Cytoprotection of granulosa cells by glycine against lytic cell death following nigericin-mediated pyroptosis

Pyroptosis, a pro-inflammatory cell death mechanism, is characterized by gasdermin mediated cell membrane pore formation, cell swelling, and eventual lytic cell membrane rupture. Increased pyroptotic cell death of granulosa cells is observed in PCOS (polycystic ovary syndrome). Granulosa cells are critical for oocyte development and hormone production. Previous studies have shown that glycine, an amino acid, suppresses cell membrane rupture and is cytoprotective against lytic cell death by inhibiting the oligomerization of NINJ1, a cell membrane-rupturing protein, without inhibiting the preceding pyroptotic steps. Previous studies have also determined that glycine levels are low in the blood serum of women with PCOS. This low glycine level may increase the sensitivity of various cells to NINJ1-mediated cell membrane rupture. In this context, glycine deficiency may cause granulosa cells to become more susceptible to lytic cell death. Suppressing the lytic death of granulosa cells by increasing glycine levels (e.g., in follicular fluid) may constitute a novel therapeutic strategy in PCOS. By inhibiting NINJ1 oligomerization, glycine can protect granulosa cells from excessive lytic cell death, while allowing the persistence of pyroptosis signaling (e.g., by not blocking gasdermin pores), potentially positively impacting follicular integrity and ovarian function. In this study, the cytoprotective potential of glycine and its effects on the inflammatory response in cells where pyroptosis was induced by nigericin using the KGN human granulosa cell line were evaluated by analyzing the amounts of PGE₂ released from the cells. The findings showed that when glycine and nigericin applications were used together, PGE₂ release was significantly increased, whereas neither produced a significant change individually. In addition, total protein release from granulosa cells was analyzed comparatively across all experimental conditions using SDS-PAGE and Commassie staining analyses. In granulosa cells where glycine suppresses NINJ1-mediated cell membrane disruption, the increased PGE2 release following the initiation of pyroptosis is hypothetically linked to a possible increase in the number of gasdermin pores in cells where lytic cell death is prevented. This study provides preliminary data for a better understanding of lytic cell death in granulosa cells and the role of glycine in this process. Keywords: Glycine, Nigericin, NINJ1, PCOS, Granulosa Cell