Division of Gastroenterology, Department of Internal Medicine, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan, ROC.
Biochem J. 2013 Feb 1;449(3):761-70. doi: 10.1042/BJ20121055.
Jaundice or hyperbilirubinaemia is a common complication of sepsis. UGT1A1 (UDP-glucuronosyltransferase 1A1) is a critical gene for bilirubin metabolism and irinotecan detoxification. However, the molecular pathogenesis of hyperbilirubinaemia during inflammation needs to be further clarified. Human hepatic UGT1A1 expression was analysed by RT (reverse transcription)-PCR, qRT-PCR (quantitative real-time PCR) and Western blotting in response to LPS (lipopolysaccharide) stimulation. Transcription regulatory elements in the upstream promoter region of the human UGT1A1 gene were determined using EMSA (electrophoretic mobility-shift assay) and ChIP (chromatin immunoprecipitation). The important role of the transcription regulatory element was examined using a luciferase assay, and was determined by qRT-PCR using a transcription factor activation inhibitor. LPS down-regulated the UGT1A1 mRNA expression in human hepatoma cell lines. A newly identified NF-κB (nuclear factor κB)-binding site was located on the upstream promoter region (-725/-716) of the human UGT1A1 gene. LPS-induced NF-κB activation and specific binding to the NF-κB-binding site can suppress human UGT1A1 promoter activity and human UGT1A1 expression. We demonstrated that LPS mediates the suppression of human UGT1A1 expression through specific binding of NF-κB to this newly identified NF-κB-binding site in the upstream promoter of the human UGT1A1 gene. The present study may partly explain the molecular pathogenesis of inflammation-associated hyperbilirubinaemia.
黄疸或高胆红素血症是败血症的常见并发症。UGT1A1（UDP-葡萄糖醛酸基转移酶 1A1）是胆红素代谢和伊立替康解毒的关键基因。然而，炎症时高胆红素血症的分子发病机制仍需进一步阐明。采用 RT-PCR、qRT-PCR 和 Western blot 分析 LPS 刺激后人肝 UGT1A1 的表达。采用 EMSA 和 ChIP 分析人 UGT1A1 基因上游启动子区的转录调节元件。通过荧光素酶测定和使用转录因子激活抑制剂的 qRT-PCR 检测转录调节元件的重要作用。LPS 下调人肝癌细胞系中的 UGT1A1 mRNA 表达。在人 UGT1A1 基因的上游启动子区（-725/-716）发现了一个新的 NF-κB（核因子 κB）结合位点。LPS 诱导的 NF-κB 激活和特异性结合 NF-κB 结合位点可抑制人 UGT1A1 启动子活性和人 UGT1A1 表达。我们证明 LPS 通过 NF-κB 与 UGT1A1 基因上游启动子中这个新鉴定的 NF-κB 结合位点的特异性结合来介导人 UGT1A1 表达的抑制。本研究部分阐明了炎症相关高胆红素血症的分子发病机制。
