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Interleukin-35 mitigates the function of murine transplanted islet cells via regulation of Treg/Th17 ratio.

ABSTRACT: Pancreatic islet transplantation is a promising treatment for type 1 diabetes (T1D). Interleukin-35 (IL-35) is a recently discovered cytokine that exhibits potent immunosuppressive functions. However, the role of IL-35 in islet transplant rejection remains to be elucidated. In this study, we isolated islet cells of BALB/c mouse and purified CD4+ T cell subsets of a C57BL/6 mouse. The model for islet transplantation was established in vitro by co-culture of the islet cells and CD4+ T cells. IL-35 (20 ng/ml) was administered every other day. Following co-culture, the islet function and Treg/Th17 ratio were analyzed on days 1, 3, and 5. Furthermore, the Th17/Treg ratio was modulated (1:0-2), and the function of islet cells as well as proliferation of Th17 cells were analyzed. T cell sorting was performed using the magnetic bead sorting method; Treg and Th17 count using flow cytometry; cell proliferation detection using the carboxyfluorescein diacetate succinimidyl ester (CFSE) method, and islet function test using the sugar stimulation test. Results showed that Th17 counts increased in the co-culture system. However, after administration of IL-35, the number of Treg cells increased significantly compared to that in the control group (50.7% of total CD4+ T cells on day 5 in IL-35 group vs. 9.5% in control group) whereas the proliferation rate of Th17 cells was significantly inhibited (0.3% in IL-35 group vs. 7.2% in control group on day 5). Reducing the Th17/Treg ratio significantly improved the function of transplanted islets. Treg inhibited Th17 proliferation and IL-35 enhanced this inhibitory effect. IL-35 mitigates the function of murine transplanted islet cells via regulation of the Treg/Th17 ratio. This might serve as a potential therapeutic strategy for in-vivo islet transplant rejection and T1D.

SUBMITTER: Zongyi Y

PROVIDER: S-EPMC5728515 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Interleukin-35 mitigates the function of murine transplanted islet cells via regulation of Treg/Th17 ratio.

Zongyi Yin Y Funian Zou Z Hao Li L Xin Wang W Ying Cheng C Jialin Zhang Z Yongfeng Liu L Baifeng Li L

PloS one 20171213 12

Pancreatic islet transplantation is a promising treatment for type 1 diabetes (T1D). Interleukin-35 (IL-35) is a recently discovered cytokine that exhibits potent immunosuppressive functions. However, the role of IL-35 in islet transplant rejection remains to be elucidated. In this study, we isolated islet cells of BALB/c mouse and purified CD4+ T cell subsets of a C57BL/6 mouse. The model for islet transplantation was established in vitro by co-culture of the islet cells and CD4+ T cells. IL-35 ...[more]

PMID: 29236782

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Project description:AIM:Atherosclerosis (AS) characterized as a chronic inflammatory disease. Multiple immune cells and inflammatory cytokines, such as high mobility group protein (HMGB1), regulatory T (Treg) cells, T helper (Th17) cells, and inflammation-related cytokines, play a key role in its pathophysiology. A large number of studies report that HMGB1 and Th17 cells may promote atherosclerosis progression, whereas Treg cells may play a protective role in atherosclerosis; thus, alterations in the Treg/Th17 ratio may exist in atherosclerosis diseases. Up till now, the relationships between HMGB1 levels and the Treg/Th17 ratio remain incompletely understood. The major purpose of this study was to investigate the relationship between HMGB1 levels and the Treg/Th17 ratio in patients with coronary artery atherosclerotic plaques. METHODS:We enrolled patients with coronary atherosclerosis and normal coronary artery as the research subjects. Flow cytometry was used to analyze the Treg cells, the Th17 cells frequency, and the Treg/Th17 ratio. Otherwise, real-time polymerase chain reaction was used for assays the mRNA expressions of HMGB1, retinoic acid-related orphan nuclear receptor C (RORC), and forkhead-winged helix transcription factor (Foxp3). Moreover, enzyme-linked immunosorbent assays were used to detect the level of protein and cytokines, such as HMGB1, IL-10, TGF-?1, IL-17A, and IL-23. RESULTS:Using flow cytometry, we observed a significantly increased of Th17 cell frequency, whereas Treg cell frequency significantly decreased in atherosclerotic patients. Consistently, the levels of RORC mRNA were significantly increased in coronary atherosclerosis (AS) group compared to normal coronary artery (NCA) group (P?0.01). In contrast, the expression of Foxp3 mRNA was markedly lower in the AS group than in the NCA group (P?0.01). Furthermore, we observed the serum concentrations of HMGB1, IL-17A, and IL-23 were significantly higher in the AS group than in the NCA group (P?0.01, respectively), whereas the concentrations of serum IL-10 and TGF-?1 were significantly lower in the AS group than in the NCA group (P?0.01, respectively). In addition, we also found that HMGB1 levels showed negative correlation with the Treg/Th17 ratio in the two groups (r=?0.6984, P?0.01). CONCLUSIONS:The data in our study indicated that HMGB1 may promote atherosclerosis progression via modulating the imbalance in the Treg/Th17 ratio.

