补肾调泡周期法对多囊卵巢模型大鼠卵巢AMH及AMHR Ⅱ表达的调控

The Regulation of Bushen-Follicle Recuperation Therapy on Expression of AMH Protein and Receptor in PCO Model Rats

目的研究补肾调泡周期法对多囊卵巢(PCO)模型大鼠卵巢抗苗勒氏管激素(AMH)及Ⅱ型受体(AMHR Ⅱ)蛋白表达的调控,初步探讨补肾调泡周期法治疗多囊卵巢综合征(PCOS)的作用机制。方法110只SD雌性大鼠随机选取10只作为正常组,剩余100只采用来曲唑溶液灌胃法构建PCO大鼠模型成功后,设立模型组、阳性对照组、A-逍遥散(高、低剂量)组、B-三子汤(高、低剂量)组、A+B-逍遥散+三子汤周期用药4组,每组10只。统计分析各组大鼠治疗前后体质量增加量;卵巢脏器系数;HE染色法观察卵巢组织病理形态;免疫组织化学法检测PCO大鼠卵巢AMH、AMHRⅡ蛋白表达。结果体质量增加量:与正常组比较,PCO模型组大鼠体质量增加量明显增多(P<0.01);与模型组比较,A低B高组、B低组及阳性对照组大鼠体质量增加量减少(P<0.05)。卵巢系数:与正常组比较,PCO模型组大鼠卵巢系数明显升高(P<0.01);与模型组比较,A低组大鼠卵巢系数降低(P<0.05),A高组、B高组、A低B低组、A低B高组及阳性对照组大鼠卵巢系数明显降低(P<0.01)。AMH蛋白表达:与正常组比较,PCO模型组大鼠卵巢AMH蛋白表达明显升高(P<0.01);与模型组比较,所有治疗组及阳性对照组大鼠卵巢AMH蛋白表达明显降低(P<0.01);治疗组间两两比较:与A高B高组比较,A(高、低)组、B(高、低)组、A低B低组、A低B高组大鼠卵巢AMH蛋白表达明显升高(P<0.01),A高B低组大鼠卵巢AMH蛋白表达升高(P<0.05)。AMHRⅡ蛋白表达:与正常组比较,PCO模型组大鼠卵巢组织AMHRⅡ蛋白表达明显升高(P<0.01);与模型组比较,A低剂量组AMHRⅡ蛋白表达降低(P<0.05),余治疗组及阳性对照组AMHRⅡ蛋白表达明显降低(P<0.01);治疗组间两两比较:与B高组比较,A低组、B低组大鼠卵巢组织AMHRⅡ蛋白表达明显升高(P<0.01),A高组大鼠卵巢组织AMHRⅡ蛋白表达升高(P<0.05),A高B高组AMHRⅡ蛋白表达明显降低(P<0.01);与B低组比较,B高组、A+B周期用药4组及阳性对照组AMHRⅡ蛋白表达明显降低(P<0.01);与A低B低组比较,A(低、高)组、B低组、A高B低组AMHRⅡ蛋白表达明显升高(P<0.01);与A低B高组比较,A(低、高)组、B低组AMHRⅡ蛋白表达明显升高(P<0.01),A高B高组AMHRⅡ蛋白表达明显降低(P<0.01),A高B低组AMHRⅡ蛋白表达降低(P<0.05);与A高B高组比较,A(低、高)组、B(低、高)组、A低B高组、A高B低组AMHRⅡ蛋白表达明显升高(P<0.01);A+B周期用药4组间大鼠卵巢AMHRⅡ蛋白表达差异比较:A高B高组<A低B高组<A高B低组。结论补肾调泡周期法能明显降低PCO模型大鼠治疗前后体质量增加量;可明显降低PCO模型大鼠的卵巢系数;改善卵巢组织病理学形态;明显降低PCO模型大鼠卵巢AMH及AMHRⅡ蛋白表达,补肾调泡周期法可能通过抑制AMH信号通路,影响卵泡募集和选择,促进卵泡发育和成熟,进而恢复卵巢形态及功能。

