Cellular and Molecular Endocrinology Laboratory, School of Life Science, Visva-Bharati University, Santiniketan 731 235, India.
Biochem Pharmacol. 2011 Nov 1;82(9):1186-97. doi: 10.1016/j.bcp.2011.07.069. Epub 2011 Jul 27.
Accumulation of bilirubin, primarily because of its insolubility, has been found to be associated with liver diseases including jaundice. Free bilirubin is insoluble; its glucuronidation by bilirubin-UGT enzyme (UGT1A1) makes it soluble and eliminates it through urine and faeces. Taking CCl(4) induced rat liver dysfunction model, we demonstrated that suppression of UGT1A1 activity in rat liver increased serum bilirubin level which could be reversed by carlinoside (Cln), a flavone glycoside. Although Cln is a flavone compound, it escaped self-glucuronidation in the intestine and readily absorbed. Kinetic study of microsomal UGT1A1 from HepG2 cells suggested that Cln enhanced enzyme activity by increasing V(max) without altering K(m). This altered V(max) was found to be due to UGT1A1 overexpression by Cln which was observed in both HepG2 and rat primary hepatocytes. Since Nrf2 is the transcription factor of UGT1A1, we examined whether Cln effect on UGT1A1 overexpression is mediated through Nrf2. In Nrf2 knock-out cells, Cln could not elevate UGT1A1 activity indicating Nrf2 to be its target. Cln significantly increased Nrf2 gene expression in HepG2 cells which was subsequently localized in nuclear region. Results from ChIP assay showed that Cln markedly augmented Nrf2 binding to UGT1A1 promoter that consequently enhanced reporter activity. Our findings therefore show that Cln upregulated Nrf2 gene expression, increased its nuclear translocation and stimulated UGT1A1 promoter activity. Total outcome of these events brought about a significant increase of bilirubin glucuronidation. Cln therefore could be a worthy choice to intervene hyperbilirubinemia due to liver dysfunction.
胆红素的积累，主要是由于其不溶性，已被发现与包括黄疸在内的肝脏疾病有关。游离胆红素不溶；其与胆红素-UGT 酶（UGT1A1）的葡萄糖醛酸化使其可溶，并通过尿液和粪便排出。采用 CCl(4)诱导的大鼠肝损伤模型，我们证明大鼠肝内 UGT1A1 活性的抑制会增加血清胆红素水平，而姜黄素（Cln），一种黄酮糖苷，可以逆转这种情况。尽管 Cln 是一种黄酮类化合物，但它在肠道中不会发生自身葡萄糖醛酸化，而是很容易被吸收。从 HepG2 细胞的微粒体 UGT1A1 的动力学研究表明，Cln 通过增加 V(max)而不改变 K(m)来增强酶活性。这种改变的 V(max)是由于 Cln 导致 UGT1A1 过表达所致，在 HepG2 和大鼠原代肝细胞中都观察到了这种情况。由于 Nrf2 是 UGT1A1 的转录因子，我们研究了 Cln 对 UGT1A1 过表达的影响是否通过 Nrf2 介导。在 Nrf2 敲除细胞中，Cln 不能提高 UGT1A1 的活性，表明 Nrf2 是其靶点。Cln 显著增加了 HepG2 细胞中 Nrf2 的基因表达，随后定位于核区。ChIP 检测结果表明，Cln 明显增加了 Nrf2 与 UGT1A1 启动子的结合，从而增强了报告基因的活性。我们的研究结果表明，Cln 上调了 Nrf2 基因的表达，增加了其核转位，并刺激了 UGT1A1 启动子的活性。这些事件的总结果导致胆红素葡萄糖醛酸化显著增加。因此，Cln 可以作为治疗因肝功能障碍引起的高胆红素血症的一种选择。
