The Potential Role Of "Liver-gut" Axis In The Development Of Rat Model Of Nonalcoholic Fatty Liver Disease

Objective:In this study,a high-fat diet combined with lipopolysaccharide(LPS)was used to induce nonalcoholic fatty liver disease(NAFLD)in rats,to investigate the potential role of liver-gut axis in the development of NAFLD.Methods:1.Establishment of NAFLD model: Wistar rats were randomly divided into normal control group and NAFLD model group(high-fat diet+ LPS).The model group was given high-fat diet and intraperitoneal injection of LPS at the end of 4 weeks(w).The experiment lasted for 12 w.Changes in rat body weight,wet liver weight,serum transaminase,and blood lipids were observed at the end of 6,8,10,and 12 w,respectively.The histomorphological changes of the rat liver were observed in hematoxylin-eosin(HE)staining.2.The role of liver-gut axis in the development of NAFLD: The changes of liver and intestine related indexes in NAFLD model rats were observed during the development of NAFLD.The contents of malondialdehyde(MDA),superoxide dismutase(SOD)and glutathione(GSH)in rat liver homogenates were determined by commercial kits.Serum levels of D-lactic acid(D-LA)and tumor necrosis factor-α(TNF-α)were determined by ELISA kits.The endotoxin content in intestine and liver were determined by limulus lysate method.The contents of free fatty acids(FFA)in intestine and serum of rats were determined by ELISA kits.Cytochrome P450(CYP450)activity in rats liver was also determined by ELISA assay.Results:1.Compared with the normal control group,the weight and wet liver weight in NAFLD model group were significantly increased,the serum ALT level were significantly increased(P﹤0.01),and the AST level also increased to a certain extent.The liver pathological results showed that the number of lipid droplets in the liver of the NAFLD model group was significantly increased at the end of 10 weeks,and a large number of lipid vacuolar degeneration was observed in hepatocytes,which was consistent with the clinical manifestations of NAFLD.2.Compared with the normal control group,the MDA content in the liver tissues in NAFLD model group were significantly increased(P﹤0.01),while SOD and GSH content were significantly decreased(P﹤0.01),indicating that the liver of rats in NAFLD model group appeared oxidative stress injury.3.Compared with the normal control group,the serum D-LA content in NAFLD model group were significantly increased(P ﹤ 0.01),suggesting that the intestinal integrity and permeability of rats in NAFLD model group were damaged,causing D-LA,a metabolite of bacteria in the intestine,to enter the blood circulation.4.Compared with normal control group,the endotoxin content in liver in the NAFLD model group were significantly increased(P﹤0.01),and the content of serum TNF-α also were significantly increased(P﹤0.05).It is possible that high levels of endotoxin caused excessive activation of liver immune cells,causing the release of the pro-inflammatory factor TNF-α,and exacerbating oxidative stress and inflammatory damage in the liver.5.Compared with the normal control group,the FFA content in the intestines and serum in NAFLD model group was significantly increased(P﹤0.01),indicating that the intestine and liver of the NAFLD model group had lipid metabolism disorders,which caused the increase of FFA production in vivo.6.Compared with the normal control group,the content of CYP450 in the liver in NAFLD model group were significantly increased(P﹤0.05),suggesting that the occurrence and development of NAFLD in rats may change the content of liver CYP450,which may affect the metabolism of drugs,fatty acids and other substances.Conclusion:High-fat diet combined with LPS induced significant oxidative stress response in the liver in the development of NAFLD model in rats.Meanwhile,the integrity and permeability of the intestine were changed by detecting markers of intestinal barrier.Damage to the intestinal barrier may contribute to increased levels of endotoxin in the liver.This study found that FFA levels in the intestine and serum in the NAFLD model group were significantly increased,indicating that lipid metabolism in rats was disordered.Increased levels of liver endotoxin and lipid metabolism disorders in rat liver may contribute to the release of pro-inflammatory factor TNF-α,which exacerbates oxidative stress and inflammatory damage.These results suggest that the interaction between liver and intestine may play an important role in the formation and occurrence of NAFLD in rats.