julie; Kassem, Mohamad; Le Borgne, Pierrick; Delabranche, Xavier; Charles, Anne-Laure; Geny, Bernard; Meziani, Ferhat; Bilbault, Pascal

2018-02-14

In septic shock patients, post-septic immunosuppression state following the systemic inflammatory response syndrome is responsible for nosocomial infections, with subsequent increased mortality. The aim of the present study was to assess the underlying cellular mechanisms of the post-septic immunosuppression state, by investigating mitochondrial functions of peripheral blood mononuclear cells (PBMCs) from septic shock patients over 7 days. Eighteen patients admitted to a French intensive care unit for septic shock were included. At days 1 and 7, PBMCs were isolated by Ficoll density gradient centrifugation. Mitochondrial respiration of intact septic PBMCs was assessed versus control group PBMCs, by measuring O2 consumption in plasma, using high-resolution respirometry. Mitochondrial respiration was then compared between septic plasmas and control plasmas for control PBMCs, septic PBMCs and lymphoid cell-line (CEM). To investigate the role of plasma, we measured several plasma cytokines, among them HMGB1, by ELISA. Basal O2 consumption of septic shock PBMCs was of 8.27 ± 3.39 and 10.48 ± 3.99 pmol/s/10 cells at days 1 and 7 respectively, significantly higher than in control PBMCs (5.37 ± 1.46 pmol/s/10 cells, p < 0.05). Septic patient PBMCs showed a lower response to oligomycin, suggesting a reduced ATP-synthase activity, as well as an increased response to FCCP suggesting an increased mitochondrial respiratory capacity. At 6 hours, septic plasmas showed a decreased O2 consumption of CEM (4.73 ± 1.46 vs. 6.58 ± 1.53, p < 0.05) as well as in control group PBMCs (1.76 ± 0.36 vs. 2.70 ± 0.42, p < 0.05), and triggered a decreased ATP-synthase activity but an increased response to FCCP. These differences are not explained by different cell survival. High HMGB1 levels were significantly associated with reduced PBMCs mitochondrial respiration. Septic plasma impairs mitochondrial respiration in immune cells, with a

[Cloning of human CD45 gene and its expression in Hela cells].

PubMed

Li, Jie; Xu, Tianyu; Wu, Lulin; Zhang, Liyun; Lu, Xiao; Zuo, Daming; Chen, Zhengliang

2015-11-01

To clone human CD45 gene PTPRC and establish Hela cells overexpressing recombinant human CD45 protein. The intact cDNA encoding human CD45 amplified using RT-PCR from the total RNA extracted from peripheral blood mononuclear cells (PBMCs) of a healthy donor was cloned into pMD-18T vector. The CD45 cDNA fragment amplified from the pMD-18T-CD45 by PCR was inserted to the coding region of the PcDNA3.1-3xflag vector, and the resultant recombinant expression vector PcDNA3.1-3xflag-CD45 was transfected into Hela cells. The expression of CD45 in Hela cells was detected by flow cytometry and Western blotting, and the phosphastase activity of CD45 was quantified using an alkaline phosphatase assay kit. The cDNA fragment of about 3 900 bp was amplified from human PBMCs and cloned into pMD-18T vector. The recombinant expression vector PcDNA3.1-3xflag-CD45 was constructed, whose restriction maps and sequence were consistent with those expected. The expression of CD45 in transfected Hela cells was detected by flow cytometry and Western blotting, and the expressed recombinant CD45 protein in Hela cells showed a phosphastase activity. The cDNA of human CD45 was successfully cloned and effectively expressed in Hela cells, which provides a basis for further exploration of the functions of CD45.

Existence of a soluble form of CD50 (intercellular adhesion molecule-3) produced upon human lymphocyte activation. Present in normal human serum and levels are increased in the serum of systemic lupus erythematosus patients.

PubMed

Pino-Otín, M R; Viñas, O; de la Fuente, M A; Juan, M; Font, J; Torradeflot, M; Pallarés, L; Lozano, F; Alberola-Ila, J; Martorell, J

1995-03-15

CD50 (ICAM-3) is a leukocyte differentiation Ag expressed almost exclusively on hemopoietic cells, with a key role in the first steps of immune response. To develop a specific sandwich ELISA to detect a soluble CD50 form (sCD50), two different mAbs (140-11 and 101-1D2) recognizing non-overlapping epitopes were used. sCD50 was detected in the supernatant of stimulated PBMCs, with the highest levels after CD3 triggering. Simultaneously, the CD50 surface expression diminished during the first 24 h. sCD50 isolated from culture supernatant and analyzed by immunoblotting showed an apparent m.w. of 95 kDa, slightly smaller than the membrane form. These data, together with Northern blot kinetics analysis, suggest that sCD50 is cleaved from cell membrane. Furthermore, we detect sCD50 in normal human sera and higher levels in sera of systemic lupus erythematosus (SLE) patients, especially in those in active phase. The sCD50 levels showed a positive correlation with sCD27 levels (r = 0.4213; p = 0.0026). Detection of sCD50, both after in vitro CD3 triggering of PBMCs and increased in SLE sera, suggests that sCD50 could be used as a marker of lymphocyte stimulation.

UV-inactivated HSV-1 potently activates NK cell killing of leukemic cells

PubMed Central

Samudio, Ismael; Rezvani, Katayoun; Shaim, Hila; Hofs, Elyse; Ngom, Mor; Bu, Luke; Liu, Guoyu; Lee, Jason T. C.; Imren, Suzan; Lam, Vivian; Poon, Grace F. T.; Ghaedi, Maryam; Takei, Fumio; Humphries, Keith; Jia, William

2016-01-01

Herein we demonstrate that oncolytic herpes simplex virus-1 (HSV-1) potently activates human peripheral blood mononuclear cells (PBMCs) to lyse leukemic cell lines and primary acute myeloid leukemia samples, but not healthy allogeneic lymphocytes. Intriguingly, we found that UV light–inactivated HSV-1 (UV-HSV-1) is equally effective in promoting PBMC cytolysis of leukemic cells and is 1000- to 10 000-fold more potent at stimulating innate antileukemic responses than UV-inactivated cytomegalovirus, vesicular stomatitis virus, reovirus, or adenovirus. Mechanistically, UV-HSV-1 stimulates PBMC cytolysis of leukemic cells, partly via Toll-like receptor-2/protein kinase C/nuclear factor-κB signaling, and potently stimulates expression of CD69, degranulation, migration, and cytokine production in natural killer (NK) cells, suggesting that surface components of UV-HSV-1 directly activate NK cells. Importantly, UV-HSV-1 synergizes with interleukin-15 (IL-15) and IL-2 in inducing activation and cytolytic activity of NK cells. Additionally, UV-HSV-1 stimulates glycolysis and fatty acid oxidation–dependent oxygen consumption in NK cells, but only glycolysis is required for their enhanced antileukemic activity. Last, we demonstrate that T cell–depleted human PBMCs exposed to UV-HSV-1 provide a survival benefit in a murine xenograft model of human acute myeloid leukemia (AML). Taken together, our results support the preclinical development of UV-HSV-1 as an adjuvant, alone or in combination with IL-15, for allogeneic donor mononuclear cell infusions to treat AML. PMID:26941401

Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment.

PubMed

Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

2015-03-01

The aim of this study was to quantify the proportion of regulatory T cells (Treg ) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)(+) Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4(+) T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3(+) Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3(+) Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3(+) Treg cells. © 2014 British Society for Immunology.

Combustible and non-combustible tobacco product preparations differentially regulate human peripheral blood mononuclear cell functions.

PubMed

Arimilli, Subhashini; Damratoski, Brad E; Prasad, G L

2013-09-01

Natural killer (NK) cells and T cells play essential roles in innate and adaptive immune responses in protecting against microbial infections and in tumor surveillance. Although evidence suggests that smoking causes immunosuppression, there is limited information whether the use of smokeless tobacco (ST) products affects immune responses. In this study, we assessed the effects of two preparations of cigarette smoke, ST extract and nicotine on T cell and NK cell responses using Toll-like receptor-ligand stimulated human peripheral blood mononuclear cells (PBMCs). The tobacco product preparations (TPPs) tested included whole smoke conditioned media (WS-CM), total particulate matter (TPM) and a ST product preparation in complete artificial saliva (ST/CAS). The PBMCs were stimulated with polyinosinic:polycytidylic acid (poly I:C) and lipopolysaccharide (LPS). A marked reduction of the expression of intracellular IFN-γ and TNF-α was evident in NK cells and T cells treated with WS-CM and TPM. Consistently, attenuation of ligand-induced secretion of cytokines (IL-1β, IL-10, IL-12 and TNF-α) from PBMCs treated with WS-CM and TPM were observed. While the treatment with TPPs did not alter the expression of the maturation marker CD69, WS-CM and TPM inhibited the cytolytic activity of human PBMCs. Suppression of perforin by WS-CM was also detected. Although interference from the vehicle confounded the interpretation of effects of ST/CAS, some effects were evident only at high concentrations. Nicotine treatment minimally impacted expression of cytokines and cytolytic activity. Data presented herein suggests that the function of NK cells and T cells is influenced by exposure to TPPs (based on equi-nicotine units) in the following order: WS-CM>TPM>ST/CAS. These findings are consistent with the hypothesis put forward by others that chronic smoking leads to immunosuppression, an effect that may contribute to increased microbial infections and cancer incidence among smokers

Chicken cathelicidin-2-derived peptides with enhanced immunomodulatory and antibacterial activities against biological warfare agents.

PubMed

Molhoek, E Margo; van Dijk, Albert; Veldhuizen, Edwin J A; Dijk-Knijnenburg, Helma; Mars-Groenendijk, Roos H; Boele, Linda C L; Kaman-van Zanten, Wendy E; Haagsman, Henk P; Bikker, Floris J

2010-09-01

Host defence peptides (HDPs) are considered to be excellent candidates for the development of novel therapeutic agents. Recently, it was demonstrated that the peptide C1-15, an N-terminal segment of chicken HDP cathelicidin-2, exhibits potent antibacterial activity while lacking cytotoxicity towards eukaryotic cells. In the present study, we report that C1-15 is active against bacteria such as Bacillus anthracis and Yersinia pestis that may potentially be used by bioterrorists. Substitution of single and multiple phenylalanine (Phe) residues to tryptophan (Trp) in C1-15 resulted in variants with improved antibacterial activity against B. anthracis and Y. pestis as well as decreased salt sensitivity. In addition, these peptides exhibited enhanced neutralisation of lipopolysaccharide (LPS)-induced release of pro-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs). The antibacterial and LPS-neutralising activities of these C1-15-derived peptides are exerted at concentrations far below the concentrations that are toxic to human PBMCs. Taken together, we show that Phe-->Trp substitutions in C1-15 variants enhances the antibacterial and LPS-neutralising activities against pathogenic bacteria, including those that may potentially be used as biological warfare agents. Copyright (c) 2010 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Tax posttranslational modifications and interaction with calreticulin in MT-2 cells and human peripheral blood mononuclear cells of human T cell lymphotropic virus type-I-associated myelopathy/tropical spastic paraparesis patients.

PubMed

Medina, Fernando; Quintremil, Sebastian; Alberti, Carolina; Barriga, Andres; Cartier, Luis; Puente, Javier; Ramírez, Eugenio; Ferreira, Arturo; Tanaka, Yuetsu; Valenzuela, Maria Antonieta

2014-04-01

The human retrovirus human T cell lymphotropic virus type-I (HTLV-1) is the etiologic agent of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Axonal degeneration in HAM/TSP patients occurs without neuron infection, with the secreted viral Tax protein proposed to be involved. We previously found that Tax secreted into the culture medium of MT-2 cells (HTLV-1-infected cell line) produced neurite retraction in neuroblastoma cells differentiated to neuronal type. To assess the relevance of Tax posttranslational modifications on this effect, we addressed the question of whether Tax secreted by MT-2 cells and peripheral blood mononuclear cells (PBMCs) of HTLV-1-infected subjects is modified. The interaction of Tax with calreticulin (CRT) that modulates intracellular Tax localization and secretion has been described. We studied Tax localization and modifications in MT-2 cells and its interaction with CRT. Intracellular Tax in MT-2 cells was assessed by flow cytometry, corresponding mainly to a 71-kDa protein followed by western blot. This protein reported as a chimera with gp21 viral protein-confirmed by mass spectrometry-showed no ubiquitination or SUMOylation. The Tax-CRT interaction was determined by confocal microscopy and coimmunoprecipitation. Extracellular Tax from HAM/TSP PBMCs is ubiquitinated according to western blot, and its interaction with CRT was shown by coimmunoprecipitation. A positive correlation between Tax and CRT secretion was observed in HAM/TSP PBMCs and asymptomatic carriers. For both proteins inhibitors and activators of secretion showed secretion through the endoplasmic reticulum-Golgi complex. Tax, present in PBMC culture medium, produced neurite retraction in differentiated neuroblastoma cells. These results suggest that Tax, whether ubiquitinated or not, is active for neurite retraction.

A shift to glycolysis accompanies the inflammatory changes in PBMCs from individuals with an IQ-discrepant memory.

PubMed

Wolfe, Hannah; Hannigan, Caoimhe; O'Sullivan, Michael; Carroll, Liam Barry; Brennan, Sabina; Lawlor, Brian; Robertson, Ian H; Lynch, Marina

2018-04-15

Identification of a blood-based biomarker that can detect early cognitive decline presents a significant healthcare challenge. We prepared peripheral blood mononuclear cells (PBMCs) from individuals who had a poorer than predicted performance in their delayed recall performance on the Logical Memory II Subtest of the Wechsler Memory Scale (WMS) relative to their IQ estimated by the National Adult Reading Test (NART); we described these individuals as IQ-discrepant, compared with IQ-consistent, individuals. Stimulation with Aβ + LPS increased production of TNFα to a greater extent in cells from IQ-discrepant, compared with IQ-consistent, individuals. This was associated with a shift towards glycolysis and the evidence indicates that 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase (PFKFB)3 plays a role in driving glycolysis. A similar shift towards glycolysis was observed in MDMs prepared from IQ-discrepant, compared with IQ-consistent, individuals. The important finding here is that we have established an increased sensitivity to Aβ + LPS stimulation in PBMCs from individuals that under-perform on a memory task, relative to their estimated premorbid IQ, which may be an indicator of early cognitive decline. This may be a useful tool in determining the presence of early cognitive dysfunction. Copyright © 2018 Elsevier B.V. All rights reserved.

Tax Posttranslational Modifications and Interaction with Calreticulin in MT-2 Cells and Human Peripheral Blood Mononuclear Cells of Human T Cell Lymphotropic Virus Type-I-Associated Myelopathy/Tropical Spastic Paraparesis Patients

PubMed Central

Medina, Fernando; Quintremil, Sebastian; Alberti, Carolina; Barriga, Andres; Cartier, Luis; Puente, Javier; Ramírez, Eugenio; Ferreira, Arturo; Tanaka, Yuetsu

2014-01-01

Abstract The human retrovirus human T cell lymphotropic virus type-I (HTLV-1) is the etiologic agent of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Axonal degeneration in HAM/TSP patients occurs without neuron infection, with the secreted viral Tax protein proposed to be involved. We previously found that Tax secreted into the culture medium of MT-2 cells (HTLV-1-infected cell line) produced neurite retraction in neuroblastoma cells differentiated to neuronal type. To assess the relevance of Tax posttranslational modifications on this effect, we addressed the question of whether Tax secreted by MT-2 cells and peripheral blood mononuclear cells (PBMCs) of HTLV-1-infected subjects is modified. The interaction of Tax with calreticulin (CRT) that modulates intracellular Tax localization and secretion has been described. We studied Tax localization and modifications in MT-2 cells and its interaction with CRT. Intracellular Tax in MT-2 cells was assessed by flow cytometry, corresponding mainly to a 71-kDa protein followed by western blot. This protein reported as a chimera with gp21 viral protein—confirmed by mass spectrometry—showed no ubiquitination or SUMOylation. The Tax–CRT interaction was determined by confocal microscopy and coimmunoprecipitation. Extracellular Tax from HAM/TSP PBMCs is ubiquitinated according to western blot, and its interaction with CRT was shown by coimmunoprecipitation. A positive correlation between Tax and CRT secretion was observed in HAM/TSP PBMCs and asymptomatic carriers. For both proteins inhibitors and activators of secretion showed secretion through the endoplasmic reticulum–Golgi complex. Tax, present in PBMC culture medium, produced neurite retraction in differentiated neuroblastoma cells. These results suggest that Tax, whether ubiquitinated or not, is active for neurite retraction. PMID:24321043

Comparative genotypic and phenotypic analysis of human peripheral blood monocytes and surrogate monocyte-like cell lines commonly used in metabolic disease research.

PubMed

Riddy, Darren M; Goy, Emily; Delerive, Philippe; Summers, Roger J; Sexton, Patrick M; Langmead, Christopher J

2018-01-01

Monocyte-like cell lines (MCLCs), including THP-1, HL-60 and U-937 cells, are used routinely as surrogates for isolated human peripheral blood mononuclear cells (PBMCs). To systematically evaluate these immortalised cells and PBMCs as model systems to study inflammation relevant to the pathogenesis of type II diabetes and immuno-metabolism, we compared mRNA expression of inflammation-relevant genes, cell surface expression of cluster of differentiation (CD) markers, and chemotactic responses to inflammatory stimuli. Messenger RNA expression analysis suggested most genes were present at similar levels across all undifferentiated cells, though notably, IDO1, which encodes for indoleamine 2,3-dioxygenase and catabolises tryptophan to kynureninase (shown to be elevated in serum from diabetic patients), was not expressed in any PMA-treated MCLC, but present in GM-CSF-treated PBMCs. There was little overall difference in the pattern of expression of CD markers across all cells, though absolute expression levels varied considerably and the correlation between MCLCs and PBMCs was improved upon MCLC differentiation. Functionally, THP-1 and PBMCs migrated in response to chemoattractants in a transwell assay, with varying sensitivity to MCP-1, MIP-1α and LTB-4. However, despite similar gene and CD expression profiles, U-937 cells were functionally impaired as no migration was observed to any chemoattractant. Our analysis reveals that the MCLCs examined only partly replicate the genotypic and phenotypic properties of human PBMCs. To overcome such issues a universal differentiation protocol should be implemented for these cell lines, similar to those already used with isolated monocytes. Although not perfect, in our hands the THP-1 cells represent the closest, simplified surrogate model of PBMCs for study of inflammatory cell migration.

DNA damage and methylation induced by glyphosate in human peripheral blood mononuclear cells (in vitro study).

PubMed

Kwiatkowska, Marta; Reszka, Edyta; Woźniak, Katarzyna; Jabłońska, Ewa; Michałowicz, Jaromir; Bukowska, Bożena

2017-07-01

Glyphosate is a very important herbicide that is widely used in the agriculture, and thus the exposure of humans to this substance and its metabolites has been noted. The purpose of this study was to assess DNA damage (determination of single and double strand-breaks by the comet assay) as well as to evaluate DNA methylation (global DNA methylation and methylation of p16 (CDKN2A) and p53 (TP53) promoter regions) in human peripheral blood mononuclear cells (PBMCs) exposed to glyphosate. PBMCs were incubated with the compound studied at concentrations ranging from 0.1 to 10 mM for 24 h. The study has shown that glyphosate induced DNA lesions, which were effectively repaired. However, PBMCs were unable to repair completely DNA damage induced by glyphosate. We also observed a decrease in global DNA methylation level at 0.25 mM of glyphosate. Glyphosate at 0.25 mM and 0.5 mM increased p53 promoter methylation, while it did not induce statistically significant changes in methylation of p16 promoter. To sum up, we have shown for the first time that glyphosate (at high concentrations from 0.5 to 10 mM) may induce DNA damage in leucocytes such as PBMCs and cause DNA methylation in human cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

Human T cell leukaemia virus type 2 tax protein mediates CC-chemokine expression in peripheral blood mononuclear cells via the nuclear factor kappa B canonical pathway.

PubMed

Barrios, C S; Castillo, L; Zhi, H; Giam, C-Z; Beilke, M A

2014-01-01

Retroviral co-infections with human immunodeficiency virus type-1 (HIV-1) and human T cell leukaemia virus type 1 (HTLV-1) or type 2 (HTLV-2) are prevalent in many areas worldwide. It has been observed that HIV-1/HTLV-2 co-infections are associated with slower rates of CD4(+) T cell decline and delayed progression to AIDS. This immunological benefit has been linked to the ability of Tax2, the transcriptional activating protein of HTLV-2, to induce the expression of macrophage inflammatory protein (MIP)-1α/CCL3, MIP-1β/CCL4 and regulated upon activation normal T cell expressed and secreted (RANTES)/CCL5 and to down-regulate the expression of the CCR5 co-receptor in peripheral blood mononuclear cells (PBMCs). This study aimed to assess the role of Tax2-mediated activation of the nuclear factor kappa B (NF-κB) signalling pathway on the production of the anti-viral CC-chemokines MIP-1α, MIP-1β and RANTES. Recombinant Tax1 and Tax2 proteins, or proteins expressed via adenoviral vectors used to infect cells, were tested for their ability to activate the NF-κB pathway in cultured PBMCs in the presence or absence of NF-κB pathway inhibitors. Results showed a significant release of MIP-1α, MIP-1β and RANTES by PBMCs after the activation of p65/RelA and p50. The secretion of these CC-chemokines was significantly reduced (P < 0·05) by canonical NF-κB signalling inhibitors. In conclusion, Tax2 protein may promote innate anti-viral immune responses through the activation of the canonical NF-κB pathway. © 2013 British Society for Immunology.

Response to Rhinovirus Infection by Human Airway Epithelial Cells and Peripheral Blood Mononuclear Cells in an In Vitro Two-Chamber Tissue Culture System

PubMed Central

Rajan, Devi; Gaston, Kelsey A.; McCracken, Courtney E.; Erdman, Dean D.; Anderson, Larry J.

2013-01-01

Human rhinovirus (HRV) infections are associated with the common cold, occasionally with more serious lower respiratory tract illnesses, and frequently with asthma exacerbations. The clinical features of HRV infection and its association with asthma exacerbation suggest that some HRV disease results from virus-induced host immune responses to infection. To study the HRV-infection-induced host responses and the contribution of these responses to disease, we have developed an in vitro model of HRV infection of human airway epithelial cells (Calu-3 cells) and subsequent exposure of human peripheral blood mononuclear cells (PBMCs) to these infected cells in a two-chamber trans-well tissue culture system. Using this model, we studied HRV 14 (species B) and HRV 16 (species A) induced cytokine and chemokine responses with PBMCs from four healthy adults. Infection of Calu-3 cells with either virus induced HRV-associated increases in FGF-Basic, IL-15, IL-6, IL-28A, ENA-78 and IP-10. The addition of PBMCs to HRV 14-infected cells gave significant increases in MIP-1β, IL-28A, MCP-2, and IFN-α as compared with mock-infected cells. Interestingly, ENA-78 levels were reduced in HRV 14 infected cells that were exposed to PBMCs. Addition of PBMCs to HRV 16-infected cells did not induce MIP-1β, IL-28A and IFN-α efficiently nor did it decrease ENA-78 levels. Our results demonstrate a clear difference between HRV 14 and HRV 16 and the source of PBMCs, in up or down regulation of several cytokines including those that are linked to airway inflammation. Such differences might be one of the reasons for variation in disease associated with different HRV species including variation in their link to asthma exacerbations as suggested by other studies. Further study of immune responses associated with different HRVs and PBMCs from different patient groups, and the mechanisms leading to these differences, should help characterize pathogenesis of HRV disease and generate novel approaches to

Activated glycoprotein A repetitions predominant (GARP)-expressing regulatory T cells inhibit allergen-induced intestinal inflammation in humanized mice.

PubMed

Eschborn, Melanie; Weigmann, Benno; Reissig, Sonja; Waisman, Ari; Saloga, Joachim; Bellinghausen, Iris

2015-07-01

Recently, we developed a humanized mouse model of allergen-induced IgE-dependent gut inflammation in PBMC-engrafted immunodeficient mice. In the present study, we wanted to investigate the role of regulatory T (Treg) cells and their activation status in this model. Nonobese diabetic-severe combined immunodeficiency-γc(-/-) mice were injected intraperitoneally with human PBMCs from allergic donors together with the respective allergen or NaCl as control in the presence or absence of different concentrations of CD4(+)CD25(+) Treg cells of the same donor. After an additional allergen boost 1 week later, mice were challenged with the allergen rectally on day 21 and gut inflammation was monitored by a high-resolution video mini-endoscopic system evaluating translucency, granularity, fibrin production, vascularity, and stool. Allergen-specific human IgE in mouse sera, which was detectable only in PBMC plus allergen-treated mice, was strongly inhibited by coinjection of Treg cells at a ratio of at least 1:10. Consequently, the presence of Treg cells significantly decreased IgE-dependent allergen-induced gut inflammation after rectal allergen challenge. In addition, Treg cells reduced allergen-specific proliferation and cytokine production of recovered human CD4(+) T cells in vitro. Activation of Treg cells before injection further increased all inhibitory effects. Prevention of gut inflammation also occurred by the administration of glycoprotein A repetitions predominant, a molecule expressed by activated Treg cells, whereas its blockade completely abrogated inhibition by Treg cells. These results demonstrate that allergen-specific gut inflammation in human PBMC-engrafted mice can be avoided by enhancing the numbers or activity of autologous Treg cells, which is of great interest for therapeutic intervention of allergic diseases of the intestine. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Severe necroinflammatory reaction caused by natural killer cell-mediated Fas/Fas ligand interaction and dendritic cells in human hepatocyte chimeric mouse.

PubMed

Okazaki, Akihito; Hiraga, Nobuhiko; Imamura, Michio; Hayes, C Nelson; Tsuge, Masataka; Takahashi, Shoichi; Aikata, Hiroshi; Abe, Hiromi; Miki, Daiki; Ochi, Hidenori; Tateno, Chise; Yoshizato, Katsutoshi; Ohdan, Hideki; Chayama, Kazuaki

2012-08-01

The necroinflammatory reaction plays a central role in hepatitis B virus (HBV) elimination. Cluster of differentiation (CD)8-positive cytotoxic T lymphocytes (CTLs) are thought to be a main player in the elimination of infected cells, and a recent report suggests that natural killer (NK) cells also play an important role. Here, we demonstrate the elimination of HBV-infected hepatocytes by NK cells and dendritic cells (DCs) using urokinase-type plasminogen activator/severe combined immunodeficiency mice, in which the livers were highly repopulated with human hepatocytes. After establishing HBV infection, we injected human peripheral blood mononuclear cells (PBMCs) into the mice and analyzed liver pathology and infiltrating human immune cells with flow cytometry. Severe hepatocyte degeneration was observed only in HBV-infected mice transplanted with human PBMCs. We provide the first direct evidence that massive liver cell death can be caused by Fas/Fas ligand (FasL) interaction provided by NK cells activated by DCs. Treatment of mice with anti-Fas antibody completely prevented severe hepatocyte degeneration. Furthermore, severe hepatocyte death can be prevented by depletion of DCs, whereas depletion of CD8-positive CTLs did not disturb the development of massive liver cell apoptosis. Our findings provide the first direct evidence that DC-activated NK cells induce massive HBV-infected hepatocyte degeneration through the Fas/FasL system and may indicate new therapeutic implications for acute severe/fulminant hepatitis B. Copyright © 2012 American Association for the Study of Liver Diseases.

Significantly Accelerated Wound Healing of Full-Thickness Skin Using a Novel Composite Gel of Porcine Acellular Dermal Matrix and Human Peripheral Blood Cells.

PubMed

Kuna, Vijay K; Padma, Arvind M; Håkansson, Joakim; Nygren, Jan; Sjöback, Robert; Petronis, Sarunas; Sumitran-Holgersson, Suchitra

2017-02-16

Here we report the fabrication of a novel composite gel from decellularized gal-gal-knockout porcine skin and human peripheral blood mononuclear cells (hPBMCs) for full-thickness skin wound healing. Decellularized skin extracellular matrix (ECM) powder was prepared via chemical treatment, freeze drying, and homogenization. The powder was mixed with culture medium containing hyaluronic acid to generate a pig skin gel (PSG). The effect of the gel in regeneration of full-thickness wounds was studied in nude mice. We found significantly accelerated wound closure already on day 15 in animals treated with PSG only or PSG + hPBMCs compared to untreated and hyaluronic acid-treated controls (p < 0.05). Addition of the hPBMCs to the gel resulted in marked increase of host blood vessels as well as the presence of human blood vessels. At day 25, histologically, the wounds in animals treated with PSG only or PSG + hPBMCs were completely closed compared to those of controls. Thus, the gel facilitated generation of new skin with well-arranged epidermal cells and restored bilayer structure of the epidermis and dermis. These results suggest that porcine skin ECM gel together with human cells may be a novel and promising biomaterial for medical applications especially for patients with acute and chronic skin wounds.

The effect of N-nitrosodimethylamine (NDMA) on Bax and Mcl-1 expression in human neutrophils.

PubMed

Jablonski, Jakub; Jablonska, Ewa; Leonik, Agnieszka

2011-12-01

In the present study we examined a role of pro-apoptotic Bax and anti-apoptotic Mcl-1 proteins, participating in the regulation of intrinsic apoptosis pathway in human neutrophils (PMNs) exposed to N-nitrosodimethylamine (NDMA), the environmental xenobiotic. For the purpose comparison, the same studies were conducted in autologous peripheral blood mononuclear cells (PBMCs). The production of cytochrome c by PMNs was also determined. A deficit of anti-apoptotic Mcl-1 and overexpression of the pro-apoptotic protein Bax suggest that the apoptosis process in human neutrophils exposed to NDMA is dependent on changes in the expression of these proteins. PMNs were more sensitive to NDMA than PBMCs.

Antiproliferative Activity of Xanthones Isolated from Artocarpus obtusus

PubMed Central

Hashim, Najihah Mohd; Rahmani, Mawardi; Ee, Gwendoline Cheng Lian; Sukari, Mohd Aspollah; Yahayu, Maizatulakmal; Oktima, Winda; Ali, Abd Manaf; Go, Rusea

2012-01-01

An investigation of the chemical constituents in Artocarpus obtusus species led to the isolation of three new xanthones, pyranocycloartobiloxanthone A (1), dihydroartoindonesianin C (2), and pyranocycloartobiloxanthone B (3). The compounds were subjected to antiproliferative assay against human promyelocytic leukemia (HL60), human chronic myeloid leukemia (K562), and human estrogen receptor (ER+) positive breast cancer (MCF7) cell lines. Pyranocycloartobiloxanthone A (1) consistently showed strong cytotoxic activity against the three cell lines compared to the other two with IC50 values of 0.5, 2.0 and 5.0 μg/mL, respectively. Compound (1) was also observed to exert antiproliferative activity and apoptotic promoter towards HL60 and MCF7 cell lines at respective IC50 values. The compound (1) was not toxic towards normal cell lines human nontumorigenic breast cell line (MCF10A) and human peripheral blood mononuclear cells (PBMCs) with IC50 values of more than 30 μg/mL. PMID:21960741

Identification, Characterization, Immunolocalization, and Biological Activity of Lucilin Peptide.

PubMed

Alberto, Tellez German; Alejandra, Zapata Jesica; Johanna, Toro Lily; Carolina, Henao Diana; Pablo, Bedoya Juan; David, Rivera Juan; Valentin, Trujillo Juan; Bruno, Rivas; Lopez, Richard Onalbi Hoyos; Carlos, Castano Jhon

2018-06-08

Maggots from the Lucilia sp. genus are used for debridement of infected and necrotic wounds. Broad-spectrum antimicrobial activity has been described in the excretion/secretions (ES 1 ) of these larvae. This study identifies the genetic sequence of a cecropin-like antimicrobial peptide from Lucilia eximia. Total RNA was extracted and used for PCR-RACE amplification of a cecropin, the native peptide was immunolocalized in the tissues and secretions of the larvae, and a synthetic analog was used to explore its antimicrobial, cytotoxic, LPS neutralizing and wound-healing activities in vitro. The genetic cDNA sequence of a cecropin-like antimicrobial peptide in L. eximia called "Lucilin" was amplified, corresponding to 63 aa completed protein and 40 aa mature peptide; the structure of the mature peptide was predicted as an α-helix. The peptide was immunolocalized in the salivary glands, fat body, the ES, and hemolymph of the maggots. Lucilin synthetic peptide analog was active against E. coli DH10B with a MIC 2 of 7.8 µg/mL, E. coli extended spectrum b-lactamase (ESBL) (MIC: 15.6 µg/mL), and Enterobacter cloacae (MIC: 125 µg/mL), but it was not active against Pseudomonas aeruginosa and Staphylococcus epidermidis; and had no cytotoxic or hemolytic activity. It showed immunomodulatory activity against human peripheral blood mononuclear cells (PBMCs) stimulated with LPS, reducing the TNF-α production when treated at 17 µg/mL and induces cell migration of Hacat at 5 and 50 µg/mL. Lucilin is a cecropin-like peptide from L. eximia with antimicrobial activity against Gram negative bacteria and immunomodulatory activities, decreasing the TNF-α production in PBMCs and inducing cellular migration in human keratinocytes. Copyright © 2018. Published by Elsevier B.V.

Low-concentration exposure to BPA, BPF and BPAF induces oxidative DNA bases lesions in human peripheral blood mononuclear cells.

PubMed

Mokra, Katarzyna; Woźniak, Katarzyna; Bukowska, Bożena; Sicińska, Paulina; Michałowicz, Jaromir

2018-06-01

Because bisphenol A (BPA) and some of its analogs have been supposed to influence development of cancer, we have assessed the effect of BPA, bisphenol S (BPS), bisphenol F (BPF) and bisphenol AF (BPAF) on DNA bases oxidation, which is a key process in cancer initiation. The analysis was conducted on human peripheral blood mononuclear cells (PBMCs), which are very useful model to assess genotoxic potential of various toxicants in different cell types. In order to determine oxidative damage to DNA pyrimidines and purines, alkaline version of the comet assay with DNA glycosylases, i.e. endonuclease III (Nth) and human 8-oxoguanine DNA glycosylase (hOGG1) was used. PBMCs were exposed to BPA or its analogs in the concentrations of 0.01, 0.1 and 1 μg/mL for 4 h and 0.001, 0.01 and 0.1 μg/mL for 48 h. We have observed that BPA, BPS, BPF and particularly BPAF caused oxidative damage to DNA pyrimidines and more strongly to purines in human PBMCs. The results have also shown that BPS, which is the most commonly used as a substitute for BPA in the manufacture induced definitely the smallest oxidative DNA bases lesions in PBMCs. Moreover, we have noticed that BPA, BPF and BPAF caused DNA damage at very low concentration of 1 ng/mL. Copyright © 2018 Elsevier Ltd. All rights reserved.

Bisphenol A and its analogs induce morphological and biochemical alterations in human peripheral blood mononuclear cells (in vitro study).

PubMed

Michałowicz, Jaromir; Mokra, Katarzyna; Bąk, Agata

2015-10-01

Few studies have addressed the cellular effects of bisphenol S (BPS) and bisphenol AF (BPAF) on cells, and no study has been conducted to analyze the mechanism of action of bisphenols in blood cells. In this study, the effect of bisphenol A (BPA), bisphenol F (BPF), BPS and BPAF on human peripheral blood mononuclear cells (PBMCs) was analyzed. It was shown that BPA, BPF and BPAF in particular, decreased cell viability, which was associated with depletion of intracellular ATP level and alterations in PBMCs size and granulation. Bisphenols enhanced ROS (including OH˙) formation, which led to damage to lipids and proteins in PBMCs. The most significant alterations in ROS level were induced by BPF, and particularly BPAF. Moreover, it was shown that BPAF most strongly provoked lipid peroxidation, while BPA and BPS caused the greatest damage to proteins. It may be concluded that BPA and its analogs were capable of inducing oxidative stress and damage in PBMCs in the concentrations ranging from 0.06 to 0.5 μM (0.02-0.1 μg/ml), which may be present in human blood as a result of environmental exposure. Although, most of bisphenols studied decreased cell viability, size and ATP level at higher concentrations, BPAF exhibited its cytotoxic potential at low concentrations ranging from 0.3 to 3 μM (0.1-1.0 μg/ml) that may correspond to concentrations in humans following occupational exposure. Copyright © 2015 Elsevier Ltd. All rights reserved.

Compartmentalized bronchoalveolar IFN-gamma and IL-12 response in human pulmonary tuberculosis.

PubMed

Herrera, Maria Teresa; Torres, Martha; Nevels, Denarra; Perez-Redondo, Carlos Núñez; Ellner, Jerrold J; Sada, Eduardo; Schwander, Stephan K

2009-01-01

Human tuberculosis (TB) principally involves the lungs, where local immunity impacts on the load of Mycobacterium tuberculosis (M.tb). Because concomitants of local Th1 immunity are still under-explored in humans, we characterized immune responses in bronchoalveolar cells (BACs) and systemically in peripheral blood mononuclear cells (PBMCs) in persons with active pulmonary TB and in healthy community controls. PPD- and live M.tb-induced IFN-gamma-production were observed in CD4(+), CD8(+), gammadeltaTCR(+), and CD56(+) alveolar T cell subpopulations and NK cells (CD3(-)CD56(+)). IFN-gamma-producing CD4(+) T cells (mostly CD45RO(+)) were more abundant (p<0.05). M.tb-induced IL-12p70, but interestingly also IL-4, was increased (p<0.05) in BACs from TB patients. Constitutive expression of IL-12Rbeta1 and IL-12Rbeta2 mRNA in BACs and PBMCs and IFN-gammaR1 in BACs was similar in both study groups. Data were normalized to account for differences in proportions of alveolar T cells and macrophages in the study groups. IFN-gamma-production and its induction by IL-12R engagement occur virtually unimpaired in the bronchoalveolar spaces of patients with pulmonary TB. The reasons for the apparent failure to control M. tuberculosis growth during active pulmonary TB disease is unknown but could be the expression of locally acting immunosuppressive mechanisms that subvert the antimycobacterial effects of IFN-gamma.

Activation of peripheral blood mononuclear cells by dengue virus infection depotentiates balapiravir.

PubMed

Chen, Yen-Liang; Abdul Ghafar, Nahdiyah; Karuna, Ratna; Fu, Yilong; Lim, Siew Pheng; Schul, Wouter; Gu, Feng; Herve, Maxime; Yokohama, Fumiaki; Wang, Gang; Cerny, Daniela; Fink, Katja; Blasco, Francesca; Shi, Pei-Yong

2014-02-01

In a recent clinical trial, balapiravir, a prodrug of a cytidine analog (R1479), failed to achieve efficacy (reducing viremia after treatment) in dengue patients, although the plasma trough concentration of R1479 remained above the 50% effective concentration (EC(50)). Here, we report experimental evidence to explain the discrepancy between the in vitro and in vivo results and its implication for drug development. R1479 lost its potency by 125-fold when balapiravir was used to treat primary human peripheral blood mononuclear cells (PBMCs; one of the major cells targeted for viral replication) that were preinfected with dengue virus. The elevated EC(50) was greater than the plasma trough concentration of R1479 observed in dengue patients treated with balapiravir and could possibly explain the efficacy failure. Mechanistically, dengue virus infection triggered PBMCs to generate cytokines, which decreased their efficiency of conversion of R1479 to its triphosphate form (the active antiviral ingredient), resulting in decreased antiviral potency. In contrast to the cytidine-based compound R1479, the potency of an adenosine-based inhibitor of dengue virus (NITD008) was much less affected. Taken together, our results demonstrate that viral infection in patients before treatment could significantly affect the conversion of the prodrug to its active form; such an effect should be calculated when estimating the dose efficacious for humans.

Base-modified UDP-sugars reduce cell surface levels of P-selectin glycoprotein 1 (PSGL-1) on IL-1β-stimulated human monocytes

PubMed Central

Kanabar, Varsha; Tedaldi, Lauren; Jiang, Jingqian; Nie, Xiaodan; Panina, Irina; Descroix, Karine; Man, Francis; Pitchford, Simon C; Page, Clive P; Wagner, Gerd K

2016-01-01

P-selectin glycoprotein ligand-1 (PSGL-1, CD162) is a cell-surface glycoprotein that is expressed, either constitutively or inducibly, on all myeloid and lymphoid cell lineages. PSGL-1 is implicated in cell–cell interactions between platelets, leukocytes and endothelial cells, and a key mediator of inflammatory cell recruitment and transmigration into tissues. Here, we have investigated the effects of the β-1,4-galactosyltransferase inhibitor 5-(5-formylthien-2-yl) UDP-Gal (5-FT UDP-Gal, compound 1) and two close derivatives on the cell surface levels of PSGL-1 on human peripheral blood mononuclear cells (hPBMCs). PSGL-1 levels were studied both under basal conditions, and upon stimulation of hPBMCs with interleukin-1β (IL-1β). Between 1 and 24 hours after IL-1β stimulation, we observed initial PSGL-1 shedding, followed by an increase in PSGL-1 levels on the cell surface, with a maximal window between IL-1β-induced and basal levels after 72 h. All three inhibitors reduce PSGL-1 levels on IL-1β-stimulated cells in a concentration-dependent manner, but show no such effect in resting cells. Compound 1 also affects the cell surface levels of adhesion molecule CD11b in IL-1β-stimulated hPBMCs, but not of glycoproteins CD14 and CCR2. This activity profile may be linked to the inhibition of global Sialyl Lewis presentation on hPBMCs by compound 1, which we have also observed. Although this mechanistic explanation remains hypothetical at present, our results show, for the first time, that small molecules can discriminate between IL-1β-induced and basal levels of cell surface PSGL-1. These findings open new avenues for intervention with PSGL-1 presentation on the cell surface of primed hPBMCs and may have implications for anti-inflammatory drug development. PMID:27233805

Inhibition of IL-17 and IL-23 in Human Keratinocytes by the A3 Adenosine Receptor Agonist Piclidenoson

PubMed Central

Barer, Faina; Itzhak, Inbal; Silverman, Michael H.

2018-01-01

Interleukin-17 and interleukin-23 play major roles in the inflammatory process in psoriasis. The Gi protein-associated A3 adenosine receptor (A3AR) is known to be overexpressed in inflammatory cells and in peripheral blood mononuclear cells (PBMCs) of patients with autoimmune inflammatory conditions. Piclidenoson, a selective agonist at the A3AR, induces robust anti-inflammatory effect in psoriasis patients. In this study, we aimed to explore A3AR expression levels in psoriasis patients and its role in mediating the anti-inflammatory effect of piclidenoson in human keratinocyte cells. A3AR expression levels were evaluated in skin tissue and PBMCs derived from psoriasis patients and healthy subjects. Proliferation assay and the expression of signaling proteins were used to evaluate piclidenoson effect on human keratinocytes (HaCat). High A3AR expression levels were found in a skin biopsy and in PBMCs from psoriasis patients in comparison to healthy subjects. Piclidenoson inhibited the proliferation of HaCat cells through deregulation of the NF-κB signaling pathway, leading to a decrease in interleukin-17 and interleukin-23 expression levels. This effect was counteracted by the specific antagonist MRS 1523. A3AR overexpression in skin and PBMCs of psoriasis patients may be used as a target to inhibit pathological cell proliferation and the production of interleukin-17 and interleukin-23. PMID:29862305

Methamidophos induces cytotoxicity and oxidative stress in human peripheral blood mononuclear cells.

PubMed

Ramirez-Vargas, Marco Antonio; Huerta-Beristain, Gerardo; Guzman-Guzman, Iris Paola; Alarcon-Romero, Luz Del Carmen; Flores-Alfaro, Eugenia; Rojas-Garcia, Aurora Elizabeth; Moreno-Godinez, Ma Elena

2017-01-01

Previous studies have shown that organophosphate pesticide (OP) exposure is associated with oxidative stress. Methamidophos (MET) is an OP widely used in agriculture, which is regarded as a highly toxic pesticide and it is a potent inhibitor of acetylcholinesterase. The aim of this study was to evaluate whether MET can induce oxidative stress at low concentrations in primary cultures of human peripheral blood mononuclear cells (PBMCs). PBMCs from healthy individuals were exposed to MET (0-80 mg/L) for 0-72 h. We performed the MTT and neutral-red assays to assess the cytotoxicity. As indicators of oxidative stress, the levels of reactive oxygen species (ROS) were assessed using flow cytometry, and the malondialdehyde (MDA) and reduced glutathione (GSH) levels were determined. MET decreased the viability of PBMCs in a dose-dependent manner. At concentrations of 3, 10, or 20 mg/L for 24 h, MET increased the ROS production significantly compared with the vehicle control. Similarly, MET increased the levels of MDA at the same concentrations that increased ROS (10 and 20 mg/L); however, no changes in GSH levels were observed. These results suggest that MET increased the generation of oxidative stress in PBMCs. © 2015 Wiley Periodicals, Inc. Environ Toxicol 32: 147-155, 2017. © 2015 Wiley Periodicals, Inc.

IN VITRO STUDIES ON HEME OXYGENASE-1 AND P24 ANTIGEN HIV-1 LEVEL AFTERHYPERBARIC OXYGEN TREATMENTOFHIV-1 INFECTED ON PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS).

PubMed

Budiarti, Retno; Kuntaman; Nasronudin; Suryokusumo; Khairunisa, Siti Qamariyah

2018-01-01

Heme oxygenase-1 (HO-1) is a protein secreted by immune cells as a part of immune response mechanism.HO-1 can be induced by variety agents that causingoxidative stress, such as exposure to 100% oxygenat2,4 ATA pressure.It plays a vital role in maintaining cellular homeostasis.This study was conducted to identify the effect of hyperbaric oxygen exposure in cultured ofPBMCthat infected by HIV-1. Primary culture of PBMCs were isolated from 16 healthy volunteers and HIV-1 infected MT4 cell line by co-culture. The PBMCs were aliquoted into two wells as control group and treatment group. The 16 samples of HIV-1 infected PBMCwere exposed to oxygen at 2,4 ATA in animal hyperbaric chamber forthree times in 30 minutes periods with 5 minutes spacing period, that called 1 session.The Treatment done on 5 sessions within 5 days. 16 samples of HIV-1 infected PMBCs that have no hyperbaric treatment became control group.The supernatant were measured the HO-1 production by ELISA andmRNA expression of HO-1 by real time PCR and the number ofantigen p24 HIV-1by ELISA. The result showed that there was no increasing of HO-1 at both mRNA level and protein level, there was a decreasing number of antigen p24 HIV-1 at the treatment group. In addition, hyperbaric exposure could not increase the expression of HO-1, more over the viral replication might be reduced by other mechanism. Hyperbaric oxygen could increases cellular adaptive response of PBMCs infected HIV-1 through increased expression of proteins that can inhibit HIV viralreplication.

IN VITRO STUDIES ON HEME OXYGENASE-1 AND P24 ANTIGEN HIV-1 LEVEL AFTERHYPERBARIC OXYGEN TREATMENTOFHIV-1 INFECTED ON PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMCS)

PubMed Central

Budiarti, Retno; Kuntaman; Nasronudin; Suryokusumo; Khairunisa, Siti Qamariyah

2018-01-01

Background: Heme oxygenase-1 (HO-1) is a protein secreted by immune cells as a part of immune response mechanism.HO-1 can be induced by variety agents that causingoxidative stress, such as exposure to 100% oxygenat2,4 ATA pressure.It plays a vital role in maintaining cellular homeostasis.This study was conducted to identify the effect of hyperbaric oxygen exposure in cultured ofPBMCthat infected by HIV-1. Material and Methods: Primary culture of PBMCs were isolated from 16 healthy volunteers and HIV-1 infected MT4 cell line by co-culture. The PBMCs were aliquoted into two wells as control group and treatment group. The 16 samples of HIV-1 infected PBMCwere exposed to oxygen at 2,4 ATA in animal hyperbaric chamber forthree times in 30 minutes periods with 5 minutes spacing period, that called 1 session. The Treatment done on 5 sessions within 5 days. 16 samples of HIV-1 infected PMBCs that have no hyperbaric treatment became control group.The supernatant were measured the HO-1 production by ELISA andmRNA expression of HO-1 by real time PCR and the number ofantigen p24 HIV-1by ELISA. Results: The result showed that there was no increasing of HO-1 at both mRNA level and protein level, there was a decreasing number of antigen p24 HIV-1 at the treatment group. In addition, hyperbaric exposure could not increase the expression of HO-1, more over the viral replication might be reduced by other mechanism. Conclusions: Hyperbaric oxygen could increases cellular adaptive response of PBMCs infected HIV-1 through increased expression of proteins that can inhibit HIV viralreplication. PMID:29619425

Distribution and Metabolism of Lipocurc™ (Liposomal Curcumin) in Dog and Human Blood Cells: Species Selectivity and Pharmacokinetic Relevance.

PubMed

Bolger, Gordon T; Licollari, Albert; Tan, Aimin; Greil, Richard; Vcelar, Brigitta; Majeed, Muhammad; Helson, Lawrence

2017-07-01

The aim of this study was to investigate the distribution of curcumin (in the form of Lipocurc™) and its major metabolite tetrahydrocurcumin (THC) in Beagle dog and human red blood cells, peripheral blood mononuclear cells (PBMC) and hepatocytes. Lipocurc™ was used as the source of curcumin for the cell distribution assays. In vitro findings with red blood cells were also compared to in vivo pharmacokinetic data available from preclinical studies in dogs and phase I clinical studies in humans. High levels of curcumin were measured in PBMCs (625.5 ng/g w.w. cell pellet or 7,297 pg/10 6 cells in dog and 353.7 ng/g w.w. cell pellet or 6,809 pg/10 6 cells in human) and in hepatocytes (414.5 ng/g w.w. cell pellet or 14,005 pg/10 6 cells in dog and 813.5 ng/g w.w. cell pellet or 13,780 pg/10 6 cells in human). Lower curcumin levels were measured in red blood cells (dog: 78.4 ng/g w.w. cell pellet or 7.2 pg/10 6 cells, human: 201.5 ng/g w.w. cell pellet or 18.6 pg/10 6 cells). A decrease in the medium concentration of curcumin was observed in red blood cells and hepatocytes, but not in PBMCs. Red blood cell levels of THC were ~5-fold higher in dog compared to human and similar between dog and human for hepatocytes and PBMCs. The ratio of THC to curcumin found in the red blood cell medium following incubation was 6.3 for dog compared to 0.006 for human, while for PBMCs and hepatocytes the ratio of THC to curcumin in the medium did not display such marked species differences. There was an excellent correlation between the in vitro disposition of curcumin and THC following incubation with red blood cells and in vivo plasma levels of curcumin and THC in dog and human following intravenous infusion. The disposition of curcumin in blood cells is, therefore, species-dependent and of pharmacokinetic relevance. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Cell surface expression of CCR5 and other host factors influence the inhibition of HIV-1 infection of human lymphocytes by CCR5 ligands

PubMed Central

Ketas, Thomas J.; Kuhmann, Shawn E.; Palmer, Ashley; Zurita, Juan; He, Weijing; Ahuja, Sunil K.; Klasse, Per Johan; Moore, John P.

2007-01-01

Several CCR5 ligands, including small molecules and monoclonal antibodies (MAbs), are being developed as therapies for infection with strains of human immunodeficiency virus type 1 (HIV-1) that use CCR5 for entry (R5 viruses). The efficacy of such therapies could be influenced by inter-individual differences in host factors, such as CCR5 expression levels. To study this, we used peripheral blood mononuclear cells (PBMCs) from humans and rhesus macaques. The half-maximal inhibitory concentrations (IC50) of the small-molecule CCR5 ligands CMPD167, UK427,857 and SCH-D, and of the PRO 140 MAb, differ by >2 logs in a donor-dependent manner. We studied this variation by using flow cytometry to measure CCR5 expression on PBMCs from six of the human donors: the IC50 values of both SCH-D and PRO 140 correlated with CCR5 expression (R2 = 0.64 and 0.99, respectively). We also determined the efficacy of the CCR5 ligands against HIV-1 infection of HeLa-derived cell lines that express CD4 at the same level but vary 2-fold in CCR5 expression (JC.48 and JC.53 cells). The moderately greater CCR5 expression on the JC.53 than the JC.48 cells was associated with proportionately higher median IC50 values for all four CCR5 ligands but not for a soluble CD4-based inhibitor or a non-nucleoside reverse transcriptase inhibitor. We conclude that differences in CCR5 expression on human PBMCs, which can be affected by CCL3L1 gene dose, may influence the antiviral potency of CCR5 ligands in vitro, but other host factors are also likely to be involved. These host factors may affect the clinical activity of CCR5 inhibitors, including their use as topical microbicides to prevent HIV-1 transmission. PMID:17428518

Bauhinia variegata candida Fraction Induces Tumor Cell Death by Activation of Caspase-3, RIP, and TNF-R1 and Inhibits Cell Migration and Invasion In Vitro

PubMed Central

Santos, K. M.; Silva-Oliveira, R. J.; Pinto, F. E.; Oliveira, B. G.; Chagas, R. C. R.; Romão, W.; Reis, R. M. V.

2018-01-01

Metastasis remains the most common cause of death in cancer patients. Inhibition of metalloproteinases (MMPs) is an interesting approach to cancer therapy because of their role in the degradation of extracellular matrix (ECM), cell-cell, and cell-ECM interactions, modulating key events in cell migration and invasion. Herein, we show the cytotoxic and antimetastatic effects of the third fraction (FR3) from Bauhinia variegata candida (Bvc) stem on human cervical tumor cells (HeLa) and human peripheral blood mononuclear cells (PBMCs). FR3 inhibited MMP-2 and MMP-9 activity, indicated by zymogram. This fraction was cytotoxic to HeLa cells and noncytotoxic to PBMCs and decreased HeLa cell migration and invasion. FR3 is believed to stimulate extrinsic apoptosis together with necroptosis, assessed by western blotting. FR3 inhibited MMP-2 activity in the HeLa supernatant, differently from the control. The atomic mass spectrometry (ESI-MS) characterization suggested the presence of glucopyranosides, D-pinitol, fatty acids, and phenolic acid. These findings provide insight suggesting that FR3 contains components with potential tumor-selective cytotoxic action in addition to the action on the migration of tumor cells, which may be due to inhibition of MMPs. PMID:29770331

The expression and activation of the AIM2 inflammasome correlates with inflammation and disease severity in patients with acute pancreatitis.

PubMed

Algaba-Chueca, Francisco; de-Madaria, Enrique; Lozano-Ruiz, Beatriz; Martínez-Cardona, Claudia; Quesada-Vázquez, Noé; Bachiller, Victoria; Tarín, Fabián; Such, José; Francés, Rubén; Zapater, Pedro; González-Navajas, José M

Acute pancreatitis is an inflammatory disorder of the pancreas that is responsible for significant morbidity and mortality. The inflammasome pathway has acquired significant relevance in the pathogenesis of many inflammatory disorders, but its role in patients with acute pancreatitis still awaits clarification. We performed a prospective study in which 27 patients with acute pancreatitis and 16 healthy controls were included. We isolated peripheral blood mononuclear cells (PBMCs) and we assessed the expression and activation of different inflammasomes as well as their association with the clinical course of the disease. Our results show that PBMCs from patients with acute pancreatitis have elevated expression of several components of the inflammasome complex, including the inflammasome-forming receptor absent in melanoma 2 (AIM2), early during the onset of the disease. Activation of the AIM2 or NLRP3 inflammasomes in PBMCs from patients with acute pancreatitis results in exacerbated IL-1β and IL-18 production compared with PBMCs from healthy controls. Furthermore, both AIM2 mRNA expression and AIM2-mediated production of IL-1β by PBMCs correlated with increased systemic inflammation in these patients. Last, AIM2 expression was further increased in those patients that developed transient or persistent organ failure (moderate or severe acute pancreatitis). Our data demonstrates that AIM2 inflammasome expression and activation is increased early during the course of acute pancreatitis, and suggests that AIM2 activation may affect systemic inflammation and organ failure in these patients. Copyright © 2017 IAP and EPC. Published by Elsevier B.V. All rights reserved.

Humoral and cellular immune responses to Yersinia pestis Pla antigen in humans immunized with live plague vaccine.

PubMed

Feodorova, Valentina A; Lyapina, Anna M; Khizhnyakova, Maria A; Zaitsev, Sergey S; Sayapina, Lidiya V; Arseneva, Tatiana E; Trukhachev, Alexey L; Lebedeva, Svetlana A; Telepnev, Maxim V; Ulianova, Onega V; Lyapina, Elena P; Ulyanov, Sergey S; Motin, Vladimir L

2018-06-01

To establish correlates of human immunity to the live plague vaccine (LPV), we analyzed parameters of cellular and antibody response to the plasminogen activator Pla of Y. pestis. This outer membrane protease is an essential virulence factor that is steadily expressed by Y. pestis. PBMCs and sera were obtained from a cohort of naïve (n = 17) and LPV-vaccinated (n = 34) donors. Anti-Pla antibodies of different classes and IgG subclasses were determined by ELISA and immunoblotting. The analysis of antibody response was complicated with a strong reactivity of Pla with normal human sera. The linear Pla B-cell epitopes were mapped using a library of 15-mer overlapping peptides. Twelve peptides that reacted specifically with sera of vaccinated donors were found together with a major cross-reacting peptide IPNISPDSFTVAAST located at the N-terminus. PBMCs were stimulated with recombinant Pla followed by proliferative analysis and cytokine profiling. The T-cell recall response was pronounced in vaccinees less than a year post-immunization, and became Th17-polarized over time after many rounds of vaccination. The Pla protein can serve as a biomarker of successful vaccination with LPV. The diagnostic use of Pla will require elimination of cross-reactive parts of the antigen.

Phenotyping of peripheral blood mononuclear cells during acute dengue illness demonstrates infection and increased activation of monocytes in severe cases compared to classic dengue fever

PubMed Central

Durbin, Anna P.; Vargas, Maria José; Wanionek, Kimberli; Hammond, Samantha N.; Gordon, Aubree; Rocha, Crisanta; Balmaseda, Angel; Harris, Eva

2008-01-01

In vitro studies have attempted to identify dengue virus (DEN) target cells in peripheral blood; however, extensive phenotyping of peripheral blood mononuclear cells (PBMCs) from dengue patients has not been reported. PBMCs collected from hospitalized children suspected of acute dengue were analyzed for DEN prM, CD32, CD86, CD14, CD11c, CD16, CD209, CCR7, CD4, and CD8 by flow cytometry to detect DEN antigen in PBMCs and to phenotype DEN-positive cells. DEN prM was detected primarily in activated monocytes (CD14+, CD32+, CD86+, CD11c+). A subset of samples analyzed for DEN nonstructural protein 3 (NS3) confirmed that approximately half of DEN antigen-positive cells contained replicating virus. A higher percentage of PBMCs from DHF patients expressed prM, CD86, CD32, and CD11c than did those from DF patients. Increased activation of monocytes and greater numbers of DEN-infected cells were associated with more severe dengue, implicating a role for monocyte activation in dengue immunopathogenesis. PMID:18452966

ADMA induces monocyte adhesion via activation of chemokine receptors in cultured THP-1 cells.

PubMed

Chen, Meifang; Li, Yuanjian; Yang, Tianlun; Wang, Yongjin; Bai, Yongping; Xie, Xiumei

2008-08-01

Asymmetric dimethylarginine (ADMA), an endogenous NOS inhibitor, is also an important inflammatory factor contributing to the development of atherosclerosis (AS). The present study was to test the effect of ADMA on angiotensin (Ang) II-induced monocytic adhesion. Human monocytoid cells (THP-1) or isolated peripheral blood monocyte cells (PBMCs) were incubated with Ang II (10(-6)M) or exogenous ADMA (30 microM) for 4 or 24h in the absence or presence of losartan or antioxidant PDTC. In cultured THP-1 cells, Ang II (10(-6)M) for 24h elevated the level of ADMA in the medium, upregulated the protein expression of protein arginine methyltransferase (PRMT) and decreased the activity of dimethylarginine dimethylaminohydrolase (DDAH). Both of Ang II and ADMA increased monocytic adhesion to human umbilical vein endothelial cells (HUVECs), elevated the levels of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-8 and tumor necrosis factor (TNF)-alpha and upregulated CCR(2) and CXCR(2) mRNA expression, concomitantly with increase in reactive oxygen species (ROS) generation and activation of nuclear factor (NF)-kappaB. Pretreatment with losartan (10 microM) or PDTC (10 microM) abolished the effects mediated by Ang II or ADMA. In isolated PBMCs from healthy individuals, ADMA upregulated the expression of CXCR(2) mRNA, which was attenuated by losartan (10 microM), however, ADMA had no effect on surface protein expression of CCR(2). The present results suggest that ADMA may be involved in monocytic adhesion induced by Ang II via activation of chemokine receptors by ROS/NF-kappaB pathway.

Inflammatory Transcriptome Profiling of Human Monocytes Exposed Acutely to Cigarette Smoke

PubMed Central

Wright, William R.; Parzych, Katarzyna; Crawford, Damian; Mein, Charles; Mitchell, Jane A.; Paul-Clark, Mark J.

2012-01-01

Background Cigarette smoking is responsible for 5 million deaths worldwide each year, and is a major risk factor for cardiovascular and lung diseases. Cigarette smoke contains a complex mixture of over 4000 chemicals containing 1015 free radicals. Studies show smoke is perceived by cells as an inflammatory and xenobiotic stimulus, which activates an immune response. The specific cellular mechanisms driving cigarette smoke-induced inflammation and disease are not fully understood, although the innate immune system is involved in the pathology of smoking related diseases. Methodology/Principle findings To address the impact of smoke as an inflammagen on the innate immune system, THP-1 cells and Human PBMCs were stimulated with 3 and 10% (v/v) cigarette smoke extract (CSE) for 8 and 24 hours. Total RNA was extracted and the transcriptome analysed using Illumina BeadChip arrays. In THP-1 cells, 10% CSE resulted in 80 genes being upregulated and 37 downregulated by ≥1.5 fold after 8 hours. In PBMCs stimulated with 10% CSE for 8 hours, 199 genes were upregulated and 206 genes downregulated by ≥1.5 fold. After 24 hours, the number of genes activated and repressed by ≥1.5 fold had risen to 311 and 306 respectively. The major pathways that were altered are associated with cell survival, such as inducible antioxidants, protein chaperone and folding proteins, and the ubiquitin/proteosome pathway. Conclusions Our results suggest that cigarette smoke causes inflammation and has detrimental effects on the metabolism and function of innate immune cells. In addition, THP-1 cells provide a genetically stable alternative to primary cells for the study of the effects of cigarette smoke on human monocytes. PMID:22363418

Human Memory CD4+ T Cell Immune Responses against Giardia lamblia

PubMed Central

Sørnes, Steinar; Peirasmaki, Dimitra; Svärd, Staffan; Langeland, Nina

2015-01-01

The intestinal protozoan parasite Giardia lamblia may cause severe prolonged diarrheal disease or pass unnoticed as an asymptomatic infection. T cells seem to play an important role in the immune response to Giardia infection, and memory responses may last years. Recently, TH17 responses have been found in three animal studies of Giardia infection. The aim of this study was to characterize the human CD4+ T cell responses to Giardia. Peripheral blood mononuclear cells (PBMCs) were obtained from 21 returning travelers with recent or ongoing giardiasis and 12 low-risk healthy controls and stimulated in vitro with Giardia lamblia proteins. Production of tumor necrosis factor alpha (TNF-α), gamma interferon, interleukin-17A (IL-17A), IL-10, and IL-4 was measured in CD4+ effector memory (EM) T cells after 24 h by flow cytometry. After 6 days of culture, activation and proliferation were measured by flow cytometry, while an array of inflammatory cytokine levels in supernatants were measured with multiplex assays. We found the number of IL-17A-producing CD4+ EM T cells, as well as that of cells simultaneously producing both IL-17A and TNF-α, to be significantly elevated in the Giardia-exposed individuals after 24 h of antigen stimulation. In supernatants of PBMCs stimulated with Giardia antigens for 6 days, we found inflammation-associated cytokines, including 1L-17A, as well as CD4+ T cell activation and proliferation, to be significantly elevated in the Giardia-exposed individuals. We conclude that symptomatic Giardia infection in humans induces a CD4+ EM T cell response of which IL-17A production seems to be an important component. PMID:26376930

Base-modified UDP-sugars reduce cell surface levels of P-selectin glycoprotein 1 (PSGL-1) on IL-1β-stimulated human monocytes.

