Jahan Asmeen, Chiang John Y L
Dept. of Biochemistry and Molecular Pathology, Northeastern Ohio Universities College of Medicine, 4209 State Route 44, P. O Box 95, Rootstown, OH 44272, USA.
Am J Physiol Gastrointest Liver Physiol. 2005 Apr;288(4):G685-95. doi: 10.1152/ajpgi.00207.2004. Epub 2004 Nov 18.
Sterol 12alpha-hydroxylase (CYP8B1) catalyzes cholic acid synthesis in the liver and is feedback inhibited by bile acids. In addition to activating farnesoid X receptor (nuclear receptor subfamily 1H4), bile acids also induce inflammatory cytokines in hepatocytes. The objective of this study was to investigate the mechanism by which inflammatory cytokines inhibit human CYP8B1 gene transcription. Real-time PCR assays revealed that both chenodeoxycholic acid (CDCA) and interleukin-1beta (IL-1beta) markedly reduced CYP8B1, cholesterol 7alpha-hydroxylase CYP7A1 and hepatic nuclear factor 4alpha (HNF4alpha) mRNA expression levels in human primary hepatocytes. However, CDCA induced, but IL-1beta reduced, small heterodimer partner (SHP) mRNA expression. IL-1beta inhibited human CYP8B1 reporter activity only in liver cells, and a c-Jun NH(2)-terminal kinase (JNK)-specific inhibitor-blocked IL-1beta inhibition. Activated JNK1 or c-Jun inhibited, whereas their dominant negative forms blocked, IL-1beta inhibition of CYP8B1 transcription. Mutagenesis analyses mapped an IL-1beta response element to a previously identified bile acid response element, which contains an HNF4alpha binding site. A dominant negative HNF4alpha inhibited CYP8B1 gene transcription and ectopically expressed HNF4alpha blocked IL-1beta inhibition. Furthermore, IL-1beta inhibited HNF4alpha gene transcription, protein expression, and binding to the CYP8B1 gene. JNK1 phosphorylated HNF4alpha and a JNK-specific inhibitor blocked the IL-1beta inhibition of HNF4alpha expression. These results suggest that IL-1beta inhibits CYP8B1 gene transcription via a mitogen-activated protein kinase/JNK pathway that inhibits HNF4alpha gene expression and its DNA-binding ability. This mechanism may play an important role in the adaptive response to inflammatory cytokines and in the protection of the liver during cholestasis.
固醇12α-羟化酶（CYP8B1）催化肝脏中胆酸的合成，并受到胆汁酸的反馈抑制。除了激活法尼醇X受体（核受体亚家族1H4）外，胆汁酸还能诱导肝细胞中的炎性细胞因子。本研究的目的是探讨炎性细胞因子抑制人CYP8B1基因转录的机制。实时PCR分析显示，鹅去氧胆酸（CDCA）和白细胞介素-1β（IL-1β）均显著降低人原代肝细胞中CYP8B1、胆固醇7α-羟化酶CYP7A1和肝细胞核因子4α（HNF4α）的mRNA表达水平。然而，CDCA诱导小异源二聚体伴侣（SHP）的mRNA表达，而IL-1β则降低其表达。IL-1β仅在肝细胞中抑制人CYP8B1报告基因活性，且c-Jun氨基末端激酶（JNK）特异性抑制剂可阻断IL-1β的抑制作用。激活的JNK1或c-Jun抑制CYP8B1转录的IL-1β抑制作用，而它们的显性负性形式则阻断该抑制作用。诱变分析将IL-1β反应元件定位到先前鉴定的胆汁酸反应元件，该元件包含一个HNF4α结合位点。显性负性HNF4α抑制CYP8B1基因转录，异位表达的HNF4α阻断IL-1β的抑制作用。此外，IL-1β抑制HNF4α基因转录、蛋白质表达及其与CYP8B1基因的结合。JNK1使HNF4α磷酸化，JNK特异性抑制剂阻断IL-1β对HNF4α表达的抑制作用。这些结果表明，IL-1β通过丝裂原活化蛋白激酶/JNK途径抑制CYP8B1基因转录，该途径抑制HNF4α基因表达及其DNA结合能力。这一机制可能在对炎性细胞因子的适应性反应以及胆汁淤积期间肝脏的保护中发挥重要作用。
