Mechanistic study on nonylphenol-induced liver fibrosis via Pink1/Parkin-mediated mitophagy and lipid droplet degradation in hepatic stellate cells

Background: Liver fibrosis (LF) is a common pathological feature in several chronic liver diseases. Nonylphenol (NP) accumulates in the liver and impairs its function. The mechanism by which NP exposure induces LF remains to be elucidated.

Objective: This study aimed to determine whether NP activates Pink1/Parkin-mediated Mitophagy to promote lipid droplet degradation in hepatic stellate cells, which then contributes to the development of LF.

Methods: Human hepatic stellate cells (LX-2) were categorized into six groups: control, Quizartinib (a Parkin inhibitor), NP, Quizartinib + NP, Mdivi-1 (a Mitophagy inhibitor), and Mdivi-1 + NP. Sixty male C57BL/6 mice were randomly assigned to six groups of 10 mice each: control (corn oil), low-dose NP (25 mg/kg), medium-dose NP (50 mg/kg), high-dose NP (100 mg/kg), Parkin knockout (KO, corn oil), and Parkin KO + NP (100 mg/kg). An additional four mice were set aside as a LF model group (Model, received 10 % CCl₄ intraperitoneally at 5 mL/kg, three times per week). All treatments were administered for 35 days.

Results: In vitro, NP exposure caused LX-2 cells to lose their stellate morphology and become elongated. Furthermore, treatment with 40 μM NP decreased the expression of the lipid droplet-coating protein Perilipin 5 (Plin5) and enhanced the expression of fibrosis markers (alpha-smooth muscle actin [α-SMA], Collagen Ⅰ) and mitophagy-related proteins (Pink1, Parkin, Beclin1, LC3 Ⅱ) (P < 0.05). However, the inhibition of Parkin or Mitophagy reversed NP-induced downregulation of Plin5 and upregulation of fibrosis markers and mitophagy-related proteins (P < 0.05). In vivo, NP exposure significantly increased the liver index and serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels in mice. However, Parkin KO reduced liver dysfunction (P < 0.05). As the NP dose increased, the Plin5 expression decreased in a dose-dependent manner, whereas fibrosis markers (α-SMA, Collagen Ⅰ) and mitophagy-related proteins (Pink1, Parkin, Beclin1, LC3 Ⅱ) increased. Nonetheless, Parkin KO mitigated the reduction of Plin5 and the elevation of fibrosis and Mitophagy markers (P < 0.05). NP exposure considerably augmented hepatic Collagen deposition in a dose-dependent manner, disrupted mitochondrial integrity, increased autophagosome numbers, and reduced the hepatic lipid droplet content. Nevertheless, Parkin KO reduced these pathological alterations (P < 0.05).

Conclusion: NP induces LF by activating Pink1/Parkin-mediated Mitophagy, which promotes lipid droplet degradation in hepatic stellate cells.

Hepatic stellate cells; Liver fibrosis; Mitophagy; Nonylphenol; Pink1/Parkin.