A novel estrogen receptor, G protein‐coupled receptor 30 (GPR30) plays a critical role, through a non‐transcriptional regulatory mode, in promoting the formation of estrogen (E2)‐induced cholesterol (Ch) gallstones in female mice

Abstract

BackgroundCh gallstones are more common in women than in men in every population studied. We have found that the classical estrogen receptor α (ERα), but not ERβ, in the liver plays a key role in E2‐induced gallstones in female mice. We recently discovered that GPR30 is a new gallstone gene, Lith18, and is expressed in the endoplasmic reticulum, but not the nucleus, of hepatocytes. Thus, the molecular mechanisms of elucidating the lithogenic role of E2 in gallstone formation became more complicated. Our aim is to explore whether GPR30 produces an independent lithogenic action to promote hepatic Ch hypersecretion and gallstone formation in female mice. We also investigated whether the epidermal growth factor receptor (EGFR) is required for GPR30 to produce a non‐transcriptional regulatory role in inhibiting hepatic Ch 7α‐hydroxylase (CYP7A1) and the classical pathway of bile acid (BA) synthesis, leading to the availability of excess amounts of Ch for hepatic hypersecretion and gallstone formation.MethodsThe role of GPR30 in Ch crystallization and gallstone formation were investigated in ovariectomized (OVX) GPR30(−/−)/ERα(−/−) and GPR30(+/+)/ERα(−/−) mice implanted subcutaneously with pellets releasing E2 at 6 μg/day and fed a lithogenic diet for 8 wk. To study whether the metabolic abnormalities underlying the major source of the excess Ch molecules lead to Ch‐supersaturated bile as activated by GPR30, we examined the role of GPR30 in regulating CYP7A1 and the classical pathway of BA synthesis through the EGFR signaling.ResultsE2 induced rapid Ch crystallization through the liquid crystalline pathway and promoted gallstone formation in OVX GPR30(+/+)/ERα(−/−) mice fed the lithogenic diet. In contrast, the prevalence of gallstones was drastically reduced to 10% in OVX GPR30(−/−)/ERα(−/−) mice. Hepatic secretion of biliary BA was significantly reduced in mice expressing GPR30, leading to an increased ratio of Ch to BA. After E2 treatment, there was a significant increase in mRNA and protein levels of EGFR and a significant reduction in expression of liver receptor homolog‐1 (LRH‐1), coupled with reduced CYP7A1 mRNA and protein levels in OVX GPR30(+/+)/ERα(−/−), but not GPR30(−/−)/ERα(−/−), mice. In vitro studies of primary hepatocytes isolated from GPR30(+/+)/ERα(−/−) mice found that when EGFR was inhibited by AG1478, a highly potent EGFR kinase inhibitor, the mRNA and protein levels of CYP7A1 were unchanged even though GPR30 was activated by E2. However, this was not the case in GPR30(−/−)/ERα(−/−) hepatocytes.ConclusionsGPR30 regulates Cyp7a1 expression through the EGFR. In the lithogenic state, reduced hepatic synthesis of BA from the classical pathway inhibits the conversion of Ch to BA due to GPR30 repression of CYP7A1 in E2‐treated mice. As a result, a significant increase in the ratio of Ch to BA leads to the imbalance of biliary lipids for keeping Ch solubilized in bile, contributing to the formation of E2‐induced gallstones.Support or Funding InformationThis work was supported in part by research grants DK101793, DK106249 and AA03557, all from the National Institutes of Health (US Public Health Service).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.