Integrating lipid genotyes and phenotypes in iPS derived hepatocytes/adipocytes

描述（由申请人提供）： 人类心血管疾病新的有效治疗方法的发现需要新的疾病机制的识别和验证。近年来，基因组变异研究进入了一个新的阶段。其中无偏全基因组关联研究（GWAS）可以识别与常见疾病相关的新的遗传位点。我们最近描述了95个与血脂水平相关的基因座-LDL胆固醇（LDL-C）、HDL胆固醇（HDL-C）或甘油三酯，这些基因座在NHLB 1心脏病研究（FHS）和其他人群队列中与心肌梗死（MI）的风险密切相关。将需要大量的工作来将新的关联转化为功能性见解，并最终转化为降低MI风险的疗法。关键的一步是确定这些基因位点如何影响与脂质代谢相关的人类组织类型（主要是肝脏和脂肪）的表型。我们已经在来自患者的手术肝脏和脂肪组织样品中进行了基因型与基因表达的表达数量性状基因座（eQTL）分析;从这项工作中，我们发现染色体1 p13-rs 12740374-上的LDL-C-和Ml-相关SNP与S 0 RT 1基因的肝脏表达之间存在强相关性。然而，这些研究受到组织稀缺和无法解决关键细胞表型如脂蛋白分泌的限制。因此，需要从特定基因型的患者中建立无限可再生的肝细胞和脂肪细胞来源。我们已经开发了通过外周血细胞（PBC）收集获得重编程诱导多能干细胞（IPS）的能力。我们还开发了一种新技术，使用编码重编程因子的体外转录RNA，在没有任何基因组改变的情况下加速和扩大IPS的产生。最后，我们已经建立了将iPS细胞分化为功能性肝细胞和脂肪细胞的方案。我们的联盟包括来自FHS，哈佛干细胞研究所（HSCI），马萨诸塞州总医院（MGH）和哈佛医学院（HMS）的研究人员。我们处于独特的地位，可以开发来自数百名FHS参与者的IPS细胞系和iPS衍生的肝细胞和脂肪细胞库，这些参与者具有全基因组基因型数据以及许多心血管表型。我们建议利用这些资源：（1）开发有效的方案以从血液样品中获得IPS细胞系，然后分化成功能性肝细胞和脂肪细胞;（2）扩大这些方案，以使得能够从Fragrance后代队列中的约400个个体高通量产生IPS细胞系、肝细胞和脂肪细胞;和（3）从这些肝细胞和脂肪细胞进行基因表达和代谢组学分析，使得能够对基因型与基因表达和代谢物水平以及现有表型数据（例如动脉粥样硬化的亚临床测量）进行综合统计分析。 (End摘要）

DESCRIPTION (provided by applicant): The discovery of new and effective treatments for human cardiovascular diseases requires the Identification and validation of novel disease mechanisms. Recently, studies of genomic variation entered a new phase. In which unbiased genome-wide association studies (GWAS) can identify novel genetic loci associated with common diseases. We have recently described 95 loci associate with blood lipid levels-LDL cholesterol (LDL-C), HDL cholesterol (HDL-C), or triglycerides, which are strongly associated with risk for myocardial infarction (Ml) In the NHLBl Framingham Heart Study (FHS) and other population cohorts. Much work will be needed to convert the novel associations into functional insights and, ultimately, therapies to reduce the risk of Ml. A key step is to determine how these genetic loci affect phenotypes in human tissue types relevant to lipid metabolism, principally liver and adipose. We have performed expression quantitative trait locus (eQTL) analyses of genotype vs. gene expression in surgical liver and adipose tissue samples from patients; from this work, we found a strong association between an LDL-C- and Ml-associated SNP on chromosome 1p13-rs12740374-and hepatic expression of the S0RT1 gene. However, these studies were limited by scarcity of tissue and the inability to address key cellular phenotypes such as lipoprotein secretion. Thus, there is a need to establish infinitely renewable sources of hepatocytes and adipocytes from patients of defined genotypes. We have developed the ability to obtain reprogrammed induced pluripotent stem (IPS) cells via peripheral blood cell (PBC) collection. We have also developed a novel technology to accelerate and scale up IPS generation, without any genomic alteration, using in vitro transcribed RNAs encoding reprogramming factors. Finally, we have established protocols to differentiate iPS cells into functional hepatocytes and adipocytes. Our consortium comprises investigators from the FHS, the Harvard Stem Cell Institute (HSCI), Massachusetts General Hospital (MGH), and Harvard Medical School (HMS). We are uniquely positioned to develop a library of IPS cell lines and iPS- derived hepatocytes and adipocytes from several hundred FHS participants, who have genome-wide genotype data as well as many cardiovascular phenotypes. We propose to leverage these resources to: (1) develop efficient protocols to obtain IPS cell lines from blood samples, followed by differentiation into functional hepatocytes and adipocytes; (2) scale up these protocols to enable high-throughput generation of IPS cell lines, hepatocytes, and adipocytes from ~400 Individuals In the Framingham Offspring Cohort; and (3) perform gene expression and metaboiomic profiling from these hepatocytes and adipocytes, enabling Integrative statistical analyses of genotypes with gene expression and metabolite levels, as well as existing phenotype data such as subclinical measures of atherosclerosis. (End of Abstract)