Th17 Cells, AIH and PBC

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From: The Liver Meeting® 2008 (http://www.aasld.org/Pages/Default.aspx)

Abstract: 1597 Year: 2008 Title: The induction of Th17 cells and reduction of regulatory T cells could participate in the onset of autoimmune hepatitis Category: J03. Adaptive Immunity Session: Inflammation and Immunity in Liver Injury Author(s): H. Ebinuma, S. Tada, K. Ojiro, H. Saito, T. Hibi, Internal Medicine, Keio University, Tokyo, JAPAN; T. Masuda, Internal Medicine, Nihonkoukan Hospital, Kawasaki, JAPAN; Date: Monday, November 03, 2008 Abstract: Background & Aim: The pathogenesis of autoimmune hepatitis (AIH) has not fully understood and this will make it difficult how to diagnose or treat. Although it has been reported that the frequency of regulatory T cells (Tregs) was decreased in the peripheral blood mononuclear cells (PBMC) of AIH patients, this association with the decrease of Tregs and disease activity was obscure. Recently, a subset of helper T cell, Th17 was newly proposed and has been reported to participate in the pathogenesis of some autoimmune or infectious disease. This study was performed to investigate the role of Tregs and Th17 cells in the PBMC and liver tissue and resolve the pathogenesis of AIH. Methods: The PBMC obtained from the patients with untreated AIH (n=6) or chronic hepatitis C (CH-C, n=26) were stained with anti-FOXP3 (eBioscience), IL17 (eBioscience), CD4, and CD25. The formalin-fixed tissues of liver needle biopsy obtained from the patients with untreated AIH (n=28) were stained with anti-FOXP3 (eBioscience) or IL17 (R & D), and the number of their positive staining per 1 visual field was counted. These frequencies in PBMC or the positive number in liver specimen were compared among the patients' characteristics, especially between their types of onset. Result: The frequency of CD4+FOXP3+ cells in the PBMC of AIH patients was lower than that of CH-C or healthy control, while the frequency of CD4+IL17+ cells was higher. FOXP3+ cells in the liver specimen of AIH patients were observed in the regions with more inflammatory cell infiltration, while IL17+ cells were observed only in acute-onset AIH patients. Although there was no association between the number of FOXP3+ or IL17+ cells and the levels of ALT or IgG, the ratio of IL17+ to FOXP3+ was significantly larger in acute-onset group of AIH compared to that in chronic group of AIH. (Acute-onset vs Chronic: 1.08±2.56 vs 0.22±0.32, p=0.0415 Mann-Whitney's test) Conclusion: The induction of Th17 cells and the decrease of Tregs was thought to participate in the onset of AIH. ____________________________________

Abstract: 1755 Year: 2008 Title: Reciprocal changes of Tr1 and Th17 are involved in the pathogenesis of autoimmune liver diseases. Category: I02. Autoimmune Liver Disease Session: Autoimmune Liver Disease Author(s): H. Takahashi, M. Nakano, C. Saeki, T. Oikawa, Y. Kuniyasu, M. Zeniya, Gastroenterology and Hepatology, JIKEI University School of Medicine, Tokyo, JAPAN; Date: Tuesday, November 04, 2008 Abstract: Background and Aim: Recently, Th17 cell had been identified as a newly effecter cell which participates in autoimmune reaction at effecter phase. On the other hand, IL-10 producing Tr1 that is induced during inflammation may regulate autoimmune reaction at effecter phase. The aim of this study is to evaluate the status of Th17 and Tr1 in autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) to reveal their role in the pathogenesis of autoimmune liver diseases. Patients and Method: Clinically and pathologically diagnosed 16 AIH, 70 PBC and 38 healthy volunteers (NC) were enrolled. CD4 cells were obtained from peripheral blood by MACS. To evaluate IL-17 production, CD4 cells were activated by rIL-23 which stimulate Th17 selectively (50 ng/ml) or rIL-12 (1 ng/ml) with anti CD3 Ab (5 mg/ml) and anti CD28 Ab (1 mg/ml) for 72 hrs. To evaluate