Effect of Cangfu Daotan Pill Containing Serum on Autophagy of Ovarian Granulosa Cells of Rats / 中国药房

ABSTRACT

OBJECTIVE：To study the effect of Cangfu daotan pill （CDP）containing serum on autophagy of ovarian granulosa cells（GCs）of rat. METHODS ：Three-month-old SD rat were divided into normal saline group （normal saline ，ig），FSH injection group（10.71 IU/kg，ih），CDP irrigation high-dose ，medium-dose and low-dse groups [ 0.5，1，2 mg/g（by crude drug ），ig]，with 6 rats in each group. They were given relevant medicine subcutaneously/intragastrically ，once a day ，for consecutive 3 days. After last medication ，blood sample was collected from the abdominal aorta to obtain drug-containing serum. GCs of rat were divided into blank control group ，model group ，FSH group （positive control ）and CDP high-dose ，medium-dose and low-dose groups. The autophagy model was induced by giving testosterone propionate ，except that the blank control group was directly added with 100 μL serum of normal saline group. Then model group was given 100 μL serum of normal saline group，and administration groups were given 100 μL drug-containing serum of corresponding drug group. The contents of estradiol（E2）and progesterone （P）in supernatant of cells were determined by ELISA. Western blot assay was used to detect protein expression of PI 3K，Akt and mTOR in cells. The mRNA expression of PI 3K，Akt，mTOR，Beclin 1，LC3Ⅰ，LC3Ⅱ and p 62 were detected by RT-PCR. RESULTS ： Compared with blank control group ，the content of E 2 in supernatant ，relative mRNA and protein expression of PI 3K，Akt and mTOR were decreased significantly in model group （P＜0.01），while relative mRNA expression of Beclin 1，LC3Ⅰ and LC 3Ⅱ were increased significantly （P＜0.01）. Compared with model group ，the content of E 2 in supernatant were significantly increased in FSH group ，CDP medium-dose and high-dose groups ，while relative mRNA expression of Beclin 1，LC3Ⅰ，LC3Ⅱ and p 62 were decreased significantly （P＜0.05 or P＜0.01）；relative mRNA and protein expression of PI 3K，Akt and mTOR were increased significantly in administration groups （P＜0.05 or P＜0.01）. CONCLUSIONS ：CDP can inhibit autophagy of GCs by activating related protein and mRNA expression of PI 3K/Akt/ mTOR signaling pathway.