PubMed

Kanabar, Varsha; Tedaldi, Lauren; Jiang, Jingqian; Nie, Xiaodan; Panina, Irina; Descroix, Karine; Man, Francis; Pitchford, Simon C; Page, Clive P; Wagner, Gerd K

2016-10-01

P-selectin glycoprotein ligand-1 (PSGL-1, CD162) is a cell-surface glycoprotein that is expressed, either constitutively or inducibly, on all myeloid and lymphoid cell lineages. PSGL-1 is implicated in cell-cell interactions between platelets, leukocytes and endothelial cells, and a key mediator of inflammatory cell recruitment and transmigration into tissues. Here, we have investigated the effects of the β-1,4-galactosyltransferase inhibitor 5-(5-formylthien-2-yl) UDP-Gal (5-FT UDP-Gal, compound 1: ) and two close derivatives on the cell surface levels of PSGL-1 on human peripheral blood mononuclear cells (hPBMCs). PSGL-1 levels were studied both under basal conditions, and upon stimulation of hPBMCs with interleukin-1β (IL-1β). Between 1 and 24 hours after IL-1β stimulation, we observed initial PSGL-1 shedding, followed by an increase in PSGL-1 levels on the cell surface, with a maximal window between IL-1β-induced and basal levels after 72 h. All three inhibitors reduce PSGL-1 levels on IL-1β-stimulated cells in a concentration-dependent manner, but show no such effect in resting cells. Compound 1: also affects the cell surface levels of adhesion molecule CD11b in IL-1β-stimulated hPBMCs, but not of glycoproteins CD14 and CCR2. This activity profile may be linked to the inhibition of global Sialyl Lewis presentation on hPBMCs by compound 1: , which we have also observed. Although this mechanistic explanation remains hypothetical at present, our results show, for the first time, that small molecules can discriminate between IL-1β-induced and basal levels of cell surface PSGL-1. These findings open new avenues for intervention with PSGL-1 presentation on the cell surface of primed hPBMCs and may have implications for anti-inflammatory drug development. © The Author 2016. Published by Oxford University Press.

Effect of citral on the cytotoxicity of doxorubicin in human B-lymphoma cells.

PubMed

Dangkong, Darinee; Limpanasithikul, Wacharee

2015-02-01

Doxorubicin is a chemotherapy agent used in non-Hodgkin's lymphoma but side effects limit its use. Citral is a mixture of neral and geranial found in essential oils of lemon grass. We evaluated the activity of citral, doxorubicin, and combination on cytotoxicity, apoptosis, and anti-proliferative effects using human lymphoma Ramos cells. Cells were treated with doxorubicin alone or in combination with citral (10, 20, and 40 μM). Cytotoxic and apoptosis studies were done after 24 and 18 h incubations, respectively. Cytotoxic effects of citral on normal human peripheral blood mononuclear cells (PBMCs) were also investigated for its safety. Changes in the expression of BCL-2 family genes were analyzed by quantitative RT-PCR. Citral had cytotoxicity on cells with an IC50 value of 77.19 ± 4.95 µM. Citral at concentrations of 10, 20, and 40 µM additively increased the cytotoxic and apoptotic effects of doxorubicin, leading to decreased IC50 (µM) of the drug from 2.50 ± 0.01 to 2.16 ± 0.03, 1.90 ± 0.04, and 1.23 ± 0.04, respectively. Enhanced cytotoxicity was not observed in normal human PBMCs. Citral (40 µM) in combination with doxorubicin (1.5 µM) increased the expression of pro-apoptotic protein BAK but significantly decreased the expression of anti-apoptotic protein BCL-XL to 5.26-fold compared with doxorubicin-treated cells. It did not change the anti-proliferative activity of drug. Citral potentiated cytotoxicity of doxorubicin by increasing apoptotic effects. We conclude that citral may have beneficial effects in patients with B cell lymphoma treated with chemotherapy.

Human Innate Immune Responses to Hexamethylene Diisocyanate (HDI) and HDI-Albumin Conjugates

PubMed Central

Wisnewski, Adam V.; Liu, Qing; Liu, Jian; Redlich, Carrie A.

2011-01-01

Background Isocyanates, a leading cause of occupational asthma, are known to induce adaptive immune responses; however, innate immune responses, which generally precede and regulate adaptive immunity, remain largely uncharacterized. Objective Identify and characterize cellular, molecular and systemic innate immune responses induced by hexamethylene diisocyanate (HDI). Methods Human peripheral blood mononuclear cells (PBMCs) were stimulated in vitro with HDI-albumin conjugates or control antigen, and changes in phenotype, gene, and protein expression were characterized by flow cytometry, microarray, Western blot and ELISA. Cell uptake of isocyanate was visualized microscopically using HDI-albumin conjugates prepared with fluorescently-labeled albumin. In vivo, human HDI exposure was performed via specific inhalation challenge, and subsequent changes in PBMCs and serum proteins were measured by flow cytometry and ELISA. Genotypes were determined by PCR. Results Human monocytes take-up HDI-albumin conjugates and undergo marked changes in morphology and gene/protein expression in vitro. The most significant (p 0.007 – 0.05) changes in mircoarray gene expression were noted in lysosomal genes, especially peptidases and proton pumps involved in antigen processing. Chemokines that regulate monocyte/macrophage trafficking (MIF, MCP-1), and pattern recognition receptors that bind chitin (chitinases) and oxidized low-density lipoprotein (CD68) were also increased following isocyanate-albumin exposure. In vivo, HDI exposed subjects exhibited an acute increase in the percentage of PBMCs with the same HDI-albumin responsive phenotype characterized in vitro (HLA-DR+/CD11c+ with altered light scatter properties). An exposure-dependent decrease (46±11%; p<0.015) in serum concentrations of chitinase-3-like-1 was also observed, in individuals that lack the major (type 1) human chitinase (due to genetic polymorphism), but not in individuals possessing at least one functional

Recombinant Brugia malayi pepsin inhibitor (rBm33) exploits host signaling events to regulate inflammatory responses associated with lymphatic filarial infections.

PubMed

Sreenivas, Kirthika; Kalyanaraman, Haripriya; Babu, Subash; Narayanan, Rangarajan Badri

2017-11-01

Prolonged existence of filarial parasites and their molecules within the host modulate the host immune system to instigate their survival and induce inflammatory responses that contribute to disease progression. Recombinant Brugia malayi pepsin inhibitor (rBm33) modulates the host immune responses by skewing towards Th1 responses characterized by secretion of inflammatory molecules such as TNF-α, IL-6, nitric oxide (NO). Here we also specified the molecular signaling events triggered by rBm33 in peripheral blood mononuclear cells (PBMCs) of filarial endemic normals (EN). rBm33 predominantly enhanced the levels of nitric oxide in cultured PBMCs but did not result in oxidative stress to the host cells. Further, rBm33 treatment of human PBMCs resulted in higher GSH/GSSG levels. MYD88 dependent activation was found to be associated with rBm33 specific inflammatory cytokine production. rBm33 triggered intracellular signaling events also involved JNK activation in host PBMCs. In addition, c-Fos and not NF-κB was identified as the transcription factor regulating the expression of inflammatory cytokines in rBm33 stimulated PBMCs. rBm33 marked its role in filarial pathology by altered levels of growth factors but did not have a significant impact on matrix metalloproteinases (MMPs), tissue inhibitors of matrix metalloproteinases (TIMPs) activity of host PBMCs. Thus, the study outlines the signaling network of rBm33 induced inflammatory responses within the host immune cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

Macrophage Migration Inhibitory Factor Enzymatic Activity, Lung Inflammation, and Cystic Fibrosis

PubMed Central

Adamali, Huzaifa; Armstrong, Michelle E.; McLaughlin, Anne Marie; Cooke, Gordon; McKone, Edward; Costello, Christine M.; Gallagher, Charles G.; Leng, Lin; Baugh, John A.; Fingerle-Rowson, Günter; Bucala, Richard J.; McLoughlin, Paul

2012-01-01

Rationale: Macrophage migration inhibitory factor (MIF) is a proinflammatory mediator with unique tautomerase enzymatic activity; the precise function has not been clearly defined. We previously demonstrated that individual patients with cystic fibrosis (CF) who are genetically predisposed to be high MIF producers develop accelerated end-organ injury. Objectives: To characterize the effects of the MIF-CATT polymorphism in patients with CF ex vivo. To investigate the role of MIF’s tautomerase activity in a murine model of Pseudomonas aeruginosa infection. Methods: MIF and tumor necrosis factor (TNF)-α protein levels were assessed in plasma or peripheral blood mononuclear cell (PBMC) supernatants by ELISA. A murine pulmonary model of chronic Pseudomonas infection was used in MIF wild-type mice (mif+/+) and in tautomerase-null, MIF gene knockin mice (mif P1G/P1G). Measurements and Main Results: MIF protein was measured in plasma and PBMCs from 5- and 6-CATT patients with CF; LPS-induced TNF-α production from PBMCs was also assessed. The effect of a specific inhibitor of MIF-tautomerase activity, ISO-1, was investigated in PBMCs. In the murine infection model, total weight loss, differential cell counts, bacterial load, and intraacinar airspace/tissue volume were measured. MIF and TNF-α levels were increased in 6-CATT compared with 5-CATT patients with CF. LPS-induced TNF-α production from PBMCs was attenuated in the presence of ISO-1. In a murine model of Pseudomonas infection, significantly less pulmonary inflammation and bacterial load was observed in mifP1G/P1G compared with mif+/+ mice. Conclusions: MIF-tautomerase activity may provide a novel therapeutic target in patients with chronic inflammatory diseases such as CF, particularly those patients who are genetically predisposed to produce increased levels of this cytokine. PMID:22592805

Tax and Semaphorin 4D Released from Lymphocytes Infected with Human Lymphotropic Virus Type 1 and Their Effect on Neurite Growth

PubMed Central

Quintremil, Sebastián; Alberti, Carolina; Rivera, Matías; Medina, Fernando; Puente, Javier; Cartier, Luis; Ramírez, Eugenio; Tanaka, Yuetsu

2016-01-01

Abstract Human lymphotropic virus type 1 (HTLV-1) is a retrovirus causing HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a neurodegenerative central nervous system (CNS) axonopathy. This virus mainly infects CD4+ T lymphocytes without evidence of neuronal infection. Viral Tax, secreted from infected lymphocytes infiltrated in the CNS, is proposed to alter intracellular pathways related to axonal cytoskeleton dynamics, producing neurological damage. Previous reports showed a higher proteolytic release of soluble Semaphorin 4D (sSEMA-4D) from CD4+ T cells infected with HTLV-1. Soluble SEMA-4D binds to its receptor Plexin-B1, activating axonal growth collapse pathways in the CNS. In the current study, an increase was found in both SEMA-4D in CD4+ T cells and sSEMA-4D released to the culture medium of peripheral blood mononuclear cells (PBMCs) from HAM/TSP patients compared to asymptomatic carriers and healthy donors. After a 16-h culture, infected PBMCs showed significantly higher levels of CRMP-2 phosphorylated at Ser522. The effect was blocked either with anti-Tax or anti-SEMA-4D antibodies. The interaction of Tax and sSEMA-4D was found in secreted medium of PBMCs in patients, which might be associated with a leading role of Tax with the SEMA-4D-Plexin-B1 signaling pathway. In infected PBMCs, the migratory response after transwell assay showed that sSEMA-4D responding cells were CD4+Tax+ T cells with a high CRMP-2 pSer522 content. In the present study, the participation of Tax-sSEMA-4D in the reduction in neurite growth in PC12 cells produced by MT2 (HTLV-1-infected cell line) culture medium was observed. These results lead to the participation of plexins in the reported effects of infected lymphocytes on neuronal cells. PMID:26389656

Tax and Semaphorin 4D Released from Lymphocytes Infected with Human Lymphotropic Virus Type 1 and Their Effect on Neurite Growth.

PubMed

Quintremil, Sebastián; Alberti, Carolina; Rivera, Matías; Medina, Fernando; Puente, Javier; Cartier, Luis; Ramírez, Eugenio; Tanaka, Yuetsu; Valenzuela, M Antonieta

2016-01-01

Human lymphotropic virus type 1 (HTLV-1) is a retrovirus causing HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a neurodegenerative central nervous system (CNS) axonopathy. This virus mainly infects CD4(+) T lymphocytes without evidence of neuronal infection. Viral Tax, secreted from infected lymphocytes infiltrated in the CNS, is proposed to alter intracellular pathways related to axonal cytoskeleton dynamics, producing neurological damage. Previous reports showed a higher proteolytic release of soluble Semaphorin 4D (sSEMA-4D) from CD4(+) T cells infected with HTLV-1. Soluble SEMA-4D binds to its receptor Plexin-B1, activating axonal growth collapse pathways in the CNS. In the current study, an increase was found in both SEMA-4D in CD4(+) T cells and sSEMA-4D released to the culture medium of peripheral blood mononuclear cells (PBMCs) from HAM/TSP patients compared to asymptomatic carriers and healthy donors. After a 16-h culture, infected PBMCs showed significantly higher levels of CRMP-2 phosphorylated at Ser(522). The effect was blocked either with anti-Tax or anti-SEMA-4D antibodies. The interaction of Tax and sSEMA-4D was found in secreted medium of PBMCs in patients, which might be associated with a leading role of Tax with the SEMA-4D-Plexin-B1 signaling pathway. In infected PBMCs, the migratory response after transwell assay showed that sSEMA-4D responding cells were CD4(+)Tax(+) T cells with a high CRMP-2 pSer(522) content. In the present study, the participation of Tax-sSEMA-4D in the reduction in neurite growth in PC12 cells produced by MT2 (HTLV-1-infected cell line) culture medium was observed. These results lead to the participation of plexins in the reported effects of infected lymphocytes on neuronal cells.

A chimeric human-mouse model of Sjögren's syndrome.

PubMed

Young, Nicholas A; Wu, Lai-Chu; Bruss, Michael; Kaffenberger, Benjamin H; Hampton, Jeffrey; Bolon, Brad; Jarjour, Wael N

2015-01-01

Despite recent advances in the understanding of Sjögren's Syndrome (SjS), the pathogenic mechanisms remain elusive and an ideal model for early drug discovery is not yet available. To establish a humanized mouse model of SjS, peripheral blood mononuclear cells (PBMCs) from healthy volunteers or patients with SjS were transferred into immunodeficient NOD-scid IL-2rγ(null) mouse recipients to produce chimeric mice. While no difference was observed in the distribution of cells, chimeric mice transferred with PBMCs from SjS patients produced enhanced cytokine levels, most significantly IFN-γ and IL-10. Histological examination revealed enhanced inflammatory responses in the lacrimal and salivary glands of SjS chimeras, as measured by digital image analysis and blinded histopathological scoring. Infiltrates were primarily CD4+, with minimal detection of CD8+ T-cells and B-cells. These results demonstrate a novel chimeric mouse model of human SjS that provides a unique in vivo environment to test experimental therapeutics and investigate T-cell disease pathology. Copyright © 2014. Published by Elsevier Inc.

Host T-cell primary allosensitization to MHC class-I- and class-II-expressing human cardiac myocytes requires the presence of a second signal.

PubMed

Ansari, A A; Wang, Y C; Kanter, K; Villinger, F; Mayne, A; Sell, K W; Herskowitz, A

1993-06-01

Normal FHCMs, or transformed cell lines derived from FHCMs, such as W1, even after induction of MHC antigens by pretreatment with IFN-gamma, failed to induce proliferation of allogeneic human PBMCs in vitro. To test the hypothesis that antigen-specific T-cell activation and proliferation require not only the binding of the TCR with its ligand, the MHC molecule, but also a second signal that involves the interaction of T-cell surface molecules with their natural ligands on the stimulating cells, a mAb against CD28 was used. Cocultures of allogeneic PBMCs with IFN-gamma-pretreated irradiated FHCMs or the W1 cell line in microtiter plates containing immobilized anti-CD28 mAb induced marked stimulator cells MHC class-II-specific proliferative responses. The W1 cell line and FHCMs failed to express detectable levels of the BB1/B7 molecule (the natural ligand for CD28) as determined by flow microfluorometry or mRNA levels coding for BB1/B7 as determined by RT-PCR. These data suggest that one of the probably reasons for the failure of MHC-expressing cardiac myocytes to induce allogeneic activation is the absence of costimulatory signals.

Age-related change in the retinoid X receptor beta gene expression in peripheral blood mononuclear cells of healthy volunteers: effect of 13-cis retinoic acid supplementation.

PubMed

Brtko, J; Rock, E; Nezbedova, P; Krizanova, O; Dvorcakova, M; Minet-Quinard, R; Farges, M-C; Ribalta, J; Winklhofer-Roob, B M; Vasson, M-P; Macejova, D

2007-01-01

The regulation of cell growth and differentiation and also expression of a number of genes by retinoids are mediated by nuclear retinoid receptors (RARs and/or RXRs). In this study we investigated age-related alteration in both RAR and RXR receptor subtypes gene expression and tissue transglutaminase (tTG) activity before and after supplementation with 13-cis retinoic acid (13cRA) in human peripheral blood mononuclear cells (PBMCs). Healthy men (40) were divided in two groups according to their age (young group: 26.1+/-4.1 years and old group: 65.4+/-3.8 years). Each volunteer received 13cRA (Curacné), 0.5mg/(kgday)) during a period of 4 weeks. We have shown that RXRbeta expression was decreased significantly (p=0.0108) in PBMCs of elderly men when compared to that of young volunteers. Distribution of retinoic acid receptor subtype expression in PBMCs was found in the order: RXRbeta>RARgamma>RXRalpha>RARalpha. The tTG activity in PBMCs reflected a trend to be enhanced after 13-cis retinoic acid supplementation. In conclusion, we demonstrate a significant decrease in the expression of RXRbeta subtype of rexinoid receptors in PBMCs of healthy elderly men. Our data suggest that in healthy elderly men reduction of RXRbeta expression in PBMCs might be a common feature of physiological senescence.

Anti-inflammatory compound resveratrol suppresses homocysteine formation in stimulated human peripheral blood mononuclear cells in vitro.

PubMed

Schroecksnadel, Katharina; Winkler, Christiana; Wirleitner, Barbara; Schennach, Harald; Weiss, Günter; Fuchs, Dietmar

2005-01-01

Inflammation, immune activation and oxidative stress play a major role in the pathogenesis of cardiovascular disorders. In addition to markers of inflammation, moderate hyperhomocysteinemia is an independent risk factor for cardiovascular disease, and there is a link between the activation of immunocompetent cells and the enhanced formation of homocysteine in vitro. Likewise, anti-inflammatory drugs and nutrients rich in antioxidant vitamins are able to reduce cardiovascular risk and to slow down the atherogenic process. Resveratrol, a phenolic antioxidant synthesized in grapes and vegetables and present in wine, has also been supposed to be beneficial for the prevention of cardiovascular events. Apart from its strong antioxidant properties, resveratrol has also been demonstrated to act as an anti-inflammatory agent. In this study the influence of resveratrol on the production of homocysteine by stimulated human peripheral blood mononuclear cells (PBMCs) was investigated. Results were compared to earlier described effects of the anti-inflammatory compounds aspirin and salicylic acid and of the lipid-lowering drug atorvastatin. Stimulation of PBMCs with the mitogens concanavalin A and phytohemagglutinin induced significantly higher homocysteine accumulation in supernatants compared with unstimulated cells. Treatment with 10-100 muM resveratrol suppressed homocysteine formation in a dose-dependent manner. Resveratrol did not influence the release of homocysteine from resting PBMCs. The data suggest that resveratrol may prevent homocysteine accumulation in the blood by suppressing immune activation cascades and the proliferation of mitogen-driven T-cells. The effect of resveratrol to down-regulate the release of homo-cysteine was comparable to the decline of neopterin concentrations in the same experiments. The suppressive effect of resveratrol was very similar to results obtained earlier with aspirin, salicylic acid and atorvastatin; however, it appeared that doses

Volcanic Ash Activates the NLRP3 Inflammasome in Murine and Human Macrophages.

PubMed

Damby, David E; Horwell, Claire J; Baxter, Peter J; Kueppers, Ulrich; Schnurr, Max; Dingwell, Donald B; Duewell, Peter

2017-01-01

Volcanic ash is a heterogeneous mineral dust that is typically composed of a mixture of amorphous (glass) and crystalline (mineral) fragments. It commonly contains an abundance of the crystalline silica (SiO 2 ) polymorph cristobalite. Inhalation of crystalline silica can induce inflammation by stimulating the NLRP3 inflammasome, a cytosolic receptor complex that plays a critical role in driving inflammatory immune responses. Ingested material results in the assembly of NLRP3, ASC, and caspase-1 with subsequent secretion of the interleukin-1 family cytokine IL-1β. Previous toxicology work suggests that cristobalite-bearing volcanic ash is minimally reactive, calling into question the reactivity of volcanically derived crystalline silica, in general. In this study, we target the NLRP3 inflammasome as a crystalline silica responsive element to clarify volcanic cristobalite reactivity. We expose immortalized bone marrow-derived macrophages of genetically engineered mice and primary human peripheral blood mononuclear cells (PBMCs) to ash from the Soufrière Hills volcano as well as representative, pure-phase samples of its primary componentry (volcanic glass, feldspar, cristobalite) and measure NLRP3 inflammasome activation. We demonstrate that respirable Soufrière Hills volcanic ash induces the activation of caspase-1 with subsequent release of mature IL-1β in a NLRP3 inflammasome-dependent manner. Macrophages deficient in NLRP3 inflammasome components are incapable of secreting IL-1β in response to volcanic ash ingestion. Cellular uptake induces lysosomal destabilization involving cysteine proteases. Furthermore, the response involves activation of mitochondrial stress pathways leading to the generation of reactive oxygen species. Considering ash componentry, cristobalite is the most reactive pure-phase with other components inducing only low-level IL-1β secretion. Inflammasome activation mediated by inhaled ash and its potential relevance in chronic pulmonary

Volcanic ash activates the NLRP3 inflammasome in murine and human macrophages

USGS Publications Warehouse

Damby, David; Horwell, Claire J.; Baxter, Peter J.; Kueppers, Ulrich; Schnurr, Max; Dingwell, Donald B.; Duewell, Peter

2018-01-01

Volcanic ash is a heterogeneous mineral dust that is typically composed of a mixture of amorphous (glass) and crystalline (mineral) fragments. It commonly contains an abundance of the crystalline silica (SiO2) polymorph cristobalite. Inhalation of crystalline silica can induce inflammation by stimulating the NLRP3 inflammasome, a cytosolic receptor complex that plays a critical role in driving inflammatory immune responses. Ingested material results in the assembly of NLRP3, ASC, and caspase-1 with subsequent secretion of the interleukin-1 family cytokine IL-1β. Previous toxicology work suggests that cristobalite-bearing volcanic ash is minimally reactive, calling into question the reactivity of volcanically derived crystalline silica, in general. In this study, we target the NLRP3 inflammasome as a crystalline silica responsive element to clarify volcanic cristobalite reactivity. We expose immortalized bone marrow-derived macrophages of genetically engineered mice and primary human peripheral blood mononuclear cells (PBMCs) to ash from the Soufrière Hills volcano as well as representative, pure-phase samples of its primary componentry (volcanic glass, feldspar, cristobalite) and measure NLRP3 inflammasome activation. We demonstrate that respirable Soufrière Hills volcanic ash induces the activation of caspase-1 with subsequent release of mature IL-1β in a NLRP3 inflammasome-dependent manner. Macrophages deficient in NLRP3 inflammasome components are incapable of secreting IL-1β in response to volcanic ash ingestion. Cellular uptake induces lysosomal destabilization involving cysteine proteases. Furthermore, the response involves activation of mitochondrial stress pathways leading to the generation of reactive oxygen species. Considering ash componentry, cristobalite is the most reactive pure-phase with other components inducing only low-level IL-1β secretion. Inflammasome activation mediated by inhaled ash and its potential relevance in chronic pulmonary

Volcanic Ash Activates the NLRP3 Inflammasome in Murine and Human Macrophages

PubMed Central

Damby, David E.; Horwell, Claire J.; Baxter, Peter J.; Kueppers, Ulrich; Schnurr, Max; Dingwell, Donald B.; Duewell, Peter

2018-01-01

Volcanic ash is a heterogeneous mineral dust that is typically composed of a mixture of amorphous (glass) and crystalline (mineral) fragments. It commonly contains an abundance of the crystalline silica (SiO2) polymorph cristobalite. Inhalation of crystalline silica can induce inflammation by stimulating the NLRP3 inflammasome, a cytosolic receptor complex that plays a critical role in driving inflammatory immune responses. Ingested material results in the assembly of NLRP3, ASC, and caspase-1 with subsequent secretion of the interleukin-1 family cytokine IL-1β. Previous toxicology work suggests that cristobalite-bearing volcanic ash is minimally reactive, calling into question the reactivity of volcanically derived crystalline silica, in general. In this study, we target the NLRP3 inflammasome as a crystalline silica responsive element to clarify volcanic cristobalite reactivity. We expose immortalized bone marrow-derived macrophages of genetically engineered mice and primary human peripheral blood mononuclear cells (PBMCs) to ash from the Soufrière Hills volcano as well as representative, pure-phase samples of its primary componentry (volcanic glass, feldspar, cristobalite) and measure NLRP3 inflammasome activation. We demonstrate that respirable Soufrière Hills volcanic ash induces the activation of caspase-1 with subsequent release of mature IL-1β in a NLRP3 inflammasome-dependent manner. Macrophages deficient in NLRP3 inflammasome components are incapable of secreting IL-1β in response to volcanic ash ingestion. Cellular uptake induces lysosomal destabilization involving cysteine proteases. Furthermore, the response involves activation of mitochondrial stress pathways leading to the generation of reactive oxygen species. Considering ash componentry, cristobalite is the most reactive pure-phase with other components inducing only low-level IL-1β secretion. Inflammasome activation mediated by inhaled ash and its potential relevance in chronic pulmonary

Defense Human Resources Activity > PERSEREC

Science.gov Websites

Skip to main content (Press Enter). Toggle navigation Defense Human Resources Activity Search Search Defense Human Resources Activity: Search Search Defense Human Resources Activity: Search Defense Human Resources Activity U.S. Department of Defense Defense Human Resources Activity Overview

4α-Methylated steroids with cytotoxic activity from the soft coral Litophyton mollis.

PubMed

Zovko Končić, Marijana; Ioannou, Efstathia; Sawadogo, Wamtinga Richard; Abdel-Razik, Ayman F; Vagias, Constantinos; Diederich, Marc; Roussis, Vassilios

2016-11-01

Seven new (1-3, 5 and 8-10) and three previously reported (4, 6 and 7) 4α-methylated steroids were isolated from the organic extract of the gorgonian Litophyton mollis. The structures and the relative configurations of the isolated natural products were determined on the basis of extensive analyses of their NMR and MS data. Metabolites 1 and 5-8 exhibited cytotoxic activity against K562 human chronic myelogenous leukemia cells with IC 50 values below 10μM, while at the same time displaying low toxicity against healthy PBMCs. Copyright © 2016 Elsevier Inc. All rights reserved.

Proteomic analysis of Fasciola hepatica excretory and secretory products (FhESPs) involved in interacting with host PBMCs and cytokines by shotgun LC-MS/MS.

PubMed

Liu, Qing; Huang, Si-Yang; Yue, Dong-Mei; Wang, Jin-Lei; Wang, Yujian; Li, Xiangrui; Zhu, Xing-Quan

2017-02-01

Fasciola hepatica is a helminth parasite with a worldwide distribution, which can cause chronic liver disease, fasciolosis, leading to economic losses in the livestock and public health in many countries. Control is mostly reliant on the use of drugs, and as a result, drug resistance has now emerged. The identification of F. hepatica genes involved in interaction between the parasite and host immune system is utmost important to elucidate the evasion mechanisms of the parasite and develop more effective strategies against fasciolosis. In this study, we aimed to identify molecules in F. hepatica excretory and secretory products (FhESPs) interacting with the host peripheral blood mononuclear cells (PBMCs), Th1-like cytokines (IL2 and IFN-γ), and Th17-like cytokines (IL17) by Co-IP combined with tandem mass spectrometry. The results showed that 14, 16, and 9 proteins in FhESPs could bind with IL2, IL17, and IFN-γ, respectively, which indicated that adult F. hepatica may evade the host immune responses through directly interplaying with cytokines. In addition, nine proteins in FhESPs could adhere to PBMCs. Our findings provided potential targets as immuno-regulators, and will be helpful to elucidate the molecular basis of host-parasite interactions and search for new potential proteins as vaccine and drug target candidates.

Stabilizing short-lived Schiff base derivatives of 5-aminouracils that activate mucosal-associated invariant T cells

NASA Astrophysics Data System (ADS)

Mak, Jeffrey Y. W.; Xu, Weijun; Reid, Robert C.; Corbett, Alexandra J.; Meehan, Bronwyn S.; Wang, Huimeng; Chen, Zhenjun; Rossjohn, Jamie; McCluskey, James; Liu, Ligong; Fairlie, David P.

2017-03-01

Mucosal-associated invariant T (MAIT) cells are activated by unstable antigens formed by reactions of 5-amino-6-D-ribitylaminouracil (a vitamin B2 biosynthetic intermediate) with glycolysis metabolites such as methylglyoxal. Here we show superior preparations of antigens in dimethylsulfoxide, avoiding their rapid decomposition in water (t1/2 1.5 h, 37 °C). Antigen solution structures, MAIT cell activation potencies (EC50 3-500 pM), and chemical stabilities are described. Computer analyses of antigen structures reveal stereochemical and energetic influences on MAIT cell activation, enabling design of a water stable synthetic antigen (EC50 2 nM). Like native antigens, this antigen preparation induces MR1 refolding and upregulates surface expression of human MR1, forms MR1 tetramers that detect MAIT cells in human PBMCs, and stimulates cytokine expression (IFNγ, TNF) by human MAIT cells. These antigens also induce MAIT cell accumulation in mouse lungs after administration with a co-stimulant. These chemical and immunological findings provide new insights into antigen properties and MAIT cell activation.

A High-Throughput Assay for Screening of Natural Products that Enhanced Tumoricidal Activity of NK Cells.

PubMed

Gong, Chenyuan; Ni, Zhongya; Yao, Chao; Zhu, Xiaowen; Ni, Lulu; Wang, Lixin; Zhu, Shiguo

2015-01-01

Recently, immunotherapy has shown a lot of promise in cancer treatment and different immune cell types are involved in this endeavor. Among different immune cell populations, NK cells are also an important component in unleashing the therapeutic activity of immune cells. Therefore, in order to enhance the tumoricidal activity of NK cells, identification of new small-molecule natural products is important. Despite the availability of different screening methods for identification of natural products, a simple, economic and high-throughput method is lacking. Hence, in this study, we have developed a high-throughput assay for screening and indentifying natural products that can enhance NK cell-mediated killing of cancer cells. We expanded human NK cell population from human peripheral blood mononuclear cells (PBMCs) by culturing these PBMCs with membrane-bound IL-21 and CD137L engineered K562 cells. Next, expanded NK cells were co-cultured with non-small cell lung cancer (NSCLC) cells with or without natural products and after 24 h of co-culturing, harvested supernatants were analyzed for IFN-γ secretions by ELISA method. We screened 502 natural products and identified that 28 candidates has the potential to induce IFN-γ secretion by NK cells to varying degrees. Among the 28 natural product candidates, we further confirmed and analyzed the potential of one molecule, andrographolide. It actually increased IFN-γ secretion by NK cells and enhanced NK cell-mediated killing of NSCLC cells. Our results demonstrated that this IFN-γ based high-throughput assay for screening of natural products for NK cell tumoricidal activity is a simple, economic and reliable method.

CXCL16/CXCR6-mediated adhesion of human peripheral blood mononuclear cells to inflamed endothelium.

PubMed

Linke, Bona; Meyer Dos Santos, Sascha; Picard-Willems, Bettina; Keese, Michael; Harder, Sebastian; Geisslinger, Gerd; Scholich, Klaus

2017-06-21

The endothelial chemokine CXC motif ligand 16 (CXCL16) is involved in the recruitment and firm adhesion of CXCR6 + cells to the atherosclerosis-prone aortic vessel wall. Recently we showed that CXCR6 + platelets from flowing blood attach to CXCL16 expressed by activated endothelium on the luminal side of the blood vessel. With this study we supplement these findings with the observation that platelets bound to the inflamed endothelium are presenting CXCR6 to CXCL16-positive peripheral blood mononuclear cells (PBMCs) and, thus, are mediating an increased adhesion of PBMCs to the arterial wall. Furthermore we identified endothelial CXCL16 as an important adhesion molecule promoting the firm adhesion of CXCR6-positive PBMCs to inflamed endothelium. Our results demonstrate that endothelial CXCL16 as well as platelet CXCR6 are acting as potent PBMC-adhesion ligands, inducing PBMC-adhesion to the atherosclerosis-prone vessel wall and thus promoting the progression of atherosclerosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

HIV-1 isolation from infected peripheral blood mononuclear cells.

PubMed

Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A; Schuitemaker, Hanneke; Scarlatti, Gabriella

2014-01-01

Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and not in tumor derived cell lines. The procedure involves culture of PBMCs from an infected patient with phytohemagglutinin (PHA)-stimulated PBMC from seronegative donors, which provide susceptible target cells for HIV replication. HIV can be isolated from the bulk population of PBMCs or after cloning of the cells to obtain viral biological clones. Viral production is determined with p24 antigen (Ag) detection assays or with reverse transcriptase (RT) activity assay. Once isolated, HIV-1 can be propagated by infecting PHA-stimulated PBMCs from healthy donors. Aliquots from culture with a high production of virus are stored for later use.

Olive Oil Phenolics Prevent Oxysterol‐Induced Proinflammatory Cytokine Secretion and Reactive Oxygen Species Production in Human Peripheral Blood Mononuclear Cells, Through Modulation of p38 and JNK Pathways

PubMed Central

Deiana, Monica; Spencer, Jeremy P. E.; Corona, Giulia

2017-01-01

Scope The aim of the present study was to investigate the ability of extra virgin olive oil (EVOO) polyphenols to counteract the proinflammatory effects induced by dietary and endogenous oxysterols in ex vivo immune cells. Methods and results Peripheral blood mononuclear cells (PBMCs), separated from the whole blood of healthy donors, were utilized and were stimulated with an oxysterols mixture, in the presence of physiologically relevant concentrations of the EVOO polyphenols, hydroxytyrosol, tyrosol, and homovanillic alcohol. Oxysterols significantly increased the production of proinflammatory cytokines, interleukin‐1β, regulated on activation, normal T‐cell expressed and secreted and macrophage migration inhibitory factor in ex vivo cultured PBMCs. Increased levels of reactive oxygen species (ROS) were also detected along with increased phosphorylation of the p38 and JNK. All phenolic compounds significantly reduced cytokine secretion induced by the oxysterols and inhibited ROS production and mitogen activated protein kinase phosphorylation. Conclusions These results suggest that extra virgin olive oil polyphenols modulate the immune response induced by dietary and endogenous cholesterol oxidation products in human immune cells and may hold benefit in controlling chronic immune and/or inflammatory processes. PMID:28815947

The circular RNA of peripheral blood mononuclear cells: Hsa_circ_0005836 as a new diagnostic biomarker and therapeutic target of active pulmonary tuberculosis.

PubMed

Zhuang, Ze-Gang; Zhang, Jun-Ai; Luo, Hou-Long; Liu, Gan-Bin; Lu, Yuan-Bin; Ge, Nan-Hai; Zheng, Bi-Ying; Li, Rui Xi; Chen, Chen; Wang, Xin; Liu, Yu-Qing; Liu, Feng-Hui; Zhou, Yong; Cai, Xiao-Zhen; Chen, Zheng W; Xu, Jun-Fa

2017-10-01

It has been reported that circular RNA (circRNA) is associated with human cancer. However, few studies have been reported in active pulmonary tuberculosis (APTB). The global circRNA expression was detected in the peripheral blood mononuclear cells (PBMCs) of APTB patients (n=5) and health controls (HC) (n=5) by using high-throughput sequencing. According to the systematical bioinformatics analysis, the basic content of circRNAs and their fold changes in the two groups were calculated. We selected 6 significant differentially expressed circRNAs, hsa_circ_0005836, hsa_circ_0009128, hsa_circ_0003519, hsa_circ_0023956, hsa_circ_0078768, and hsa_circ_0088452 and validated the expression in PBMCs from APTB (n=10) and HC (n=10) by real-time quantitative reverse transcription-polymerase chain reactions (qRT-PCRs). Further, the verification of these specific circRNAs (hsa_circ_0005836 and hsa_circ_0009128) between APTB (n=34) and HC (n=30) in PBMCs was also conducted by qRT-PCRs. The RNA-seq data showed the significant differential expression of the 523 circRNAs between the APTB and HC groups (199 circRNAs were significantly up-regulated and 324 circRNAs were down-regulated). Hsa_circ_0005836 and hsa_circ_0009128 expression was significantly down-regulated in the PBMCs of APTB (P<0.05) in the samples of APTB compared to HC in our study. The gene ontology based enrichment analysis of the circRNA-miRNA-mRNAs network showed that cellular catabolic process (P=7.10E-08), regulation of metabolic process (P=2.10E-06), catalytic activity (P=3.67E-08), protein binding (P=1.71E-07), cell part (P=3.46E-06), intracellular part (P=1.71E-07), and intracellular (P=3.67E-08) were recognized in the comparisons between APTB and HC. Based on KEGG analysis, HTLV-I infection, regulation of actin cytoskeleton, neurotrophin signaling pathway and mTOR signaling pathway were relevant during tuberculosis bacillus infection. We found for the first time that hsa_circ_0005836 and hsa_circ_0009128 were

HUMAN EXPOSURE ACTIVITY PATTERNS

EPA Science Inventory

Human activity/uptake rate data are necessary to estimate potential human exposure and intake dose to environmental pollutants and to refine human exposure models. Personal exposure monitoring studies have demonstrated the critical role that activities play in explaining and pre...

Transcription of PR3 and Related Myelopoiesis Genes in Peripheral Blood Mononuclear Cells in Active Wegener's Granulomatosis

PubMed Central

Cheadle, Chris; Berger, Alan E.; Andrade, Felipe; James, Regina; Johnson, Kristen; Watkins, Tonya; Park, Jin Kyun; Chen, Yu-Chi; Ehrlich, Eva; Mullins, Marissa; Chrest, Francis; Barnes, Kathleen C.; Levine, Stuart M.

2010-01-01

Objective Wegener's granulomatosis (WG) is a systemic inflammatory disease causing substantial morbidity. This study seeks to understand the biology underlying WG, and to discover markers of disease activity useful in prognosis and treatment guidance. Methods Gene expression profiling was performed using total RNA from PBMC and granulocyte fractions from 41 WG patients and 23 healthy controls. Gene set enrichment analysis (GSEA) was performed to search for candidate WG-associated molecular pathways and disease activity biomarkers. Principal component analysis (PCA) was used to visualize relationships between subgroups of WG patients and controls. Longitudinal changes in PR3 expression were evaluated using RT-PCR, and clinical outcomes including remission status and disease activity were determined using the BVAS-WG. Results We identified 86 genes significantly up-regulated in WG PBMCs and 40 in WG PMNs relative to controls. Genes up-regulated in WG PBMCs were involved in myeloid differentiation, and included the WG autoantigen, PR3. The coordinated regulation of myeloid differentiation genes was confirmed by gene set analysis. Median expression values of the 86 WG PBMC genes were associated with disease activity (p=1.3 × 10−4), and patients expressing these genes at a lower level were only modestly different from healthy controls (p=0.07). PR3 transcription was significantly up-regulated in the PBMCs (p=1.3 ×10−5, FDR=0.002), but not in the PMNs (p=0.03, FDR=0.28) of WG patients, and changes in BVAS-WG tracked with PBMC PR3 RNA levels in a preliminary longitudinal analysis. Conclusion Transcription of PR3 and related myeloid differentiation genes in PBMCs may represent novel markers of disease activity in WG. PMID:20155833

An in vitro model of Mycobacterium leprae induced granuloma formation.

PubMed

Wang, Hongsheng; Maeda, Yumi; Fukutomi, Yasuo; Makino, Masahiko

2013-06-20

Leprosy is a contagious and chronic systemic granulomatous disease caused by Mycobacterium leprae. In the pathogenesis of leprosy, granulomas play a key role, however, the mechanisms of the formation and maintenance of M. leprae granulomas are still not clearly understood. To better understand the molecular physiology of M. leprae granulomas and the interaction between the bacilli and human host cells, we developed an in vitro model of human granulomas, which mimicked the in vivo granulomas of leprosy. Macrophages were differentiated from human monocytes, and infected with M. leprae, and then cultured with autologous human peripheral blood mononuclear cells (PBMCs). Robust granuloma-like aggregates were obtained only when the M. leprae infected macrophages were co-cultured with PBMCs. Histological examination showed M. leprae within the cytoplasmic center of the multinucleated giant cells, and these bacilli were metabolically active. Macrophages of both M1 and M2 types co-existed in the granuloma like aggregates. There was a strong relationship between the formation of granulomas and changes in the expression levels of cell surface antigens on macrophages, cytokine production and the macrophage polarization. The viability of M. leprae isolated from granulomas indicated that the formation of host cell aggregates benefited the host, but the bacilli also remained metabolically active. A simple in vitro model of human M. leprae granulomas was established using human monocyte-derived macrophages and PBMCs. This system may be useful to unravel the mechanisms of disease progression, and subsequently develop methods to control leprosy.

Human milk miRNAs primarily originate from the mammary gland resulting in unique miRNA profiles of fractionated milk

PubMed Central

Alsaweed, Mohammed; Lai, Ching Tat; Hartmann, Peter E.; Geddes, Donna T.; Kakulas, Foteini

2016-01-01

Human milk (HM) contains regulatory biomolecules including miRNAs, the origin and functional significance of which are still undetermined. We used TaqMan OpenArrays to profile 681 mature miRNAs in HM cells and fat, and compared them with maternal peripheral blood mononuclear cells (PBMCs) and plasma, and bovine and soy infant formulae. HM cells and PBMCs (292 and 345 miRNAs, respectively) had higher miRNA content than HM fat and plasma (242 and 219 miRNAs, respectively) (p < 0.05). A strong association in miRNA profiles was found between HM cells and fat, whilst PBMCs and plasma were distinctly different to HM, displaying marked inter-individual variation. Considering the dominance of epithelial cells in mature milk of healthy women, these results suggest that HM miRNAs primarily originate from the mammary epithelium, whilst the maternal circulation may have a smaller contribution. Our findings demonstrate that unlike infant formulae, which contained very few human miRNA, HM is a rich source of lactation-specific miRNA, which could be used as biomarkers of the performance and health status of the lactating mammary gland. Given the recently identified stability, uptake and functionality of food- and milk-derived miRNA in vivo, HM miRNA are likely to contribute to infant protection and development. PMID:26854194

Human milk miRNAs primarily originate from the mammary gland resulting in unique miRNA profiles of fractionated milk.

PubMed

Alsaweed, Mohammed; Lai, Ching Tat; Hartmann, Peter E; Geddes, Donna T; Kakulas, Foteini

2016-02-08

Human milk (HM) contains regulatory biomolecules including miRNAs, the origin and functional significance of which are still undetermined. We used TaqMan OpenArrays to profile 681 mature miRNAs in HM cells and fat, and compared them with maternal peripheral blood mononuclear cells (PBMCs) and plasma, and bovine and soy infant formulae. HM cells and PBMCs (292 and 345 miRNAs, respectively) had higher miRNA content than HM fat and plasma (242 and 219 miRNAs, respectively) (p < 0.05). A strong association in miRNA profiles was found between HM cells and fat, whilst PBMCs and plasma were distinctly different to HM, displaying marked inter-individual variation. Considering the dominance of epithelial cells in mature milk of healthy women, these results suggest that HM miRNAs primarily originate from the mammary epithelium, whilst the maternal circulation may have a smaller contribution. Our findings demonstrate that unlike infant formulae, which contained very few human miRNA, HM is a rich source of lactation-specific miRNA, which could be used as biomarkers of the performance and health status of the lactating mammary gland. Given the recently identified stability, uptake and functionality of food- and milk-derived miRNA in vivo, HM miRNA are likely to contribute to infant protection and development.

Basic aminopeptidase activity is an emerging biomarker in collagen-induced rheumatoid arthritis.

PubMed

Mendes, Mariana Trivilin; Murari-do-Nascimento, Stephanie; Torrigo, Isis Rossetti; Alponti, Rafaela Fadoni; Yamasaki, Simone Cristina; Silveira, Paulo Flavio

2011-04-11

The objective of this study was to investigate the catalytic activity of basic aminopeptidase (APB) and its association with periarticular edema and circulating tumor necrosis factor (TNF)-alpha and type II collagen (CII) antibodies (AACII) in a rat model of rheumatoid arthritis (RA) induced by CII (CIA). Edema does not occur in part of CII-treated, even when AACII is higher than in control. TNF-alpha is detectable only in edematous CII-treated. APB in synovial membrane is predominantly a membrane-bound activity also present in soluble form and with higher activity in edematous than in non-edematous CII-treated or control. Synovial fluid and blood plasma have lower APB in non-edematous than in edematous CII-treated or control. In peripheral blood mononuclear cells (PBMCs) the highest levels of APB are found in soluble form in control and in membrane-bound form in non-edematous CII-treated. CII treatment distinguishes two categories of rats: one with arthritic edema, high AACII, detectable TNF-alpha, high soluble and membrane-bound APB in synovial membrane and low APB in the soluble fraction of PBMCs, and another without edema and with high AACII, undetectable TNF-alpha, low APB in the synovial fluid and blood plasma and high APB in the membrane-bound fraction of PBMCs. Data suggest that APB and CIA are strongly related. 2011 Elsevier B.V. All rights reserved.

Advanced feeder-free generation of induced pluripotent stem cells directly from blood cells.

PubMed

Trokovic, Ras; Weltner, Jere; Nishimura, Ken; Ohtaka, Manami; Nakanishi, Mahito; Salomaa, Veikko; Jalanko, Anu; Otonkoski, Timo; Kyttälä, Aija

2014-12-01

Generation of validated human induced pluripotent stem cells (iPSCs) for biobanking is essential for exploring the full potential of iPSCs in disease modeling and drug discovery. Peripheral blood mononuclear cells (PBMCs) are attractive targets for reprogramming, because blood is collected by a routine clinical procedure and is a commonly stored material in biobanks. Generation of iPSCs from blood cells has previously been reported using integrative retroviruses, episomal Sendai viruses, and DNA plasmids. However, most of the published protocols require expansion and/or activation of a specific cell population from PBMCs. We have recently collected a PBMC cohort from the Finnish population containing more than 2,000 subjects. Here we report efficient generation of iPSCs directly from PBMCs in feeder-free conditions in approximately 2 weeks. The produced iPSC clones are pluripotent and transgene-free. Together, these properties make this novel method a powerful tool for large-scale reprogramming of PBMCs and for iPSC biobanking. ©AlphaMed Press.

Recombinant Human T-Cell Leukemia Virus Types 1 and 2 Tax Proteins Induce High Levels of CC-Chemokines and Downregulate CCR5 in Human Peripheral Blood Mononuclear Cells

PubMed Central

Barrios, Christy S.; Abuerreish, Muna; Lairmore, Michael D.; Castillo, Laura; Giam, Chou-Zen

2011-01-01

Abstract Human T-cell leukemia viruses types 1 (HTLV-1) and 2 (HTLV-2) produce key transcriptional regulatory gene products, known as Tax1 and Tax2, respectively. Tax1 and Tax2 transactivate multiple host genes involved in cellular immune responses within the cellular microenvironment, including induction of genes encoding expression of CC-chemokines. It is speculated that HTLV Tax proteins may act as immune modulators. In this study, recombinant Tax1 and Tax2 proteins were tested for their effects on the viability of cultured peripheral blood mononuclear cells (PBMCs), and their ability to induce expression of CC-chemokines and to downregulate the level of CCR5 expression in PBMCs. PBMCs obtained from uninfected donors were cultured in a range of Tax1 and Tax2 concentrations (10–100 pM), and supernatant fluids were harvested at multiple time points for quantitative determinations of MIP-1α/CCL3, MIP-1β/CCL4, and RANTES/CCL5. Treatment of PBMCs with Tax1 and Tax2 proteins (100 pM) resulted in a significant increase in viability over a 7-d period compared to controls (p<0.01). Both Tax1 and Tax2 induced high levels of all three CC-chemokines over the dosing range compared to mock-treated controls (p<0.05). The gated population of lymphocytes treated with Tax2, as well as lymphocytes from HTLV-2-infected donors, showed a significantly lower percentage of CCR5-positive cells compared to those of uninfected donors and from mock-treated lymphocytes, respectively (p<0.05). These results suggest that Tax1 and Tax2 could promote innate immunity in the extracellular environment during HTLV-1 and HTLV-2 infections via CC-chemokine ligands and receptors. PMID:22111594

Evaluation of DNA-damaging potential of bisphenol A and its selected analogs in human peripheral blood mononuclear cells (in vitro study).

PubMed

Mokra, Katarzyna; Kuźmińska-Surowaniec, Agnieszka; Woźniak, Katarzyna; Michałowicz, Jaromir

2017-02-01

In the present study, we have investigated DNA-damaging potential of BPA and its analogs, i.e. bisphenol S (BPS), bisphenol F (BPF) and bisphenol AF (BPAF) in human peripheral blood mononuclear cells (PBMCs) using the alkaline and neutral versions of the comet assay, which allowed to evaluate DNA single strand-breaks (SSBs) and double strand-breaks (DSBs). The use of the alkaline version of comet assay made also possible to analyze the kinetics of DNA repair in PBMCs after exposure of the cells to BPA or its analogs. We have observed an increase in DNA damage in PBMCs treated with BPA or its analogs in the concentrations ranging from 0.01 to 10 μg/ml after 1 and 4 h incubation. It was noted that bisphenols studied caused DNA damage mainly via SSBs, while DNA fragmentation via double DSBs was low. The strongest changes in DNA damage were provoked by BPA and particularly BPAF, which were capable of inducing SSBs even at 0.01 μg/ml, while BPS caused the lowest changes (only at 10 μg/ml). We have also observed that PBMCs significantly repaired bisphenols-induced DNA damage but they were unable (excluding cells treated with BPS) to repair totally DNA breaks. Copyright © 2016 Elsevier Ltd. All rights reserved.

Multiple correlation analyses revealed complex relationship between DNA methylation and mRNA expression in human peripheral blood mononuclear cells.

PubMed

Xie, Fang-Fei; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Hong; Wu, Jian; Guo, Yu-Fan; Zeng, Ke-Qin; Wang, Ming-Jun; Zhu, Xiao-Wei; Xia, Wei; Wang, Lan; He, Pei; Bing, Peng-Fei; Lu, Xin; Zhang, Yong-Hong; Lei, Shu-Feng

2018-01-01

DNA methylation is an important regulator on the mRNA expression. However, a genome-wide correlation pattern between DNA methylation and mRNA expression in human peripheral blood mononuclear cells (PBMCs) is largely unknown. The comprehensive relationship between mRNA and DNA methylation was explored by using four types of correlation analyses and a genome-wide methylation-mRNA expression quantitative trait locus (eQTL) analysis in PBMCs in 46 unrelated female subjects. An enrichment analysis was performed to detect biological function for the detected genes. Single pair correlation coefficient (r T1 ) between methylation level and mRNA is moderate (-0.63-0.62) in intensity, and the negative and positive correlations are nearly equal in quantity. Correlation analysis on each gene (T4) found 60.1% genes showed correlations between mRNA and gene-based methylation at P < 0.05 and more than 5.96% genes presented very strong correlation (R T4  > 0.8). Methylation sites have regulation effects on mRNA expression in eQTL analysis, with more often observations in region of transcription start site (TSS). The genes under significant methylation regulation both in correlation analysis and eQTL analysis tend to cluster to the categories (e.g., transcription, translation, regulation of transcription) that are essential for maintaining the basic life activities of cells. Our findings indicated that DNA methylation has predictive regulation effect on mRNA with a very complex pattern in PBMCs. The results increased our understanding on correlation of methylation and mRNA and also provided useful clues for future epigenetic studies in exploring biological and disease-related regulatory mechanisms in PBMC.

Modulation of T Cell Activation by Malignant Melanoma Initiating Cells

PubMed Central

Schatton, Tobias; Schütte, Ute; Frank, Natasha Y.; Zhan, Qian; Hoerning, André; Robles, Susanne C.; Zhou, Jun; Hodi, F. Stephen; Spagnoli, Giulio C.; Murphy, George F.; Frank, Markus H.

2010-01-01

Highly immunogenic cancers such as malignant melanoma are capable of inexorable tumor growth despite the presence of antitumor immunity. This raises the possibility that only a restricted minority of tumorigenic malignant cells might possess the phenotypic and functional characteristics to modulate tumor-directed immune activation. Here we provide evidence supporting this hypothesis, by demonstrating that tumorigenic ABCB5+ malignant melanoma-initiating cells (MMICs) possess the capacity to preferentially inhibit interleukin (IL)-2-dependent T cell activation and to support, in a B7.2-dependent manner, regulatory T (Treg) cell induction. Compared to melanoma bulk populations, ABCB5+ MMICs expressed lower levels of the major histocompatibility complex (MHC) class I, showed aberrant positivity for MHC class II, and exhibited lower expression levels of the melanoma-associated antigens (MAAs) MART-1, ML-IAP, NY-ESO-1, and MAGE-A. In addition, tumorigenic ABCB5+ subpopulations preferentially expressed the costimulatory molecules B7.2 and PD-1 in both established melanoma xenografts and clinical tumor specimens in vivo. In immune activation assays, ABCB5+ melanoma cells inhibited mitogen-dependent human peripheral blood mononuclear cell (PBMC) proliferation and IL-2 production more efficiently than ABCB5− populations. Moreover, coculture with ABCB5+ MMICs increased, in a B7.2 signalling-dependent manner, CD4+CD25+FoxP3+ Treg cell abundance and IL-10 production by mitogen-activated PBMCs. Consistent with these findings, ABCB5+ melanoma subsets also preferentially inhibited IL-2 production and induced IL-10 secretion by cocultured patient-derived, syngeneic PBMCs. Our findings identify novel T cell-modulatory functions of ABCB5+ melanoma subpopulations and suggest specific roles for MMICs in the evasion of antitumor immunity and in cancer immunotherapeutic resistance. PMID:20068175

Comparison of Diabetic and Non-diabetic Human Leukocytic Responses to Different Capsule Types of Klebsiella pneumoniae Responsible for Causing Pyogenic Liver Abscess

PubMed Central

Lee, I. Russel; Sng, Ethel; Lee, Kok-Onn; Molton, James S.; Chan, Monica; Kalimuddin, Shirin; Izharuddin, Ezlyn; Lye, David C.; Archuleta, Sophia; Gan, Yunn-Hwen

2017-01-01

The major risk factor for Klebsiella liver abscess (KLA) is type 2 diabetes mellitus (DM), but the immunological mechanisms involved in the increased susceptibility are poorly defined. We investigated the responses of neutrophils and peripheral blood mononuclear cells (PBMCs) to hypervirulent Klebsiella pneumoniae (hvKP), the causative agent of KLA. DNA and myeloperoxidase levels were elevated in the plasma of KLA patients compared to uninfected individuals indicating neutrophil activation, but diabetic status had no effect on these neutrophil extracellular trap (NET) biomarkers in both subject groups. Clinical hvKP isolates universally stimulated KLA patient neutrophils to produce NETs ex vivo, regardless of host diabetic status. Ability of representative capsule types (K1, K2, and non-K1/K2 strains) to survive intra- and extra-cellular killing by type 2 DM and healthy neutrophils was subsequently examined. Key findings were: (1) type 2 DM and healthy neutrophils exhibited comparable total, phagocytic, and NETs killing against hvKP, (2) phagocytic and NETs killing were equally effective against hvKP, and (3) hypermucoviscous K1 and K2 strains were more resistant to total, phagocytic, and NETs killing compared to the non-mucoviscous, non-K1/K2 strain. The cytokine response and intracellular killing ability of type 2 DM as well as healthy PBMCs upon encounter with the different capsule types was also examined. Notably, the IL-12–IFNγ axis and its downstream chemokines MIG, IP-10, and RANTES were produced at slightly lower levels by type 2 DM PBMCs than healthy PBMCs in response to representative K1 and non-K1/K2 strains. Furthermore, type 2 DM PBMCs have a mild defect in its ability to control hvKP replication relative to healthy PBMCs. In summary, our work demonstrates that type 2 DM does not overtly impact neutrophil intra- and extra-cellular killing of hvKP, but may influence cytokine/chemokine production and intracellular killing by PBMCs. PMID:28936426

Exposure of sea bream (Sparus aurata) to toxic concentrations of benzo[a]pyrene: possible human health effect.

PubMed

Zena, R; Speciale, A; Calabrò, C; Calò, M; Palombieri, D; Saija, A; Cimino, F; Trombetta, D; Lo Cascio, P

2015-12-01

Polycyclic aromatic hydrocarbons (PAHs) can accumulate in the food chain, due to their lipophilic properties. Fish can accumulate contaminants including PAHs and frequent consumption of such contaminated fish can pose risk to human health. The aim of this study was to clarify if acute exposure of sea bream (Sparus aurata, a fish species of great economic importance in the Atlantic and Mediterranean areas) to a PAH, benzo[a]pyrene (B[a]P), at a dose that can induce CYP1A and pathological changes in fish gills, liver and muscle, can induce accumulation in muscle. We investigated the cytotoxic effects (as changes in cell viability, DNA laddering and glutathione content) of in vitro exposure of human peripheral blood mononuclear cells (PBMCs) to organic extracts obtained from muscle of sea breams previously exposed via water to B[a]P (2mg/l, for 12, 24 and 72 h). At this level of exposure, B[a]P caused morphological changes, inflammatory response and CYP1A induction not only in sea bream gills and liver but also in muscle; furthermore, in fish muscle we observed a substantial B[a]P accumulation, which may be associated with the increased CYP1A activity in liver and especially in muscle. However, when PBMCs were exposed to organic extracts obtained from sea bream muscle contaminated with B[a]P, a toxic, although modest effect was revealed, consisting in a significant decrease in cell glutathione levels without alterations in cell viability and DNA laddering. This suggests that consumption of sea breams from B[a]P contaminated waters might represent a risk for human health. Copyright © 2015 Elsevier Inc. All rights reserved.

Anti-inflammatory and cytoprotective effects of a squalene synthase inhibitor, TAK-475 active metabolite-I, in immune cells simulating mevalonate kinase deficiency (MKD)-like condition.

PubMed

Suzuki, Nobutaka; Ito, Tatsuo; Matsui, Hisanori; Takizawa, Masayuki

2016-01-01

TAK-475 (lapaquistat acetate) and its active metabolite-I (TAK-475 M-I) inhibit squalene synthase, which catalyzes the conversion of farnesyl diphosphate (FPP) to squalene. FPP is a substrate for synthesis of other mevalonate-derived isoprenoids (MDIs) such as farnesol (FOH), geranlygeranyl diphosphate (GGPP), and geranylgeraniol. In patients with MKD, a rare autosomal recessive disorder, defective activity of mevalonate kinase leads to a shortage of MDIs. MDIs especially GGPP are required for prenylation of proteins, which is a posttranslation modification necessary for proper functioning of proteins like small guanosine triphosphatases. Malfunction of prenylation of proteins results in upregulation of the inflammatory cascade, leading to increased production of proinflammatory cytokines like interleukin-1β (IL-1β), eventually leading to episodic febrile attacks. In vitro, TAK-475 M-I incubation in a concentration dependent manner increased levels of FPP, GGPP, and FOH in human monocytic THP-1 cells. In subsequent experiments, THP-1 cells or human peripheral blood mononuclear cells (PBMCs) were incubated with simvastatin, which inhibits hydroxymethylglutaryl-coenzyme A reductase and thereby decreases levels of the precursors of MDIs, leading to the depletion of MDIs as expected in MKD patients. Increased levels of GGPP and FPP attenuated lipopolysaccharide (LPS)-induced IL-1β production in THP-1 cells and human PBMCs in statin-treated conditions. The MDIs also significantly reduced the damaged cell ratio in this active MKD-like condition. Moreover, TAK-475 M-I directly inhibited LPS-induced IL-1β production from statin-treated THP-1 cells. These results show anti-inflammatory and cytoprotective effects of MDIs via TAK-475 M-I treatment in statin-treated immune cells, suggesting that possible therapeutic effects of TAK-475 treatment in MKD patients.

The Staphyloccous aureus Eap protein activates expression of proinflammatory cytokines.

PubMed

Scriba, Thomas J; Sierro, Sophie; Brown, Eric L; Phillips, Rodney E; Sewell, Andrew K; Massey, Ruth C

2008-05-01

The extracellular adhesion protein (Eap) secreted by the major human pathogen Staphylococcus aureus is known to have several effects on human immunity. We have recently added to knowledge of these roles by demonstrating that Eap enhances interactions between major histocompatibility complex molecules and human leukocytes. Several studies have indicated that Eap can induce cytokine production by human peripheral blood mononuclear cells (PBMCs). To date, there has been no rigorous attempt to identify the breadth of cytokines produced by Eap stimulation or to identify the cell subsets that respond. Here, we demonstrate that Eap induces the secretion of the proinflammatory cytokines interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) by CD14(+) leukocytes (monocytes and macrophages) within direct ex vivo PBMC populations (note that granulocytes are also CD14(+) but are largely depleted from PBMC preparations). Anti-intercellular adhesion molecule 1 (CD54) antibodies inhibited this induction and implicated a role for this known Eap binding protein in cellular activation. IL-6 and TNF-alpha secretion by murine cells exposed to Eap was also observed. The activation of CD14(+) cells by Eap suggests that it could play a significant role in both septic shock and fever, two of the major pathological features of S. aureus infections.

