Idiopathic and familial syndromes of pulmonary arterial hypertension (IPAH/FPAH) typically are associated with muscularization and obstruction of pulmonary arterial microperfusion circuits in the lung. We propose that the pathobiology of PAH represents a dysfunctional, peri-vascular wound healing response based on a functional deficit in the ability of the recently discovered Pur ? DNA-binding protein to repress TGF?1 signaling in the lung. Excessive transcriptional activation of wound-healing genes due to unchecked collaborative interaction between serum response factor (SRF) and TGF?1-regulated Smad proteins 2 and 3 results in accelerated peri-arteriolar myofibroblast (MFB) differentiation and adventitial fibrosis with loss of pulmonary arterial compliance and eventual right heart failure. Smads 2 and 3 normally dissociate gene-inhibitory SRF-Pur ? protein complexes to allow activation of the smooth muscle ?-actin (SM?A) and type I collagen ?2-subunit promoters as a first step in the MFB differentiation process. We will test the hypothesis that the SRF-Pur? inhibitory complex is unstable in PAH-derived MFBs due to over-active PI3K/Akt feed-forward signaling kinases and/or impaired feed-back inhibition mediated by sub-optimal MEK1/Erk1,2/Egr-1 signaling.
In Aim 1, we propose to characterize the sub-cellular compartmentalization of transcriptional activators and repressors implicated in peri-arteriolar myofibroblast differentiation and remodeling in IPAH/FPAH syndromes using an immunocytochemistry approach.
For Aim 2, we will define the biochemical dysfunction that causes excess peri-arteriolar myofibroblast differentiation in IPAH/FPAH syndromes using epigenetic/metabolic approaches that target SRF-Pur ? physical interplay in pulmonary artery adventitial fibroblasts isolated from normal or disease-affected donors. We have developed solid-phase ELISA tools to quantitatively evaluate protein:protein and protein:DNA interactions that uniquely regulate the process of adventitial MFB differentiation. The assembly of a specialized transcriptional regulatory complex capable of triggering prototypical gene responses in MFBs represents a convergence point for complex vascular-disease signaling consisting of multiple compensatory and patient-specific layers of control. We expect that knowledge gained could further basic understanding of rate-limiting interactions that foster loss of arterial compliance typically associated with the most devastating IPAH/FPAH disease syndromes. Future detailed analysis of the protein biochemistry of activator-repressor dynamic interplay could reveal novel targets for therapeutic management of pulmonary arterial disease and right heart failure that may ultimately improve patient long-term survival.

Public Health Relevance

Gene activation in the blood vessel wall is controlled by protein:protein and protein:DNA complexes that are necessary for rapid wound healing and tissue repair after infection or physical trauma. When these processes become inoperative or dysfunctional in the cardiopulmonary system, excess scar tissue can accumulate and stiffen very small blood vessels that eventually slows normal blood flow leading to clot formation, stroke, tissue infarction (degradation), and sudden death. Protein complexes that control wound healing in the lung may be faulty in patients with pulmonary hypertension which if untreated will eventually damage the right side of the heart that functions as the blood pump to the lung. The proposed research will examine the protein constituents of these regulatory complexes and determine if they are different in normal individuals compared to patients diagnosed with end-stage pulmonary hypertension. The proposed research may lead to the design of new treatment strategies for this devastating lung vascular disease that has no cure and only treatable by performing lung transplantation surgery.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Small Research Grants (R03)
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Special Emphasis Panel (ZHL1-CSR-Q (S1))
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Moore, Timothy M
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Ohio State University
Schools of Medicine
United States
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Hariharan, Seethalakshmi; Kelm Jr, Robert J; Strauch, Arthur Roger (2014) The Pur?/Pur? single-strand DNA-binding proteins attenuate smooth-muscle actin gene transactivation in myofibroblasts. J Cell Physiol 229:1256-71
Willis, William L; Hariharan, Seethalakshmi; David, Jason J et al. (2013) Transglutaminase-2 mediates calcium-regulated crosslinking of the Y-box 1 (YB-1) translation-regulatory protein in TGF?1-activated myofibroblasts. J Cell Biochem 114:2753-69