| Bile acid (BA) recirculation and synthesis are tightly regulated via communication along the gut-liver axis and assist in the regulation of triglyceride (TG) and cholesterol homeostasis. Serum TGs and cholesterol are considered to be treatable risk factors for cardiovascular disease, which is the leading cause of death both globally and in the United States. While pharmaceuticals are common treatment strategies, nearly one-third of the population use complementary and alternative (CAM) therapy alone or in conjunction with medications, consequently it is important that we understand the mechanisms by which CAM treatments function at the molecular level. It was previously demonstrated that one such CAM therapy, namely a grape seed procyanidin extract (GSPE), reduces serum TGs via the farnesoid X receptor (Fxr). GSPE treatment also induces the expression of hepatic cholesterol 7alpha-hydroxylase (Cyp7a1), the rate limiting enzyme for de novo BA synthesis. Herein, we demonstrate that both gene and protein expression of Cyp7a1 is increased due to the fact that GSPE selectively regulates intestinal Fxr target genes involved in BA uptake and transport. Apical sodium dependent bile acid transporter (Asbt) expression is decreased with a concomitant reduction in fibroblast growth factor 15 (Fgf15), leading to a lack of repression on hepatic Cyp7a1. The subsequent 47% decrease in serum BAs and 69% increase in fecal BA excretion results in a significant reduction in serum TG and cholesterol. These Fxr dependent effects are lost in Fxr-/- mice, clearly demonstrating the critical role of this nuclear receptor. In a subsequent study we confirm that GSPE represses Asbt expression, while the BA sequestrant cholestyramine (CHY) induces expression. Treatment with either GSPE or CHY increases expression of Cyp7a1, with co-administration augmenting the increase. In the liver, GSPE and CHY independently induce expression of genes regulating cholesterol and lipid synthesis; however, when combined the expression of cholesterogenic and lipogenic genes induced by CHY is attenuated. Taken together these data indicate that GSPE has the potential for use either alone or as a complementary therapy in the treatment of hypertriglyceridemia and hypercholesterolemia. These findings, combined with the ability of low molecular weight procyanidins (LMW-PCNs) to modify intracellular proteins and signaling pathways led us to optimize a protocol for isolating LMW-PCNs from the seeds of grapes grown at the University of Nevada, Reno (UNR) vineyard. An ethyl acetate based extraction process utilizing whole seeds was found to be both time and cost effective, while preserving the anti-oxidant properties of the procyanidin-rich extract. This protocol will provide the basis for further extractions in order to conduct in vitro and in vivo testing, potentially allowing for the development of a value added product from the UNR vineyards. |