IL-10 production, CD4 cells were activated by anti CD28 Ab (10 mg/ml) or anti CD46 Ab which stimulate IL-10 production of Tr1 selectively (5 mg/ml) and rIL-2 (50 IU/ml) with anti CD3 Ab (10 mg/ml) for 72 hrs. IL-17 and IL-10 in supernatant were assayed by ELISA. Result: IL-17 production stimulated by rIL-23 was significantly higher in AIH and PBC (2317±643, 1953±1049 vs. 1226±535, p<0.01) compared with NC. There was no significant difference of IL-17 production stimulated by rIL-12. In AIH, a positive correlation between ALT and IL-17 was observed (r=0.39). Though there was no correlation between ALP and IL-17 in PBC, ALP level of the patients who produce IL-17 more than two-fold by rIL-23 stimulation compared with rIL-12 stimulation was significantly higher than the patients who produce less IL-17 by rIL-23 stimulation compared with rIL-12 stimulation (387±175 vs. 248±92, p<0.05). IL-10 production stimulated by anti CD46 Ab was significantly lower in AIH and PBC (2717±801, 3189±1516 vs. 5250±1334 pg/ml, p<0.01) compared with NC. In AIH, a significant negative correlation between ALT and IL-10 was observed (r=-0.57, p<0.05). There was no correlation between ALP and IL-10 in PBC, The levels of IL-17 and IL-10 production of AIH patients who are treated with predonisolon or azathioprine were not different from those of patients who are treated with only UDCA. There was no correlation between IL-17 production and IL-10 production in any situations. Conclusion: Increased IL-17 production of CD4 cells stimulated by rIL-23 and decreased IL-10 production of CD4 cells stimulated by anti CD46 Ab were observed in AIH and PBC. These results indicated that reciprocal changes of Tr1 and Th17 participated in the pathogenesis of autoimmune liver diseases. ____________________________________

Abstract: 674 Year: 2008 Title: Preferential Hepatic Induction Of Th17 Cells In IL-2 Receptor á Knockout (IL-2Rá KO) Mice: A Murine Model Of Primary Biliary Cirrhosis (PBC) Category: I01. PBC/PSC Session: PBC/PSC Author(s): R.Y. Lan, Z. Lian, G. Yang, Y. Moritoki, M. Gershwin, Internal Medicine, University of California, , , CA; K. Tsuneyama, Graduate School of Medicine and Pharmaceutical Science, University of Toyama, Toyama, JAPAN; A.A. Ansari, Department of Pathology, Emory University School of Medicine, Atlanta, GA; R.L. Coppel, Department of Biochemistry and Molecular Biology, Monash University, Melbourne, , AUSTRALIA; Date: Saturday, November 01, 2008 Full Text: Unavailable. Abstract: PBC is characterized histologically by autoreactive CD4 and CD8 T cells surrounding damaged bile ducts. The cellular and molecular events which lead to such accumulation have not been defined, in part due to the difficulty in accessing and studying human liver. Recently, the role of IL-17 has been investigated in a number of autoimmune diseases, including rheumatoid arthritis, multiple sclerosis, and systemic lupus erythematosus. CD4+ T cells are a major source of IL-17, which compose a distinct T helper subset (Th17). IL-17 can induce the expression of several proinflammatory cytokines and chemokines such as IL-1, IL-6, TNF, GM-CSF and CXCL8 in a variety of target cells including epithelial cells, endothelial cells, fibroblasts, osteoblasts, and macrophages. However, the involvement of IL-17 in PBC remains to be clarified. IL-2 has recently been found to inhibit Th17 induction, as evidenced by the increased frequency of Th17 cells and serum IL-17 levels in IL-2 and IL-2 receptor deficient mice. We examined the IL-17 profile in the sera, livers, and spleens of IL-2Rá KO mice using ELISA, flow cytometric analysis, in vitro cultures of CD4+ T cells, and immunohistochemical staining. IL-2Rá KO mice exhibit a peak in serum IL-17 around 8-13 wks of age, which decreases over time. Histologically, marked aggregations of IL-17 positive cells were observed in portal tracts. Notably, the frequency of Th17 cells from IL-2Rá KO livers was