TEC directly improved the LPS-induced insults on FXR and LXR in hepatocytes
The molecular basis for LPS-induced cholestasis has mainly been examined in hepatocytes and has been attributed to decreased expression of several transporters important for canalicular bile formation[34, 35]. Mechanically, it was previously reported that NFκb-p65 can bind to FXR and LXR promoters, inhibit the expression of FXR and LXR, and lead to the decreased expression of bile transporters, resulting in the retention of bile acids and phytosterol. Phytosterol would then further inhibit the activity of FXR[36]. Therefore, we investigated whether TEC could directly suppress LPS-induced activation of NFκb-p65 in hepatocytes. Primary hepatocytes were treated with LPS and/or TEC, and it was found that TEC reversed LPS-induced NFκb-p65 activation, as shown by decreased phosphorylation of NFκb-p65 and reduced NFκb-p65 binding to the FXR and LXR promoter (Figure 5A-C). In addition, TEC reversed the LPS-inhibited expression of FXR and LXR, as well as the expression of Bsep and ABCG5 (Figure 5D). To determine whether TEC regulates the expression and activity of FXR and LXR via PPARγ, hepatocytes were treated with LPS, shPPARγ and TEC as indicated in Figure 5E. The results showed that PPARγ expression deficiency in hepatocytes blocked the regulatory effects of TEC on NFκb-p65 activity, the expression of FXR, Bsep, LXR and ABCG5 (Figure 5E and 5F). Furthermore, when NFκb-p65 was inhibited, TEC treatment could not further enhance the expression of FXR and LXR as well as FXR-regulated and LXR-regulated genes in the presence of LPS (Figure S3A-C). Finally, FXR or LXR deficiency abolished the influence of TEC on the expression of Mrp2, ABCG5 and ABCG8, but not Bsep (Figure S4A-B). Combined with the results of Bsep in Figure 5D and 5F, this suggested that TEC regulated the expression of Bsep, which was partly dependent on FXR and completely relied on PPARγ.
TEC directly restored LPS-induced dysfunction of hepatocytes by activating PPARγ.