Experimental Research On Intervention Mechanism Of Yishen Tune-periods Method To POF Model Rats

Objective:Observation the effect of Yishen tune-periods method to oestrous cycle in POF model rats and ovarian steroid hormone generating capacity in POF model rats (E2,P,T); Observation the effect of Yishen tune-periods method to ovarian reserve capacity (FSH,LH, INHB,AMH); Observation the moderate effect of Yishen tune-periods method to humoral immunity (AOAB) and neuroimmunology (P-EP); Observation the effect of Yishen tune-periods method to the apoptosis related proteins of granule cells (Bcl-2,Fas,Fas-1); Observation the moderate effect of Yishen tune-periods method to cellular immune related factors (VEGF,TNF-α INF-γ); Observation the change of Yishen tune-periods method to serum lipid (TC,TG) and Observation the effect of Yishen tune-periods method to ovarian morphology,in order to discuss interaction mechanism between POF and Yishen tune-periods method considering integrity and coordination of nerve-endocrine-immune network,and dose-effect relationship,it can provide the scientific basis in prevention and treatment POF with Chinese medicine.Method:choose SPF female healthy8weeks SD rats, the vaginal exfoliated cytology of each rat was inspected to elect rats with normal estrous cycle for animal subjects, except the blank group (A), the rest of animal subjects are given by the injection of cyclophosphamide to establish POF model. POF model rats were randomly divided into namely model group (B),Climen group (C), Zishentianjing soup high-dose group (D1), Zishentianjing soup low-dose group (D2),Wenshentianjing soup high-dose group (E1),Wenshentianjing soup low-dose group(E2),Yishen tune-periods method high-dose group (F1),Yishen tune-periods method low-dose group (F2), Yishen tune-periods method high-low dose group (G1) and Yishen tune-periods method low-high dose group (G2). A, B group were poured distilled water into stomach twice daily, each Chinese traditional medicine group was poured into stomach with the cold water for required concentration, low dose, high doses were equivalent to the clinical equivalent quantity2,4times respectively, Climen group (C) is equivalent to the clinical equivalent4times of quantity, each group was poured once daily, Consecutive medication last20days. The vaginal exfoliated cytology of each rat was inspected and record.After the last medication, all rats were taken blood by decollation.after doing it,measure level of E2,P,T in serum using calculating instrument of radio-immunity method; detect level of FSH,LH,β-EP,AOAB and expression of VEGF,TNF-α IFN-yin serum using ELISA; detect level of AMH, INHB in serum using double antibody sandwich of ELISA; measure level of TC,TG in serum using CHO. Decollation to separate hypothalamus and measure content of β-EP with hypothalamus using ELISA;spaying and fixation with paraffins,slicing,red dying to observe quantity of follicle at all levels and pathological change of follicle;measure protein expression of Bcl-2,Fas,Fas-L in serum using immunohistochemical methodResult:1. The effect of Yishen tune-periods method to oestrous cycle:Disorder oestrous cycle was improved in Yishen tune-periods method high-lowdose group (G1) dramatically.2. The effect of Yishen tunc-periods method to ovarian steroid hormone generating capacity:The level of E2, P in serum of Yishen tune-periods method high-lowdose group (G1) was elevated dramatically and the level of T in same group was reduced dramatically.3. The effect of Yishen tune-periods method to ovarian reserve capacity:The level of FSH,LH in serum of Yishen tune-periods method high-low dose group (G1) drops dramatically,the expression of INHB, AMH in serum was strengthened apparently.4. The moderate effect of Yishen tune-periods method to cellular immune related factors:The expression of TNF-αand INF-ydrops apparently in serum of Yishen tune-periods method high-low dose group (G1), and the expression of VEGF are strengthened in same group indistinctively.5.The effect of Yishen tune-periods method to the apoptosis related proteins of granule cells:Not only the expression of Bcl-2in serum of Yishen tune-periods method high-low dose group (G1) was strengthened distinctively, but also Fas did, in the contrary, the expression of Fas-1drops apparently.6.The moderate effect of Yishen tune-periods method to humoral immunityand neuroimmunology:The concentration of AOAB in serum of Yishen tune-periods method high-low dose group (G1) drop apparently,and the concentration of (3-EP in serum and hypothalamus are elevated apparently in same group.7. The change of Yishen tune-periods method to serum lipid:The level of both TC and TG drop apparently in serum of Yishen tune-periods method high-low dose group (G1).8.The effect of Yishen tune-periods method to ovarian morphology:The quantity of primary follicle,growing follicle and corpus luteum are incr-eased apparently in ovary of Yishen tune-periods method high-low dose group (G1),and inflammatory assault embellish are redused apparently in same group.Conclusion:1. It is succeeding by the injection of cyclophosphamide to establish POF model.2. Yishen tune-periods method with high-low dose can improve the disorder oestrous cycle.3. Yishen tune-periods method with high-low dose can adjust ovarian steroid hormone generating capacity.4. Yishen tune-periods method with high-low dose can improve ovarian reserve capacity.5. Yishen tune-periods method with high-low dose can participate in immune adjustment and follicle growth.6. Yishen tune-periods method with high-low dose can control the apoptosis of ovarian granulose cell.7. Yishen tune-periods method with high-low dose can weaken immune response and enhance neuroregulation act on ovary.8. Yishen tune-periods method with high-low dose can reduce the contect of serum lipid.9. Yishen tune-periods method with high-low dose can encourage follicular development.Therefore, Yishen tune-periods method can adjust the NEI network through multiple links in order to realize the comprehensive effect on the intervention of POF.