Abnormal Granulosa Cell Apoptosis In Polycystic Ovary Syndrome

Polycystic ovary syndrome (PCOS) is a heterogeneity of clinical syndrome, which is caused by a wide range of metabolic abnormalities, mainly endocrine disorders. Both ovaries of PCOS patients contain a large number of antral follicle, however they are not able to produce mature follicle periodically. The pathological mechanism of PCOS has not yet been made clear. Presently the primary, intrinsic abnormalities of follicle growth are considered to be the possible pathological mechanism basis which causes PCOS non-ovulation and clinical endocrine abnormality. Studies show that the intrinsic follicular dysplasia may be associated with the ovarian granulosa cell apoptosis regulation disorders. Recent studies point to the role of the endoplasmic reticulum (ER) in the sensing and transduction of apoptotic signals. Studies have established that expression of mutant, folding incompetent proteins causes ER stress and elicits an ER stress response, called the unfolded protein response (UPR). The initial intent of the UPR is adaptation and restoration of the normal ER function. The failure of such adaptative mechanisms leads to alarm signaling and finally to cell suicide, usually in the form of apoptosis, as a last resort to do away with dysfunctional cells.CHOP expressed in DNA-damaged ovarian granulosa cells. On the basic of these results, here we investigated test our hypothesis that differences in granulosa cell apoptosis may underlie abnormalities that affect follicular development, and to explore the pathogenesis of PCOS.OBJECTIVE:The purposes of this study were to score the composition of follicles in normal and PCOS ovaries, and to investigate apoptosis and the expressions of Bcl - 2 , Caspase3 , Bip and CHOP in granulosa cells of PCOS .Based on these experiments, we tried to make some exploration on the pathogenesis of PCOS.METHODS:We took 21 cases of ovarian wedge resection of ovarian pathology due to PCOS, and 15 cases of normal (non-PCOS) ovarian tissue from age-matched patients at early stage of the menstrual cycle as control group. Ovarian follicles of two groups were counted by morphometric analysis.Granulosa cells apoptosis in ovaries was observed by TdT-mediated dUTP-biotin nick end-labeling (TUNEL).The expression of apoptosis regulatory protein caspase3, Bcl-2 and endoplasmic reticulum stress protein Bip, CHOP protein was analysed with immunohistochemical staining.RESULTS:1.Stockpiling of growing follicles in ovaries of women with PCOSTwo pathology physicians observed the pathological structure in the optical microscope double-blindly. 10 visual fields (10*10) were randomly selected, and recorded the number of follicles at all levels. The total number of growing follicles was significantly greater in PCOS ovaries than normal, but the number of nongrowing primordial follicles did not differ. The transitional primary follicles, typical primary follicles, secondary follicles and antral follicle are more than control group, the differences are statistically significant. (P<0.05).Differential counts showed that the number of growing follicles at each stage of development was significantly greater: PCOS had 1.8-fold more primary, 1.6-fold more secondary, and 1.6-fold more Graafian follicles than normal. The greatest effect was on the classic primary follicles where the number was almost 3-fold greater in PCOS ovaries.2. Granulosa cells from PCOS patients have higher apoptotic ratesTen visual fields were randomly chosen for each slice, and observed the percentage of TUNEL-positive granulosa cells of the total granulosa cell number, that is, apoptotic index (AI). The comparison of ovarian tissue biopsy revealed that the apoptosis rate of primordial follicle, primary follicle and secondary follicle granulosa cells of the two groups are not significantly different. The antral follicular granulosa cell apoptosis rate of PCOS group is higher than control group. The difference is statistically significant. (P<0.05)3. Expression of apoptotic regulators in granulosa cellsCaspase3 and Bcl-2 protein was mainly expressed in the cytoplasm and membrane of granulosa cells, showing brown yellow. Significantly higher apoptotic rates were associated with stronger intensities immunostaining of the apoptotic effector caspase-3 (P<0.05) and weaker intensities immunostaining of the anti-apoptotic survival factor cellular Bcl -2 in the PCOS group (P<0.05).4. Expression of endoplasmic reticulum stress protein in granulosa cellsHallmark of endoplasmic reticulum stress protein CHOP protein was mainly expressed in the nuclei of granulosa cells.BIP protein was mainly expressed in the cytoplasm and membrane of granulosa cells, showing brown yellow. The intensities of CHOP immunostaining in PCOS ovaries were significantantly stronger compared to human normal ovaries.No significant difference of Bip expression was found between follicle of PCOS and the normal control group.CONCLUSIONS:PCOS patients have higher numbers of primary follicles, secondary follicles and antral follicle in ovarian compared with normal women. Significantly higher apoptotic rates were found in granulosa cells of patients with PCOS, compared with women with regular ovulatory cycles.The intrinsic follicular dysplasia may be associated with the ovarian granulosa cell apoptosis regulation disorders. Endoplasmic reticulum stress might be involved in the pathogenesis of PCOS.