【作者】 姚晖； 姜晓刚； 梁磊； 梁颖； 黄志恩； 张继平； 邓虹珠；

【Author】 Yao Hui 1,Jiang xiaogang 2,Liang lei 3,Liang ying 1,Huang zhien 1,Zhang jiping 1,Deng Hongzhu 4,(1 The second people’s hospital of Foshan,Foshan 528000;2 Department of pathology,Jining medical college,Jining 272013;3 Department of Pharmacy,Guangzhou General Hospital of Guangzhou Military Commad,Guangzhou,510010;4 Traditional Medicine College of Southern Medical University,Guangzhou 510515,P.R.China)

【机构】 佛山市第二人民医院药剂科； 济宁医学院病理教研室； 广州军区广州总医院药剂科； 南方医科大学中医药学院；

【摘要】 目的:探讨哈蟆油(OR)对D-半乳糖所致雌性衰老大鼠肝脏组织p16、p21和cyclinD1蛋白表达的影响,进一步探讨OR延缓雌性大鼠机体衰老机制。方法SPF级SD雌性青年大鼠40只随机分为模型组(D-gal组)、维生素E(VE组)、哈蟆油高剂量组(OR-H组)、中剂量组(OR-M组)、低剂量组(OR-L组),每组8只,D-半乳糖颈背部皮下注42d,建立亚急性衰老模型。另取雌性青年大鼠8只,同样部位每日注射生理盐水,作为空白组。第3周开始灌胃给药,给药时间30天。给药结束后,免疫组化法检测衰老大鼠肝脏组织p16和p21蛋白的表达情况,免疫印迹法(Westernblotting)检测OR对雌性衰老大鼠肝脏组织cyclinD1表达的影响。结果雌性衰老大鼠肝脏组织免疫组化结果表明,p16和p21多为胞浆表达,弥漫性、灶性分布均有。D-gal组p16和p21阳性细胞积分与空白组比较升高,差异有显著性(P<0.01)。与D-gal组相比,OR-M和OR-L组,p16阳性细胞积分降低,差异有显著性(P值均<0.01),OR各剂量组p21阳性细胞积分降低,差异有显著性(P值均<0.01)。肝脏组织Westernblotting结果表明,D-gal组肝脏组织cyclinD1蛋白表达与空白组比较降低,差异有显著性(P<0.01)。OR各剂量组cyclinD1蛋白表达与D-gal组比较,表达均升高(P值均<0.01),OR-H组尤为明显。结论OR可降低肝脏组织p16、p21蛋白的高表达。同时显著提高细胞增殖正性调控因子cyclinD1蛋白的表达,促进衰老雌性大鼠肝细胞增殖。哈蟆油延缓雌性衰老作用可能通过调控肝脏p16,p21,cyclinD1信号通路来促进有关增殖调控蛋白表达发挥延缓衰老作用。更多还原

【Abstract】 Objective To investigate the expressions of liver tissue p16,p21 and cyclinD1 in aged female rats with Oviductus Ranae(OR) by D-galactose.Methods:40 male SD rats(SPF grade) were randomly divided into 5 equal groups,namely the OR-H group,OR-M group,OR-L group,VE group and D-gal group.The rats received subcutaneous injection of D-Galactose for 6 weeks to establish aging models.Another 8 rats were injected daily with normal saline (NS) to serve as the normal control group.From the third week of the experiment,the rats were given oral OR (in OR group) or Vitamin E (in VE group) till 30 days.After completion of drug administration,all the rats were sacrificed and p16 and p21 were detected in these groups liver tissues by immuno-histochemistry(IHC) and cyclinD1 was detected by Western-blotting method.Results:The result of IHC had shown the positive production of p16 and p21 were mainly multifocal located in endochylema of liver cell and their distribution were diffuse and focus.Positive cell integral of p16 and p21 in liver tissue of D-gal female aged rats were significantly increased than normal group (P<0.01).After taking OR,the integral of positive cell of p16 in OR-M and OR-L groups was significantly decreased than D-gal group(P<0.01).The integral of positive cell of p21 in OR-H,OR-M and OR-L groups was significantly decreased than D-gal group (P< 0.01).The result of Western-blotting had shown that expression of cyclinD1 protein in liver of D-gal female rats was decreased than normal group and the difference was significant (P<0.01).Expression of cyclinD1 in all OR groups was increased compared with D-gal group and the difference was significant (P <0.01).Expression of cyclinD1 in OR-H group was the highest in all OR groups.Conclusions:OR can decrease over-expression of p16 and p21 protein in liver.Meanwhile,OR can enhance expression of cyclinD1 in liver of female rats.OR can promote liver cell proliferation of aging female rats.Anti-aging effects of OR is possibly through regulation of p16,p21,cyclinD1 signaling pathways to promote expression of the proliferation regulatory protein and the signaling pathways play a role in delaying aging.更多还原