Loss of fibrocystin promotes interleukin-8-dependent proliferation and CTGF production of biliary epithelium

Abstract Background & Aims Congenital hepatic fibrosis (CHF) is a genetic liver disease with abnormal proliferation of cholangiocyte and progressive hepatic fibrosis. Mutation of polycystic kidney and hepatic disease 1 ( PKHD1 ) causes CHF and the dysfunction of its coding protein, fibrocystin, is associated with ductal plate malformation during fetal development and further pathological changes in CHF patients. However, the underlying molecular mechanism of CHF remains unclear, which is quite different from liver cirrhosis. This study investigated the molecular mechanism of CHF-pathophysiology using a genetically engineered human induced pluripotent stem (iPS) cell model to aid the discovery of novel therapeutic agents for CHF. Methods PKHD1-deficient (PKHD1-KO) and heterozygously mutated PKHD1 iPS clones were established by RNA-guided genome editing using CRISPR/Cas9 system. The iPS clones were differentiated into cholangiocyte-like cells in cysts (cholangiocytic cysts, CCs) in a 3D-culture system. Results The CCs were composed of monolayer cholangiocyte-like cells. The proliferation of PKHD1-KO CCs was significantly increased by interleukin-8 (IL-8) secreted in an autocrine manner. IL-8 production was significantly elevated in PKHD1-KO CCs due to mitogen-activated protein kinase pathway activation caused by fibrocystin deficiency. The production of connective tissue growth factor (CTGF) was also increased in PKHD1-KO CCs in an IL-8-dependent manner. Furthermore, validation analysis demonstrated that both the serum IL-8 level and the expression of IL-8 and CTGF in the liver samples were significantly increased in CHF patients, consistent with our in vitro human iPS-disease model of CHF. Conclusions Loss of fibrocystin function promotes IL-8-dependent proliferation and CTGF production of human cholangiocytes, suggesting that IL-8 and CTGF are essential for the pathogenesis of CHF. IL-8 and CTGF are candidate molecular targets for the treatment of CHF. Lay summary: Congenital hepatic fibrosis (CHF) is a genetic liver disease due to the mutation of PKHD1 gene and the dysfunction of its coding protein, fibrocystin, is closely associated with CHF pathogenesis. This study showed that the loss of fibrocystin function promotes proliferation and connective tissue growth factor (CTGF) production of cholangiocytes in an IL-8-dependent manner, using a disease in vitro model of human induced pluripotent stem (iPS) cells. The expressions of IL-8 and CTGF in the liver were significantly promoted in CHF patients consistent with the results of iPS cell-model, suggesting that IL-8 and CTGF are essential for the pathogenesis of CHF.