Molecular mechanism of Mfn2 alleviating endoplasmic reticulum stress and inhibiting apoptosis of sheep follicular granulosa cells.

Abstract

Follicle development is a crucial step in mammalian reproductive processes, the specific role of Mfn2 in regulating mitochondrial function and endoplasmic reticulum stress in this process is still unclear, this study aimed to investigate the role of Mfn2 in the follicular development of adult sheep. Large, medium, and small follicles were collected, and granulosa cells (GCs) were isolated from large follicles. The expression levels of Mfn2 in different follicles were detected using qRT-PCR and Western blot, and the localization of Mfn2 in follicles was determined through immunofluorescence. Additionally, the expression levels of the mitochondrial autophagy-related protein Pink1, endoplasmic reticulum stress proteins (Grp78, Perk, Chop), and apoptosis-related proteins (Bcl2 and BAX) were detected. Furthermore, siRNAs were transfected into GCs to knock down Mfn2 expression, and changes in intracellular Ca²⁺ accumulation and mitochondrial membrane potential were evaluated, along with the expression levels of the aforementioned proteins. The results showed that Mfn2 expression was significantly higher in large follicles compared to small follicles and was primarily localized in GCs. Compared to small follicles, the expression levels of Pink1, Grp78, Perk, Chop, and BAX were significantly lower in large follicles, while Bcl2 expression was significantly increased (P<0.01). After Mfn2 knockdown, intracellular Ca²⁺ levels and mitochondrial membrane potential were significantly reduced, while the expression levels of Pink1, Grp78, Perk, Chop, and BAX were significantly increased, and Bcl2 expression was significantly decreased (P<0.01). Mfn2 may influence cell apoptosis during sheep follicular development by regulating mitochondrial function and endoplasmic reticulum stress.