Novel transcriptional regulators of bile acid metabolism and hepatotoxicity

描述（由申请人提供）：法尼索- x核受体（FXR）的激活诱导许多影响多种代谢途径调节的基因的转录。FXR在正常生理中的重要性可以在动物中看到，FXR的失调已被证明与胆汁酸代谢异常、高脂血症、动脉粥样硬化、高血糖、对肝毒素的易感性增加、肝癌和肠屏障通透性增加有关。尽管有如此多的作用，但FXR在调节胆汁酸代谢和肠肝循环的几乎所有方面的中心作用尤为重要。鹅去氧胆酸（CDCA）和胆酸（CA）的肝脏合成分别受Cyp7a1和Cyp8b1酶的调控。大量研究支持肝脏和肠道FXR激活导致Cyp7a1和/或Cyp8b1抑制的模型。目前的理论表明，这两个基因的抑制依赖于肝脏中SHP水平的增加和/或肠源性Fgf15/19与其在肝细胞质膜上的同源受体的相互作用。然而，SHP和Fgf15/19的相对重要性，以及参与急性或长期抑制Cyp7a1和Cyp8b1的机制仍有待确定。事实上，目前的数据表明，其他的未知途径可能也参与了这两个基因的抑制。在此，我们确定了两种参与抑制Cyp7a1和Cyp8b1的新途径。首先，我们确定了两个转录抑制因子作为新的FXR靶基因。这两个基因都不参与胆汁酸或脂质代谢。然后，我们使用功能获得和功能丧失研究来证明这些阻遏物分别抑制Cyp8b1和Cyp7a1。重要的是，这种抑制改变了胆汁酸池中特定胆汁酸的比例。此外，我们证明了谷胱甘肽转移酶和相关基因的数量调节，其功能是保护肝脏免受毒性挑战。总之，这些初步研究i)挑战了当前的范式，即SHP和Fgf15/19是Cyp7a1和Cyp8b1的主要/唯一调节因子；ii)提供了一种机制，解释了FXR激活可能导致的肝保护。提出的实验将扩展这些新发现，并确定所涉及的分子机制。更好地了解FXR激活的影响显得尤为重要，因为FXR激动剂目前正在临床试验中进行测试。

DESCRIPTION (provided by applicant): Activation of the farnesoid-X-nuclear receptor (FXR) induces the transcription of numerous genes that affect the regulation of multiple metabolic pathways. The importance of FXR in normal physiology can be seen in animals where dysregulation of FXR has been shown to be associated with abnormal bile acid metabolism, hyperlipidemia, atherosclerosis, hyperglycemia, increased susceptibility to hepatotoxins, hepatocarcinoma and increased permeability of the intestinal barrier. Despite this plethora of effects, the central role of FXR in regulating virtually all aspects of bile acid metabolism and th enterohepatic circulation is particularly important. The hepatic synthesis of chenodeoxycholic acid (CDCA) and cholic acid (CA) is regulated by the enzymes Cyp7a1 and Cyp8b1, respectively. Numerous studies have supported a model in which activation of hepatic and intestinal FXR results in repression of Cyp7a1 and/or Cyp8b1. Current dogma suggests that repression of these two genes is dependent upon increased levels of SHP in the liver and/or the interaction of intestinally- derived Fgf15/19 with its cognate receptor on the hepatocyte plasma membrane. However, the relative importance of SHP and Fgf15/19, and the mechanisms involved in acute or long-term repression of Cyp7a1 and Cyp8b1 remains to be established. Indeed, the current data suggest that additional as yet unknown, pathways may also be involved in repression of these two genes. Herein, we identify two novel pathways that are involved in the repression of Cyp7a1 and Cyp8b1. First, we identified two transcriptional repressors as novel FXR target genes. Neither gene was known to be involved in bile acid or lipid metabolism. We then used gain-of-function and loss-of-function studies to demonstrate that these repressors function to repress Cyp8b1 and Cyp7a1, respectively. Importantly, such repression altered the ratios of specific bile acids in the bile acid pool. In addition, we demonstrate the regulation of number of glutathione transferases, and related genes, that function to protect the liver from toxic challenges. Together, these preliminary studies i) challenge the current paradigm that SHP and Fgf15/19 are the major/sole regulators of Cyp7a1 and Cyp8b1 and ii) provide a mechanism that explains the hepato-protection that can result from FXR activation. The proposed experiments will extend these novel findings and identify the molecular mechanisms involved. A better understanding of the effects of FXR activation appears particularly important, as FXR agonists are currently being tested in clinical trials.