SEW2871 attenuates ANIT-induced hepatotoxicity by protecting liver barrier function via sphingosine 1-phosphate receptor-1-mediated AMPK signaling pathway.

Cholestatic liver injury, a group of diseases characterized with dysregulated bile acid (BA) homeostasis, was partly resulted from BA circulation disorders, which is commonly associated with the damage of hepatocyte barrier function. However, the underlying hepatocyte barrier-protective molecular mechanisms of cholestatic liver injury remain poorly understood. Interestingly, recent studies have shown that sphingosine-1-phosphate (S1P) participated in the process of cholestasis by activating its G protein-coupled receptors S1PRs, regaining the integrity of hepatocyte tight junctions (TJs). Here, we showed that SEW2871, a selective agonist of sphingosine-1-phosphate receptor 1(S1PR1), alleviated ANIT-induced TJs damage in 3D-cultured mice primary hepatocytes. Molecular mechanism studies indicated that AMPK signaling pathways was involved in TJs protection of SEW2871 in ANIT-induced hepatobiliary barrier function deficiency. AMPK antagonist compound C (CC) and agonist AICAR were all used to further identify the important role of AMPK signaling pathway in SEW2871's TJs protection of ANIT-treated mice primary hepatocytes. The in vivo data showed that SEW2871 ameliorated ANIT-induced cholestatic hepatotoxicity. Further protection mechanism research demonstrated that SEW2871 not only regained hepatocyte TJs by the upregulated S1PR1 via AMPK signaling pathway, but also recovered hepatobiliary barrier function deficiency, which was verified by the restored BA homeostasis by using of high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). These results revealed that the increased expression of S1PR1 induced by SEW2871 could ameliorate ANIT-induced cholestatic liver injury through improving liver barrier function via AMPK signaling and subsequently reversed the disrupted BA homeostasis. Our study provided strong evidence that S1PR1 may be a promising therapeutic approach for treating intrahepatic cholestatic liver injury. Graphical abstract.

Amelioration of non-alcoholic fatty liver disease by sodium butyrate is linked to the modulation of intestinal tight junctions in db/db mice.

The intestinal microenvironment, a potential factor that contributes to the development of non-alcoholic fatty liver disease (NALFD) and type 2 diabetes (T2DM), has a close relationship with intestinal tight junctions (TJs). Here, we show that the disruption of intestinal TJs in the intestines of 16-week-old db/db mice and in high glucose (HG)-cultured Caco-2 cells can both be improved by sodium butyrate (NaB) in a dose-dependent manner in vitro and in vivo. Accompanying the improved intestinal TJs, NaB not only relieved intestine inflammation of db/db mice and HG and LPS co-cultured Caco-2 cells but also restored intestinal Takeda G-protein-coupled (TGR5) expression, resulting in up-regulated serum GLP-1 levels. Subsequently, the GLP-1 analogue Exendin-4 was used to examine the improvement of lipid accumulation in HG and free fatty acid (FFA) co-cultured HepG2 cells. Finally, we used 16-week-old db/db mice to examine the hepatoprotective effects of NaB and its producing strain Clostridium butyricum. Our data showed that NaB and Clostridium butyricum treatment significantly reduced the levels of blood glucose and serum transaminase and markedly reduced T2DM-induced histological alterations of the liver, together with improved liver inflammation and lipid accumulation. These findings suggest that NaB and Clostridium butyricum are a potential adjuvant treatment strategy for T2DM-induced NAFLD; their hepatoprotective effect was linked to the modulation of intestinal TJs, causing the restoration of glucose and lipid metabolism and the improvement of inflammation in hepatocytes.

Sphingosine 1-phosphate receptor-1 specific agonist SEW2871 ameliorates ANIT-induced dysregulation of bile acid homeostasis in mice plasma and liver.

Dysregulated bile acid (BA) homeostasis is an extremely significant pathological phenomenon of intrahepatic cholestasis, and the accumulated BA could further trigger hepatocyte injury. Here, we showed that the expression of sphingosine-1-phosphate receptor 1 (S1PR1) was down-regulated by α-naphthylisothiocyanate (ANIT) in vivo and in vitro. The up-regulated S1PR1 induced by SEW2871 (a specific agonist of S1PR1) could improve ANIT-induced deficiency of hepatocyte tight junctions (TJs), cholestatic liver injury and the disrupted BA homeostasis in mice. BA metabolic profiles showed that SEW2871 not only reversed the disruption of plasma BA homeostasis, but also alleviated BA accumulation in the liver of ANIT-treated mice. Further quantitative analysis of 19 BAs showed that ANIT increased almost all BAs in mice plasma and liver, all of which were restored by SEW2871. Our data demonstrated that the top performing BAs were taurine conjugated bile acids (T-), especially taurocholic acid (TCA). Molecular mechanism studies indicated that BA transporters, synthetase, and BAs nuclear receptors (NRs) might be the important factors that maintained BA homeostasis by SEW2871 in ANIT-induced cholestasis. In conclusion, these results demonstrated that S1PR1 selective agonists might be the novel and potential effective agents for the treatment of intrahepatic cholestasis by recovering dysregulated BA homeostasis.