An in vitro model of Mycobacterium leprae induced granuloma formation

PubMed Central

2013-01-01

Background Leprosy is a contagious and chronic systemic granulomatous disease caused by Mycobacterium leprae. In the pathogenesis of leprosy, granulomas play a key role, however, the mechanisms of the formation and maintenance of M. leprae granulomas are still not clearly understood. Methods To better understand the molecular physiology of M. leprae granulomas and the interaction between the bacilli and human host cells, we developed an in vitro model of human granulomas, which mimicked the in vivo granulomas of leprosy. Macrophages were differentiated from human monocytes, and infected with M. leprae, and then cultured with autologous human peripheral blood mononuclear cells (PBMCs). Results Robust granuloma-like aggregates were obtained only when the M. leprae infected macrophages were co-cultured with PBMCs. Histological examination showed M. leprae within the cytoplasmic center of the multinucleated giant cells, and these bacilli were metabolically active. Macrophages of both M1 and M2 types co-existed in the granuloma like aggregates. There was a strong relationship between the formation of granulomas and changes in the expression levels of cell surface antigens on macrophages, cytokine production and the macrophage polarization. The viability of M. leprae isolated from granulomas indicated that the formation of host cell aggregates benefited the host, but the bacilli also remained metabolically active. Conclusions A simple in vitro model of human M. leprae granulomas was established using human monocyte-derived macrophages and PBMCs. This system may be useful to unravel the mechanisms of disease progression, and subsequently develop methods to control leprosy. PMID:23782413

Fractalkine in human inflammatory cardiomyopathy.

PubMed

Escher, F; Vetter, R; Kühl, U; Westermann, D; Schultheiss, H-P; Tschöpe, C

2011-05-01

Cardiac inflammation is important for the prognosis of patients with inflammatory cardiomyopathy (CMi), but the mechanisms leading to it are not fully elucidated. To study the role of fractalkine (CX3CL1) in chemotactic and adhesive properties of peripheral blood mononuclear cells (PBMCs) in patients with CMi. Patients with enterovirus (EV)-positive CMi, patients with virus-negative CMi, patients with parvovirus B19 (B19) genomes with low intramyocardial inflammation and patients without cardiac inflammation and viral infection in the endomyocardial biopsy (EMB) were enrolled (n=10/group). The expression of CX3CL1 and monocyte chemoattractant protein (MCP-1) in EMBs was significantly increased in EV-positive and virus-negative patients with CMi in contrast to controls and B19-positive patients (EV+ vs controls: CX3CL1-area fraction (AF) % 0.078±0.012 vs 0.009±0.003 p<0.05; MCP-1-AF % 0.093±0.023 vs 0.011±0.009). The receptor (CX3CR1)-mediated chemotaxis was increased twofold in PBMCs in comparison with those of controls. The MCP-1 secretion was 3.1-fold higher in PBMCs from EV-positive patients compared with controls, and this elevation was further increased by CX3CL1 in EV-positive patients. No significant CX3CL1-mediated MCP-1 increase was seen in PBMCs from healthy controls. Moreover, spontaneously beating neonatal rat cardiomyocytes exposed to CX3CL1 exhibited an attenuated positive chronotropic response to β-adrenergic stimulation with isoproterenol. The cardiac and plasma CX3CL1/CX3CR1 system is upregulated in CMi and this affects the functional potential of PBMCs. Moreover, a direct cardiodepressive effect of CX3CL1 in cardiac tissue was demonstrated since neonatal cardiomyocytes exhibited an attenuated positive chronotropic response to β-adrenergic stimulation.

Methamphetamine induces trace amine-associated receptor 1 (TAAR1) expression in human T lymphocytes: role in immunomodulation.

PubMed

Sriram, Uma; Cenna, Jonathan M; Haldar, Bijayesh; Fernandes, Nicole C; Razmpour, Roshanak; Fan, Shongshan; Ramirez, Servio H; Potula, Raghava

2016-01-01

The novel transmembrane G protein-coupled receptor, trace amine-associated receptor 1 (TAAR1), represents a potential, direct target for drugs of abuse and monoaminergic compounds, including amphetamines. For the first time, our studies have illustrated that there is an induction of TAAR1 mRNA expression in resting T lymphocytes in response to methamphetamine. Methamphetamine treatment for 6 h significantly increased TAAR1 mRNA expression (P < 0.001) and protein expression (P < 0.01) at 24 h. With the use of TAAR1 gene silencing, we demonstrate that methamphetamine-induced cAMP, a classic response to methamphetamine stimulation, is regulated via TAAR1. We also show by TAAR1 knockdown that the down-regulation of IL-2 in T cells by methamphetamine, which we reported earlier, is indeed regulated by TAAR1. Our results also show the presence of TAAR1 in human lymph nodes from HIV-1-infected patients, with or without a history of methamphetamine abuse. TAAR1 expression on lymphocytes was largely in the paracortical lymphoid area of the lymph nodes with enhanced expression in lymph nodes of HIV-1-infected methamphetamine abusers rather than infected-only subjects. In vitro analysis of HIV-1 infection of human PBMCs revealed increased TAAR1 expression in the presence of methamphetamine. In summary, the ability of methamphetamine to activate trace TAAR1 in vitro and to regulate important T cell functions, such as cAMP activation and IL-2 production; the expression of TAAR1 in T lymphocytes in peripheral lymphoid organs, such as lymph nodes; and our in vitro HIV-1 infection model in PBMCs suggests that TAAR1 may play an important role in methamphetamine -mediated immune-modulatory responses. © Society for Leukocyte Biology.

iNKT cells ameliorate human autoimmunity: Lessons from alopecia areata.

PubMed

Ghraieb, Amal; Keren, Aviad; Ginzburg, Alex; Ullmann, Yehuda; Schrum, Adam G; Paus, Ralf; Gilhar, Amos

2018-04-18

Alopecia areata (AA) is understood to be a CD8+/NKG2D+ T cell-dependent autoimmune disease. Here, we demonstrate that human AA pathogenesis of is also affected by iNKT10 cells, an unconventional T cell subtype whose number is significantly increased in AA compared to healthy human skin. AA lesions can be rapidly induced in healthy human scalp skin xenotransplants on Beige-SCID mice by intradermal injections of autologous healthy-donor PBMCs pre-activated with IL-2. We show that in this in vivo model, the development of AA lesions is prevented by recognized the iNKT cell activator, α-galactosylceramide (α-GalCer), which stimulates iNKT cells to expand and produce IL-10. Moreover, in pre-established humanized mouse AA lesions, hair regrowth is promoted by α-GalCer treatment through a process requiring both effector-memory iNKT cells, which can interact directly with CD8+/NKG2D+ T cells, and IL-10. This provides the first in vivo evidence in a humanized model of autoimmune disease that iNKT10 cells are key disease-protective lymphocytes. Since these regulatory NKT cells can both prevent the development of AA lesions and promote hair re-growth in established AA lesions, targeting iNKT10 cells may have preventive and therapeutic potential also in other autoimmune disorders related to AA. Copyright © 2018 Elsevier Ltd. All rights reserved.

In Vitro Anticancer Effect of Gedunin on Human Teratocarcinomal (NTERA-2) Cancer Stem-Like Cells

PubMed Central

Tharmarajah, Luxmiga; Ediriweera, Meran Keshawa; Piyathilaka, Poorna; Senathilake, Kanishka Sithira; Rajagopalan, Umapriyatharshini; Galhena, Prasanna Bandula

2017-01-01

Gedunin is one of the major compounds found in the neem tree (Azadirachta indica). In the present study, antiproliferative potential of gedunin was evaluated in human embryonal carcinoma cells (NTERA-2, a cancer stem cell model) and peripheral blood mononuclear cells (PBMCs), using Sulforhodamine (SRB) and WST-1 assays, respectively. The effects of gedunin on expression of heat shock protein 90 (HSP90), its cochaperone Cdc37, and HSP client proteins (AKT, ErbB2, and HSF1) were evaluated by real-time PCR. Effects of gedunin on apoptosis were evaluated by (a) apoptosis associated morphological changes, (b) caspase 3/7 expression, (c) DNA fragmentation, (d) TUNEL assay, and (e) real-time PCR of apoptosis related genes (Bax, p53, and survivin). Gedunin showed a promising antiproliferative effect in NTERA-2 cells with IC50 values of 14.59, 8.49, and 6.55 μg/mL at 24, 48, and 72 h after incubations, respectively, while exerting a minimal effect on PBMCs. Expression of HSP90, its client proteins, and survivin was inhibited and Bax and p53 were upregulated by gedunin. Apoptosis related morphological changes, DNA fragmentation, and increased caspase 3/7 activities confirmed the proapoptotic effects of gedunin. Collectively, results indicate that gedunin may be a good drug lead for treatment of chemo and radiotherapy resistant cancer stem cells. PMID:28680880

Mycobacterium tuberculosis Transfer RNA Induces IL-12p70 via Synergistic Activation of Pattern Recognition Receptors within a Cell Network.

PubMed

Keegan, Caroline; Krutzik, Stephan; Schenk, Mirjam; Scumpia, Philip O; Lu, Jing; Pang, Yan Ling Joy; Russell, Brandon S; Lim, Kok Seong; Shell, Scarlet; Prestwich, Erin; Su, Dan; Elashoff, David; Hershberg, Robert M; Bloom, Barry R; Belisle, John T; Fortune, Sarah; Dedon, Peter C; Pellegrini, Matteo; Modlin, Robert L

2018-05-01

Upon recognition of a microbial pathogen, the innate and adaptive immune systems are linked to generate a cell-mediated immune response against the foreign invader. The culture filtrate of Mycobacterium tuberculosis contains ligands, such as M. tuberculosis tRNA, that activate the innate immune response and secreted Ags recognized by T cells to drive adaptive immune responses. In this study, bioinformatics analysis of gene-expression profiles derived from human PBMCs treated with distinct microbial ligands identified a mycobacterial tRNA-induced innate immune network resulting in the robust production of IL-12p70, a cytokine required to instruct an adaptive Th1 response for host defense against intracellular bacteria. As validated by functional studies, this pathway contained a feed-forward loop, whereby the early production of IL-18, type I IFNs, and IL-12p70 primed NK cells to respond to IL-18 and produce IFN-γ, enhancing further production of IL-12p70. Mechanistically, tRNA activates TLR3 and TLR8, and this synergistic induction of IL-12p70 was recapitulated by the addition of a specific TLR8 agonist with a TLR3 ligand to PBMCs. These data indicate that M. tuberculosis tRNA activates a gene network involving the integration of multiple innate signals, including types I and II IFNs, as well as distinct cell types to induce IL-12p70. Copyright © 2018 by The American Association of Immunologists, Inc.

Experimental exposure of blue mussels (Mytilus galloprovincialis) to high levels of benzo[a]pyrene and possible implications for human health.

PubMed

Speciale, A; Zena, R; Calabrò, C; Bertuccio, C; Aragona, M; Saija, A; Trombetta, D; Cimino, F; Lo Cascio, P

2018-04-15

Polycyclic aromatic hydrocarbons (PAHs) are lipophilic compounds able to accumulate in the food chain. Mussels showed to bioaccumulate contaminants, such as PAHs, so that recurrent consumption of such contaminated food represents a risk for human health. This study was aimed to elucidate if acute exposure of Mediterranean blue mussel (Mytilus galloprovincialis), a bivalve of great economic importance in several countries, to a PAH, benzo[a]pyrene (B[a]P), at doses able to induce cytochrome P450 1A (CYP1A) and pathological changes in mussel gills, can produce accumulation in soft tissue. We explored the cytotoxic effects (cell viability, DNA laddering, and glutathione levels) of in vitro exposure of human peripheral blood mononuclear cells (PBMCs) to organic extracts obtained from blue mussels previously exposed for 12 and 72h via water to B[a]P (0.5-1mg/L). In our experimental conditions, B[a]P induced CYP1A induction and morphological changes in mussel gills and a significant B[a]P accumulation in soft tissue. Conversely, exposing PBMCs to organic extracts obtained from contaminated mussels, resulted in a significant reduction of cell viability and cell glutathione content, and in an increase in DNA laddering. This confirms that consumption of mussels from B[a]P polluted waters might affect human health. Our data lead us to suggest that CYP1A activity in mussel gills may be useful (more than the amount of detected PAHs in the mussel edible tissue) as a marker in assessment of risk for health of consumers exposed to PAHs through ingestion of shellfish. Copyright © 2017 Elsevier Inc. All rights reserved.

An in vitro evaluation of anti-aging effect of guluronic acid (G2013) based on enzymatic oxidative stress gene expression using healthy individuals PBMCs.

PubMed

Taeb, Mahsa; Mortazavi-Jahromi, Seyed Shahabeddin; Jafarzadeh, Abdollah; Mirzaei, Mohammad Reza; Mirshafiey, Abbas

2017-06-01

Aging is usually associated with increased levels of oxidants, and may result in damages caused by oxidative stress. There is a direct relationship between aging and increased incidence of inflammatory diseases. The present research intended to study the anti-aging and anti-inflammatory effects of the drug G2013 (guluronic acid) at low and high doses on the genes expression of a number of enzymes involved in oxidative stress (including SOD2, GPX1, CAT, GST, iNOS, and MPO) in peripheral blood mononuclear cells (PBMCs) of healthy individuals under in vitro conditions. Venous blood samples were taken from 20 healthy individuals, the PBMCs were isolated and their RNAs extracted and their cDNAs were synthesized, and the genes expression levels were measured using the qRT-PCR technique. Our results indicated that this drug could, at both low and high doses, significantly reduce the expression of the genes for SOD2, GPX1, CAT, and GST compared to the LPS group (p<0.0001). Moreover, it was noticed that the drug is able to significantly reduce gene expression levels at the high dose and at both doses (low and high), for iNOS and MPO compared to the LPS group (p<0.0001), respectively. The present research showed that G2013, as a novel NSAID drug with immunomodulatory properties, could modulate the expression levels of the genes for SOD2, GPX1, CAT, GST, iNOS, and MPO, to the level of healthy gene expression, and possibly it might reduce the pathological process of aging and age-related inflammatory diseases. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Attenuation of LPS-induced inflammation by ICT, a derivate of icariin, via inhibition of the CD14/TLR4 signaling pathway in human monocytes.

PubMed

Wu, Jinfeng; Zhou, Junmin; Chen, Xianghong; Fortenbery, Nicole; Eksioglu, Erika A; Wei, Sheng; Dong, Jingcheng

2012-01-01

To evaluate the anti-inflammatory potential of ICT in LPS stimulated human innate immune cells. 3, 5, 7-Trihydroxy-4'-methoxy-8-(3-hydroxy-3- methylbutyl)-flavone (ICT) is a novel derivative of icariin, the major active ingredient of Herba Epimedii, an herb used in traditional Chinese medicine. We previously demonstrated its anti-inflammatory potential in a murine macrophage cell line as well as in mouse models. We measured TNF-α production by ELISA, TLR4/CD14 expression by flow cytometry, and NF-κB and MAPK activation by western blot all in LPS-stimulated PBMC, human monocytes, or THP-1 cells after treatment with ICT. ICT inhibited LPS-induced TNF-α production in THP-1 cells, PBMCs and human monocytes in a dose-dependent manner. ICT treatment resulted in down-regulation of the expression of CD14/TLR4 and attenuated NF-κB and MAPK activation induced by LPS. We illustrate the anti-inflammatory property of ICT in human immune cells, especially in monocytes. These effects were mediated, at least partially, via inhibition of the CD14/TLR4 signaling pathway. Copyright © 2011 Elsevier B.V. All rights reserved.

Less immune activation following social stress in rural vs. urban participants raised with regular or no animal contact, respectively

PubMed Central

Böbel, Till S.; Hackl, Sascha B.; Jarczok, Marc N.; Rohleder, Nicolas; Rook, Graham A.; Gündel, Harald; Waller, Christiane

2018-01-01

Urbanization is on the rise, and environments offering a narrow range of microbial exposures are linked to an increased prevalence of both physical and mental disorders. Human and animal studies suggest that an overreactive immune system not only accompanies stress-associated disorders but might even be causally involved in their pathogenesis. Here, we show in young [mean age, years (SD): rural, 25.1 (0.78); urban, 24.5 (0.88)] healthy human volunteers that urban upbringing in the absence of pets (n = 20), relative to rural upbringing in the presence of farm animals (n = 20), was associated with a more pronounced increase in the number of peripheral blood mononuclear cells (PBMCs) and plasma interleukin 6 (IL-6) concentrations following acute psychosocial stress induced by the Trier social stress test (TSST). Moreover, ex vivo-cultured PBMCs from urban participants raised in the absence of animals secreted more IL-6 in response to the T cell-specific mitogen Con A. In turn, antiinflammatory IL-10 secretion was suppressed following TSST in urban participants raised in the absence of animals, suggesting immunoregulatory deficits, relative to rural participants raised in the presence of animals. Questionnaires, plasma cortisol, and salivary α-amylase, however, indicated the experimental protocol was more stressful and anxiogenic for rural participants raised in the presence of animals. Together, our findings support the hypothesis that urban vs. rural upbringing in the absence or presence of animals, respectively, increases vulnerability to stress-associated physical and mental disorders by compromising adequate resolution of systemic immune activation following social stress and, in turn, aggravating stress-associated systemic immune activation. PMID:29712842

Tumor Necrosis Factor-Like Weak Inducer of Apoptosis Activates Type I Interferon Signals in Lupus Nephritis.

PubMed

Xue, Leixi; Liu, Lei; Huang, Jun; Wen, Jian; Yang, Ru; Bo, Lin; Tang, Mei; Zhang, Yi; Liu, Zhichun

2017-01-01

Type I interferon (IFN) plays a central role in pathogenesis of systemic lupus erythematosus (SLE); tumor necrosis factor-like weak inducer of apoptosis (TWEAK) has been associated with a pathogenic role in lupus nephritis (LN). Thus we investigated whether TWEAK could induce the activation of type I IFN pathway in LN. We examined this in patient-derived peripheral blood mononuclear cells (PBMCs) as well as MRL/lpr mice, a murine LN model. Relative to the control cohorts, MRL/lpr mice showed severe histological changes, high index levels of renal damage, and elevated expression of type I IFN-inducible genes. After shRNA suppression of TWEAK, we observed that renal damage was significantly attenuated and expression of type I IFN-inducible genes was reduced in MRL/lpr mice. In parallel, siRNA of TWEAK also significantly reduced the expression of type I IFN-inducible genes in PBMCs relative to control transfections. In PBMCs, TWEAK stimulation also led to expression of type I IFN-inducible genes. Our results illustrate a novel regulatory role of TWEAK, in which its activity positively regulates type I IFN pathway in LN based on preclinical models. Our findings suggest TWEAK could act as a critical target in preventing renal damage in patients with LN.

Tumor necrosis factor-α inhibits effects of aryl hydrocarbon receptor ligands on cell death in human lymphocytes.

PubMed

Ghatrehsamani, Mahdi; Soleimani, Masoud; Esfahani, Behjat A Moayedi; Shirzad, Hedayatollah; Hakemi, Mazdak G; Mossahebimohammadi, Majid; Eskandari, Nahid; Adib, Minoo

2015-01-01

Activation of aryl hydrocarbon receptor (AhR) leads to diverse outcome in various kinds of cells. AhR activation may induce apoptosis or prevent of apoptosis and cell death. Recent studies suggest that apoptosis effects of AhR can be modulated by inflammatory cytokine like tumor necrosis factor alpha (TNF-α). In this study, we try to investigate the possible interaction of TNF-α with the 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD), a ligand of AhR, on peripheral lymphocytes. Human peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood by discontinuous density gradient centrifugation on ficoll. Isolated PBMCs were divided into four groups: Control group, TNF-α administered group, TCDD administered group, co-administered group with TCDD and TNF-α. Cells were maintained for a week in lymphocyte culture condition. Then, TNF-α was added to group 2 and 4. Finally, apoptosis and necrosis were analyzed in all samples using flowcytometry. In group 4, the mean percent of necrosis and apoptosis in TCDD treatment groups was significantly larger than other groups; (P < 0.05). Furthermore, there was no significant difference between the mean percent of cell death in TNF-α administered group and TCDD administered group (P > 0.05). However, the mean percent of cell death in co-administered group with TCDD and TNF-α was significantly lower than other groups; (P < 0.05). TNF-α could significantly inhibit effects of TCDD on lymphocytes apoptosis. Combination effects of TNF-α and TCDD on lymphocyte increase cell survival.

CD4-veCD8-ve CD30+ve T cells are detectable in human lung transplant patients and their proportion of the lymphocyte population after in vitro stimulation with donor spleen cells correlates with preservation of lung physiology.

PubMed

Polster, K; Walker, A; Fildes, J; Entwistle, G; Yonan, N; Hutchinson, I V; Leonard, C T

2005-06-01

Survival following lung transplantation is less than 50% at 5 years, mainly due to immune-mediated chronic rejection. Recently a novel subset of T cells, CD4-veCD8-ve CD30+ve, so-called double negative (DN) CD30+ve T cells, has been described and shown to be responsible for tolerance in an animal model of skin transplantation. We investigated 18 lung transplant recipients for the presence of DN CD30+ve T cells in resting peripheral blood and also following in vitro stimulation of recipient peripheral blood mononuclear cells (PBMCs) with donor spleen cells. Small percentages (0.2% to 6%) of DN T cells are detectable in resting PBMCs of human transplant patients (n = 18), but these did not correlate with allograft function, acute rejection episodes, HLA mismatch, or CMV status. On repeated stimulation of recipient PBMCs (two exposures) in vitro by donor spleen cells (2:1 ratio stimulators to responders) the percentage of DN CD30+ve T cells within the lymphocyte pool correlated with preservation of allograft lung function (both for FEV(1), P = .009, and FEF(25-75), P = .036) and was inversely correlated with grade of chronic rejection. On repeated exposure of recipient PBMCs to donor spleen cells with a 1:1 ratio the percentage of DN CD30+ve T cells correlated with the number of acute rejection episodes of grade 2 or greater. The total number of HLA mismatches correlated with the percentage DN CD30+ve T cells present after primary stimulation of recipient PBMCs with donor spleen cells (1:1 ratio). The number of mismatches at the B locus inversely correlated with the percentage of DN CD30+ve T cells after primary stimulation of recipient PBMCs with donor spleen cells (1:1 ratio; P = .031, n = 18). Percentages of DN CD30+ve T cells present following repeated stimulation of recipient PBMCs by donor spleen cells correlated with preservation of graft function following lung transplantation.

Oxytetracycline induces DNA damage and epigenetic changes: a possible risk for human and animal health?

PubMed

Gallo, Adriana; Landi, Rosaria; Rubino, Valentina; Di Cerbo, Alessandro; Giovazzino, Angela; Palatucci, Anna Teresa; Centenaro, Sara; Guidetti, Gianandrea; Canello, Sergio; Cortese, Laura; Ruggiero, Giuseppina; Alessandrini, Andrea; Terrazzano, Giuseppe

2017-01-01

Oxytetracycline (OTC), which is largely employed in zootechnical and veterinary practices to ensure wellness of farmed animals, is partially absorbed within the gastrointestinal tract depositing in several tissues. Therefore, the potential OTC toxicity is relevant when considering the putative risk derived by the entry and accumulation of such drug in human and pet food chain supply. Despite scientific literature highlights several OTC-dependent toxic effects on human and animal health, the molecular mechanisms of such toxicity are still poorly understood. Here, we evaluated DNA damages and epigenetic alterations by quantitative reverse transcription polymerase chain reaction, quantitative polymerase chain reaction, chromatin immuno-precipitation and Western blot analysis. We observed that human peripheral blood mononuclear cells (PBMCs) expressed DNA damage features (activation of ATM and p53, phosphorylation of H2AX and modifications of histone H3 methylation of lysine K4 in the chromatin) after the in vitro exposure to OTC. These changes are linked to a robust inflammatory response indicated by an increased expression of Interferon (IFN)- γ and type 1 superoxide dismutase (SOD1). Our data reveal an unexpected biological in vitro activity of OTC able to modify DNA and chromatin in cultured human PBMC. In this regard, OTC presence in foods of animal origin could represent a potential risk for both the human and animal health.

Activity of SHIP, Which Prevents Expression of Interleukin 1β, Is Reduced in Patients With Crohn's Disease.

PubMed

Ngoh, Eyler N; Weisser, Shelley B; Lo, Young; Kozicky, Lisa K; Jen, Roger; Brugger, Hayley K; Menzies, Susan C; McLarren, Keith W; Nackiewicz, Dominika; van Rooijen, Nico; Jacobson, Kevan; Ehses, Jan A; Turvey, Stuart E; Sly, Laura M

2016-02-01

Crohn's disease (CD) is associated with a dysregulated immune response to commensal micro-organisms in the intestine. Mice deficient in inositol polyphosphate 5'-phosphatase D (INPP5D, also known as SHIP) develop intestinal inflammation resembling that of patients with CD. SHIP is a negative regulator of PI3Kp110α activity. We investigated mechanisms of intestinal inflammation in Inpp5d(-/-) mice (SHIP-null mice), and SHIP levels and activity in intestinal tissues of subjects with CD. We collected intestines from SHIP-null mice, as well as Inpp5d(+/+) mice (controls), and measured levels of cytokines of the interleukin 1 (IL1) family (IL1α, IL1β, IL1ra, and IL6) by enzyme-linked immunosorbent assay. Macrophages were isolated from lamina propria cells of mice, IL1β production was measured, and mechanisms of increased IL1β production were investigated. Macrophages were incubated with pan-phosphatidylinositol 3-kinase inhibitors or PI3Kp110α-specific inhibitors. Some mice were given an antagonist of the IL1 receptor; macrophages were depleted from ilea of mice using clodronate-containing liposomes. We obtained ileal biopsies from sites of inflammation and peripheral blood mononuclear cells (PBMCs) from treatment-naïve subjects with CD or without CD (controls), and measured SHIP levels and activity. PBMCs were incubated with lipopolysaccharide and adenosine triphosphate, and levels of IL1β production were measured. Inflamed intestinal tissues and intestinal macrophages from SHIP-null mice produced higher levels of IL1B and IL18 than intestinal tissues from control mice. We found PI3Kp110α to be required for macrophage transcription of Il1b. Macrophage depletion or injection of an IL1 receptor antagonist reduced ileal inflammation in SHIP-null mice. Inflamed ileal tissues and PBMCs from patients with CD had lower levels of SHIP protein than controls (P < .0001 and P < .0002, respectively). There was an inverse correlation between levels of SHIP activity in

Antibody-dependent cellular cytotoxicity (ADCC) activity of a novel anti-PD-L1 antibody avelumab (MSB0010718C) on human tumor cells

PubMed Central

Fantini, Massimo; Heery, Christopher R.; Gulley, James L.; Tsang, Kwong Yok; Schlom, Jeffrey

2015-01-01

Several anti-PD1/PD-L1 monoclonal antibodies (MAb) are currently providing evidence of clinical benefit in subsets of cancer patients. The mode of action of these MAbs is to inhibit PD1 on immune cells interacting with PD-L1 on tumor cells. These MAbs are either designed or engineered to eliminate antibody-dependent cell-mediated cytotoxicity (ADCC), which, however, has been implicated as an important mechanism in several highly effective MAb-mediated cancer therapies. A fully human anti-PD-L1 MAb would potentially be able to block PD-L1/PD1 interactions and also mediate the ADCC lysis of tumor cells. MSB0010718C (designated avelumab) is a fully human IgG1 anti-PD-L1 MAb. The studies reported here demonstrate (a) the ability of avelumab to lyse a range of human tumor cells in the presence of PBMC or NK effectors; (b) IFNγ can enhance tumor cell PD-L1 expression and in some cases enhance ADCC tumor cell lysis; (c) purified NK cells are potent effectors for avelumab; (d) similar levels of avelumab-mediated ADCC lysis of tumor cells are seen using purified NK as effectors from either healthy donors or cancer patients; (e) very low levels of avelumab-mediated lysis are seen using whole PBMCs as targets; this finding complements results seen in analyses of PBMC subsets of patients receiving avelumab; and (f) the addition of IL12 to NK cells greatly enhances avelumab-mediated ADCC. These studies thus provide an additional mode of action for an anti-PD-L1 MAb and support the rationale for further studies to enhance avelumab-mediated ADCC activity. PMID:26014098

Amniotic membrane extract differentially regulates human peripheral blood T cell subsets, monocyte subpopulations and myeloid dendritic cells.

PubMed

Laranjeira, Paula; Duque, Marta; Vojtek, Martin; Inácio, Maria J; Silva, Isabel; Mamede, Ana C; Laranjo, Mafalda; Pedreiro, Susana; Carvalho, Maria J; Moura, Paulo; Abrantes, Ana M; Maia, Cláudio J; Domingues, Pedro; Domingues, Rosário; Martinho, António; Botelho, Maria F; Trindade, Hélder; Paiva, Artur

2018-03-26

The discovery of the immunoregulatory potential of human amniotic membrane (hAM) propelled several studies focusing on its application for the treatment of immunological disorders. However, there is little information regarding the effects of hAM on distinct activation and differentiation stages of immune cells. Here, we aim to investigate the effect of human amniotic membrane extract (hAME) on the pattern of cytokine production by T cells, monocytes and myeloid dendritic cells (mDCs). For this purpose, peripheral blood mononuclear cells (PBMCs) from eight healthy individuals were stimulated in vitro in the presence or absence of hAME. Mitogen-induced proliferation of PBMCs and cytokine production among the distinct T cell functional compartments, monocyte subpopulations and mDCs were evaluated. hAME displayed an anti-proliferative effect and decreased the frequency of T cells producing tumor necrosis factor (TNF)α, interferon (IFN)γ and interleukin (IL)-2, for all T cell functional compartments. The frequency of IL-17 and IL-9-producing T cells was also reduced. The inhibition of mRNA expression of granzyme B, perforin and NKG2D by CD8 + T cells and γδ T cells and the augment of FoxP3 and IL-10 in CD4 + T cells and IL-10 in regulatory T cells were also observed. Furthermore, hAME inhibited IFNγ-induced protein (IP)-10 expression by classical and non-classical monocytes, without hampering the production of TNFα and IL-6 by monocytes and mDCs. These results suggest that hAME exerts an anti-inflammatory effect on T cells, still at a different extent for distinct T cell functional compartments.

Tumour Necrosis Factor-alpha (TNF-α) and its soluble receptor type 1 (sTNFR I) in human active and healed leishmaniases.

PubMed

Nateghi Rostami, M; Seyyedan Jasbi, E; Khamesipour, A; Mohammadi, A M

2016-04-01

The role of tumour necrosis factor-alpha (TNF-α) is not fully understood in human leishmaniasis. We analysed the alterations in the levels of TNF-α, soluble TNF receptor type 1 (sTNFR I), IL-17 and IL-22 productions in active and healed leishmaniases. Blood samples were collected from volunteers with active cutaneous leishmaniasis (ACL), the same subjects after lesion healing (healed CL = HCL), volunteers with active visceral leishmaniasis (AVL), healed VL (HVL) and healthy controls. Levels of cytokines were titrated on Leishmania Ag-stimulated PBMC culture. The mean level of TNF-α production from stimulated cells was significantly higher in ACL than controls (P < 0·001) and significantly reduced after treatment in HCL volunteers (P < 0·05). The mean level of sTNFR I production was significantly higher in ACL than controls (P < 0·001) and significantly reduced after treatment in HCL volunteers (P < 0·05). The mean level of IL-22 production in AVL was significantly higher than controls (P < 0·05) and was significantly lower in HVL compared with AVL (P < 0·001) and controls (P < 0·05). The levels of TNF-α (P = 0·0025) and sTNFR I (P < 0·01) productions from PBMCs showed significant decreasing trend after treatment in each CL volunteer. Reduction in TNF-α is associated with clinical response to treatment and healing of CL lesions due to L. major. © 2016 John Wiley & Sons Ltd.

An Enzyme-Linked Immunosorbent Spot Assay Measuring Borrelia burgdorferi B31-Specific Interferon Gamma-Secreting T Cells Cannot Discriminate Active Lyme Neuroborreliosis from Past Lyme Borreliosis: a Prospective Study in the Netherlands.

PubMed

van Gorkom, T; Sankatsing, S U C; Voet, W; Ismail, D M; Muilwijk, R H; Salomons, M; Vlaminckx, B J M; Bossink, A W J; Notermans, D W; Bouwman, J J M; Kremer, K; Thijsen, S F T

2018-04-01

Two-tier serology testing is most frequently used for the diagnosis of Lyme borreliosis (LB); however, a positive result is no proof of active disease. To establish a diagnosis of active LB, better diagnostics are needed. Tests investigating the cellular immune system are available, but studies evaluating the utility of these tests on well-defined patient populations are lacking. Therefore, we investigated the utility of an enzyme-linked immunosorbent spot (ELISpot) assay to diagnose active Lyme neuroborreliosis. Peripheral blood mononuclear cells (PBMCs) of various study groups were stimulated by using Borrelia burgdorferi strain B31 and various recombinant antigens, and subsequently, the number of Borrelia -specific interferon gamma (IFN-γ)-secreting T cells was measured. We included 33 active and 37 treated Lyme neuroborreliosis patients, 28 healthy individuals treated for an early manifestation of LB in the past, and 145 untreated healthy individuals. The median numbers of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs did not differ between active Lyme neuroborreliosis patients (6.0; interquartile range [IQR], 0.5 to 14.0), treated Lyme neuroborreliosis patients (4.5; IQR, 2.0 to 18.6), and treated healthy individuals (7.4; IQR, 2.3 to 14.9) ( P = 1.000); however, the median number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs among untreated healthy individuals was lower (2.0; IQR, 0.5 to 3.9) ( P ≤ 0.016). We conclude that the Borrelia ELISpot assay, measuring the number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 10 5 PBMCs, correlates with exposure to the Borrelia bacterium but cannot be used for the diagnosis of active Lyme neuroborreliosis. Copyright © 2018 van Gorkom et al.

An Enzyme-Linked Immunosorbent Spot Assay Measuring Borrelia burgdorferi B31-Specific Interferon Gamma-Secreting T Cells Cannot Discriminate Active Lyme Neuroborreliosis from Past Lyme Borreliosis: a Prospective Study in the Netherlands

PubMed Central

van Gorkom, T.; Sankatsing, S. U. C.; Voet, W.; Ismail, D. M.; Muilwijk, R. H.; Salomons, M.; Vlaminckx, B. J. M.; Bossink, A. W. J.; Notermans, D. W.; Bouwman, J. J. M.; Kremer, K.

2018-01-01

ABSTRACT Two-tier serology testing is most frequently used for the diagnosis of Lyme borreliosis (LB); however, a positive result is no proof of active disease. To establish a diagnosis of active LB, better diagnostics are needed. Tests investigating the cellular immune system are available, but studies evaluating the utility of these tests on well-defined patient populations are lacking. Therefore, we investigated the utility of an enzyme-linked immunosorbent spot (ELISpot) assay to diagnose active Lyme neuroborreliosis. Peripheral blood mononuclear cells (PBMCs) of various study groups were stimulated by using Borrelia burgdorferi strain B31 and various recombinant antigens, and subsequently, the number of Borrelia-specific interferon gamma (IFN-γ)-secreting T cells was measured. We included 33 active and 37 treated Lyme neuroborreliosis patients, 28 healthy individuals treated for an early manifestation of LB in the past, and 145 untreated healthy individuals. The median numbers of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 105 PBMCs did not differ between active Lyme neuroborreliosis patients (6.0; interquartile range [IQR], 0.5 to 14.0), treated Lyme neuroborreliosis patients (4.5; IQR, 2.0 to 18.6), and treated healthy individuals (7.4; IQR, 2.3 to 14.9) (P = 1.000); however, the median number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 105 PBMCs among untreated healthy individuals was lower (2.0; IQR, 0.5 to 3.9) (P ≤ 0.016). We conclude that the Borrelia ELISpot assay, measuring the number of B. burgdorferi B31-specific IFN-γ-secreting T cells/2.5 × 105 PBMCs, correlates with exposure to the Borrelia bacterium but cannot be used for the diagnosis of active Lyme neuroborreliosis. PMID:29367297

Increase in activated Treg in TIL in lung cancer and in vitro depletion of Treg by ADCC using an antihuman CCR4 mAb (KM2760).

PubMed

Kurose, Koji; Ohue, Yoshihiro; Sato, Eiichi; Yamauchi, Akira; Eikawa, Shingo; Isobe, Midori; Nishio, Yumi; Uenaka, Akiko; Oka, Mikio; Nakayama, Eiichi

2015-01-01

Tregs infiltrate tumors and inhibit immune responses against them. We investigated subpopulations of Foxp3 CD4 T cells previously defined by Miyara et al. (Immunity 30, 899-911, 2009) in peripheral blood mononuclear cells (PBMCs) and tumor infiltrating lymphocytes (TILs) in lung cancer. We also showed that Tregs in healthy donors that express CCR4 could be efficiently eliminated in vitro by cotreatment with antihuman (h) CCR4 mAb (KM2760) and NK cells. In lung cancer, the number of activated/effector Tregs and non-Tregs, but not resting/naive Tregs, was increased in TILs compared with the number of those cells in PBMCs. The non-Treg population contained Th2 and Th17. CCR4 expression on activated/effector Tregs and non-Tregs in TILs was down-regulated compared with that on those cells in PBMCs. Chemokinetic migration of CD25 CD4 T cells containing the Treg population sorted from the PBMCs of healthy donors to CCL22/MDC was abrogated by pretreatment with anti-hCCR4 mAb (KM2760). The inhibitory activity of CD25 CD127 CD4 Tregs on the proliferative response of CD4 and CD8 T cells stimulated with anti-CD3/CD28 coated beads was abrogated by adding an anti-hCCR4 mAb (KM2760) and CD56 NK cells to the culture. The findings suggested the CCR4 on activated/effector Tregs and non-Tregs was functionally involved in the chemokinetic migration and accumulation of those cells to the tumor site. In vitro findings of efficient elimination of Tregs may give the basis for implementation of a clinical trial to investigate Treg depletion by administration of an anti-hCCR4 mAb to solid cancer patients.

Arginase activity in peripheral blood of patients with intestinal schistosomiasis, Wonji, Central Ethiopia.

PubMed

Getaneh, A; Tamrat, A; Tadesse, K

2015-07-01

Morbidity and mortality caused by schistosomiasis usually results from immunopathology. But the underlying mechanisms are not yet clearly understood. Th2-type immune response is thought to be dominant during chronic schistosomiasis, and upregulation of arginase-I is one component of this milieu. A cohort study was conducted to assess arginase activity in peripheral blood of humans with intestinal schistosomiasis in Wonji-Shoa Sugar Estate, Central Ethiopia. Laboratory-confirmed 30 Schistosoma mansoni-infected patients and 18 apparently healthy controls were recruited. Faecal egg count was carried out by Kato-Katz technique. Plasma and peripheral blood mononuclear cells (PBMCs) were isolated from whole blood. Activity of arginase in plasma and PBMC lysates was measured, and results were compared with that of controls. Twenty-one of 30 patients had light infection, whereas moderate and heavy intensity infections were observed in eight and only one patient(s), respectively. A significant increase in both PBMC (patients: 59.96 + 82.99, controls: 25.44 + 24.6 mU/mg protein, P < 0.0001) and plasma (patients: 1.61 + 2.19, controls: 0.31 + 0.73 mU/mL plasma, P < 0.0001) arginase activity was observed during human S. mansoni infection. Arginase activity increases in peripheral blood of patients with intestinal schistosomiasis. © 2015 John Wiley & Sons Ltd.

Secretion of interferon gamma from human immune cells is altered by exposure to tributyltin and dibutyltin.

PubMed

Lawrence, Shanieek; Reid, Jacqueline; Whalen, Margaret

2015-05-01

Tributyltin (TBT) and dibutyltin (DBT) are widespread environmental contaminants found in food, beverages, and human blood samples. Both of these butyltins (BTs) interfere with the ability of human natural killer (NK) cells to lyse target cells and alter secretion of the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) from human immune cells in vitro. The capacity of BTs to interfere with secretion of other pro-inflammatory cytokines has not been examined. Interferon gamma (IFNγ) is a modulator of adaptive and innate immune responses, playing an important role in overall immune competence. This study shows that both TBT and DBT alter secretion of IFNγ from human immune cells. Peripheral blood cell preparations that were increasingly reconstituted were used to determine if exposures to either TBT or DBT affected IFNγ secretion and how the makeup of the cell preparation influenced that effect. IFNγ secretion was examined after 24 h, 48 h, and 6 day exposures to TBT (200 - 2.5 nM) and DBT (5 - 0.05 µM) in highly enriched human NK cells, a monocyte-depleted preparation of PBMCs, and monocyte-containing PBMCs. Both BTs altered IFNγ secretion from immune cells at most of the conditions tested (either increasing or decreasing secretion). However, there was significant variability among donors as to the concentrations and time points that showed changes as well as the baseline secretion of IFNγ. The majority of donors showed an increase in IFNγ secretion in response to at least one concentration of TBT or DBT at a minimum of one length of exposure. © 2013 Wiley Periodicals, Inc.

Selective reactivation of human herpesvirus 6 in patients with autoimmune connective tissue diseases.

PubMed

Broccolo, Francesco; Drago, Francesco; Cassina, Giulia; Fava, Andrea; Fusetti, Lisa; Matteoli, Barbara; Ceccherini-Nelli, Luca; Sabbadini, Maria Grazia; Lusso, Paolo; Parodi, Aurora; Malnati, Mauro S

2013-11-01

Viral infections have been associated with autoimmune connective tissue diseases. To evaluate whether active infection by Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpesvirus (HHV)-6, -7, -8, as well as parvovirus B19 (B19V) occur in patients with autoimmune connective tissue diseases, viral DNA loads were assessed in paired samples of serum and peripheral blood mononuclear cells (PBMCs) of 115 patients affected by different disorders, including systemic sclerosis, systemic, and discoid lupus erythematosus, rheumatoid arthritis, and dermatomyositis. Two additional groups, patients affected by inflammatory diseases (n=51) and healthy subjects (n=58) were studied as controls. The titers of anti-HHV-6 and anti-EBV antibodies were also evaluated. Cell-free HHV-6 serum viremia was detected in a significantly higher proportion of connective tissue diseases patients compared to controls (P<0.0002); a significant association between HHV-6 reactivation and the active disease state was found only for lupus erythematosus (P=0.021). By contrast, the rate of cell-free EBV viremia was similar in patients and controls groups. Cell-free CMV, HHV-8, and B19V viremia was not detected in any subject. Anti-HHV-6 and anti-EBV early antigen IgG titers were both significantly higher in autoimmune diseases patients as compared to healthy controls, although they were not associated with the presence of viremia. EBV, HHV-6, -7 prevalence and viral load in PBMCs of patients with connective tissue diseases and controls were similar. These data suggest that HHV-6 may act as a pathogenic factor predisposing patients to the development of autoimmune connective tissue diseases or, conversely, that these disorders may predispose patients to HHV-6 reactivation. © 2013 Wiley Periodicals, Inc.

Peripheral blood lymphocytes express recombination-activating genes 1 and 2 during Epstein-Barr virus-induced infectious mononucleosis.

PubMed

Wagner, Hans-Joachim; Scott, Rona S; Buchwald, Dedra; Sixbey, John W

2004-09-01

Implicit in the persistence of Epstein-Barr virus (EBV) in B lymphocytes is the successful circumvention of ongoing cell selection for competence of B cell receptors (BCRs). Because the EBV infection of B cells in vitro induces enzymatic machinery that is responsible for secondary immunoglobulin gene rearrangement, we examined the expression of the recombination-activating genes (RAGs) in peripheral blood mononuclear cells (PBMCs) from 26 patients with infectious mononucleosis (IM). RAG1 and/or RAG2 RNA was detected in PBMCs from 42% of patients with IM but not from healthy control subjects. EBV may usurp the cellular mechanism that diversifies the BCR, to guarantee a level of survival signaling sufficient for its own persistence.

Antibody-Dependent Cellular Cytotoxicity Activity of a Novel Anti-PD-L1 Antibody Avelumab (MSB0010718C) on Human Tumor Cells.

PubMed

Boyerinas, Benjamin; Jochems, Caroline; Fantini, Massimo; Heery, Christopher R; Gulley, James L; Tsang, Kwong Yok; Schlom, Jeffrey

2015-10-01

Several anti-PD-1/PD-L1 monoclonal antibodies (mAb) are currently providing evidence of clinical benefit in subsets of cancer patients. The mode of action of these mAbs is to inhibit PD-1 on immune cells interacting with PD-L1 on tumor cells. These mAbs are either designed or engineered to eliminate antibody-dependent cell-mediated cytotoxicity (ADCC), which, however, has been implicated as an important mechanism in several highly effective mAb-mediated cancer therapies. A fully human anti-PD-L1 mAb would potentially be able to block PD-1/PD-L1 interactions and also mediate the ADCC lysis of tumor cells. MSB0010718C (designated avelumab) is a fully human IgG1 anti-PD-L1 mAb. The studies reported here demonstrate (i) the ability of avelumab to lyse a range of human tumor cells in the presence of PBMC or NK effectors; (ii) IFNγ can enhance tumor cell PD-L1 expression and, in some cases, enhance ADCC tumor cell lysis; (iii) purified NK cells are potent effectors for avelumab; (iv) similar levels of avelumab-mediated ADCC lysis of tumor cells are seen using purified NK as effectors from either healthy donors or cancer patients; (v) very low levels of avelumab-mediated lysis are seen using whole PBMCs as targets; this finding complements results seen in analyses of PBMC subsets of patients receiving avelumab; and (vi) the addition of IL12 to NK cells greatly enhances avelumab-mediated ADCC. These studies thus provide an additional mode of action for an anti-PD-L1 mAb and support the rationale for further studies to enhance avelumab-mediated ADCC activity. ©2015 American Association for Cancer Research.

Plumbagin exerts an immunosuppressive effect on human T-cell acute lymphoblastic leukemia MOLT-4 cells

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

Bae, Kyoung Jun; Lee, Yura; Kim, Soon Ae

2016-04-22

Of the hematological disorders typified by poor prognoses and survival rates, T-cell acute lymphoblastic leukemia (T-ALL) is one of the most commonly diagnosed. Despite the development of new therapeutic agents, the treatment options for this cancer remain limited. In this manuscript, we investigated the anti-proliferative effects of plumbagin, mediated by the activation of mitogen-activated protein kinase (MAPK) pathways, and inhibition of NF-κB signaling; the human T-ALL MOLT-4 cell line was used as our experimental system. Plumbagin is a natural, plant derived compound, which exerts an anti-proliferative activity against many types of human cancer. Our experiments confirm that plumbagin induces a caspase-dependentmore » apoptosis of MOLT-4 cells, with no significant cytotoxicity seen for normal peripheral blood mononuclear cells (PBMCs). Plumbagin also inhibited LPS-induced phosphorylation of p65, and the transcription of NF-κB target genes. Our results now show that plumbagin is a potent inhibitor of the NF-κB signaling pathway, and suppressor of T-ALL cell proliferation. - Highlights: • Plumbagin induces caspase-dependent apoptosis in T-ALL MOLT-4 cells. • Plumbagin activates phosphorylation of stress-activated protein kinase (SAPK) JNK and p38. • Plumbagin inhibits LPS-mediated NF-κB signaling cascade. • Plumbagin inhibits LPS-mediated transcriptional activity of pro-inflammatory cytokines.« less

Quantification of HTLV-1 reverse transcriptase activity in ATL patients treated with zidovudine and interferon-α.

PubMed

Macchi, Beatrice; Balestrieri, Emanuela; Frezza, Caterina; Grelli, Sandro; Valletta, Elena; Marçais, Ambroise; Marino-Merlo, Francesca; Turpin, Jocelyn; Bangham, Charles R; Hermine, Olivier; Mastino, Antonio; Bazarbachi, Ali

2017-05-09

The therapeutic efficacy of the AZT and IFN combination in ATL presumably reflects the inhibition of RT-related functions.HTLV-1-RT activity from short-term cultured PBMCs may represent a predictive correlate of clinical response to AZT/IFN in ATL patients.

Intrinsic Patterns of Human Activity

NASA Astrophysics Data System (ADS)

Hu, Kun; Ivanov, Plamen Ch.; Chen, Zhi; Hilton, Michael; Stanley, H. Eugene; Shea, Steven

2003-03-01

Activity is one of the defining features of life. Control of human activity is complex, being influenced by many factors both extrinsic and intrinsic to the body. The most obvious extrinsic factors that affect activity are the daily schedule of planned events, such as work and recreation, as well as reactions to unforeseen or random events. These extrinsic factors may account for the apparently random fluctuations in human motion observed over short time scales. The most obvious intrinsic factors are the body clocks including the circadian pacemaker that influences our sleep/wake cycle and ultradian oscillators with shorter time scales [2, 3]. These intrinsic rhythms may account for the underlying regularity in average activity level over longer periods of up to 24 h. Here we ask if the known extrinsic and intrinsic factors fully account for all complex features observed in recordings of human activity. To this end, we measure activity over two weeks from forearm motion in subjects undergoing their regular daily routine. Utilizing concepts from statistical physics, we demonstrate that during wakefulness human activity possesses previously unrecognized complex dynamic patterns. These patterns of activity are characterized by robust fractal and nonlinear dynamics including a universal probability distribution and long-range power-law correlations that are stable over a wide range of time scales (from minutes to hours). Surprisingly, we find that these dynamic patterns are unaffected by changes in the average activity level that occur within individual subjects throughout the day and on different days of the week, and between subjects. Moreover, we find that these patterns persist when the same subjects undergo time-isolation laboratory experiments designed to account for the phase of the circadian pacemaker, and control the known extrinsic factors by restricting behaviors and manipulating scheduled events including the sleep/wake cycle. We attribute these newly

Brain Activity and Human Unilateral Chewing

PubMed Central

Quintero, A.; Ichesco, E.; Myers, C.; Schutt, R.; Gerstner, G.E.

2012-01-01

Brain mechanisms underlying mastication have been studied in non-human mammals but less so in humans. We used functional magnetic resonance imaging (fMRI) to evaluate brain activity in humans during gum chewing. Chewing was associated with activations in the cerebellum, motor cortex and caudate, cingulate, and brainstem. We also divided the 25-second chew-blocks into 5 segments of equal 5-second durations and evaluated activations within and between each of the 5 segments. This analysis revealed activation clusters unique to the initial segment, which may indicate brain regions involved with initiating chewing. Several clusters were uniquely activated during the last segment as well, which may represent brain regions involved with anticipatory or motor events associated with the end of the chew-block. In conclusion, this study provided evidence for specific brain areas associated with chewing in humans and demonstrated that brain activation patterns may dynamically change over the course of chewing sequences. PMID:23103631

Tryptamine and dimethyltryptamine inhibit indoleamine 2,3 dioxygenase and increase the tumor-reactive effect of peripheral blood mononuclear cells.

PubMed

Tourino, Melissa Cavalheiro; de Oliveira, Edson Mendes; Bellé, Luziane Potrich; Knebel, Franciele Hinterholz; Albuquerque, Renata Chaves; Dörr, Felipe Augusto; Okada, Sabrina Sayori; Migliorini, Silene; Soares, Irene Silva; Campa, Ana

2013-07-01

Indoleamine 2,3-dioxygenase (IDO) is an interferon-γ (IFN-γ)-induced tryptophan-degrading enzyme, producing kynurenine (KYN) that participates in the mechanism of tumor immune tolerance. Thus, IDO inhibition has been considered a strategy for anticancer therapy. The aim of this study was to identify whether the metabolites originated from the competitive routes of tryptophan metabolism, such as the serotonergic or N, N-dimethyltryptamine (DMT) pathways, have inhibitory effects on recombinant human IDO (rhIDO) activity. Serotonin and melatonin had no effect; on the other hand, tryptamine (TRY) and DMT modulated the activity of rhIDO as classical non-competitive inhibitors, with Ki values of 156 and 506 μM, respectively. This inhibitory effect was also observed on constitutively expressed or IFN-γ-induced IDO in the A172 human glioma cell line. TRY and DMT increased the cytotoxic activity of peripheral blood mononuclear cells (PBMCs) in co-culture assays. We conclude that the IDO inhibition by TRY and DMT contributed to a more effective tumor-reactive response by the PBMCs. Copyright © 2013 John Wiley & Sons, Ltd.

The aminobisphosphonate pamidronate controls influenza pathogenesis by expanding a γδ T cell population in humanized mice

PubMed Central

Zheng, Jian; Liu, Yinping; Sia, Sin Fun; Liu, Ming; Qin, Gang; Xiang, Zheng; Lam, Kwok-Tai

2011-01-01

There are few antiviral drugs for treating influenza, and the emergence of antiviral resistance has further limited the available therapeutic options. Furthermore, antivirals are not invariably effective in severe influenza, such as that caused by H5N1 viruses. Thus, there is an urgent need to develop alternative therapeutic strategies. Here, we show that human Vγ9Vδ2 T cells expanded by the aminobisphosphonate pamidronate (PAM) kill influenza virus–infected cells and inhibit viral replication in vitro. In Rag2−/−γc−/− immunodeficient mice reconstituted with human peripheral mononuclear cells (huPBMCs), PAM reduces disease severity and mortality caused by human seasonal H1N1 and avian H5N1 influenza virus, and controls the lung inflammation and viral replication. PAM has no such effects in influenza virus–infected Rag2−/−γc−/− mice reconstituted with Vγ9Vδ2 T cell–depleted huPBMCs. Our study provides proof-of-concept of a novel therapeutic strategy for treating influenza by targeting the host rather than the virus, thereby reducing the opportunity for the emergence of drug-resistant viruses. As PAM has been commonly used to treat osteoporosis and Paget’s disease, this new application of an old drug potentially offers a safe and readily available option for treating influenza. PMID:21708931

Anti-LINGO-1 has no detectable immunomodulatory effects in preclinical and phase 1 studies

PubMed Central

Ranger, Ann; Ray, Soma; Szak, Suzanne; Dearth, Andrea; Allaire, Norm; Murray, Ronald; Gardner, Rebecca; Cadavid, Diego

2017-01-01

Objective: To evaluate whether the anti-LINGO-1 antibody has immunomodulatory effects. Methods: Human peripheral blood mononuclear cells (hPBMCs), rat splenocytes, and rat CD4+ T cells were assessed to determine whether LINGO-1 was expressed and was inducible. Anti-LINGO-1 Li81 (0.1–30 μg/mL) effect on proliferation/cytokine production was assessed in purified rat CD4+ T cells and hPBMCs stimulated with antibodies to CD3 +/– CD28. In humans, the effect of 2 opicinumab (anti-LINGO-1/BIIB033; 30, 60, and 100 mg/kg) or placebo IV administrations was evaluated in RNA from blood and CSF samples taken before and after administration in phase 1 clinical trials; paired samples were assessed for differentially expressed genes by microarray. RNA from human CSF cell pellets was analyzed by quantitative real-time PCR for changes in transcripts representative of cell types, activation markers, and soluble proteins of the adaptive/innate immune systems. ELISA quantitated the levels of CXCL13 protein in human CSF supernatants. Results: LINGO-1 is not expressed in hPBMCs, rat splenocytes, or rat CD4+ T cells; LINGO-1 blockade with Li81 did not affect T-cell proliferation or cytokine production from purified rat CD4+ T cells or hPBMCs. LINGO-1 blockade with opicinumab resulted in neither significant changes in immune system gene expression in blood and CSF, nor changes in CXCL13 CSF protein levels (clinical studies). Conclusions: These data support the hypothesis that LINGO-1 blockade does not affect immune function. Classification of evidence: This study provides Class II evidence that in patients with MS, opicinumab does not have immunomodulatory effects detected by changes in immune gene transcript expression. PMID:29259995

Utility of Survival Motor Neuron ELISA for Spinal Muscular Atrophy Clinical and Preclinical Analyses

PubMed Central

Kobayashi, Dione T.; Olson, Rory J.; Sly, Laurel; Swanson, Chad J.; Chung, Brett; Naryshkin, Nikolai; Narasimhan, Jana; Bhattacharyya, Anuradha; Mullenix, Michael; Chen, Karen S.

2011-01-01

Objectives Genetic defects leading to the reduction of the survival motor neuron protein (SMN) are a causal factor for Spinal Muscular Atrophy (SMA). While there are a number of therapies under evaluation as potential treatments for SMA, there is a critical lack of a biomarker method for assessing efficacy of therapeutic interventions, particularly those targeting upregulation of SMN protein levels. Towards this end we have engaged in developing an immunoassay capable of accurately measuring SMN protein levels in blood, specifically in peripheral blood mononuclear cells (PBMCs), as a tool for validating SMN protein as a biomarker in SMA. Methods A sandwich enzyme-linked immunosorbent assay (ELISA) was developed and validated for measuring SMN protein in human PBMCs and other cell lysates. Protocols for detection and extraction of SMN from transgenic SMA mouse tissues were also developed. Results The assay sensitivity for human SMN is 50 pg/mL. Initial analysis reveals that PBMCs yield enough SMN to analyze from blood volumes of less than 1 mL, and SMA Type I patients' PBMCs show ∼90% reduction of SMN protein compared to normal adults. The ELISA can reliably quantify SMN protein in human and mouse PBMCs and muscle, as well as brain, and spinal cord from a mouse model of severe SMA. Conclusions This SMN ELISA assay enables the reliable, quantitative and rapid measurement of SMN in healthy human and SMA patient PBMCs, muscle and fibroblasts. SMN was also detected in several tissues in a mouse model of SMA, as well as in wildtype mouse tissues. This SMN ELISA has general translational applicability to both preclinical and clinical research efforts. PMID:21904622

Variables influencing anti-human immunodeficiency virus type 1 neutralizing human monoclonal antibody (NhMAb) production among infected Thais.

PubMed

Akapirat, Siriwat; Avihingsanon, Anchalee; Ananworanich, Jintanat; Schuetz, Alexandra; Ramasoota, Pongrama; Luplertlop, Natthanej; Ono, Ken-Ichiro; Ikuta, Kazuyoshi; Utachee, Piraporn; Kameoka, Masanori; Leaungwutiwong, Pornsawan

2013-09-01

We conducted this study to determine the clinical variables associated with the production of human immunodeficiency virus type 1 (HIV-1) circulating recombinant form (CRF) 01_AE neutralizing human monoclonal antibodies (NhMAbs) using a hybridoma technique. This cross sectional study was performed in 20 asymptomatic HIV-1-infected Thais. Peripheral blood mononuclear cells (PBMCs) were obtained from each study participant and fused with SPYMEG cells. Culture supernatant collected from growing hybridomas was tested for neutralizing activity against HIV-1 CRF01_AE Env-recombinant viruses. Fifty hybridomas expressing anti-HIV-1 NhMAbs with strong neutralizing activity against at least 1 CRF01_AE Env-recombinant virus were found. A positive association between the numbers of hybridomas produced and the CD4 counts of study participants (p = 0.019) was observed. NhMAb-producing hybridomas with strong neutralizing activity were mostly found in participans diagnosed with HIV-1 infection within the previous 1 year. The HIV-1 viral load was not significantly correlated with the numbers of either established hybridomas or clones expressing anti-HIV-1 NhMAbs with strong neutralizing activity. To our knowledge, this is the first study of NhMAb-producing hybridomas obtained from HIV-1 CRF01_AE-infected populations identified by antibody binding to HIV-1 V3 loop peptide enzyme-linked immunosorbent assay (ELISA) or TRUGENE HIV-1 Genotyping Assay (HIV-1 pol sequence). It provides important criterion to slect study participants with high CD4 counts who produce large numbers of hybridoma clones. The results are valuable for further studies related to nurtalizing antibodies production and HIV-1 vaccine development.

Comparison of RAW264.7, human whole blood and PBMC assays to screen for immunomodulators.

PubMed

Elisia, Ingrid; Pae, Han Bee; Lam, Vivian; Cederberg, Rachel; Hofs, Elyse; Krystal, Gerald

2018-01-01

The RAW264.7 mouse macrophage cell line is used extensively to carry out in vitro screens for immunomodulators. Compounds that are effective at reducing the expression of pro-inflammatory cytokines or nitric oxide (NO) from lipopolysaccharide (LPS)-stimulated RAW264.7 cells are often considered candidate anti-inflammatory agents for humans. There is, however, very little data on the reliability of this screen to identify bona fide human immunomodulators. We compared the efficacy of 37 purported immunomodulators to modulate LPS or E. coli-induced inflammatory responses in RAW264.7 cell, whole human blood and human peripheral blood mononuclear cell (PBMC) assays. Interestingly, there was no significant correlation (R=0.315) between the responses obtained with RAW264.7 cells and the whole blood assay (WBA), suggesting that compounds demonstrating efficacy in RAW264.7 cells may be ineffective in humans, and, more importantly, compounds that are effective in humans may be missed with a RAW264.7 screen. Interestingly, there was also no significant correlation between the WBA and human PBMCs when the latter were cultured with 10% FCS, but a moderate correlation was seen when the PBMCs were cultured with 25% autologous plasma. The presence of plasma thus contributes to the overall inflammatory response observed in the WBA. We then asked if RAW264.7 cells, given that they are mouse macrophage-like cells, respond in a manner similar to primary murine derived macrophages. Intriguingly, there was no significant correlation (R=0.012) with the 37 putative immunomodulators, pointing to distinct inflammatory response mechanisms in the two model systems. We conclude that the use of a WBA to confirm potential immunomodulators obtained from high throughput screening with RAW264.7 cells is advisable and that future screens be carried out using a WBA. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of Eucommia ulmoides Oliv., Gynostemma pentaphyllum (Thunb.) Makino, and Curcuma longa L. on Th1- and Th2-cytokine responses and human leukocyte antigen-DR expression in peripheral blood mononuclear cells of septic patients.

PubMed

Wu, Huang-Pin; Lin, Yin-Ku

2018-05-10

Many traditional Chinese medicines (TCM), such as Eucommia ulmoides Oliv., Gynostemma pentaphyllum (Thunb.) Makino, and Curcuma longa L., have been reported to have various immune-modulatory effects. To determine the effects of extracts from these three TCM on type 1 T help (Th1)- and Th2-cytokine responses and human leukocyte antigen (HLA)-DR expression in peripheral blood mononuclear cells (PBMCs) obtained from septic patients. Lipopolysaccharide (LPS)-stimulated PBMCs of healthy controls and septic patients were cultured for 48 hs with or without 0.05/0.1 mg/ml of TCM extract. HLA-DR expression in monocytes was detected using flow cytofluorimetry. The interferon [IFN]-γ, tumor necrosis factor [TNF]-α, interleukin (IL)- 2, IL-5, IL-10, and IL-13 levels in supernatants were measured with a human enzyme-linked immunosorbent assay. Treatment with either 0.05 or 0.1 mg/ml of C. longa L. extract significantly restored the percentage of HLA-DR-positive monocytes, which was decreased by LPS in control and patient groups. Treatment with 0.05 or 0.1 mg/ml E. ulmoides Oliv. and C.longa L. extract decreased IL-10 production from LPS-stimulated PBMCs of controls and patients. In patients with sepsis, C. longa L. extract decreased IL-10 production to a greater degree than did E. ulmoides Oliv extract. Although IFN-γ, TNF-α, or IL-13 productions from LPS-stimulated PBMCs were influenced by E. ulmoides Oliv., G. pentaphyllum (Thunb.) Makino, or C. longa L. in control or sepsis groups in this study, only the influence of IL-10 was consistent in both control and sepsis groups. By enhancing monocyte HLA-DR expression and decreasing IL-10 production, C. longa L. might help restore inflammatory responses in septic patients to eradicate pathogens. Copyright © 2018 Elsevier B.V. All rights reserved.

