[Extracellular metabolome remodeling mediates CART-suppressed bovine follicular development]

This study aimed to investigate the effects of cocaine- and amphetamine-regulated transcript peptide (CART) on the secretory function of bovine ovarian granulosa cells (GCs) and the metabolic micro-environment of follicular fluid. We employed untargeted metabolomics to analyze the metabolite profile of GC culture medium after treatment with CART_55-102, and integrated these data with transcriptomic and intracellular metabolomic data from the same experimental model. In the positive and negative ion modes, we identified 77 and 193 differential metabolites, respectively. Among these, we observed a significant decrease in glucose concentration and a significant increase in cholesterol concentration in the culture medium. Joint pathway enrichment analysis revealed that 6 and 17 Kyoto encyclopedia of genes and genomes (KEGG) pathways were commonly enriched by both differential genes and differential metabolites in the positive and negative ion modes, respectively. KEGG markup language (KGML) network analysis showed direct associations of LRRC51 with cholesterol and 16α-hydroxyestrone in the steroid biosynthesis pathway, ATP2B2 and CALML4 with cholesterol in the aldosterone synthesis and secretion pathway, and UROC1 with thiourocanic acid in the histidine metabolism pathway. On the basis of our previous intracellular metabolomic data (increased intracellular glucose and decreased intracellular cholesterol), the results of this study indicate that CART promotes glucose uptake by GCs but inhibits its intracellular oxidative utilization, leading to an increase in intracellular glucose content. At the same time, CART suppresses de novo cholesterol synthesis and promotes cholesterol release into the extracellular space, resulting in elevated extracellular cholesterol levels. In conclusion, this study proposes a potential novel mechanism hypothesis for CART inhibition of follicular development from a metabolic secretion perspective. That is, CART may remodel GC energy metabolism and cholesterol homeostasis to reshape the follicular fluid microenvironment. Our findings provide a new theoretical basis for understanding the regulatory mechanisms of bovine follicular development and have important reference value for improving the reproductive efficiency of cows and promoting the sustainable development of the cattle industry.