Identification of Genes Activated by Bile Acids

性状（由申请方提供）：鹅去氧胆酸（CDCA），一种主要 胆汁酸，最近已显示激活法尼醇X受体（FXR）， 核激素受体超家族的一员。最近的数据表明， 激活的FXR控制胆汁酸生物合成和血浆脂质水平。作为 因此，FXR可能影响胆结石和/或动脉粥样硬化的发展。 然而，目前对靶基因知之甚少， 受激活的FXR影响的代谢途径。在第一个具体的 目的：利用抑制性消减杂交技术和DNA 微阵列来鉴定由CDCA激活的FXR调控的基因。这些 研究将利用稳定过表达高水平的HepG 2和Caco 2细胞， 为了更容易地识别FXR靶基因。我们将使用正常的， 用FXR配体处理的FXR-/-或VP 16-FXR转基因小鼠，以证明 (i)这些相同基因在体内被诱导和（ii）FXR的激活 导致血浆脂质降低。在第二个具体目标中，我们将 确定FXRE和其他关键的顺式元件在启动子中的一些 选择目标1中已鉴定的基因，以确认这些基因 基因是FXR/CDCA的直接靶点。在第三个目标中，我们将产生小鼠， 其在肝脏中过表达大鼠VP 16-FXR（参见目的1）。最后，具体而言， 目的4，我们将分离来自HepG 2和Caco 2细胞的细胞系， 表达FXR 1、FXR 2或FXR 3。目的基因的诱导，鉴定 在目的1中，将确定每种FXR亚型，以检测 假设特定基因/代谢途径被每个FXR激活 同种型。

DESCRIPTION (provided by applicant): Chenodeoxycholic acid (CDCA), a primary bile acid, has recently been shown to activate the farnesoid X receptor (FXR), a member of the nuclear hormone receptor superfamily. Recent data suggest that activated FXR controls both bile acid biosynthesis and plasma lipid levels. As such, FXR may affect the development of gallstones and/or atherosclerosis. However at the current time, very little is known about the target genes and metabolic pathways that are affected by activated FXR. In the first specific aim, we propose to use Suppression Subtractive Hybridization and DNA microarrays to identify genes that are regulated by CDCA-activated FXR. These studies will utilize HepG2 and Caco2 cells that stably overexpress high levels of FXR in order to more easily identify FXR-target genes. We will use normal, FXR-/- or VP16-FXR transgenic mice, treated with FXR ligands, to demonstrate (i) that these same genes are induced in vivo and (ii) that activation of FXR results in a decrease in plasma lipids. In the second specific aim, we will identify FXREs and other critical cis elements in the promoters of a few selected genes that have been identified in aim 1, so as to confirm that these genes are direct targets of FXR/CDCA. In the third aim, we will generate mice that overexpress rat VP16-FXR in their livers (see aim 1). Finally, in specific aim 4, we will isolate cell lines derived from HepG2 and Caco2 cells that express either FXR1, FXR2, or FXR3. The induction of target genes, identified in aim 1, by each FXR isoform will be determined in order to test the hypothesis that specific genes/metabolic pathways are activated by each FXR isoforms.