补肾填精中药仙子益真胶囊对卵巢早衰模型小鼠卵巢PI3K/AKT通路的影响

Effect of Shen-tonifying Traditional Chinese Herbs Xianzi Yizhen Capsules on PI3K/AKT Signaling Pathway of Mice with Premature Ovarian Failure

目的研究补肾填精中药仙子益真胶囊对卵巢早衰小鼠磷酯酰肌醇3激酶(PI3K)、蛋白激酶B(AKT)、Ki67蛋白表达及各级卵泡数量的影响。方法选用动情周期规律的57只SPF级雌性C57BL/6J型小鼠随机分为空白对照组(7只)、模型组(50只)。模型组使用雷公藤多苷片造模,将成功造模的35只小鼠随机分为模型组、益真低剂量组[仙子益真胶囊0. 75 g/(kg·d)]、益真中剂量组[仙子益真胶囊1. 5 g/(kg·d)]、益真高剂量组[仙子益真胶囊3. 0 g/(kg·d)]和西药组[戊酸雌二醇片0. 15 mg/(kg·d)],每组7只。给予相应药物治疗,治疗21天后采用HE染色法检测卵巢组织的形态变化,计算各级卵泡数目,免疫组化方法检测Ki67蛋白表达,Western blot法检测小鼠卵巢AKT及PI3K蛋白表达。结果与空白对照组比较,模型组原始卵泡、次级卵泡、窦状卵泡百分比降低,Ki67表达、AKT及PI3K蛋白表达降低(P <0. 05,P <0. 01),闭锁卵泡百分比升高(P <0. 01)。与模型组比较,益真低、中、高剂量组及西药组原始卵泡、次级卵泡、窦状卵泡百分比升高,闭锁卵泡百分比降低(P <0. 05,P <0. 01),Ki67表达、AKT及PI3K蛋白表达增高(P <0. 05)。结论补肾填精中药仙子益真胶囊改善卵巢功能的机制可能与提高了卵巢细胞中PI3K/AKT通路活性及细胞增殖状态相关,从而增加了原始卵泡、次级卵泡、窦状卵泡的含量,降低了闭锁卵泡的含量。

Objective To explore the effect of Shen-tonifying traditional Chinese herbs Xianzi Yizhen（ XZYZ） Capsules on the express of phosphatidynositol 3 kinase（ PI3 K）,protein kinase B（ AKT）,Ki67 protein and follicle numbers at all levels of mice with premature ovarian failure. Methods Totally 57 female C57 BL/6 J mice of SPF level with regular estrous cycles were randomly divided into blank-control group（ n = 7） and model group（ n =50）. After the premature ovarian failure model was established by Glucosidorum Tripterygll Totorum,the modeled mice were divided into placebo group,low-dose XZYZ group[XZYZ capsules 0.75 g/（ kg · d） ],medium-dose XZYZ group [XZYZ capsules 1. 5 g/（ kg·d） ],and high-dose XZYZ group [XZYZ capsules 3. 0 g/（ kg·d） ] and Western medicine group [Estradiol Valerate tablets 0. 15 mg/（ kg·d） ]. Each intervention group contains 7 mice. After 21 days of duration,follicle numbers at all levels were counted by HE staining of ovarian tissues,the express of Ki67 was tested by immunohistochemistry methord and the express of AKT and PI3 were detected by Western blot method. Results The percentage of primordial follicle,secondary follicle and antral follicle of placebo group were lower than those of the blank-control group（ P〈0. 05）,as well as the express of Ki67,AKT and PI3 protein（ P〈0. 01）. The percentage of atretic follicle of placebo group was higer than that of the blank-control group（ P〈0. 01）.The percentage of primordial follicle,secondary follicle and antral follicle of intervention groups were higher than thoseof the placebo group（ P〈0. 05）,as well as the express of Ki67,AKT and PI3 protein（ P〈0. 05）. The percentage of atretic follicle of intervention groups was lower than that of the placebo group（ P〈0. 01）. Conclusion Shen-tonifying traditional Chinese herbs XZYZ capsules can improve the activity of PI3 K/AKT signaling pathway,increase ovarian reserve of primordial follicle,secondary follicle and antral follicle and reduce the reserve of atretic follicle,thus to promote ovarian function.