Objective To study the regulation of Bushen-follicle recuperation therapy on the expression of AMH Protein and type Ⅱ receptor(AMHRⅡ)expression in PCO rat models,to investigate the mechanism of Bushenfollicle recuperation therapy for polycystic ovary syndrome(PCOS).Methods Ten of 110 female SD rats were randomly selected as normal group,the remained 100 rats were divided into model group,positive control group,A-Xiaoyao pulvis high-,and low-dose groups,B-Sanzi decoction high-,and low-dose groups,A+B-Xiaoyao pulvis+Sanzi decoction group,10 rats in each group.The increment of body mass was statistically analyzed.Ovarian organ coefficient were measured.The pathological morphology of ovarian tissue was observed by HE staining.AMH and AMHRⅡprotein expression in PCO rats’ovarian were detected by immunohistochemisty method.Results Body mass increment:compared with the normal group,the body mass increment of PCO model group was increased significantly(P<0.01).Compared with the model group,the body mass increment of A low+B high group,B low group and positive control group were reduced(P<0.05).Ovarian coefficient:compared with the normal group,the ovarian coefficient of PCO model group was increased significantly(P<0.01).Compared with the model group,the ovarian coefficient of A low group reduced(P<0.05),and those of A high group,B high group,A low+B low group,A low+B low group,and the positive control group reduced significantly(P<0.01).AMH protein expression:compared with the normal group,the expression of AMH protein in the ovaries of PCO model group was increased significantly(P<0.01).Compared with the model group,the expression of AMH protein in ovary of all the treatment groups and positive control groups were reduced significantly(P<0.01).Comparison between treatment groups:compared with A high+B high group,the expression of AMH protein in the ovary of rats in A high and low groups,B high and low groups,A low+B low group,and A low+B high group were increased significantly(P<0.01),and the expression of AMH protein in the ovaries of rats in A high+B low group were increased significantly(P<0.05).AMHRⅡprotein expression:compared with normal group,AMHRⅡprotein expression of the PCO model group rats’ovarian tissue were increased significantly(P<0.01).Compared with model group,AMHRⅡprotein expression of A low dose group was reduced(P<0.05),those of the other treatment groups and positive control group were reduced significantly(P<0.01).Comparison between treatment group:compared with group B high group,AMHRⅡprotein expression of A low group and B low group rats’ovarian tissues were increased significantly(P<0.01),that of A high group rats’ovarian tissue were increased(P<0.05);AMHRⅡprotein expression of A high+B high group was reduced significantly(P<0.01).Compared with B low group,AMHRⅡprotein expression of B high group,A+B cycle groups and positive control group significantly were reduced(P<0.01).Compared with A low+B low group,AMHRⅡprotein expression of A low and high groups,B low group,A high+B low group were increased significantly(P<0.01).Compared with A low+B high group,AMHRⅡprotein expression of A low and high groups,B low group were increased significantly(P<0.01),that of A high+B high group were reduced significantly(P<0.01),and that of A high+B low group were reduced(P<0.05).Compared with A high+B high group,AMHRⅡprotein expression of A low and high groups,and B low and high groups,A low+B high,A high+B low group were increased significantly(P<0.01).Comparison of AMHRⅡprotein expression between A+B cycle groups:A high+B high<A low+B high group<A high+B low group.Conclusion The regulation of Bushen-follicle recuperation therapy significantly reduce the increment of body mass.The ovarian coefficient of PCO model rats was significantly reduced.The histopathological morphology of ovary was improved,and the AMH and AMHRⅡprotein expression of PCO model rats were obviously reduced.The regulation of Bushen-follicle recuperation therapy may affect follicle recruitment and selection by inhibiting the AMH signaling pathway,promote follicle development and maturation,and thereby restore ovarian morphology and function.