Activation of G Protein-coupled Receptor 30 Promotes Proliferation of Goat Mammary Epithelial Cells via MEK/Erk&PI3K/Akt Signaling Pathway

Abstract BackgroundGoat is an important dairy animal. During lactation, maintaining a high proliferative activity in goat mammary epithelial cells (GMECs) is significant to improve the yield and composition of goat milk. Estrogen is an essential hormone in epithelial cell proliferation and ductal morphogenesis of mammary gland. G protein-coupled receptor 30 (GPR30) is a novel membrane receptor of estrogen. However, the relationship between estrogen/GPR30 signaling and proliferation of goat mammary epithelial cells has not been reported. And the molecular mechanisms underlying the proliferative effect of estrogen via GPR30 on GMECs remain unclear.ResultsTo investigate the effect of estrogen/GPR30 signaling on GMECs proliferation, goat mammary epithelial cells, which expressed cytokeratin 18 and β-casein, were isolated and identified, defining their mammary alveolar epithelium origination. Estrogen and GPR30 agonist G1 obviously promoted the proliferation of GEMCs, in contrast, GPR30 antagonist G15 partly abolished estrogen-induced cell proliferation. Remarkably, the stimulatory effect of estrogen and G1 on GMECs growth was suppressed by GPR30 knockdown detected by cell counting assay, CCK-8 assay, and BrdU assay, suggesting that estrogen/GPR30 signaling was involved in GMECs proliferation. Additionally, G15 decreased cyclin D1, cyclin B1, CDK1, and p-CDK1 expression, resulting in cell cycle arrest in the G2/M phase via a down-regulated phosphorylation of Erk1/2 and Akt compared with estrogen alone. What’s more, knock-down GPR30 led to an accumulation in the G2/M phase and inhibition of cyclin D1, cyclin B1, CDK1, and p-CDK1 expression via a down-regulation of phosphorylated Erk1/2 and Akt despite the presence of estrogen and G1. Furthermore, MEK inhibitor and PI3K inhibitor decreased the expression of cyclin D1, cyclin B1, CDK1, and p-CDK1, and repressed estrogen-induced and G1-driven promotion of cell growth. It indicated that estrogen/GPR30 signaling played an important role in GMECs proliferation by affecting cell cycle progression via MEK/Erk&PI3K/Akt signaling pathway.ConclusionThis study may provide a new insight into the effect of estrogen/GPR30 signaling on the regulatory action of goat mammary gland development.