Genetic Regulation of Nonalcoholic Fatty Liver Disease

非酒精性脂肪肝的基因调控

基本信息

批准号：
10424579
负责人：
Aras Nikodemas Mattis
金额：
$ 56.41万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-08 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10424579
关键词：
Adult Age Alternative Splicing American Animal Model Biology Cell Line Cell model Cells Development Diagnostic tests Disease model Electronic Health Record Fatty Liver Fatty acid glycerol esters Genes Genetic Genetic Diseases Genetic Models Genetic Predisposition to Disease Genetic Risk Genetic Variation Haplotypes Hepatic Hepatocyte Human Hypertriglyceridemia Impairment Individual Lead Linkage Disequilibrium Lipids Liver Liver Failure Liver diseases Measures Mediating Modeling Mus Non obese Oleates Patients Phenotype Pilot Projects Plasma Proteins Race Regulation Research Testing Therapeutic Time Triglycerides Undifferentiated Variant base case control causal variant clinical diagnosis cost cytokine disease diagnosis disease phenotype endoplasmic reticulum stress genetic variant hepatoma cell in vivo induced pluripotent stem cell knock-down lipid biosynthesis lipid metabolism mouse model non-alcoholic fatty liver disease nonalcoholic steatohepatitis novel novel strategies overexpression precision medicine risk variant screening sex tool transcriptome sequencing

项目摘要

SUMMARY Between 30-40% of American adults have nonalcoholic fatty liver disease (NAFLD), which currently has no therapeutic options and is predicted to become the leading cause of the liver failure in the US. NAFLD is initiated by excess lipid accumulation in the liver (i.e. hepatic steatosis). The well-established risk variants TM6SF2 rs58542926 (E167K) and PNPLA3 rs738409 (I148M) both lead to increased cellular lipid accumulation. While patient-derived cell-based models are a valuable tool for the discovery of novel genetic factors, most NAFLD models are based on liver-derived or hepatocyte-like cells, which are time-intensive and costly to generate. Since steatosis is a cell autonomous phenotype, we propose an entirely novel approach of using patient-derived induced pluripotent stem cells (iPSCs) in their undifferentiated state as a NAFLD genetic model. In preliminary studies we found that oleate-treated iPSC both before and after differentiation into hepatocyte-like cells (iHeps) with the TM6SF2 rs58542926-T risk allele have greater lipid accumulation compared to non-carriers, differences that were not completely rescued after gene editing to revert rs58542926 to the non-risk allele. This finding highlights the potential for iPSCs and iHeps to interrogate the genetic predisposition to NAFLD, as well as suggests that the association between rs58542926 and NAFLD is not driven by TM6SF2 alone. rs58542926 is in near perfect linkage disequilibrium with rs10401969, which regulates SUGP1 protein levels. In pilot studies, SUGP1 knock-down increased triglyceride levels in human hepatoma cell lines, while knock-down in vivo resulted in hepatic steatosis. While our findings implicate SUGP1 rs10401069 as an additional causal variant for NAFLD, to date, mechanistic studies of this haplotype have been focused on TM6SF2 alone. In Aim 1 we propose to test whether SUGP1 rs10401969 modulates NAFLD in cellular and animal models by: i) Identifying NAFLD relevant cellular phenotypes that differ between iHeps from SUGP1 and rs10401969 double-carriers vs. non-carriers; ii) Introduce the rs10401969 and rs58542926 risk-alleles in non-carriers, and test for recapitulation of cellular phenotypes; iii) Verify that SUGP1 knock-down and overexpression modulates NAFLD in vivo; and iv) Identify SUGP1 target genes that mediate NAFLD- phenotypes. In Aim 2, we will establish undifferentiated iPSCs as a cellular model for NAFLD genetic risk variants by: i) Comparing NAFLD-relevant cellular phenotypes in iPSCs compared to iHeps; ii) Confirming that iPSCs can model single SNP effects, and testing whether phenotypes differ in iPSCs selected from the extremes of the NAFLD genetic risk score distribution; and iii) testing for differences in cellular phenotypes in iPSCs from NAFLD cases vs. controls. Together these studies will significantly advance the field of NAFLD genetics by defining SUGP1 as a genetic contributor to NAFLD, and establishing iPSCs as a novel cellular model to identify and functionalize NAFLD genetic variants. The successful implementation of these aims will reveal new biology underlying genetic drivers of NAFLD that may ultimately be used to develop precision medicine standards for NAFLD diagnosis, screening and management.

