Hepatic NFAT signaling regulates the expression of inflammatory cytokines in cholestasis

The inflammatory response plays an important role in cholestatic liver injury where bile acid (BA) induction of proinflammatory cytokines in hepatocytes is an initial pathophysiologic event. However, the signaling pathways involving BA stimulation of cytokine production remain to be elucidated. In this report, we examined the functional role of the Nuclear Factor of Activated T-cells (NFAT) in BA-induction of inflammatory genes in hepatic cells and cholestatic livers. We found that NFAT isoform c1 and c3 were expressed in human and mouse hepatocytes. When treated with cholestatic levels of BA, both human and mouse hepatocytes but not cholangiocytes increased NFATc3 nuclear translocation, associated with elevated mRNA levels of IL-8, Cxcl2, and Cxcl10 in these cells. Blocking NFAT activation with pathway-specific inhibitors (i.e. cyclosporine A, FK-506, KN-62 and Inca-6) or knocking down Nfatc3, significantly repressed BA-induction of these cytokines in mouse hepatocytes, including Ccl2, Cxcl2, Cxcl10, Icam1 and Egr1. Nuclear expression of NFATc3/Nfatc3 protein was also increased in cholestatic livers after bile duct ligation or in Abcb4-/- mice and in patients with primary biliary cholangitis and primary sclerosing cholangitis in association with tissue elevations of Cxcl2 and IL-8. Gene reporter assays and ChIP-PCR demonstrated that the NFAT response element in its promoter played a key role in BA-induced human IL-8 expression. Together our findings indicate that NFAT plays an important role in BA stimulation of hepatic cytokines in cholestasis and is a mechanism that may provide novel targets to reduce cholestatic liver injury.