The Bile Acid Synthesis Pathway Is Present and Functional in the Human Ovary Laura P. Smith1,3, Maik Nierstenhoefer2,4, Sang Wook Yoo1,2, Alan S. Penzias1,3, Edda Tobiasch4, Anny Usheva1,2* 1 Division of Reproductive Endocrinology & Infertility, Department of Obstetrics & Gynecology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, United States of America, 2 Division of Endocrinology, Department of Medicine, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, United States of America, 3 Boston IVF, Waltham, Massachusetts, United States of America, 4 University of Applied Sciences Bonn-Rhein-Sieg, Rheinbach, Germany Abstract Background: Bile acids, end products of the pathway for cholesterol elimination, are required for dietary lipid and fat- soluble vitamin absorption and maintain the balance between cholesterol synthesis in the liver and cholesterol excretion. They are composed of a steroid structure and are primarily made in the liver by the oxidation of cholesterol. Cholesterol is also highly abundant in the human ovarian follicle, where it is used in the formation of the sex steroids. Methodology/Principal Findings: Here we describe for the first time evidence that all aspects of the bile acid synthesis pathway are present in the human ovarian follicle, including the enzymes in both the classical and alternative pathways, the nuclear receptors known to regulate the pathway, and the end product bile acids. Furthermore, we provide functional evidence that bile acids are produced by the human follicular granulosa cells in response to cholesterol presence in the culture media. Conclusions/Significance: These findings establish a novel pathway present in the human ovarian follicle that has the capacity to compete directly with sex steroid synthesis. Citation: Smith LP, Nierstenhoefer M, Yoo SW, Penzias AS, Tobiasch E, et al. (2009) The Bile Acid Synthesis Pathway Is Present and Functional in the Human Ovary. PLoS ONE 4(10): e7333. doi:10.1371/journal.pone.0007333 Editor: Wen-Liang Zhou, Sun Yat-Sen University, China Received June 23, 2009; Accepted September 16, 2009; Published October 6, 2009 Copyright: 2009 Smith et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Fellowship funding to Laura Smith, NIH funding to Anny Usheva. The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Alan Penzias MD: Ferring pharmaceuticals: Speakers bureau; research grants, EMD Serono: research grants, Repromedix: research grants, Organon Inc: research grants * E-mail: ausheva@bidmc.harvard.edu Introduction Bile acids are powerful detergents with essential functions in lipid and cholesterol processing. They play key roles in the gastrointestinal and hepatobiliary systems, are required for dietary lipid and fat-soluble vitamin absorption, and maintain the balance between cholesterol synthesis in the liver and cholesterol excretion [1–3]. Bile acids are composed of a steroid structure and are made primarily in the liver by the oxidation of cholesterol. The synthesis of bile acids and their localization are very tightly regulated due to their membrane and epithelial toxicity [3,4]. Bile acid synthesis can occur through two pathways: the classic (neutral) pathway or the alternative (acidic) pathway [5]. The classic pathway occurs in the liver, and accounts for approximately 90% of bile acid synthesis. The alternative pathway produces the remaining 10% of bile acids and is primarily extrahepatic. The classical pathway begins with the rate-limiting 7-a hydroxylation of cholesterol by cholesterol 7-a hydroxylase (CYP7A1) and the 12-a hydroxylation of the intermediates by sterol 12-a hydroxylase (CYP8B1), followed by side chain oxidation by sterol 27 hydroxylase (CYP27A1) [2,6–8]. The alternative pathway begins with the hydroxylation of the cholesterol side chain by sterol 27 hydroxylase (CYP27A1) in extrahepatic sites including vascular endothelium and macrophages, followed by 7-a hydroxylation of the oxysterol intermediates by oxysterol 7-a hydroxylase (CYP7B1) [6,9]. There are well-known feedback loops controlling gene tran- scription of the major enzymes in bile acid synthesis, especially CYP7A1 and CYP8B1 [6,7,10–12]. The nuclear receptors that have been demonstrated to regulate the bile acid synthesis cascade include the Farnesoid X Receptor (FXR), Retinoid X Receptor a (RXRa), Liver X Receptor a (LXRa), and Liver Receptor Homologue 1 (LRH-1) [6,13–15]. Only LRH-1 has been previously studied in steroidogenic tissue, including ovaries, from mice and rats [16–18]. In addition to its presence in the gastrointestinal and hepatobiliary systems, cholesterol is known to be highly abundant in the human ovarian follicle, where it is used in the formation of the sex steroids [19–21]. The human ovarian follicle is the fluid- filled sac that contains the oocyte (egg), and is lined with granulosa cells which house the enzymatic machinery to participate in sex steroid synthesis. Given the high content of both cholesterol and the sex steroids in the human ovarian follicles, we searched for the presence of bile acids in the ovarian follicular fluid [22]. Here we describe for the first time evidence that all aspects of the bile acid synthesis pathway are present in the human ovarian follicle, including the enzymes in both the classical and alternative PLoS ONE | www.plosone.org 1 October 2009 | Volume 4 | Issue 10 | e7333