补肾调经方对小鼠体外培养卵泡卵母细胞BMPRⅡ/ALK6-Smads表达的影响  

Bushen Tiaojing Recipe Regulated Expressions of BMPRⅡ/ALK6-Smads in Mouse Oocytes Culturein vitro

作　　者：孙晓换[1] 范丽洁[1] 李丽[1] 魏学聪 刘亚华[1] 耿丹丹[1] 谷向向 杜惠兰[1,2] 

机构地区：[1]河北医科大学研究生学院,石家庄050017 [2]河北中医学院中西医结合学院中西医结合生殖疾病协同创新中心,石家庄050091

出　　处：《中国中西医结合杂志》2016年第10期1241-1246,共6页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金资助项目(No.81173294)

摘　　要：目的观察补肾调经方对小鼠体外培养窦前卵泡成活数及卵母细胞骨形成蛋白Ⅱ型受体(bone morphogenetic protein receptorⅡ,BMPRⅡ)/激活素受体样激酶6-Smads(activin receptor-like kinase6-drosophila mothers against decapentaplegic proteins,ALK6-Smads)信号通路的影响,探讨其提高体外生长卵泡卵母细胞质量的作用机制。方法 65只12日龄健康雌性昆明小鼠,采用机械法分离其窦前卵泡,分别以正常大鼠血清、高、中、低剂量补肾调经方大鼠含药血清、高剂量补肾调经方大鼠含药血清+K02288孵育,分为对照组、补肾高、中、低剂量组(即补肾组)、抑制剂组,普通法体外培养。培养第6天比较补肾组与对照组窦前卵泡成活数,实时荧光定量PCR法检测卵母细胞中BMPRⅡ、ALK6、Smad1、Smad5、Smad8mRNA表达;细胞免疫荧光法检测上述指标和磷酸化的Smad1/5/8(phospho-Smad1/5/8,p-Smad1/5/8)蛋白表达水平。结果与对照组比较,补肾高剂量组窦前卵泡成活数增加,补肾各剂量组BMPRⅡ、ALK6、Smad5、Smad8 mRNA表达水平升高,上述指标及p-Smad1/5/8蛋白表达水平升高,补肾高、中剂量组Smad1 mRNA及其蛋白表达水平升高(P<0.05)。与补肾高剂量组比较,抑制剂组p-Smad1/5/8蛋白表达降低(P<0.05)。结论补肾调经方能增加体外培养窦前卵泡成活数,提高卵母细胞质量,其作用机制可能与调控卵母细胞中BMPRⅡ/ALK6-Smads信号通路有关。Objective To observe the effects of Bushen Tiaojing Recipe（ BTR） on the counts of survival preantral follicles and the bone morphogenetic protein receptor Ⅱ（ BMPR Ⅱ） / activin receptor-like kinase 6-drosophila mothers against decapentaplegic proteins（ ALK6-Smads） signal pathway in oocytes culturedin vitro,and to study its mechanism for improving the quality of oocytes. Methods Preantral follicles were mechanically isolated from 65 female 12-day old healthy Kunming mice,which were inoculated by normal rats＇ serum（ as the control group）,high,medium,low dose BTR containing serums（ as Shen-supplementing groups）,high dose BTR containing serum ＋ K02288（ as the inhibitor group）,respectively. All were cultured by common methodin vitro. On the6 th day the counts of survival preantral follicles were compared between each Shen-supplementing group and the control group respectively. mRNA expressions of BMPRⅡ,ALK6,Smad1,Smad5,and Smad8 were detected by Real-time fluorescence quantitative PCR. The protein expressions of indices mentioned above and phospho-Smad1 /5 / 8（ p-Smad1 /5 /8） were detected by cellular immunofluorescence test. Results Compared with the control group,the quantity of survival preantral follicles increased in the high dose BTR containing serum group; mRNA expres-sions of BMPRⅡ,ALK6,Smad5,and Smad8 were elevated,protein expressions of indices mentioned above and pSmad1 /5 /8 were increased in the 3 Shen-supplementing groups（ P〈0. 05）; mRNA and protein expressions of Smad1 were increased in high and medium dose BTR containing serum groups（P〈0. 05）. Compared with the high dose BTR containing serum group,protein expressions of Smad1 /5 /8 were reduced in the inhibitor group（P〈0. 05）. Conclusion BTR could elevate the quantity of survival preantral follicles culturedin vitro and improve the quality of oocytes,which might be possibly associated to regulating the BMPR Ⅱ / ALK6-Smads signal pathway in oocytes.

关 键 词：补肾调经方 窦前卵泡 卵母细胞 骨形成蛋白Ⅱ型受体 激活素受体样激酶6 信号通路 

分 类 号：R285.5[医药卫生—中药学]