Transforming growth factor beta1, microRNAs and diabetic nephropathy

描述（由申请人提供）：糖尿病肾病（DN）是糖尿病的主要并发症之一，可导致终末期肾脏疾病。DN的发病率正在上升，需要采取新的战略来对抗这种使人衰弱的疾病。DN的主要特征包括由于细胞外基质（ECM）蛋白的积累而增加的肾小球硬化和系膜细胞扩张。虽然几种生化途径和关键的促纤维化因子，如转化生长因子-b1 （TGFb1）和ECM蛋白胶原蛋白，都与DN的发病有关，但调控它们的微妙分子机制尚不清楚。在之前的资助期内，我们确定了肾脏microRNAs （miRNAs）在DN发病机制中的新作用。我们证明了miR-192可以介导TGF b1诱导的系膜细胞（MCs）中胶原蛋白的表达，并且在小鼠模型中miR-192缺乏可以保护DN的关键特征。从那时起，肾脏mirna在各种肾脏疾病中得到了越来越多的关注，也被认为是DN的有前途的临床生物标志物。然而，我们对改变DN进展的mirna谱及其治疗潜力的了解仍然有限。我们的目标是通过确定新发现的mirna及其宿主基因在DN发病机制中的作用，以及利用它们的潜力来满足更好的DN治疗的关键需求的转化方法来解决这一知识空白。我们将继续研究大量新的初步数据，这些数据表明，在体外MCs和糖尿病小鼠肾小球中，TGFb1和高糖共同上调了一种新的“Mega Cluster”mirna。此外，我们发现该簇嵌入在一个长转录本中，并且都受应激响应转录因子的调节，而多组分mirna的靶基因则调节蛋白质翻译、肥大和细胞应激。核心假设是，糖尿病条件下mirna大簇的上调抑制其关键功能基因靶点的表达，诱导肾小球MC肥大、蛋白质合成和纤维化，从而增加DN的进展。这一假设将通过三个具体目标进行测试，这将：i)检查大型集群中关键mirna的调节分子机制；ii)确定这些miRNA和关键共同靶基因在MCs中的功能作用，iii)最终评估新的基因靶向以及在小鼠DN模型中下调该miRNA簇基因组区域的翻译方法。这些具有概念创新和临床意义的研究结果可以确定糖尿病肾脏中目前未知的调节因子，从而可能导致识别急需的DN新治疗靶点，从而对该领域的发展产生积极影响。

DESCRIPTION (provided by applicant): Diabetic nephropathy (DN) is one of the major complications of diabetes that can lead to end stage renal disease. The rates of DN are escalating and new strategies are needed to combat this debilitating condition. Key features of DN include increased glomerulosclerosis and mesangial cell expansion due to the accumulation of extracellular matrix (ECM) proteins. Although several biochemical pathways and key profibrotic factors, such as transforming growth factor-b1 (TGFb1) and the ECM protein collagen, have been implicated in the pathogenesis of DN, the subtle molecular mechanisms regulating them are unclear. In the previous funding period we identified new roles for renal microRNAs (miRNAs) in the pathogenesis of DN. We demonstrated that miR-192 can mediate TGF b1 induced collagen expression in mesangial cells (MCs), and that miR-192 deficiency can protect against key features of DN in mouse models. Since then, renal miRNAs have gained increased attention in various renal diseases, and are also being recognized as promising clinical biomarkers for DN. However, we still have only limited knowledge about the spectrum of miRNAs that modify DN progression and their therapeutic potential. Our objective is to address this gap in knowledge by identifying the roles of newly identified miRNAs and their host genes in the pathogenesis of DN, and translational approaches to harness their potential to meet the critical need for better therapies for DN. We will follow up on extensive new preliminary data showing that a novel "Mega Cluster" of miRNAs is collectively up-regulated by TGFb1 and high glucose in vitro in MCs, and in diabetic mice glomeruli in vivo. Furthermore, we find that this cluster is embedded within a long transcript and both are regulated by stress responsive transcription factors, whereas target genes of multiple component miRNAs regulate protein translation, hypertrophy and cellular stress. The central hypothesis is that up-regulation of the mega cluster of miRNAs diabetic conditions suppresses the expression of their key functional gene targets, inducing glomerular MC hypertrophy, protein synthesis and fibrosis, and thereby augmenting DN progression. This hypothesis will be tested via three Specific Aims which will: i) examine the molecular mechanisms of regulation of key miRNAs within the mega cluster; ii) identify the functional roles of these miRNAs and key common target genes in MCs, and iii) finally evaluate novel gene targeting as well as translational approaches to down-regulate this miRNA cluster genomic region in mouse models of DN. The results of these conceptually innovative and clinically significant studies can define currently unknown regulatory factors in the diabetic kidney that could lead to the identification of critically needed new therapeutic targets for DN and thus have a positive impact to advance the field.