Survival role of superoxide dismutase 1 on human granulosa luteinized cells in vitro.

OBJECTIVE: Deleterious effects of free radicals do not only result from the amount of free radicals produced but also are related to the efficiency and to the activities of enzymatic antioxidant systems. We investigated the effect of exogenous superoxide dismutase (SOD1) or Cu-chelating agent diethyldithiocarbamate (DDC) on the apoptosis (caspase-3 activity) of human granulosa luteinized cells (hGLC) in vitro. METHODS: The effects of SOD1 and DDC were studied using in vitro culture system, caspase-3 and the total SOD activity in hGLCs were measured using AcDEVD-PNA substrate and Beaushap and Fridovich methods, respectively, after 48 h of the culture period. RESULTS: The activity of SOD1 was the lowest in GLCs treated with 100 µM DDC as compared to control cells and to the cells supplemented with Cu, Zn-SOD or DDC (10 µM). The effect of DDC was associated with elevated caspase-3 activity as compared to control cells. CONCLUSIONS: It was demonstrated for the first time that the supplementation of cultured hGLCs with Cu2+, Zn2+-SOD (200 U/ml) maintains the viability of hGLCs via caspase-3 suppression.

Antibodies against granulosa luteinized cells and their targets in women attending IVF program.

PROBLEM: The study was conducted to investigate the presence and the possible role of anti-granulosa luteinized cells (GLCs) antibodies in infertile patients, attending in vitro fertilization (IVF) cycles, as well as to attempt the identification of putative target antigens. METHOD OF STUDY: A total of 101 sera from infertile women (patient group) and 33 sera from fertile women (control group) were investigated. The patients were divided in subgroups according to: ovarian response to controlled ovarian hyperstimulation (COHS)--poor responders and good responders; fertilization rate--low fertilization rate and high fertilization rate. The relation between embryo transfer (ET) outcome and prevalence of anti-GLC antibodies was investigated too. NONIDET P-40 (NP-40) and whole-GLC lysates, as antigenic materials, were used in enzyme-linked immunosorbent assay (ELISA) and immunoblotting, respectively. RESULTS: Based on the cut-off value of 2 S.D. above the control mean value, the prevalence of patients positive for anti-GLC antibodies was found to be 28.7% (29/101) compared with 9.1% (3/33) in control fertile women (P < 0.05). Anti-GLC antibodies were detected in 54.8% (17/31) of women with low fertilization rate compared with 15.7% (11/70) of women with high fertilization rate (P = 0.003) and it was established that women positive for anti-GLC antibodies had lower fertilization rate, than those negative for anti-GLC antibodies. Immunoblotting has demonstrated a number of GLC proteins with molecular masses (MM) of 110, 70-80, 47 and 37 kDa, frequently reacting with anti-GLC antibodies in patient's sera, as possible targets. CONCLUSION: The presented results on the prevalence and role of anti-GLC antibodies, as well as the data on the target antigens confirmed our hypothesis about the potential involvement of GLC antigens in some cases of immunological infertility in IVF patients.