Protective Effect Of Salidroside Against CCl₄-induced Liver Injury Through Resist Oxidative Stress In Mice

Rhodiola,known as"plateau ginseng",has the functions of anti-hypoxia,anti-oxidation,anti-fatigue,anti-lactic acid content increase and liver protection.Its main active ingredient salidroside has many pharmacological effects,such as protecting nerve and cardiovascular system,protecting liver and liver,regulating metabolism,increasing immune system function and intervening aging.The antioxidant effect of salidroside has been confirmed,but whether the protective effect of salidroside on liver injury in mice through antioxidant effect has not been reported.In this study,salidroside was applied to mice with liver injury induced by carbon tetrachloride,and its protective effect and antioxidant mechanism were studied in order to develop a natural hepatoprotective drug.The experiment was divided into blank group,model group,salidroside group,salidroside prevention group and glycyrrhizin prevention group.The mice were injected with 60 mg/kg salidroside intraperitoneally for 14 d.Then the mice were given 10 mL/kg1%CCl₄ oil solution intraperitoneally to induce liver injury.After 24 h,the mice were killed by anesthesia.11.25 mg/kg glycyrrhizin injection was used as positive drug.By comparing the body weight and liver index of mice,the pathological changes of liver were observed,the pathological sections of liver tissues were observed,the serum transaminase level was determined,and the hepatoprotective effect of salidroside was preliminarily determined.In addition,the structure and function of mitochondria were observed by observing the ultrastructure of liver and MMP,and the level of ROS in liver was measured to observe the level of ROS,the main culprit of oxidative stress.The basic biochemical indexes of liver homogenate were determined to observe whether the antioxidant system in mice was broken by ROS,and to further explore the antioxidant and hepatoprotective effects of salidroside.Finally,the expression of Gadd45a,Mapk7 and Rras2,the major downstream genes of each branch pathway of MAPK,was quantitatively determined by fluorescence.The expression of Gadd45a and Rras2 was semi-localized by immunohistochemical method.The antioxidant and hepatoprotective effects of salidroside were demonstrated from the basic molecular mechanism.The results are as follows:1 Preliminary study on protective effect of salidroside on acute liver injury induced by CCl₄ in miceThe results showed that there was no significant difference in liver index among groups.The liver of blank group and salidroside group was normal,red-brown,bright and soft.In the model group,the appearances of liver were slightly enlarged,the envelope was tense,the edge was blunt and round,and the gloss of the surface was decreased.There were obvious fatty degeneration,yellowish-brown spots,and the whole was yellowish-brown.In salidroside preventive group,the gloss of liver surface was slightly reduced and slightly steatosis was observed,which was significantly alleviated compared with model group.The liver of Ganlixin prevention group was slightly enlarged,dim in color,slightly steatosis and slightly yellowish-brown,which was better than that of model group.Pathological observation showed that there were typical pathological changes in the model group.More than 70%of the hepatocytes had severe steatosis,forming different size lipid droplets vacuoles,focal necrosis around the central vein,nuclear concentration,fragmentation or dissolution,and accompanied by a large number of inflammatory cell infiltration.Salidroside preventive group and glycyrrhizin preventive group can significantly reduce the symptoms of steatosis,and salidroside effect is better.The results of GOT and GPT in serum of mice showed that compared with model group,salidroside preventive group had significantly decreased enzymes and improved liver injury?P<0.05?,while glycyrrhizin reduced GPT level,and the effect was not as good as salidroside.2 Antioxidant and hepatoprotective effects of salidroside on CCl₄-induced acute liver injury in miceUltrastructural observation showed that the decrease of nuclear chromatin in the model group resulted in the decrease of electron density,organelle and some of the cytoplasm stained lightly.Mitochondria were swollen,cristae broken and some adventitia dissolved.Compared with model group,the structure of bilayer nuclear membrane was clear and complete,some mitochondrial cristae were broken,some endoplasmic reticulum were dilated,and the content density of cytoplasm was between salidroside prevention group and model group.Salidroside preventive group had regular organelle structure,neat mitochondrial cristae,intact membrane and slightly lower electron density than the blank group.The results of MMP and ROS showed that MMP decreased significantly and ROS increased significantly in the model group?P<0.05?.Compared with model group,salidroside preventive group and glycyrrhizin preventive group increased MMP and decreased ROS significantly?P<0.05?.Liver biochemical indicators showed that the SOD levels of model group and glycyrrhizin prevention group were significantly lower than those of blank group,and the SOD levels of salidroside prevention group were slightly higher than those of model group?P<0.05?.Salidroside pretreatment effectively inhibited the decline of CAT induced by the model.Salidroside also has a good effect on the recovery of GSH level.Salidroside can effectively reduce MDA production,but glycyrrhizin can not.3 Study on the basic molecular mechanism of salidroside's antioxidant and hepatoprotective effects on CCl₄-induced acute liver injury in miceReal-time fluorescence quantitative PCR and immunohistochemical results showed that salidroside could significantly reduce the expression of oxidative stress-activated Gadd45a,Mapk7 and Rras2 in the MAPK signaling pathway,which were 0.56,0.80 and0.49 times higher than those in the model group,respectively.The expression of Gadd45a was not decreased in the prophylaxis group,which was significantly different from that in the blank group?P<0.05?.Gadd45a positive protein was mainly expressed in the cytoplasm of hepatocytes around the central vein in the model group,while almost no other histones were expressed.Rras2 protein was expressed in patches in model group and glycyrrhizin prophylaxis group,and only a small amount in other groups.The optical density of positive proteins was counted by software.Gadd45a and Rras2 proteins were significantly expressed in model group,which were 2.67 and 3.31 times higher than those in high blank group,respectively.Salidroside restored these stress proteins to normal level.This experiment proved that salidroside can effectively resist carbon tetrachloride-induced acute liver injury in mice,protect the integrity of mitochondrial function and structure,reduce ROS production,regulate antioxidant genes related to MAPK pathway,and achieve antioxidant and hepatoprotective effects,but more in-depth specific mechanisms need to be further studied.