Cyr61 Alleviates Cholangitis by Inhibiting Cytotoxic Effects of CD8⁺ T Cells on Biliary Epithelial Cells

Abstract

Objective

Primary biliary cholangitis (PBC) is a chronic progressive cholestatic liver disease. In recent years, researchers have found that cysteine-rich angiogenic inducer 61 (Cyr61, also known as CCN1) has a potential role in reducing portal inflammation in patients with PBC. This study aimed to explore the relationship between Cyr61 and PBC to provide new ideas and an experimental basis for the clinical treatment of PBC.

Methods

After induction of the overexpression of Cyr61 in a mouse model of PBC using recombinant adenovirus, hematoxylin and eosin staining and pathological scores were used to indicate intrahepatic inflammation and bile duct damage. Real-time PCR was used to detect changes in inflammation-related cytokines in the liver. To further study the mechanism, we assessed whether Cyr61 protects bile duct epithelial cells from cytotoxic effects.

Results

Serum and hepatic Cyr61 levels were increased in the murine model of PBC. Overexpression of Cyr61 alleviated hepatic inflammation and bile duct injury in vivo. Cyr61 inhibited the cytotoxic effects of CD8⁺ T cells by acting on biliary epithelial cells (BECs) in vitro.

Conclusion

Our results provide novel insight into the pathogenesis of PBC and suggest that Cyr61 plays a dominant role in the cytotoxic effects on BECs in PBC. Consequently, therapeutic strategies targeting Cyr61 could be a potent therapy for PBC.

Keywords

cysteine-rich angiogenic inducer 61 / primary biliary cholangitis / inflammation / bile duct damage / cytotoxicity

References

[1] YounossiZM, BernsteinD, ShiffmanML, et al.. Diagnosis and Management of Primary Biliary Cholangitis. Am J Gastroenterol, 2019, 114(1): 48-63

[2] KennedyL, FrancisH, InvernizziP, et al.. Secretin/ secretin receptor signaling mediates biliary damage and liver fibrosis in early-stage primary biliary cholangitis. FASEB J, 2019, 33(9): 10 269-10 279

[3] Beretta-PiccoliBT, Mieli-VerganiG, VerganiD, et al.. The challenges of primary biliary cholangitis: What is new and what needs to be done. J Autoimmun, 2019, 105: 102328

[4] HondaA, TanakaA, KanekoT, et al.. Bezafibrate Improves GLOBE and UK-PBC Scores and Long-Term Outcomes in Patients With Primary Biliary Cholangitis. Hepatology (Baltimore, Md.), 2019, 70(6): 2035-2046

[5] HirschfieldGM, GershwinME. The immunobiology and pathophysiology of primary biliary cirrhosis. Annu Rev Pathol, 2013, 8: 303-330

[6] NevensF, AndreoneP, MazzellaG, et al.. A Placebo-Controlled Trial of Obeticholic Acid in Primary Biliary Cholangitis. N Engl J Med, 2016, 375(7): 631-643

[7] TanakaA, TakikawaH, MochidaS, et al.. Changing Nomenclature for PBC from “Primary Biliary Cirrhosis” to “Primary Biliary Cholangitis”. J Gastroenterol, 2016, 51(7): 748-749

[8] JuL, SunY, XueH, et al.. CCN1 promotes hepatic steatosis and inflammation in non-alcoholic steatohepatitis. Sci Rep, 2020, 10(1): 3201

[9] TsudaM, AmbrosiniYM, ZhangW, et al.. Fine phenotypic and functional characterization of effector cluster of differentiation 8 positive T cells in human patients with primary biliary cirrhosis. Hepatology (Baltimore, Md.), 2011, 54(4): 1293-1302

[10] TwiggSM. Regulation and bioactivity of the CCN family of genes and proteins in obesity and diabetes. J Cell Commun Signal, 2018, 12(1): 359-368

[11] YangR, ChenY, ChenD. Biological functions and role of CCN1/Cyr61 in embryogenesis and tumorigenesis in the female reproductive system (Review). Mol Med Rep, 2018, 17(1): 3-10

[12] ParkYS, HwangS, JinYM, et al.. CCN1 secreted by tonsil-derived mesenchymal stem cells promotes endothelial cell angiogenesis via integrin alphav beta3 and AMPK. J Cell Physiol, 2015, 230(1): 140-149

[13] SchoberJM, ChenN, GrzeszkiewiczTM, et al.. Identification of integrin alpha(M)beta(2) as an adhesion receptor on peripheral blood monocytes for Cyr61 (CCN1) and connective tissue growth factor (CCN2): immediate-early gene products expressed in atherosclerotic lesions. Blood, 2002, 99(12): 4457-4465

[14] YangY, QiQ, WangY, et al.. Cysteine-rich protein 61 regulates adipocyte differentiation from mesenchymal stem cells through mammalian target of rapamycin complex 1 and canonical Wnt signaling. FASEB J, 2018, 32(6): 3096-3107

