Role of Gut Bacterial Side-Chain Cleavage of Cortisol in Host 11Beta-Hydroxyandrostenedione Formation

在内分泌学文献中，关于雄激素原11β-11的生物合成存在重大争议。 羟基雄烯二酮(11β-OHAD)。11-β-OHAD的合成被认为主要是在 肾上腺经11β-羟化雄烯二酮(A4)。另一种途径是通过侧链裂解 皮质醇，尽管它被认为是次要的。17-羟基酶/C17，20-裂解酶(CYP17A1)缺乏患者 无法合成皮质醇或A4。当给予外源性皮质醇时，尿液轮廓呈衍生性变化。 在11个β-Ohad中。这表明要么是一种新的宿主酶，要么是负责侧链的肠道微生物酶。 皮质醇的裂解。我们已经在一种高度活跃的肠道微生物途径中确定了可能与此有关的基因 用于皮质醇的侧链裂解。我们假设肠道微生物区系是一种重要的、未被充分研究的 宿主内分泌系统的组成部分，产生大量的11β-Ohad。这个项目将 通过比较无菌猪和定居猪的稳定同位素标记的皮质醇代谢来验证这一假设 有着不同复杂程度的肠道微生物。我们将通过单次检测确定对宿主结肠生理的影响。 免疫细胞的细胞RNA-Seq、免疫组织化学和流式细胞术。理解生理作用 肠道细菌对侧链的裂解也是决定未来调节亲 雄激素的形成。我们将在体外测定细菌对皮质醇和11β-Ohad的转录反应。 在活体内。此外，为了证明负责皮质醇侧链的基因之间的因果关系 裂解(DesAB)和宿主类固醇代谢组谱，我们将利用合成生物学方法来工程 将desAB途径(野生型和非活性突变体)导入大肠杆菌，并在生猪体内定植。这些 研究有望解开一个存在了几十年的谜团，如果它发生变化，可能会导致范式的转变。 可以证明，肠道微生物区系对一种数量上主要的宿主类固醇和亲 雄激素。

A major controversy exists in the endocrinology literature regarding the biosynthesis of the pro-androgen 11β- hydroxyandrostenedione (11β-OHAD). The synthesis of 11β-OHAD is thought mainly to be synthesized in the adrenal gland by 11β-hydroxylation of androstenedione (A4). Another route is through the side-chain cleavage of cortisol, although it is thought to be minor. Patients with 17-hydroxylase/C17,20-lyase (CYP17A1) deficiency fail to synthesize either cortisol or A4. When given cortisol exogenously, the urinary profile exhibits derivatives of 11β-OHAD. This suggests either a novel host enzyme, or a gut microbial enzyme responsible for side-chain cleavage of cortisol. We have identified genes in a highly active gut microbial pathway that may be responsible for the side-chain cleavage of cortisol. We hypothesize that gut microbiota are an important, and understudied component of the host endocrine system, which generate significant quantities of 11β-OHAD. This project will test this hypothesis by comparing stable isotope-labeled cortisol metabolism in germ-free pigs and pigs colonized with gut microbiomes of different complexity. We will determine effects on host colonic physiology through single- cell RNA-Seq, immunohistochemistry, and flow cytometry of immune cells. Understanding the physiological role of side-chain cleavage by gut bacteria is also important in determining future strategies to modulate pro- androgen formation. We will determine bacterial transcriptomic responses to cortisol and 11β-OHAD both in vitro and in vivo. Furthermore, to demonstrate causation between the genes responsible for cortisol side-chain cleavage (desAB) and host steroid metabolome profile, we will utilize a synthetic biology approach to engineer the desAB pathway (both wild type and inactive mutant) into E. coli and colonize the gnotobiotic pigs. These studies are expected to resolve an enigma that has existed for decades, and may result in a paradigm-shift if it can be shown that the gut microbiota contributes significantly to a quantitatively major host steroid and pro- androgen.