YTHDF2 influences hepatic fibrosis by regulating ferroptosis in hepatic stellate cells by mediating the expression of ACSL4 in an m 6A-dependent manner.

Abstract

Hepatic fibrosis (HF) is an abnormal reparative response of the liver to chronic injury and is histologically reversible. In recent years, increasing interest has been given to changes in m ⁶A in liver disease. In this study, we explore the role of the m ⁶A-modified reading protein YTHDF2 in HF and its regulatory mechanism. The HF mouse model is generated through CCl ₄ injection, and the cell model is via TGF-β stimulation. The liver tissues are subjected to hematoxylin-eosin, Masson, and α-SMA immunohistochemical staining. Reactive oxygen species (ROS) and iron levels are examined via relevant kits. Quantitative real-time PCR, immunofluorescence staining, and western blot analysis were conducted to measure the YTHDF2 and ACSL4 levels. RNA immunoprecipitation, methylated RNA immunoprecipitation, RNA pull-down, and polysome fractionation were performed to understand the regulatory mechanism by which YTHDF2 affects ACSL4. The results show that YTHDF2 is highly expressed after HF induction, and the inhibition of YTHDF2 reduces fibrosis as well as ROS and iron levels. In vitro, overexpression of YTHDF2 increases hepatic stellate cell activation, as well as ROS and iron levels, and this effect is blocked by the silencing of ACSL4. YTHDF2 acts as a regulator of ACSL4 expression and is involved in m ⁶A modification. In addition, in vivo experiments indicate that overexpression of ACSL4 reverses the attenuating effect of YTHDF2 interference on HFs. Therefore, YTHDF2 mediates the expression of the ferroptosis marker protein ACSL4 in an m ⁶A-dependent manner, thereby affecting HF.