Protective Effect of Water-Soluble Acacetin Prodrug on APAP-Induced Acute Liver Injury Is Associated with Upregulation of PPARγ and Alleviation of ER Stress.

A water-soluble acacetin prodrug has been synthesized and reported by our group previously. Acetaminophen (APAP) overdose is a leading cause of acute liver injury. We found that subcutaneous injection of acacetin prodrug (5, 10, 20 mg/kg) decreased serum ALT, AST, and ALP, corrected the abnormal MDA and GSH in liver, and improved intrahepatic hemorrhage and destruction of liver structures in APAP (300 mg/kg)-treated mice. Molecular mechanism analysis revealed that the expressions of endoplasmic reticulum (ER) stress markers ATF6, CHOP, and p-PERK, apoptosis-related protein BAX, and cleaved caspase 3 were decreased by acacetin in a dose-dependent manner in vivo and in vitro. Moreover, via the acacetin-upregulated peroxisome-proliferator-activated receptor gamma (PPARγ) of HepG2 cells and liver, the suppressive effect of acacetin on ER stress and apoptosis was abolished by PPARγ inhibitor (GW9662) or PPARγ-siRNA. Molecular docking revealed that acacetin can bind to three active pockets of PPARγ, mainly by hydrogen bond. Our results provide novel evidence that acacetin prodrug exhibits significant protective effect against APAP-induced liver injury by targeting PPARγ, thereby suppressing ER stress and hepatocyte apoptosis. Acacetin prodrug is likely a promising new drug candidate for treating patients with acute liver injury induced by APAP.

The natural flavone acacetin protects against high-fat diet-induced lipid accumulation in the liver via the endoplasmic reticulum stress/ferroptosis pathway.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. To date, no medication has been approved to treat NAFLD. In this study, we evaluated the therapeutic effect of the natural flavone acacetin on high-fat diet (HFD)-induced NAFLD in mice and the underlying mechanisms. We found that acacetin (10, 20, 50 mg/kg/day) suppressed the increase in body weight, serum total cholesterol, triglycerides, low-density lipoprotein, aspartate aminotransferase, and alanine aminotransferase levels in mice fed with HFD with a dose-dependent manner. Hepatic lipid accumulation, iron overload, and lipid peroxidation were significantly alleviated by acacetin. Quantitative PCR and western blotting revealed that acacetin inhibited endoplasmic reticulum (ER) stress, ferroptosis, and expressions of lipid acid synthesis-related genes in the livers of HFD mice. Similar results were observed in HepG2 cells treated with oleic acid and lipopolysaccharide. The suppressive effects of acacetin on triglycerides and expression of lipid acid synthesis genes were abolished by ER stress and the ferroptosis activators, erastin or TU. Interestingly, the action of TU was more potent than that of erastin. Treatment with the ER stress inhibitor GSK and the ferroptosis inhibitor Fer-1 revealed that ER stress was the upstream signal of ferroptosis for hepatic lipid accumulation. These findings suggest the protective effect of acacetin against lipid accumulation via suppressing ER stress and ferroptosis and provide evidence that ER stress is an upstream signal of ferroptosis in lipid accumulation. Acacetin may be a promising candidate agent for NAFLD treatment.

Kir2.1 channel regulates macrophage polarization via the Ca2+/CaMK II/ERK/NF-κB signaling pathway.

Macrophage polarization plays a key role in the inflammatory response. Various ion channels expressed in macrophages have been documented, but very little is known about their roles in macrophage polarization. We found that knockdown or blockade of the Kir2.1 (also known as KCNJ2) channel significantly inhibited M1 macrophage polarization, but promoted M2 macrophage polarization. Lipopolysaccharide (LPS)-induced M1 polarization was also remarkably suppressed in high extracellular K+ solutions (70 mM K+), and this inhibition was partially abolished by adding Ca2+ to the culture medium. Ca2+ imaging showed that Ca2+ influx was dependent on the hyperpolarized membrane potential generated by the Kir2.1 channel. The upregulation of phospho (p)-CaMK II, p-ERK, and p-NF-κB proteins in macrophages from the RAW264.7 cell line that were stimulated with LPS was significantly reversed by blocking the Kir2.1 channel or culturing the cells with 70 mM K+ medium. Furthermore, in vivo studies showed that mice treated with a Kir2.1 channel blocker were protected from LPS-induced peritonitis. In summary, our data reveal the essential role of the Kir2.1 channel in regulating macrophage polarization via the Ca2+/CaMK II/ERK/NF-κB signaling pathway.