Follicular atresia in buffalo: Cocaine- and Amphetamine-Regulated Transcript (CART) the underlying mechanisms

Abstract

Abstract Background: Atresia, a prevailing phenomenon within the majority of ovarian follicles, is a process regulated by hormone-induced apoptosis and influenced by changes in both endocrine and paracrine factors. Amongthese alterations, granulosa cell (GC) apoptosis is a key mechanism orchestrated through diverse signaling pathways. Cocaine- and amphetamine-regulated transcript (CART) signaling within ovarian GCs has been demonstrated to play a key role in the regulation of follicular atresia in cattle, pigs and sheep. In the present study, we aimed to investigate the potential local regulatory role of CART in the process of GCapoptosis-induced follicular atresia in buffalo, focusing on the modulation of the AKT/GSK3β/β-catenin pathways, the intracellular signaling pathways involved in cell viability. To address our objective, we firstinvestigated the association between CART gene (CARTPT) expression levels and follicular atresia by conducting gene expression analyses for CARTPT and related genes in both healthy and atretic follicles. Next, we investigated the influence of CART supplementation, with or without FSH, on GCestradiol production and apoptosis, exploring its potential regulatory role in the AKT/GSK3β/β-catenin pathways. Results: Our findings revealed increased expression of the CARTPT and BAX genes in atretic follicles, accompanied by decreased levels of AKT, β-catenin, and CYP19A1, compared to those in healthy follicles. Subsequently, CART treatment in the presence of FSH potently inhibited the FSH-induced rise in GC viability by reducing estradiol production and increasing apoptosis. This change was accompanied by an increase in the gene expression levels of both CARTPT and BAX, while the expression levels of AKT and CYP19A1 weresignificantly decreased. Moreover, at the protein level, treatment with CART in the presence of FSH negatively affects the activity of AKT, β-catenin and LEF1, while the activity of GSK3β was enhanced. Conclusion: In conclusion, our study illuminates how CART negatively influences buffalo GC viability by affecting estradiol production and promoting apoptosis—a key factor in follicular atresia. This regulatory mechanism involves modulation of the AKT/GSK3β/β-catenin pathway, providing valuable insights into the intricate mechanisms governing ovarian follicle development and granulosa cell function. These findings have implications for reproductive biology not only for buffalo but also for different species.