Endometriosis affects the mouse oocyte quality by altering the molecular machinery involved in cortical reaction

KLINSKY, OMAR G.; WETTEN, PAULA A.; CASAIS, MARILINA ; SAYEGH, FABIANA C. ; MICHAUT, MARCELA A.

Mendoza

Congreso; LVIII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research; 2022

Sociedad Argentina de Investigaciones en Bioquímica y Biología Molecular

Endometriosis is a chronic gynecological disease characterized by the presence of endometrial-like tissue outside the uterine cavity, for example, in the pelvic cavity. Among the symptoms generated by the disease, infertility is one of the most prevalent and oocyte quality has been identified as the most important factor. Nevertheless, the factors that define this poor quality are unknown. Previous studies carried out in our laboratory showed that oocytes obtained from female mice with experimentally induced endometriosis (endo oocytes) presented some parameters related to oocyte quality altered, indicating that endometriosis might affect the oocyte competence to be fertilized. It was observed that matured metaphase II (MII) endo oocytes were not able to secrete the cortical granules content during parthenogenetic activation when compared with control oocytes. This secretory event -also known as cortical reaction- involves the fusion of cortical granules with the oocyte plasma membrane, and it is physiologically activated by the sperm during fertilization avoiding the fusion of new sperm (polyspermy). Therefore, this work aimed to determine possible factors that may affect the cortical granules exocytosis (CGE) in endo oocytes. Considering that CGE is regulated by the actin cytoskeleton and specific exocytic protein machinery -that we have previously characterized-, we hypothesized that the expression of F-actin and two key proteins for CGE -alphaSNAP and NSF- were altered in endo oocytes. First, an autologous endometriosis model was induced in female CF-1 mice (8 weeks of age). After 4 weeks, endo, sham, and control females were hormonally stimulated to obtain matured MII oocytes. Cells from three groups were fixed and permeabilized to perform indirect immunofluorescence assays. Fluorescent phalloidin was used to identify F-actin and specific primary antibodies were used to detect alphaSNAP and NSF. The analysis of fluorescence intensity by confocal microscopy showed that cortical F-actin was thickened in endo oocytes when compared with sham and control oocytes, while there was no difference between these last groups. On the other hand, when the fluorescence intensity corresponding to alphaSNAP and NSF were studied, we observed that both protein expressions were reduced in endo oocytes compared to sham and control oocytes. These results confirm our hypothesis and suggest that the molecular machinery involved in cortical granule exocytosis is altered in endo oocytes affecting their quality and competence to be fertilized.