Activation of GPR30 Inhibits Cardiac Fibroblast Proliferation

The incidence of left ventricular diastolic dysfunction significantly increases in postmenopausal women suggesting the association between estrogen loss and diastolic dysfunction. The in vivo activation of G protein-coupled estrogen receptor (GPR30) attenuates the adverse effects of estrogen loss on cardiac fibrosis and diastolic dysfunction in mRen2.Lewis rats. This study was designed to address the effects of GPR30 on cardiac fibroblast proliferation in rats. The expression of GPR30 in cardiac fibroblasts isolated from adult Sprague-Dawley rats was confirmed by RT-PCR, Western blot analysis, and immunofluorescence staining. Results from BrdU incorporation assays, cell counting, carboxyfluorescein diacetate succinimidyl ester labeling in conjunction with flow cytometry, and Ki-67 staining showed that treatment with G1, a specific agonist of GPR30, inhibited cardiac fibroblast proliferation in a dose-dependent manner, which was associated with decreases in CDK1 and cyclin B1 protein expressions. In the GPR30-KO cells, BrdU incorporation, and CDK1 and cyclin B1 expressions significantly increased when compared to GPR30-intact cells. G1 had no effect on BrdU incorporation, CDK1 and cyclin B1 mRNA levels in GPR30-KO cells. In vivo studies showed increases in CDK1 and cyclin B1 mRNA levels, Ki-67-positive cells, and the immunohistochemistry staining of vimentin, a fibroblast marker, in the left ventricles from ovariectomized mRen2.Lewis rats versus hearts from ovary-intact littermates; 2 weeks of G1 treatment attenuated these adverse effects of estrogen loss. This study demonstrates that GPR30 is expressed in rat cardiac fibroblasts, and activation of GPR30 limits proliferation of these cells likely via suppression of the cell cycle proteins, cyclin B1, and CDK1.