Heart failure is a progressive disease characterized by cardiomyocyte (CM) loss, interstitial fibrosis, loss of ventricular compliance and chamber remodeling. CMs change in response to pathological stimuli, altering their cell morphology, increasing protein synthesis and upregulating fetal genes. Though initially compensatory, these changes eventually become maladaptive, inducing fibrosis and adverse cardiac responses. Cardiac fibroblasts (FBs) are implicated in regulating aspects of this deleterious profile, but specific molecular mechanisms regulating their function remain unclear. FBs are, however, known to be significant effectors of cardiac function, where they act as principal determinants of ventricular remodeling and fibrosis in response to stress and injury. In chronic disease states, however, this fibrotic response leads to reduced wall compliance, decreased diffusion efficiency, and arrhythmias. Thus, limiting fibrosis and the activity of myofibroblasts under conditions of chronic stress would be beneficial in preventing heart failure. As part of our original grant, we generated mice with CM- specific deletion of RhoA, a Ras-related small G protein, to determine the molecular mechanisms of its activity specifically in myocardium. In response to chronic stress, we found that hearts from these mice developed accelerated dilation, with significant loss of contractile function. However, and despite the heart failure pathology, they also had significantly reduced cardiac fibrosis, with a demonstrated decrease in transcriptional activation of genes involved in the fibrotic response, including the serum response factor (SRF) and myocardin related transcription factors (MRTF). Together, our data suggest that RhoA is a critical and nodal enzyme in cardiac injury, functioning to both preserve contractility and to mediate activation of profibrotic genes. Here, in this renewal application, we propose to specifically interrogate the functional and mechanistic role(s) for RhoA signaling in fibrosis. We hypothesize that RhoA modulates FB activity both directly, within the activated FB, as well as indirectly, through CM-specific paracrine signals, to mediate myofibroblast activation and/or response to cardiac stress and injury. We propose to 1) examine the CM-specific RhoA-mediated paracrine signals that drive myofibroblast transformation and activation; 2) determine if RhoA signaling is necessary and sufficient for myofibroblast propagation in activated FBs; 3) utilize novel nanoparticle targeted technology to deliver cell specific inhibitors of RhoA effectors to ameliorate fibrosis and to prevent cardiac disease progression. These data will verify, for the first time, the primacy of the RhoA pathway in the fibrotic response in vivo and will identify novel targets and therapeutic strategies for the treatment of cardiac fibrosis and heart failure.
Relevance: Using a combined, comprehensive set of biochemical, proteomic and genetic approaches, we will 1) examine RhoA-mediated paracrine signals that drive myofibroblast transformation and activation; 2) determine if RhoA signaling is necessary and sufficient for direct myofibroblast function both in vitro and in vivo using a novel inducible, fibroblast-specific transgenic approach; 3) utilize novel nanoparticle targeting technology to deliver cell specific inhibitors of RhoA effectors to ameliorate fibrosis and to prevent cardiac disease progression. These data will verify, for the first time, the primacy of the RhoA pathway in the fibrotic response in vivo and will identify novel targets and therapeutic strategies for treatment of cardiac fibrosis and heart failure.
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