Toxic effects of Tripterygium glycoside tablets on the reproductive system of male rats by metabolomics, cytotoxicity, and molecular docking.

BACKGROUND: Tripterygium glycoside tablets (TGT) is the most common preparation from Tripterygium wilfordii Hook F, which is widely used in clinical for treating rheumatoid arthritis (RA) and other autoimmune diseases. However, its serious reproductive toxicity limits its application. PURPOSE: This study aimed to elucidate the toxic effects of TGT on the reproductive system of male RA rats and its potential toxic components and mechanism. METHODS: Collagen-induced arthritis (CIA) rat model was established, and TGT suspension was given at low, medium, and high doses. Gonadal index, pathological changes, and the number of spermatogenic cells were used to evaluate the toxic effects of TGT on the reproductive system. Non-targeted metabolomics of testicular tissue was conducted by UHPLC-QTOF/MS. Combined with network toxicology, the key targets of TGT-induced reproductive toxicity were screened and RT-qPCR was used to validation. In vitro toxicity of 19 components of TGT was evaluated using TM3 and TM4 cell lines. Molecular docking was used to predict the interaction between toxic components and key targets. RESULTS: TGT reduced testicular and epididymis weight. Pathology analysis showed a lot of deformed and atrophic spermatogenic tubules. The number of spermatogenic cells decreased significantly (P<0.0001). A total of 58 different metabolites including platelet-activating factor (PAF), lysophosphatidylcholine (Lyso PC), phosphatidylinositol (PI), glutathione (GSH), and adenosine monophosphate (AMP) were identified by testicular metabolomics. Glycerophospholipid metabolism, ether lipid metabolism, and glutathione metabolism were key pathways responsible for the reproductive toxicity of TGT. Ten key reproductive toxicity targets were screened by network toxicology. The cytotoxicity test showed that triptolide, triptonide, celastrol, and demethylzeylasteral could significantly reduce the viability of TM3 and TM4 cells. Alkaloids had no apparent toxic effects. Molecular docking showed that the four toxic components had a good affinity with 10 key targets. All binding energies were less than -7 kcal/mol. The RT-qPCR results showed the Cyp19a1 level was significantly up-regulated. Pik3ca and Pik3cg levels were significantly down-regulated. CONCLUSION: Through testicular metabolomics, we found that TGT may cause reproductive toxicity through CYP19A1, PIK3CA, and PIK3CG three target, which was preliminarily revealed. This study laid the foundation for elucidating the toxicity mechanism of TGT and evaluating its safety and quality.

Effects of total flavonids of astragalus on arrhythmia,endoplasmic reticulum stress in mice with viral myocarditis / 中国应用生理学杂志

OBJECTIVE@#To investigate the effects of total flavonids of astragalus(TFA) on arrhythmia, endoplasmic reticulum stress and connexcin in mice with viral myocarditis and to clarify the mechanisms of TFA against viral myocarditis complicated with arrhythmia.@*METHODS@#Thirty-six male Balb/c mice were randomly divided into control group, viral myocarditis group and total flavonoids group (=12). The mice of viral myocarditis were intraperitonealy injected with 0.1 ml/day 10-950 TCID CVB3 for 3 days. The mice of TFA group were intraperitoneal injected with 0.1 ml/day 10-950 TCID CVB3 for 3 days and treated with 0.1ml, 20 mg/L TFA by tail vein injection. At the end of the experiment, arrhythmia was detected by electrocardiogram, the heart of mice were stained by HE, the expressions of glucose-regulated protein 78(GRP78), endoplasmic reticulum stress signaling pathway factor activating transcription factor 4(ATF4) and connexcin 43(Cx43) were detected by Western blot.@*RESULTS@#The expressions of GRP78 and ATF4 were increased and the expression of Cx43 was decreased in viral myocarditis, while TFA inhibited these effect of viral myocarditis in heart of mice.@*CONCLUSIONS@#The antiarrhythmic effect of TFA may be related to the alleviation of endoplasmic reticulum stress and the increase of Cx43 expression.

Effect of Astragalus injection on cardiomyocyte apoptosis, endoplasmic reticulum stress and expression of connexin in cardiomyopathy rats induced by adriamycin / 中国应用生理学杂志

OBJECTIVES@#To investigate the effect of Astragalus injection on cardiomyocyte apoptosis, endoplasmic reticulum stress and connexin protein in cardiomyopathy rats induced by adriamycin.@*METHODS@#Thirty-six male Wister rats were randomly divided into control group (=12), adriamycin(ADR) group (=12) and Astragalus group (=12). The normal saline (10 ml/kg body weight) was injected intraperitoneally in control group rats, ADR (2 mg/kg body weight) was injected intraperitoneally in ADR group rats, ADR (10 ml/kg body weight) and Astragalus injection (10 ml/kg body weight) were injected intraperitoneally in rats of astragalus group, one time a week, totle 3 times. By the end of the 7 week, the left ventricular end-diastolic diameter(LVEDD), left ventricular end-systolic diameter (LVESD) and left ventricular ejection fraction (LVEF) were measured by echocardiography. Then the rats in the three groups were sacrificed and the left ventricle section was stained by HE, Masson, uranyl acetate/lead citrate respectively, the cardiomyopathy and ultrastructural changes were observed under light microscope and transmission electron microscope. The apoptosis of rat cardiomyocyte were analyzed by TUNEL. The expression of connexin Cx43 and p-Cx43 was detected by immunohistochemistry. The expression of glucose-regulated protein 78 (Grp78),activating transcription factor 4 (ATF-4) and C/EBP homologous protein (CHOP) were detected by real time PCR.@*RESULTS@#Compared with control group, LVEDD, LVESD increased and LVEF decreased, myocardial fibers were disordered and edematous, infiltrated by lymphocytes, the mitochondria were destroyed and vacuolized, and the number of cardiomyocyte apoptosis was increased(<0.01) in ADR group. The expression of Grp78, ATF-4, CHOP and p-Cx43 were increased, and the expression of Cx43 was decreased in ADR group. However, compared with ADR group, LVEDD, LVESD decreased and LVEF increased, the cardiomyopathy and ultrastructural changes were significantly improved, the number of cardiomyocyte apoptosis was significantly decreased (<0. 01); the expression of Grp78, ATF-4, CHOP and p-Cx43 decreased (<0.01); the expression of Cx43 increased in Astragalus group (<0.01).@*CONCLUSIONS@#Astragalus injection may effectively improve the myocardial damage induced by adriamycin, its mechanism may be related to the inhibition of endoplasmic reticulum stress (ERS) and the decrease of phosphorylation of CX43 in cardiomyopathy rats induced by adriamycin.