Project description:ObjectiveTreg cells need to acquire an effector phenotype to function in settings of inflammation. Whether effector Treg cells can limit disease severity in lupus is unknown. Interferon regulatory factor 4 (IRF-4) is an essential controller of effector Treg cells and regulates their ability to express interleukin-10 (IL-10). In non-Treg cells, IRF-4 activity is modulated by interactions with DEF-6 and its homolog switch-associated protein 70 (SWAP-70). Although mice lacking both DEF-6 and SWAP-70 (double-knockout [DKO] mice) develop lupus, they display normal survival, suggesting that in DKO mice, Treg cells can moderate disease development. The purpose of this study was to investigate whether Treg cells from DKO mice have an increased capacity to become effector Treg cells due to the ability of DEF-6 and SWAP-70 to restrain IRF-4 activity.MethodsTreg cells were evaluated by fluorescence-activated cell sorting. The B lymphocyte-induced maturation protein 1 (BLIMP-1)/IL-10 axis was assessed by crossing DKO mice with BLIMP-1-YFP-10BiT dual-reporter mice. Deletion of IRF-4 in Treg cells from DKO mice was achieved by generating FoxP3(Cre) IRF-4(fl/fl) DKO mice.ResultsThe concomitant absence of DEF-6 and SWAP-70 led to increased numbers of Treg cells, which acquired an effector phenotype in a cell-intrinsic manner. In addition, Treg cells from DKO mice exhibited enhanced expression of the BLIMP-1/IL-10 axis. Notably, DKO effector Treg cells survived and expanded as disease progressed. The accumulation of Treg cells from DKO mice was associated with the up-regulation of genes controlling autophagy. IRF-4 was required for the expansion and function of effector Treg cells from DKO mice.ConclusionThis study revealed the existence of mechanisms that, by acting on IRF-4, can fine-tune the function and survival of effector Treg cells in lupus. These findings suggest that the existence of a powerful effector Treg cell compartment that successfully survives in an unfavorable inflammatory environment could limit disease development.

Project description:Introduction: The role of the immune response in the pathogenesis of antiphospholipid syndrome (APS) remains elusive. It is possible that differences in the frequencies of Th17 cells and/or defects in the immunoregulatory mechanisms are involved in the pathogenesis of APS. Our aim was to determine the peripheral blood Th cells phenotype and the circulating cytokine profile in patients with primary APS (pAPS) and compare it with systemic lupus erythemathosus (SLE) as disease control group. Methods: The frequencies of circulating regulatory T cells (Tregs) were determined in PBMCs from 36 patients with pAPS by flow cytometry. As control groups we included 21 age- and gender-matched healthy controls (HC) and 11 patients with SLE. The suppressive capacity of Tregs was evaluated in vitro by coculture assay. On the other hand, intracellular cytokine production was assessed in Th1, Th2, and Th17 cells and circulating IL-6, IL-10, and IL-35 were measured by Cytometric Bead Array and ELISA. The quantification of Th master gene expression levels was performed by real time quantitative PCR. Results: pAPS patients and SLE patients did not show differences in the percentage or number of Tregs compared to HC. The suppressive capacity of Tregs was also similar in the three study group. Instead, we found higher FoxP3·mRNA expression levels in pAPS patients and HC than SLE patients. Regarding the Th17 response, patients with pAPS and HC showed a significantly lower frequency of circulating Th17 cells than SLE. However, no differences were observed in the Th1 response between patients and controls. Thus, increased Th17/Th1 and Th17/Treg ratios were found in SLE patients but not in pAPS patients. pAPS and SLE patients had higher serum IL-6 levels than HC but there was not difference between both disease groups. Besides, a significant increase in the immunosuppressive cytokine levels was observed only in pAPS as compared to HC. Conclusions: Our data demonstrate an increased inflammatory profile of peripheral blood CD4+ T cells from SLE as compared with pAPS mostly due to an increased Th17 response. In conclusion, there seems not to be a direct pathogenic role for Th cells in pAPS but in SLE.

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Interleukin-35 mitigates the function of murine transplanted islet cells via regulation of Treg/Th17 ratio.

Publications

Interleukin-35 mitigates the function of murine transplanted islet cells via regulation of Treg/Th17 ratio.

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