Bile Acid and Sphingosine-1-phosphate Receptor-mediated Signaling in Cholestasis

胆汁酸和 1-磷酸鞘氨醇受体介导的胆汁淤积信号传导

• 批准号: 9024718
• 负责人: PHILLIP B HYLEMON
• 金额: $ 41.17万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-12-01 至 2020-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9024718
• 关键词: Apical; Bile Acids; Bile fluid; Biology; Cell Proliferation; Cell membrane; Cells; Chemicals; Cholestasis; Chronic; Data; Diffusion; Disease; Disease Progression; Down-Regulation; Etiology; Functional disorder; G-Protein-Coupled Receptors; Gene Expression; Genes; Growth; Hepatocyte; Histone Acetylation; Histones; Homeostasis; Human; Impairment; Inflammation; Inflammatory; Ligation; Link; Liver; Liver Fibrosis; Liver diseases; MAP3K1 gene; MAPK3 gene; Mediating; Mitogen-Activated Protein Kinases; Modeling; Molecular; Morbidity - disease rate; Mus; Nuclear; Pathway interactions; Patients; Phosphotransferases; Physiological; Play; Primary biliary cirrhosis; Protein Kinase C; Proteins; Proto-Oncogene Proteins c-akt; Reporting; Role; Serum; Signal Pathway; Signal Transduction; Sodium; Sphingosine; Sphingosine-1-Phosphate Receptor; Staging; System; Taurine Cholate; Testing; Therapeutic; Up-Regulation; apical membrane; base; basolateral membrane; bile acid transporter; bile duct; bile formation; bile salts; cell motility; cholangiocyte; clinical application; clinically significant; cytokine; effective therapy; genetic inhibitor; inhibitor/antagonist; liver injury; liver transplantation; migration; mortality; mouse model; novel; novel strategies; primary sclerosing cholangitis; public health relevance; receptor; receptor coupling; receptor-mediated signaling; small hairpin RNA; small molecule; sphingosine kinase; theories; tool

 DESCRIPTION (provided by applicant): Cholangiocytes, which form the bile duct system in the liver, are the major target cells in a number of human cholestatic liver diseases. Although there has been significant improvement in the understanding of the pathophysiology of disease progression over the past two decades, the underlying cellular/molecular mechanisms remain largely unknown. Cholangiocytes are continuously exposed to high concentrations of bile salts at their apical membranes. Bile acids taken up by an apical sodium bile acid transporter (ASBT) on cholangiocytes are unidirectionally transported across the cell and secreted via specific transporters (MDR3 and Ostα/Ostβ) on the basolateral membrane. ASBT has been reported to be regulated by changes in bile acid concentration and inflammatory cytokines. Bile acids taken up by cholangiocytes have been reported to activate a number of intracellular signaling pathways including: PKC, PI3K, MAP Kinase, and ERK 1/2 allowing for normal physiological homeostasis. Loss of ASBT allows only conjugated bile acids (CBAs) to activate plasma membrane receptors as hydrophilic bile acids cannot easily enter cells by simple diffusion. We have recently reported that CBAs activate the AKT and ERK1/2 signaling pathways via the G protein coupled receptor (GPCR) sphingosine-1-phosphate receptor 2 (S1PR2) in hepatocytes and cholangiocytes. The levels of CBAs in serum and liver are significantly elevated in chronic cholestasis, which is correlated with bile duct obstruction. Our preliminary data indicates that: 1 S1PR2 is the predominant S1P receptor expressed in cholangiocytes; 2) taurocholate (TCA)-induced cell proliferation and migration are inhibited by a specific shRNA and an antagonist of S1PR2 in cholangiocytes; 3) bile duct ligation (BDL) induces the up-regulation of S1PR2 gene expression and down-regulation of ASBT expression in mouse primary cholangiocytes; 4) BDL- induced cholangiocyte proliferation and liver fibrosis are significantly reduced in S1PR2-/- mice; 5) both S1PR2 and SphK2 are up-regulated in the liver of mdr2-/- mice (a PSC mouse model). In addition, it has been reported that TCA concentration was dramatically elevated in the liver and serum after BDL in mice. Based on these studies and our preliminary results, we HYPOTHESIZE that CBA-mediated activation of the S1PR2/SphK2 signaling cascades plays a critical role in promoting chronic cholangiopathy in cholestatic liver diseases. Three specific aims are proposed to test our central hypothesis. 1) To define the role of S1PR2 and SphK2 in CBA-mediated cholestatic liver injury using the BDL mouse model; 2) To identify the molecular/cellular mechanisms by which CBA-mediated S1PR2/SphK2 activation promotes cholestatic liver injury; 3) To test the therapeutic strategy for cholestatic liver injury by targeing S1PR2/SphK2 using chemical inhibitors and genetic tools in mdr2-/- mice, a cholestasis model of PSC. Completion of these specific aims will not only identify the potential cellular/molecular mechanisms involved in the initiation and progression of cholestatic liver diseases, but will also establish a novel theory in bile acid and S1P biology.