Rapid resetting of human peripheral clocks by phototherapy during simulated night shift work.

PubMed

Cuesta, Marc; Boudreau, Philippe; Cermakian, Nicolas; Boivin, Diane B

2017-11-24

A majority of night shift workers have their circadian rhythms misaligned to their atypical schedule. While bright light exposure at night is known to reset the human central circadian clock, the behavior of peripheral clocks under conditions of shift work is more elusive. The aim of the present study was to quantify the resetting effects of bright light exposure on both central (plasma cortisol and melatonin) and peripheral clocks markers (clock gene expression in peripheral blood mononuclear cells, PBMCs) in subjects living at night. Eighteen healthy subjects were enrolled to either a control (dim light) or a bright light group. Blood was sampled at baseline and on the 4 th day of simulated night shift. In response to a night-oriented schedule, the phase of PER1 and BMAL1 rhythms in PBMCs was delayed by ~2.5-3 h (P < 0.05), while no shift was observed for the other clock genes and the central markers. Three cycles of 8-h bright light induced significant phase delays (P < 0.05) of ~7-9 h for central and peripheral markers, except BMAL1 (advanced by +5h29; P < 0.05). Here, we demonstrate in humans a lack of peripheral clock adaptation under a night-oriented schedule and a rapid resetting effect of nocturnal bright light exposure on peripheral clocks.

Virus-specific cytotoxic T cells in chronic active Epstein-Barr virus infection.

PubMed

Shibayama, Haruna; Imadome, Ken-Ichi; Onozawa, Erika; Tsuzura, Akiho; Miura, Osamu; Koyama, Takatoshi; Arai, Ayako

2017-01-01

Chronic active Epstein-Barr virus infection (CAEBV) is a disease characterized by clonally proliferating and activated EBV-infected T or NK cells accompanied by chronic inflammation and T- or NK-cell neoplasms. However, the mechanism for developing CAEBV has not been clarified to date. Because the decreased number or inactivation of EBV-specific cytotoxic T lymphocytes (CTLs) resulted in the development of EBV-positive B-cell neoplasms, we investigated the number of CTLs in CAEBV patients using the tetrameric complexes of HLA-restricted EBV-specific peptides. Among the seven patients examined, EBV-specific CTLs were detected in the peripheral blood mononuclear cells (PBMCs) of four cases but were not detected in three cases. The ratio of EBV-specific CTLs in PBMCs tended to be higher in the patients with active disease than in those with inactive disease. In two patients in whom EBV-specific CTLs had not been detected, CTLs appeared after the eradication of EBV-infected T cells by allogeneic bone marrow transplantation. These results suggested that the failure of CTLs had a role in developing CAEBV, although the induction number and function of EBV-specific CTLs might vary in each patient.

Diagnostic values for the viral load in peripheral blood mononuclear cells of patients with chronic active Epstein-Barr virus disease.

PubMed

Ito, Yoshinori; Suzuki, Michio; Kawada, Jun-ichi; Kimura, Hiroshi

2016-04-01

Chronic active Epstein-Barr virus disease (CAEBV) is a distinct EBV-associated lymphoproliferative disease with a poor prognosis. Although the viral load in blood samples has been widely used for diagnosing CAEBV, well-defined viral load thresholds to guide clinicians are currently lacking. The aim of the present study was to determine standardized diagnostic values for EBV load in blood samples of CAEBV patients using the World Health Organization international standard for reporting. Levels of EBV DNA in 103 peripheral blood mononuclear cells (PBMCs) and 95 plasma/serum samples from 107 cases with CAEBV were quantified and expressed in international units. Receiver operating characteristic curves were analyzed to assess the most appropriate cut-off values for levels of EBV DNA to distinguish CAEBV from EBV-associated infectious mononucleosis (IM) and controls with past EBV infection. Levels of EBV DNA in PBMCs were significantly higher in the CAEBV group (median, 10(4.2) IU/μgDNA) compared to the IM (median, 10(2.1) IU/μgDNA) and control groups. An inconsistent qualitative result was seen in 13 of 86 CAEBV patients; in these, EBV-DNA was positive in PBMCs, but negative in plasma. Diagnostic cut-off values for viral load in PBMCs from CAEBV patients, as compared to those of healthy controls and IM patients, were 10(2.0) IU/μgDNA and 10(3.2) IU/μgDNA, respectively. For diagnostic purposes, the viral load of PBMCs was better than of plasma/serum. A diagnostic cut-off EBV load for CAEBV may be useful for the management of CAEBV patients. Copyright © 2015 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Repeated exposure to Lutzomyia intermedia sand fly saliva induces local expression of interferon-inducible genes both at the site of injection in mice and in human blood.

PubMed

Weinkopff, Tiffany; de Oliveira, Camila I; de Carvalho, Augusto M; Hauyon-La Torre, Yazmin; Muniz, Aline C; Miranda, Jose Carlos; Barral, Aldina; Tacchini-Cottier, Fabienne

2014-01-01

During a blood meal, Lutzomyia intermedia sand flies transmit Leishmania braziliensis, a parasite causing tegumentary leishmaniasis. In experimental leishmaniasis, pre-exposure to saliva of most blood-feeding sand flies results in parasite establishment in absence of any skin damages in mice challenged with dermotropic Leishmania species together with saliva. In contrast, pre-immunization with Lu. intermedia salivary gland sonicate (SGS) results in enhanced skin inflammatory exacerbation upon co-inoculation of Lu. intermedia SGS and L. braziliensis. These data highlight potential unique features of both L. braziliensis and Lu. intermedia. In this study, we investigated the genes modulated by Lu. intermedia SGS immunization to understand their potential impact on the subsequent cutaneous immune response following inoculation of both SGS and L. braziliensis. The cellular recruitment and global gene expression profile was analyzed in mice repeatedly inoculated or not with Lu. intermedia. Microarray gene analysis revealed the upregulation of a distinct set of IFN-inducible genes, an immune signature not seen to the same extent in control animals. Of note this INF-inducible gene set was not induced in SGS pre-immunized mice subsequently co-inoculated with SGS and L. braziliensis. These data suggest the parasite prevented the upregulation of this Lu. intermedia saliva-related immune signature. The presence of these IFN-inducible genes was further analyzed in peripheral blood mononuclear cells (PBMCs) sampled from uninfected human individuals living in a L. braziliensis-endemic region of Brazil thus regularly exposed to Lu. intermedia bites. PBMCs were cultured in presence or absence of Lu. intermedia SGS. Using qRT-PCR we established that the IFN-inducible genes induced in the skin of SGS pre-immunized mice, were also upregulated by SGS in PBMCs from human individuals regularly exposed to Lu. intermedia bites, but not in PBMCs of control subjects. These data demonstrate

Repeated Exposure to Lutzomyia intermedia Sand Fly Saliva Induces Local Expression of Interferon-Inducible Genes Both at the Site of Injection in Mice and in Human Blood

PubMed Central

Weinkopff, Tiffany; de Oliveira, Camila I.; de Carvalho, Augusto M.; Hauyon-La Torre, Yazmin; Muniz, Aline C.; Miranda, Jose Carlos; Barral, Aldina; Tacchini-Cottier, Fabienne

2014-01-01

During a blood meal, Lutzomyia intermedia sand flies transmit Leishmania braziliensis, a parasite causing tegumentary leishmaniasis. In experimental leishmaniasis, pre-exposure to saliva of most blood-feeding sand flies results in parasite establishment in absence of any skin damages in mice challenged with dermotropic Leishmania species together with saliva. In contrast, pre-immunization with Lu. intermedia salivary gland sonicate (SGS) results in enhanced skin inflammatory exacerbation upon co-inoculation of Lu. intermedia SGS and L. braziliensis. These data highlight potential unique features of both L. braziliensis and Lu. intermedia. In this study, we investigated the genes modulated by Lu. intermedia SGS immunization to understand their potential impact on the subsequent cutaneous immune response following inoculation of both SGS and L. braziliensis. The cellular recruitment and global gene expression profile was analyzed in mice repeatedly inoculated or not with Lu. intermedia. Microarray gene analysis revealed the upregulation of a distinct set of IFN-inducible genes, an immune signature not seen to the same extent in control animals. Of note this INF-inducible gene set was not induced in SGS pre-immunized mice subsequently co-inoculated with SGS and L. braziliensis. These data suggest the parasite prevented the upregulation of this Lu. intermedia saliva-related immune signature. The presence of these IFN-inducible genes was further analyzed in peripheral blood mononuclear cells (PBMCs) sampled from uninfected human individuals living in a L. braziliensis-endemic region of Brazil thus regularly exposed to Lu. intermedia bites. PBMCs were cultured in presence or absence of Lu. intermedia SGS. Using qRT-PCR we established that the IFN-inducible genes induced in the skin of SGS pre-immunized mice, were also upregulated by SGS in PBMCs from human individuals regularly exposed to Lu. intermedia bites, but not in PBMCs of control subjects. These data demonstrate

Segmentation and Recognition of Continuous Human Activity

DTIC Science & Technology

2001-01-01

This paper presents a methodology for automatic segmentation and recognition of continuous human activity . We segment a continuous human activity into...commencement or termination. We use single action sequences for the training data set. The test sequences, on the other hand, are continuous sequences of human ... activity that consist of three or more actions in succession. The system has been tested on continuous activity sequences containing actions such as

The efficacy and pharmacokinetics of brincidofovir for the treatment of lethal rabbitpox virus infection: a model of smallpox disease.

PubMed

Trost, Lawrence C; Rose, Michelle L; Khouri, Jody; Keilholz, Laurie; Long, James; Godin, Stephen J; Foster, Scott A

2015-05-01

Brincidofovir (BCV) has broad-spectrum in vitro activity against dsDNA viruses, including smallpox, and is being developed as a treatment for smallpox as well as infections caused by other dsDNA viruses. BCV has previously been shown to be active in multiple animal models of smallpox. Here we present the results of a randomized, blinded, placebo-controlled study of the efficacy and pharmacokinetics of a novel, "humanized" regimen of BCV for treatment of New Zealand White rabbits infected with a highly lethal inoculum of rabbitpox virus, a well characterized model of smallpox. Compared with placebo, a dose-dependent increase in survival was observed in all BCV-treatment groups. Concentrations of cidofovir diphosphate (CDV-PP), the active antiviral, in rabbit peripheral blood mononuclear cells (PBMCs) were determined for comparison to those produced in humans at the dose proposed for treatment of smallpox. CDV-PP exposure in PBMCs from rabbits given BCV scaled to human exposures at the dose proposed for treatment of smallpox, which is also currently under evaluation for other indications. The results of this study demonstrate the activity of BCV in the rabbitpox model of smallpox and the feasibility of scaling doses efficacious in the model to a proposed human dose and regimen for treatment of smallpox. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Effect of cryopreservation on IL-4, IFNgamma and IL-6 production of porcine peripheral blood lymphocytes.

PubMed

Li, Xiujin; Zhong, Zhenyu; Liang, Shuang; Wang, Xingxing; Zhong, Fei

2009-12-01

Cryopreservation of animal or human peripheral blood mononuclear cells (PBMC) is a commonly used technique. Effects of cryopreservation on functional capacity, especially the cytokine production of human PBMCs, have been extensively defined. However, certain animals, such as livestock, are a shortage of these information. Here we investigated the effects of cryopreservation on cytokine (IL-4, IFNgamma and IL-6) production of porcine PBMC. The porcine PBMCs were cryopreserved at -196 degrees C for a variety time periods for 2, 5, 25 and 50 days. Viability and cytokine production of the porcine PBMCs were measured before and after cryopreservation. The results showed that about 90% cell recovery rate was obtained at each storage time, indicating that about 10% loss of PBMCs in this short-term cryopreservation was due to the freezing process rather than the duration of cryopreservation. The fresh or frozen resting porcine PBMCs produced little cytokines in the absence of stimulation. However, three cytokines were apparently increased after PMA stimulation in both fresh and frozen porcine PBMCs. The sensitivity of frozen cells to PMA simulation for IFNgamma and IL-6 production was different from that of the fresh ones. IFNgamma production from the frozen PBMCs was significantly higher than that from the fresh ones (P<0.01). In contrast, IL-6 level from the frozen sample was significantly lower than that from the fresh one (P<0.05). Those results indicate that cryopreservation can increase the sensitivity of porcine PBMCs stimulated by PMA for IFNgamma production but not for IL-6 production. There was no significant difference of IL-4 production between fresh and frozen cells either stimulated (P>0.05) or un-stimulated (P>0.05).

BJ-TSA-9, a novel human tumor-specific gene, has potential as a biomarker of lung cancer.

PubMed

Li, Yunyan; Dong, Xueyuan; Yin, Yanhui; Su, Yanrong; Xu, Qingwen; Zhang, Yuxia; Pang, Xuewen; Zhang, Yu; Chen, Weifeng

2005-12-01

Using bioinformatics, we have identified a novel tumor-specific gene BJ-TSA-9, which has been validated by Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR). BJ-TSA-9 mRNA was expressed in 52.5% (21 of 40) of human lung cancer tissues and was especially higher in lung adenocarcinoma (68.8%). To explore the potential application of BJ-TSA-9 for the detection of circulating cancer cells in lung cancer patients, nested RT-PCR was performed. The overall positive detection rate was 34.3% (24 of 70) in peripheral blood mononuclear cells (PBMCs) of patients with various types of lung cancers and was 53.6% (15 of 28) in PBMCs of lung adenocarcinoma patients. In combination with the detection of two known marker genes SCC and LUNX, the detection rate was increased to 81.4%. A follow-up study was performed in 37 patients after surgical removal of tumor mass. Among nine patients with persistent detection of two to three tumor marker transcripts in PBMCs, six patients had recurrence/metastasis. In contrast, 28 patients with transient detection of one tumor marker or without detection of any tumor marker were all in remission. Thus, BJ-TSA-9 may serve as a marker for lung cancer diagnosis and as a marker, in combination with two other tumor markers, for the prediction of the recurrence and prognosis of lung cancer patients.

A methionine-free diet associated with nitrosourea treatment down-regulates methylguanine-DNA methyl transferase activity in patients with metastatic cancer.

PubMed

Thivat, Emilie; Durando, Xavier; Demidem, Aïcha; Farges, Marie-Chantal; Rapp, Maryse; Cellarier, Eric; Guenin, Samuel; D'Incan, Michel; Vasson, Marie-Paule; Chollet, Philippe

2007-01-01

Methionine (MET) depletion used in association with chemotherapy improves the therapeutic index in animal models. This potentiating effect may be due to tumor cell sensitization to chloroethylnitrosoureas through their MET dependency and the down-regulation of O6- methylguanine-DNA methyltransferase (MGMT). Our purpose was to evaluate the impact of the association of a dietary MET restriction with nitrosourea treatment on MGMT activity in peripheral blood mononuclear cells (PBMCs). Six patients with metastatic cancer (melanoma and glioma) received 4 cycles of a MET-free diet with cystemustine (60 mg/m2). MGMT activity in PBMCs decreased by an average of 13% from 553+/-90 fnol/mg before the diet to 413+/-59 fmol/mg after the diet + chemotherapy period (p=0.029). The decrease of MGMT activity was not affected by the duration of the MET-free diet period but seems to be correlated to the plasma MET depletion induced by the MET-free diet.

Phosphorylation of Extracellular Signal-Regulated Kinase (ERK)-1/2 Is Associated with the Downregulation of Peroxisome Proliferator–Activated Receptor (PPAR)-γ during Polymicrobial Sepsis

PubMed Central

Kaplan, Jennifer M; Hake, Paul W; Denenberg, Alvin; Nowell, Marchele; Piraino, Giovanna; Zingarelli, Basilia

2010-01-01

Peroxisome proliferator–activated receptor (PPAR)-γ is a ligand-activated transcription factor and regulates inflammation. Posttranslational modifications regulate the function of PPARγ, potentially affecting inflammation. PPARγ contains a mitogen-activated protein kinase (MAPK) site, and phosphorylation by extracellular signal-regulated kinase (ERK)-1/2 leads to inhibition of PPARγ. This study investigated the kinetics of PPARγ expression and activation in parenchymal and immune cells in sepsis using the MAPK/ERK kinase (MEK)-1 inhibitor, an upstream kinase of ERK1/2. Adult male Sprague Dawley rats were subjected to polymicrobial sepsis by cecal ligation and puncture. Rats received intraperitoneal injection of vehicle or the MEK1 inhibitor PD98059 (5 mg/kg) 30 min before cecal ligation and puncture. Rats were euthanized at 0, 1, 3, 6 and 18 h after cecal ligation and puncture. Control animals used were animals at time 0 h. Lung, plasma and peripheral blood mononuclear cells (PBMCs) were collected for biochemical assays. In vehicle-treated rats, polymicrobial sepsis resulted in significant lung injury. In the lung and PBMCs, nuclear levels of PPARγ were decreased and associated with an increase in phosphorylated PPARγ and phosphorylated ERK1/2 levels. Treatment with the MEK1 inhibitor increased the antiinflammatory plasma adipokine adiponectin, restored PPARγ expression in PBMCs and lung, and decreased lung injury. The inflammatory effects of sepsis cause changes in PPARγ expression and activation, in part, because of phosphorylation of PPARγ by ERK1/2. This phosphorylation can be reversed by ERK1/2 inhibition, thereby improving lung injury. PMID:20809049

Human NK Cell Subset Functions Are Differentially Affected by Adipokines

PubMed Central

Huebner, Lena; Engeli, Stefan; Wrann, Christiane D.; Goudeva, Lilia; Laue, Tobias; Kielstein, Heike

2013-01-01

Background Obesity is a risk factor for various types of infectious diseases and cancer. The increase in adipose tissue causes alterations in both adipogenesis and the production of adipocyte-secreted proteins (adipokines). Since natural killer (NK) cells are the host’s primary defense against virus-infected and tumor cells, we investigated how adipocyte-conditioned medium (ACM) affects functions of two distinct human NK cell subsets. Methods Isolated human peripheral blood mononuclear cells (PBMCs) were cultured with various concentrations of human and murine ACM harvested on two different days during adipogenesis and analyzed by fluorescent-activated cell sorting (FACS). Results FACS analyses showed that the expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), granzyme A (GzmA) and interferon (IFN)-γ in NK cells was regulated in a subset-specific manner. ACM treatment altered IFN-γ expression in CD56dim NK cells. The production of GzmA in CD56bright NK cells was differentially affected by the distinct adipokine compositions harvested at different states of adipogenesis. Comparison of the treatment with either human or murine ACM revealed that adipokine-induced effects on NK cell expression of the leptin receptor (Ob-R), TRAIL and IFN-γ were species-specific. Conclusion Considering the growing prevalence of obesity and the various disorders related to it, the present study provides further insights into the roles human NK cell subsets play in the obesity-associated state of chronic low-grade inflammation. PMID:24098717

Secretion of Interferon gamma (IFNγ) from Human Immune Cells is Altered by Exposure to Tributyltin (TBT) and Dibutyltin (DBT)

PubMed Central

Lawrence, Shanieek; Reid, Jacqueline; Whalen, Margaret

2013-01-01

Tributyltin (TBT) and dibutyltin (DBT) are widespread environmental contaminants found in food, beverages, and human blood samples. Both of these butyltins (BTs) interfere with the ability of human natural killer (NK) cells to lyse target cells and also alter secretion of the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) from human immune cells in vitro. The capacity of BTs to interfere with secretion of other pro-inflammatory cytokines has not been examined. Interferon gamma (IFNγ) is a modulator of adaptive and innate immune responses, playing an important role in overall immune competence. This study shows that both TBT and DBT alter secretion of IFNγ from human immune cells. Peripheral blood cell preparations that were increasingly reconstituted were used to determine if exposures to either TBT or DBT affected IFNγ secretion and how the makeup of the cell preparation influenced that effect. IFNγ secretion was examined after 24 h, 48 h and 6 day exposures to TBT (200- 2.5 nM) and DBT (5- 0.05 μM) in highly enriched human NK cells, a monocyte-depleted preparation of PBMCs, and monocyte-containing PBMCs. Both BTs altered IFNγ secretion from NK cells at most of the conditions tested (either increasing or decreasing secretion). However, there was significant variability among donors as to the concentrations and time points that showed changes as well as the baseline secretion of IFNγ. The majority of donors showed an increase in IFNγ secretion in response to at least one concentration of TBT or DBT at a minimum of one length of exposure. PMID:24357260

Brief Report: Blockade of TANK-Binding Kinase 1/IKKɛ Inhibits Mutant Stimulator of Interferon Genes (STING)-Mediated Inflammatory Responses in Human Peripheral Blood Mononuclear Cells.

PubMed

Frémond, Marie-Louise; Uggenti, Carolina; Van Eyck, Lien; Melki, Isabelle; Bondet, Vincent; Kitabayashi, Naoki; Hertel, Christina; Hayday, Adrian; Neven, Bénédicte; Rose, Yoann; Duffy, Darragh; Crow, Yanick J; Rodero, Mathieu P

2017-07-01

Gain-of-function mutations in TMEM173, encoding the stimulator of interferon (IFN) genes (STING) protein, underlie a novel type I interferonopathy that is minimally responsive to conventional immunosuppressive therapies and associated with high frequency of childhood morbidity and mortality. STING gain-of-function causes constitutive oversecretion of IFN. This study was undertaken to determine the effects of a TANK-binding kinase 1 (TBK-1)/IKKɛ inhibitor (BX795) on secretion and signaling of IFN in primary peripheral blood mononuclear cells (PBMCs) from patients with mutations in STING. PBMCs from 4 patients with STING-associated disease were treated with BX795. The effect of BX795 on IFN pathways was assessed by Western blotting and an IFNβ reporter assay, as well as by quantification of IFNα in cell lysates, staining for STAT-1 phosphorylation, and measurement of IFN-stimulated gene (ISG) messenger RNA (mRNA) expression. Treatment of PBMCs with BX795 inhibited the phosphorylation of IFN regulatory factor 3 and IFNβ promoter activity induced in HEK 293T cells by cyclic GMP-AMP or by genetic activation of STING. In vitro exposure to BX795 inhibited IFNα production in PBMCs of patients with STING-associated disease without affecting cell survival. In addition, BX795 decreased STAT-1 phosphorylation and ISG mRNA expression independent of IFNα blockade. These findings demonstrate the effect of BX795 on reducing type I IFN production and IFN signaling in cells from patients with gain-of-function mutations in STING. A combined inhibition of TBK-1 and IKKɛ therefore holds potential for the treatment of patients carrying STING mutations, and may also be relevant in other type I interferonopathies. © 2017, American College of Rheumatology.

Amoebal Endosymbiont Protochlamydia Induces Apoptosis to Human Immortal HEp-2 Cells

PubMed Central

Ito, Atsushi; Matsuo, Junji; Nakamura, Shinji; Yoshida, Asahi; Okude, Miho; Hayashi, Yasuhiro; Sakai, Haruna; Yoshida, Mitsutaka; Takahashi, Kaori; Yamaguchi, Hiroyuki

2012-01-01

Protochlamydia, an environmental chlamydia and obligate amoebal endosymbiotic bacterium, evolved to survive within protist hosts, such as Acanthamobae, 700 million years ago. However, these bacteria do not live in vertebrates, including humans. This raises the possibility that interactions between Protochlamydia and human cells could induce a novel cytopathic effect, leading to new insights into host-parasite relationships. Therefore, we studied the effect of Protochlamydia on the survival of human immortal cell line, HEp-2 cells and primary peripheral blood mononuclear cells (PBMC). Using mainly 4′,6-diamidino-2-phenylindole staining, fluorescent in situ hybridization, transmission electron microscopy, and also TUNEL and Transwell assays, we demonstrated that the Protochlamydia induced apoptosis in HEp-2 cells. The attachment of viable bacterial cells, but not an increase of bacterial infectious progenies within the cells, was required for the apoptosis. Other chlamydiae [Parachlamydia acanthamoebae and Chlamydia trachomatis (serovars D and L2)] did not induce the same phenomena, indicating that the observed apoptosis may be specific to the Protochlamydia. Furthermore, the bacteria had no effect on the survival of primary PBMCs collected from five volunteers, regardless of activation. We concluded that Protochlamydia induces apoptosis in human-immortal HEp-2 cells and that this endosymbiont could potentially be used as a biological tool for the elucidation of novel host-parasite relationships. PMID:22276171

Amoebal endosymbiont Protochlamydia induces apoptosis to human immortal HEp-2 cells.

PubMed

Ito, Atsushi; Matsuo, Junji; Nakamura, Shinji; Yoshida, Asahi; Okude, Miho; Hayashi, Yasuhiro; Sakai, Haruna; Yoshida, Mitsutaka; Takahashi, Kaori; Yamaguchi, Hiroyuki

2012-01-01

Protochlamydia, an environmental chlamydia and obligate amoebal endosymbiotic bacterium, evolved to survive within protist hosts, such as Acanthamobae, 700 million years ago. However, these bacteria do not live in vertebrates, including humans. This raises the possibility that interactions between Protochlamydia and human cells could induce a novel cytopathic effect, leading to new insights into host-parasite relationships. Therefore, we studied the effect of Protochlamydia on the survival of human immortal cell line, HEp-2 cells and primary peripheral blood mononuclear cells (PBMC). Using mainly 4',6-diamidino-2-phenylindole staining, fluorescent in situ hybridization, transmission electron microscopy, and also TUNEL and Transwell assays, we demonstrated that the Protochlamydia induced apoptosis in HEp-2 cells. The attachment of viable bacterial cells, but not an increase of bacterial infectious progenies within the cells, was required for the apoptosis. Other chlamydiae [Parachlamydia acanthamoebae and Chlamydia trachomatis (serovars D and L2)] did not induce the same phenomena, indicating that the observed apoptosis may be specific to the Protochlamydia. Furthermore, the bacteria had no effect on the survival of primary PBMCs collected from five volunteers, regardless of activation. We concluded that Protochlamydia induces apoptosis in human-immortal HEp-2 cells and that this endosymbiont could potentially be used as a biological tool for the elucidation of novel host-parasite relationships.

p38 Mitogen-Activated Protein Kinase-γ Inhibition by Long-Acting β2 Adrenergic Agonists Reversed Steroid Insensitivity in Severe Asthma

PubMed Central

Mercado, Nicholas; To, Yasuo; Kobayashi, Yoshiki; Adcock, Ian M.; Barnes, Peter J.

2011-01-01

Corticosteroid insensitivity (CI) is a major barrier to treating severe asthma. Despite intensive research, the molecular mechanism of CI remains uncertain. The aim of this study was to determine abnormality in corticosteroid action in severe asthma and to identify the molecular mechanism of the long-acting β2-adrenergic agonists (LABAs) formoterol and salmeterol on restoration of corticosteroid sensitivity in severe asthma in vitro. Peripheral blood mononuclear cells (PBMCs) were obtained from 16 subjects with severe corticosteroid-insensitive asthma, 6 subjects with mild corticosteroid-sensitive asthma, and 11 healthy volunteers. Corticosteroid (dexamethasone) sensitivity was determined on tumor necrosis factor-α (TNF-α)-induced interleukin (IL)-8 production. Glucocorticoid receptor (GR) phosphorylation and kinase phosphorylation were evaluated by immunoprecipitation-Western blotting analysis and kinase phosphorylation array in IL-2/IL-4-treated corticosteroid insensitive model in PBMCs. In vitro corticosteroid sensitivity on TNF-α-induced IL-8 production was significantly lower in patients with severe asthma than in healthy volunteers and patients with mild asthma. This CI seen in severe asthma was associated with reduced GR nuclear translocation and with hyperphosphorylation of GR, which were reversed by LABAs. In IL-2/IL-4-treated PBMCs, LABAs inhibited phosphorylation of Jun-NH2-terminal kinase and p38 mitogen-activated protein kinase-γ (p38MAPK-γ) as well as GR. In addition, cells with p38MAPK-γ knockdown by RNA interference did not develop CI in the presence of IL-2/IL-4. Furthermore, p38MAPK-γ protein expression was up-regulated in PBMCs from some patients with severe asthma. In conclusion, p38 MAPK-γ activation impairs corticosteroid action and p38 MAPK-γ inhibition by LABAs has potential for the treatment of severe asthma. PMID:21917909

A benzenediamine derivative fc-99 attenuates lupus-like syndrome in MRL/lpr mice related to suppression of pDC activation.

PubMed

Ji, Jianjian; Fan, Hongye; Li, Fanlin; Li, Xiaojing; Dong, Guanjun; Gong, Wei; Song, Yuxian; Liu, Fei; Hua, Chunyan; Tan, Renxiang; Dou, Huan; Hou, Yayi

2015-12-01

Systemic lupus erythematosus (SLE) is an autoimmune disease with prominent chronic inflammatory aspects. Plasmacytoid dendritic cells (pDCs), which are the principal interferon-α (IFN-α)-producing cells, have known to be critically involved in SLE pathogenesis. Our previous research demonstrated that a benzenediamine derivative FC-99 possessed anti-inflammatory activities. However, the effects of FC-99 on SLE have not been investigated to date. In this study, we found that FC-99 attenuated lupus-like pathological symptoms and lupus nephritis as well as the expression of pro-inflammatory cytokines in kidneys of MRL/lpr mice. FC-99 also decreased both the total IgM, total IgG and anti-dsDNA IgG levels in sera and the activation of B cells in the PBMCs and spleens of MRL/lpr mice. Moreover, FC-99 inhibited the abnormal activation and number of pDCs from PBMCs and spleens and levels of IFN-α in MRL/lpr mice. Notably, FC-99 significantly suppressed the expression of IFN-inducible genes in peripheral blood mononuclear cells (PBMCs) and spleens from MRL/lpr mice. As expected, in vitro experiments demonstrated that FC-99 decreased both the activation and IFN-α production of pDCs and inhibited IRAK4 phosphorylation in pDCs upon TLR7 and TLR9 stimulation. We further confirm that the inhibition of FC-99 on B cell activation depended on level of pDCs-secreting IFN-α. These data indicate that FC-99 attenuated lupus-like syndrome in MRL/lpr mice related to suppression of pDC activation, especially pDCs-secreting IFN-α. This study suggests that FC-99 may be a potential therapeutic candidate for the treatment of SLE. Copyright © 2015 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Integrated Extravehicular Activity Human Research Plan: 2017

NASA Technical Reports Server (NTRS)

Abercromby, Andrew

2017-01-01

Multiple organizations within NASA as well as industry and academia fund and participate in research related to extravehicular activity (EVA). In October 2015, representatives of the EVA Office, the Crew and Thermal Systems Division (CTSD), and the Human Research Program (HRP) at NASA Johnson Space Center agreed on a formal framework to improve multi-year coordination and collaboration in EVA research. At the core of the framework is an Integrated EVA Human Research Plan and a process by which it will be annually reviewed and updated. The over-arching objective of the collaborative framework is to conduct multi-disciplinary cost-effective research that will enable humans to perform EVAs safely, effectively, comfortably, and efficiently, as needed to enable and enhance human space exploration missions. Research activities must be defined, prioritized, planned and executed to comprehensively address the right questions, avoid duplication, leverage other complementary activities where possible, and ultimately provide actionable evidence-based results in time to inform subsequent tests, developments and/or research activities. Representation of all appropriate stakeholders in the definition, prioritization, planning and execution of research activities is essential to accomplishing the over-arching objective. A formal review of the Integrated EVA Human Research Plan will be conducted annually. Coordination with stakeholders outside of the EVA Office, CTSD, and HRP is already in effect on a study-by-study basis; closer coordination on multi-year planning with other EVA stakeholders including academia is being actively pursued. Details of the preliminary Integrated EVA Human Research Plan are presented including description of ongoing and planned research activities in the areas of: physiological and performance capabilities; suit design parameters; EVA human health and performance modeling; EVA tasks and concepts of operations; EVA informatics; human-suit sensors; suit

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

NASA Technical Reports Server (NTRS)

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

1999-01-01

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

Persistent Alterations of Gene Expression Profiling of Human Peripheral Blood Mononuclear Cells From Smokers

PubMed Central

Weng, Daniel Y.; Chen, Jinguo; Taslim, Cenny; Hsu, Ping-Ching; Marian, Catalin; David, Sean P.; Loffredo, Christopher A.; Shields, Peter G.

2016-01-01

The number of validated biomarkers of tobacco smoke exposure is limited, and none exist for tobacco-related cancer. Additional biomarkers for smoke, effects on cellular systems in vivo are needed to improve early detection of lung cancer, and to assist the Food and Drug Administration in regulating exposures to tobacco products. We assessed the effects of smoking on the gene expression using human cell cultures and blood from a cross-sectional study. We profiled global transcriptional changes in cultured smokers’ peripheral blood mononuclear cells (PBMCs) treated with cigarette smoke condensate (CSC) in vitro (n = 7) and from well-characterized smokers’ blood (n = 36). ANOVA with adjustment for covariates and Pearson correlation were used for statistical analysis in this study. CSC in vitro altered the expression of 1 178 genes (177 genes with > 1.5-fold-change) at P < 0.05. In vivo, PBMCs of heavy and light smokers differed for 614 genes (29 with > 1.5-fold-change) at P < 0.05 (309 remaining significant after adjustment for age, race, and gender). Forty-one genes were persistently altered both in vitro and in vivo, 22 having the same expression pattern reported for non-small cell lung cancer. Our data provides evidence that persistent alterations of gene expression in vitro and in vivo may relate to carcinogenic effects of cigarette smoke, and the identified genes may serve as potential biomarkers for cancer. The use of an in vitro model to corroborate results from human studies provides a novel way to understand human exposure and effect. PMID:26294040

Detection of circulating tumor cells from cryopreserved human sarcoma peripheral blood mononuclear cells.

PubMed

Li, Heming; Meng, Qing H; Noh, Hyangsoon; Batth, Izhar Singh; Somaiah, Neeta; Torres, Keila E; Xia, Xueqing; Wang, Ruoyu; Li, Shulin

2017-09-10

Circulating tumor cells (CTCs) enter the vasculature or lymphatic system after shedding from the primary tumor. CTCs may serve as "seed" cells for tumor metastasis. The utility of CTCs in clinical applications for sarcoma is not fully investigated, partly owing to the necessity for fresh blood samples and the lack of a CTC-specific antibody. To overcome these drawbacks, we developed a technique for sarcoma CTCs capture and detection using cryopreserved peripheral blood mononuclear cells (PBMCs) and our proprietary cell-surface vimentin (CSV) antibody 84-1, which is specific to tumor cells. This technique was validated by sarcoma cell spiking assay, matched CTCs comparison between fresh and cryopreserved PBMCs, and independent tumor markers in multiple types of sarcoma patient blood samples. The reproducibility was maximized when cryopreserved PBMCs were prepared from fresh blood samples within 2 h of the blood draw. In summary, as far as we are aware, ours is the first report to capture and detect CTCs from cryopreserved PBMCs. Further validation in other types of tumor may help boost the feasibility and utility of CTC-based diagnosis in a centralized laboratory. Copyright © 2017 Elsevier B.V. All rights reserved.

Immune activation underlies a sustained clinical response to Yttrium-90 radioembolisation in hepatocellular carcinoma.

PubMed

Chew, Valerie; Lee, Yun Hua; Pan, Lu; Nasir, Nurul J M; Lim, Chun Jye; Chua, Camillus; Lai, Liyun; Hazirah, Sharifah Nur; Lim, Tony Kiat Hon; Goh, Brian K P; Chung, Alexander; Lo, Richard H G; Ng, David; Filarca, Rene L F; Albani, Salvatore; Chow, Pierce K H

2018-02-13

Yttrium-90 (Y90)-radioembolisation (RE) significantly regresses locally advanced hepatocellular carcinoma and delays disease progression. The current study is designed to deeply interrogate the immunological impact of Y90-RE, which elicits a sustained therapeutic response. Time-of-flight mass cytometry and next-generation sequencing (NGS) were used to analyse the immune landscapes of tumour-infiltrating leucocytes (TILs), tumour tissues and peripheral blood mononuclear cells (PBMCs) at different time points before and after Y90-RE. TILs isolated after Y90-RE exhibited signs of local immune activation: higher expression of granzyme B (GB) and infiltration of CD8 + T cells, CD56 + NK cells and CD8 + CD56 + NKT cells. NGS confirmed the upregulation of genes involved in innate and adaptive immune activation in Y90-RE-treated tumours. Chemotactic pathways involving CCL5 and CXCL16 correlated with the recruitment of activated GB + CD8 + T cells to the Y90-RE-treated tumours. When comparing PBMCs before and after Y90-RE, we observed an increase in tumour necrosis factor-α on both the CD8 + and CD4 + T cells as well as an increase in percentage of antigen-presenting cells after Y90-RE, implying a systemic immune activation. Interestingly, a high percentage of PD-1 + /Tim-3 + CD8 + T cells coexpressing the homing receptors CCR5 and CXCR6 denoted Y90-RE responders. A prediction model was also built to identify sustained responders to Y90-RE based on the immune profiles from pretreatment PBMCs. High-dimensional analysis of tumour and systemic immune landscapes identified local and systemic immune activation that corresponded to the sustained response to Y90-RE. Potential biomarkers associated with a positive clinical response were identified and a prediction model was built to identify sustained responders prior to treatment. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is

Effects of Arsenic Trioxide on INF-gamma Gene Expression in MRL/lpr Mice and Human Lupus.

PubMed

Hu, Hongye; Chen, Enjiu; Li, Yongji; Zhu, Xiaochun; Zhang, Ting; Zhu, Xiaofang

2017-11-20

Arsenic trioxide (As2O3; ATO), a traditional Chinese medicine, is used to treat patients with acute promye-locytic leukemia, while its application for treatment of systemic lupus erythematosus (SLE) is still under evaluation. The high expression of INF-gamma (INF-γ) is a primary pathogenic factor in SLE. It is found that ATO can reduce INF-γ expression levels in lupus-prone mice, whereas it is not clear whether ATO has the same effect on SLE patients. Therefore, this study was to investigate the underlying mechanism of the effects of ATO on the expression of INF-γ in splenocytes of MRL/lpr mice and PBMCs of human lupus. The mRNA and protein expression levels of INF-γ were assessed by real-time RT-PCR and ELISA, respectively. The histone acetylation status of the INF-γ promoter and the binding of RNA polymerase II (RNA Pol II) to the INF-γ promoter were detected using a chromatin immunoprecipitation (ChIP) technique. The mRNA and protein expression levels of INF-γ decreased in both splenocytes of MRL/lpr mice and PBMCs of SLE patients with ATO treatment, which were accompanied by reduced histone H4 and H3 acetylation in INF-γ promoter and decreased combination of RNA Pol II to the INF-γ promoter. Therefore, ATO may reduce the expression level of the INF-γ by altering the levels of INF-γ promoter acetylation and the combination of RNA Pol II to the INF-γ promoter in splenocytes of MRL/lpr mice and PBMCs of SLE patients.

Mucorales spores induce a proinflammatory cytokine response in human mononuclear phagocytes and harbor no rodlet hydrophobins.

PubMed

Wurster, Sebastian; Thielen, Vanessa; Weis, Philipp; Walther, Paul; Elias, Johannes; Waaga-Gasser, Ana Maria; Dragan, Mariola; Dandekar, Thomas; Einsele, Hermann; Löffler, Jürgen; Ullmann, Andrew J

2017-11-17

Mucormycoses are life-threatening infections in immunocompromised patients. This study characterizes the response of human mononuclear cells to different Mucorales and Ascomycota. PBMC, monocytes, and monocyte derived dendritic cells (moDCs) from healthy donors were stimulated with resting and germinated stages of Mucorales and Ascomycota. Cytokine response and expression of activation markers were studied. Both inactivated germ tubes and resting spores of Rhizopus arrhizus and other human pathogenic Mucorales species significantly stimulated mRNA synthesis and secretion of proinflammatory cytokines. Moreover, R. arrhizus spores induced the upregulation of co-stimulatory molecules on moDCs and a specific T-helper cell response. Removal of rodlet hydrophobins by hydrofluoric acid treatment of A. fumigatus conidia resulted in enhanced immunogenicity, whereas the cytokine response of PBMCs to dormant R. arrhizus spores was not influenced by hydrofluoric acid. Scanning electron micrographs of Mucorales spores did not exhibit any morphological correlates of rodlet hydrophobins. Taken together, this study revealed striking differences in the response of human mononuclear cells to resting stages of Ascomycota and Mucorales, which may be explained by absence of an immunoprotective hydrophobin layer in Mucorales spores.

Cocoa Consumption Alters the Global DNA Methylation of Peripheral Leukocytes in Humans with Cardiovascular Disease Risk Factors: A Randomized Controlled Trial

PubMed Central

Crescenti, Anna; Solà, Rosa; Valls, Rosa M.; Caimari, Antoni; del Bas, Josep M.; Anguera, Anna; Anglés, Neus; Arola, Lluís

2013-01-01

DNA methylation regulates gene expression and can be modified by different bioactive compounds in foods, such as polyphenols. Cocoa is a rich source of polyphenols, but its role in DNA methylation is still unknown. The objective was to assess the effect of cocoa consumption on DNA methylation and to determine whether the enzymes involved in the DNA methylation process participate in the mechanisms by which cocoa exerts these effects in humans. The global DNA methylation levels in the peripheral blood were evaluated in 214 volunteers who were pre-hypertensive, stage-1 hypertensive or hypercholesterolemic. The volunteers were divided into two groups: 110 subjects who consumed cocoa (6 g/d) for two weeks and 104 control subjects. In addition, the peripheral blood mononuclear cells (PBMCs) from six subjects were treated with a cocoa extract to analyze the mRNA levels of the DNA methyltransferases (DNMTs), methylenetetrahydrofolate reductase (MTHFR), and methionine synthase reductase (MTRR) genes. Cocoa consumption significantly reduced the DNA methylation levels (2.991±0.366 vs. 3.909±0.380, p<0.001). Additionally, we found an association between the cocoa effects on DNA methylation and three polymorphisms located in the MTHFR, MTRR, and DNMT3B genes. Furthermore, in PBMCs, the cocoa extract significantly lowered the mRNA levels of the DNMTs, MTHFR, and MTRR. Our study demonstrates for the first time that the consumption of cocoa decreases the global DNA methylation of peripheral leukocytes in humans with cardiovascular risk factors. In vitro experiments with PBMCs suggest that cocoa may exert this effect partially via the down-regulation of DNMTs, MTHFR and MTRR, which are key genes involved in this epigenetic process. Trial Registration Clinicaltrials.gov NCT00511420 and NCT00502047 PMID:23840361

Cocoa Consumption Alters the Global DNA Methylation of Peripheral Leukocytes in Humans with Cardiovascular Disease Risk Factors: A Randomized Controlled Trial.

PubMed

Crescenti, Anna; Solà, Rosa; Valls, Rosa M; Caimari, Antoni; Del Bas, Josep M; Anguera, Anna; Anglés, Neus; Arola, Lluís

2013-01-01

DNA methylation regulates gene expression and can be modified by different bioactive compounds in foods, such as polyphenols. Cocoa is a rich source of polyphenols, but its role in DNA methylation is still unknown. The objective was to assess the effect of cocoa consumption on DNA methylation and to determine whether the enzymes involved in the DNA methylation process participate in the mechanisms by which cocoa exerts these effects in humans. The global DNA methylation levels in the peripheral blood were evaluated in 214 volunteers who were pre-hypertensive, stage-1 hypertensive or hypercholesterolemic. The volunteers were divided into two groups: 110 subjects who consumed cocoa (6 g/d) for two weeks and 104 control subjects. In addition, the peripheral blood mononuclear cells (PBMCs) from six subjects were treated with a cocoa extract to analyze the mRNA levels of the DNA methyltransferases (DNMTs), methylenetetrahydrofolate reductase (MTHFR), and methionine synthase reductase (MTRR) genes. Cocoa consumption significantly reduced the DNA methylation levels (2.991±0.366 vs. 3.909±0.380, p<0.001). Additionally, we found an association between the cocoa effects on DNA methylation and three polymorphisms located in the MTHFR, MTRR, and DNMT3B genes. Furthermore, in PBMCs, the cocoa extract significantly lowered the mRNA levels of the DNMTs, MTHFR, and MTRR. Our study demonstrates for the first time that the consumption of cocoa decreases the global DNA methylation of peripheral leukocytes in humans with cardiovascular risk factors. In vitro experiments with PBMCs suggest that cocoa may exert this effect partially via the down-regulation of DNMTs, MTHFR and MTRR, which are key genes involved in this epigenetic process. Clinicaltrials.govNCT00511420 and NCT00502047.

Expression of 11beta-hydroxysteroid-dehydrogenase type 2 in human thymus.

PubMed

Almanzar, Giovanni; Mayerl, Christina; Seitz, Jan-Christoph; Höfner, Kerstin; Brunner, Andrea; Wild, Vanessa; Jahn, Daniel; Geier, Andreas; Fassnacht, Martin; Prelog, Martina

2016-06-01

11beta-hydroxysteroid-dehydrogenase type 2 (11β-HSD2) is a high affinity dehydrogenase which rapidly inactivates physiologically-active glucocorticoids to protect key tissues. 11β-HSD2 expression has been described in peripheral cells of the innate and the adaptive immune system as well as in murine thymus. In absence of knowledge of 11β-HSD2 expression in human thymus, the study aimed to localize 11β-HSD2 in human thymic tissue. Thymic tissue was taken of six healthy, non-immunologically impaired male infants below 12months of age with congenital heart defects who had to undergo correction surgery. 11β-HSD2 protein expression was analyzed by immunohistochemistry and Western blot. Kidney tissue, peripheral blood mononuclear cells (PBMCs) and human umbilical vein endothelial cells (HUVEC) were taken as positive controls. Significant expression of 11β-HSD2 protein was found at single cell level in thymus parenchyma, at perivascular sites of capillaries and small vessels penetrating the thymus lobuli and within Hassall's bodies. The present study demonstrates that 11β-HSD2 is expressed in human thymus with predominant perivascular expression and also within Hassall's bodies. To our knowledge, this is the first report confirming 11β-HSD2 expression at the protein level in human thymic tissue underlining a potential role of this enzyme in regulating glucocorticoid function at the thymic level. Copyright © 2016 Elsevier Inc. All rights reserved.

Human Activity Modeling and Simulation with High Biofidelity

DTIC Science & Technology

2013-01-01

Human activity Modeling and Simulation (M&S) plays an important role in simulation-based training and Virtual Reality (VR). However, human activity M...kinematics and motion mapping/creation; and (e) creation and replication of human activity in 3-D space with true shape and motion. A brief review is

Ci8 short, a novel LPS-induced peptide from the ascidian Ciona intestinalis, modulates responses of the human immune system.

PubMed

Bonura, Angela; Vizzini, Aiti; Vlah, Sara; Gervasi, Francesco; Longo, Alessandra; Melis, Mario R; Schildberg, Frank A; Colombo, Paolo

2018-02-01

The selective modulation of immunity is an emerging concept driven by the vast advances in our understanding of this crucial host defense system. Invertebrates have raised researchers' interest as potential sources of new bioactive molecules owing to their antibacterial, anticancer and immunomodulatory activities. A LipoPolySaccharide (LPS) challenge in the ascidian Ciona intestinalis generates the transcript, Ci8 short, with cis-regulatory elements in the 3' UTR region that are essential for shaping innate immune responses. The derived amino acidic sequence in silico analysis showed specific binding to human Major Histocompatibility Complex (MHC) Class I and Class II alleles. The role of Ci8 short peptide was investigated in a more evolved immune system using human Peripheral Blood Mononuclear Cells (PBMCs) as in vitro model. The biological activities of this molecule include the activation of 70kDa TCR ζ chain Associated Protein kinase (ZAP-70) and T Cell Receptor (TCR) Vβ oligo clonal selection on CD4 + T lymphocytes as well as increased proliferation and IFN-γ secretion. Furthermore Ci8 short affects CD4 + /CD25 high induced regulatory T cells (iTreg) subset selection which co-expressed the functional markers TGF-β1/Latency Associated Protein (LAP) and CD39/CD73. This paper describes a new molecule that modulates important responses of the human adaptive immune system. Copyright © 2017 Elsevier GmbH. All rights reserved.

Anti-complement activities of human breast-milk.

PubMed

Ogundele, M O

1999-08-01

It has long been observed that the human milk possesses significant anti-inflammatory properties, while simultaneously protecting the infant against many intestinal and respiratory pathogens. There is, however, a paucity of information on the degree and extent of this anti-inflammatory activity. In the present study, the inhibitory effects of different fractions of human milk on serum complement activity were analysed. Colostrum and milk samples from healthy voluntary lactating donors at different postpartum ages were obtained and pooled normal human serum was used as source of complement in a modified CH50 assay. Inherent complement activity in human milk was also investigated by measuring the deposition of an activated C3 fragment on a serum-sensitive bacteria, and by haemolytic assays. Most whole- and defatted-milk samples consistently showed a dose-dependent inhibition of the serum complement activity. This inhibition was greater in mature milk compared to transitional milk samples. It was enhanced by inactivation of milk complement, and diminished by centrifugation of milk samples, which partly removed fat and larger protein components including casein micelles. Inherent complement activity in human milk was also demonstrated by haemolysis of sensitised sheep erythrocytes and deposition of C3 fragments on solid-phase bacteria. These activities were highest in the colostrum and gradually decreased as lactation proceeded. Several natural components abundant in the fluid phase of the human breast-milk have been shown to be inhibitors of complement activation in vitro. Their physiological significance probably reside in their ability to prevent inflammatory-induced tissue damage of the delicate immature gastrointestinal tract of the new-born as well as the mammary gland itself, which may arise from ongoing complement activation.

The IDO1 selective inhibitor epacadostat enhances dendritic cell immunogenicity and lytic ability of tumor antigen-specific T cells.

PubMed

Jochems, Caroline; Fantini, Massimo; Fernando, Romaine I; Kwilas, Anna R; Donahue, Renee N; Lepone, Lauren M; Grenga, Italia; Kim, Young-Seung; Brechbiel, Martin W; Gulley, James L; Madan, Ravi A; Heery, Christopher R; Hodge, James W; Newton, Robert; Schlom, Jeffrey; Tsang, Kwong Y

2016-06-21

Epacadostat is a novel inhibitor of indoleamine-2,3-dioxygenase-1 (IDO1) that suppresses systemic tryptophan catabolism and is currently being evaluated in ongoing clinical trials. We investigated the effects of epacadostat on (a) human dendritic cells (DCs) with respect to maturation and ability to activate human tumor antigen-specific cytotoxic T-cell (CTL) lines, and subsequent T-cell lysis of tumor cells, (b) human regulatory T cells (Tregs), and (c) human peripheral blood mononuclear cells (PBMCs) in vitro. Simultaneous treatment with epacadostat and IFN-γ plus lipopolysaccharide (LPS) did not change the phenotype of matured human DCs, and as expected decreased the tryptophan breakdown and kynurenine production. Peptide-specific T-cell lines stimulated with DCs pulsed with peptide produced significantly more IFN-γ, TNFα, GM-CSF and IL-8 if the DCs were treated with epacadostat. These T cells also displayed higher levels of tumor cell lysis on a per cell basis. Epacadostat also significantly decreased Treg proliferation induced by IDO production from IFN-γ plus LPS matured human DCs, although the Treg phenotype did not change. Multicolor flow cytometry was performed on human PBMCs treated with epacadostat; analysis of 123 discrete immune cell subsets revealed no changes in major immune cell types, an increase in activated CD83+ conventional DCs, and a decrease in immature activated Tim3+ NK cells. These studies show for the first time several effects of epacadostat on human DCs, and subsequent effects on CTL and Tregs, and provide a rationale as to how epacadostat could potentially increase the efficacy of immunotherapeutics, including cancer vaccines.

Real-time Human Activity Recognition

NASA Astrophysics Data System (ADS)

Albukhary, N.; Mustafah, Y. M.

2017-11-01

The traditional Closed-circuit Television (CCTV) system requires human to monitor the CCTV for 24/7 which is inefficient and costly. Therefore, there’s a need for a system which can recognize human activity effectively in real-time. This paper concentrates on recognizing simple activity such as walking, running, sitting, standing and landing by using image processing techniques. Firstly, object detection is done by using background subtraction to detect moving object. Then, object tracking and object classification are constructed so that different person can be differentiated by using feature detection. Geometrical attributes of tracked object, which are centroid and aspect ratio of identified tracked are manipulated so that simple activity can be detected.

Anti-leukemia activity of a bacterial toxin with natural specificity for LFA-1 on white blood cells

PubMed Central

Kachlany, Scott C.; Schwartz, Amy B.; Balashova, Nataliya V.; Hioe, Catarina E.; Tuen, Michael; Le, Amy; Kaur, Manpreet; Mei, Yongyi; Rao, Jia

2009-01-01

The oral bacterium, Aggregatibacter actinomycetemcomitans, produces a leukotoxin (LtxA) that is specific for white blood cells (WBCs) from humans and Old World primates by interacting with lymphocyte function antigen-1 (LFA-1) on susceptible cells. To determine if LtxA could be used as a therapeutic agent for the treatment of WBC diseases, we tested the in vitro and in vivo anti-leukemia activity of the toxin. LtxA kills human malignant WBC lines and primary leukemia cells from acute myeloid leukemia patients, but healthy peripheral blood mononuclear cells (PBMCs) are relatively resistant to LtxA-mediated cytotoxicity. Levels of LFA-1 on cell lines correlated with killing by LtxA and the toxin preferentially killed cells expressing the activated form of LFA-1. In a SCID mouse model for human leukemia, LtxA had potent therapeutic value resulting in long-term survival in LtxA-treated mice. Intravenous infusion of LtxA into a rhesus macaque resulted in a drop in WBC counts at early times post-infusion; however, red blood cells, platelets, hemoglobin and blood chemistry values remained unaffected. Thus, LtxA may be an effective and safe novel therapeutic agent for the treatment of hematologic malignancies. PMID:19747730

Understanding human dynamics in microblog posting activities

NASA Astrophysics Data System (ADS)

Jiang, Zhihong; Zhang, Yubao; Wang, Hui; Li, Pei

2013-02-01

Human activity patterns are an important issue in behavior dynamics research. Empirical evidence indicates that human activity patterns can be characterized by a heavy-tailed inter-event time distribution. However, most researchers give an understanding by only modeling the power-law feature of the inter-event time distribution, and those overlooked non-power-law features are likely to be nontrivial. In this work, we propose a behavior dynamics model, called the finite memory model, in which humans adaptively change their activity rates based on a finite memory of recent activities, which is driven by inherent individual interest. Theoretical analysis shows a finite memory model can properly explain various heavy-tailed inter-event time distributions, including a regular power law and some non-power-law deviations. To validate the model, we carry out an empirical study based on microblogging activity from thousands of microbloggers in the Celebrity Hall of the Sina microblog. The results show further that the model is reasonably effective. We conclude that finite memory is an effective dynamics element to describe the heavy-tailed human activity pattern.

Human body contour data based activity recognition.

PubMed

Myagmarbayar, Nergui; Yuki, Yoshida; Imamoglu, Nevrez; Gonzalez, Jose; Otake, Mihoko; Yu, Wenwei

2013-01-01

This research work is aimed to develop autonomous bio-monitoring mobile robots, which are capable of tracking and measuring patients' motions, recognizing the patients' behavior based on observation data, and providing calling for medical personnel in emergency situations in home environment. The robots to be developed will bring about cost-effective, safe and easier at-home rehabilitation to most motor-function impaired patients (MIPs). In our previous research, a full framework was established towards this research goal. In this research, we aimed at improving the human activity recognition by using contour data of the tracked human subject extracted from the depth images as the signal source, instead of the lower limb joint angle data used in the previous research, which are more likely to be affected by the motion of the robot and human subjects. Several geometric parameters, such as, the ratio of height to weight of the tracked human subject, and distance (pixels) between centroid points of upper and lower parts of human body, were calculated from the contour data, and used as the features for the activity recognition. A Hidden Markov Model (HMM) is employed to classify different human activities from the features. Experimental results showed that the human activity recognition could be achieved with a high correct rate.

Bioavailability of herbs and spices in humans as determined by ex vivo inflammatory suppression and DNA strand breaks.

PubMed

Percival, Susan S; Vanden Heuvel, John P; Nieves, Carmelo J; Montero, Cindy; Migliaccio, Andrew J; Meadors, Joanna

2012-08-01

The aim of this work was to determine the bioavailability of herbs and spices after human consumption by measuring the ability to protect lymphocytes from an oxidative injury and by examining the impact on inflammatory biomarkers in activated THP-1 cells. Ten to 12 subjects in each of 13 groups consumed a defined amount of herb or spice for 7 days. Blood was drawn from subjects before consumption and 1 hour after taking the final herb or spice capsules. Subject serum and various extractions of the herbs and spices were analyzed for antioxidant capacity by oxygen radical absorbance capacity (ORAC) analysis or by 1,1-diphenyl-2-picrylhydrzyl (DPPH). Subject peripheral blood mononuclear cells (PBMCs) in medium with10% autologous serum were incubated with hydrogen peroxide to induce DNA strand breaks. Subject serum was also used to treat activated THP-1 cells to determine relative quantities of 3 inflammatory cytokine (tumor necrosis factor-α [TNF-α], interleukin-1α [IL-1α], and IL-6) mRNAs. Herbs and spices that protected PBMCs against DNA strand breaks were paprika, rosemary, ginger, heat-treated turmeric, sage, and cumin. Paprika also appeared to protect cells from normal apoptotic processes. Of the 3 cytokine mRNAs studied (TNF-α, IL-1α, and IL-6), TNF-α was the most sensitive responder to oxidized LDL-treated macrophages. Clove, ginger, rosemary, and turmeric were able to significantly reduce oxidized LDL-induced expression of TNF-α. Serum from those consuming ginger reduced all three inflammatory biomarkers. Ginger, rosemary, and turmeric showed protective capacity by both oxidative protection and inflammation measures. DNA strand breaks and inflammatory biomarkers are a good functional measure of a food's bioavailability.

Pegylated interferons Lambda-1a and alfa-2a display different gene induction and cytokine and chemokine release profiles in whole blood, human hepatocytes and peripheral blood mononuclear cells.

PubMed

Freeman, J; Baglino, S; Friborg, J; Kraft, Z; Gray, T; Hill, M; McPhee, F; Hillson, J; Lopez-Talavera, J C; Wind-Rotolo, M

2014-06-01

Pegylated interferon-lambda-1a (Lambda), a type III interferon (IFN) in clinical development for the treatment of chronic HCV infection, has shown comparable efficacy and an improved safety profile to a regimen based on pegylated IFN alfa-2a (alfa). To establish a mechanistic context for this improved profile, we investigated the ex vivo effects of Lambda and alfa on cytokine and chemokine release, and on expression of IFN-stimulated genes (ISGs) in primary human hepatocytes and peripheral blood mononuclear cells (PBMCs) from healthy subjects. Our findings were further compared with changes observed in blood analysed from HCV-infected patients treated with Lambda or alfa in clinical studies. mRNA transcript and protein expression of the IFN-λ-limiting receptor subunit was lower compared with IFN-α receptor subunits in all cell types. Upon stimulation, alfa and Lambda induced ISG expression in hepatocytes and PBMCs, although in PBMCs Lambda-induced ISG expression was modest. Furthermore, alfa and Lambda induced release of cytokines and chemokines from hepatocytes and PBMCs, although differences in their kinetics of induction were observed. In HCV-infected patients, alfa treatment induced ISG expression in whole blood after single and repeat dosing. Lambda treatment induced modest ISG expression after single dosing and showed no induction after repeat dosing. Alfa and Lambda treatment increased IP-10, iTAC, IL-6, MCP-1 and MIP-1β levels in serum, with alfa inducing higher levels of all mediators compared with Lambda. Overall, ex vivo and in vivo induction profiles reported in this analysis strongly correlate with clinical observations of fewer related adverse events for Lambda vs those typically associated with alfa. © 2014 John Wiley & Sons Ltd.

Human T-cell leukemia virus type I (HTLV-1) proviral load and disease progression in asymptomatic HTLV-1 carriers: a nationwide prospective study in Japan.

PubMed

Iwanaga, Masako; Watanabe, Toshiki; Utsunomiya, Atae; Okayama, Akihiko; Uchimaru, Kaoru; Koh, Ki-Ryang; Ogata, Masao; Kikuchi, Hiroshi; Sagara, Yasuko; Uozumi, Kimiharu; Mochizuki, Manabu; Tsukasaki, Kunihiro; Saburi, Yoshio; Yamamura, Masaomi; Tanaka, Junji; Moriuchi, Yukiyoshi; Hino, Shigeo; Kamihira, Shimeru; Yamaguchi, Kazunari

2010-08-26

Definitive risk factors for the development of adult T-cell leukemia (ATL) among asymptomatic human T-cell leukemia virus type I (HTLV-1) carriers remain unclear. Recently, HTLV-1 proviral loads have been evaluated as important predictors of ATL, but a few small prospective studies have been conducted. We prospectively evaluated 1218 asymptomatic HTLV-1 carriers (426 males and 792 females) who were enrolled during 2002 to 2008. The proviral load at enrollment was significantly higher in males than females (median, 2.10 vs 1.39 copies/100 peripheral blood mononuclear cells [PBMCs]; P < .001), in those 40 to 49 and 50 to 59 years of age than that of those 40 years of age and younger (P = .02 and .007, respectively), and in those with a family history of ATL than those without the history (median, 2.32 vs 1.33 copies/100 PBMCs; P = .005). During follow-up, 14 participants progressed to overt ATL. Their baseline proviral load was high (range, 4.17-28.58 copies/100 PBMCs). None developed ATL among those with a baseline proviral load lower than approximately 4 copies. Multivariate Cox analyses indicated that not only a higher proviral load, advanced age, family history of ATL, and first opportunity for HTLV-1 testing during treatment for other diseases were independent risk factors for progression of ATL.

Spectrum and mechanisms of inflammasome activation by chitosan.

PubMed

Bueter, Chelsea L; Lee, Chrono K; Wang, Jennifer P; Ostroff, Gary R; Specht, Charles A; Levitz, Stuart M

2014-06-15

Chitosan, the deacetylated derivative of chitin, can be found in the cell wall of some fungi and is used in translational applications. We have shown that highly purified preparations of chitosan, but not chitin, activate the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome in primed mouse bone marrow-derived macrophages (BMMΦ), inducing a robust IL-1β response. In this article, we further define specific cell types that are activated and delineate mechanisms of activation. BMMΦ differentiated to promote a classically activated (M1) phenotype released more IL-1β in response to chitosan than intermediate or alternatively activated macrophages (M2). Chitosan, but not chitin, induced a robust IL-1β response in mouse dendritic cells, peritoneal macrophages, and human PBMCs. Three mechanisms for NLRP3 inflammasome activation may contribute: K(+) efflux, reactive oxygen species, and lysosomal destabilization. The contributions of these mechanisms were tested using a K(+) efflux inhibitor, high extracellular potassium, a mitochondrial reactive oxygen species inhibitor, lysosomal acidification inhibitors, and a cathepsin B inhibitor. These studies revealed that each of these pathways participated in optimal NLRP3 inflammasome activation by chitosan. Finally, neither chitosan nor chitin stimulated significant release from unprimed BMMΦ of any of 22 cytokines and chemokines assayed. This study has the following conclusions: 1) chitosan, but not chitin, stimulates IL-1β release from multiple murine and human cell types; 2) multiple nonredundant mechanisms appear to participate in inflammasome activation by chitosan; and 3) chitin and chitosan are relatively weak stimulators of inflammatory mediators from unprimed BMMΦ. These data have implications for understanding the nature of the immune response to microbes and biomaterials that contain chitin and chitosan. Copyright © 2014 by The American Association of Immunologists, Inc.

