| ObjectiveTo study the effects of different doses of Jianpi Shugan decoction on liver function,blood lipids,liver histopathological changes,ultrastructure of hepatocyte mitochondria and mitochondrial related functional factors TFAM and AIF in NASH rats induced by high fat diet,and to explore the action target and mechanism of Jianpi Shugan decoction at subcellular level.Methods30 male SD rats were randomly divided into normal group(n=5),model group(n=5),low dose group(n=5),middle dose group(n=5),high dose group(n=5)and Essentiale group(n=5).Except for the normal group,the other 5 groups were fed with high-fat diet(88%basic diet+2%cholesterol+10%lard)to induce rat NASH model.Each treatment group was given drugs during the modeling period.The drug concentration was 31.5 g/kg in the high dose group,15.75 g/kg in the middle dose group,7.875 g/kg in the low dose group and 137.3 mg/kg in the Essentiale group,and deionized water was given to the normal group and the model group according to their body weight once a day.The animals were killed after 8 weeks of modeling.The serum was taken to detect liver enzymes and blood lipids,while the liver tissues were made into paraffin sections and frozen sections.The HE staining sections were used to observed pathological changes of liver tissues and the oil red O staining sections were used to observed the lipid deposition of hepatocytes.The liver tissues were also made into electron microscope sections to observe the ultrastructure of liver cells and count the number of mitochondria.Besides,the liver tissues were taken for Western Blot detection of TFAM and AIF.Results1.General condition:The hair color of the model group was rough,the diet decreased,the stool was yellow and soft with smelly smell,and some rats in the model group were grumpy.There was no significant difference in body weight among all rats.The liver index of rats in the model group was significantly higher than that in the normal group(P<0.05).There was no difference in liver index between the other treatment groups and the model group.2.Serum enzyme:serum ALT and AST in the model group were significantly higher than those in the normal group(P<0.01),while serum ALT in the low and middle dose groups and Essentiale group were lower than those in the model group(P<0.05).Serum AST in each treatment group was lower than that in the model group(P<0.05),and the middle dose of Jianpi Shugan decoction group and Essentiale group reduced the serum AST more significantly(P<0.01).3.Blood lipids:serum TG,TC and LDL-C in the model group were significantly higher than those in the normal group(P<0.01),while serum HDL-C in the model group was significantly lower than that in the normal group(P<0.01).Serum TG in low-dose group,high-dose group and Essentiale group was significantly lower than that in model group(P<0.01).Serum TC in each traditional Chinese medicine treatment group and Essentiale group was lower than that in the model group(P<0.05).Serum LDL-C in the high dose group was lower than that in the model group(P<0.05),and serum LDL-C in the middle dose group and Essentiale group was significantly lower than that in the model group(P<0.01).Serum HDL-C in middle and high dose groups and Essentiale group were significantly higher than those in model group(P<0.01).In addition,the effect of improving serum HDL-C of the middle dose group and Essentiale group was significantly higher than that in the high dose group(P=0.00<0.01).4.Pathology:compared with the normal group,there were obvious hepatic steatosis,intralobular inflammation and hepatocyte balloon degeneration in the model group,and the total score of NAS was more than 5,which was significantly different from that in the normal group(P<0.01).The total score of NAS in each treatment group was significantly lower than that in the model group(P<0.01).The low,middle and high dose groups of Jianpi Shugan decoction could significantly reduce the scores of intrahepatic lobular inflammation and hepatocyte balloon degeneration(P<0.05),while Essentiale group could significantly reduce the scores of hepatic steatosis,intralobular inflammation and hepatocyte balloon degeneration(P<0.05).5.Ultrastructure and number of mitochondria:the morphology of hepatocytes in the normal group was normal,which the mitochondrial bilayer membrane was complete and clear and the mitochondrial crest was neatly arranged with the largest number.In the model group,hepatocytes enlarged and contained a large number of fat vesicles,the number of organelles such as mitochondria and endoplasmic reticulum decreased,mitochondria enlarged,the bilayer membrane blurred or disappeared,and the arrangement of mitochondrial crest was disordered.Lipid droplets were still deposited in each treatment group but with smaller number than that in the model group,which may be related to the division of hepatic acini.The number of mitochondria in the treatment group was higher than that in the model group(P>0.05),and the morphology of most mitochondria returned to normal.Part of the mitochondrial bilayer membrane structure in the treatment group is clear,but there were still a small number of mitochondrial bilayer membrane structure destruction,and mitochondrial crest arrangement disorder.The number of endoplasmic reticulum in the treatment group increased compared with the model group,and the structure is clearer.The lipid droplet deposition of hepatocytes in the Essentiale group was less than that in the model group and the Jianpi Shugan decoction group,and the number of mitochondria in the Essentiale group was significantly higher than that in the model group and other traditional Chinese medicine treatment groups(P<0.01).6.Mitochondrial functional factors:the expression of TFAM in the model group was lower than that in the normal group(P>0.05),while the expression of TFAM in the high dose group and Essentiale group was significantly higher than that in the model group(P<0.05).The expression of AIF in the model group was higher than that in the normal group(P>0.05),while the expression of AIF protein in the high dose group and Essentiale group was lower than that in the model group(P<0.05).Conclusions1.Jianpi Shugan decoction can significantly reduce the serum liver enzymes,blood lipids and the score of liver histopathology in NASH rats.There may be a dose-effect relationship of Jianpi Shugan decoction in improving liver histopathological,that is,the curative effect increases with the increase of drug dosage.2.Jianpi Shugan decoction can repair the ultrastructural damage of hepatocyte mitochondria,restore the structural integrity of mitochondrial bilayer membrane,improve the disorder of mitochondrial crest arrangement,reduce mitochondrial swelling,increase the number of normal mitochondria,restore the structure and number of endoplasmic reticulum and reduce endoplasmic reticulum stress in NASH rats.3.Jianpi Shugan decoction can regulate the production of mitochondria by up-regulating the expression of TFAM,reduce the abnormal outflow of AIF by regulating the opening of MPTP,and reduce the apoptosis induced by AIF by inhibiting the expression of AIF. |
PDF Full Text Request