养阴舒肝颗粒对VCD诱导卵巢功能下降大鼠Bcl-2,Bax及Caspase-3蛋白表达的影响

目的:探讨养阴舒肝颗粒对卵巢功能下降模型大鼠卵巢功能及;B淋巴细胞瘤-2(Bcl-2),Bcl-2相关X蛋白(Bax),半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)蛋白的影响。方法:选取动情周期规则的雌性SD大鼠,取10只作为正常组,其余大鼠均采用160 mg·kg~(-1)去氧乙烯基环己烯(VCD)腹腔注射诱导卵巢功能下降模型,取造模成功的56只大鼠按体质量随机分为模型组,养阴舒肝颗粒低(2.1 g·kg~(-1)),中(4.2 g·kg~(-1)),高剂量组(8.4 g·kg~(-1))和坤泰胶囊组,每组11只(模型组12只)。采用养阴舒肝颗粒和坤泰胶囊进行灌胃给药,干预治疗4周后进行卵巢称质量,计算各级卵泡的数目,酶联免疫吸附测定法(ELISA)检测血清性激素水平卵泡刺激素(FSH),雌二醇(E_2),抗缪勒氏管激素(AMH),蛋白免疫印迹法(Western blot)检测卵巢组织Bcl-2,Bax,Caspase-3的表达量。结果:与正常组比较,模型组大鼠的卵巢质量和卵巢指数下降(P0.01),闭锁卵泡数目增多(P0.01),FSH水平升高(P0.01),E_2和AMH水平下降(P0.05),卵巢组织Bcl-2蛋白的表达减少(P0.01),Bax,Caspase-3蛋白的表达(P0.01);与模型组比较,养阴舒肝颗粒可以增加模型大鼠的双侧卵巢质量(P0.05,P0.01);养阴舒肝颗粒和坤泰胶囊可以增加卵巢指数(P0.05,P0.01);养阴舒肝颗粒可以减少闭锁卵泡的数目(P0.05);养阴舒肝颗粒和坤泰胶囊均可以降低FSH水平(P0.05),升高E_2和AMH水平(P0.05),增加卵巢组织Bcl-2的表达(P0.01),同时抑制Bax,Caspase-3的表达(P0.01)。结论:养阴舒肝颗粒可以提高卵巢功能下降模型大鼠的卵巢功能,其机制可能与抗细胞凋亡有关。

Effect of Yangyin Shugan Granule on Expression of Bcl-2, Bax and Caspase-3 Protein in Rats with VCD-induced Diminished Ovarian Function

Objective: To investigate the effects of Yangyin Shugan granule(YYSG) on ovarian function in rats with diminished ovarian function, and explore its effect on B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), and cysteine-aspartic acid protease-3(Caspase-3) protein expression.Method: SD female rats with estrous cycle rules were selected, 10 used as blank group, and the others were used to establish diminished ovarian function model by injecting 160 mg·kg^-1 4-vinylcyclohexenediepoxide (VCD) into enterocoelia. 56 model rats with diminished ovarian function were randomly divided into 5 groups:model group, low-, middle-, high-dosage YYSG groups, and Kuntai capsule group, n=11 on each group (n=12 in model group). The YYSG groups (2.1, 4.2, 8.4 g·kg^-1) and Kuntai capsule group had intragastric administration for 4 weeks after the model establishment. Then the ovarian weight was measured and follicle count was calculated. The levels of serum sex hormonesfollicule-stimulating hormone (FSH), estradiol (E₂), and anti-mullerian hormone (AMH)] were detected by enzyme-linked immunosorbent assay (ELISA). The expression levels of Bcl-2, Bax, Caspase-3 in ovarian tissue were detected by Western blot.Result: As compared with the normal group, the ovary weight and ovary index were decreased (P<0.01), Atresia follicle count was decreased (P<0.01), FSH level was increased (P<0.01), E₂ and AMH levels were decreased (P<0.05), Bcl-2 protein expressions were decreased (P<0.01), and Bax and Caspase-3 protein expressions were increased(P<0.01) in model group. As compared with the model group, the ovary weights of YYSG groups were increased(P<0.05,P<0.01); YYSG granule and Kuntai capsule could increase the ovarian index(P<0.05,P<0.01); YYSG granule could reduce the numbers of atresia follicles(P<0.05); YYSG granule and Kuntai capsule could decrease FSH level but increase E₂ and AMH levels(P<0.05), increase the expression of Bcl-2 protein(P<0.01), and decrease the expression levels of Bax, Caspase-3 protein(P<0.01).Conclusion: YYSG could improve the ovarian function of rats with diminished ovarian function and the mechanisms may be associated with anti-apoptosis.