Red ginseng and vitamin C increase immune cell activity and decrease lung inflammation induced by influenza A virus/H1N1 infection.

PubMed

Kim, Hyemin; Jang, Mirim; Kim, Yejin; Choi, Jiyea; Jeon, Jane; Kim, Jihoon; Hwang, Young-Il; Kang, Jae Seung; Lee, Wang Jae

2016-03-01

Because red ginseng and vitamin C have immunomodulatory function and anti-viral effect, we investigated whether red ginseng and vitamin C synergistically regulate immune cell function and suppress viral infection. Red ginseng and vitamin C were treated to human peripheral blood mononuclear cells (PBMCs) or sarcoma-associated herpesvirus (KSHV)-infected BCBL-1, and administrated to Gulo(-/-) mice, which are incapable of synthesizing vitamin C, with or without influenza A virus/H1N1 infection. Red ginseng and vitamin C increased the expression of CD25 and CD69 of PBMCs and natural killer (NK) cells. Co-treatment of them decreased cell viability and lytic gene expression in BCBL-1. In Gulo(-/-) mice, red ginseng and vitamin C increased the expression of NKp46, a natural cytotoxic receptor of NK cells and interferon (IFN)-γ production. Influenza infection decreased the survival rate, and increased inflammation and viral plaque accumulation in the lungs of vitamin C-depleted Gulo(-/-) mice, which were remarkably reduced by red ginseng and vitamin C supplementation. Administration of red ginseng and vitamin C enhanced the activation of immune cells like T and NK cells, and repressed the progress of viral lytic cycle. It also reduced lung inflammation caused by viral infection, which consequently increased the survival rate. © 2016 Royal Pharmaceutical Society, Journal of Pharmacy and Pharmacology.

Biphasic Elimination of Tenofovir Diphosphate and Nonlinear Pharmacokinetics of Zidovudine Triphosphate in a Microdosing Study

PubMed Central

Chen, Jianmeng; Flexner, Charles; Liberman, Rosa G.; Skipper, Paul L.; Louissaint, Nicolette; Tannenbaum, Steven R.; Hendrix, Craig; Fuchs, Edward

2012-01-01

Objective Phase 0 studies can provide initial pharmacokinetics (PK) data in humans and help to facilitate early drug development, but their predictive value for standard dosing is controversial. To evaluate the prediction of microdosing for active intracellular drug metabolites, we compared the PK profile of two antiretroviral drugs, zidovudine (ZDV) and tenofovir (TFV), in microdose and standard dosing regimens. Study Design We administered a microdose (100 μg) of 14C-labeled drug (ZDV or tenofovir disoproxil fumarate (TDF)) with or without a standard unlabelled dose (300 mg) to healthy volunteers. Both the parent drug in plasma and the active metabolite, ZDV-triphosphate (ZDV-TP) or TFV-diphosphate (TFV-DP) in PBMCs and CD4+ cells were measured by AMS. Results The intracellular ZDV-TP concentration increased less than proportionally over the dose range studied (100 μg to 300 mg), while the intracellular TFV-DP PK were linear over the same dose range. ZDV-TP concentrations were lower in CD4+ cells versus total peripheral blood mononuclear cells (PBMCs), while TFV-DP concentrations were not different in CD4+ cells and PBMCs. Conclusion Our data were consistent with a rate-limiting step in the intracellular phosphorylation of ZDV but not TFV. AMS shows promise for predicting the PK of active intracellular metabolites of nucleosides, but nonlinearity of PK may be seen with some drugs. PMID:23187888

Scaling behavior of online human activity

NASA Astrophysics Data System (ADS)

Zhao, Zhi-Dan; Cai, Shi-Min; Huang, Junming; Fu, Yan; Zhou, Tao

2012-11-01

The rapid development of the Internet technology enables humans to explore the web and record the traces of online activities. From the analysis of these large-scale data sets (i.e., traces), we can get insights about the dynamic behavior of human activity. In this letter, the scaling behavior and complexity of human activity in the e-commerce, such as music, books, and movies rating, are comprehensively investigated by using the detrended fluctuation analysis technique and the multiscale entropy method. Firstly, the interevent time series of rating behaviors of these three types of media show similar scaling properties with exponents ranging from 0.53 to 0.58, which implies that the collective behaviors of rating media follow a process embodying self-similarity and long-range correlation. Meanwhile, by dividing the users into three groups based on their activities (i.e., rating per unit time), we find that the scaling exponents of the interevent time series in the three groups are different. Hence, these results suggest that a stronger long-range correlations exist in these collective behaviors. Furthermore, their information complexities vary in the three groups. To explain the differences of the collective behaviors restricted to the three groups, we study the dynamic behavior of human activity at the individual level, and find that the dynamic behaviors of a few users have extremely small scaling exponents associated with long-range anticorrelations. By comparing the interevent time distributions of four representative users, we can find that the bimodal distributions may bring forth the extraordinary scaling behaviors. These results of the analysis of the online human activity in the e-commerce may not only provide insight into its dynamic behaviors but may also be applied to acquire potential economic interest.

Physical environment virtualization for human activities recognition

NASA Astrophysics Data System (ADS)

Poshtkar, Azin; Elangovan, Vinayak; Shirkhodaie, Amir; Chan, Alex; Hu, Shuowen

2015-05-01

Human activity recognition research relies heavily on extensive datasets to verify and validate performance of activity recognition algorithms. However, obtaining real datasets are expensive and highly time consuming. A physics-based virtual simulation can accelerate the development of context based human activity recognition algorithms and techniques by generating relevant training and testing videos simulating diverse operational scenarios. In this paper, we discuss in detail the requisite capabilities of a virtual environment to aid as a test bed for evaluating and enhancing activity recognition algorithms. To demonstrate the numerous advantages of virtual environment development, a newly developed virtual environment simulation modeling (VESM) environment is presented here to generate calibrated multisource imagery datasets suitable for development and testing of recognition algorithms for context-based human activities. The VESM environment serves as a versatile test bed to generate a vast amount of realistic data for training and testing of sensor processing algorithms. To demonstrate the effectiveness of VESM environment, we present various simulated scenarios and processed results to infer proper semantic annotations from the high fidelity imagery data for human-vehicle activity recognition under different operational contexts.

Indoleamine 2,3-Dioxygenase Is Dispensable for The Immunomodulatory Function of Stem Cells from Human Exfoliated Deciduous Teeth.

PubMed

Alipour, Razieh; Masoumi Karimi, Masoumeh; Hashemi-Beni, Batool; Adib, Minoo; Sereshki, Nasrin; Sadeghi, Farzaneh

2017-01-01

In this study, we sought to better understand the immunoregulatory function of stem cells derived from human exfoliated deciduous teeth (SHED). We studied the role of the interferon gamma (IFN-γ)-indoleamine 2,3-dioxygenase (IDO)-axis in immunoregulation of SHED compared to bone marrow derived mesenchymal stem cells (BMMSCs) under the same conditions. In this cross-sectional study, recently isolated human T cells were stimulated either by mitogen or inactivated allogeneic peripheral blood mononuclear cells (PBMCs). These T cells were subsequently co-cultured with, either SHED or BMMSCs in the presence or absence of 1-methyl-tryptophan (1-MT) or neutralizing anti- human-IFN-γ antibodies. In all co-cultures we evaluated lymphocyte activation as well as IDO activity. SHED, similar to conventional BMMSCs, had anti-proliferative effects on stimulated T cells and reduced their cytokine production. This property of SHED and BMMSCs was changed by IFN-γ neutralization. We detected IDO in the immunosuppressive supernatant of all co-cultures. Removal of IDO decreased the immunosuppression of BMMSCs. SHED, like BMMSCs, produced the IDO enzyme. Although IFN-γ is one of inducer of IDO production in SHED, these cells were not affected by IFN-γ in the same manner as BMMSCs. Unlike BMMSCs, the IDO enzyme did not contribute to their immunosuppression and might have other cell-type specific roles.

Cockroach protease allergen induces allergic airway inflammation via epithelial cell activation

PubMed Central

Kale, Sagar L.; Agrawal, Komal; Gaur, Shailendra Nath; Arora, Naveen

2017-01-01

Protease allergens are known to enhance allergic inflammation but their exact role in initiation of allergic reactions at mucosal surfaces still remains elusive. This study was aimed at deciphering the role of serine protease activity of Per a 10, a major cockroach allergen in initiation of allergic inflammation at mucosal surfaces. We demonstrate that Per a 10 increases epithelial permeability by disruption of tight junction proteins, ZO-1 and occludin, and enhances the migration of Monocyte derived dendritic cell precursors towards epithelial layer as exhibited by trans-well studies. Per a 10 exposure also leads to secretion of IL-33, TSLP and intracellular Ca2+ dependent increase in ATP levels. Further, in vivo experiments revealed that Per a 10 administration in mice elevated allergic inflammatory parameters along with high levels of IL-33, TSLP, IL-1α and uric acid in the mice lungs. We next demonstrated that Per a 10 cleaves CD23 (low affinity IgE receptor) from the surface of PBMCs and purified B cells and CD25 (IL-2 receptor) from the surface of PBMCs and purified T cells in an activity dependent manner, which might favour Th2 responses. In conclusion, protease activity of Per a 10 plays a significant role in initiation of allergic airway inflammation at the mucosal surfaces. PMID:28198394

Biofidelic Human Activity Modeling and Simulation with Large Variability

DTIC Science & Technology

2014-11-25

A systematic approach was developed for biofidelic human activity modeling and simulation by using body scan data and motion capture data to...replicate a human activity in 3D space. Since technologies for simultaneously capturing human motion and dynamic shapes are not yet ready for practical use, a...that can replicate a human activity in 3D space with the true shape and true motion of a human. Using this approach, a model library was built to

Effect of Spaceflight on the Functions of NK and LAK Cells

NASA Technical Reports Server (NTRS)

Pierson, Duane L.; Grimm, Elizabeth A.; Pierson, Duane L.; Paloski, W. H. (Technical Monitor)

1999-01-01

Spaceflight-associated stress alters some aspects of the human immune response. In this study, we determined the effects of 10 days aboard the Space Shuttle on the cytotoxic activity of NK and LAK cells. The subjects of this study were crewmembers of two 10-day shuttle flights. Ten-ml blood specimens were obtained from ten astronauts 10 days before launch, immediately after landing, and 3 days after landing. PBMCs were separated from the blood specimens and stored at -800 C. All PBMCs were thawed simultaneously, and the cytotoxic activities of NK and LAK cells were measured by a 4-hour Cr-51 release assay. K562 cells were used to assess NK-cell cytotoxicity. After 4 days of IL-2 activation, the LAK cell cytotoxic activity was determined using K562 and Daudi cells as the target cells. NK-cell cytotoxicity was decreased at landing (p less than 0.0005) in 9/10 astronauts, and in most cases recovered to preflight levels by 3 days after landing; NK-cell cytotoxicity was increased in one astronaut at landing. LAK cytotoxic activity against K562 cells was decreased at landing in 6/10 astronauts (p=0.018), and activity against Daudi cells was decreased in 7/10 astronauts (p=0.01). Phenotyping of PBMCs and LAK cells showed alterations in some surface markers and adhesion molecules (CD1 1 b, CD1 1 c, CD1 1 a, CD1 6, L-Selectin and CD3). Thus spaceflight leads to a decrease in the functions of NK and LAK cells in most astronauts.

Immunosuppressive function of mesenchymal stem cells from human umbilical cord matrix in immune thrombocytopenia patients.

PubMed

Ma, Li; Zhou, Zeping; Zhang, Donglei; Yang, Shaoguang; Wang, Jinhong; Xue, Feng; Yang, Yanhui; Yang, Renchi

2012-05-01

Human umbilical cord matrix/Wharton's jelly (hUC)-derived mesenchymal stem cells (MSC) have been shown to have marked therapeutic effects in a number of inflammatory diseases and autoimmune diseases in humans based on their potential for immunosuppression and their low immunogenicity. Currently, no data are available on the effectiveness of UC-MSC transplantation in immune thrombocytopenia (ITP) patients. It was the objective of this study to assess the effect of allogeneic UC-MSCs on ITP patients in vitro and in vivo. Peripheral blood mononuclear cells (PBMCs) and bone marrow mononuclear cells (BM-MNCs) from ITP patients and healthy controls were co-cultured with UC-MSCs for three days and seven days, respectively. Flow cytometry and ELISA were applied to assess the various parameters. In PBMCs from ITP patients, the proliferation of autoreactive T, B lymphocytes and destruction of autologous platelets were dramatically suppressed by UC-MSCs. UC-MSCs not only suppressed co-stimulatory molecules CD80, CD40L and FasL expression but also in shifting Th1/Th2/Treg cytokines profile in ITP patients. UC-MSCs obviously reversed the dysfunctions of megakaryocytes by promoting platelet production and decreasing the number of living megakaryocytes as well as early apoptosis. In addition, the level of thrombopoietin was increased significantly. Our clinical study showed that UC-MSCs play a role in alleviating refractory ITP by increasing platelet numbers. These findings suggested that UC-MSCs transplantation might be a potential therapy for ITP.

Human Umbilical Cord Mesenchymal Stem Cells Inhibit the Function of Allogeneic Activated Vγ9Vδ2 T Lymphocytes In Vitro

PubMed Central

Liu, Xiaohuan; Feng, Ting; Gong, Tianxiang; Shen, Chongyang; Zhu, Tingting; Wu, Qihong; Li, Qiang; Li, Hong

2015-01-01

Background. Human umbilical cord mesenchymal stem cells (UC-MSCs) can regulate the function of immune cells. However, whether and how UC-MSCs can modulate the function of Vγ9Vδ2 T cells has not been fully understood. Methods. The PBMCs or Vγ9Vδ2 T cells were activated and expanded with pamidronate (PAM) and interleukin-2 (IL-2) with or without the presence UC-MSCs. The effects of UC-MSCs on the proliferation, cytokine expression, and cytotoxicity of Vγ9Vδ2 T cells were determined by flow cytometry. The effects of UC-MSCs on Fas-L, TRAIL-expressing Vγ9Vδ2 T cells, and Vγ9Vδ2 T cell apoptosis were determined by flow cytometry. Results. UC-MSCs inhibited Vγ9Vδ2 T cell proliferation in a dose-dependent but cell-contact independent manner. Coculture with UC-MSCs reduced the frequency of IFNγ+ but increased granzyme B+ Vγ9Vδ2 T cells. UC-MSCs inhibited the cytotoxicity of Vγ9Vδ2 T cells against influenza virus H1N1 infected A549 cells and also reduced the frequency of Fas-L+, TRAIL+ Vγ9Vδ2 T cells but failed to modulate the apoptosis of Vγ9Vδ2 T cells. Conclusions. These results indicated that UC-MSCs efficiently suppressed the proliferation and cytotoxicity of Vγ9Vδ2 T cells and modulated their cytokine production. Fas-L and TRAIL were involved in the regulation. Cell contact and apoptosis of Vγ9Vδ2 T cells were not necessary for the inhibition. PMID:25984529

In vitro antiretroviral activity and in vivo toxicity of the potential topical microbicide copper phthalocyanine sulfate.

PubMed

Styczynski, Ashley R; Anwar, Khandaker N; Sultana, Habiba; Ghanem, Abdelhamid; Lurain, Nell; Chua, Aishi; Ghassemi, Mahmood; Novak, Richard M

2015-08-30

Copper has antimicrobial properties and has been studied for its activity against viruses, including HIV. Copper complexed within a phthalocyanine ring, forming copper (II) phthalocyanine sulfate (CuPcS), may have a role in microbicide development when used intravaginally. CuPcS toxicity was tested against cervical epithelial cells, TZM-BL cells, peripheral blood mononuclear cells (PBMC), and cervical explant tissues using cell viability assays. In vivo toxicity was assessed following intravaginal administration of CuPcS in female BALB/C mice and measured using a standardized histology grading system on reproductive tract tissues. Efficacy studies for preventing infection with HIV in the presence of various non-toxic concentrations of CuPcS were carried out in TZM-BL, PBMC, and cervical explant cultures using HIV-1BAL and various pseudovirus subtypes. Non-linear regression was applied to the data to determine the EC50/90 and CC50/90. CuPcS demonstrated inhibition of HIV infection in PBMCs at concentrations that were non-toxic in cervical epithelial cells and PBMCs with EC50 values of approximately 50 μg/mL. Reproductive tract tissue analysis revealed no toxicity at 100 mg/mL. Human cervical explant tissues challenged with HIV in the presence of CuPcS also revealed a dose-response effect at preventing HIV infection at non-toxic concentrations with an EC50 value of 65 μg/mL. These results suggest that CuPcS may be useful as a topical microbicide in concentrations that can be achieved in the female genital tract.

Viability of primary osteoblasts after treatment with tenofovir alafenamide: Lack of cytotoxicity at clinically relevant drug concentrations

PubMed Central

Callebaut, Christian; Liu, Yang; Babusis, Darius; Ray, Adrian; Miller, Michael; Kitrinos, Kathryn

2017-01-01

Tenofovir alafenamide (TAF) is a phosphonoamidate prodrug of the nucleotide HIV reverse transcriptase inhibitor tenofovir (TFV). TAF is approved for the treatment of HIV-1 infection as part of the single-tablet regimen containing elvitegravir, cobicistat, emtricitabine, and TAF. When dosed once-daily, TAF results in approximately 90% lower levels of plasma TFV and a 4-fold increase in intracellular TFV-diphosphate (TFV-DP) in PBMCs compared with the TFV prodrug tenofovir disoproxil fumarate (TDF). Several antiretrovirals, including TDF, have been associated with bone mineral density decreases in patients; the effect of clinically relevant TAF concentrations on primary osteoblast viability was therefore assessed in vitro. Studies in PBMCs determined that a 2-hour TAF exposure at concentrations similar to human plasma Cmax achieved intracellular TFV-DP levels comparable to those observed after the maximum recommended human dose of 25 mg TAF. Comparable intracellular TFV-DP levels were achieved in primary osteoblasts with 2-hour TAF exposure daily for 3 days at concentrations similar to those used for PBMCs (100–400 nM). No change in cell viability was observed in either primary osteoblasts or PBMCs. The mean TAF CC50 in primary osteoblasts after 3 days of daily 2-hour pulses was >500 μM, which is >1033 times higher than the TAF maximum recommended human dose plasma Cmax. In summary, primary osteoblasts were not preferentially loaded by TAF compared with PBMCs, with comparable TFV-DP levels achieved in both cell types. Furthermore, there was no impact on osteoblast cell viability at clinically relevant TAF concentrations. PMID:28182625

Viability of primary osteoblasts after treatment with tenofovir alafenamide: Lack of cytotoxicity at clinically relevant drug concentrations.

PubMed

Callebaut, Christian; Liu, Yang; Babusis, Darius; Ray, Adrian; Miller, Michael; Kitrinos, Kathryn

2017-01-01

Tenofovir alafenamide (TAF) is a phosphonoamidate prodrug of the nucleotide HIV reverse transcriptase inhibitor tenofovir (TFV). TAF is approved for the treatment of HIV-1 infection as part of the single-tablet regimen containing elvitegravir, cobicistat, emtricitabine, and TAF. When dosed once-daily, TAF results in approximately 90% lower levels of plasma TFV and a 4-fold increase in intracellular TFV-diphosphate (TFV-DP) in PBMCs compared with the TFV prodrug tenofovir disoproxil fumarate (TDF). Several antiretrovirals, including TDF, have been associated with bone mineral density decreases in patients; the effect of clinically relevant TAF concentrations on primary osteoblast viability was therefore assessed in vitro. Studies in PBMCs determined that a 2-hour TAF exposure at concentrations similar to human plasma Cmax achieved intracellular TFV-DP levels comparable to those observed after the maximum recommended human dose of 25 mg TAF. Comparable intracellular TFV-DP levels were achieved in primary osteoblasts with 2-hour TAF exposure daily for 3 days at concentrations similar to those used for PBMCs (100-400 nM). No change in cell viability was observed in either primary osteoblasts or PBMCs. The mean TAF CC50 in primary osteoblasts after 3 days of daily 2-hour pulses was >500 μM, which is >1033 times higher than the TAF maximum recommended human dose plasma Cmax. In summary, primary osteoblasts were not preferentially loaded by TAF compared with PBMCs, with comparable TFV-DP levels achieved in both cell types. Furthermore, there was no impact on osteoblast cell viability at clinically relevant TAF concentrations.

The chromatin accessibility signature of human immune aging stems from CD8+ T cells.

PubMed

Ucar, Duygu; Márquez, Eladio J; Chung, Cheng-Han; Marches, Radu; Rossi, Robert J; Uyar, Asli; Wu, Te-Chia; George, Joshy; Stitzel, Michael L; Palucka, A Karolina; Kuchel, George A; Banchereau, Jacques

2017-10-02

Aging is linked to deficiencies in immune responses and increased systemic inflammation. To unravel the regulatory programs behind these changes, we applied systems immunology approaches and profiled chromatin accessibility and the transcriptome in PBMCs and purified monocytes, B cells, and T cells. Analysis of samples from 77 young and elderly donors revealed a novel and robust aging signature in PBMCs, with simultaneous systematic chromatin closing at promoters and enhancers associated with T cell signaling and a potentially stochastic chromatin opening mostly found at quiescent and repressed sites. Combined analyses of chromatin accessibility and the transcriptome uncovered immune molecules activated/inactivated with aging and identified the silencing of the IL7R gene and the IL-7 signaling pathway genes as potential biomarkers. This signature is borne by memory CD8 + T cells, which exhibited an aging-related loss in binding of NF-κB and STAT factors. Thus, our study provides a unique and comprehensive approach to identifying candidate biomarkers and provides mechanistic insights into aging-associated immunodeficiency. © 2017 Ucar et al.

The chromatin accessibility signature of human immune aging stems from CD8+ T cells

PubMed Central

Marches, Radu; Rossi, Robert J.; Uyar, Asli; Wu, Te-Chia; Stitzel, Michael L.; Palucka, A. Karolina

2017-01-01

Aging is linked to deficiencies in immune responses and increased systemic inflammation. To unravel the regulatory programs behind these changes, we applied systems immunology approaches and profiled chromatin accessibility and the transcriptome in PBMCs and purified monocytes, B cells, and T cells. Analysis of samples from 77 young and elderly donors revealed a novel and robust aging signature in PBMCs, with simultaneous systematic chromatin closing at promoters and enhancers associated with T cell signaling and a potentially stochastic chromatin opening mostly found at quiescent and repressed sites. Combined analyses of chromatin accessibility and the transcriptome uncovered immune molecules activated/inactivated with aging and identified the silencing of the IL7R gene and the IL-7 signaling pathway genes as potential biomarkers. This signature is borne by memory CD8+ T cells, which exhibited an aging-related loss in binding of NF-κB and STAT factors. Thus, our study provides a unique and comprehensive approach to identifying candidate biomarkers and provides mechanistic insights into aging-associated immunodeficiency. PMID:28904110

Cytomegalovirus cultured from different major leukocyte subpopulations: association with clinical features in CMV immunoglobulin G-positive renal allograft recipients.

PubMed

Schäfer, P; Tenschert, W; Cremaschi, L; Schröter, M; Gutensohn, K; Laufs, R

2000-08-01

Cytomegalovirus (CMV) cultured from peripheral blood mononuclear cells (PBMCs) was shown to be associated more closely with clinical manifestations than infectious CMV in polymorphonuclear leukocytes (PMNLs) of renal allograft recipients with secondary CMV infection. Shell vial culture was carried out with ficoll-purified PBMCs and PMNLs of 71 CMV IgG-positive patients after kidney transplantation. Thirty-six patients experienced active CMV infections. Of these, 17 developed clinical symptoms. The diagnostic value of PMNLs and PBMCs viremia was determined in comparison to pp65 antigenemia, leukoDNAemia, plasma DNAemia, and detection of cytomegalic endothelial cells. In both PMNLs and PBMCs (with or without detectable endothelial cells), frequencies and levels of viremia were significantly higher among symptomatic patients. Regarding the occurrence of clinical CMV manifestations, the sensitivity of culture from PMNLs and from PBMCs fractions was 100%. Viremia in PBMCs, however, was far more specific (94%) than in PMNLs (74%). Cutoff values established previously for pp65 antigenemia and leukoDNAemia, standard markers in the laboratory, had similar specificity (96% each) to PBMCs viremia, but were less sensitive (88% each). Plasma DNA-emia was both less sensitive (82%) and less specific (69%) than PBMCs viremia. Detection of endothelemia showed maximal specificity (100%), but inferior sensitivity (47%). All patients had PBMCs viremia before the onset of symptoms. In conclusion, infectious CMV present in PBMCs may prove to be a determinant of clinical CMV manifestations in seropositive immunocompromised individuals. Factors involved in PBMCs tropism may help to understand the pathogenetic mechanisms of CMV dissemination in this group of patients. Copyright 2000 Wiley-Liss, Inc.

The development of a modified human IFN-alpha2b linked to the Fc portion of human IgG1 as a novel potential therapeutic for the treatment of hepatitis C virus infection.

PubMed

Jones, Tim D; Hanlon, Marian; Smith, Beverley J; Heise, Charles T; Nayee, Prakash D; Sanders, Deborah A; Hamilton, Anita; Sweet, Clive; Unitt, Esther; Alexander, Graeme; Lo, Kin-Ming; Gillies, Stephen D; Carr, Frank J; Baker, Matthew P

2004-09-01

Interferon-alpha (IFN-alpha), in conjunction with ribavirin, is the current standard for the treatment of chronic hepatitis C virus (HCV) infection. This treatment requires frequent dosing, with a significant risk of the development of anti-IFN-alpha neutralizing antibodies that correlates with lack of efficacy or relapse. We have developed an IFN-alpha linked to the Fc region of human IgG1 for improved half-life and less frequent dosing. We have also identified, using a human T cell proliferation assay, three regions of IFN-alpha2b that are potentially immunogenic, and a variant containing a total of six mutations within these regions was made. This variant was made as a fusion to Fc either with or without a flexible linker between the fusion partners. Both configurations of the variant were less active than native IFN-alpha alone, although the variant containing the flexible linker had in vitro antiviral activity within the range of other modified IFN-alphas currently in clinical use. Peptides spanning the modified regions were tested in T cell proliferation assays and found to be less immunogenic than native controls when using peripheral blood mononuclear cells (PBMCs) from both healthy individuals and HCV-infected patients who had been treated previously with IFN-alpha2b. Copyright Mary Ann Liebert, Inc.

Human activity discrimination for maritime application

NASA Astrophysics Data System (ADS)

Boettcher, Evelyn; Deaver, Dawne M.; Krapels, Keith

2008-04-01

The US Army RDECOM CERDEC Night Vision and Electronic Sensors Directorate (NVESD) is investigating how motion affects the target acquisition model (NVThermIP) sensor performance estimates. This paper looks specifically at estimating sensor performance for the task of discriminating human activities on watercraft, and was sponsored by the Office of Naval Research (ONR). Traditionally, sensor models were calibrated using still images. While that approach is sufficient for static targets, video allows one to use motion cues to aid in discerning the type of human activity more quickly and accurately. This, in turn, will affect estimated sensor performance and these effects are measured in order to calibrate current target acquisition models for this task. The study employed an eleven alternative forced choice (11AFC) human perception experiment to measure the task difficulty of discriminating unique human activities on watercrafts. A mid-wave infrared camera was used to collect video at night. A description of the construction of this experiment is given, including: the data collection, image processing, perception testing and how contrast was defined for video. These results are applicable to evaluate sensor field performance for Anti-Terrorism and Force Protection (AT/FP) tasks for the U.S. Navy.

Mitochondrial Toxicity Studied with the PBMC of Children from the Chinese National Pediatric Highly Active Antiretroviral Therapy Cohort

PubMed Central

Liu, Daojie; Yin, Jiming; Qiao, Luxin; Shi, Ying; Dong, Yaowu; Li, Ning; Zhang, Fujie; Chen, Dexi

2013-01-01

As the backbone of highly active antiretroviral therapy (HAART), nucleoside reverse transcriptase inhibitors (NRTIs) have effectively improved outcomes for HIV-infected patients. However, long-term treatment with NRTIs can cause a series of pathologies associated with mitochondrial toxicity. To date, the status and mechanism of mitochondrial toxicity induced by NRTIs are still not clear, especially in HIV-infected children. As part of the national pediatric HAART program in China, our study focused on mitochondrial toxicity and its potential mechanism in HIV-1-infected children who were divided into two groups based on their duration of treatment with NRTIs: one group received treatment for less than 36 months and one group was treated for 36 to 72 months. The control group comprised age-matched non-HIV-infected children. Blood lactic acid and ATP levels in peripheral blood mononuclear cells (PBMCs) were measured to evaluate mitochondrial function, and mtDNA copies and mutations in PBMCs were determined for detecting mtDNA lesions. Simultaneously, TK2 and P53R2 gene expression in PBMC was measured. As compared with the control group, blood lactic acid levels in both NRTI treatment groups were significantly higher, whereas ATP levels and mtDNA mutation rates in PBMCs did not differ between the control and the two NRTI treatment groups. Both NRTI treatment groups exhibited significant mtDNA loss. N Moreover, we found that P53R2 mRNA expression and protein levels were significantly reduced in both treatment groups and that TK2 mRNA expression and protein levels were induced in the long-term NRTI treatment group. These results suggest that mitochondrial toxicity occurs in long-term HAART patients and that P53R2 and TK2 levels in PBMCs are useful biomarkers for detecting mitochondrial toxicity in patients on long-term treatment with NRTIs. PMID:23468942

LYATK1 potently inhibits LPS-mediated pro-inflammatory response

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

Xi, Feng; Liu, Yuan; Wang, Xiujuan

Lipopolysaccharide (LPS)-primed monocytes/macrophages produce pro-inflammatory cytokines, which could lead to endotoxin shock. TGF-β-activated kinase1 (TAK1) activation is involved in the process. In the current study, we studied the potential effect of a selective TAK1 inhibitor, LYTAK1, on LPS-stimulated response both in vitro and in vivo. We demonstrated that LYTAK1 inhibited LPS-induced mRNA expression and production of several pro-inflammatory cytokines [interleukin 1β (IL-1β), tumor necrosis factor-α (TNFα) and interleukin-6 (IL-6)] in RAW 264.7 macrophages. LYTAK1's activity was almost nullified with TAK1 shRNA-knockdown. Meanwhile, in both primary mouse bone marrow derived macrophages (BMDMs) and human peripheral blood mononuclear cells (PBMCs), LPS-induced pro-inflammatory cytokine productionmore » was again attenuated with LYTAK1 co-treatment. Molecularly, LYTAK1 dramatically inhibited LPS-induced TAK1-nuclear factor kappa B (NFκB) and mitogen-activated protein kinase (Erk, Jnk and p38) activation in RAW 264.7 cells, mouse BMDMs and human PBMCs. In vivo, oral administration of LYTAK1 inhibited LPS-induced activation of TAK1-NFκB-p38 in ex-vivo cultured PBMCs, and cytokine production and endotoxin shock in mice. Together, these results demonstrate that LYTAK1 inhibits LPS-induced production of several pro-inflammatory cytokines and endotoxin shock probably through blocking TAK1-regulated signalings. - Highlights: • LYTAK1 inhibits LPS-induced pro-inflammatory cytokine production in RAW 264.7 cells. • The effect by LYTAK1 is more potent than other known TAK1 inhibitors. • LYTAK1 inhibits LPS-induced cytokine production in primary macrophages/monocytes. • LYTAK1 inhibits LPS-induced TAK1-NFκB and MAPK activation in macrophages/monocytes. • LYTAK1 gavage inhibits LPS-induced endotoxin shock and cytokine production in mice.« less

Structural characterization and immunomodulatory activity of a pectic polysaccharide (CALB-4) from Fructus aurantii.

PubMed

Shu, Zunpeng; Yang, Yanni; Xing, Na; Wang, Yi; Wang, Qiuhong; Kuang, Haixue

2018-02-01

A purified polysaccharide, designated CALB-4, was acquired from Fructus aurantii that is the traditional edible/medicina plant in China. The present study was performed to characterize the CALB-4 and to evaluate its immunomodulatory activities on human peripheral blood mononuclear cells (PBMCs). The structure of CALB-4 was characterized by partial acid hydrolysis, periodate oxidation, Smith degradation, and methylation analysis combined with gas chromatography-mass spectrometry (GC-MS), Infrared Spectroscopy (IR) and scanning electron microscopy (SEM). The results indicated that CALB-4 was elucidated as a pectic polysaccharide and its main chain is composed of Man, Gal UA and Gal, interspersed with Ara, Rha, Man and Gal. Furthermore, immunological tests showed that CALB-4 exhibits the immunoenhancement effects. The mechanism for this action might be attributed to the increase of the cytoplasmic concentration of pro-IL-1 via the up-regulation of several mitogen-activated protein kinases (MAPKs) and the nuclear translocation of p65. This study clarified that CALB-4 could be as an efficacious biological response modifier in immunotherapy. Copyright © 2018. Published by Elsevier B.V.

In vivo expression of human cytomegalovirus (HCMV) microRNAs during latency.

PubMed

Meshesha, Mesfin K; Bentwich, Zvi; Solomon, Semaria A; Avni, Yonat Shemer

2016-01-01

Viral encoded microRNAs play key roles in regulating gene expression and the life cycle of human herpes viruses. Latency is one of the hallmarks of the human cytomegalovirus (HCMV or HHV5) life cycle, and its control may have immense practical applications. The present study aims to identify HCMV encoded microRNAs during the latency phase of the virus. We used a highly sensitive real time PCR (RTPCR) assay that involves a pre-amplification step before RTPCR. It can detect HCMV encoded microRNAs (miRNAs) during latency in purified monocytes and PBMCs from HCMV IgG positive donors and in latently infected monocytic THP-1 cell lines. During the latency phase, only eight HCMV encoded microRNAs were detected in PBMCs, monocytes and in the THP-1 cells. Five originated from the UL region of the virus genome and three from the US region. Reactivation of the virus from latency, in monocytes obtained from the same donor, using dexamethasone restored the expression of all known HCMV encoded miRNAs including those that were absent during latency. We observed a shift in the abundance of the two arms of mir-US29 between the productive and latency stages of the viral life cycle, suggesting that the star "passenger" form of this microRNA is preferentially expressed during latency. As a whole, our study demonstrates that HCMV expresses during the latency phase, both in vivo and in vitro, only a subset of its microRNAs, which may indicate that they play an important role in maintenance and reactivation of latency. Copyright © 2015 Elsevier B.V. All rights reserved.

A Simple Mouse Model for the Study of Human Immunodeficiency Virus.

PubMed

Kim, Kang Chang; Choi, Byeong-Sun; Kim, Kyung-Chang; Park, Ki Hoon; Lee, Hee Jung; Cho, Young Keol; Kim, Sang Il; Kim, Sung Soon; Oh, Yu-Kyoung; Kim, Young Bong

2016-02-01

Humanized mouse models derived from immune-deficient mice have been the primary tool for studies of human infectious viruses, such as human immunodeficiency virus (HIV). However, the current protocol for constructing humanized mice requires elaborate procedures and complicated techniques, limiting the supply of such mice for viral studies. Here, we report a convenient method for constructing a simple HIV-1 mouse model. Without prior irradiation, NOD/SCID/IL2Rγ-null (NSG) mice were intraperitoneally injected with 1 × 10(7) adult human peripheral blood mononuclear cells (hu-PBMCs). Four weeks after PBMC inoculation, human CD45(+) cells, and CD3(+)CD4(+) and CD3(+)CD8(+) T cells were detected in peripheral blood, lymph nodes, spleen, and liver, whereas human CD19(+) cells were observed in lymph nodes and spleen. To examine the usefulness of hu-PBMC-inoculated NSG (hu-PBMC-NSG) mice as an HIV-1 infection model, we intravenously injected these mice with dual-tropic HIV-1DH12 and X4-tropic HIV-1NL4-3 strains. HIV-1-infected hu-PBMC-NSG mice showed significantly lower human CD4(+) T cell counts and high HIV viral loads in the peripheral blood compared with noninfected hu-PBMC-NSG mice. Following highly active antiretroviral therapy (HAART) and neutralizing antibody treatment, HIV-1 replication was significantly suppressed in HIV-1-infected hu-PBMC-NSG mice without detectable viremia or CD4(+) T cell depletion. Moreover, the numbers of human T cells were maintained in hu-PBMC-NSG mice for at least 10 weeks. Taken together, our results suggest that hu-PBMC-NSG mice may serve as a relevant HIV-1 infection and pathogenesis model that could facilitate in vivo studies of HIV-1 infection and candidate HIV-1 protective drugs.

The study on specific umbilical blood Dc vaccine for Beige nude mice loaded human colorectal carcinoma to induce anti-tumor immunity.

PubMed

Fu, Z-X; Han, J-S; Liu, F; Zhao, Z-L; Li, D-B; Shi, L; Dong, J-T; Zhou, Y; Cai, J-H

2017-05-01

This study is to observe the immunosuppression of CD137L transfected umbilical blood Dcs (Dendritic cell) vaccine to tumor development of SCID/ Beige nude mice. Samples of umbilical blood in the childbirth pregnant women were collected by density gradient centrifugation. Umbilical cord blood dendritic cells (Dcs) were transfected by specific CD137L via LipofectamineTM method and cells were harvested. Meanwhile, the peripheral blood of volunteers was collected to isolate Dcs, the Dcs were cultured for 5 days and hatched with SW-1116 cells antigen. The mature Dcs were harvested. The male SCID/Beige nude mice were subcutaneously injected with human SW-1116 cells in axillary to build colorectal carcinoma model as blank control (Blank). The naked peripheral blood Dc vaccine group (cPBMCs), the SW-1116 antigen-specific peripheral blood Dc vaccine group (pDcs) and the CD137L specific umbilical blood Dc vaccine group (tuDcs) were injected 24 h before tumor cells injection, respectively to recur the humanized immune reconstruction. The general life, living habits changes, tumor growing time and tumor size were observed. The nude mice were sacrificed 18 days after tumor formation. The tumor size, mice weight, in vitro tumor weight, liver weight and spleen weight of mice were recorded to evaluate the anti-tumor effect of the specific immune cells. The nude mice in pDcs group showed better general living condition, slower tumor growth, smaller tumor volume and no ulceration, necrosis, and death in nude mice. The tumor formation time in different groups was 4.71 ± 0.18 ds (blank), 7.71 ± 0.29 ds (cPBMCs), 7.86 ± 0.26 ds (pDcs) and 8.14 ± 0.69 ds (tuDcs) respectively. There were significant differences between blank and other three groups (F = 40.96, p < 0.01). Compared to mice in blank group, the tumor volume of cPBMCs group was significantly smaller (201.43 ± 69.84 mm³ vs. 436.04 ± 54.50 mm³, p < 0.01) and the tumor weight were significantly smaller (1.25 ± 0.12 g vs

Integrin activation controls metastasis in human breast cancer

NASA Astrophysics Data System (ADS)

Felding-Habermann, Brunhilde; O'Toole, Timothy E.; Smith, Jeffrey W.; Fransvea, Emilia; Ruggeri, Zaverio M.; Ginsberg, Mark H.; Hughes, Paul E.; Pampori, Nisar; Shattil, Sanford J.; Saven, Alan; Mueller, Barbara M.

2001-02-01

Metastasis is the primary cause of death in human breast cancer. Metastasis to bone, lungs, liver, and brain involves dissemination of breast cancer cells via the bloodstream and requires adhesion within the vasculature. Blood cell adhesion within the vasculature depends on integrins, a family of transmembrane adhesion receptors, and is regulated by integrin activation. Here we show that integrin v3 supports breast cancer cell attachment under blood flow conditions in an activation-dependent manner. Integrin v3 was found in two distinct functional states in human breast cancer cells. The activated, but not the nonactivated, state supported tumor cell arrest during blood flow through interaction with platelets. Importantly, activated αvβ3 was expressed by freshly isolated metastatic human breast cancer cells and variants of the MDA-MB 435 human breast cancer cell line, derived from mammary fat pad tumors or distant metastases in severe combined immunodeficient mice. Expression of constitutively activated mutant αvβ3D723R, but not αvβ3WT, in MDA-MB 435 cells strongly promoted metastasis in the mouse model. Thus breast cancer cells can exhibit a platelet-interactive and metastatic phenotype that is controlled by the activation of integrin αvβ3. Consequently, alterations within tumors that lead to the aberrant control of integrin activation are expected to adversely affect the course of human breast cancer.

Activation of peroxisome proliferator-activated receptor-{alpha} enhances fatty acid oxidation in human adipocytes

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

Lee, Joo-Young; Hashizaki, Hikari; Goto, Tsuyoshi

2011-04-22

Highlights: {yields} PPAR{alpha} activation increased mRNA expression levels of adipocyte differentiation marker genes and GPDH activity in human adipocytes. {yields} PPAR{alpha} activation also increased insulin-dependent glucose uptake in human adipocytes. {yields} PPAR{alpha} activation did not affect lipid accumulation in human adipocytes. {yields} PPAR{alpha} activation increased fatty acid oxidation through induction of fatty acid oxidation-related genes in human adipocytes. -- Abstract: Peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}) is a key regulator for maintaining whole-body energy balance. However, the physiological functions of PPAR{alpha} in adipocytes have been unclarified. We examined the functions of PPAR{alpha} using human multipotent adipose tissue-derived stem cells as a humanmore » adipocyte model. Activation of PPAR{alpha} by GW7647, a potent PPAR{alpha} agonist, increased the mRNA expression levels of adipocyte differentiation marker genes such as PPAR{gamma}, adipocyte-specific fatty acid-binding protein, and lipoprotein lipase and increased both GPDH activity and insulin-dependent glucose uptake level. The findings indicate that PPAR{alpha} activation stimulates adipocyte differentiation. However, lipid accumulation was not changed, which is usually observed when PPAR{gamma} is activated. On the other hand, PPAR{alpha} activation by GW7647 treatment induced the mRNA expression of fatty acid oxidation-related genes such as CPT-1B and AOX in a PPAR{alpha}-dependent manner. Moreover, PPAR{alpha} activation increased the production of CO{sub 2} and acid soluble metabolites, which are products of fatty acid oxidation, and increased oxygen consumption rate in human adipocytes. The data indicate that activation of PPAR{alpha} stimulates both adipocyte differentiation and fatty acid oxidation in human adipocytes, suggesting that PPAR{alpha} agonists could improve insulin resistance without lipid accumulation in adipocytes. The

Fire, Climate, and Human Activity: A Combustive Combination

NASA Astrophysics Data System (ADS)

Kehrwald, N. M.; Battistel, D.; Argiriadis, E.; Barbante, C.; Barber, L. B.; Fortner, S. K.; Jasmann, J.; Kirchgeorg, T.; Zennaro, P.

2017-12-01

Ice and lake core records demonstrate that fires caused by human activity can dominate regional biomass burning records in the Common Era. These major increases in fires are often associated with extensive land use change such as an expansion in agriculture. Regions with few humans, relatively stable human populations and/or unvarying land use often have fire histories that are dominated by climate parameters such as temperature and precipitation. Here, we examine biomass burning recorded in ice cores from northern Greenland (NEEM, (77°27'N; 51°3.6'W), Alaska (Juneau Icefield, 58° 35' N; 134° 29'W) and East Antarctica (EPICA DOME C; 75°06'S; 123°21'E), along with New Zealand lake cores to investigate interactions between climate, fire and human activity. Biomarkers such as levoglucosan, and its isomers mannosan and galactosan, can only be produced by cellulose combustion and therefore are specific indicators of past fire activity archived in ice and lake cores. These fire histories add another factor to climate proxies from the same core, and provide a comparison to regional fire syntheses from charcoal records and climate models. For example, fire data from the JSBACH-Spitfire model for the past 2000 years demonstrates that a climate-only scenario would not increase biomass burning in high northern latitudes for the past 2000 years, while NEEM ice core and regional pollen records demonstrate both increased fire activity and land use change that may be ascribed to human activity. Additional biomarkers such as fecal sterols in lake sediments can determine when people were in an area, and can help establish if an increased human presence in an area corresponds with intensified fire activity. This combination of specific biomarkers, other proxy data, and model output can help determine the relative impact of humans versus climate factors on regional fire activity.

Brain Activation During Singing: "Clef de Sol Activation" Is the "Concert" of the Human Brain.

PubMed

Mavridis, Ioannis N; Pyrgelis, Efstratios-Stylianos

2016-03-01

Humans are the most complex singers in nature, and the human voice is thought by many to be the most beautiful musical instrument. Aside from spoken language, singing represents a second mode of acoustic communication in humans. The purpose of this review article is to explore the functional anatomy of the "singing" brain. Methodologically, the existing literature regarding activation of the human brain during singing was carefully reviewed, with emphasis on the anatomic localization of such activation. Relevant human studies are mainly neuroimaging studies, namely functional magnetic resonance imaging and positron emission tomography studies. Singing necessitates activation of several cortical, subcortical, cerebellar, and brainstem areas, served and coordinated by multiple neural networks. Functionally vital cortical areas of the frontal, parietal, and temporal lobes bilaterally participate in the brain's activation process during singing, confirming the latter's role in human communication. Perisylvian cortical activity of the right hemisphere seems to be the most crucial component of this activation. This also explains why aphasic patients due to left hemispheric lesions are able to sing but not speak the same words. The term clef de sol activation is proposed for this crucial perisylvian cortical activation due to the clef de sol shape of the topographical distribution of these cortical areas around the sylvian fissure. Further research is needed to explore the connectivity and sequence of how the human brain activates to sing.

Synthesis of curcumin-loaded chitosan phosphate nanoparticle and study of its cytotoxicity and antimicrobial activity.

PubMed

Deka, C; Aidew, L; Devi, N; Buragohain, A K; Kakati, D K

2016-11-01

Curcumin has acquired an important position in the treatment of various diseases. But its use, as a chemotherapeutic agent, is limited due to its low water solubility, poor bioavailability, and its sensitive nature at the physiological pH. To overcome this, curcumin was loaded into chitosan phosphate nanoparticles (CPNs). The loading efficiency was found to be 84%. DLS studies revealed the average particle size of CPNs and curcumin-loaded CPNs as 53 and 91 nm, respectively, and TEM results supplemented these values. A sustained release pattern was noticed and the amount of curcumin released in acidic pH was higher than at physiological pH. The curcumin nanoformulation exhibited proficient activity against both Gram-positive and Gram-negative bacteria as well as fungus. Cytocompatibility of the nanoformulations against peripheral blood mononuclear cells (PBMCs) and murine monocyte-macrophage cell line was confirmed by incubating with PBMCs and murine monocyte-macrophage cell line.

Nitric oxide modulates the immunological response of bovine PBMCs in an in vitro BRDc infection model.

PubMed

Sheridan, Michael Peter; Regev-Shoshani, Gilly; Martins, James; Vimalanathan, Selvarani; Miller, Chris

2016-12-01

Bovine respiratory disease complex (BRDc) is a multi-factorial disease, involving both viral and bacterial pathogens, that negatively impacts the cattle feedlot industry. A nitric oxide releasing solution (NORS) has been developed and shown to have potential in the prevention of BRDc. This study investigated the underlying immunological mechanisms through which the nitroslyating agent NORS provides protection against the development of BRDc in susceptible cattle. An in vitro BRDc experimental model was designed using bovine peripheral blood mononuclear cells (PBMCs) which were infected with bovine herpesvirus 1 (BHV-1) and subsequently cultured with lipopolysaccharides (LPS) extracted from Mannheimia haemolytica bacteria. The cells were treated with NORS following viral infection to reflect the timing of administering the NORS treatment in feedlots during initial processing. An expression and protein analysis of key genes involved in the innate immune response was carried out. The BRDc model produced significant increases in gene expression (p<0.01) and protein release (p<0.05) of the proinflammatory cytokines IL-1β and TNF. Treatment with NORS reduced the protein levels of IL-1β (0.39-fold↓) (p<0.05) and TNF (0.48-fold↓) (p<0.01) in the BRDc experimental group when compared against the non-treatment BRDc controls. TLR4 expression, having been significantly reduced under the BRDc experimental conditions (0.33-fold↓) (p<0.05), increased significantly (0.76-fold↑) (p<0.05) following NORS treatment. This study provides evidence suggesting that NO may protect against the development of BRDc by limiting deleterious inflammation while simultaneously increasing TLR4 expression and enhancing the ability of the host to detect and respond to bacterial pathogens. Copyright © 2016 Elsevier Ltd. All rights reserved.

Human Activity Recognition in AAL Environments Using Random Projections.

PubMed

Damaševičius, Robertas; Vasiljevas, Mindaugas; Šalkevičius, Justas; Woźniak, Marcin

2016-01-01

Automatic human activity recognition systems aim to capture the state of the user and its environment by exploiting heterogeneous sensors attached to the subject's body and permit continuous monitoring of numerous physiological signals reflecting the state of human actions. Successful identification of human activities can be immensely useful in healthcare applications for Ambient Assisted Living (AAL), for automatic and intelligent activity monitoring systems developed for elderly and disabled people. In this paper, we propose the method for activity recognition and subject identification based on random projections from high-dimensional feature space to low-dimensional projection space, where the classes are separated using the Jaccard distance between probability density functions of projected data. Two HAR domain tasks are considered: activity identification and subject identification. The experimental results using the proposed method with Human Activity Dataset (HAD) data are presented.

Human Activity Recognition in AAL Environments Using Random Projections

PubMed Central

Damaševičius, Robertas; Vasiljevas, Mindaugas; Šalkevičius, Justas; Woźniak, Marcin

2016-01-01

Automatic human activity recognition systems aim to capture the state of the user and its environment by exploiting heterogeneous sensors attached to the subject's body and permit continuous monitoring of numerous physiological signals reflecting the state of human actions. Successful identification of human activities can be immensely useful in healthcare applications for Ambient Assisted Living (AAL), for automatic and intelligent activity monitoring systems developed for elderly and disabled people. In this paper, we propose the method for activity recognition and subject identification based on random projections from high-dimensional feature space to low-dimensional projection space, where the classes are separated using the Jaccard distance between probability density functions of projected data. Two HAR domain tasks are considered: activity identification and subject identification. The experimental results using the proposed method with Human Activity Dataset (HAD) data are presented. PMID:27413392

Human Dental Pulp Stem Cells Suppress Alloantigen-induced Immunity by Stimulating T Cells to Release Transforming Growth Factor Beta.

PubMed

Kwack, Kyu Hwan; Lee, Jung Min; Park, Sang Hyuk; Lee, Hyeon Woo

2017-01-01

Human dental pulp stem cells (hDPSCs) are ideal candidates for regenerating damaged dental tissue. To examine the possibility that hDPSCs may be used to regenerate pulp, we tested their in vitro effects on acute allogeneic immune responses. A peripheral blood mononuclear cell (PBMC) proliferation assay and immunoglobulin (Ig) production assay were performed to evaluate the immunosuppressive properties of hDPSCs. The mixed lymphocyte reaction was suppressed by incubation with hDPSCs. Transforming growth factor beta (TGF-β) was the major soluble factor responsible for inhibiting the allogeneic proliferation of PBMCs. The production of IgM and IgG by allogeneic activation of responder B lymphocytes was also completely abrogated by TGF-β released from hDPSCs via interferon gamma in response to activation of the responder T lymphocytes. hDPSCs inhibit acute allogeneic immune responses by their release of TGF-β as a result of allogeneic stimulation of T lymphocytes. This study provides an insight into the potential clinical use of hDPSCs for allogeneic transplantation. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Dual Inhibition of Rip2 and IRAK1/4 Regulates IL-1β and IL-6 in Sarcoidosis Alveolar Macrophages and Peripheral Blood Mononuclear Cells

PubMed Central

Talreja, Jaya; Talwar, Harvinder; Ahmad, Nisar; Rastogi, Ruchi; Samavati, Lobelia

2016-01-01

Sarcoidosis is a multisystem granulomatous disease of unknown etiology that primarily affects the lungs. Our previous work indicates that activation of p38 plays a pivotal role in sarcoidosis inflammatory response. Therefore, we investigated the upstream kinase responsible for activation of p38 in sarcoidosis alveolar macrophages (AMs) and peripheral mononuclear cells (PBMCs). We identified that sustained p38 phosphorylation in sarcoidosis AMs and PBMCs is associated with active MKK4 but not with MKK3/6. Additionally, we found that sarcoidosis AMs exhibit a higher expression of IRAK1, IRAK-M and Rip2. Surprisingly, ex vivo treatment of sarcoidosis AMs or PBMCs with IRAK1/4 inhibitor led to a significant increase in IL-1β mRNA expression both spontaneously and in response to TLR2 ligand. However, a combination of Rip2 and IRAK-1/4 inhibitors significantly decreased both IL-1β and IL-6 production in sarcoidosis PBMCs and moderately in AMs. Importantly, a combination of Rip2 and IRAK-1/4 inhibitors led to decreased IFN-γ and IL-6, and decreased percentage of activated CD4+CD25+ cells in PBMCs. These data suggest that in sarcoidosis both pathways, namely IRAK and Rip2 are deregulated. Targeted modulation of Rip2 and IRAK pathways may prove to be a novel treatment for sarcoidosis. PMID:27402699

Degalactosylated/desialylated human serum containing GcMAF induces macrophage phagocytic activity and in vivo antitumor activity.

PubMed

Kuchiike, Daisuke; Uto, Yoshihiro; Mukai, Hirotaka; Ishiyama, Noriko; Abe, Chiaki; Tanaka, Daichi; Kawai, Tomohito; Kubo, Kentaro; Mette, Martin; Inui, Toshio; Endo, Yoshio; Hori, Hitoshi

2013-07-01

The group-specific component protein-derived macrophage-activating factor (GcMAF) has various biological activities, such as macrophage activation and antitumor activity. Clinical trials of GcMAF have been carried out for metastatic breast cancer, prostate cancer, and metastatic colorectal cancer. In this study, despite the complicated purification process of GcMAF, we used enzymatically-treated human serum containing GcMAF with a considerable macrophage-stimulating activity and antitumor activity. We detected GcMAF in degalactosylated/desialylated human serum by western blotting using an anti-human Gc globulin antibody, and Helix pomatia agglutinin lectin. We also found that GcMAF-containing human serum significantly enhanced the phagocytic activity of mouse peritoneal macrophages and extended the survival time of mice bearing Ehrlich ascites tumors. We demonstrated that GcMAF-containing human serum can be used as a potential macrophage activator for cancer immunotherapy.

Role of activity in human dynamics

NASA Astrophysics Data System (ADS)

Zhou, T.; Kiet, H. A. T.; Kim, B. J.; Wang, B.-H.; Holme, P.

2008-04-01

The human society is a very complex system; still, there are several non-trivial, general features. One type of them is the presence of power-law-distributed quantities in temporal statistics. In this letter, we focus on the origin of power laws in rating of movies. We present a systematic empirical exploration of the time between two consecutive ratings of movies (the interevent time). At an aggregate level, we find a monotonous relation between the activity of individuals and the power law exponent of the interevent time distribution. At an individual level, we observe a heavy-tailed distribution for each user, as well as a negative correlation between the activity and the width of the distribution. We support these findings by a similar data set from mobile phone text-message communication. Our results demonstrate a significant role of the activity of individuals on the society-level patterns of human behavior. We believe this is a common character in the interest-driven human dynamics, corresponding to (but different from) the universality classes of task-driven dynamics.

Distributed Neural Activity Patterns during Human-to-Human Competition

PubMed Central

Piva, Matthew; Zhang, Xian; Noah, J. Adam; Chang, Steve W. C.; Hirsch, Joy

2017-01-01

Interpersonal interaction is the essence of human social behavior. However, conventional neuroimaging techniques have tended to focus on social cognition in single individuals rather than on dyads or groups. As a result, relatively little is understood about the neural events that underlie face-to-face interaction. We resolved some of the technical obstacles inherent in studying interaction using a novel imaging modality and aimed to identify neural mechanisms engaged both within and across brains in an ecologically valid instance of interpersonal competition. Functional near-infrared spectroscopy was utilized to simultaneously measure hemodynamic signals representing neural activity in pairs of subjects playing poker against each other (human–human condition) or against computer opponents (human–computer condition). Previous fMRI findings concerning single subjects confirm that neural areas recruited during social cognition paradigms are individually sensitive to human–human and human–computer conditions. However, it is not known whether face-to-face interactions between opponents can extend these findings. We hypothesize distributed effects due to live processing and specific variations in across-brain coherence not observable in single-subject paradigms. Angular gyrus (AG), a component of the temporal-parietal junction (TPJ) previously found to be sensitive to socially relevant cues, was selected as a seed to measure within-brain functional connectivity. Increased connectivity was confirmed between AG and bilateral dorsolateral prefrontal cortex (dlPFC) as well as a complex including the left subcentral area (SCA) and somatosensory cortex (SS) during interaction with a human opponent. These distributed findings were supported by contrast measures that indicated increased activity at the left dlPFC and frontopolar area that partially overlapped with the region showing increased functional connectivity with AG. Across-brain analyses of neural coherence

Interleukin-7 (IL-7) enhances class switching to IgE and IgG4 in the presence of T cells via IL-9 and sCD23.

PubMed

Jeannin, P; Delneste, Y; Lecoanet-Henchoz, S; Gretener, D; Bonnefoy, J Y

1998-02-15

Interleukin-7 (IL-7) is a B-cell growth factor produced by both bone marrow stroma cells and follicular dendritic cells (FDCs) located in primary lymphoid follicles and germinal centers. In this study, we have evaluated the role of IL-7 on human Ig class switching. IL-7 was added to peripheral blood mononuclear cells (PBMCs) or tonsillar B cells in the absence or presence of IL-4 and/or anti-CD40 monoclonal antibody (MoAb). Alone, IL-7 did not affect Ig production by PBMCs or by anti-CD40 MoAb-stimulated B cells. Rather, IL-7 potentiated IL-4-induced IgE and IgG4 production by PBMCs. In parallel, IgG3 production was also enhanced but to a lesser extent, whereas the production of the other isotypes was unaltered. The activity of IL-2, IL-9, or IL-15, which share usage of the common gamma chain for signaling, was also assessed. IL-9, like IL-7, potentiated mainly IgE and IgG4 production by IL-4-stimulated PBMCs. IL-15, in contrast, was ineffective, whereas IL-2 enhanced the production of all isotypes. More precisely, IL-7 potentiation of IgE and IgG4 production required the presence of T cells and was accompanied by an increase of the expression of two soluble molecules favoring preferentially IgE and IgG4 synthesis: CD23 (sCD23) and IL-9. Moreover, neutralizing anti-CD23 and anti-IL-9 antibodies partly inhibited the increase of IgE synthesis induced by IL-7. Thus, IL-7 produced locally in the germinal centers by FDCs may interact with T cells and potentiate human IgE and IgG4 switching by favoring IL-9 and sCD23 production.

River Networks and Human Activities: Global Fractal Analysis Using Nightlight Data

NASA Astrophysics Data System (ADS)

McCurley, K. 4553; Fang, Y.; Ceola, S.; Paik, K.; McGrath, G. S.; Montanari, A.; Rao, P. S.; Jawitz, J. W.

2016-12-01

River networks hold an important historical role in affecting human population distribution. In this study, we link the geomorphological structure of river networks to the pattern of human activities at a global scale. We use nightlights as a valuable proxy for the presence of human settlements and economic activity, and we employ HydroSHEDS as the main data source on river networks. We test the hypotheses that, analogous to Horton's laws, human activities (magnitude of nightlights) also show scaling relationship with stream order, and that the intensity of human activities decrease as the distance from the basin outlet increase. Our results demonstrate that the distribution of human activities shows a fractal structure, with power-law scaling between human activities and stream order. This relationship is robust among global river basins. Human activities are more concentrated in larger order basins, but show large variation in equivalent order basins, with higher population density emergent in the basins connected with high-order rivers. For all global river basins longer than 400km, the average intensity of human activities decrease as the distance to the outlets increases, albeit with signatures of large cities at varied distances. The power spectrum of human width (area) function is found to exhibit power law scaling, with a scaling exponent that indicates enrichment of low frequency variation. The universal fractal structure of human activities may reflect an optimum arrangement for humans in river basins to better utilize the water resources, ecological assets, and geographic advantages. The generalized patterns of human activities could be applied to better understand hydrologic and biogeochemical responses in river basins, and to advance catchment management.

Activation of human T cells in hypertension: Studies of Humanized Mice and Hypertensive Humans

PubMed Central

Itani, Hana A.; McMaster, William G.; Saleh, Mohamed A.; Nazarewicz, Rafal R.; Mikolajczyk, Tomasz P.; Kaszuba, Anna; Konior, Anna; Prejbisz, Aleksander; Januszewicz, Andrzej; Norlander, Allison E.; Chen, Wei; Bonami, Rachel H.; Marshall, Andrew F.; Poffenberger, Greg; Weyand, Cornelia M.; Madhur, Meena S.; Moore, Daniel J.; Harrison, David G.; Guzik, Tomasz J.

2016-01-01

Emerging evidence supports an important role for T cells in the genesis of hypertension. Because this work has predominantly been performed in experimental animals, we sought to determine whether human T cells are activated in hypertension. We employed a humanized mouse model in which the murine immune system is replaced by the human immune system. Angiotensin II increased systolic pressure to 162 mm Hg vs. 116 mm Hg for sham treated animals. Flow cytometry of thoracic lymph nodes, thoracic aorta and kidney revealed increased infiltration of human leukocytes (CD45+) and T lymphocytes (CD3+ and CD4+) in response to angiotensin II infusion. Interestingly, there was also an increase in the memory T cells (CD3+/CD45RO+) in the aortas and lymph nodes. Prevention of hypertension using hydralazine and hydrochlorothiazide prevented the accumulation of T cells in these tissues. Studies of isolated human T cells and monocytes indicated that angiotensin II had no direct effect on cytokine production by T cells or the ability of dendritic cells to drive T cell proliferation. We also observed an increase in circulating IL-17A producing CD4+ T cells and both CD4+ and CD8+ T cells that produce IFN-γ in hypertensive compared to normotensive humans. Thus, human T cells become activated and invade critical end-organ tissues in response to hypertension in a humanized mouse model. This response likely reflects the hypertensive milieu encountered in vivo and is not a direct effect of the hormone angiotensin II. PMID:27217403

Activation of Human T Cells in Hypertension: Studies of Humanized Mice and Hypertensive Humans.

