Evaluating the role of Exogenous administration of micro RNA 29b in preventing Chronic Kidney Disease induced cardiac Fibrosis

描述（由申请方提供）：由于心血管组织的结构重塑增加，慢性肾病（CKD）患者和动物模型的心血管功能下降恶化。先前的研究评估了这些心血管变化的分子机制，揭示了Na/K-ATP酶（NKA）受体复合物激活在心脏纤维化中的核心作用。通过NKA受体复合物的信号传导由强心类固醇（CTS）、海蟾毒配基（MBG）的增加的血浆水平诱导。因此，Src或哺乳动物雷帕霉素靶蛋白（mTOR）通路抑制剂抑制NKA信号传导导致心脏纤维化减弱。然而，当与升高水平的CTS结合使用时，这些抑制剂也导致心脏细胞死亡增加。然后，预防纤维化的策略转向寻求更具体的靶点，直接调节心脏中的纤维化生长。使用来自5/6部分肾切除（PNx）或MBG输注动物的心脏组织，数据显示PNx和MBG输注显著降低了microRNA 29 b（miR 29 b）表达，其直接靶向胶原蛋白的mRNA，胶原蛋白是纤维化的必需组分。miR 29 b在体外胶原合成和纤维化中的作用已经得到很好的确立。 在动物模型中，miR-29 b的显著下调与心脏组织中胶原mRNA水平的显著增加相关。鉴于来自文献的体内数据和证据，假设miR 29 b是心脏纤维化的重要调节剂，对心脏细胞中的促纤维化信号传导具有特异性抑制作用，并且miR 29 b模拟物可用作潜在的体内治疗剂以预防由CKD诱导的心脏纤维化。 具体地，在体外，其旨在测试miR 29 b的过表达是否阻止心脏成纤维细胞和肌细胞中由MBG诱导的胶原合成。在体内，目的是测试慢性miR 29 b模拟物输注是否预防CKD的PNx模型中的心脏纤维化。研究表明，PNx和MBG输注会导致心脏纤维化和心肌病。两者均增加血浆MBG水平、NKA-受体复合物信号传导激活并降低心脏组织中的miR-29 b表达。为了确定外源性增加的miR 29 b水平是否防止CKD的PNx模型中的心脏纤维化，将通过渗透微型泵向经历PNx手术的大鼠输注miR 29 b模拟物或乱序miRNA。在PNx后四周，将处死大鼠并评价左心室的miR 29 b水平和心脏纤维化和左心室功能的生理测量。这项研究如果成功，将为预防CKD诱导的心血管衰退相关的心脏纤维化提供一种新的治疗策略，同时也为利用miRNA模拟物治疗疾病提供了概念验证方法。

DESCRIPTION (provided by applicant): Cardiovascular functional decline is worsened in patients and animals models with Chronic Kidney Disease (CKD) due to increased structural remodeling of cardiovascular tissues. Previous studies evaluating molecular mechanisms underlying these cardiovascular changes have revealed a central role for Na/K-ATPase (NKA) receptor complex activation in cardiac fibrosis. Signaling by the NKA receptor complex is induced by increased plasma levels of the cardiotonic steroid (CTS), Marinobufagenin (MBG). In accordance, inhibition of NKA signaling by Src or mammalian target of Rapamycin (mTOR) pathway inhibitors led to attenuation of cardiac fibrosis. However, these inhibitors also caused increased cardiac cell death when used in conjunction with elevated levels of CTS. Strategies to prevent fibrosis were then shifted to seek a more specific target that directly regulates fibrotic growth in the heart. Using heart tissue from 5/6th partial nephrectomized (PNx) or MBG-infused animals, data shows that PNx and MBG infusion significantly decreased microRNA 29b (miR29b) expression, which directly targets mRNA of collagen, an essential component for fibrosis. The role of miR29b in collagen synthesis and fibrosis in vitro has been well established. In animal models, significant down- regulation of miR-29b correlates with significantly increased collagen mRNA levels in cardiac tissues. Given the in vivo data and evidence from literature, it is hypothesized that miR29b is an important regulator of cardiac fibrosis with specific inhibitory effects on profibrotic signaling in cardiac cells and that miR29b mimics can be used as a potential in vivo therapeutic agents to prevent cardiac fibrosis induced by CKD. Specifically, in vitro it is aimed to test if over-expression of miR29b prevents collagen synthesis induced by MBG in cardiac fibroblasts and myocytes. In vivo, the aim is to test if chronic miR29b mimic infusion prevents cardiac fibrosis in the PNx model of CKD. Research has shown that PNx and MBG-infusion cause cardiac fibrosis and cardiomyopathy. Both increase plasma MBG-Levels, NKA-receptor complex signaling activation and decrease miR-29b expression in cardiac tissues. To determine whether exogenously increasing miR29b levels prevents cardiac fibrosis in the PNx model of CKD, rats subjected to PNx-surgery will be infused with either miR29b mimic or scramble miRNA through an osmotic minipump. Four weeks following PNx, rats will be sacrificed and the left ventricle evaluated for miR29b levels and physiological measures of cardiac fibrosis and left ventricular function. This study if successful will provide a novel therapeutic strategy for preventing CKD induced cardiovascular decline associated cardiac fibrosis, while also providing proof of concept approaches to utilizing miRNA mimics to treat diseases.