[Mechanism of Kehuang Capsules in ameliorating metabolic-associated fatty liver disease in rats by inhibiting ferroptosis].

This study aimed to investigate the mechanism by which Kehuang Capsules ameliorate metabolic-associated fatty liver disease(MAFLD) in rats through inhibiting ferroptosis. Thirty-six SPF-grade Wistar rats were randomly divided into six groups: a normal group, a normal administration group(0.3 g·kg~(-1)), a model group, low-dose and high-dose groups of Kehuang Capsules(0.3 and 0.6 g·kg~(-1)), and a Dongbaogantai group(0.1 g·kg~(-1)). The normal and normal administration groups were fed a standard diet, while the model, Kehuang Capsules, and Dongbaogantai groups were fed a high-fat diet. Starting from the 13th week of modeling, the corresponding drug treatment was given. The normal and model groups were given equal volumes of distilled water by gavage. After four weeks of continuous intervention, the rats were sacrificed, and serum and liver tissue samples were collected for subsequent index determination. The levels or activities of total cholesterol(TC), triglyceride(TG), catalase(CAT), superoxide dismutase(SOD), reduced glutathione(GSH), and malondialdehyde(MDA) in serum and liver were detected by biochemical methods. Serum levels or activities of low-density lipoprotein cholesterol(LDL-C), high-density lipoprotein cholesterol(HDL-C), aspartate aminotransferase(AST), and alanine aminotransferase(ALT) were also detected by biochemical methods. The levels of interleukin(IL)-1β, IL-6, and tumor necrosis factor alpha(TNF-α) in serum and liver were detected using enzyme-linked immunosorbent assay(ELISA). The levels of total iron and ferrous ions in the liver were determined by colorimetry. Hematoxylin-eosin(HE), Oil Red O, and Prussian blue staining were performed to assess liver histopathology, lipid deposition, and iron deposition, respectively. Western blot was used to detect the expression of ferroptosis-related proteins, including glutathione peroxidase 4(GPX4), solute carrier family 7 member 11(SLC7A11), acyl-coenzyme A synthetase long-chain family member 4(ACSL4), transferrin receptor 1(TFR1), and ferroportin 1(FPN1). The results show that Kehuang Capsules can significantly reduce the levels of TC, TG, IL-1β, IL-6, TNF-α, and MDA in serum and liver, while increasing the levels of CAT, SOD, and GSH. The levels and activities of LDL-C, AST, and ALT in serum are significantly decreased, while HDL-C levels are increased. The contents of total iron ions and ferrous ions in the liver are also decreased. HE staining results reveal varying degrees of improvement in liver cord arrangement and cell structures across all treatment groups. Oil red O staining results show that the red lipid droplets in the liver are decreased, and the color becomes lighter in each treatment group. Prussian blue staining results show a marked reduction in iron particles in hepatocytes and alleviated iron deposition. The protein expression levels of GPX4 and SLC7A11 in liver tissue are significantly increased, while TFR1 expression is significantly decreased. The protein expression of ACSL4 is decreased in the low-dose group of Kehuang Capsules, while no significant change is observed in the high-dose group. In addition, no significant change is observed in FPN1 protein expression across all treatment groups. In conclusion, Kehuang Capsules may alleviate hepatic steatosis and inflammatory injury in MAFLD rats by regulating iron metabolism and oxidative stress to inhibit ferroptosis.