PubMed

Itani, Hana A; McMaster, William G; Saleh, Mohamed A; Nazarewicz, Rafal R; Mikolajczyk, Tomasz P; Kaszuba, Anna M; Konior, Anna; Prejbisz, Aleksander; Januszewicz, Andrzej; Norlander, Allison E; Chen, Wei; Bonami, Rachel H; Marshall, Andrew F; Poffenberger, Greg; Weyand, Cornelia M; Madhur, Meena S; Moore, Daniel J; Harrison, David G; Guzik, Tomasz J

2016-07-01

Emerging evidence supports an important role for T cells in the genesis of hypertension. Because this work has predominantly been performed in experimental animals, we sought to determine whether human T cells are activated in hypertension. We used a humanized mouse model in which the murine immune system is replaced by the human immune system. Angiotensin II increased systolic pressure to 162 versus 116 mm Hg for sham-treated animals. Flow cytometry of thoracic lymph nodes, thoracic aorta, and kidney revealed increased infiltration of human leukocytes (CD45(+)) and T lymphocytes (CD3(+) and CD4(+)) in response to angiotensin II infusion. Interestingly, there was also an increase in the memory T cells (CD3(+)/CD45RO(+)) in the aortas and lymph nodes. Prevention of hypertension using hydralazine and hydrochlorothiazide prevented the accumulation of T cells in these tissues. Studies of isolated human T cells and monocytes indicated that angiotensin II had no direct effect on cytokine production by T cells or the ability of dendritic cells to drive T-cell proliferation. We also observed an increase in circulating interleukin-17A producing CD4(+) T cells and both CD4(+) and CD8(+) T cells that produce interferon-γ in hypertensive compared with normotensive humans. Thus, human T cells become activated and invade critical end-organ tissues in response to hypertension in a humanized mouse model. This response likely reflects the hypertensive milieu encountered in vivo and is not a direct effect of the hormone angiotensin II. © 2016 American Heart Association, Inc.

Human monocyte-derived suppressor cells control graft-versus-host disease by inducing regulatory forkhead box protein 3-positive CD8+ T lymphocytes.

PubMed

Janikashvili, Nona; Trad, Malika; Gautheron, Alexandrine; Samson, Maxime; Lamarthée, Baptiste; Bonnefoy, Francis; Lemaire-Ewing, Stéphanie; Ciudad, Marion; Rekhviashvili, Khatuna; Seaphanh, Famky; Gaugler, Béatrice; Perruche, Sylvain; Bateman, Andrew; Martin, Laurent; Audia, Sylvain; Saas, Philippe; Larmonier, Nicolas; Bonnotte, Bernard

2015-06-01

Adoptive transfer of immunosuppressive cells has emerged as a promising strategy for the treatment of immune-mediated disorders. However, only a limited number of such cells can be isolated from in vivo specimens. Therefore efficient ex vivo differentiation and expansion procedures are critically needed to produce a clinically relevant amount of these suppressive cells. We sought to develop a novel, clinically relevant, and feasible approach to generate ex vivo a subpopulation of human suppressor cells of monocytic origin, referred to as human monocyte-derived suppressive cells (HuMoSCs), which can be used as an efficient therapeutic tool to treat inflammatory disorders. HuMoSCs were generated from human monocytes cultured for 7 days with GM-CSF and IL-6. The immune-regulatory properties of HuMoSCs were investigated in vitro and in vivo. The therapeutic efficacy of HuMoSCs was evaluated by using a graft-versus-host disease (GvHD) model of humanized mice (NOD/SCID/IL-2Rγc(-/-) [NSG] mice). CD33+ HuMoSCs are highly potent at inhibiting the proliferation and activation of autologous and allogeneic effector T lymphocytes in vitro and in vivo. The suppressive activity of these cells depends on signal transducer and activator of transcription 3 activation. Of therapeutic relevance, HuMoSCs induce long-lasting memory forkhead box protein 3-positive CD8+ regulatory T lymphocytes and significantly reduce GvHD induced with human PBMCs in NSG mice. Ex vivo-generated HuMoSCs inhibit effector T lymphocytes, promote the expansion of immunosuppressive forkhead box protein 3-positive CD8+ regulatory T cells, and can be used as an efficient therapeutic tool to prevent GvHD. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Citrus flavone tangeretin inhibits leukaemic HL-60 cell growth partially through induction of apoptosis with less cytotoxicity on normal lymphocytes.

PubMed

Hirano, T; Abe, K; Gotoh, M; Oka, K

1995-12-01

Certain anti-cancer agents are known to induce apoptosis in human tumour cells. However, these agents are intrinsically cytotoxic against cells of normal tissue origin, including myelocytes and immunocytes. Here we show that a naturally occurring flavone of citrus origin, tangeretin (5,6,7,8,4'-pentamethoxyflavone), induces apoptosis in human promyelocytic leukaemia HL-60 cells, whereas the flavone showed no cytotoxicity against human peripheral blood mononuclear cells (PBMCs). The growth of HL-60 cells in vitro assessed by [3H]thymidine incorporation or tetrazolium crystal formation was strongly suppressed in the presence of tangeretin; the IC50 values range between 0.062 and 0.173 microM. Apoptosis of HL-60 cells, assessed by cell morphology and DNA fragmentation, was demonstrated in the presence of > 2.7 microM tangeretin. Flow cytometric analysis of tangeretin-treated HL-60 cells also demonstrated apoptotic cells with low DNA content and showed a decrease of G1 cells and a concomitant increase of S and/or G2/M cells. Apoptosis was evident after 24 h of incubation with tangeretin, and the tangeretin effect as assessed by DNA fragmentation or growth inhibition was significantly attenuated in the presence of Zn2+, which is known to inhibit Ca(2+)-dependent endonuclease activity. Ca2+ and Mg2+, in contrast, promoted the effect of tangeretin. Cycloheximide significantly decreased the tangeretin effect on HL-60 cell growth, suggesting that protein synthesis is required for flavonoid-induced apoptosis. Tangeretin showed no cytotoxicity against either HL-60 cells or mitogen-activated PBMCs even at high concentration (27 microM) as determined by a dye exclusion test. Moreover, the flavonoid was less effective on growth of human T-lymphocytic leukaemia MOLT-4 cells or on blastogenesis of PBMCs. These results suggest that tangeretin inhibits growth of HL-60 cells in vitro, partially through induction of apoptosis, without causing serious side-effects on immune cells.

48 CFR 3452.224-72 - Research activities involving human subjects.

Code of Federal Regulations, 2012 CFR

2012-10-01

... involving human subjects. 3452.224-72 Section 3452.224-72 Federal Acquisition Regulations System DEPARTMENT... Text of Provisions and Clauses 3452.224-72 Research activities involving human subjects. As prescribed... human subjects covered under 34 CFR part 97: Research Activities Involving Human Subjects (MAR 2011) (a...

48 CFR 3452.224-72 - Research activities involving human subjects.

Code of Federal Regulations, 2011 CFR

2011-10-01

... involving human subjects. 3452.224-72 Section 3452.224-72 Federal Acquisition Regulations System DEPARTMENT... Text of Provisions and Clauses 3452.224-72 Research activities involving human subjects. As prescribed... human subjects covered under 34 CFR part 97: Research Activities Involving Human Subjects (MAR 2011) (a...

48 CFR 3452.224-72 - Research activities involving human subjects.

Code of Federal Regulations, 2014 CFR

2014-10-01

... involving human subjects. 3452.224-72 Section 3452.224-72 Federal Acquisition Regulations System DEPARTMENT... Text of Provisions and Clauses 3452.224-72 Research activities involving human subjects. As prescribed... human subjects covered under 34 CFR part 97: Research Activities Involving Human Subjects (MAR 2011) (a...

48 CFR 3452.224-72 - Research activities involving human subjects.

Code of Federal Regulations, 2013 CFR

2013-10-01

... involving human subjects. 3452.224-72 Section 3452.224-72 Federal Acquisition Regulations System DEPARTMENT... Text of Provisions and Clauses 3452.224-72 Research activities involving human subjects. As prescribed... human subjects covered under 34 CFR part 97: Research Activities Involving Human Subjects (MAR 2011) (a...

Multilevel depth and image fusion for human activity detection.

PubMed

Ni, Bingbing; Pei, Yong; Moulin, Pierre; Yan, Shuicheng

2013-10-01

Recognizing complex human activities usually requires the detection and modeling of individual visual features and the interactions between them. Current methods only rely on the visual features extracted from 2-D images, and therefore often lead to unreliable salient visual feature detection and inaccurate modeling of the interaction context between individual features. In this paper, we show that these problems can be addressed by combining data from a conventional camera and a depth sensor (e.g., Microsoft Kinect). We propose a novel complex activity recognition and localization framework that effectively fuses information from both grayscale and depth image channels at multiple levels of the video processing pipeline. In the individual visual feature detection level, depth-based filters are applied to the detected human/object rectangles to remove false detections. In the next level of interaction modeling, 3-D spatial and temporal contexts among human subjects or objects are extracted by integrating information from both grayscale and depth images. Depth information is also utilized to distinguish different types of indoor scenes. Finally, a latent structural model is developed to integrate the information from multiple levels of video processing for an activity detection. Extensive experiments on two activity recognition benchmarks (one with depth information) and a challenging grayscale + depth human activity database that contains complex interactions between human-human, human-object, and human-surroundings demonstrate the effectiveness of the proposed multilevel grayscale + depth fusion scheme. Higher recognition and localization accuracies are obtained relative to the previous methods.

Human activities change marine ecosystems by altering predation risk.

PubMed

Madin, Elizabeth M P; Dill, Lawrence M; Ridlon, April D; Heithaus, Michael R; Warner, Robert R

2016-01-01

In ocean ecosystems, many of the changes in predation risk - both increases and decreases - are human-induced. These changes are occurring at scales ranging from global to local and across variable temporal scales. Indirect, risk-based effects of human activity are known to be important in structuring some terrestrial ecosystems, but these impacts have largely been neglected in oceans. Here, we synthesize existing literature and data to explore multiple lines of evidence that collectively suggest diverse human activities are changing marine ecosystems, including carbon storage capacity, in myriad ways by altering predation risk. We provide novel, compelling evidence that at least one key human activity, overfishing, can lead to distinct, cascading risk effects in natural ecosystems whose magnitude exceeds that of presumed lethal effects and may account for previously unexplained findings. We further discuss the conservation implications of human-caused indirect risk effects. Finally, we provide a predictive framework for when human alterations of risk in oceans should lead to cascading effects and outline a prospectus for future research. Given the speed and extent with which human activities are altering marine risk landscapes, it is crucial that conservation and management policy considers the indirect effects of these activities in order to increase the likelihood of success and avoid unfortunate surprises. © 2015 John Wiley & Sons Ltd.

Correlation analyses revealed global microRNA-mRNA expression associations in human peripheral blood mononuclear cells.

PubMed

Wang, Lan; Zhu, Jiang; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Xiao-Wei; Xia, Wei; Xie, Fang-Fei; He, Pei; Bing, Peng-Fei; Qiu, Ying-Hua; Lin, Xiang; Lu, Xin; Zhang, Lei; Yi, Neng-Jun; Zhang, Yong-Hong; Lei, Shu-Feng

2018-02-01

MicroRNAs (miRNAs) can regulate gene expression through binding to complementary sites in the 3'-untranslated regions of target mRNAs, which will lead to existence of correlation in expression between miRNA and mRNA. However, the miRNA-mRNA correlation patterns are complex and remain largely unclear yet. To establish the global correlation patterns in human peripheral blood mononuclear cells (PBMCs), multiple miRNA-mRNA correlation analyses and expression quantitative trait locus (eQTL) analysis were conducted in this study. We predicted and achieved 861 miRNA-mRNA pairs (65 miRNAs, 412 mRNAs) using multiple bioinformatics programs, and found global negative miRNA-mRNA correlations in PBMC from all 46 study subjects. Among the 861 pairs of correlations, 19.5% were significant (P < 0.05) and ~70% were negative. The correlation network was complex and highlighted key miRNAs/genes in PBMC. Some miRNAs, such as hsa-miR-29a, hsa-miR-148a, regulate a cluster of target genes. Some genes, e.g., TNRC6A, are regulated by multiple miRNAs. The identified genes tend to be enriched in molecular functions of DNA and RNA binding, and biological processes such as protein transport, regulation of translation and chromatin modification. The results provided a global view of the miRNA-mRNA expression correlation profile in human PBMCs, which would facilitate in-depth investigation of biological functions of key miRNAs/mRNAs and better understanding of the pathogenesis underlying PBMC-related diseases.

Recognition of human activity characteristics based on state transitions modeling technique

NASA Astrophysics Data System (ADS)

Elangovan, Vinayak; Shirkhodaie, Amir

2012-06-01

Human Activity Discovery & Recognition (HADR) is a complex, diverse and challenging task but yet an active area of ongoing research in the Department of Defense. By detecting, tracking, and characterizing cohesive Human interactional activity patterns, potential threats can be identified which can significantly improve situation awareness, particularly, in Persistent Surveillance Systems (PSS). Understanding the nature of such dynamic activities, inevitably involves interpretation of a collection of spatiotemporally correlated activities with respect to a known context. In this paper, we present a State Transition model for recognizing the characteristics of human activities with a link to a prior contextbased ontology. Modeling the state transitions between successive evidential events determines the activities' temperament. The proposed state transition model poses six categories of state transitions including: Human state transitions of Object handling, Visibility, Entity-entity relation, Human Postures, Human Kinematics and Distance to Target. The proposed state transition model generates semantic annotations describing the human interactional activities via a technique called Casual Event State Inference (CESI). The proposed approach uses a low cost kinect depth camera for indoor and normal optical camera for outdoor monitoring activities. Experimental results are presented here to demonstrate the effectiveness and efficiency of the proposed technique.

Khat (Catha edulis) alters the phenotype and anti-microbial activity of peripheral blood mononuclear cells.

PubMed

Murdoch, Craig; Aziz, Hesham Abdul; Fang, Hsin-Yu; Jezan, Hussun; Musaid, Raga; Muthana, Munitta

2011-12-08

The habit of khat chewing has been associated with increased risk of systemic and oral disease. Although research has been conducted on the affects of khat on oral epithelial cells, little is known about its influence on immune cells. This study examined the biological effects of khat on the phenotype and function of peripheral blood mononuclear cells (PBMCs). Khat-stimulated PBMCs were examined for signs of cytotoxicity, apoptosis and changes in cell surface receptor and cytokine expression. Khat-induced regulation of transcription factors and stress-related factors were examined, as was PBMC phagocytic activity against oral bacteria. Khat was cytotoxic to PBMC in a dose- and time-dependent manner and cell death was mediated by apoptosis. Khat-treated PBMC showed increased expression of co-stimulatory molecules (CD80, CD86 and MHC II) and pattern recognition receptors (TLR-2, TLR-4 and TREM-1) but secretion of inflammatory cytokines (TNFα, IL-6, CCL5, CXCL8) was inhibited. In contrast, khat induced an increase in the anti-inflammatory cytokine IL-10 as well as IL-2, IFN-γ, FasL and HSP70. These khat-induced alterations were accompanied by increased expression of transcription factors p38 MAPK and HIF-1α, whilst expression of NFκB p65 was inhibited. Although the ability of PBMC to phagocytose dextran and oral bacteria was inhibited, production of reactive oxygen species was increased. These data suggest that khat may severely influence the effectiveness of immune surveillance and anti-microbial capacity of PBMCs. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Understanding human activity patterns based on space-time-semantics

NASA Astrophysics Data System (ADS)

Huang, Wei; Li, Songnian

2016-11-01

Understanding human activity patterns plays a key role in various applications in an urban environment, such as transportation planning and traffic forecasting, urban planning, public health and safety, and emergency response. Most existing studies in modeling human activity patterns mainly focus on spatiotemporal dimensions, which lacks consideration of underlying semantic context. In fact, what people do and discuss at some places, inferring what is happening at the places, cannot be simple neglected because it is the root of human mobility patterns. We believe that the geo-tagged semantic context, representing what individuals do and discuss at a place and a specific time, drives a formation of specific human activity pattern. In this paper, we aim to model human activity patterns not only based on space and time but also with consideration of associated semantics, and attempt to prove a hypothesis that similar mobility patterns may have different motivations. We develop a spatiotemporal-semantic model to quantitatively express human activity patterns based on topic models, leading to an analysis of space, time and semantics. A case study is conducted using Twitter data in Toronto based on our model. Through computing the similarities between users in terms of spatiotemporal pattern, semantic pattern and spatiotemporal-semantic pattern, we find that only a small number of users (2.72%) have very similar activity patterns, while the majority (87.14%) show different activity patterns (i.e., similar spatiotemporal patterns and different semantic patterns, similar semantic patterns and different spatiotemporal patterns, or different in both). The population of users that has very similar activity patterns is decreased by 56.41% after incorporating semantic information in the corresponding spatiotemporal patterns, which can quantitatively prove the hypothesis.

Human Interferon Regulatory Factor 2 Gene Expression is Induced in Chronic Hepatitis C Virus Infection—A Possible Mode of Viral Persistence

PubMed Central

Mukherjee, Rathindra M; Bansode, Budhapriyavilas; Gangwal, Puja; Jakkampudi, Aparna; Reddy, Panyala B; Rao, Padaki N; Gupta, Rajesh; Reddy, D Nageshwar

2012-01-01

Background The interferon regulatory factors (IRFs) are a family of transcription factors known to be involved in the modulation of cellular responses to interferons (IFNs) and viral infection. While IRF-1 acts as a positive regulator, IRF-2 is known to repress IFN-mediated gene expression. The increase in the IRF-1/IRF-2 ratio is considered as an important event in the transcriptional activation of IFN-α gene toward development of the cellular antiviral response. Objective This study was performed to assess the expression of IRF mRNAs along with the expression level of IFN-α, its receptor (IFNAR-1), and the signal transduction factor (STAT-1) in treatment naive hepatitis C virus (HCV)-infected subjects. Materials Thirty-five chronically infected (CHC) patients and 39 voluntary blood donors as controls were included in the study. Quantification of HCV-RNA (ribonucleic acid) and genotyping were done by real-time polymerase chain reaction (PCR) and hybridization assays, respectively, using patient's serum/plasma. In both controls and patients, the serum level of IFN-α and IFN-α was measured by flow cytometry. Target gene expressions were studied by retro-transcription of respective mRNAs extracted from peripheral blood mononuclear cells (PBMCs) followed by PCR amplification and densitometry. Minus-strand HCV-RNA as a marker of viral replication in PBMCs was detected by an inhouse PCR assay. Results Both IRF-1 and IRF-2 genes were significantly enhanced in CHC than in control subjects (P < 0.001). A significant positive correlation (r2 = 0.386, P <0.01) was obtained between higher IRF-2 gene expression and increasing level of HCV-RNA. Chronically infected subjects (13%) harboring replicating HCV in PBMCs showed no significant differences in gene expressions than the subjects without HCV in PBMCs. Conclusion Our findings indicate that HCV modulates host immunity by inducing IRF-2 gene to counteract IRF-1-mediated IFN-α gene expression. Since the IRF-2 gene is

A different representation of natural T cells and natural killer cells between tumor-infiltrating and periphery lymphocytes in human hepatocellular carcinoma.

PubMed

Li, Xiao-Feng; Dai, Dong; Song, Xiu-Yu; Liu, Jian-Jing; Zhu, Lei; Zhu, Xiang; Ma, Wenchao; Xu, Wengui

2017-05-01

Natural T cells [cluster of differentiation (CD) 3 + CD56 + ] and natural killer (NK) cells (CD3 - CD56 + ) are particularly abundant in the human liver and serve an important role in immune responses in the liver. The aim of the present study was to extensively determine the phenotypic and functional characteristics of natural T and NK cells in human hepatocellular carcinoma (HCC). Tumorous and non-tumorous tissue infiltrating lymphocytes (TILs and NILs, respectively) and peripheral blood mononuclear cells (PBMCs) from patients with hepatocellular carcinoma (HCC) were obtained to determine the frequency and phenotype of natural T/NK cells by a multicolor fluorescence activated cell sorting analysis. The abundance of natural T cells and NK cells was decreased in TILs vs. NILs (natural T cells, 6.315±1.002 vs. 17.16±1.804; NK cells, 6.324±1.559 vs. 14.52±2.336, respectively). However such results were not observed in PBMCs from HCC patients vs. that of healthy donors. Notably, a substantial fraction of the natural T cells (21.96±5.283) in TILs acquired forkhead box P3 (FOXP3) expression, and the FOXP3 + natural T cells lost the expression of interferon-γ and perforin. Conversely, being similar to the conventional FOXP3 + regulatory T cells, the FOXP3 + natural T cells assumed a specific phenotype that was characteristic of CD25 + , CD45RO + and cytotoxic T-lymphocyte-associated protein 4 + . Consistent with the phenotypic conversion, the present functional results indicate that FOXP3 expression in natural T cells contributes to the acquisition of a potent immunosuppressive capability. In conclusion, the present study describes a different representation of natural T cells and NK cells in local tumor tissues and in the periphery blood of patients with HCC, and identified a new type of FOXP3-expressing natural T cell spontaneously arising in the TILs of HCC.

Quantitative Comparison of Active and Latent Tuberculosis in the Cynomolgus Macaque Model▿ †

PubMed Central

Lin, Philana Ling; Rodgers, Mark; Smith, Le'kneitah; Bigbee, Matthew; Myers, Amy; Bigbee, Carolyn; Chiosea, Ion; Capuano, Saverio V.; Fuhrman, Carl; Klein, Edwin; Flynn, JoAnne L.

2009-01-01

We previously described that low-dose Mycobacterium tuberculosis infection in cynomolgus macaques results in a spectrum of disease similar to that of human infection: primary disease, latent infection, and reactivation tuberculosis (S. V. Capuano III, D. A. Croix, S. Pawar, A. Zinovik, A. Myers, P. L. Lin, S. Bissel, C. Fuhrman, E. Klein, and J. L. Flynn, Infect. Immun. 71:5831-5844, 2003). This is the only established model of latent infection, and it provides a unique opportunity to understand host and pathogen differences across of range of disease states. Here, we provide a more extensive and detailed characterization of the gross pathology, microscopic histopathology, and immunologic characteristics of monkeys in each clinical disease category. The data underscore the similarities between human and nonhuman primate M. tuberculosis infection. Furthermore, we describe novel methods of quantifying gross pathology and bacterial burden that distinguish between active disease and latent infection, and we extend the usefulness of this model for comparative studies. Early in infection, an abnormal chest X ray, M. tuberculosis growth by gastric aspirate, and increased mycobacterium-specific gamma interferon (IFN-γ) in peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage (BAL) cells were associated with the development of active disease. At necropsy, disease was quantified with respect to pathology and bacterial numbers. Microscopically, a spectrum of granuloma types are seen and differ with disease type. At necropsy, monkeys with active disease had more lung T cells and more IFN-γ from PBMC, BAL, and mediastinal lymph nodes than monkeys with latent infection. Finally, we have observed a spectrum of disease not only in monkeys with active disease but also in those with latent infection that provides insight into human latent tuberculosis. PMID:19620341

[Construction of dengue virus-specific full-length fully human antibody libraries by mammalian display technology].

PubMed

Wen, Yangming; Lan, Kaijian; Wang, Junjie; Yu, Jingyi; Qu, Yarong; Zhao, Wei; Zhang, Fuchun; Tan, Wanlong; Cao, Hong; Zhou, Chen

2013-06-01

To construct dengue virus-specific full-length fully human antibody libraries using mammalian cell surface display technique. Total RNA was extracted from peripheral blood mononuclear cells (PBMCs) from convalescent patients with dengue fever. The reservoirs of the light chain and heavy chain variable regions (LCκ and VH) of the antibody genes were amplified by RT-PCR and inserted into the vector pDGB-HC-TM separately to construct the light chain and heavy chain libraries. The library DNAs were transfected into CHO cells and the expression of full-length fully human antibodies on the surface of CHO cells was analyzed by flow cytometry. Using 1.2 µg of the total RNA isolated from the PBMCs as the template, the LCκ and VH were amplified and the full-length fully human antibody mammalian display libraries were constructed. The kappa light chain gene library had a size of 1.45×10(4) and the heavy chain gene library had a size of 1.8×10(5). Sequence analysis showed that 8 out of the 10 light chain clones and 7 out of the 10 heavy chain clones randomly picked up from the constructed libraries contained correct open reading frames. FACS analysis demonstrated that all the 15 clones with correct open reading frames expressed full-length antibodies, which could be detected on CHO cell surfaces. After co-transfection of the heavy chain and light chain gene libraries into CHO cells, the expression of full-length antibodies on CHO cell surfaces could be detected by FACS analysis with an expressible diversity of the antibody library reaching 1.46×10(9) [(1.45×10(4)×80%)×(1.8×10(5)×70%)]. Using 1.2 µg of total RNA as template, the LCκ and VH full-length fully human antibody libraries against dengue virus have been successfully constructed with an expressible diversity of 10(9).

Solution-Phase Crosstalk and Regulatory Interactions Between Multipotent Adult Progenitor Cells and Peripheral Blood Mononuclear Cells

PubMed Central

van’t Hof, Wouter; Reddy, Ashok P.; Wilmarth, Phillip A.; David, Larry L.; Raber, Amy; Bogaerts, Annelies; Timmerman, Lien; Pinxteren, Jef; Roobrouck, Valerie D.; Deans, Robert J.; Maziarz, Richard T.

2015-01-01

Multipotent adult progenitor cells (MAPCs) are adult adherent stromal stem cells currently being assessed in clinical trials for acute graft versus host disease with demonstrated immunomodulatory capabilities and the potential to ameliorate detrimental autoimmune and inflammation-related processes. Anti-CD3/anti-CD28 (3/28) activation of T cells within the peripheral blood mononuclear cell (PBMC) compartment was performed in the presence or absence of MAPCs. Liquid chromatography-coupled tandem mass spectrometry was used to characterize the differential secretion of proteins, and transcriptional profiling was used to monitor mRNA expression changes in both cell populations. Overall, 239 secreted and/or ectodomain-shed proteins were detected in the secretomes of PBMCs and MAPCs. In addition, 3/28 activation of PBMCs induced differential expression of 2,925 genes, and 22% of these transcripts were differentially expressed on exposure to MAPCs in Transwell. MAPCs exposed to 3/28-activated PBMCs showed differential expression of 1,247 MAPC genes. Crosstalk was demonstrated by reciprocal transcriptional regulation. Secretome proteins and transcriptional signatures were used to predict molecular activities by which MAPCs could dampen local and systemic inflammatory responses. These data support the hypothesis that MAPCs block PBMC proliferation via cell cycle arrest coupled to metabolic stress in the form of tryptophan depletion, resulting in GCN2 kinase activation, downstream signaling, and inhibition of cyclin D1 translation. These data also provide a plausible explanation for the immune privilege reported with administration of donor MAPCs. Although most components of the major histocompatibility complex class II antigen presentation pathway were markedly transcriptionally upregulated, cell surface expression of human leukocyte antigen-DR is minimal on MAPCs exposed to 3/28-activated PBMCs. Significance This study documents experiments quantifying solution

Comparative study of different procedures for the separation of peripheral blood mononuclear cells in cytokine-induced killer cell immunotherapy for hepatocarcinoma.

PubMed

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

2015-04-01

Cytokine-induced killer (CIK) cell immunotherapy exhibits significant advantages in the clinical treatment of tumors. This study was designed to compare the biological characteristics of autologous CIK cells from patients with hepatocarcinoma following different procedures for the separation of peripheral blood mononuclear cells (PBMCs). Forty-four hepatocarcinoma patients were enrolled and distributed into two groups. PBMCs were isolated either using a blood cell separator (apheresis method) or Ficoll lymphocyte separation medium (Ficoll method). The total amount, collection efficacy, and cell status of PBMCs in the two groups were determined. According to the number and status of collected PBMCs, different cultivation procedures were used for their amplification and activation and the proliferation ability, phenotype, and killing activity of CIK cells in the two groups were evaluated. Our results indicated that the number of collected PBMCs in the apheresis group was far more than that in the Ficoll group. However, the isolation rate was lower, and more cellular debris was observed in the apheresis group, which may be the cause of some untoward effects. Following in vitro culture, the enrichment time of CIK cells was longer in the Ficoll group, and the percentages of CD3(+)CD4(+) (Th) and CD4(+)CD25(+) (Treg) cells were higher. In the apheresis group, the percentages of CD3(-)CD56(+) (NK) and CD3(+)CD56(+) (NKT) cells were higher, and the CIK cells exhibited a higher cytolytic activity against HepG2 hepatoma cells. In conclusion, different procedures for PBMCs separation can influence the biological activities of CIK cells, and the apheresis method is more effective at enhancing the antitumor efficacy of CIK cells. However, significant attention should be paid to the possibility of adverse reactions in apheresis donors.

ANALYSIS OF HUMAN ACTIVITY DATA FOR USE IN MODELING ENVIRONMENTAL EXPOSURES

EPA Science Inventory

Human activity data are a critical part of exposure models being developed by the US EPA's National Exposure Research Laboratory (NERL). An analysis of human activity data within NERL's Consolidated Human Activity Database (CHAD) was performed in two areas relevant to exposure ...

Docosahexaenoyl serotonin emerges as most potent inhibitor of IL-17 and CCL-20 released by blood mononuclear cells from a series of N-acyl serotonins identified in human intestinal tissue.

PubMed

Wang, Ya; Balvers, Michiel G J; Hendriks, Henk F J; Wilpshaar, Tessa; van Heek, Tjarda; Witkamp, Renger F; Meijerink, Jocelijn

2017-09-01

Fatty acid amides (FAAs), conjugates of fatty acids with ethanolamine, mono-amine neurotransmitters or amino acids are a class of molecules that display diverse functional roles in different cells and tissues. Recently we reported that one of the serotonin-fatty acid conjugates, docosahexaenoyl serotonin (DHA-5-HT), previously found in gut tissue of mouse and pig, attenuates the IL-23-IL-17 signaling axis in LPS-stimulated mice macrophages. However, its presence and effects in humans remained to be elucidated. Here, we report for the first time its identification in human intestinal (colon) tissue, along with a series of related N-acyl serotonins. Furthermore, we tested these fatty acid conjugates for their ability to inhibit the release of IL-17 and CCL-20 by stimulated human peripheral blood mononuclear cells (PBMCs). Serotonin conjugates with palmitic acid (PA-5-HT), stearic acid (SA-5-HT) and oleic acid (OA-5-HT) were detected in higher levels than arachidonoyl serotonin (AA-5-HT) and DHA-5-HT, while eicosapentaenoyl serotonin (EPA-5-HT) could not be quantified. Among these, DHA-5-HT was the most potent in inhibiting IL-17 and CCL-20, typical Th17 pro-inflammatory mediators, by Concanavalin A (ConA)-stimulated human PBMCs. These results underline the idea that DHA-5-HT is a gut-specific endogenously produced mediator with the capacity to modulate the IL-17/Th17 signaling response. Our findings may be of relevance in relation to intestinal inflammatory diseases like Crohn's disease and Ulcerative colitis. Copyright © 2017. Published by Elsevier B.V.

Simulation-Based Mission Rehearsal as a Human Activity System.

DTIC Science & Technology

1996-09-01

explain this demonstrated importance of the people involved in MR, a human activity system model of simulation-based rehearsal was developed. It provides...Implications of this human activity system view are discussed, including: places in the mission preparation process where simulation can benefit operations

THE NATIONAL EXPOSURE RESEARCH LABORATORY'S COMPREHENSIVE HUMAN ACTIVITY DATABASE

EPA Science Inventory

EPA's National Exposure Research Laboratory (NERL) has combined data from nine U.S. studies related to human activities into one comprehensive data system that can be accessed via the world-wide web. The data system is called CHAD-Consolidated Human Activity Database-and it is ...

THE NATIONAL EXPOSURE RESEARCH LABORATORY'S CONSOLIDATED HUMAN ACTIVITY DATABASE

EPA Science Inventory

EPA's National Exposure Research Laboratory (NERL) has combined data from 12 U.S. studies related to human activities into one comprehensive data system that can be accessed via the Internet. The data system is called the Consolidated Human Activity Database (CHAD), and it is ...

Anti-inflammatory and anti-bacterial properties of tetramethylhexadecenyl succinyl cysteine (TSC): a skin-protecting cosmetic functional ingredient.

PubMed

Fernandéz, J R; Rouzard, K; Voronkov, M; Huber, K L; Stock, J B; Stock, M; Gordon, J S; Pérez, E

2015-02-01

The skin is the first line of defence against exposure to microbial, physical, environmental and chemical insults. In mobilizing a protective response, several different cell types located in our skin release and respond to pro-inflammatory cytokines ensuring skin homeostasis and health. However, chronic activation of this response eventually causes damage resulting in premature ageing. Diosodium tetramethylhexadecenyl succinyl cysteine (TSC or SIG1273), an isoprenylcysteine small molecule, down modulates these inflammatory signalling pathways in various cell types (keratinocytes, peripheral blood mononuclear cells (PBMCs) and endothelial cells) and possesses anti-bacterial properties. Thus, TSC represents a novel cosmetic functional ingredient that provides a broad spectrum of benefits for the skin. To assess the anti-inflammatory properties of TSC in several cutaneous cell types and further investigate its anti-microbial activity. Cultured normal human epidermal keratinocytes were exposed to chemical irritant phorbol 12-myrisate 13-acetate (TPA) or ultraviolet-B light (UVB) to induce pro-inflammatory cytokine (IL-6, IL-8 and TNF-α) production. T-cell receptor (TCR) activation of PBMCs and nickel (Ni(2+) ) treatments of human dermal microvascular endothelial cells (HDMECs) were performed resulting in IL-4, IL-6, IL-8 and IL-17 production. Streptococcus pyogenes were cultured to determine minimal inhibitory concentration values. In vitro studies demonstrate TSC blocks TPA and UVB-induced cytokine production in cultured keratinocytes. Similarly, TSC inhibits overproduction of IL-4 and IL-17 in T-cell receptor (TCR)-activated PBMCs as well as nickel induction of IL-6 and IL-8 in HDMECs. Lastly, TSC demonstrated anti-microbial properties, inhibiting cell growth of S. pyogenes. Tetramethylhexadecenyl succinyl cysteine represents a novel cosmetic functional ingredient that provides a dual modulating benefit of skin protection to individuals by reducing inflammation in

Coffee extracts suppress tryptophan breakdown in mitogen-stimulated peripheral blood mononuclear cells.

PubMed

Gostner, Johanna M; Schroecksnadel, Sebastian; Jenny, Marcel; Klein, Angela; Ueberall, Florian; Schennach, Harald; Fuchs, Dietmar

2015-01-01

Coffee consumption is considered to exert an influence on mood, the immune system, cardiovascular disease, and cancer development, but the mechanisms of action of coffee and its compounds are only partly known and understood. Immunomodulatory effects of filtered extracts of coffee and decaffeinated coffee as well as coffee compounds were investigated in human peripheral blood mononuclear cells (PBMCs) stimulated with mitogen phytohemagglutinin (PHA). The activation of PBMCs was monitored by the breakdown of tryptophan to kynurenine via enzyme indoleamine 2,3-dioxygenase (IDO) and the production of the immune activation marker neopterin by GTP-cyclohydrolase I (GCH1). Both of these biochemical pathways are induced during cellular immune activation in response to the Th1-type cytokine interferon-γ (IFN-γ). Filtered extracts of coffee and decaffeinated coffee both suppressed tryptophan breakdown and neopterin formation in mitogen-stimulated PBMCs efficiently and in a dose-dependent manner. Of 4 coffee compounds tested individually, only gallic acid and less strong also caffeic acid had a consistent suppressive influence but also affected cell viability, whereas pure caffeine and chlorogenic acid exerted no relevant effect in the PBMC assay. The parallel influence of extracts on tryptophan breakdown and neopterin production shows an anti-inflammatory and immunosuppressive property of coffee extracts and some of its compounds. When extrapolating the in vitro results to in vivo, IFN-γ-mediated breakdown of tryptophan could be counteracted by the consumption of coffee or decaffeinated coffee. This may increase tryptophan availability for the biosynthesis of the neurotransmitter 5-hydroxytryptamine (serotonin) and thereby improve mood and quality of life.

Injection drug use facilitates hepatitis C virus infection of peripheral blood mononuclear cells.

PubMed

Resti, Massimo; Azzari, Chiara; Moriondo, Maria; Betti, Letizia; Sforzi, Idanna; Novembre, Elio; Vierucci, Alberto

2002-08-01

Infection of peripheral blood mononuclear cells (PBMCs) with hepatitis C virus (HCV) has been demonstrated and has been found to play a role in relapse of HCV disease and vertical transmission of HCV. Injection drug use is thought to impair function of the immune system and induce tolerance to viruses; therefore, HCV infection of PBMCs could be more likely to occur in injection drug users (IDUs) with HCV infection. Of 108 women who tested negative for human immunodeficiency virus type 1 and positive for HCV RNA, 51 had a history of injection drug use and 57 had no known risk factor for HCV infection. HCV infection was found, by nested reverse-transcription polymerase chain reaction analysis, in the PBMCs of 33 IDUs and of 13 non-IDUs (P=.00003). No correlation was found between infection of the PBMCs and HCV genotype or virus load. Route of transmission and viral factors, as well as immunologic dysfunction, may play a role in viral tropism.

Human Activity Helps Prey Win the Predator-Prey Space Race

PubMed Central

Muhly, Tyler B.; Semeniuk, Christina; Massolo, Alessandro; Hickman, Laura; Musiani, Marco

2011-01-01

Predator-prey interactions, including between large mammalian wildlife species, can be represented as a “space race”, where prey try to minimize and predators maximize spatial overlap. Human activity can also influence the distribution of wildlife species. In particular, high-human disturbance can displace large carnivore predators, a trait-mediated direct effect. Predator displacement by humans could then indirectly benefit prey species by reducing predation risk, a trait-mediated indirect effect of humans that spatially decouples predators from prey. The purpose of this research was to test the hypothesis that high-human activity was displacing predators and thus indirectly creating spatial refuge for prey species, helping prey win the “space race”. We measured the occurrence of eleven large mammal species (including humans and cattle) at 43 camera traps deployed on roads and trails in southwest Alberta, Canada. We tested species co-occurrence at camera sites using hierarchical cluster and nonmetric multidimensional scaling (NMS) analyses; and tested whether human activity, food and/or habitat influenced predator and prey species counts at camera sites using regression tree analysis. Cluster and NMS analysis indicated that at camera sites humans co-occurred with prey species more than predator species and predator species had relatively low co-occurrence with prey species. Regression tree analysis indicated that prey species were three times more abundant on roads and trails with >32 humans/day. However, predators were less abundant on roads and trails that exceeded 18 humans/day. Our results support the hypothesis that high-human activity displaced predators but not prey species, creating spatial refuge from predation. High-human activity on roads and trails (i.e., >18 humans/day) has the potential to interfere with predator-prey interactions via trait-mediated direct and indirect effects. We urge scientist and managers to carefully consider and quantify

Diisopropylfluorophosphate-sensitive aryl acylamidase activity of fatty acid free human serum albumin.

PubMed

Manoharan, Indumathi; Boopathy, Rathnam

2006-08-15

Butyrylcholinesterase in human plasma and acetylcholinesterase in human red blood cells have aryl acylamidase activity toward o-nitroacetanilide, hydrolyzing the amide bond to produce o-nitroaniline and acetate. People with a genetic variant of butyrylcholinesterase that had no detectable activity with butyrylthiocholine, nevertheless had aryl acylamidase activity in their plasma. To determine the source of this aryl acylamidase activity we tested fatty acid free human albumin for activity. We found that albumin had aryl acylacylamidase activity and that this activity was inhibited by diisopropylfluorophosphate. Since the esterase activity of albumin is also inhibited by diisopropylfluorophosphate, and since it is known that diisopropylfluorophosphate covalently binds to Tyr 411 of human albumin, we conclude that the active site for aryl acylamidase activity of albumin is Tyr 411. Albumin accounts for about 10% of the aryl acylamidase activity in human plasma.

Rebamipide, an Amino Acid Analog of 2(1H)-Quinolinone, Inhibits the Formation of Human Osteoclasts

PubMed Central

Kobashigawa, Tsuyoshi

2016-01-01

Objectives. Drug repositioning or drug reprofiling (DR) has recently been growing in importance. DR has a significant advantage over traditional drug development because the repositioned drug has already passed toxicity tests; its safety is known, and the risk of adverse toxicology is reduced. In the current study, we investigated the role of rebamipide, a mucosa-protecting agent, with recently reported anti-inflammatory function, in human osteoclastogenesis. Methods. Peripheral blood mononuclear cells (PBMCs) were cultured in the presence of M-CSF and sRANKL. Osteoclast formation was evaluated by immunohistological staining for CD51/61 (vitronectin receptors). Osteoclast formation, in the presence or absence of rebamipide (0, 1, and 3 mM), was observed by time-lapse photography and actin ring formation. The number of absorption sites and area of absorption were calculated using Osteologic™ plates. Pit formation was studied by 3D-SEM. Results. Rebamipide inhibited human osteoclast formation at 3 mM, a pharmacological concentration, and inhibited resorbing activity dose-dependently. Rebamipide induced the degradation of actin rings in mature osteoclasts. This mechanism may involve inhibiting the osteoclast fusion pathway through reducing the expression of DC-specific transmembrane protein (DC-STAMP). Conclusions. The present study suggests that rebamipide would be useful as a novel agent for osteoporosis and rheumatoid arthritis. PMID:27965978

Rebamipide, an Amino Acid Analog of 2(1H)-Quinolinone, Inhibits the Formation of Human Osteoclasts.

PubMed

Nanke, Yuki; Kobashigawa, Tsuyoshi; Yago, Toru; Kawamoto, Manabu; Yamanaka, Hisashi; Kotake, Shigeru

2016-01-01

Objectives . Drug repositioning or drug reprofiling (DR) has recently been growing in importance. DR has a significant advantage over traditional drug development because the repositioned drug has already passed toxicity tests; its safety is known, and the risk of adverse toxicology is reduced. In the current study, we investigated the role of rebamipide, a mucosa-protecting agent, with recently reported anti-inflammatory function, in human osteoclastogenesis. Methods. Peripheral blood mononuclear cells (PBMCs) were cultured in the presence of M-CSF and sRANKL. Osteoclast formation was evaluated by immunohistological staining for CD51/61 (vitronectin receptors). Osteoclast formation, in the presence or absence of rebamipide (0, 1, and 3 mM), was observed by time-lapse photography and actin ring formation. The number of absorption sites and area of absorption were calculated using Osteologic™ plates. Pit formation was studied by 3D-SEM. Results . Rebamipide inhibited human osteoclast formation at 3 mM, a pharmacological concentration, and inhibited resorbing activity dose-dependently. Rebamipide induced the degradation of actin rings in mature osteoclasts. This mechanism may involve inhibiting the osteoclast fusion pathway through reducing the expression of DC-specific transmembrane protein (DC-STAMP). Conclusions . The present study suggests that rebamipide would be useful as a novel agent for osteoporosis and rheumatoid arthritis.

The effect of ex vivo CDDO-Me activation on nuclear factor erythroid 2-related factor 2 pathway in white blood cells from patients with septic shock.

PubMed

Noel, Sanjeev; Zheng, Laura; Navas-Acien, Ana; Fuchs, Ralph J

2014-11-01

Nuclear factor erythroid 2-related factor 2 (NRF2) has been shown to protect against experimental sepsis in mice and lipopolysaccharide (LPS)-induced inflammation in ex vivo white blood cells from healthy subjects by upregulating cellular antioxidant genes. The objective of this study was to test the hypothesis that ex vivo methyl 2-cyano-3,12-dioxoolean-1,9-dien-28-oate (CDDO-Me) activates NRF2-regulated antioxidant genes in white blood cells from patients with septic shock and protects against LPS-induced inflammation and reactive oxidative species production. Peripheral blood was collected from 18 patients with septic shock who were being treated in medical and surgical intensive care units. Real-time polymerase chain reaction was used to quantify the expression of NRF2 target genes (NQO1, HO-1, GCLM, and FTL) and IL-6 in peripheral blood mononuclear cells (PBMCs), monocytes, and neutrophils after CDDO-Me treatment alone or after subsequent LPS exposure. Superoxide anion (O2) was measured to assess the effect of CDDO-Me pretreatment on subsequent LPS exposure. Treatment with CDDO-Me increased the gene expression of NQO1 (P = 0.04) and decreased the expression of HO-1 (P = 0.03) in PBMCs from patients with septic shock. Purified monocytes exhibited significant increases in the expression of NQO1 (P = 0.01) and GCLM (P = 0.003) after CDDO-Me treatment. Levels of other NRF2 target genes (HO-1 and FTL) remained similar to those of vehicle-treated cells. Peripheral blood mononuclear cells showed a trend toward increased IL-6 gene expression after CDDO-Me treatment, whereas purified monocytes showed a trend toward decreased IL-6. There was no discernible trend in the IL-6 expression subsequent to LPS treatment in either vehicle-treated or CDDO-Me-treated PBMCs and monocytes. Treatment with CDDO-Me significantly increased O2 production in PBMCs (P = 0.04). Although CDDO-Me pretreatment significantly attenuated O2 production to subsequent LPS exposure (P = 0.03), the

Kefir induces apoptosis and inhibits cell proliferation in human acute erythroleukemia.

PubMed

Jalali, Fatemeh; Sharifi, Mohammadreza; Salehi, Rasoul

2016-01-01

Acute erythroleukemia is an uncommon subtype of acute myeloid leukemia which has been considered to be a subtype of AML with a worse prognosis. Intensive chemotherapy is the first line of treatment. In recent years, the effect of kefir on some malignancies has been experimented. Kefir is a kind of beverage, which obtained by incubation of kefir grains with raw milk. Kefir grains are a symbiotic complex of different kinds of yeasts and bacteria, especially lactic acid bacteria which gather in a mostly carbohydrate matrix, named kefiran. We investigated the effect of kefir on acute erythroleukemia cell line (KG-1) and peripheral blood mononuclear cells (PBMCs). The cell line and PBMCs were treated with different doses of kefir and milk and incubated for three different times. We used Polymixin B to block the lipopolysaccharide and NaOH (1 mol/l) to neutralize the acidic media. Viability was detected by MTT assay. Apoptosis and necrosis were assessed by annexin-propidium iodide staining. Our results showed that kefir induced apoptosis and necrosis in KG-1 cell line. It was revealed that kefir decreased proliferation in erythroleukemia cell line. We did not observe a remarkable effect of kefir on PBMCs. Our study suggested that kefir may have potential to be an effective treatment for erythroleukemia.

Activity-Dependent Human Brain Coding/Noncoding Gene Regulatory Networks

PubMed Central

Lipovich, Leonard; Dachet, Fabien; Cai, Juan; Bagla, Shruti; Balan, Karina; Jia, Hui; Loeb, Jeffrey A.

2012-01-01

While most gene transcription yields RNA transcripts that code for proteins, a sizable proportion of the genome generates RNA transcripts that do not code for proteins, but may have important regulatory functions. The brain-derived neurotrophic factor (BDNF) gene, a key regulator of neuronal activity, is overlapped by a primate-specific, antisense long noncoding RNA (lncRNA) called BDNFOS. We demonstrate reciprocal patterns of BDNF and BDNFOS transcription in highly active regions of human neocortex removed as a treatment for intractable seizures. A genome-wide analysis of activity-dependent coding and noncoding human transcription using a custom lncRNA microarray identified 1288 differentially expressed lncRNAs, of which 26 had expression profiles that matched activity-dependent coding genes and an additional 8 were adjacent to or overlapping with differentially expressed protein-coding genes. The functions of most of these protein-coding partner genes, such as ARC, include long-term potentiation, synaptic activity, and memory. The nuclear lncRNAs NEAT1, MALAT1, and RPPH1, composing an RNAse P-dependent lncRNA-maturation pathway, were also upregulated. As a means to replicate human neuronal activity, repeated depolarization of SY5Y cells resulted in sustained CREB activation and produced an inverse pattern of BDNF-BDNFOS co-expression that was not achieved with a single depolarization. RNAi-mediated knockdown of BDNFOS in human SY5Y cells increased BDNF expression, suggesting that BDNFOS directly downregulates BDNF. Temporal expression patterns of other lncRNA-messenger RNA pairs validated the effect of chronic neuronal activity on the transcriptome and implied various lncRNA regulatory mechanisms. lncRNAs, some of which are unique to primates, thus appear to have potentially important regulatory roles in activity-dependent human brain plasticity. PMID:22960213

Detection of cardiac activity changes from human speech

NASA Astrophysics Data System (ADS)

Tovarek, Jaromir; Partila, Pavol; Voznak, Miroslav; Mikulec, Martin; Mehic, Miralem

2015-05-01

Impact of changes in blood pressure and pulse from human speech is disclosed in this article. The symptoms of increased physical activity are pulse, systolic and diastolic pressure. There are many methods of measuring and indicating these parameters. The measurements must be carried out using devices which are not used in everyday life. In most cases, the measurement of blood pressure and pulse following health problems or other adverse feelings. Nowadays, research teams are trying to design and implement modern methods in ordinary human activities. The main objective of the proposal is to reduce the delay between detecting the adverse pressure and to the mentioned warning signs and feelings. Common and frequent activity of man is speaking, while it is known that the function of the vocal tract can be affected by the change in heart activity. Therefore, it can be a useful parameter for detecting physiological changes. A method for detecting human physiological changes by speech processing and artificial neural network classification is described in this article. The pulse and blood pressure changes was induced by physical exercises in this experiment. The set of measured subjects was formed by ten healthy volunteers of both sexes. None of the subjects was a professional athlete. The process of the experiment was divided into phases before, during and after physical training. Pulse, systolic, diastolic pressure was measured and voice activity was recorded after each of them. The results of this experiment describe a method for detecting increased cardiac activity from human speech using artificial neural network.

Winter to summer change in vitamin D status reduces systemic inflammation and bioenergetic activity of human peripheral blood mononuclear cells.

PubMed

Calton, Emily K; Keane, Kevin N; Raizel, Raquel; Rowlands, Jordan; Soares, Mario J; Newsholme, Philip

2017-08-01

Vitamin D status [25(OH)D] has recently been reported to be associated with altered cellular bioenergetic profiles of peripheral blood mononuclear cells (PBMCs). No study has tracked the seasonal variation of 25(OH)D and its putative influence on whole body energy metabolism, cellular bioenergetic profiles, inflammatory markers and clinical chemistry. Whole body energy metabolism and substrate utilisation were measured by indirect calorimetry. PBMCs obtained from the same subjects were isolated from whole blood, counted and freshly seeded. Bioenergetic analysis (mitochondrial stress test and glycolysis stress test) was performed using the Seahorse XF e 96 flux analyser. 25(OH)D was assessed using the Architect immunoassay method. 25(OH)D increased by a median (IQR) of 14.40 (20.13)nmol/L (p<0.001) from winter to summer and was accompanied by significant improvements in indices of insulin sensitivity, McAuley's index (p=0.019) and quantitative insulin sensitivity check index (p=0.028). PBMC mitochondrial parameters basal respiration, non-mitochondrial respiration, ATP production, proton leak, and maximal respiration decreased in summer compared to winter. Similarly, PBMC glycolytic parameters glycolytic activity, glucose response, and glycolytic capacity were all reduced in summer compared to winter. There was also a trend for absolute resting metabolic rate (RMR) to decrease (p=0.066). Markers of systemic inflammation MCP-1, IL-6, IL-8, IL-10, and IL-12p70 decreased significantly in summer compared to winter. Participants who entered winter with a low 25(OH)D (<50nmol/L), had the greatest alteration in bioenergetic parameters in summer, relative to those with winter 25(OH)D concentrations of 50-75nmol/L or >75nmol/L. The absolute change in 25(OH)D was not associated with altered bioenergetics. Seasonal improvements in 25(OH)D was associated with reduced systemic inflammation, PBMC bioenergetic profiles and whole body energy metabolism. These observational changes

Human milk proteins: key components for the biological activity of human milk.

PubMed

Lönnerdal, Bo

2004-01-01

Human milk contains a wide array of proteins that provide biologic activities ranging from antimicrobial effects to immunostimulatory functions. Proteins like lactoferrin, secretory IgA, kappa-casein, lactoperoxidase, haptocorrin, lactadherin and peptides formed from human milk proteins during digestion can inhibit the growth of pathogenic bacteria and viruses and therefore protect against infection. At the same time, proteins like lactoferrin, bile-salt stimulated lipase, haptocorrin, kappa-casein, and folate-binding protein can facilitate the absorption of nutrients in the neonatal gut. However, the proteins in human milk themselves also provide adequate amounts of essential amino acids to the growing infant. This suggests a highly adapted digestive system, which allows the survival of some proteins and peptides in the upper gastrointestinal tract, while still allowing amino acid utilization from these proteins further down in the gut. It is now possible to produce recombinant human milk proteins in transgenic plants and animals, which makes it possible to further study the bioactivity of these proteins. Provided these proteins can be produced in large scale at low cost, that they show biologic activity and pose no safety concerns, it may be possible to add some human milk proteins to infant diets, such as formula and complementary foods. Human milk proteins produced in rice or potatoes, for example, could be added without much purification, because these staples commonly are used in weaning foods. Thus, some qualities provided by human milk may be included into other diets, although it is highly unlikely that all unique components of human milk can be copied this way.

Human telomerase: biogenesis, trafficking, recruitment, and activation.

PubMed

Schmidt, Jens C; Cech, Thomas R

2015-06-01

Telomerase is the ribonucleoprotein enzyme that catalyzes the extension of telomeric DNA in eukaryotes. Recent work has begun to reveal key aspects of the assembly of the human telomerase complex, its intracellular trafficking involving Cajal bodies, and its recruitment to telomeres. Once telomerase has been recruited to the telomere, it appears to undergo a separate activation step, which may include an increase in its repeat addition processivity. This review covers human telomerase biogenesis, trafficking, and activation, comparing key aspects with the analogous events in other species. © 2015 Schmidt and Cech Published by Cold Spring Harbor Laboratory Press.

Evaluation of Antigen-Specific IgM and IgG Production during an In Vitro Peripheral Blood Mononuclear Cell Culture Assay.

PubMed

Matsuda, Yoshiko; Imamura, Ryoichi; Takahara, Shiro

2017-01-01

The recent attention given to diseases associated with memory B-cell (mBC)-produced antibodies (Abs) suggests the need for a similar in vitro assay to evaluate the functions of mBCs. Here, we cultured peripheral blood mononuclear cells (PBMCs) with the intent to collect mBC-derived Abs in vitro and maintain their cell-cell contact-dependent interactions with helper T-cells. PBMCs were cultured with interleukin (IL)-21, CpG-oligodeoxynucleotides (ODN), phorbol myristate acetate (PMA), and phytohemagglutinin/leucoagglutinin (PHA-L) in 24-well flat-bottom plates (5 × 10 5  cells/well). A culture supernatant analysis of PBMCs from healthy donors ( n  = 10) indicated that antigen-specific IgM Ab levels in a PBMC culture supernatant might be better able to demonstrate the antigen sensitization status in a smaller peripheral blood sample, compared to IgG because Epstein-Barr virus-specific IgM mBCs circulate peripherally at a significantly higher frequency once antiviral humoral immunity has stabilized. Thus, our in vitro assay demonstrated the potential significance of antigen-specific IgM Ab production in the culture supernatants. Furthermore, an analysis of cultured PBMCs from allograft kidney recipients ( n  = 16) sensitized with de novo donor-specific human leukocyte antigen (HLA)-specific Abs (DSAs) showed that IgM-type HLA-specific Abs were detected mainly from the culture supernatants from PBMCs of patients with stable graft function, whereas IgG isotype HLA Abs were detectable only from patients with biopsy-proven antibody-mediated rejection. In other words, these IgG isotype Abs also represented an activated humoral immune response in vivo . Additionally, IgM- and IgG-expressing mBCs from healthy donors ( n  = 5) were cultured with IL-21, CpG-ODN, and a supernatant produced by stimulating CD19 + B-cell-depleted PBMCs with PHA-L and PMA in 24-well flat-bottom plates (1 × 10 5  cells/well), and the resulting in vitro analysis provided

Cell–specific Variation in E-selectin Ligand Expression Among Human Peripheral Blood Mononuclear Cells: Implications for Immunosurveillance and Pathobiology

PubMed Central

Silva, Mariana; Fung, Ronald Kam Fai; Donnelly, Conor Brian; Videira, Paula Alexandra; Sackstein, Robert

2017-01-01

Both host defense and immunopathology are shaped by the ordered recruitment of circulating leukocytes to affected sites, a process initiated by binding of blood-borne cells to E-selectin displayed at target endothelial beds. Accordingly, knowledge of the expression and function of leukocyte E-selectin ligands is key to understanding the tempo and specificity of immunoreactivity. Here, we performed E-selectin adherence assays under hemodynamic flow conditions coupled with flow cytometry and western blot analysis to elucidate the function and structural biology of glycoprotein E-selectin ligands expressed on human peripheral blood mononuclear cells (PBMCs). Circulating monocytes uniformly express high levels of the canonical E-selectin binding determinant sLeX and display markedly greater adhesive interactions with E-selectin than do circulating lymphocytes, which exhibit variable E-selectin binding among CD4+ and CD8+ T-cells but no binding by B-cells. Monocytes prominently present sLeX decorations on an array of protein scaffolds including PSGL-1, CD43, and CD44 (rendering the E-selectin ligands CLA, CD43E, and HCELL, respectively), and B-cells altogether lack E-selectin ligands. Quantitative PCR gene expression studies of glycosyltransferases that regulate display of sLeX reveal high transcript levels among circulating monocytes and low levels among circulating B-cells, and, commensurately, cell surface α(1,3)-fucosylation reveals that acceptor sialyllactosaminyl glycans convertible into sLeX are abundantly expressed on human monocytes yet are relatively deficient on B-cells. Collectively, these findings unveil distinct cell-specific patterns of E-selectin ligand expression among human PBMCs, indicating that circulating monocytes are specialized to engage E-selectin and providing key insights into the molecular effectors mediating recruitment of these cells at inflammatory sites. PMID:28330896

Drug Modulators of B Cell Signaling Pathways and Epstein-Barr Virus Lytic Activation.

PubMed

Kosowicz, John G; Lee, Jaeyeun; Peiffer, Brandon; Guo, Zufeng; Chen, Jianmeng; Liao, Gangling; Hayward, S Diane; Liu, Jun O; Ambinder, Richard F

2017-08-15

Epstein-Barr virus (EBV) is a ubiquitous human gammaherpesvirus that establishes a latency reservoir in B cells. In this work, we show that ibrutinib, idelalisib, and dasatinib, drugs that block B cell receptor (BCR) signaling and are used in the treatment of hematologic malignancies, block BCR-mediated lytic induction at clinically relevant doses. We confirm that the immunosuppressive drugs cyclosporine and tacrolimus also inhibit BCR-mediated lytic induction but find that rapamycin does not inhibit BCR-mediated lytic induction. Further investigation shows that mammalian target of rapamycin complex 2 (mTORC2) contributes to BCR-mediated lytic induction and that FK506-binding protein 12 (FKBP12) binding alone is not adequate to block activation. Finally, we show that BCR signaling can activate EBV lytic induction in freshly isolated B cells from peripheral blood mononuclear cells (PBMCs) and that activation can be inhibited by ibrutinib or idelalisib. IMPORTANCE EBV establishes viral latency in B cells. Activation of the B cell receptor pathway activates lytic viral expression in cell lines. Here we show that drugs that inhibit important kinases in the BCR signaling pathway inhibit activation of lytic viral expression but do not inhibit several other lytic activation pathways. Immunosuppressant drugs such as cyclosporine and tacrolimus but not rapamycin also inhibit BCR-mediated EBV activation. Finally, we show that BCR activation of lytic infection occurs not only in tumor cell lines but also in freshly isolated B cells from patients and that this activation can be blocked by BCR inhibitors. Copyright © 2017 American Society for Microbiology.

Neuropeptides activate human mast cell degranulation and chemokine production

PubMed Central

Kulka, Marianna; Sheen, Cecilia H; Tancowny, Brian P; Grammer, Leslie C; Schleimer, Robert P

2008-01-01

During neuronal-induced inflammation, mast cells may respond to stimuli such as neuropeptides in an FcεRI-independent manner. In this study, we characterized human mast cell responses to substance P (SP), nerve growth factor (NGF), calcitonin gene-related peptide (CGRP) and vasoactive intestinal polypeptide (VIP) and compared these responses to human mast cell responses to immunoglobulin E (IgE)/anti-IgE and compound 48/80. Primary cultured mast cells, generated from CD34+ progenitors in the presence of stem cell factor and interleukin-6 (IL-6), and human cultured mast cells (LAD2) were stimulated with these and other stimuli (gastrin, concanavalin A, radiocontrast media, and mannitol) and their degranulation and chemokine production was assessed. VIP and SP stimulated primary human mast cells and LAD cells to degranulate; gastrin, concanavalin A, radiocontrast media, mannitol, CGRP and NGF did not activate degranulation. While anti-IgE stimulation did not induce significant production of chemokines, stimulation with VIP, SP or compound 48/80 potently induced production of monocyte chemoattractant protein-1, inducible protein-10, monokine induced by interferon-γ (MIG), RANTES (regulated on activation, normal, T-cell expressed, and secreted) and IL-8. VIP, SP and compound 48/80 also activated release of tumour necrosis factor, IL-3 and granulocyte–macrophage colony-stimulating factor, but not IL-4, interferon-γ or eotaxin. Human mast cells expressed surface neurokinin 1 receptor (NK1R), NK2R, NK3R and VIP receptor type 2 (VPAC2) but not VPAC1 and activation of human mast cells by IgE/anti-IgE up-regulated expression of VPAC2, NK2R, and NK3R. These studies demonstrate the pattern of receptor expression and activation of mast cell by a host of G-protein coupled receptor ligands and suggest that SP and VIP activate a unique signalling pathway in human mast cells. These results are likely to have direct relevance to neuronally induced inflammatory diseases. PMID

A novel method for evaluating antibody-dependent cell-mediated cytotoxicity by flowcytometry using cryopreserved human peripheral blood mononuclear cells

PubMed Central

Yamashita, Makiko; Kitano, Shigehisa; Aikawa, Hiroaki; Kuchiba, Aya; Hayashi, Mitsuhiro; Yamamoto, Noboru; Tamura, Kenji; Hamada, Akinobu

2016-01-01

Analyzing the cytotoxic functions of effector cells, such as NK cells against target cancer cells, is thought to be necessary for predicting the clinical efficacy of antibody-dependent cellular cytotoxicity (ADCC) -dependent antibody therapy. The 51Cr release assay has long been the most widely used method for quantification of ADCC activity. However, the reproducibilities of these release assays are not adequate, and they do not allow evaluation of the lysis susceptibilities of distinct cell types within the target cell population. In this study, we established a novel method for evaluating cytotoxicity, which involves the detection and quantification of dead target cells using flowcytometry. CFSE (carboxyfluorescein succinimidyl ester) was used as a dye to specifically stain and thereby label the target cell population, allowing living and dead cells, as well as both target and effector cells, to be quantitatively distinguished. Furthermore, with our new approach, ADCC activity was more reproducibly, sensitively, and specifically detectable, not only in freshly isolated but also in frozen human peripheral blood mononuclear cells (PBMCs), than with the calcein-AM release assay. This assay, validated herein, is expected to become a standard assay for evaluating ADCC activity which will ultimately contribute the clinical development of ADCC dependent-antibody therapies. PMID:26813960

A novel method for evaluating antibody-dependent cell-mediated cytotoxicity by flowcytometry using cryopreserved human peripheral blood mononuclear cells.

