杞蛎还少方改善D-半乳糖致亚急性衰老小鼠性激素水平紊乱与睾丸氧化损伤的效用评价研究

Study on Efficacy Evaluation of Qili Huanshao Formula in Ameliorating Sex Hormone Disturbance and Oxidative Damage in Testicular of D-Galactose-Induced Subacute Aging Mice

目的 评价杞蛎还少方对D-半乳糖(D-galactose,D-gal)致亚急性衰老小鼠性激素水平紊乱与睾丸组织氧化损伤的改善作用与机制研究。方法 将105只雄性小鼠随机分成正常组,模型组,杞蛎还少方低、中、高剂量组,维生素E组和金匮肾气丸组,通过连续腹腔注射D-gal建立亚急性衰老模型小鼠,同时各组分别给予杞蛎还少方提取物、维生素E和金匮肾气丸。通过HE染色法检测睾丸组织病理损伤情况,ELISA法检测小鼠血清中相关性激素水平,试剂盒检测氧化应激因子表达,TUNEL法检测睾丸细胞凋亡,Western blot法检测小鼠睾丸组织中氧化应激与凋亡相关蛋白表达。结果 相较于衰老模型小鼠,杞蛎还少方各剂量组均能改善睾丸组织病理损伤;提高血清T、E_2、GnRH含量,降低LH、FSH含量(P<0.01);增强血清与睾丸组织中SOD活力(P<0.01)、GSH含量(P<0.01),降低MDA含量(P<0.01)。杞蛎还少方中剂量组能够显著抑制睾丸细胞凋亡(P<0.01),提高模型组小鼠睾丸组织Nrf2、HO-1蛋白表达及Bcl-2/Bax比值(P<0.05),下调Cleaved-Caspase-3/Caspase-3、Cleaved-Caspase-9/Caspase-9凋亡蛋白表达水平(P<0.05)。结论 杞蛎还少方可有效改善衰老小鼠的性激素紊乱,保护睾丸组织,其作用机制可能与抑制睾丸组织慢性氧化应激诱导的细胞凋亡有关。研究结果将为枸杞子及其组方临床应用与抗衰老功能性产品开发提供借鉴和奠定基础。

OBJECTIVE To evaluate the effect and mechanism of Qili Huanshao Formula(QLHSF)in ameliorating sex hormone disturbance and oxidative damage in testicular tissues of D-galactose-induced subacute aging mice.METHODS 105 male mice were randomly divided into the normal group,model group,low,medium and high doses of QLHSF group,vitamin E(VE)group and Jin Gui Shen Qi Wan(JGSQW)group.The subacute senescence model was established by continuous intraperitoneal injection of D-galactose(D-gal)and treated by intragastric administration,as well.The testicular histopathological damage was detected by HE staining.The levels of relevant sex hormones in serum were detected by ELISA.The expression of oxidative stress factors was detected by related kits.The apoptotic cells in testicular tissue were detected by TUNEL assay,and the expressions of oxidative stress and apoptotic related proteins in the testicular tissues of mice were detected by Western blot.RESULTS Compared to the aging model mice,all dose groups of QLHSF could obviously improve testicular tissue pathological damage.The levels of T,E 2 and GnRH levels were increased,while LH and FSH were decreased in serum(P<0.01).Meanwhile,the SOD activity(P<0.01)and the levels of GSH(P<0.01)were increased,while MDA levels(P<0.01)were decreased in serum and testicular tissues.The medium dose group of QLHSF significantly inhibited testicular cell apoptosis(P<0.01),increased the expression of Nrf2,HO-1 protein and Bcl-2/Bax ratio(P<0.05),and down-regulated the expressions of Cleaved-Caspase-3/Caspase-3,Cleaved-Caspase-9/Caspase-9 apoptotic protein in the testis of mice(P<0.05).CONCLUSION QLHSF can effectively improve sex hormone disturbance and protect testicular tissue in aging mice,and the mechanism of action might be related to the inhibition of oxidative stress-induced apoptosis in testicular tissues.The study will provide reference and lay the foundation for the clinical application and functional anti-aging product development of Lycium barbarum and its formulations.