[15] MoonHG, QinZ, QuanT, et al.. Matrix protein CCN1 induced by bacterial DNA and CpG ODN limits lung inflammation and contributes to innate immune homeostasis. Mucosal Immunol, 2015, 8(2): 243-253

[16] FanY, YangX, ZhaoJ, et al.. Cysteine-rich 61 (Cyr61): a biomarker reflecting disease activity in rheumatoid arthritis. Arthritis Res Ther, 2019, 21(1): 123

[17] LinJ, LiN, ChenH, et al.. Serum Cyr61 is associated with clinical disease activity and inflammation in patients with systemic lupus erythematosus. Medicine, 2015, 94(19): e834

[18] WooYJ, SeoY, KimJJ, et al.. Serum CYR61 Is Associated with Disease Activity in Graves’ Orbitopathy. Ocul Immunol Inflamm, 2018, 26(7): 1094-1100

[19] ChoiJS, KimKH, LauLF. The matricellular protein CCN1 promotes mucosal healing in murine colitis through IL-6. Mucosal Immunol, 2015, 8(6): 1285-1296

[20] LöbelM, BauerS, MeiselC, et al.. CCN1: a novel inflammation-regulated biphasic immune cell migration modulator. Cell Mol Life Sci, 2012, 69(18): 3101-3113

[21] RotherM, KrohnS, KaniaG, et al.. Matricellular signaling molecule CCN1 attenuates experimental autoimmune myocarditis by acting as a novel immune cell migration modulator. Circulation, 2010, 122(25): 2688-2698

[22] ZhangH, LianM, ZhangJ, et al.. A functional characteristic of cysteine-rich protein 61: Modulation of myeloid-derived suppressor cells in liver inflammation. Hepatology (Baltimore, Md.), 2018, 67(1): 232-246

[23] WakabayashiK, LianZX, LeungPS, et al.. Loss of tolerance in C57BL/6 mice to the autoantigen E2 subunit of pyruvate dehydrogenase by a xenobiotic with ensuing biliary ductular disease. Hepatology (Baltimore, Md.), 2008, 48(2): 531-540

[24] YoshidaK, YangGX, ZhangW, et al.. Deletion of interleukin-12p40 suppresses autoimmune cholangitis in dominant negative transforming growth factor beta receptor type II mice. Hepatology (Baltimore, Md.), 2009, 50(5): 1494-1500

[25] GrazioliS, GilS, AnD, et al.. CYR61 (CCN1) overexpression induces lung injury in mice. Am J Physiol Lung Cell Mol Physiol, 2015, 308(8): L759-765

[26] MigitaK, IlyassovaB, KovzelEF, et al.. Serum BAFF and APRIL levels in patients with PBC. Clinical immunology (Orlando, Fla.), 2010, 134(2): 217-225

[27] MaWT, ChenDK. Immunological abnormalities in patients with primary biliary cholangitis. Clin Sci (Lond), 2019, 133(6): 741-760

[28] JuL, SunY, XueH, et al.. CCN1 promotes hepatic steatosis and inflammation in non-alcoholic steatohepatitis. Sci Rep, 2020, 10(1): 3201

[29] BianZ, PengY, YouZ, et al.. CCN1 expression in hepatocytes contributes to macrophage infiltration in nonalcoholic fatty liver disease in mice. J Lipid Res, 2013, 54(1): 44-54

[30] LiZQ, WuWR, ZhaoC, et al.. CCN1/Cyr61 enhances the function of hepatic stellate cells in promoting the progression of hepatocellular carcinoma. Int J Mol Med, 2018, 41(3): 1518-1528

[31] KimKH, ChenCC, AlpiniG, et al.. CCN1 induces hepatic ductular reaction through integrin αvβ5-mediated activation of NF-κB. J Clin Invest, 2015, 125(5): 1886-1900

[32] YouZ, WangQ, BianZ, et al.. The immunopathology of liver granulomas in primary biliary cirrhosis. J Autoimmun, 2012, 39(3): 216-221

[33] BenselerV, WarrenA, VoM, et al.. Hepatocyte entry leads to degradation of autoreactive CD8 T cells. Proc Natl Acad Sci USA, 2011, 108(40): 16 735-16 740

[34] ZhangW, ZhangR, ZhangJ, et al.. Proteomic analysis reveals distinctive protein profiles involved in CD8 T cell-mediated murine autoimmune cholangitis. Cell Mol Immunol, 2018, 15(8): 756-767

[35] YangGX, LianZX, ChuangYH, et al.. Adoptive transfer of CD8(+) T cells from transforming growth factor beta receptor type II (dominant negative form) induces autoimmune cholangitis in mice. Hepatology, 2008, 47(6): 1974-1982