PubMed

Yamashita, Makiko; Kitano, Shigehisa; Aikawa, Hiroaki; Kuchiba, Aya; Hayashi, Mitsuhiro; Yamamoto, Noboru; Tamura, Kenji; Hamada, Akinobu

2016-01-27

Analyzing the cytotoxic functions of effector cells, such as NK cells against target cancer cells, is thought to be necessary for predicting the clinical efficacy of antibody-dependent cellular cytotoxicity (ADCC) -dependent antibody therapy. The (51)Cr release assay has long been the most widely used method for quantification of ADCC activity. However, the reproducibilities of these release assays are not adequate, and they do not allow evaluation of the lysis susceptibilities of distinct cell types within the target cell population. In this study, we established a novel method for evaluating cytotoxicity, which involves the detection and quantification of dead target cells using flowcytometry. CFSE (carboxyfluorescein succinimidyl ester) was used as a dye to specifically stain and thereby label the target cell population, allowing living and dead cells, as well as both target and effector cells, to be quantitatively distinguished. Furthermore, with our new approach, ADCC activity was more reproducibly, sensitively, and specifically detectable, not only in freshly isolated but also in frozen human peripheral blood mononuclear cells (PBMCs), than with the calcein-AM release assay. This assay, validated herein, is expected to become a standard assay for evaluating ADCC activity which will ultimately contribute the clinical development of ADCC dependent-antibody therapies.

In vitro effects of ATG-Fresenius on immune cell adhesion.

PubMed

Kanzler, I; Seitz-Merwald, I; Schleger, S; Kaczmarek, I; Kur, F; Beiras-Fernandez, A

2013-06-01

ATG-Fresenius, a purified rabbit polyclonal anti-human T-lymphocyte immunoglobulin is used for induction immunosuppression as well as prevention and treatment of acute rejection episodes among patients receiving solid organ transplants. The aim of this study was to investigate the in vitro activity of ATG-Fresenius upon immune cell adhesion, which may explain its activity to mitigate ischemia-reperfusion injury. Human vascular endothelial cells (HUVEC) and peripheral blood mononuclear cells (PBMCs) isolated from umbilical vein or peripheral blood were incubated 20 to 24 hours before analysis. HUVEC were incubated with 10 and 100 μg/mL ATG-Fresenius or reference polyclonal rabbit immunoglobulin G. Analysis of immune cell adhesion to endothelial cells was studied in cocultures of PBMCs and adherent HUVEC. Endothelial cell expression of adhesion molecules CD62E and CD54 was determined by flow cytometry. The numbers of T-, B- and natural killer cells attached to HUVEC were also determined by flow cytometry. Groups were compared using one-way analysis of variance. We showed that ATG-Fresenius binds to endothelial cells particularly activated ones expressing increased levels of E-selectin and ICAM-1. The increased binding of ATG-Fresenius to activated endothelial cells was consistent with its known binding to Intercellular Adhesion Molecule 1 (ICAM-1) and selectins. We also showed that ATG-Fresenius inhibited adhesion of prestimulated immune cells to activated endothelium. We demonstrated dose-dependent binding of ATG-Fresenius to activated endothelial cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Suppression of human monocyte tissue factor induction by red wine phenolics and synthetic derivatives of resveratrol.

PubMed

Kaur, Gurjeet; Roberti, Marinella; Raul, Francis; Pendurthi, Usha R

2007-01-01

Prevention of cardiovascular disease through nutritional supplements is growing in popularity throughout the world. Multiple epidemiologic studies found that moderate consumption of alcohol, particularly red wine, lowers mortality rates from coronary heart diseases (CHD). Chronic inflammation and atherosclerosis associated with CHD culminate in aberrant intravascular expression of tissue factor (TF), which triggers blood coagulation leading to thrombosis, a major cause for heart attack. We showed earlier that two red wine phenolics, resveratrol and quercetin, suppressed TF induction in endothelial cells. In the present study, we investigated efficacy of seven resveratrol derivatives, which were shown to be effective in regulating cancer cell growth in vitro at much lower concentrations than the parent compound resveratrol, in inhibiting TF induction in peripheral blood mononuclear cells (PBMCs). We also tested possible synergistic effects of resveratrol and quercetin with the other major red wine phenolics in suppression of lipopolysaccharide-induced TF expression in human PBMCs. We found that several resveratrol derivatives were 2- to 10-fold more efficient than resveratrol in inhibiting TF induction. Our study found no evidence for synergism among red wine polyphenolics. These data suggest that structural alterations of resveratrol can be effective in producing potent antithrombotic agents that will have therapeutic potential in the improvement of cardiovascular health and prevention of CHD. Among major red wine phenolics, quercetin appears to be the predominant suppressor of TF induction.

Influence of Fragrances on Human Psychophysiological Activity: With Special Reference to Human Electroencephalographic Response

PubMed Central

Sowndhararajan, Kandhasamy; Kim, Songmun

2016-01-01

The influence of fragrances such as perfumes and room fresheners on the psychophysiological activities of humans has been known for a long time, and its significance is gradually increasing in the medicinal and cosmetic industries. A fragrance consists of volatile chemicals with a molecular weight of less than 300 Da that humans perceive through the olfactory system. In humans, about 300 active olfactory receptor genes are devoted to detecting thousands of different fragrance molecules through a large family of olfactory receptors of a diverse protein sequence. The sense of smell plays an important role in the physiological effects of mood, stress, and working capacity. Electrophysiological studies have revealed that various fragrances affected spontaneous brain activities and cognitive functions, which are measured by an electroencephalograph (EEG). The EEG is a good temporal measure of responses in the central nervous system and it provides information about the physiological state of the brain both in health and disease. The EEG power spectrum is classified into different frequency bands such as delta (0.5–4 Hz), theta (4–8 Hz), alpha (8–13 Hz), beta (13–30 Hz) and gamma (30–50 Hz), and each band is correlated with different features of brain states. A quantitative EEG uses computer software to provide the topographic mapping of the brain activity in frontal, temporal, parietal and occipital brain regions. It is well known that decreases of alpha and beta activities and increases of delta and theta activities are associated with brain pathology and general cognitive decline. In the last few decades, many scientific studies were conducted to investigate the effect of inhalation of aroma on human brain functions. The studies have suggested a significant role for olfactory stimulation in the alteration of cognition, mood, and social behavior. This review aims to evaluate the available literature regarding the influence of fragrances on the

Towards discrete wavelet transform-based human activity recognition

NASA Astrophysics Data System (ADS)

Khare, Manish; Jeon, Moongu

2017-06-01

Providing accurate recognition of human activities is a challenging problem for visual surveillance applications. In this paper, we present a simple and efficient algorithm for human activity recognition based on a wavelet transform. We adopt discrete wavelet transform (DWT) coefficients as a feature of human objects to obtain advantages of its multiresolution approach. The proposed method is tested on multiple levels of DWT. Experiments are carried out on different standard action datasets including KTH and i3D Post. The proposed method is compared with other state-of-the-art methods in terms of different quantitative performance measures. The proposed method is found to have better recognition accuracy in comparison to the state-of-the-art methods.

Potential ex vivo immunomodulatory effects of San-Huang-Xie-Xin-Tang and its component herbs on mice and humans.

PubMed

Li, Chia-Yang; Hou, Yu-Chi; Lee Chao, Pei-Dawn; Shia, Chi-Sheng; Hsu, Ian C; Fang, Shih-Hua

2010-02-03

San-Huang-Xie-Xin-Tang (SHXXT), an important Chinese medicine formula, contains Rhei Rhizoma (RR), Scutellariae Radix (SR) and Coptidis Rhizoma (CR). RR and SR are abundant in anthraquinone and flavonoid polyphenols. Pharmacokinetic study of SHXXT indicated that glucuronides were the predominant forms of polyphenols in rats. As an extension of pharmacokinetic study, the serum metabolites of SHXXT, RR, SR and CR were prepared from rats and quantitated, then the immunomodulation effects were examined by culturing these serum metabolites with murine and human immune cells. The results indicated that the inhibitions on nitric oxide (NO) and cytokine production from mitogen-activated peritoneal macrophages by the serum metabolites of SHXXT, RR, SR and CR were through reducing the protein expression of inducible NO synthase (iNOS) and the IC(50) were 0.8%, 1.5%, 3.0% and 0.8% of their blood concentrations, respectively. In addition, the serum metabolites of SHXXT, RR, SR and CR significantly decreased the ratios of interferon-gamma (IFN-gamma) to interleukin (IL)-4 in mitogen-stimulated mice spleen cells and human peripheral blood mononuclear cells (PBMCs). Moreover, the serum metabolites of SHXXT and SR significantly arrested the mitogen-stimulated mice spleen cells at G2/M stage. In conclusion, the serum metabolites of SHXXT and the component herbs exerted promising modulation activities on the immune functions and the cell cycle distribution of mice and human immune cells. We suggest that SHXXT is a promising remedy for immunomodulation through Th1/Th2 regulation. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

Brain activation during human male ejaculation revisited.

PubMed

Georgiadis, Janniko R; Reinders, A A T Simone; Van der Graaf, Ferdinand H C E; Paans, Anne M J; Kortekaas, Rudie

2007-04-16

In a prior [O]-H2O positron emission tomographic study we reported brain regions involved in human male ejaculation. Here, we used another, more recently acquired data set to evaluate the methodological approach of this previous study, and discovered that part of the reported activation pattern was not related to ejaculation. With a new analysis of these ejaculation data, we now demonstrate ejaculation-related activations in the deep cerebellar nuclei (dentate nucleus), anterior vermis, pons, and ventrolateral thalamus, and, most importantly, ejaculation-related deactivations throughout the prefrontal cortex. This revision offers a new and more accurate insight into the brain regions involved in human male ejaculation.

Predicting human activities in sequences of actions in RGB-D videos

NASA Astrophysics Data System (ADS)

Jardim, David; Nunes, Luís.; Dias, Miguel

2017-03-01

In our daily activities we perform prediction or anticipation when interacting with other humans or with objects. Prediction of human activity made by computers has several potential applications: surveillance systems, human computer interfaces, sports video analysis, human-robot-collaboration, games and health-care. We propose a system capable of recognizing and predicting human actions using supervised classifiers trained with automatically labeled data evaluated in our human activity RGB-D dataset (recorded with a Kinect sensor) and using only the position of the main skeleton joints to extract features. Using conditional random fields (CRFs) to model the sequential nature of actions in a sequence has been used before, but where other approaches try to predict an outcome or anticipate ahead in time (seconds), we try to predict what will be the next action of a subject. Our results show an activity prediction accuracy of 89.9% using an automatically labeled dataset.

Antiviral interactions of combinations of highly potent 2,4(1H,3H)-pyrimidinedione congeners and other anti-HIV agents.

PubMed

Hartman, Tracy L; Yang, Lu; Buckheit, Robert W

2011-12-01

Structure-activity relationship evaluation of seventy-four 2,4(1H,3H)-pyrimidinedione derivatives identified seven lead compounds based on anti-HIV-1 potency, extended range of action to include HIV-2, virus entry inhibition, reverse transcriptase inhibition, and lack of cytotoxicity to human cells. The selected pyrimidinedione congeners are highly active inhibitors of HIV-1 with EC(50) values ranging from 0.6 to 2 nM in CEM-SS cells infected with laboratory derived viruses, 11-20 nM in fresh human PBMCs infected with subtype B (HT/92/599) virus, and 2-7 nM in PBMCs infected with the clinical subtype C (ZA/97/003) virus. Combination antiviral assays were performed using the laboratory adapted RF strain of HIV-1 in CEM-SS cells and with a clade B and C low passage clinical isolate in fresh human peripheral mononuclear cells and the compound interactions were analyzed using MacSynergy II. The seven pyrimidinedione compounds resulted in additive to synergistic interactions in combination with entry and fusion inhibitors, nonnucleoside and nucleoside reverse transcriptase inhibitors, and the protease inhibitors. No evidence of antagonistic antiviral activity or synergistic cytotoxicity was detected with the combinations of compounds tested. The dual mechanism of action of the pyrimidinediones resulting in inhibition of both virus entry and reverse transcription suggests excellent potential of these lead pyrimidinediones as candidates for combination therapy with other approved HIV inhibitors of varying mechanism of action. Copyright © 2011. Published by Elsevier B.V.

Immune Modulation in Normal Human Peripheral Blood Mononuclear Cells (PBMCs) (Lymphocytes) in Response to Benzofuran-2-Carboxylic Acid Derivative KMEG during Spaceflight

NASA Astrophysics Data System (ADS)

Okoro, Elvis; Mann, Vivek; Ellis, Ivory; Mansoor, Elvedina; Olamigoke, Loretta; Marriott, Karla Sue; Denkins, Pamela; Williams, Willie; Sundaresan, Alamelu

2017-08-01

Microgravity and radiation exposure during space flight have been widely reported to induce the suppression of normal immune system function, and increase the risk of cancer development in humans. These findings pose a serious risk to manned space missions. Interestingly, recent studies have shown that benzofuran-2-carboxylic acid derivatives can inhibit the progression of some of these devastating effects on earth and in modeled microgravity. However, these studies had not assessed the impacts of benzofuran-2- carboxylic acid and its derivatives on global gene expression under spaceflight conditions. In this study, the ability of a specific benzofuran-2-carboxylic acid derivative (KMEG) to confer protection from radiation and restore normal immune function was investigated following exposure to space flight conditions on the ISS. Normal human peripheral blood mononuclear cells (lymphocytes) treated with 10 µ g/ml of KMEG together with untreated control samples were flown on Nanoracks hardware on Spacex-3 flight. The Samples were returned one month later and gene expression was analyzed. A 1g-ground control experiment was performed in parallel at the Kennedy spaceflight center. The first overall subtractive unrestricted analysis revealed 78 genes, which were differentially expressed in space flight KMEG, untreated lymphocytes as compared to the corresponding ground controls. However, in KMEG-treated space flight lymphocytes, there was an increased expression of a group of genes that mediate increased transcription, translation and innate immune system mediating functions of lymphocytes as compared to KMEG-untreated samples. Analysis of genes related to T cell proliferation in spaceflight KMEG-treated lymphocytes compared to 1g-ground KMEG- treated lymphocytes revealed six T cell proliferation and signaling genes to be significantly upregulated (p < 0.001) and five related genes were found to be significantly down-regulated. These genes play a significant role in

Regulated expression of telomerase activity in human T lymphocyte development and activation

PubMed Central

1996-01-01

Telomerase, a ribonucleoprotein that is capable of synthesizing telomeric repeats, is expressed in germline and malignant cells, and is absent in most normal human somatic cells. The selective expression of telomerase has thus been proposed to be a basis for the immortality of the germline and of malignant cells. In the present study, telomerase activity was analyzed in normal human T lymphocytes. It was found that telomerase is expressed at a high level in thymocyte subpopulations, at an intermediate level in tonsil T lymphocytes, and at a low to undetectable level in peripheral blood T lymphocytes. Moreover, telomerase activity is highly inducible in peripheral T lymphocytes by activation through CD3 with or without CD28 costimulation, or by stimulation with phorbol myristate acetate (PMA)/ionomycin. The induction of telomerase by anti-CD3 plus anti-CD28 (anti-CD3/CD28) stimulation required RNA and protein synthesis, and was blocked by herbimycin A, an inhibitor of S pi protein tyrosine kinases. The immunosuppressive drug cyclosporin A selectively inhibited telomerase induction by PMA/ionomycin and by anti-CD3, but not by anti-CD3/CD28. Although telomerase activity in peripheral T lymphocytes was activation dependent and correlated with cell proliferation, it was not cell cycle phase restricted. These results indicate that the expression of telomerase in normal human T lymphocytes is both developmentally regulated and activation induced. Telomerase may thus play a permissive role in T cell development and in determining the capacity of lymphoid cells for cell division and clonal expansion. PMID:8676067

5' Rapid Amplification of cDNA Ends and Illumina MiSeq Reveals B Cell Receptor Features in Healthy Adults, Adults With Chronic HIV-1 Infection, Cord Blood, and Humanized Mice.

PubMed

Waltari, Eric; Jia, Manxue; Jiang, Caroline S; Lu, Hong; Huang, Jing; Fernandez, Cristina; Finzi, Andrés; Kaufmann, Daniel E; Markowitz, Martin; Tsuji, Moriya; Wu, Xueling

2018-01-01

Using 5' rapid amplification of cDNA ends, Illumina MiSeq, and basic flow cytometry, we systematically analyzed the expressed B cell receptor (BCR) repertoire in 14 healthy adult PBMCs, 5 HIV-1+ adult PBMCs, 5 cord blood samples, and 3 HIS-CD4/B mice, examining the full-length variable region of μ, γ, α, κ, and λ chains for V-gene usage, somatic hypermutation (SHM), and CDR3 length. Adding to the known repertoire of healthy adults, Illumina MiSeq consistently detected small fractions of reads with high mutation frequencies including hypermutated μ reads, and reads with long CDR3s. Additionally, the less studied IgA repertoire displayed similar characteristics to that of IgG. Compared to healthy adults, the five HIV-1 chronically infected adults displayed elevated mutation frequencies for all μ, γ, α, κ, and λ chains examined and slightly longer CDR3 lengths for γ, α, and λ. To evaluate the reconstituted human BCR sequences in a humanized mouse model, we analyzed cord blood and HIS-CD4/B mice, which all lacked the typical SHM seen in the adult reference. Furthermore, MiSeq revealed identical unmutated IgM sequences derived from separate cell aliquots, thus for the first time demonstrating rare clonal members of unmutated IgM B cells by sequencing.

Humpback Dolphin (Genus Sousa) Behavioural Responses to Human Activities.

PubMed

Piwetz, Sarah; Lundquist, David; Würsig, Bernd

2015-01-01

Humpback dolphins (genus Sousa) use shallow, near-shore waters throughout their range. This coastal distribution makes them vulnerable to recreational and commercial disturbances, especially near heavily populated and industrialized areas. Most research focusing on Sousa and human activities has emphasized direct impacts and threats, involving injury and death, with relatively little focus on indirect effects on dolphins, such as changes in behaviour that may lead to deleterious effects. Understanding behaviour is important in resolving human-wildlife conflict and is an important component of conservation. This chapter gives an overview of animal behavioural responses to human activity with examples from diverse taxa; reviews the scientific literature on behavioural responses of humpback dolphins to human activity throughout their range, including marine vessel traffic, dolphin tourism, cetacean-fishery interactions, noise pollution, and habitat alteration; and highlights information and data gaps for future humpback dolphin research to better inform behaviour-based management decisions that contribute to conservation efforts. © 2015 Elsevier Ltd All rights reserved.

The Evolution of Lineage-Specific Regulatory Activities in the Human Embryonic Limb

PubMed Central

Cotney, Justin; Leng, Jing; Yin, Jun; Reilly, Steven K.; DeMare, Laura E.; Emera, Deena; Ayoub, Albert E.; Rakic, Pasko; Noonan, James P.

2013-01-01

SUMMARY The evolution of human anatomical features likely involved changes in gene regulation during development. However, the nature and extent of human-specific developmental regulatory functions remain unknown. We obtained a genome-wide view of cis-regulatory evolution in human embryonic tissues by comparing the histone modification H3K27ac, which provides a quantitative readout of promoter and enhancer activity, during human, rhesus, and mouse limb development. Based on increased H3K27ac, we find that 13% of promoters and 11% of enhancers have gained activity on the human lineage since the human-rhesus divergence. These gains largely arose by modification of ancestral regulatory activities in the limb or potential co-option from other tissues and are likely to have heterogeneous genetic causes. Most enhancers that exhibit gain of activity in humans originated in mammals. Gains at promoters and enhancers in the human limb are associated with increased gene expression, suggesting they include molecular drivers of human morphological evolution. PMID:23827682

Inhibition by sulfonamides of the candidacidal activity of human neutrophils.

PubMed

Lehrer, R I

1971-12-01

Sulfonamides reduced substantially the ability of normal human neutrophils to kill strains of Candida albicans and Candida tropicalis, and impaired to a lesser extent their activity against Staphylococcus aureus 502A and Serratia marcescens. Sulfonamides also inhibited (a) iodination of Candida cells by normal neutrophils; (b) candidacidal activity in cell-free systems containing purified human myeloperoxidase, hydrogen peroxide, and potassium iodide; and (c) accumulation of molecular iodine in analogously constructed cell-free systems. In contrast to these effects on reactions catalyzed by myeloperoxidase, sulfonamides exerted relatively little effect on the levels of microbicidal activity manifested by human neutrophils that lacked myeloperoxidase. Sulfonamides appear to influence the function of human neutrophils predominantly by interfering with myeloperoxidase-mediated pathways. Certain basic and clinical implications of these data are discussed.

Selective induction of apoptosis in human gastric cancer cells by Lactobacillus kefiri (PFT), a novel kefir product

PubMed Central

GHONEUM, MAMDOOH; FELO, NOURAN

2015-01-01

The present study was undertaken to evaluate the effect of Lactobacillus kefiri (PFT), a novel kefir product, on apoptosis of gastric cancer cells (AGS), breast cancer cells (4T1), and human peripheral blood mononuclear cells (PBMCs). Cells were cultured with PFT and apoptosis was determined by flow cytometry using 7-AAD dye and cytospin preparation. Mitochondrial dysfunction and expression of Bcl2 were monitored by flow cytometry. Results showed that PFT induced apoptosis in AGS gastric cancer cells in a dose-dependent manner. Apoptosis was detected at a concentration of 0.3 mg/ml (20.8%), increased to 25.8% at 0.6 mg/ml, 37% at 1.2 mg/ml, 53.1% at 2.5 mg/ml, and peaked at 66.3% at 5.0 mg/ml. Apoptosis is associated with the decreased polarization of mitochondrial membrane potential (MMP) and decreased Bcl2 expression. PFT-treated AGS cells manifested membrane blebbing, nuclear condensation, and fragmentation as identified in cytospin cytocentrifuge Giemsa stained preparations. On the other hand, flow cytometry analysis showed that PFT did not induce apoptosis in 4T1 breast cancer cells nor in PBMCs. These results suggest that PFT is safe for white blood cells and selectively induces apoptotic effects in gastric cancer cells. Hence, it may have potential as a therapeutic agent for the treatment of gastric cancers. PMID:26251956

Selective induction of apoptosis in human gastric cancer cells by Lactobacillus kefiri (PFT), a novel kefir product.

PubMed

Ghoneum, Mamdooh; Felo, Nouran

2015-10-01

The present study was undertaken to evaluate the effect of Lactobacillus kefiri (PFT), a novel kefir product, on apoptosis of gastric cancer cells (AGS), breast cancer cells (4T1), and human peripheral blood mononuclear cells (PBMCs). Cells were cultured with PFT and apoptosis was determined by flow cytometry using 7-AAD dye and cytospin preparation. Mitochondrial dysfunction and expression of Bcl2 were monitored by flow cytometry. Results showed that PFT induced apoptosis in AGS gastric cancer cells in a dose-dependent manner. Apoptosis was detected at a concentration of 0.3 mg/ml (20.8%), increased to 25.8% at 0.6 mg/ml, 37% at 1.2 mg/ml, 53.1% at 2.5 mg/ml, and peaked at 66.3% at 5.0 mg/ml. Apoptosis is associated with the decreased polarization of mitochondrial membrane potential (MMP) and decreased Bcl2 expression. PFT-treated AGS cells manifested membrane blebbing, nuclear condensation, and fragmentation as identified in cytospin cytocentrifuge Giemsa stained preparations. On the other hand, flow cytometry analysis showed that PFT did not induce apoptosis in 4T1 breast cancer cells nor in PBMCs. These results suggest that PFT is safe for white blood cells and selectively induces apoptotic effects in gastric cancer cells. Hence, it may have potential as a therapeutic agent for the treatment of gastric cancers.

A novel cyclohexene derivative, ethyl (6R)-6-[N-(2-Chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (TAK-242), selectively inhibits toll-like receptor 4-mediated cytokine production through suppression of intracellular signaling.

PubMed

Ii, Masayuki; Matsunaga, Naoko; Hazeki, Kaoru; Nakamura, Kazuyo; Takashima, Katsunori; Seya, Tsukasa; Hazeki, Osamu; Kitazaki, Tomoyuki; Iizawa, Yuji

2006-04-01

Proinflammatory mediators such as cytokines and NO play pivotal roles in various inflammatory diseases. To combat inflammatory diseases successfully, regulation of proinflammatory mediator production would be a critical process. In the present study, we investigated the in vitro effects of ethyl (6R)-6-[N-(2-chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (TAK-242), a novel small molecule cytokine production inhibitor, and its mechanism of action. In RAW264.7 cells and mouse peritoneal macrophages, TAK-242 suppressed lipopolysaccharide (LPS)-induced production of NO, tumor necrosis factor-alpha (TNF-alpha), and interleukin (IL)-6, with 50% inhibitory concentration (IC50) of 1.1 to 11 nM. TAK-242 also suppressed the production of these cytokines from LPS-stimulated human peripheral blood mononuclear cells (PBMCs) at IC50 values from 11 to 33 nM. In addition, the inhibitory effects on the LPS-induced IL-6 and IL-12 production were similar in human PBMCs, monocytes, and macrophages. TAK-242 inhibited mRNA expression of IL-6 and TNF-alpha induced by LPS and interferon-gamma in RAW264.7 cells. The phosphorylation of mitogen-activated protein kinases induced by LPS was also inhibited in a concentration-dependent manner. However, TAK-242 did not antagonize the binding of LPS to the cells. It is noteworthy that TAK-242 suppressed the cytokine production induced by Toll-like receptor (TLR) 4 ligands, but not by ligands for TLR2, -3, and -9. In addition, IL-1beta-induced IL-8 production from human PBMCs was not markedly affected by TAK-242. These data suggest that TAK-242 suppresses the production of multiple cytokines by selectively inhibiting TLR4 intracellular signaling. Finally, TAK-242 is a novel small molecule TLR4 signaling inhibitor and could be a promising therapeutic agent for inflammatory diseases, whose pathogenesis involves TLR4.

Human Activity Recognition from Body Sensor Data using Deep Learning.

PubMed

Hassan, Mohammad Mehedi; Huda, Shamsul; Uddin, Md Zia; Almogren, Ahmad; Alrubaian, Majed

2018-04-16

In recent years, human activity recognition from body sensor data or wearable sensor data has become a considerable research attention from academia and health industry. This research can be useful for various e-health applications such as monitoring elderly and physical impaired people at Smart home to improve their rehabilitation processes. However, it is not easy to accurately and automatically recognize physical human activity through wearable sensors due to the complexity and variety of body activities. In this paper, we address the human activity recognition problem as a classification problem using wearable body sensor data. In particular, we propose to utilize a Deep Belief Network (DBN) model for successful human activity recognition. First, we extract the important initial features from the raw body sensor data. Then, a kernel principal component analysis (KPCA) and linear discriminant analysis (LDA) are performed to further process the features and make them more robust to be useful for fast activity recognition. Finally, the DBN is trained by these features. Various experiments were performed on a real-world wearable sensor dataset to verify the effectiveness of the deep learning algorithm. The results show that the proposed DBN outperformed other algorithms and achieves satisfactory activity recognition performance.

Characterization of Acid Sphingomyelinase Activity in Human Cerebrospinal Fluid

PubMed Central

Mühle, Christiane; Huttner, Hagen B.; Walter, Silke; Reichel, Martin; Canneva, Fabio; Lewczuk, Piotr; Gulbins, Erich; Kornhuber, Johannes

2013-01-01

Background As a key enzyme in sphingolipid metabolism, acid sphingomyelinase (ASM) is involved in the regulation of cell fate and signaling via hydrolysis of sphingomyelin to form ceramide. While increased activity of the lysosomal form has been associated with various pathological conditions, there are few studies on secretory ASM limited only to cell models, plasma or serum. Methods An optimized assay based on a fluorescent substrate was applied to measure the ASM activity in cerebrospinal fluid (CSF) collected from mice and from 42 patients who were classified as controls based on normal routine CSF values. Results We have detected ASM activity in human CSF, established a sensitive quantitative assay and characterized the enzyme’s properties. The enzyme resembles plasmatic ASM including protein stability and Zn2+-dependence but the assays differ considerably in the optimal detergent concentration. Significantly increased activities in the CSF of ASM transgenic mice and undetectable levels in ASM knock-out mice prove that the measured ASM activity originates from the ASM-encoding gene SMPD1. CSF localized ASM activities were comparable to corresponding serum ASM levels at their respective optimal reaction conditions, but no correlation was observed. The large variance in ASM activity was independent of sex, age or analyzed routine CSF parameters. Conclusions Human and mouse CSF contain detectable levels of secretory ASM, which are unrelated to serum ASM activities. Further investigations in humans and in animal models will help to elucidate the role of this enzyme in human disease and to assess its value as a potential biomarker for disease type, severity, progress or therapeutic success. PMID:23658784

Dasatinib is preclinically active against Src-overexpressing human transitional cell carcinoma of the urothelium with activated Src signaling.

PubMed

Levitt, Jonathan M; Yamashita, Hideyuki; Jian, Weiguo; Lerner, Seth P; Sonpavde, Guru

2010-05-01

Dasatinib is an orally administered multitargeted kinase inhibitor that targets Src family tyrosine kinases, Abl, c-Kit, and PDGFR. A preclinical study was conducted to evaluate dasatinib alone or combined with cisplatin for human transitional cell carcinoma (TCC). Expression of Src in a human TCC tissue microarray was evaluated by immunohistochemistry. The activity of dasatinib and/or cisplatin was evaluated in six human TCC cell lines. Western blot was done to assess Src and phosphorylated-Src (p-Src) expression. The activity of dasatinib alone and in combination with cisplatin was determined in murine subcutaneous xenografts. Sixty-two percent to 75% of human TCC expressed Src. Dasatinib displayed significant antiproliferative activity at nanomolar concentrations against two human TCC cell lines (RT4 and Hu456) that exhibited high Src and p-Src expression and were cisplatin-resistant. RT4 cells were the most sensitive and displayed the highest level of Src pathway activation (p-Src/Src ratio). Dasatinib downregulated p-Src in either sensitive or resistant cells. TCC cells that were sensitive to cisplatin (5637 and TCC-SUP) were highly resistant to dasatinib and exhibited low Src expression. Dasatinib showed antitumor activity in RT4 murine xenografts, and the combination of dasatinib and cisplatin was significantly more active than placebo. Combination dasatinib plus cisplatin significantly inhibited proliferation and promoted apoptosis in vivo. In conclusion, dasatinib displayed significant preclinical antitumor activity against Src-overexpressing human TCC with active Src signaling and was highly active in combination with cisplatin in vivo. Further clinical development might be warranted in selected human subjects.

Systems Approach to Understanding Electromechanical Activity in the Human Heart

PubMed Central

Rudy, Yoram; Ackerman, Michael J.; Bers, Donald M.; Clancy, Colleen E.; Houser, Steven R.; London, Barry; McCulloch, Andrew D.; Przywara, Dennis A.; Rasmusson, Randall L.; Solaro, R. John; Trayanova, Natalia A.; Van Wagoner, David R.; Varró, András; Weiss, James N.; Lathrop, David A.

2010-01-01

The National Heart, Lung, and Blood Institute (NHLBI) convened a workshop of cardiologists, cardiac electrophysiologists, cell biophysicists, and computational modelers on August 20 and 21, 2007, in Washington, DC, to advise the NHLBI on new research directions needed to develop integrative approaches to elucidate human cardiac function. The workshop strove to identify limitations in the use of data from nonhuman animal species for elucidation of human electromechanical function/activity and to identify what specific information on ion channel kinetics, calcium handling, and dynamic changes in the intracellular/extracellular milieu is needed from human cardiac tissues to develop more robust computational models of human cardiac electromechanical activity. This article summarizes the workshop discussions and recommendations on the following topics: (1) limitations of animal models and differences from human electrophysiology, (2) modeling ion channel structure/function in the context of whole-cell electrophysiology, (3) excitation–contraction coupling and regulatory pathways, (4) whole-heart simulations of human electromechanical activity, and (5) what human data are currently needed and how to obtain them. The recommendations can be found on the NHLBI Web site at http://www.nhlbi.nih.gov/meetings/workshops/electro.htm. PMID:18779456

Chronic active Epstein-Barr virus infection associated with mutations in perforin that impair its maturation.

PubMed

Katano, Harutaka; Ali, Mir A; Patera, Andriani C; Catalfamo, Marta; Jaffe, Elaine S; Kimura, Hiroshi; Dale, Janet K; Straus, Stephen E; Cohen, Jeffrey I

2004-02-15

Chronic active Epstein-Barr virus infection (CAEBV) is a rare disease in which previously healthy persons develop severe, life-threatening illness. Mutations in the perforin gene have been found in familial hemophagocytic lymphohistiocytosis, which shares some features with CAEBV. We studied a patient who died at age 18, 10 years after the onset of CAEBV. The patient had high titers of antibodies to EBV, EBV RNA in lymph nodes, T-cell lymphoproliferative disease, and hemophagocytic lymphohistiocytosis. DNA sequencing showed novel mutations in both alleles of the perforin gene that resulted in amino acid changes in the protein. The quantity of the native form of perforin from the patient's stimulated peripheral blood mononuclear cells (PBMCs) was extremely low and immunoblotting showed accumulation of an uncleaved precursor form of perforin. Stimulated PBMCs from the patient were defective for Fas-independent cytotoxicity. These data imply that mutations in this patient resulted in reduced perforin-mediated cytotoxicity by his lymphocytes. This is the first case in which perforin mutations have been shown to result in accumulation of the uncleaved, immature form of perforin. Mutations in the perforin gene are associated with some cases of CAEBV with hemophagocytic lymphohistiocytosis.

3D Data Acquisition Platform for Human Activity Understanding

DTIC Science & Technology

2016-03-02

address fundamental research problems of representation and invariant description of3D data, human motion modeling and applications of human activity analysis, and computational optimization of large-scale 3D data.

3D Data Acquisition Platform for Human Activity Understanding

DTIC Science & Technology

2016-03-02

address fundamental research problems of representation and invariant description of 3D data, human motion modeling and applications of human activity analysis, and computational optimization of large-scale 3D data.

MEASURING CHOLINESTERASE ACTIVITY IN HUMAN SALIVA

EPA Science Inventory

To assess the potential for using saliva in pesticide biomonitoring, the consistency of cholinesterase activity in human saliva collected over time was examined. In this pilot study, saliva was collected from 20 healthy adults once per week for 5 consecutive weeks using 2 differe...

MEASURING CHOLINESTERASE ACTIVITY IN HUMAN SALIVA.

EPA Science Inventory

To assess the potential for using saliva in pesticide biomonitoring, the consistency of cholinesterase activity in human saliva collected over time was examined. In this pilot study, saliva was collected from 20 healthy adults once per week for 5 consecutive weeks using 2 differe...

The Activation Pathway of Human Rhodopsin in Comparison to Bovine Rhodopsin*

PubMed Central

Kazmin, Roman; Rose, Alexander; Szczepek, Michal; Elgeti, Matthias; Ritter, Eglof; Piechnick, Ronny; Hofmann, Klaus Peter; Scheerer, Patrick; Hildebrand, Peter W.; Bartl, Franz J.

2015-01-01

Rhodopsin, the photoreceptor of rod cells, absorbs light to mediate the first step of vision by activating the G protein transducin (Gt). Several human diseases, such as retinitis pigmentosa or congenital night blindness, are linked to rhodopsin malfunctions. Most of the corresponding in vivo studies and structure-function analyses (e.g. based on protein x-ray crystallography or spectroscopy) have been carried out on murine or bovine rhodopsin. Because these rhodopsins differ at several amino acid positions from human rhodopsin, we conducted a comprehensive spectroscopic characterization of human rhodopsin in combination with molecular dynamics simulations. We show by FTIR and UV-visible difference spectroscopy that the light-induced transformations of the early photointermediates are very similar. Significant differences between the pigments appear with formation of the still inactive Meta I state and the transition to active Meta II. However, the conformation of Meta II and its activity toward the G protein are essentially the same, presumably reflecting the evolutionary pressure under which the active state has developed. Altogether, our results show that although the basic activation pathways of human and bovine rhodopsin are similar, structural deviations exist in the inactive conformation and during receptor activation, even between closely related rhodopsins. These differences between the well studied bovine or murine rhodopsins and human rhodopsin have to be taken into account when the influence of point mutations on the activation pathway of human rhodopsin are investigated using the bovine or murine rhodopsin template sequences. PMID:26105054

Human brain activity with functional NIR optical imager

NASA Astrophysics Data System (ADS)

Luo, Qingming

2001-08-01

In this paper we reviewed the applications of functional near infrared optical imager in human brain activity. Optical imaging results of brain activity, including memory for new association, emotional thinking, mental arithmetic, pattern recognition ' where's Waldo?, occipital cortex in visual stimulation, and motor cortex in finger tapping, are demonstrated. It is shown that the NIR optical method opens up new fields of study of the human population, in adults under conditions of simulated or real stress that may have important effects upon functional performance. It makes practical and affordable for large populations the complex technology of measuring brain function. It is portable and low cost. In cognitive tasks subjects could report orally. The temporal resolution could be millisecond or less in theory. NIR method will have good prospects in exploring human brain secret.

Herpes simplex virus antigens directly activate NK cells via TLR2, thus facilitating their presentation to CD4 T lymphocytes.

PubMed

Kim, Min; Osborne, Naomi R; Zeng, Weiguang; Donaghy, Heather; McKinnon, Kay; Jackson, David C; Cunningham, Anthony L

2012-05-01

NK cells infiltrate human herpetic lesions, but their role has been underexplored. HSV can stimulate innate immune responses via surface TLR2, which is expressed on monocyte-derived dendritic cells (DCs) and NK cells. In this study, UV-inactivated HSV1/2 and immunodominant HSV2 glycoprotein D peptides conjugated to the TLR2 agonist dipalmitoyl-S-glyceryl cysteine stimulated CD4 T lymphocyte IFN-γ responses within PBMCs or in coculture with monocyte-derived DCs. NK cells contributed markedly to the PBMC responses. Furthermore, NK cells alone were activated directly by both Ags, also upregulating HLA-DR and HLA-DQ and then they activated autologous CD4 T lymphocytes. Using Transwells, Ag-stimulated NK cells and CD4 T lymphocytes were shown to interact through both cell-to-cell contact and cytokines, differing in relative importance in different donors. A distinct immunological synapse between Ag-stimulated NK cells and CD4 T lymphocytes was observed, indicating the significance of their cell-to-cell contact. A large proportion (57%) of NK cells was also in contact with CD4 T lymphocytes in the dermal infiltrate of human recurrent herpetic lesions. Thus, NK cells stimulated by TLR2-activating HSV Ags can present Ag alone or augment the role of DCs in vitro and perhaps in herpetic lesions or draining lymph nodes. In addition to DCs, NK cells should be considered as targets for adjuvants during HSV vaccine development.

Global microRNA profiling of peripheral blood mononuclear cells in patients with Behçet's disease.

PubMed

Erre, Gian Luca; Piga, Matteo; Carru, Ciriaco; Angius, Andrea; Carcangiu, Laura; Piras, Marco; Sotgia, Salvatore; Zinellu, Angelo; Mathieu, Alessandro; Passiu, Giuseppe; Pescatori, Mario

2015-01-01

To explore the post-transcriptional regulation of the peripheral blood mononuclear cells (PBMCs) transcriptome by microRNAs in Behçet's disease (BD). Using TaqMan Low Density Array-based microRNAs expression profiling, the expression of 750 mature human microRNAs in PBMCs from 5 BD patients and 3 healthy controls (HC) was compared. The expression of deregulated microRNAs was then validated by quantitative real time-polymerase chain reaction (qRT-PCR), in 42 BD patients and 8 HC. In the initial screening, 13 microRNAs appeared deregulated in BD vs HC. Among them, the differential expression of miR-720 and miR-139-3p was confirmed by qRT-PCR, (p<0.05 and FDR<5%). Areas under the receiver operating characteristic curve for miR-139-3p, miR-720 and miR-139-3p+miR-720 in the validation cohort were 0.84, 0.87 and 0.92 respectively, indicating good discrimination between BD patients and HC. Post-hoc analysis showed that 9 out of 13 microRNAs from the discovery phase were significantly upregulated in active vs. quiescent BD, suggesting inflammation as a key regulator of microRNAs machinery in BD. In silico analysis revealed that several BD candidate susceptibility genes are predicted target of significantly deregulated microRNAs in active BD. A significant enrichment in microRNAs targeting elements of the Toll-like receptor (TLR) and T-cell receptor signalling pathways was also assumed. miR199-3p and miR720 deserve further confirmation as biomarkers of BD in larger studies. PBMCs from active BD displayed a unique signature of microRNAs which may be implicated in regulation of innate immunity activation and T-cell function.

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

Kawamoto, Eiji; Emergency and Critical Care Center, Mie University Hospital, 2-174 Edobashi, Tsu 514-8507; Okamoto, Takayuki, E-mail: okamotot@doc.medic.mie-u.ac.jp

LFA-1 (αLβ2) and Mac-1 (αMβ2) integrins regulate leukocyte trafficking in health and disease by binding primarily to IgSF ligand ICAM-1 and ICAM-2 on endothelial cells. Here we have shown that the anti-coagulant molecule thrombomodulin (TM), found on the surface of endothelial cells, functions as a potentially new ligand for leukocyte integrins. We generated a recombinant extracellular domain of human TM and Fc fusion protein (TM-domains 123-Fc), and showed that pheripheral blood mononuclear cells (PBMCs) bind to TM-domains 123-Fc dependent upon integrin activation. We then demonstrated that αL integrin-blocking mAb, αM integrin-blocking mAb, and β2 integrin-blocking mAb inhibited the binding ofmore » PBMCs to TM-domains 123-Fc. Furthermore, we show that the serine/threonine-rich domain (domain 3) of TM is required for the interaction with the LFA-1 (αLβ2) and Mac-1 (αMβ2) integrins to occur on PBMCs. These results demonstrate that the LFA-1 and Mac-1 integrins on leukocytes bind to TM, thereby establishing the molecular and structural basis underlying LFA-1 and Mac-1 integrin interaction with TM on endothelial cells. In fact, integrin-TM interactions might be involved in the dynamic regulation of leukocyte adhesion with endothelial cells. - Highlights: • LFA-1 and Mac-1 integrins bind to the anti-coagulant molecule thrombomodulin. • The serine/threonine-rich domain of thrombomodulin is essential to interact with the LFA-1 and Mac-1 integrins on PBMCs. • Integrin-TM interactions might be involved in the dynamic regulation of leukocyte adhesion with endothelial cells.« less

Human Relations Education; A Guidebook to Learning Activities.

ERIC Educational Resources Information Center

Buffalo Public Schools, NY. Human Relations Project of Western New York.

This guidebook is designed to acquaint teachers with human relations classroom materials, extracurricular activities, and an inservice approach to self-evaluation. A product of an ESEA Title III program, it contains human relations-oriented lessons--divided by grade level and subject matter--intended to supplement or enrich existing curricula and…

[gammadelta T cells stimulated by zoledronate kill osteosarcoma cells].

PubMed

Jiang, Hui; Xu, Qiang; Yang, Chao; Cao, Zhen-Guo; Li, Zhao-Xu; Ye, Zhao-Ming

2010-12-01

To investigate the cytotoxicity of human γδT cells from PBMCs stimulated by zoledronate against osteosarcoma cell line HOS in vitro and in vivo and evaluate the relavent pathways. The peripheral blood mononuclear cells (PBMCs)of healthy donors were stimulated by single dose zoledronate and cultured in the present of IL-2 for two weeks, analysising the percentage of γδT cells on a FACSCalibur cytometer.Study the cytotoxicity of γδT cells against the osteosarcoma line HOS using LDH release assay kit. Pre-treatment of γδT cells with anti-human γδTCR antibody, anti-human NKG2D antibody and concanamycin A to bolck the relavent pathways for evaluating the mechenisms of its cytotoxicity. In vivo, BALB/c mice were inoculated subcutaneously osteosarcoma cell HOS for developing hypodermal tumors. And they were randomized into two groups: unteated group, γδT cell therapy group. Tumor volume and weight of the two groups were compared. After two weeks of culture, γδT cells from zoledronate-stimulated PBMCs could reach (95±3)%. When the E:T as 6:1, 12:1, 25:1, 50:1, the percentage of osteosarcoma cell HOS killed by γδT cells was 26.8%, 31.5%, 37.8%, 40.9%, respectively.When anti-huma γδTCR antibody, anti-human NKG2D antibody and concanamycin A blocked the relavent pathways, the percentage was 32.3%, 4.7%, 16.7% ( E:T as 25:1), respectively. In vivo, the tumor inhibition rate of the group of γδT cell therapy was 42.78%. γδT cells derived from PBMCs stimulated by zoledronate can acquired pure γδT cells. And they show strong cytoxicity against osteosarcoma cell line HOS in vitro and in vivo.

An Ahemolytic Pneumolysin of Streptococcus Pneumoniae Manipulates Human Innate and CD4+ T-Cell Responses and Reduces Resistance to Colonization in Mice in a Serotype-Independent Manner

PubMed Central

Khan, M. Nadeem; Coleman, John Robert; Vernatter, Joshua; Varshney, Avanish Kumar; Dufaud, Chad; Pirofski, Liise-anne

2014-01-01

Background. Some Streptococcus pneumoniae serotypes express an ahemolytic pneumolysin (PLYa). Serotypes that commonly express PLYa, including serotype 8 (ST8) and ST1, are often associated with a low prevalence during colonization but a higher propensity to cause invasive disease. We sought to study the host response to ST8 PLYa in a homologous and heterologous capsular background. Methods. We genetically exchanged the PLYa of ST8 strain 6308 with the hemolytic PLY (PLYh) of ST3 A66.1 and vice versa and determined the impact of the exchange on nasopharyngeal colonization in mice. Then, to compare the response of human cells to PLYa-expressing and PLYh-expressing strains, we infected human peripheral blood mononuclear cells (PBMCs) with PLY-switched strains and assessed dendritic cell and CD4+ T-cell responses by intracellular cytokine staining. Result. Mice colonized with PLYa-expressing strains had significantly higher colonization densities than those colonized with PLYh-expressing strains, irrespective of capsular background. Compared with infection of PBMCs with PLYh-expressing strains, infection with PLYa-expressing strains induced diminished innate (dendritic cell cytokines, costimulatory receptor, and apoptotic) and adaptive (CD4+ T-cell proliferative and memory interleukin 17A) responses. Conclusion. Our findings demonstrate that PLYa has the potential to manipulate host immunity irrespective of capsule type. PLY exchange between STs expressing PLYa and PLYh could lead to unexpected colonization or invasion phenotypes. PMID:25001458

Activation of human B cells by phosphorothioate oligodeoxynucleotides.

PubMed Central

Liang, H; Nishioka, Y; Reich, C F; Pisetsky, D S; Lipsky, P E

1996-01-01

To investigate the potential of DNA to elicit immune responses in man, we examined the capacity of a variety of oligodeoxynucleotides (ODNs) to stimulate highly purified T cell-depleted human peripheral blood B cells. Among 47 ODNs of various sequences tested, 12 phosphorothioate oligodeoxynucleotides (sODNs) induced marked B cell proliferation and Ig production. IL-2 augmented both proliferation and production of IgM, IgG, and IgA, as well as IgM anti-DNA antibodies, but was not necessary for B cell stimulation. Similarly, T cells enhanced stimulation, but were not necessary for B cell activation. After stimulation with the active sODNs, more than 95% of B cells expressed CD25 and CD86. In addition, B cells stimulated with sODNs expressed all six of the major immunoglobulin VH gene families. These results indicate that the human B cell response to sODN is polyclonal. Active sODN coupled to Sepharose beads stimulated B cells as effectively as the free sODN, suggesting that stimulation resulted from engagement of surface receptors. These data indicate that sODNs can directly induce polyclonal activation of human B cells in a T cell-independent manner by engaging as yet unknown B cell surface receptors. PMID:8787674

S100A8 and S100A9 Induce Cytokine Expression and Regulate the NLRP3 Inflammasome via ROS-Dependent Activation of NF-κB1

PubMed Central

Simard, Jean-Christophe; Cesaro, Annabelle; Chapeton-Montes, Julie; Tardif, Mélanie; Antoine, Francis; Girard, Denis; Tessier, Philippe A.

2013-01-01

S100A8 and S100A9 are cytoplasmic proteins expressed by phagocytes. High concentrations of these proteins have been correlated with various inflammatory conditions, including autoimmune diseases such as rheumatoid arthritis and Crohn’s disease, as well as autoinflammatory diseases. In the present study, we examined the effects of S100A8 and S100A9 on the secretion of cytokines and chemokines from PBMCs. S100A8 and S100A9 induced the secretion of cytokines such as IL-6, IL-8, and IL-1β. This secretion was associated with the activation and translocation of the transcription factor NF-κB. Inhibition studies using antisense RNA and the pharmacological agent BAY-117082 confirmed the involvement of NF-κB in IL-6, IL-8, and IL-1β secretion. S100A8- and S100A9-mediated activation of NF-κB, the NLR family, pyrin domain-containing 3 (NLRP3) protein, and pro-IL-1β expression was dependent on the generation of reactive oxygen species. This effect was synergistically enhanced by ATP, a known inflammasome activator. These results suggest that S100A8 and S100A9 enhance the inflammatory response by inducing cytokine secretion of PBMCs. PMID:23977231

Detecting Human Activity Using Acoustic, Seismic, Accelerometer, Video, and E-field Sensors

DTIC Science & Technology

2011-09-01

Detecting Human Activity using Acoustic, Seismic, Accelerometer, Video, and E-field Sensors by Sarah H. Walker and Geoffrey H. Goldman...Adelphi, MD 20783-1197 ARL-TR-5729 September 2011 Detecting Human Activity using Acoustic, Seismic, Accelerometer, Video, and E-field Sensors...DD-MM-YYYY) September 2011 2. REPORT TYPE 3. DATES COVERED (From - To) 4. TITLE AND SUBTITLE Detecting Human Activity using Acoustic

A Multiscale Survival Process for Modeling Human Activity Patterns.

PubMed

Zhang, Tianyang; Cui, Peng; Song, Chaoming; Zhu, Wenwu; Yang, Shiqiang

2016-01-01

Human activity plays a central role in understanding large-scale social dynamics. It is well documented that individual activity pattern follows bursty dynamics characterized by heavy-tailed interevent time distributions. Here we study a large-scale online chatting dataset consisting of 5,549,570 users, finding that individual activity pattern varies with timescales whereas existing models only approximate empirical observations within a limited timescale. We propose a novel approach that models the intensity rate of an individual triggering an activity. We demonstrate that the model precisely captures corresponding human dynamics across multiple timescales over five orders of magnitudes. Our model also allows extracting the population heterogeneity of activity patterns, characterized by a set of individual-specific ingredients. Integrating our approach with social interactions leads to a wide range of implications.

Fatty acid synthase inhibition activates AMP-activated protein kinase in SKOV3 human ovarian cancer cells.

PubMed

Zhou, Weibo; Han, Wan Fang; Landree, Leslie E; Thupari, Jagan N; Pinn, Michael L; Bililign, Tsion; Kim, Eun Kyoung; Vadlamudi, Aravinda; Medghalchi, Susan M; El Meskini, Rajaa; Ronnett, Gabriele V; Townsend, Craig A; Kuhajda, Francis P

2007-04-01

Fatty acid synthase (FAS), the enzyme responsible for the de novo synthesis of fatty acids, is highly expressed in ovarian cancers and most common human carcinomas. Inhibition of FAS and activation of AMP-activated protein kinase (AMPK) have been shown to be cytotoxic to human cancer cells in vitro and in vivo. In this report, we explore the cytotoxic mechanism of action of FAS inhibition and show that C93, a synthetic FAS inhibitor, increases the AMP/ATP ratio, activating AMPK in SKOV3 human ovarian cancer cells, which leads to cytotoxicity. As a physiologic consequence of AMPK activation, acetyl-CoA carboxylase (ACC), the rate-limiting enzyme of fatty acid synthesis, was phosphorylated and inhibited whereas glucose oxidation was increased. Despite these attempts to conserve energy, the AMP/ATP ratio increased with worsening cellular redox status. Pretreatment of SKOV3 cells with compound C, an AMPK inhibitor, substantially rescued the cells from C93 cytotoxicity, indicating its dependence on AMPK activation. 5-(Tetradecyloxy)-2-furoic acid, an ACC inhibitor, did not activate AMPK despite inhibiting fatty acid synthesis pathway activity and was not significantly cytotoxic to SKOV3 cells. This indicates that substrate accumulation from FAS inhibition triggering AMPK activation, not end-product depletion of fatty acids, is likely responsible for AMPK activation. C93 also exhibited significant antitumor activity and apoptosis against SKOV3 xenografts in athymic mice without significant weight loss or cytotoxicity to proliferating cellular compartments such as bone marrow, gastrointestinal tract, or skin. Thus, pharmacologic FAS inhibition selectively activates AMPK in ovarian cancer cells, inducing cytotoxicity while sparing most normal human tissues from the pleiotropic effects of AMPK activation.

The Human Activity of Evaluation Theorizing.

ERIC Educational Resources Information Center

Alkin, Marvin C.; Ellett, Frederick, Jr.

Theorizing about evaluation should be conceptualized as a human activity governed by certain strategies and principles. The theories advanced by various evaluators have changed over the years, thus illustrating ten principles of evaluation. The starting point for theory development or modification is self-reflection and review of one's own…

A Novel Wearable Sensor-Based Human Activity Recognition Approach Using Artificial Hydrocarbon Networks.

PubMed

Ponce, Hiram; Martínez-Villaseñor, María de Lourdes; Miralles-Pechuán, Luis

2016-07-05

Human activity recognition has gained more interest in several research communities given that understanding user activities and behavior helps to deliver proactive and personalized services. There are many examples of health systems improved by human activity recognition. Nevertheless, the human activity recognition classification process is not an easy task. Different types of noise in wearable sensors data frequently hamper the human activity recognition classification process. In order to develop a successful activity recognition system, it is necessary to use stable and robust machine learning techniques capable of dealing with noisy data. In this paper, we presented the artificial hydrocarbon networks (AHN) technique to the human activity recognition community. Our artificial hydrocarbon networks novel approach is suitable for physical activity recognition, noise tolerance of corrupted data sensors and robust in terms of different issues on data sensors. We proved that the AHN classifier is very competitive for physical activity recognition and is very robust in comparison with other well-known machine learning methods.

Exposures to the environmental toxicants pentachlorophenol (PCP) and dichlorodiphenyltrichloroethane (DDT) modify secretion of interleukin 1-beta (IL-1β) from human immune cells.

PubMed

Martin, Tamara J; Whalen, Margaret M

2017-04-01

Pentachlorophenol (PCP) and Dichlorodiphenyltrichloroethane (DDT) are environmental contaminants found in human blood. Previous studies have shown that PCP and DDT inhibit the lytic function of highly purified human natural killer (NK) lymphocytes and decrease the expression of several surface proteins on NK cells. Interleukin-1 βeta (IL-1β) is a cytokine produced by lymphocytes and monocytes, and anything that elevates its levels inappropriately can lead to chronic inflammation, which among other consequences can increase tumor development and invasiveness. Here, PCP and DDT were examined for their ability to alter secretion of IL-1β from immune cell preparations of various complexity: NK cells; monocyte-depleted (MD) peripheral blood mononuclear cells (PBMCS); and PBMCs. Cells were exposed to concentrations of PCP ranging from 5 to 0.05 µM and DDT concentrations of 2.5-0.025 μM for 24, 48 h, and 6 days. Results showed that both PCP and DDT increased IL-1β secretion from all of the immune cell preparations. The specific concentrations of PCP and DDT that increased IL-1β secretion varied by donor. Immune cells from all donors showed compound-induced increases in IL-1β secretion at one or more concentration at one or more length of exposure. The mechanism of PCP stimulation of IL1-β secretion was also addressed, and it appears that the MAPKs, ERK1/2 and p38, may be utilized by PCP to stimulate secretion of IL-1β.

Human activity and the spread of Phytophthora ramorum

Treesearch

Hall J. Cushman; Michelle Cooper; Ross K. Meentemeyer; Shelly Benson

2008-01-01

Increasing numbers of studies are finding that humans can facilitate the spread of exotic plant species in protected wildlands. Hiking trails commonly serve as conduits for invaders and the number of exotic plant species occurring in protected areas is often correlated positively with visitation rates. Despite such evidence linking human activity to the spread of...

Comparison of Human Immunodeficiency Virus Type 1 Tropism Profiles in Clinical Samples by the Trofile and MT-2 Assays▿

PubMed Central

Coakley, Eoin; Reeves, Jacqueline D.; Huang, Wei; Mangas-Ruiz, Marga; Maurer, Irma; Harskamp, Agnes M.; Gupta, Soumi; Lie, Yolanda; Petropoulos, Christos J.; Schuitemaker, Hanneke; van 't Wout, Angélique B.

2009-01-01

The recent availability of CCR5 antagonists as anti-human immunodeficiency virus (anti-HIV) therapeutics has highlighted the need to accurately identify CXCR4-using variants in patient samples when use of this new drug class is considered. The Trofile assay (Monogram Biosciences) has become the method that is the most widely used to define tropism in the clinic prior to the use of a CCR5 antagonist. By comparison, the MT-2 assay has been used since early in the HIV epidemic to define tropism in clinical specimens. Given that there are few data from direct comparisons of these two assays, we evaluated the performance of the plasma-based Trofile assay and the peripheral blood mononuclear cell (PBMC)-based MT-2 assay for the detection of CXCR4 use in defining the tropism of HIV isolates derived from clinical samples. The various samples used for this comparison were derived from participants of the Amsterdam Cohort Studies on HIV infection and AIDS who underwent consecutive MT-2 assay testing of their PBMCs at approximately 3-month intervals. This unique sample set was specifically selected because consecutive MT-2 assays had demonstrated a shift from negative to positive in PBMCs, reflecting the first emergence of CXCR4-using virus in PBMCs above the level of detection of the assay in these individuals. Trofile testing was performed with clonal HIV type 1 (HIV-1) variants (n = 21), MT-2 cell culture-derived cells (n = 20) and supernatants (n = 42), and plasma samples (n = 76). Among the clonal HIV-1 variants and MT-2 cell culture-derived samples, the results of the Trofile and MT-2 assays demonstrated a high degree of concordance (95% to 98%). Among consecutive plasma samples, detection of CXCR4-using virus was at or before the time of first detection by the MT-2 assay in 5/10 patients by the original Trofile assay and in 9/10 patients by the enhanced-sensitivity Trofile assay. Differences in the time to the first detection of CXCR4 use between the MT-2 assay (PBMCs

Comparison of human immunodeficiency virus type 1 tropism profiles in clinical samples by the Trofile and MT-2 assays.

PubMed

Coakley, Eoin; Reeves, Jacqueline D; Huang, Wei; Mangas-Ruiz, Marga; Maurer, Irma; Harskamp, Agnes M; Gupta, Soumi; Lie, Yolanda; Petropoulos, Christos J; Schuitemaker, Hanneke; van 't Wout, Angélique B

2009-11-01

The recent availability of CCR5 antagonists as anti-human immunodeficiency virus (anti-HIV) therapeutics has highlighted the need to accurately identify CXCR4-using variants in patient samples when use of this new drug class is considered. The Trofile assay (Monogram Biosciences) has become the method that is the most widely used to define tropism in the clinic prior to the use of a CCR5 antagonist. By comparison, the MT-2 assay has been used since early in the HIV epidemic to define tropism in clinical specimens. Given that there are few data from direct comparisons of these two assays, we evaluated the performance of the plasma-based Trofile assay and the peripheral blood mononuclear cell (PBMC)-based MT-2 assay for the detection of CXCR4 use in defining the tropism of HIV isolates derived from clinical samples. The various samples used for this comparison were derived from participants of the Amsterdam Cohort Studies on HIV infection and AIDS who underwent consecutive MT-2 assay testing of their PBMCs at approximately 3-month intervals. This unique sample set was specifically selected because consecutive MT-2 assays had demonstrated a shift from negative to positive in PBMCs, reflecting the first emergence of CXCR4-using virus in PBMCs above the level of detection of the assay in these individuals. Trofile testing was performed with clonal HIV type 1 (HIV-1) variants (n = 21), MT-2 cell culture-derived cells (n = 20) and supernatants (n = 42), and plasma samples (n = 76). Among the clonal HIV-1 variants and MT-2 cell culture-derived samples, the results of the Trofile and MT-2 assays demonstrated a high degree of concordance (95% to 98%). Among consecutive plasma samples, detection of CXCR4-using virus was at or before the time of first detection by the MT-2 assay in 5/10 patients by the original Trofile assay and in 9/10 patients by the enhanced-sensitivity Trofile assay. Differences in the time to the first detection of CXCR4 use between the MT-2 assay (PBMCs

Esterase Activity and Intracellular Localization in Reconstructed Human Epidermal Cultured Skin Models.

PubMed

Tokudome, Yoshihiro; Katayanagi, Mishina; Hashimoto, Fumie

2015-06-01

Reconstructed human epidermal culture skin models have been developed for cosmetic and pharmaceutical research. This study evaluated the total and carboxyl esterase activities (i.e., Km and Vmax , respectively) and localization in two reconstructed human epidermal culture skin models (LabCyte EPI-MODEL [Japan Tissue Engineering] and EpiDerm [MatTek/Kurabo]). The usefulness of the reconstruction cultured epidermis was also verified by comparison with human and rat epidermis. Homogenized epidermal samples were fractioned by centrifugation. p-nitrophenyl acetate and 4-methylumbelliferyl acetate were used as substrates of total esterase and carboxyl esterase, respectively. Total and carboxyl esterase activities were present in the reconstructed human epidermal culture skin models and were localized in the cytosol. Moreover, the activities and localization were the same as those in human and rat epidermis. LabCyte EPI-MODEL and EpiDerm are potentially useful for esterase activity prediction in human epidermis.

Influence of vestibular activation on respiration in humans

NASA Technical Reports Server (NTRS)

Monahan, Kevin D.; Sharpe, Melissa K.; Drury, Daniel; Ertl, Andrew C.; Ray, Chester A.

2002-01-01

The purpose of this study was to determine the effects of the semicircular canals and otolith organs on respiration in humans. On the basis of animal studies, we hypothesized that vestibular activation would elicit a vestibulorespiratory reflex. To test this hypothesis, respiratory measures, arterial blood pressure, and heart rate were measured during engagement of semicircular canals and/or otolith organs. Dynamic upright pitch and roll (15 cycles/min), which activate the otolith organs and semicircular canals, increased respiratory rate (Delta2 +/- 1 and Delta3 +/- 1 breaths/min, respectively; P < 0.05). Dynamic yaw and lateral pitch (15 cycles/min), which activate the semicircular canals, increased respiration similarly (Delta3 +/- 1 and Delta2 +/- 1, respectively; P < 0.05). Dynamic chair rotation (15 cycles/min), which mimics dynamic yaw but eliminates neck muscle afferent, increased respiration (Delta3 +/- 1; P < 0.05) comparable to dynamic yaw (15 cycles/min). Increases in respiratory rate were graded as greater responses occurred during upright (Delta5 +/- 2 breaths/min) and lateral pitch (Delta4 +/- 1) and roll (Delta5 +/- 1) performed at 30 cycles/min. Increases in breathing frequency resulted in increases in minute ventilation during most interventions. Static head-down rotation, which activates otolith organs, did not alter respiratory rate (Delta1 +/- 1 breaths/min). Collectively, these data indicate that semicircular canals, but not otolith organs or neck muscle afferents, mediate increased ventilation in humans and support the concept that vestibular activation alters respiration in humans.

