Effects Of Repeated Super-ovulation On The Ovarian Reserve Function Of Rats

ObjectivesTo study the effects of repeated super-ovulation on the ovarian reserve function of SD rats and its possible mechanisms, offering the theory support for the clinical uses.Methods30 female SD rats were randomly divided into three groups, experimental groupâ… was treated with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (HCG) 6 times, experimental groupâ…¡was treated with normal saline 3 times, subsequently stimulated as the same as experimental groupâ… 3 times and the control group was treated with normal saline 6 times. The rats' body weights were weighed once every four days during the whole experiment, the change of estrous cycle was observed by vaginal smear each day. Half rats were killed on the fifteenth day after the experiment, the others were killed on the thirtieth day and the organ indices were calculate, the level of serum estrodiol-2 (E2), follicle stimulating hormone (FSH), luteinizing hormone (LH) and FSH/LH ratio in blood were detected by radioimmunoassay (RIA), the ovarian morphology and follicle number were observed through light microscope, and the expression of apoptosis inhibitory factor (Bcl-2) and apoptosis factor (Bax) in rats' ovarian tissue were detected by immunohistochemistry.Results1. The rats' body weight of experimental groupâ… andâ…¡were significantly lower than that of the control group on the thirtieth day after the experimental treatment (P<0.05).2. Vaginal smear showed that the rats' estrous cycle of experimental groupâ… andâ…¡became irregular after repeated super-ovulation. Majority of rats had no estrus and vaginal smear showed that these rats stopped at diestrus stage.3. The ovarian organ indices of experimental groupâ… rose on the fifteenth day after the experimental treatment (P<0.05), but there were no statistical significances of the ovarian organ indices among the different groups on the thirtieth day after the experimental treatment (P>0.05). There were no significantly difference of the uterine organ indices among the different groups and different time points (P>0.05).4. There were no significantly difference of the serum basal hormones among the different groups (P>0.05).5. The number of growing follicles in experimental groups was less than that of control group. The experimental groups could see much atresia follicles and could not see large diameter follicle. The control group could see lots of growth follicles which have different size. The control group had much more primordial follicles and primary follicles in the cortex than the experimental groups.6. The number of primordial follicles and primary follicles of adult rats was reduced in experimental groups (P<0.05), and there were significantly differences between the experimental groups on the fifteenth day after the experimental treatment (P<0.05), but there were no significantly difference between the experimental groups on the thirtieth day after the experimental treatment (P>0.05). There were significantly differences of the primordial follicles and primary follicles number in experimental group II between the two time points (P<0.05).7. There were significantly differences of the expression of Bcl-2 among the different groups (P<0.05), experimental groupâ… 0.05). There were no significantly difference of the expression of Bax among the different groups and different time points (P>0.05).Conclusions1. The animal experiment shows that repeated super-ovulation can accelerate the aging and lead to rats'estrous cycle disorders. The number of primordial follicles and primary follicles of adult rats are reduced in two experimental groups. However, the serum basal hormones in each group are not significantly different. The experimental results suggest that repeated super-ovulation possible role in primordial follicles and primary follicles, causing the primordial follicles and primary follicles atresia which are the manifestation of ovarian reserves function, and the more frequency used, the more seriously. 2. The animal experiment shows that repeated super-ovulation reduce the apoptosis suppressor genes Bcl-2 expression level of rat ovary, and the more frequency used, the more seriously. But it doesn't effect on apoptosis gene Bax expression level. So we don't think that the mechanism of repeated super-ovulation reduced ovarian reserve function is Bcl-2/Bax adjust follicular apoptosis which causing primordial follicles and primary follicles atresia.