Transforming growth factor beta1, microRNAs, lncRNA and diabetic nephropathy

项目摘要 糖尿病肾病（DKD）是糖尿病的主要衰弱并发症，也是重要的医疗保健问题。 问题.目前DKD的治疗方法并不完全有效，许多患者仍进展为肾衰竭。 对DKD机制的认识不足限制了有效治疗的发展， 这强调了鉴定导致DKD的新介质和机制的迫切需要。上一 在资助期间，我们通过证明非编码RNA（ncRNA）的参与产生了重大影响， 包括microRNA（miRNAs）和长ncRNA（lncRNA）在DKD发病机制中的作用。我们描述了一个 肾小球系膜细胞中的一种新的lncRNA，lncMGC，它是一个巨大的miRNAs簇，miR-379的宿主转录本。 集群我们发现lncMGC调节与DKD相关的关键因子， 靶向lncMGC的寡核苷酸改善了早期DKD的病理特征。然而，分子和 lncMGC促进DKD特征的表观遗传机制，lncMGC或miR-379的作用 缺乏对糖尿病/肥胖诱导的小鼠肾功能不全的体内影响，以及性别特异性影响尚不清楚。 目前的更新将利用创新技术和微型计算机解决这些关键的知识差距。 该提案的目的是描述下游靶点和表观遗传调节因子的作用， IncMGC-miR-379轴在导致肾功能不全和DKD的过程中的作用。大量新的初步数据 支持我们的研究目标包括鉴定：a）lncMGC的关键蛋白结合伴侣， 染色质重塑; B）与DKD病理学相关的miR-379的新靶点;和c）高度发展 通过CRISPR-Cas9编辑的创新小鼠模型。我们假设糖尿病重新编程了肾小球系膜 细胞（MC）转录组和表观基因组失调lncMGC，宿主miR-379，及其关键靶基因， 影响线粒体功能、氧化和ER应激，导致纤维化、肾功能障碍和DKD。 Specific Aim 1将在MC中使用最先进的组学分析和技术来确定新的蛋白质靶点 以及lncMGC-miR-379轴促进DKD的表观遗传机制。《Specific Aim 2》将使用新的 miR-379和lncMGC缺陷的小鼠模型，以评估lncMGC、miR-379和相关基因的体内作用。 DKD的关键目标Specific Aim 3将利用新型人源化lncMGC小鼠和GapmeR靶向 使用人lncMGC来评估人DKD治疗的翻译潜力。 由于ncRNA在疾病状态中具有重要作用，因此这些持续的研究既科学又实用。 对DKD研究具有临床意义。该项目是创新的，因为它使用尖端技术，新颖， 小鼠模型和转化方法。它们一起可以改变现有的范式， 通过发现新的ncRNA介导的DKD进展调控， 和治疗意义，特别是对目前可用的治疗没有反应的患者。

PROJECT SUMMARY Diabetic kidney disease (DKD) is a major debilitating complication of diabetes and a significant healthcare problem. Current therapies for DKD are not fully efficacious, with many patients still progressing to renal failure. Inadequate understanding of mechanisms involved in DKD has restricted the development of effective therapies, which underscores the critical need to identify novel mediators and mechanisms leading to DKD. In the previous funding period, we made a significant impact by demonstrating the involvement of non-coding RNAs (ncRNAs), including microRNAs (miRNAs) and long ncRNAs (lncRNAs), in the pathogenesis of DKD. We characterized a novel lncRNA, lncMGC, in mesangial cells, which is a host transcript of a mega cluster of miRNAs, the miR-379 cluster. We found that lncMGC modulates critical factors associated with DKD and that a GapmeR antisense oligonucleotide targeting lncMGC ameliorates pathological features of early DKD. However, the molecular and epigenetic mechanisms by which lncMGC promotes features of DKD, the effects of lncMGC, or miR-379 deficiency in vivo on diabetes/obesity-induced renal dysfunction in mice, and sex-specific effects are unclear. The current renewal will address these crucial gaps in knowledge using innovative technologies and mice. The objective of this proposal is to characterize the role of downstream targets and epigenetic regulators of the IncMGC-miR-379 axis in processes that lead to renal dysfunction and DKD. Extensive new preliminary data supporting our research goals include identification of: a) key protein binding partners of lncMGC associated with chromatin remodeling; b) new targets of miR-379 related to DKD pathology; and c) development highly innovative mouse models by CRISPR-Cas9 editing. We hypothesize that diabetes re-programs the mesangial cell (MC) transcriptome and epigenome to dysregulate lncMGC, hosted miR-379, and their key target genes that affect mitochondrial function, oxidative and ER stress resulting in fibrosis, renal dysfunction, and DKD. Specific Aim 1 will use state-of-the-art Omics profiling and technologies in MCs to determine novel protein targets and epigenetic mechanisms by which the lncMGC-miR-379 axis promotes DKD. Specific Aim 2 will use novel mouse models of miR-379 and lncMGC deficiency to evaluate the in vivo roles of lncMGC, miR-379, and related critical targets in DKD. Specific Aim 3 will utilize a novel humanized lncMGC mouse and GapmeR targeting human lncMGC to evaluate the translational potential for human DKD treatment. Because ncRNAs have essential roles in disease states, these continuing studies are both scientifically and clinically significant for DKD research. The project is innovative because it uses cutting edge technologies, novel mouse models, and translational methods. Together, they can alter existing paradigms and have a positive impact by uncovering new ncRNA-mediated regulation of DKD progression, with potentially far-reaching clinical and therapeutic implications, particularly for patients not responding to currently available treatments.