The human cytoplasmic dynein interactome reveals novel activators of motility

PubMed Central

Redwine, William B; DeSantis, Morgan E; Hollyer, Ian; Htet, Zaw Min; Tran, Phuoc Tien; Swanson, Selene K; Florens, Laurence; Washburn, Michael P; Reck-Peterson, Samara L

2017-01-01

In human cells, cytoplasmic dynein-1 is essential for long-distance transport of many cargos, including organelles, RNAs, proteins, and viruses, towards microtubule minus ends. To understand how a single motor achieves cargo specificity, we identified the human dynein interactome by attaching a promiscuous biotin ligase (‘BioID’) to seven components of the dynein machinery, including a subunit of the essential cofactor dynactin. This method reported spatial information about the large cytosolic dynein/dynactin complex in living cells. To achieve maximal motile activity and to bind its cargos, human dynein/dynactin requires ‘activators’, of which only five have been described. We developed methods to identify new activators in our BioID data, and discovered that ninein and ninein-like are a new family of dynein activators. Analysis of the protein interactomes for six activators, including ninein and ninein-like, suggests that each dynein activator has multiple cargos. DOI: http://dx.doi.org/10.7554/eLife.28257.001 PMID:28718761

a Three-Step Spatial-Temporal Clustering Method for Human Activity Pattern Analysis

NASA Astrophysics Data System (ADS)

Huang, W.; Li, S.; Xu, S.

2016-06-01

How people move in cities and what they do in various locations at different times form human activity patterns. Human activity pattern plays a key role in in urban planning, traffic forecasting, public health and safety, emergency response, friend recommendation, and so on. Therefore, scholars from different fields, such as social science, geography, transportation, physics and computer science, have made great efforts in modelling and analysing human activity patterns or human mobility patterns. One of the essential tasks in such studies is to find the locations or places where individuals stay to perform some kind of activities before further activity pattern analysis. In the era of Big Data, the emerging of social media along with wearable devices enables human activity data to be collected more easily and efficiently. Furthermore, the dimension of the accessible human activity data has been extended from two to three (space or space-time) to four dimensions (space, time and semantics). More specifically, not only a location and time that people stay and spend are collected, but also what people "say" for in a location at a time can be obtained. The characteristics of these datasets shed new light on the analysis of human mobility, where some of new methodologies should be accordingly developed to handle them. Traditional methods such as neural networks, statistics and clustering have been applied to study human activity patterns using geosocial media data. Among them, clustering methods have been widely used to analyse spatiotemporal patterns. However, to our best knowledge, few of clustering algorithms are specifically developed for handling the datasets that contain spatial, temporal and semantic aspects all together. In this work, we propose a three-step human activity clustering method based on space, time and semantics to fill this gap. One-year Twitter data, posted in Toronto, Canada, is used to test the clustering-based method. The results show that the

High seroprevalence of human herpesvirus 8 infection in patients with systemic lupus erythematosus.

PubMed

Tung, Yi-Ching; Ke, Liang-Yin; Tsai, Shih-Meng; Lu, Po-Liang; Tsai, Wen-Chan

2013-12-01

Systemic lupus erythematosus (SLE) is an autoimmune disease that is associated with genetic and environmental factors and microbial infection. With respect to microbial infection, the Epstein-Barr virus (EBV) has been a widely discussed issue in recent decades. Human herpesvirus 8 (HHV-8) belongs to the same γ-herpesvirus subfamily as EBV and is closely related to it. Until now, only one paper has reported the prevalence of HHV-8 infection in SLE patients. The goal of this study was to detect the prevalence of HHV-8 infection in SLE patients to elucidate the relationship between HHV-8 and SLE. We assessed the seroprevalence of HHV-8 in SLE patients by immunofluorescence assay. In addition, we quantified the HHV-8 DNA in peripheral blood mononuclear cells (PBMCs) from SLE patients by real-time polymerase chain reaction (PCR). The prevalence of HHV-8 antibodies in SLE and normal controls was 57.8% (26/45) and 19.2% (5/26), respectively. These data were highly significant (P = 0.001). No HHV-8 DNA was detected by real-time PCR in SLE patients' PBMCs. In addition, we analyzed the prevalence of HHV-8 infection in patients with rheumatoid arthritis and ankylosing spondylitis. There was no significant difference between the patients and normal controls. A high seroprevalence of HHV-8 infection was found in patients with SLE. The negative DNA load and high seropositive rate indicate that most patients might have latent infection. © 2013 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

The Activity of Immunoglobulin Y Anti-Mycobacterium tuberculosis on Proliferation and Cytokine Expression of Rat Peripheral Blood Mononuclear Cells.

PubMed

Sudjarwo, Sri Agus; Eraiko, Koerniasari; Sudjarwo, Giftania Wardani; Koerniasari

2017-12-01

It has long been known that chickens, like mammals, are capable of producing antigen-specific immunoglobulin Y (IgY), which functions similar to IgG. The present study was performed to investigate the activity of IgY anti- Mycobacterium tuberculosis on proliferation, interleukin (IL)-2, and interferon (IFN)-γ expression of rat peripheral blood mononuclear cells (PBMCs). The activity of IgY anti- M. tuberculosis in different doses (25, 50, and 100 μg/ml) on rat PBMCs proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The production of IL-2 and IFN-γ in the PBMC supernatant was determined using enzyme-linked immunosorbent assay. Investigation was performed on mRNA expression of IL-2 and IFN-γ by reverse transcription-polymerase chain reaction (RT-PCR). IgY anti- M. tuberculosis significantly increased the proliferation of rat PBMC. Furthermore, IgY anti-M. tuberculosis dose dependently increased IL-2 and IFN-γ production in PBMC, suggesting that pharmacological activities of IgY anti- M. tuberculosis in PBMC may be mediated by regulating the production of cytokines. In the RT-PCR, expression of cytokines such as IL-2 and IFN-γ in PBMC cultures was increased by IgY anti- M. tuberculosis . We concluded that increasing IL-2 and IFN-γ productions in PBMC was related to IgY anti- M. tuberculosis , stimulating the mRNA transcription (gene expression) of these cytokines which can induce proliferation of PBMC. Lohman laying hens immunized intramuscularly with antigens of M. tuberculosis can produce specific IgY anti- Mycobacterium tuberculosis complexIgY anti- M. tuberculosis significantly increased the proliferation of rat peripheral blood mononuclear cell (PBMC)IgY anti- M. tuberculosis dose dependently increased interleukin 2 (IL-2) and interferon (IFN)-γ production in PBMCIn the reverse transcription-polymerase chain reaction, expression of cytokines such as IL-2 and IFN-γ in PBMC cultures was increased by Ig

Transfer Learning for Improved Audio-Based Human Activity Recognition.

PubMed

Ntalampiras, Stavros; Potamitis, Ilyas

2018-06-25

Human activities are accompanied by characteristic sound events, the processing of which might provide valuable information for automated human activity recognition. This paper presents a novel approach addressing the case where one or more human activities are associated with limited audio data, resulting in a potentially highly imbalanced dataset. Data augmentation is based on transfer learning; more specifically, the proposed method: (a) identifies the classes which are statistically close to the ones associated with limited data; (b) learns a multiple input, multiple output transformation; and (c) transforms the data of the closest classes so that it can be used for modeling the ones associated with limited data. Furthermore, the proposed framework includes a feature set extracted out of signal representations of diverse domains, i.e., temporal, spectral, and wavelet. Extensive experiments demonstrate the relevance of the proposed data augmentation approach under a variety of generative recognition schemes.

The Need for a Harmonized Repository for Next-Generation Human Activity Data

EPA Science Inventory

Multi-tiered human time-activity-location data can inform many efforts to describe human exposures to air pollutants and other chemicals on a range of temporal and spatial scales. In the last decade, EPA's Consolidated Human Activity Database (CHAD) has served as a harmonized rep...

Paracetamol (acetaminophen) attenuates in vitro mast cell and peripheral blood mononucleocyte cell histamine release induced by N-acetylcysteine.

PubMed

Coulson, James; Thompson, John Paul

2010-02-01

The treatment of acute paracetamol (acetaminophen) poisoning with N-acetylcysteine (NAC) is frequently complicated by an anaphylactoid reaction to the antidote. The mechanism that underlies this reaction is unclear. We used the human mast cell line 1 (HMC-1) and human peripheral blood mononucleocytes (PBMCs) to investigate the effects of NAC and paracetamol on histamine secretion in vitro. HMC-1 and human PBMCs were incubated in the presence of increasing concentrations of NAC +/- paracetamol. Cell viability was determined by the Trypan Blue Assay, and histamine secretion was measured by ELISA. NAC was toxic to HMC-1 cells at 100 mg/mL and to PBMCs at 67 mg/mL. NAC increased HMC-1 and PBMC histamine secretion at concentrations of NAC from 20 to 50 mg/mL and 2.5 to 100 mg/mL, respectively. NAC-induced histamine secretion by both cell types was reduced by co-incubation with 2.5 mg/mL of paracetamol. Paracetamol (acetaminophen) is capable of modifying histamine secretion in vitro. This may explain the clinical observation of a lower incidence of adverse reactions to NAC in vivo when higher concentrations of paracetamol are present than when paracetamol concentrations are low. Paracetamol (acetaminophen) attenuates in vitro mast cell and PBMC cell histamine release induced by NAC.

Selective targeting of JAK/STAT signaling is potentiated by Bcl-xL blockade in IL-2–dependent adult T-cell leukemia

PubMed Central

Zhang, Meili; Mathews Griner, Lesley A.; Ju, Wei; Duveau, Damien Y.; Guha, Rajarshi; Petrus, Michael N.; Wen, Bernard; Maeda, Michiyuki; Shinn, Paul; Ferrer, Marc; Conlon, Kevin D.; Bamford, Richard N.; O’Shea, John J.; Thomas, Craig J.; Waldmann, Thomas A.

2015-01-01

Adult T-cell leukemia (ATL) develops in individuals infected with human T-cell lymphotropic virus-1 (HTLV-1). Presently there is no curative therapy for ATL. HTLV-1–encoded protein Tax (transactivator from the X-gene region) up-regulates Bcl-xL (B-cell lymphoma-extra large) expression and activates interleukin-2 (IL-2), IL-9, and IL-15 autocrine/paracrine systems, resulting in amplified JAK/STAT signaling. Inhibition of JAK signaling reduces cytokine-dependent ex vivo proliferation of peripheral blood mononuclear cells (PBMCs) from ATL patients in smoldering/chronic stages. Currently, two JAK inhibitors are approved for human use. In this study, we examined activity of multiple JAK inhibitors in ATL cell lines. The selective JAK inhibitor ruxolitinib was examined in a high-throughput matrix screen combined with >450 potential therapeutic agents, and Bcl-2/Bcl-xL inhibitor navitoclax was identified as a strong candidate for multicomponent therapy. The combination was noted to strongly activate BAX (Bcl-2-associated X protein), effect mitochondrial depolarization, and increase caspase 3/7 activities that lead to cleavage of PARP (poly ADP ribose polymerase) and Mcl-1 (myeloid cell leukemia 1). Ruxolitinib and navitoclax independently demonstrated modest antitumor efficacy, whereas the combination dramatically lowered tumor burden and prolonged survival in an ATL murine model. This combination strongly blocked ex vivo proliferation of five ATL patients’ PBMCs. These studies provide support for a therapeutic trial in patients with smoldering/chronic ATL using a drug combination that inhibits JAK signaling and antiapoptotic protein Bcl-xL. PMID:26396258

PgTeL, the lectin found in Punica granatum juice, is an antifungal agent against Candida albicans and Candida krusei.

PubMed

da Silva, Pollyanna Michelle; de Moura, Maiara Celine; Gomes, Francis Soares; da Silva Trentin, Danielle; Silva de Oliveira, Ana Patrícia; de Mello, Gabriela Souto Vieira; da Rocha Pitta, Maira Galdino; de Melo Rego, Moacyr Jesus Barreto; Coelho, Luana Cassandra Breitenbach Barroso; Macedo, Alexandre José; de Figueiredo, Regina Celia Bressan Queiroz; Paiva, Patrícia Maria Guedes; Napoleão, Thiago Henrique

2018-03-01

The pomegranate (Punica granatum) sarcotesta contains a chitin-binding lectin (PgTeL) with antibacterial activity against human pathogenic species. In this work, the structural stability of PgTeL was evaluated by fluorimetric analysis and the lectin was evaluated for cytotoxicity to human peripheral blood mononuclear cells (PBMCs) and antifungal activity against Candida albicans and Candida krusei. PgTeL folding was impaired when lectin was incubated at pH≥6.0. On the other hand, the lectin did not undergo unfolding even when heated at 100°C. PgTeL (1, 10, and 100μg/mL) was not cytotoxic to PBMCs. Antifungal activity was detected for C. albicans (MIC: 25μg/mL; MFC: 50μg/mL) and C. krusei (MIC and MFC of 12.5μg/mL). Treatment of yeast cells with PgTeL resulted in decrease of intracellular ATP content even at sub-inhibitory concentrations (½MIC and ¼MIC) and induced lipid peroxidation. In addition, PgTeL damaged the integrity of fungal cell wall of both species, with more pronounced effects in C. krusei. The lectin showed significant antibiofilm activity on C. albicans at sub-inhibitory concentrations (0.195 and 0.39μg/mL). In conclusion, PgTeL is an anti-Candida agent whose action mechanism involves oxidative stress, energetic collapse, damage to the cell wall and rupture of yeast cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Robust Indoor Human Activity Recognition Using Wireless Signals.

PubMed

Wang, Yi; Jiang, Xinli; Cao, Rongyu; Wang, Xiyang

2015-07-15

Wireless signals-based activity detection and recognition technology may be complementary to the existing vision-based methods, especially under the circumstance of occlusions, viewpoint change, complex background, lighting condition change, and so on. This paper explores the properties of the channel state information (CSI) of Wi-Fi signals, and presents a robust indoor daily human activity recognition framework with only one pair of transmission points (TP) and access points (AP). First of all, some indoor human actions are selected as primitive actions forming a training set. Then, an online filtering method is designed to make actions' CSI curves smooth and allow them to contain enough pattern information. Each primitive action pattern can be segmented from the outliers of its multi-input multi-output (MIMO) signals by a proposed segmentation method. Lastly, in online activities recognition, by selecting proper features and Support Vector Machine (SVM) based multi-classification, activities constituted by primitive actions can be recognized insensitive to the locations, orientations, and speeds.

Systems analysis of protective immune responses to RTS,S malaria vaccination in humans

PubMed Central

Kazmin, Dmitri; Nakaya, Helder I.; Lee, Eva K.; Johnson, Matthew J.; van der Most, Robbert; van den Berg, Robert A.; Ballou, W. Ripley; Jongert, Erik; Wille-Reece, Ulrike; Ockenhouse, Christian; Aderem, Alan; Zak, Daniel E.; Sadoff, Jerald; Hendriks, Jenny; Wrammert, Jens; Ahmed, Rafi; Pulendran, Bali

2017-01-01

RTS,S is an advanced malaria vaccine candidate and confers significant protection against Plasmodium falciparum infection in humans. Little is known about the molecular mechanisms driving vaccine immunity. Here, we applied a systems biology approach to study immune responses in subjects receiving three consecutive immunizations with RTS,S (RRR), or in those receiving two immunizations of RTS,S/AS01 following a primary immunization with adenovirus 35 (Ad35) (ARR) vector expressing circumsporozoite protein. Subsequent controlled human malaria challenge (CHMI) of the vaccinees with Plasmodium-infected mosquitoes, 3 wk after the final immunization, resulted in ∼50% protection in both groups of vaccinees. Circumsporozoite protein (CSP)-specific antibody titers, prechallenge, were associated with protection in the RRR group. In contrast, ARR-induced lower antibody responses, and protection was associated with polyfunctional CD4+ T-cell responses 2 wk after priming with Ad35. Molecular signatures of B and plasma cells detected in PBMCs were highly correlated with antibody titers prechallenge and protection in the RRR cohort. In contrast, early signatures of innate immunity and dendritic cell activation were highly associated with protection in the ARR cohort. For both vaccine regimens, natural killer (NK) cell signatures negatively correlated with and predicted protection. These results suggest that protective immunity against P. falciparum can be achieved via multiple mechanisms and highlight the utility of systems approaches in defining molecular correlates of protection to vaccination. PMID:28193898

Amphetamine increases activity but not exploration in humans and mice

PubMed Central

Minassian, Arpi; Young, Jared W.; Cope, Zackary A.; Henry, Brook L.; Geyer, Mark A.; Perry, William

2015-01-01

Rationale Cross-species quantification of physiological behavior enables a better understanding of the biological systems underlying neuropsychiatric diseases such as Bipolar Disorder (BD). Cardinal symptoms of manic BD include increased motor activity and goal-directed behavior, thought to be related to increased catecholamine activity, potentially selective to dopamine homeostatic dysregulation. Objectives The objective of this study was to test whether acute administration of amphetamine, a norepinephrine/dopamine transporter inhibitor and dopamine releaser, would replicate the profile of activity and exploration observed in both humans with manic BD and mouse models of mania. Methods Healthy volunteers with no psychiatric history were randomized to a one-time dose of placebo (n=25), 10 mg d-amphetamine (n=18), or 20 mg amphetamine (n=23). 80 mice were administered one of 4 doses of d-amphetamine or vehicle. Humans and mice were tested in the Behavioral Pattern Monitor (BPM), which quantifies motor activity, exploratory behavior, and spatial patterns of behavior. Results In humans, the 20-mg dose of amphetamine increased motor activity as measured by acceleration without marked effects on exploration or spatial patterns of activity. In mice, amphetamine increased activity, decreased specific exploration, and caused straighter, one-dimensional movements in a dose-dependent manner. Conclusions Consistent with mice, amphetamine increased motoric activity in humans without increasing exploration. Given that BD patients exhibit heightened exploration, these data further emphasize the limitation of amphetamine-induced hyperactivity as a suitable model for BD. Further, these studies highlight the utility of cross-species physiological paradigms in validating biological mechanisms of psychiatric diseases. PMID:26449721

Esterase Activity and Intracellular Localization in Reconstructed Human Epidermal Cultured Skin Models

PubMed Central

Katayanagi, Mishina; Hashimoto, Fumie

2015-01-01

Background Reconstructed human epidermal culture skin models have been developed for cosmetic and pharmaceutical research. Objective This study evaluated the total and carboxyl esterase activities (i.e., Km and Vmax, respectively) and localization in two reconstructed human epidermal culture skin models (LabCyte EPI-MODEL [Japan Tissue Engineering] and EpiDerm [MatTek/Kurabo]). The usefulness of the reconstruction cultured epidermis was also verified by comparison with human and rat epidermis. Methods Homogenized epidermal samples were fractioned by centrifugation. p-nitrophenyl acetate and 4-methylumbelliferyl acetate were used as substrates of total esterase and carboxyl esterase, respectively. Results Total and carboxyl esterase activities were present in the reconstructed human epidermal culture skin models and were localized in the cytosol. Moreover, the activities and localization were the same as those in human and rat epidermis. Conclusion LabCyte EPI-MODEL and EpiDerm are potentially useful for esterase activity prediction in human epidermis. PMID:26082583

Methods to Evaluate Cytotoxicity and Immunosuppression of Combustible Tobacco Product Preparations

PubMed Central

Arimilli, Subhashini; Damratoski, Brad E.; G.L., Prasad

2015-01-01

Among other pathophysiological changes, chronic exposure to cigarette smoke causes inflammation and immune suppression, which have been linked to increased susceptibility of smokers to microbial infections and tumor incidence. Ex vivo suppression of receptor-mediated immune responses in human peripheral blood mononuclear cells (PBMCs) treated with smoke constituents is an attractive approach to study mechanisms and evaluate the likely long-term effects of exposure to tobacco products. Here, we optimized methods to perform ex vivo assays using PBMCs stimulated by bacterial lipopolysaccharide, a Toll-like receptor-4 ligand. The effects of whole smoke-conditioned medium (WS-CM), a combustible tobacco product preparation (TPP), and nicotine were investigated on cytokine secretion and target cell killing by PBMCs in the ex vivo assays. We show that secreted cytokines IFN-γ, TNF, IL-10, IL-6, and IL-8 and intracellular cytokines IFN-γ, TNF-α, and MIP-1α were suppressed in WS-CM-exposed PBMCs. The cytolytic function of effector PBMCs, as determined by a K562 target cell killing assay was also reduced by exposure to WS-CM; nicotine was minimally effective in these assays. In summary, we present a set of improved assays to evaluate the effects of TPPs in ex vivo assays, and these methods could be readily adapted for testing other products of interest. PMID:25650834

Methods to evaluate cytotoxicity and immunosuppression of combustible tobacco product preparations.

PubMed

Arimilli, Subhashini; Damratoski, Brad E; G L, Prasad

2015-01-10

Among other pathophysiological changes, chronic exposure to cigarette smoke causes inflammation and immune suppression, which have been linked to increased susceptibility of smokers to microbial infections and tumor incidence. Ex vivo suppression of receptor-mediated immune responses in human peripheral blood mononuclear cells (PBMCs) treated with smoke constituents is an attractive approach to study mechanisms and evaluate the likely long-term effects of exposure to tobacco products. Here, we optimized methods to perform ex vivo assays using PBMCs stimulated by bacterial lipopolysaccharide, a Toll-like receptor-4 ligand. The effects of whole smoke-conditioned medium (WS-CM), a combustible tobacco product preparation (TPP), and nicotine were investigated on cytokine secretion and target cell killing by PBMCs in the ex vivo assays. We show that secreted cytokines IFN-γ, TNF, IL-10, IL-6, and IL-8 and intracellular cytokines IFN-γ, TNF-α, and MIP-1α were suppressed in WS-CM-exposed PBMCs. The cytolytic function of effector PBMCs, as determined by a K562 target cell killing assay was also reduced by exposure to WS-CM; nicotine was minimally effective in these assays. In summary, we present a set of improved assays to evaluate the effects of TPPs in ex vivo assays, and these methods could be readily adapted for testing other products of interest.

Cryopreservation does not alter main characteristics of Good Manufacturing Process-grade human multipotent mesenchymal stromal cells including immunomodulating potential and lack of malignant transformation.

PubMed

Luetzkendorf, Jana; Nerger, Katrin; Hering, Julian; Moegel, Angelika; Hoffmann, Katrin; Hoefers, Christiane; Mueller-Tidow, Carsten; Mueller, Lutz P

2015-02-01

The immunomodulating capacity of multipotent mesenchymal stromal cells (MSCs) qualifies them as a therapeutic tool in several diseases. However, repeated transplantation with products of reproducible characteristics may be required. This could be achieved with cryopreserved aliquots of Good Manufacturing Practice (GMP)-grade MSCs. However, the impact of cryopreservation on the characteristics of GMP-MSCs is ill defined. We produced fresh and cryopreserved MSCs from human donors with a xenogen-free GMP protocol. Immunogenicity and immunomodulating capacity were tested in co-culture with putative recipient-specific peripheral blood mononuclear cells (PBMCs). Risk of malignant transformation was assessed in vitro and in vivo. Cryopreservation had no impact on viability and consensus criteria of MSCs. In co-culture with PBMCs, MSCs showed low immunogenicity and suppressed mitogen-stimulated proliferation of PBMC irrespective of cryopreservation. Cytogenetic aberrations were not observed consistently in fresh and cryopreserved products, and no signs of malignant transformation occurred in functional assays. MSC products from an elderly pretreated donor showed reduced functional quality, but imminent failure of functional criteria could be detected by an increased population doubling time in early passages. This study is the first systematic analysis on cryopreservation of xenogen-free human bone marrow-derived GMP-MSCs. The data support that cryopreservation does not alter the characteristics of the cells and thus may allow the generation of products for serial transplantation. In addition, the protocol allowed early detection of MSC products with low functional capacity. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Exploring Techniques for Vision Based Human Activity Recognition: Methods, Systems, and Evaluation

PubMed Central

Xu, Xin; Tang, Jinshan; Zhang, Xiaolong; Liu, Xiaoming; Zhang, Hong; Qiu, Yimin

2013-01-01

With the wide applications of vision based intelligent systems, image and video analysis technologies have attracted the attention of researchers in the computer vision field. In image and video analysis, human activity recognition is an important research direction. By interpreting and understanding human activities, we can recognize and predict the occurrence of crimes and help the police or other agencies react immediately. In the past, a large number of papers have been published on human activity recognition in video and image sequences. In this paper, we provide a comprehensive survey of the recent development of the techniques, including methods, systems, and quantitative evaluation of the performance of human activity recognition. PMID:23353144

Combining users' activity survey and simulators to evaluate human activity recognition systems.

PubMed

Azkune, Gorka; Almeida, Aitor; López-de-Ipiña, Diego; Chen, Liming

2015-04-08

Evaluating human activity recognition systems usually implies following expensive and time-consuming methodologies, where experiments with humans are run with the consequent ethical and legal issues. We propose a novel evaluation methodology to overcome the enumerated problems, which is based on surveys for users and a synthetic dataset generator tool. Surveys allow capturing how different users perform activities of daily living, while the synthetic dataset generator is used to create properly labelled activity datasets modelled with the information extracted from surveys. Important aspects, such as sensor noise, varying time lapses and user erratic behaviour, can also be simulated using the tool. The proposed methodology is shown to have very important advantages that allow researchers to carry out their work more efficiently. To evaluate the approach, a synthetic dataset generated following the proposed methodology is compared to a real dataset computing the similarity between sensor occurrence frequencies. It is concluded that the similarity between both datasets is more than significant.

Quantification of cytokine mRNA in peripheral blood mononuclear cells using branched DNA (bDNA) technology.

PubMed

Shen, L P; Sheridan, P; Cao, W W; Dailey, P J; Salazar-Gonzalez, J F; Breen, E C; Fahey, J L; Urdea, M S; Kolberg, J A

1998-06-01

Changes in the patterns of cytokine expression are thought to be of central importance in human infectious and inflammatory diseases. As such, there is a need for precise, reproducible assays for quantification of cytokine mRNA that are amenable to routine use in a clinical setting. In this report, we describe the design and performance of a branched DNA (bDNA) assay for the direct quantification of multiple cytokine mRNA levels in peripheral blood mononuclear cells (PBMCs). Oligonucleotide target probe sets were designed for several human cytokines, including TNFalpha, IL-2, IL-4, IL-6, IL-10, and IFNgamma. The bDNA assay yielded highly reproducible quantification of cytokine mRNAs, exhibited a broad linear dynamic range of over 3-log10, and showed a sensitivity sufficient to measure at least 3000 molecules. The potential clinical utility of the bDNA assay was explored by measuring cytokine mRNA levels in PBMCs from healthy and immunocompromised individuals. Cytokine expression levels in PBMCs from healthy blood donors were found to remain relatively stable over a one-month period of time. Elevated levels of IFNgamma mRNA were detected in PBMCs from HIV-1 seropositive individuals, but no differences in mean levels of TNFalpha or IL-6 mRNA were detected between seropositive and seronegative individuals. By providing a reproducible method for quantification of low abundance transcripts in clinical specimens, the bDNA assay may be useful for studies addressing the role of cytokine expression in disease.

Nonlinear analysis of human physical activity patterns in health and disease.

PubMed

Paraschiv-Ionescu, A; Buchser, E; Rutschmann, B; Aminian, K

2008-02-01

The reliable and objective assessment of chronic disease state has been and still is a very significant challenge in clinical medicine. An essential feature of human behavior related to the health status, the functional capacity, and the quality of life is the physical activity during daily life. A common way to assess physical activity is to measure the quantity of body movement. Since human activity is controlled by various factors both extrinsic and intrinsic to the body, quantitative parameters only provide a partial assessment and do not allow for a clear distinction between normal and abnormal activity. In this paper, we propose a methodology for the analysis of human activity pattern based on the definition of different physical activity time series with the appropriate analysis methods. The temporal pattern of postures, movements, and transitions between postures was quantified using fractal analysis and symbolic dynamics statistics. The derived nonlinear metrics were able to discriminate patterns of daily activity generated from healthy and chronic pain states.

Activation of mouse and human peroxisome proliferator-activated receptor alpha by perfluoroalkyl acids of different functional groups and chain lengths.

PubMed

Wolf, Cynthia J; Takacs, Margy L; Schmid, Judith E; Lau, Christopher; Abbott, Barbara D

2008-11-01

Perfluoroalkyl acids (PFAAs) are surfactants used in consumer products and persist in the environment. Some PFAAs elicit adverse effects on rodent development and survival. PFAAs can activate peroxisome proliferator-activated receptor alpha (PPARalpha) and may act via PPARalpha to produce some of their effects. This study evaluated the ability of numerous PFAAs to induce mouse and human PPARalpha activity in a transiently transfected COS-1 cell assay. COS-1 cells were transfected with either a mouse or human PPARalpha receptor-luciferase reporter plasmid. After 24 h, cells were exposed to either negative controls (water or dimethyl sulfoxide, 0.1%); positive control (WY-14643, PPARalpha agonist); perfluorooctanoic acid or perfluorononanoic acid at 0.5-100 microM; perfluorobutanoic acid, perfluorohexanoic acid, perfluorohexane sulfonate, or perfluorodecanoic acid (PFDA) at 5-100 microM; or perfluorobutane sulfonate or perfluorooctane sulfonate at 1-250 microM. After 24 h of exposure, luciferase activity from the plasmid was measured. Each PFAA activated both mouse and human PPARalpha in a concentration-dependent fashion, except PFDA with human PPARalpha. Activation of PPARalpha by PFAA carboxylates was positively correlated with carbon chain length, up to C9. PPARalpha activity was higher in response to carboxylates compared to sulfonates. Activation of mouse PPARalpha was generally higher compared to that of human PPARalpha. We conclude that, in general, (1) PFAAs of increasing carbon backbone chain lengths induce increasing activity of the mouse and human PPARalpha with a few exceptions, (2) PFAA carboxylates are stronger activators of mouse and human PPARalpha than PFAA sulfonates, and (3) in most cases, the mouse PPARalpha appears to be more sensitive to PFAAs than the human PPARalpha in this model.

Gram-positive and gram-negative bacteria induce different patterns of cytokine production in human mononuclear cells irrespective of taxonomic relatedness.

PubMed

Skovbjerg, Susann; Martner, Anna; Hynsjö, Lars; Hessle, Christina; Olsen, Ingar; Dewhirst, Floyd E; Tham, Wilhelm; Wold, Agnes E

2010-01-01

Upon bacterial stimulation, tissue macrophages produce a variety of cytokines that orchestrate the immune response that clears the infection. We have shown that Gram-positives induce higher levels of interleukin-12 (IL-12), interferon-gamma (IFN-gamma), and tumor necrosis factor (TNF) from human peripheral blood mononuclear cells (PBMCs) than do Gram-negatives, which instead induce more of IL-6, IL-8, and IL-10. Here, we study whether these patterns follows or crosses taxonomic borders. PBMCs from blood donors were incubated with UV-inactivated bacteria representing 37 species from five phyla. IL-12, TNF, IL-1beta, IL-6, IL-8, and IL-10 were measured in the supernatants after 24 h and IFN-gamma after 5 days. Irrespective of phylogenetic position, Gram-positive bacteria induced much more IL-12 (nine times more on average) and IFN-gamma (seven times), more TNF (three times), and slightly more IL-1beta (1.5 times) than did Gram-negatives, which instead induced more IL-6 (1.5 times), IL-8 (1.9 times), and IL-10 (3.3 times) than did Gram-positives. A notable exception was the Gram-positive Listeria monocytogenes, which induced very little IL-12, IFN-gamma, and TNF. The results confirm the fundamental difference in innate immune responses to Gram-positive and Gram-negative bacteria, which crosses taxonomic borders and probably reflects differences in cell wall structure.

Decreasing the configurational entropy and the hydrophobicity of EBV-derived peptide 11389 increased its antigenicity, immunogenicity and its ability of inducing IL-6.

PubMed

Urquiza, Mauricio; Guevara, Tatiana; Rodriguez, Cristina; Melo-Cardenas, Johanna; Vanegas, Magnolia; Patarroyo, Manuel E

2012-06-01

Peptide 11389 from CD21-binding region of EBV-gp350/220 protein binds to PBMCs inducing IL-6 expression and inhibiting EBV-binding to PBMCs. In addition, anti-peptide 11389 antibodies recognize EBV-infected cells and inhibit both EBV infection and IL-6 production in PBMCs. We have postulated that native structure stabilization of peptide 11389 sequence can increase its biological activity. The strategy was to modify its sequence to restrict the number of structures that peptide 11389 could acquire in solution (decreasing peptide's configurational entropy) and to weaken the non-relevant intermolecular interactions (decreasing its hydrophobicity), preserving CD21-interacting residues and structure as displayed in the native protein. Thirteen analog peptides were designed and synthesized; most of them were monomers containing an intra-chain disulfide bridge. Analog peptides 34058, 34060, 34061, 34296, 34298, 34299 and 34300 inhibited EBV invasion of PBMCs. Peptides 34059, 34060, 34295 and 34297 induced IL-6 levels in PBMCs (EC50=3.4, 3.3, 0.5, 0.5 μM, respectively) at higher potency than peptide 11389 (EC50=5.8 μM). Peptides 34057, 34059, 34060, 34301 and 34302 interacted with anti-EBV antibodies with affinities from 3 to 50 times higher than peptide 11389. Most of analog peptides were highly immunogenic and elicited antibodies that cross-react with EBV. In conclusion, we have designed peptides displaying higher biological activity than peptide 11389.

Water quality degradation effects on freshwater availability: Impacts to human activities

USGS Publications Warehouse

Peters, N.E.; Meybeck, Michel

2000-01-01

The quality of freshwater at any point on the landscape reflects the combined effects of many processes along water pathways. Human activities on all spatial scales affect both water quality and quantity. Alteration of the landscape and associated vegetation has not only changed the water balance, but typically has altered processes that control water quality. Effects of human activities on a small scale are relevant to an entire drainage basin. Furthermore, local, regional, and global differences in climate and water flow are considerable, causing varying effects of human activities on land and water quality and quantity, depending on location within a watershed, geology, biology, physiographic characteristics, and climate. These natural characteristics also greatly control human activities, which will, in turn, modify (or affect) the natural composition of water. One of the most important issues for effective resource management is recognition of cyclical and cascading effects of human activities on the water quality and quantity along hydrologic pathways. The degradation of water quality in one part of a watershed can have negative effects on users downstream. Everyone lives downstream of the effects of some human activity. An extremely important factor is that substances added to the atmosphere, land, and water generally have relatively long time scales for removal or clean up. The nature of the substance, including its affinity for adhering to soil and its ability to be transformed, affects the mobility and the time scale for removal of the substance. Policy alone will not solve many of the degradation issues, but a combination of policy, education, scientific knowledge, planning, and enforcement of applicable laws can provide mechanisms for slowing the rate of degradation and provide human and environmental protection. Such an integrated approach is needed to effectively manage land and water resources.

Immunotherapy of HIV-infected patients with Gc protein-derived macrophage activating factor (GcMAF).

PubMed

Yamamoto, Nobuto; Ushijima, Naofumi; Koga, Yoshihiko

2009-01-01

Serum Gc protein (known as vitamin D3-binding protein) is the precursor for the principal macrophage activating factor (MAF). The MAF precursor activity of serum Gc protein of HIV-infected patients was lost or reduced because Gc protein is deglycosylated by alpha-N-acetylgalactosaminidase (Nagalase) secreted from HIV-infected cells. Therefore, macrophages of HIV-infected patients having deglycosylated Gc protein cannot be activated, leading to immunosuppression. Since Nagalase is the intrinsic component of the envelope protein gp120, serum Nagalase activity is the sum of enzyme activities carried by both HIV virions and envelope proteins. These Nagalase carriers were already complexed with anti-HIV immunoglobulin G (IgG) but retained Nagalase activity that is required for infectivity. Stepwise treatment of purified Gc protein with immobilized beta-galactosidase and sialidase generated the most potent macrophage activating factor (termed GcMAF), which produces no side effects in humans. Macrophages activated by administration of 100 ng GcMAF develop a large amount of Fc-receptors as well as an enormous variation of receptors that recognize IgG-bound and unbound HIV virions. Since latently HIV-infected cells are unstable and constantly release HIV virions, the activated macrophages rapidly intercept the released HIV virions to prevent reinfection resulting in exhaustion of infected cells. After less than 18 weekly administrations of 100 ng GcMAF for nonanemic patients, they exhibited low serum Nagalase activities equivalent to healthy controls, indicating eradication of HIV-infection, which was also confirmed by no infectious center formation by provirus inducing agent-treated patient PBMCs. No recurrence occurred and their healthy CD + cell counts were maintained for 7 years.

Energy monitoring system based on human activity in the workplace

NASA Astrophysics Data System (ADS)

Mustafa, Nur Hanim; Husain, Mohd Nor; Aziz, Mohamad Zoinol Abidin Abdul; Othman, Mohd Azlishah; Malek, Fareq

2015-05-01

Human behaviors always related to day routine activities in a smart house directly give the significant factor to manage energy usage in human life. An Addition that, the factor will contribute to the best efficiency of the system. This paper will focus on the monitoring efficiency based on duration time in office hours around 8am until 5pm which depend on human behavior at working place. Besides that, the correlation coefficient method is used to show the relation between energy consumption and energy saving based on the total hours of time energy spent. In future, the percentages of energy monitoring system usage will be increase to manage energy saving based on human behaviors. This scenario will help to see the human activity in the workplace in order to get the energy saving and support world green environment.

Chemical synthesis of biologically active tat trans-activating protein of human immunodeficiency virus type 1.

PubMed Central

Chun, R; Glabe, C G; Fan, H

1990-01-01

Full-length (86-residue) polypeptide corresponding to the human immunodeficiency virus type 1 tat trans-activating protein was chemically synthesized on a semiautomated apparatus, using an Fmoc amino acid continuous-flow strategy. The bulk material was relatively homogeneous, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing, and it showed trans-activating activity when scrape loaded into cells containing a human immunodeficiency virus long terminal repeat-chloramphenicol acetyl-transferase reporter plasmid. Reverse-phase high-pressure liquid chromatography yielded a rather broad elution profile, and assays across the column for biological activity indicated a sharper peak. Thus, high-pressure liquid chromatography provided for enrichment of biological activity. Fast atom bombardment-mass spectrometry of tryptic digests of synthetic tat identified several of the predicted tryptic peptides, consistent with accurate chemical synthesis. Images PMID:2186178

Effects of aircraft noise on human activities

NASA Technical Reports Server (NTRS)

Arnoult, M. D.; Gilfillan, L. G.

1983-01-01

The effects of aircrft noise on human activities was investigated by developing a battery of tasks (1) representative of a range of human activities and (2) sensitive to the disruptive effects of noise. The noise used were recordings of jet aircraft and helicopter sounds at three lvels of loudness--60, 70, and 80 dB(A). Experiment 1 investigated 12 different cognitive tasks, along with two intelligibility tasks included to validate that the noises were being effective. Interference with intelligibility was essentially the same as found in the research literature, but only inconsistent effects were found on either accuracy or latency of performance on the cognitive tasks. When the tasks were grouped into four categories (Intelligibility, Matching, Verbal, and Arithmetic), reliable differences in rated annoyingness of the noises were related to the task category and to the type of noise (jet or helicopter).

Functional assessment of human enhancer activities using whole-genome STARR-sequencing.

PubMed

Liu, Yuwen; Yu, Shan; Dhiman, Vineet K; Brunetti, Tonya; Eckart, Heather; White, Kevin P

2017-11-20

Genome-wide quantification of enhancer activity in the human genome has proven to be a challenging problem. Recent efforts have led to the development of powerful tools for enhancer quantification. However, because of genome size and complexity, these tools have yet to be applied to the whole human genome.  In the current study, we use a human prostate cancer cell line, LNCaP as a model to perform whole human genome STARR-seq (WHG-STARR-seq) to reliably obtain an assessment of enhancer activity. This approach builds upon previously developed STARR-seq in the fly genome and CapSTARR-seq techniques in targeted human genomic regions. With an improved library preparation strategy, our approach greatly increases the library complexity per unit of starting material, which makes it feasible and cost-effective to explore the landscape of regulatory activity in the much larger human genome. In addition to our ability to identify active, accessible enhancers located in open chromatin regions, we can also detect sequences with the potential for enhancer activity that are located in inaccessible, closed chromatin regions. When treated with the histone deacetylase inhibitor, Trichostatin A, genes nearby this latter class of enhancers are up-regulated, demonstrating the potential for endogenous functionality of these regulatory elements. WHG-STARR-seq provides an improved approach to current pipelines for analysis of high complexity genomes to gain a better understanding of the intricacies of transcriptional regulation.

Activation of human peroxisome-activated receptor-gamma ...

EPA Pesticide Factsheets

Obesity in children has become an epidemic and recent research suggests a possible contribution from exposure to environmental chemicals. Several chemicals, such as phthalates, brominated flame retardants, and perfluorinated chemicals, are common in house dust on floors where children play and are suspected obesogens. Obesogens can act via a mechanism that involves activation of peroxisome proliferator-activated receptor-gamma (PPARy). A previous study found that dust collected from children’s homes binds to PPARy. Here, we investigated the ability of house dust to activate PPARy in a transiently transfected cell assay. Dust samples were collected in 2012 from carpeted and hardwood floors in children’s homes using thimbles fitted into a vacuum cleaner hose (“TEO” samples), or from homes in an adult cohort NIEHS study. Dust was extracted with 50:50 hexane:acetone, sonicated, centrifuged, and the organic layer collected. This was repeated 2X. The extracts were filtered to remove particulates, dried with purified nitrogen, and reconstituted in DMS0 at 200 ug/ul. COS-1 cells were transfected for 24 hrs with a human PPARy vector containing a luciferase reporter, and exposed for 24 hrs to negative controls water or DMSO (0.1%), positive controls Troglitazone (3 uM in water) or Rosiglitazone (100 nM in DMSO), or dust extracts serially diluted in DMEM at 50, 100, and 200 ug/ml in 0.1% DMSO. Cells were lysed and luciferase activity was measured. Data were log-tra

Developing a Table of Forces for Human Activity as It Relates ...

EPA Pesticide Factsheets

Report The purpose of this project was to evaluate forces generated by human activity that may cause reaerosolization of Bacillus anthracis spores in the aftermath of an intentional release. Understanding human interaction is important to inform communities on activities that may cause the re-distribution of spores. This report presents the results of an extensive literature review of human-surface interaction forces with respect to Bacillus spore reaerosolization in the outdoor environment.

Effects of freezing on the bactericidal activity of human milk.

PubMed

Takci, Sahin; Gulmez, Dolunay; Yigit, Sule; Dogan, Ozlem; Dik, Kezban; Hascelik, Gulsen

2012-08-01

Storage of human milk by freezing has been recommended for long-term storage. The present study analyzed the bactericidal activity of human milk on Escherichia coli and Pseudomonas aeruginosa and determined the changes in bactericidal activity following freezing at -20°C and -80°C for 1 month and 3 months. Forty-eight milk samples were collected from 48 lactating mothers. Each sample was divided into 10 aliquots. Two of the samples were processed immediately and the others were stored at both -20°C and -80°C until analysis after 1 month and 3 months of freezing. All of the fresh milk samples showed bactericidal activity against E coli and P aeruginosa. Freezing at -20°C for 1 month did not cause statistically significant alteration in bactericidal activity (P > 0.017), whereas storage for 3 months lowered the degree of bactericidal activity significantly (P < 0.017) against E coli. Bactericidal activity was protected when the samples were stored at -80°C. There was no statistically significant difference in the bactericidal activity of human milk against E coli between freezing at -20°C and -80°C for 1 month (P > 0.017); however, when milk was stored for 3 months, -80°C was significantly more protective (P < 0.017). Freezing at -20°C and -80°C for 1 month and 3 months did not cause any significant change in bactericidal activity against P aeruginosa (P > 0.05). Storage by freezing at -80°C is more appropriate to keep bactericidal capacity of stored human milk >1 month if affordable and available, especially in intensive care settings.

Ensemble Manifold Rank Preserving for Acceleration-Based Human Activity Recognition.

PubMed

Tao, Dapeng; Jin, Lianwen; Yuan, Yuan; Xue, Yang

2016-06-01

With the rapid development of mobile devices and pervasive computing technologies, acceleration-based human activity recognition, a difficult yet essential problem in mobile apps, has received intensive attention recently. Different acceleration signals for representing different activities or even a same activity have different attributes, which causes troubles in normalizing the signals. We thus cannot directly compare these signals with each other, because they are embedded in a nonmetric space. Therefore, we present a nonmetric scheme that retains discriminative and robust frequency domain information by developing a novel ensemble manifold rank preserving (EMRP) algorithm. EMRP simultaneously considers three aspects: 1) it encodes the local geometry using the ranking order information of intraclass samples distributed on local patches; 2) it keeps the discriminative information by maximizing the margin between samples of different classes; and 3) it finds the optimal linear combination of the alignment matrices to approximate the intrinsic manifold lied in the data. Experiments are conducted on the South China University of Technology naturalistic 3-D acceleration-based activity dataset and the naturalistic mobile-devices based human activity dataset to demonstrate the robustness and effectiveness of the new nonmetric scheme for acceleration-based human activity recognition.

Human Activity Recognition by Combining a Small Number of Classifiers.

PubMed

Nazabal, Alfredo; Garcia-Moreno, Pablo; Artes-Rodriguez, Antonio; Ghahramani, Zoubin

2016-09-01

We consider the problem of daily human activity recognition (HAR) using multiple wireless inertial sensors, and specifically, HAR systems with a very low number of sensors, each one providing an estimation of the performed activities. We propose new Bayesian models to combine the output of the sensors. The models are based on a soft outputs combination of individual classifiers to deal with the small number of sensors. We also incorporate the dynamic nature of human activities as a first-order homogeneous Markov chain. We develop both inductive and transductive inference methods for each model to be employed in supervised and semisupervised situations, respectively. Using different real HAR databases, we compare our classifiers combination models against a single classifier that employs all the signals from the sensors. Our models exhibit consistently a reduction of the error rate and an increase of robustness against sensor failures. Our models also outperform other classifiers combination models that do not consider soft outputs and an Markovian structure of the human activities.

Recognition of human activities using depth images of Kinect for biofied building

NASA Astrophysics Data System (ADS)

Ogawa, Ami; Mita, Akira

2015-03-01

These days, various functions in the living spaces are needed because of an aging society, promotion of energy conservation, and diversification of lifestyles. To meet this requirement, we propose "Biofied Building". The "Biofied Building" is the system learnt from living beings. The various information is accumulated in a database using small sensor agent robots as a key function of this system to control the living spaces. Among the various kinds of information about the living spaces, especially human activities can be triggers for lighting or air conditioning control. By doing so, customized space is possible. Human activities are divided into two groups, the activities consisting of single behavior and the activities consisting of multiple behaviors. For example, "standing up" or "sitting down" consists of a single behavior. These activities are accompanied by large motions. On the other hand "eating" consists of several behaviors, holding the chopsticks, catching the food, putting them in the mouth, and so on. These are continuous motions. Considering the characteristics of two types of human activities, we individually, use two methods, R transformation and variance. In this paper, we focus on the two different types of human activities, and propose the two methods of human activity recognition methods for construction of the database of living space for "Biofied Building". Finally, we compare the results of both methods.

Activation of human liver 3 alpha-hydroxysteroid dehydrogenase by sulphobromophthalein.

PubMed Central

Matsuura, K; Tamada, Y; Deyashiki, Y; Miyabe, Y; Nakanishi, M; Ohya, I; Hara, A

1996-01-01

Human liver contains at least two isoenzymes (DD2 and DD4) of 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase. The NADP(H)-linked oxidoreductase activities of DD4 were activated more than 4-fold by sulphobromophthalein at concentrations above 20 microM and under physiological pH conditions. Sulphobromophthalein did not stimulate the activities of DD2 and human liver aldehyde reductase, which are functionally and/or structurally related to DD4. No stimulatory effect on the activity of DD4 was observed with other organic anions such as Indocyanine Green, haematin and Rose Bengal. The binding of sulphobromophthalein to DD4 was instantaneous and reversible, and was detected by fluorescence and ultrafiltration assays. The activation by sulphobromophthalein decreased the activation energy in the dehydrogenation reaction for the enzyme, and increased both kcat, and Km values for the coenzymes and substrates. Kinetic analyses with respect to concentrations of NADP+ and (S)-(+)-indan-1-ol indicated that sulphobromophthalein was a non-essential activator of mixed type showing a dissociation constant of 2.6 microM. Thus, the human 3 alpha-hydroxysteroid dehydrogenase isoenzyme has a binding site specific to sulphobromophthalein, and the hepatic metabolism mediated by this isoenzyme may be influenced when this drug is administered. PMID:8546681

Tax secretion from peripheral blood mononuclear cells and Tax detection in plasma of patients with human T-lymphotropic virus-type 1-associated myelopathy/tropical spastic paraparesis and asymptomatic carriers.

PubMed

Medina, Fernando; Quintremil, Sebastián; Alberti, Carolina; Godoy, Fabián; Pando, María E; Bustamante, Andrés; Barriga, Andrés; Cartier, Luis; Puente, Javier; Tanaka, Yuetsu; Valenzuela, María A; Ramírez, Eugenio

2016-03-01

Human T-lymphotropic virus-type 1 (HTLV-1) is the etiologic agent of the neurologic disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Tax viral protein plays a critical role in viral pathogenesis. Previous studies suggested that extracellular Tax might involve cytokine-like extracellular effects. We evaluated Tax secretion in 18 h-ex vivo peripheral blood mononuclear cells (PBMCs) cultures from 15 HAM/TSP patients and 15 asymptomatic carriers. Futhermore, Tax plasma level was evaluated from other 12 HAM/TSP patients and 10 asymptomatic carriers. Proviral load and mRNA encoding Tax were quantified by PCR and real-time RT-PCR, respectively. Intracellular Tax in CD4(+)CD25(+) cells occurred in 100% and 86.7% of HAM/TSP patients and asymptomatic carriers, respectively. Percentage of CD4(+)CD25(+) Tax+, proviral load and mRNA encoding Tax were significantly higher in HAM/TSP patients. Western blot analyses showed higher secretion levels of ubiquitinated Tax in HAM/TSP patients than in asymptomatic carriers. In HTLV-1-infected subjects, Western blot of plasma Tax showed higher levels in HAM/TSP patients than in asymptomatic carriers, whereas no Tax was found in non-infected subjects. Immunoprecipitated plasma Tax resolved on SDS-PAGE gave two major bands of 57 and 48 kDa allowing identification of Tax and Ubiquitin peptides by mass spectrometry. Relative percentage of either CD4(+)CD25(+) Tax+ cells, or Tax protein released from PBMCs, or plasma Tax, correlates neither with tax mRNA nor with proviral load. This fact could be explained by a complex regulation of Tax expression. Tax secreted from PBMCs or present in plasma could potentially become a biomarker to distinguish between HAM/TSP patients and asymptomatic carriers. © 2015 Wiley Periodicals, Inc.

Derivation of low flow frequency distributions under human activities and its implications

NASA Astrophysics Data System (ADS)

Gao, Shida; Liu, Pan; Pan, Zhengke; Ming, Bo; Guo, Shenglian; Xiong, Lihua

2017-06-01

Low flow, refers to a minimum streamflow in dry seasons, is crucial to water supply, agricultural irrigation and navigation. Human activities, such as groundwater pumping, influence low flow severely. In order to derive the low flow frequency distribution functions under human activities, this study incorporates groundwater pumping and return flow as variables in the recession process. Steps are as follows: (1) the original low flow without human activities is assumed to follow a Pearson type three distribution, (2) the probability distribution of climatic dry spell periods is derived based on a base flow recession model, (3) the base flow recession model is updated under human activities, and (4) the low flow distribution under human activities is obtained based on the derived probability distribution of dry spell periods and the updated base flow recession model. Linear and nonlinear reservoir models are used to describe the base flow recession, respectively. The Wudinghe basin is chosen for the case study, with daily streamflow observations during 1958-2000. Results show that human activities change the location parameter of the low flow frequency curve for the linear reservoir model, while alter the frequency distribution function for the nonlinear one. It is indicated that alter the parameters of the low flow frequency distribution is not always feasible to tackle the changing environment.

A Flexible Approach for Human Activity Recognition Using Artificial Hydrocarbon Networks

PubMed Central

Ponce, Hiram; Miralles-Pechuán, Luis; Martínez-Villaseñor, María de Lourdes

2016-01-01

Physical activity recognition based on sensors is a growing area of interest given the great advances in wearable sensors. Applications in various domains are taking advantage of the ease of obtaining data to monitor personal activities and behavior in order to deliver proactive and personalized services. Although many activity recognition systems have been developed for more than two decades, there are still open issues to be tackled with new techniques. We address in this paper one of the main challenges of human activity recognition: Flexibility. Our goal in this work is to present artificial hydrocarbon networks as a novel flexible approach in a human activity recognition system. In order to evaluate the performance of artificial hydrocarbon networks based classifier, experimentation was designed for user-independent, and also for user-dependent case scenarios. Our results demonstrate that artificial hydrocarbon networks classifier is flexible enough to be used when building a human activity recognition system with either user-dependent or user-independent approaches. PMID:27792136

A Flexible Approach for Human Activity Recognition Using Artificial Hydrocarbon Networks.

PubMed

Ponce, Hiram; Miralles-Pechuán, Luis; Martínez-Villaseñor, María de Lourdes

2016-10-25

Physical activity recognition based on sensors is a growing area of interest given the great advances in wearable sensors. Applications in various domains are taking advantage of the ease of obtaining data to monitor personal activities and behavior in order to deliver proactive and personalized services. Although many activity recognition systems have been developed for more than two decades, there are still open issues to be tackled with new techniques. We address in this paper one of the main challenges of human activity recognition: Flexibility. Our goal in this work is to present artificial hydrocarbon networks as a novel flexible approach in a human activity recognition system. In order to evaluate the performance of artificial hydrocarbon networks based classifier, experimentation was designed for user-independent, and also for user-dependent case scenarios. Our results demonstrate that artificial hydrocarbon networks classifier is flexible enough to be used when building a human activity recognition system with either user-dependent or user-independent approaches.

Differential Antifungal Activity of Human and Cryptococcal Melanins with Structural Discrepancies.

PubMed

Correa, Néstor; Covarrubias, Cristian; Rodas, Paula I; Hermosilla, Germán; Olate, Verónica R; Valdés, Cristián; Meyer, Wieland; Magne, Fabien; Tapia, Cecilia V

2017-01-01

Melanin is a pigment found in all biological kingdoms, and plays a key role in protection against ultraviolet radiation, oxidizing agents, and ionizing radiation damage. Melanin exerts an antimicrobial activity against bacteria, fungi, and parasites. We demonstrated an antifungal activity of synthetic and human melanin against Candida sp. The members of the Cryptococcus neoformans and C. gattii species complexes are capsulated yeasts, which cause cryptococcosis. For both species melanin is an important virulence factor. To evaluate if cryptococcal and human melanins have antifungal activity against Cryptococcus species they both were assayed for their antifungal properties and physico-chemical characters. Melanin extracts from human hair and different strains of C. neoformans ( n = 4) and C. gattii ( n = 4) were investigated. The following minimum inhibitory concentrations were found for different melanins against C. neoformans and C. gattii were (average/range): 13.7/(7.8-15.6) and 19.5/(15.6-31.2) μg/mL, respectively, for human melanin; 273.4/(125->500) and 367.2/(125.5->500) μg/mL for C. neoformans melanin and 125/(62.5-250) and 156.2/(62-250) μg/mL for C. gattii melanin. Using Scanning Electron Microscopy we observed that human melanin showed a compact conformation and cryptococcal melanins exposed an amorphous conformation. Infrared spectroscopy (FTIR) showed some differences in the signals related to C-C bonds of the aromatic ring of the melanin monomers. High Performance Liquid Chromatography established differences in the chromatograms of fungal melanins extracts in comparison with human and synthetic melanin, particularly in the retention time of the main compound of fungal melanin extracts and also in the presence of minor unknown compounds. On the other hand, MALDI-TOF-MS analysis showed slight differences in the spectra, specifically the presence of a minor intensity ion in synthetic and human melanin, as well as in some fungal melanin extracts. We

10th NTES Conference: Nickel and arsenic compounds alter the epigenome of peripheral blood mononuclear cells

PubMed Central

Brocato, Jason; Costa, Max

2014-01-01

The mechanisms that underlie metal carcinogenesis are the subject of intense investigation ; however, data from in vitro and in vivo studies are starting to piece together a story that implicates epigenetics as a key player. Data from our lab has shown that nickel compounds inhibit dioxygenase enzymes by displacing iron in the active site. Arsenic is hypothesized to inhibit these enzymes by diminishing ascorbate levels- an important co-factor for dioxygenases. Inhibition of histone demethylase dioxygenases can increase histone methylation levels, which also may affect gene expression. Recently, our lab conducted a series of investigations in human subjects exposed to high levels of nickel or arsenic compounds. Global levels of histone modifications in peripheral blood mononuclear cells (PBMCs) from exposed subjects were compared to low environmentally exposed controls. Results showed that nickel increased H3K4me3 and decreased H3K9me2 globally. Arsenic increased H3K9me2 and decreased H3K9ac globally. Other histone modifications affected by arsenic were sex-dependent. Nickel affected the expression of 2,756 genes in human PBMCs and many of the genes were involved in immune and carcinogenic pathways. This review will describe data from our lab that demonstrates for the first time that nickel and arsenic compounds affect global levels of histone modifications and gene expression in exposed human populations. PMID:24837610

A method for isolation of rat lymphocyte-rich mononuclear cells from lung tissue useful for determination of nucleoside triphosphate diphosphohydrolase activity.

PubMed

Jaques, Jeandre Augusto Dos S; Peres Rezer, João Felipe; Ruchel, Jader Betsch; Gutierres, Jessié; Bairros, André Valle; Gomes Farias, Iria Luiza; Almeida da Luz, Sonia Cristina; Mello Bertoncheli, Claudia de; Chitolina Schetinger, Maria Rosa; Morsch, Vera Maria; Leal, Daniela Bitencourt Rosa

2011-03-01

Methods for the isolation of peripheral blood mononuclear cells (PBMCs) and human lung mononuclear cells (LMCs) have been proposed previously. This study describes a method that allows the separation of lymphocyte-rich LMCs from rats. Trypan blue was applied to determine cell viability. White blood cell and differential cell counts were also performed. Relationships between nucleoside triphosphate diphosphohydrolase (NTPDase, EC 3.6.1.5) activities expressed in milligrams of protein, millions of cells, and millions of viable cells were examined as linear correlations. The lung tissue yielded 82.46% lymphocytes, 8.6% macrophages, 2.20% monocytes, and 1.27% polymorphonuclear cells (PMNs). In LMCs, a very strong correlation was observed as follows: between NTPDase activity, as determined using ATP or ADP as a substrate, expressed in milligrams of protein and that expressed in millions of cells (r ≥ 0.91), between that expressed in milligrams of protein and that expressed in millions of viable cells (r ≥ 0.91), and between that expressed in millions of cells and that expressed in millions of viable cells (r ≥ 0.98). Based on our results, we affirm that NTPDase activity could be expressed in millions of viable cells, millions of cells, or milligrams of protein. 2010 Elsevier Inc. All rights reserved.

Four new carbazole alkaloids from Murraya koenigii that display anti-inflammatory and anti-microbial activities.

PubMed

Nalli, Yedukondalu; Khajuria, Vidushi; Gupta, Shilpa; Arora, Palak; Riyaz-Ul-Hassan, Syed; Ahmed, Zabeer; Ali, Asif

2016-03-28

In our present study, four new, designated as murrayakonine A-D (), along with 18 known carbazole alkaloids were isolated from CHCl3 : MeOH (1 : 1) crude extracts of the stems and leaves of Murraya koenigii (Linn.) Spreng. The structures of the all isolated compounds were characterized by analysis of HR-ESI-MS and NMR (1D and 2D spectroscopy) results, and comparison of their data with the literature data. For the first time, all the isolates were evaluated for their anti-inflammatory activities, using both in vitro and in vivo experiments, against the key inflammatory mediators TNF-α and IL-6. The new compound murrayakonine A (), O-methylmurrayamine A () and mukolidine () were proven to be the most active, efficiently inhibiting TNF-α and IL-6 release in a dose-dependent manner and showing decreased LPS induced TNF-α and IL-6 production in human PBMCs. Furthermore, all the isolates were screened for their antimicrobial potential, and the compounds girinimbine () (IC50 3.4 μM) and 1-hydroxy-7-methoxy-8-(3-methylbut-2-en-1-yl)-9H-carbazole-3-carbaldehyde () (IC50 10.9 μM) displayed potent inhibitory effects against Bacillus cereus. Furthermore, compounds murrayamine J () (IC50 11.7 μM) and koenimbine () (IC50 17.0 μM) were active against Staphylococcus aureus. However, none of the compounds were found to be active against Escherichia coli or Candida albicans.

Human butyrylcholinesterase components differ in aryl acylamidase activity.

PubMed

Montenegro, María F; María, T Moral-Naranjo; de la Cadena, María Páez; Campoy, Francisco J; Muñoz-Delgado, Encarnación; Vidal, Cecilio J

2008-04-01

Apart from its esterase activity, butyrylcholinesterase (BuChE) displays aryl acylamidase (AAA) activity able to hydrolyze o-nitroacetanilide (ONA) and its trifluoro-derivative (F-ONA). We report here that, despite amidase and esterase sites residing in the same protein, in human samples depleted of acetylcholinesterase the ratio of amidase to esterase activity varied depending on the source of BuChE. The much faster degradation of ONA and F-ONA by BuChE monomers (G1) of colon and kidney than by the tetramers (G4) suggests aggregation-driven differences in the AAA site between single and polymerized subunits. The similar ratio of F-ONAto butyrylthiocholine hydrolysis by serum G1 and G4 forms support structural differences in the amidase site according to the source of BuChE. The changing ratios of amidase to esterase activities in the human sources probably arise from post-translational modifications in BuChE subunits, the specific proportion of monomers and oligomers and the variable capacity of the tetramers for degrading ONA and F-ONA. The elevated amidase activity of BuChE monomers and the scant activity of the tetramers justify the occurrence of single BuChE subunits in cells as a means to sustain the AAA activity of BuChE which otherwise could be lost by tetramerization.

Spinal Motion and Muscle Activity during Active Trunk Movements - Comparing Sheep and Humans Adopting Upright and Quadrupedal Postures.

PubMed