Granulosa cell dysfunction is associated with MCU-mediated mitochondrial calcium dyshomeostasis and reduced AKT phosphorylation in porcine cystic follicles.

Abstract

Follicular cysts represent a major cause of reproductive failure in sows, characterized by severe granulosa cell dysfunction; however, the underlying molecular mechanisms remain incompletely elucidated. This study therefore aimed to investigate the mechanisms responsible for granulosa cell dysfunction in porcine cystic follicles. Histological, ELISA, and RT-qPCR analyses revealed that cystic follicles exhibited a reduced granulosa cell layer, disorganized architecture, and significantly impaired steroidogenic capacity. Transcriptomic sequencing indicated that differentially expressed genes were primarily enriched in pathways related to calcium transport, mitochondrial function, PI3K/AKT signaling, and apoptosis. Notably, expression of the key mitochondrial calcium uniporter (MCU) was significantly downregulated in granulosa cells from cystic follicles. To further elucidate the role of MCU in granulosa cell function, MCU was knocked down in normal granulosa cells. This knockdown led to impaired mitochondrial calcium uptake, decreased membrane potential, and reduced ATP synthesis, along with suppressed expression of key steroidogenic genes and decreased estradiol secretion. Mechanistically, granulosa cells following MCU knockdown exhibited a significantly reduced level of AKT phosphorylation, which was associated with an increase in apoptosis. In summary, this study reveals that MCU-mediated disruption of mitochondrial calcium homeostasis may serve as a core mechanism underlying granulosa cell dysfunction. In normal granulosa cells, MCU knockdown exhibited an impairment of mitochondrial energy metabolism, defective steroidogenesis, and a marked reduction in AKT phosphorylation, accompanied by increased apoptosis. Furthermore, the consistency of this phenotype with that observed in granulosa cells from cystic follicles suggests that MCU may represent a potential target for intervention in cyst-associated reproductive disorders. These findings also offer novel insights into the pathological mechanisms underlying follicular cysts.