概括 30-40% 的美国成年人患有非酒精性脂肪肝 (NAFLD)，目前尚无这种疾病治疗选择，预计将成为美国肝衰竭的主要原因。 NAFLD 启动由于肝脏中过多的脂质积累（即肝脂肪变性）。成熟的风险变体 TM6SF2 rs58542926 (E167K) 和 PNPLA3 rs738409 (I148M) 都会导致细胞脂质积累增加。尽管源自患者的细胞模型是发现新遗传因素的宝贵工具，大多数 NAFLD 模型基于肝源性细胞或肝细胞样细胞，这些细胞的生成既耗时又昂贵。自从脂肪变性是一种细胞自主表型，我们提出了一种全新的方法，利用患者来源的未分化状态的诱导多能干细胞（iPSC）作为 NAFLD 遗传模型。在初步研究我们发现油酸处理的 iPSC 在分化为肝细胞样细胞 (iHeps) 之前和之后与非携带者相比，TM6SF2 rs58542926-T 风险等位基因具有更大的脂质积累，差异在基因编辑将 rs58542926 恢复为非风险等位基因后，这些基因并没有被完全拯救。这一发现强调了 iPSC 和 iHeps 探究 NAFLD 遗传易感性的潜力，以及表明 rs58542926 与 NAFLD 之间的关联并非仅由 TM6SF2 驱动。 rs58542926 是与调节 SUGP1 蛋白水平的 rs10401969 存在近乎完美的连锁不平衡。在试点研究中， SUGP1 敲低会增加人肝癌细胞系中的甘油三酯水平，而在体内敲低导致肝脏脂肪变性。虽然我们的研究结果表明 SUGP1 rs10401069 是一个额外的因果变异 NAFLD，迄今为止，对该单倍型的机制研究一直集中在 TM6SF2 上。在目标 1 中，我们建议通过以下方式测试 SUGP1 rs10401969 是否在细胞和动物模型中调节 NAFLD：i) NAFLD 相关细胞表型在 SUGP1 和 rs10401969 双携带者的 iHeps 与 rs10401969 双携带者之间存在差异。非承运人； ii) 在非携带者中引入 rs10401969 和 rs58542926 风险等位基因，并进行重演测试细胞表型； iii) 验证 SUGP1 敲低和过度表达在体内调节 NAFLD；和四）确定介导 NAFLD 表型的 SUGP1 靶基因。在目标2中，我们将建立未分化的iPSC 作为 NAFLD 遗传风险变异的细胞模型： i) 比较 NAFLD 相关细胞表型 iPSC 与 iHeps 的比较； ii) 确认 iPSC 可以模拟单一 SNP 效应，并测试是否从 NAFLD 遗传风险评分分布的极端情况中选择的 iPSC 的表型有所不同；和三）测试 NAFLD 病例与对照 iPSC 中细胞表型的差异。这些研究一起通过将 SUGP1 定义为 NAFLD 的遗传因素，将显着推进 NAFLD 遗传学领域的发展，并且建立 iPSC 作为一种新型细胞模型来识别和功能化 NAFLD 遗传变异。成功者这些目标的实现将揭示 NAFLD 遗传驱动因素的新生物学，最终可能用于制定 NAFLD 诊断、筛查和管理的精准医学标准。