The Hepatotoxicity And Mechanism Of XianLingGuBao In Sprague-Dawley Rats

XianLingGuBao(XLGB)compound is an herbal formula that has been used in clinical settings to treat orthopaedic diseases,included in the National Essential Drugs List.Since XLGB goes on sale,the therapeutic effect has been remarkable.However,its increasing reports of adverse reactions of liver damage have also attracted widespread attentions.However,there are still few studies about preclinical XLGB-induced liver injury and its hepatoxic mechanism.Given to the dimness of the characteristics,causes and mechanism of XLGB hepatoxicity,it is urgent to carry out related researches to provide experimental evidence for safety supervision and risk control of XLGB and similar prescription drugs.The 6-week-old Sprague-Dawley(SD)rats,elderly female SD rats and immune-stress rats were used to evaluate the hepatotoxicity of XLGB,and explore the toxicity mechanisms on oxidative stress and bile acid metabolism.The details are as follows:The 6-week-old SD rats were repeatedly given XLGB for 8 weeks,the highest dose equivalent to 10 times of the clinical dose.However,there was no significant change in body weights and serum biochemistry,and no drug-related lesions in pathological examination.The 12～13-month-old SD female rats were used to simulate the elderly women in the characteristic of physiological and metabolism process.The rats were repeated intragastric administration of XLGB for 90 days for studying the toxicity of XLGB,the highest dose of XLGB equivalent to 20 times of the clinical dose.The body weights and food intakes of the drug-administered group were significantly lower than the CON group;There was no drug-related difference in organ weights.There was no statistical difference between groups in serum biochemistry and hematology;the gross pathological examination showed that XLGB could promote the disappearance of thymuses,however,there was no drug-related lesions in histopathological examination.SD male rats were induced immunological stress by LPS to increase their sensitivities to the drugs,hepatotoxicity.Based on the hepatotoxicity of XLGB,the SD male rat with immune stress was optimized for the evaluation of drug hepatotoxicity.The stimulation dose of LPS was determined to be 0.1 mg/kg,and two administrations before and after LPS was used to increase the incidence of drug-induced liver injury.What’s more,the defined administration time and animal body weights,prolonged animal adaptation time were used to reduce the difference between the batches of the experiments.8-week-old SD male rats with LPS stimulation were used for acute hepatotoxicity evaluation of XLGB.The results showed that XLGB could significantly increase the liver weight,liver coefficient,serum concentration of transaminase,total bilirubin and triglyceride in immune stress rats.Histopathological assessment further showed that extensive hepatocellular necrosis and inflamnatory infiltration were evident in rats with XLGB/LPS co-treatment,and the incidence and level of lesion are dependent to the dose of XLGB.The immunofluorescent assessment showed extensive apoptosis in hepatocytes from these co-treated rats,and the immunohistochemical assessment showed the Kupffer cells and hepatic stellate cell activation in liver from these co-treated rats.Through the mechanism studies,it is suggested that oxidative stress plays an important role in the acute liver injury induced by XLGB in rats.The increased expression of oxidase induced by XLGB and immune stimulation makes contribution to the process of liver injury.The results showed that XLGB could increase the expression of NADPH oxidase and iNOS and CYP2E1 in rats.Combined with the immune stimulation of LPS,XLGB can induce acute liver injury in SD rats.Antioxidant drugs can be used to reduce the oxidative damage of liver in the treatment of acute XLGB-induced liver injury.6-week-old SD male rats administrated XLGB for 14 days and with LPS stimulation were used for repeated hepatotoxicity evaluation of XLGB.The results showed that XLGB could significantly increase serum concentration of transaminase,total bilirubin and triglyceride in immune stress rats.Histopathological assessment further showed that extensive hepatocellular necrosis and inflammatory infiltration were evident in rats with XLGB/LPS co-treatment,and the incidence and level of lesion are dependent to the dose of XLGB.Given to the changes of lipid metabolism induced by repeated administration of drugs,the metabolism of bile acids was studied.The mechanism of XLGB repeated hepatotoxicity was deduced form effects of XLGB or LPS to the transcription and the translation of rate-limiting enzyme of bile acid biosynthesis,the regulator and transporter of bile acid in rats.XLGB could promote the expression of bile acid synthesis rate-limiting enzyme CYP7A1 and inhibit the expression of bile acid negative feedback regulator FXR.The expression of NF-Kb which can lock the promoter of FXR wasinduced by LPS,and the transporters of bile acids were depressed by LPS.The biosynthesis and metabolism of bile acid were disordered by XLGB and LPS,which made contributions to the liver damage.In addition to antioxidant therapy,the cholestasis drugs were added to the treatment of repeated administration XLGB-induced liver injury to reduce the toxicity of bile acids to the liver.In this study,the hepatotoxicity of XLGB was studied in increasing the dose of XLGB,using old or immune stress rats.An animal model with liver injury induced by XLGB was established for the first time,and the toxic mechanism of acute and repeated liver injury induced by XLGB was explored.It was preliminarily proved that the oxidative stress and disorder of bile acid metabolism induced by XLGB and LPS play an important role in XLGB-induced liver injury in immune stress rats.