Intratracheal exposure to polyhexamethylene guanidine phosphate disrupts coordinate regulation of FXR-SHP-mediated cholesterol and bile acid homeostasis in mouse liver.

A public health crisis in the form of a significant incidence of fatal pulmonary disease caused by repeated use of humidifier disinfectants containing polyhexamethylene guanidine phosphate (PHMG) recently arose in Korea. Although the mechanisms of pulmonary fibrosis following respiratory exposure to PHMG are well described, distant-organ effect has not been reported. In this study, we investigated whether intratracheal administration of PHMG affects liver pathophysiology and metabolism. Our PHMG mouse model showed a significant decrease in liver cholesterol level. An mRNA-seq analysis of liver samples revealed an alteration in the gene expression associated with cholesterol biosynthesis and metabolism to bile acids. The expression of genes involved in cholesterol synthesis was decreased in a real-time PCR analysis. To our surprise, we found that the coordinate regulation of cholesterol and bile acid homeostasis was completely disrupted. Despite the decreased cholesterol synthesis and low bile acid levels, the farnesoid X receptor/small heterodimer partner pathway, which controls negative feedback of bile acid synthesis, was activated in PHMG mice. As a consequence, gene expression of Cyp7a1 and Cyp7b1, the rate-limiting enzymes of the classical and alternative pathways of bile acid synthesis, was significantly downregulated. Notably, the changes in gene expression were corroborated by the hepatic concentrations of the bile acids. These results suggest that respiratory exposure to PHMG could cause cholestatic liver injury by disrupting the physiological regulation of hepatic cholesterol and bile acid homeostasis.

SNX10-mediated degradation of LAMP2A by NSAIDs inhibits chaperone-mediated autophagy and induces hepatic lipid accumulation.

Rationale: While some non-steroidal anti-inflammatory drugs (NSAIDs) are reported to induce hepatic steatosis, the molecular mechanisms are poorly understood. This study presented the mechanism by which NSAIDs induce hepatic lipid accumulation. Methods: Mouse primary hepatocytes and HepG2 cells were used to examine the underlying mechanism of NSAID-induced hepatic steatosis. Lipid accumulation was measured using Nile-red assay and BODIPY 493/503. The activity of chaperone-mediated autophagy (CMA) was determined by western blotting, qRT-PCR, and confocal imaging. The effect of NSAID on CMA inhibition was evaluated in vivo using diclofenac and CMA activator (AR7) administered mice. Results: All tested NSAIDs in this study accumulated neutral lipids in hepatocytes, diclofenac having demonstrated the most potency in that regard. Diclofenac-induced lipid accumulation was confirmed in both mouse primary hepatocytes and the liver of mice. NSAIDs inhibited CMA, as reflected by the decreased expression of lysosome-associated membrane glycoprotein 2 isoform A (LAMP2A) protein, the increased expression of CMA substrate proteins such as PLIN2, and the decreased activity of photoactivatable KFERQ-PAmCherry reporter. Reactivation of CMA by treatment with AR7 or overexpression of LAMP2A inhibited diclofenac-induced lipid accumulation and hepatotoxicity. Upregulation of sorting nexin 10 (SNX10) via the CHOP-dependent endoplasmic reticulum stress response and thus maturation of cathepsin A (CTSA) was shown to be responsible for the lysosomal degradation of LAMP2A by diclofenac. Conclusion: We demonstrated that NSAIDs induced SNX10- and CTSA-dependent degradation of LAMP2A, thereby leading to the suppression of CMA. In turn, impaired CMA failed to degrade PLIN2 and disrupted cellular lipid homeostasis, thus leading to NSAID-induced steatosis and hepatotoxicity.