 描述（由申请人提供）：胆管细胞形成肝脏中的胆管系统，是许多人类胆汁淤积性肝病的主要靶细胞。虽然在过去的二十年中，对疾病进展的病理生理学的理解有了显着的改善，但基本的细胞/分子机制仍然很大程度上未知。胆管细胞在其顶膜处持续暴露于高浓度的胆汁盐。由胆管细胞上的顶端胆汁酸钠转运蛋白（ASBT）摄取的胆汁酸单向转运穿过细胞，并通过基底外侧膜上的特异性转运蛋白（MDR 3和Ostα/Ostβ）分泌。据报道，ASBT受胆汁酸浓度和炎性细胞因子变化的调节。据报道，胆管细胞摄取的胆汁酸激活许多细胞内信号传导途径，包括：PKC、PI 3 K、MAP激酶和ERK 1/2，从而实现正常的生理稳态。ASBT的丧失仅允许结合胆汁酸（CBAs）激活质膜受体，因为亲水性胆汁酸不能通过简单扩散轻易进入细胞。我们最近报道了CBAs通过G蛋白偶联受体（GPCR）鞘氨醇-1-磷酸受体2（S1 PR 2）激活肝细胞和胆管细胞中的AKT和ERK 1/2信号通路。慢性胆汁淤积症患者血清和肝脏中CBAs水平显著升高，与胆道梗阻有关。初步研究结果表明：1 S1 PR 2是胆管细胞表达的主要S1 P受体; 2）牛磺胆酸（taurocholate，TCA）诱导的胆管细胞增殖和迁移可被特异性shRNA和S1 PR 2拮抗剂所抑制; 3）胆管结扎（bile duct ligation，BDL）诱导小鼠原代胆管细胞S1 PR 2基因表达上调和ASBT表达下调; 4）BDL诱导的胆管细胞增殖和肝纤维化在S1 PR 2-/-小鼠中显著减少; 5）S1 PR 2和SphK 2在mdr 2-/-小鼠（PSC小鼠模型）的肝脏中均上调。此外，据报道，BDL后小鼠肝脏和血清中的TCA浓度显著升高。基于这些研究和我们的初步结果，我们假设CBA介导的S1 PR 2/SphK 2信号级联激活在胆汁淤积性肝病中促进慢性胆管病变中起着关键作用。提出了三个具体目标来检验我们的中心假设。1)目的：1）探讨S1 PR 2和SphK 2在CBA介导的胆汁淤积性肝损伤中的作用; 2）探讨CBA介导的S1 PR 2/SphK 2活化促进胆汁淤积性肝损伤的分子/细胞机制; 3）在mdr 2-/-小鼠中通过使用化学抑制剂和遗传工具靶向S1 PR 2/SphK 2来测试胆汁淤积性肝损伤的治疗策略，PSC的胆汁淤积模型。这些特定目标的完成不仅将确定参与胆汁淤积性肝病的启动和进展的潜在细胞/分子机制，而且还将建立胆汁酸和S1 P生物学的新理论。