中文摘要：

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和细胞周期蛋白D1蛋白表达的影响，进一步探讨哈蟆油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D-半乳糖组、维生素E组、哈蟆油高剂量组、中剂量组、低剂量组，每组8只，D-半乳糖颈背部皮下注射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，作为空白对照组。第15天开始灌胃给药，给药时间28d。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，蛋白免疫印迹法检测哈蟆油对雌性衰老大鼠肝脏组织细胞周期蛋白D1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-半乳糖组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-半乳糖组相比，哈蟆油中、低剂量组，p16阳性细胞积分降低，差异有显著性（P值均〈0．01），哈蟆油各剂量组p21阳性细胞积分降低，差异有显著性（P值均〈0．01）。肝脏组织蛋白免疫印迹法结果表明，D-半乳糖组肝脏组织细胞周期蛋白D1蛋白表达与空白组比较降低，差异有显著性（P〈0．01）。哈蟆油各剂量组细胞周期蛋白D1蛋白表达与D-半乳糖组比较，表达均升高（P值均〈0．01），哈蟆油高剂量组尤为明显。结论哈蟆油可降低肝脏组织p16、p21蛋白的高表达。同时显著提高细胞增殖正性调控因子细胞周期蛋白D1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，细胞周期蛋白D1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。

英文摘要：

OBJECTIVE To investigate the effect of Oviductus Ranae （OR） on the expression of p16, p21 and cyclinD1 protein in liver tissue of D-galactose-induced aging female rats and to further explore the anti-aging mechanism of OR in female rats. METH- ODS Fourty young female SD rats （ SPF grade） were randomly divided into 5 groups including OR-H group, OR-M group, OR-L group, VE group, and D-gal group with 8 in each group. The 40 rats received subcutaneous injection of D-galactose on the nape for 42 d to establish aging model. Another 8 rats were injected daily with normal saline （NS） at the same position as the normal control group. From the 15th day of the experiment,the rats were given oral OR （in OR group） or Vitamin E （in VE group） for 28 d. Then all the rats were sacrificed, p16 and p21 in liver tissue were detected by immuno-histochemistry （IHC）, and cyclinD1 was detected by West- ern-blotting method. RESULTS The result of IHC showed that p16 and p21 mainly multifocally located in endochylema of liver cells and their distribution was diffuse and focus. Positive cell integral of p16 and p21 in the liver tissue of D-gal female aged rats was signif- icantly increased than in normal control group （P 〈 0. 01 ）. After taking OR, the integral of p16- positive cell in OR-M and OR-L groups was significantly decreased than in D-gal group （P 〈 0. 01 ）. The integral of p21-positive cell in OR-H, OR-M and OR-L groups was significantly decreased than D-gal group （P 〈 0. 01 ）. The result of Western-blotting showed that the expression of cyclinD1 protein in the liver tissue of D-gal female rats was significantly decreased than in normal control group （ P 〈 0. 01 ）. The expression of cyclinD1 in all OR groups was significantly increased compared with D-gal group （ P 〈 0. 01 ）. The expression of cyclinD1 in OR-H group was the highest in all OR groups. CONCLUSION OR can decrease the over-expression of p16 and p21 protein in liver tissue. Meanwhile, OR can enhance the expression o