found to be significantly higher than that from IL-2Rá KO spleens (20.39% ± 3.70 vs 6.36% ± 2.18, respectively, P<0.05). IL-2Rá KO liver CD4+ T cells produced significantly higher levels of IL-17 compared to their splenic counterparts (1204 pg/ml ± 87.82 vs 329.4pg/ml ± 59.24, P<0.0001). Interestingly, CD4+ T cells from livers of normal C57BL/6J (B6) mice also secreted higher levels of IL-17 relative to those from spleens (35.86 pg/ml ± 3.264 vs 8.61 pg/ml ± 2.120, respectively. P<0.001), indicating a preferential induction of Th17 cells in livers. CD4+ T cells from livers of IL-2Rá KO mice demonstrate a significantly greater ratio of IL-17 to IFN-Õ production compared to those from IL-2Rá KO spleens (6.8 ± 0.79 vs 1.4 ± 0.21, respectively. P<0.01), demonstrating a clear Th17 bias in livers compared to spleen. Importantly, splenic CD4+ T cells increased IL-17 production approximately 10 fold upon coculture with liver non-parenchymal cells in B6 mice (41.02 pg/ml ± 7.57 vs 508.2 pg/ml ± 39.70, respectively), indicating a role of the liver microenvironment in Th17 induction. ____________________________________ Abstract: 785 Year: 2008 Title: Biliary innate immunity contributes Th17 dominant milieu and constitutive chronic cholangitis in primary biliary cirrhosis Category: B03. Cholangiocyte Biology Session: Cholangiocyte Biology, Bile Formation, and Cholestasis Author(s): K. Harada, Y. Sato, K. Isse, H. Ikeda, M. Enomae, Y. Nakanuma, Human Pathology, Kanazawa University, Kanazawa, JAPAN; Date: Sunday, November 02, 2008 Abstract: Aims and purpose: Primary biliary cirrhosis (PBC) is characterized by the immunopathologic destruction of interlobular bile ducts. Recently, in addition to two different types of CD4-positive helper T cells, namely Th1 and Th2, the third pathogenic helper cells, Th17 cells, are newly classified, and the association with the chronic inflammation of autoimmune diseases is noted. Human Th17 cells characterized by the production of primarily interleukin (IL)-17 are induced from Th0 by the presence of IL-6 and IL-1â and maintained by IL-23. In this study, we examined the role of Th17 cells in the mechanisms of pathogenesis of PBC. Methods: By using cultured human biliary epithelial cells (BECs), the expression of Th17-related cytokines (IL-6, IL-1â, IL-23 p19, IL-23/IL-12 p40), chemokines, and receptors for IL-17, and the alterations of these cytokines and chemokines by treatments with pathogen-associated molecular patterns (PAMPs) via Toll-like receptors (TLRs) were examined in cultured BECs by RT-PCR and real-time PCR. Moreover, the intrahepatic distribution of IL-17-positive cells and the expression of IL-6, IL-1â, and IL-23 p19 in bile ducts were examined by immunohistochemistry. Results: The expression of IL-6, IL-1â, IL-23 p19, and IL-23/IL-12 p40 mRNAs was low or not detected in cultured BECs by PCR, but induced or enhanced by treatments with PAMPs such as LPS (TLR4 ligand) and Pam3CSK4 (TLR1/2 ligand). Moreover, cultured BECs constantly expressed IL-17 receptors (IL-17RA and IL-17RC) and a treatment with IL-17 upregulated the expression of IL-6 and IL-1â, but not IL-23 p19 and IL-23/IL-12 p40. Immunohistochemistry revealed that IL-17-positive infiltrating cells were present within the inflammed portal area in PBC and control diseased livers. In particular, in PBC, IL-17-positive infiltrating cells were dense around the damaged interlobular bile ducts. IL-6, IL-1â, and IL-23 p19 were expressed in biliary epithelial cells in intrahepatic bile ducts. Especially, the expression of IL-6 and IL-1â was enhanced in damaged bile ducts of PBC. In contrast, IL-23 p19 was constantly expressed in intrahepatic bile ducts, irrespective of PBC and control diseased livers. Conclusion: This study potentially provides that Th17 cells are associated with the continuous inflammation of PBC and the induction of Th17 cells is closely associated with the biliary innate immune response.