27-Hydroxycholesterol induces liver fibrosis via down-regulation of trimethylation of histone H3 at lysine 27 by activating oxidative stress; effect of nutrient interventions

Background: Chronic liver injury caused by activation of hepatic stellate cells (HSCs) is a key event in the development of liver fibrosis (LF). A high-cholesterol diet can prompt accumulation of free cholesterol in HSCs, which promotes HSC activation and progression of LF. Objective: 27-Hydroxycholesterol (27HC) is the most abundant cholesterol metabolite. Here, we investigated whether the HSC activation and LF induced by high cholesterol is caused by its metabolite 27HC, and whether TGF8 classical signaling were involved in these processes. Methods: In vitro, LX2 and HSC-T6 cells were used to explore the effects of 27HC on activation of HSCs, while LSECs were used to observe the effects of 27HC on capillarization. In vivo, zebrafish were used to assess the effect of 27HC on LF. Results: The cholesterol metabolite 27HC promoted the proliferation of HSCs and up-regulated expression of COL1 and alpha-SMA as well as CTGF and TIMP1. Also, 27HC up-regulated expression of Smad2/3 and phosphorylated Smad2/3 in HSCs. Furthermore, 27HC-induced up-regulation of COL-1, alpha-SMA, CTGF, and TIMP1 protein levels was inhibited by Smad2/3 knockout. In addition, 27HC down-regulated H3K27me3 by inhibition of EZH2 and promotion of UTX and JMJD3 expression via the TGF8 signaling, thereby inducing activation of HSCs. Notably, 27HC significantly aggravated the pathological damage induced by DEN, and induced deposition of collagen fibers in zebrafish liver. Folic acid (FA) and resveratrol (RES) both reduced 27HC-induced production of reactive oxygen species (ROS) and inhibited the effects of TGF8 signaling on EZH2, UTX, and JMJD3, thereby increasing H3K27me3, and finally jointly inhibiting LF. Conclusion: Cholesterol is metabolized to 27HC, which mediates activation of HSCs and onset of LF. Reduced expression of H3k27me3 by TGF8 signaling is crucial to 27HC-induced LF. FA and RES ameliorated activation of HSCs and LF by reducing 27HC-induced production of ROS and regulating of H3K27me3.