苦参素对2型糖尿病雄性大鼠生殖损伤及Nrf2/HO-1信号通路的影响

Effects of oxymatrine on reproductive injury and Nrf2/HO-1 signaling pathway in type 2 diabetic male rats

关键词:  苦参素  2型糖尿病  生殖  睾丸  氧化应激  Nrf2/HO-1信号通路

Key words:  oxymatrine  type 2 diabetes mellitus  reproduction  testicle  oxidative stress  Nrf2/HO-1 signaling pathway

摘要:

[目的] 探讨苦参素（OMT）对2型糖尿病（T2DM）雄性大鼠生殖损伤及E2相关因子2（Nrf2）/血红素加氧酶1（HO-1）信号通路的影响。[方法] 采用高脂饮食结合腹腔注射链脲佐菌素的方法制备T2DM雄性大鼠模型，设正常（Normal）组、模型（Model）组、二甲双胍（Met）组和OMT低、中、高剂量组，每组8只。Met组每日1次灌胃给药200 mg/kg，OMT低、中、高剂量组分别每日1次灌胃给药25、50、100 mg/kg，Normal组和Model组每日1次灌胃给予生理盐水，疗程4周。检测空腹血糖（FBG）和血清睾酮（T）、促性腺激素释放激素（GnRH）、促黄体激素（LH）、卵泡刺激素（FSH）水平，测量睾丸质量和睾丸体积，苏木精-伊红（HE）染色法观察睾丸组织病理学改变，测量输精管直径（STD）和输精管上皮厚度（TSE），精液分析仪检测精子数量、精子存活率和精子活力，比色法检测睾丸组织抗氧化酶（SOD、GSH-Px）活性和丙二醇（MDA）含量，实时荧光定量聚合酶链式反应（RT-PCR）法检测E2相关因子2（Nrf2）、血红素加氧酶-1（HO-1） mRNA表达，蛋白免疫印迹法（Western blot）检测Nrf2、p-Nrf2、HO-1蛋白表达。[结果] 与Model组比较，Met组和OMT低、中、高剂量组FBG水平降低，血清T、GnRH、LH、FSH水平升高（P＜0.05）；睾丸质量和睾丸体积升高（P＜0.05）；睾丸组织病理学改变明显改善，STD和TSE升高（P＜0.05）；精子数量、精子存活率、（a+b）级活力精子升高（P＜0.05）；SOD、GSH-Px活性升高，MDA含量降低（P＜0.05）；Nrf2、HO-1 mRNA表达量升高，Nrf2、p-Nrf2、HO-1蛋白表达量升高（P＜0.05）。OMT低、中、高剂量组上述作用呈剂量依赖性（P＜0.05）。除FBG、GSH-Px外，OMT高剂量组对其他指标的影响优于Met组（P＜0.05）。[结论] OMT对T2DM雄性大鼠生殖损伤具有保护作用，其机制可能与激活Nrf2/HO-1信号通路，抑制氧化应激，改善下丘脑-垂体-性腺轴功能有关。

Abstract:

[Objective] To investigate the effects of oxymatrine（OMT） on reproductive injury and E2 related factor 2（Nrf2）/heme oxygenase-1（HO-1） signaling pathway in type 2 diabetic（T2DM） male rats. [Methods] The T2DM rat model was prepared by high-fat diet combined with intraperitoneal injection of streptozotocin. And the normal group，model group，metformin（Met） group and OMT low-dose，medium-dose，high-dose groups were set up，with 8 rats in each group. The Met group was given 200 mg/kg by intragastric administration once a day；the OMT low，medium，high dose groups were given 25，50 and 100 mg/kg by intragastric administration once a day；and the normal group and model group were given normal saline intragastric administration once a day；and the course of treatment was 4 weeks. The levels of fasting blood glucose（FBG） and serum testosterone（T），gonadotropin-releasing hormone（GnRH），luteinizing hormone（LH），follicle-stimulating hormone（FSH） in serum were detected；the testicular mass and volume were measured；the histopathological changes of testicle were observed by HE staining，the seminiferous tubules diameter（STD） and thickness of the seminiferous epithelium（TSE） were measured. The sperm count，sperm survival rate and sperm motility were detected through semen analyzer. The activity of antioxidant enzymes（SOD and GSH-Px） and the content of MDA in testicular tissue were detected by colorimetry. The expressions of Nrf2，HO-1 mRNA were detected by RT-PCR；the expressions of Nrf2，p-Nrf2，HO-1 protein were detected by Western blot. [Results] Compared with the model group，the level of FBG in Met group and OMT low，medium，high dose groups were decreased，and the level of T，GnRH，LH，FSH in serum were increased（P＜0.05）；the testicular mass and volume were increased（P＜0.05）. The testicular histopathological changes were significantly improved，and the STD，TSE were increased（P＜0.05）. The sperm count，sperm survival rate，sperm motile of（a+b） grade were increased（P＜0.05）. The activity of SOD，GSH-Px were increased，while the content of MDA was decreased（P＜0.05）. The expression of Nrf2，HO-1 mRNA were increased，and the expressions of Nrf2，p-Nrf2，HO-1 protein were increased（P＜0.05）. The effects of OMT low，medium，high dose groups were dose-dependent（P＜0.05）. Except for FBG and GSH-Px，the effect of OMT high dose group on other indexes were better than those of Met group（P＜0.05）. [Conclusion] OMT has protective effect on reproductive injury in T2DM male rats，and its mechanism may be related to the activation of Nrf2/HO-1 signaling pathway，inhibition of oxidative stress，and improvement of hypothalamic-pituitary-gonadal axis function.