Peripheral Artery Disease (PAD) is an occlusive disease of the lower extremity arteries leading to debilitating complications (e.g. claudication, amputation) due to defect in proper vascularization and efficient vascular remodeling. Recent attempts to promote therapeutic angiogenesis by VEGF therapies in patients with PAD have failed, perhaps because these strategies have only not targeted smooth muscle cells (SMC), cells that are essential for remodeling of capillaries and terminal arterioles (i.e. arteriogenesis) to increase blood distribution to ischemic regions. Arteriogenesis relies on the ability of SMC to be plastic and undergo a reversible phenotypic switching where they transiently downregulate their contractile apparatus, participate in capillary investment, and then re-differentiate back to the contractile state. However, the understanding of the mechanisms controlling SMC ability to re-differentiate and the retention of their lineage memory in vivo is limited. We previously identified an epigenetic signature specific to the SMC lineage consisting of the dimethylation of the lysine 4 of histone 3 (H3K4me2) on the promoters of the SMC marker genes (referred as SMC-H3K4me2) that is retained during SMC dedifferentiation. These observations suggest that SMC-H3K4me2 could play a pivotal role in maintenance of SMC identity and retention of lineage memory and could be a key mechanism controlling SMC participation in arteriogenesis. Studies in this proposal will test the central hypothesis that H3K4me2 controls the SMC differentiation state and enables SMC involvement in collateral capillary investment and muscularization during arteriogenesis by promoting the recruitment of TET2 on SMC marker genes. Consistent with this hypothesis, our preliminary studies, utilizing a novel locus-specific epigenome editing system to remove specifically H3K4me2 on the SMC marker genes, provide evidence that SMC-H3K4me2 tightly regulates SMC differentiation state and show that H3K4me2 interacts with Ten-eleven translocation (TET2), a key enzyme mediating DNA demethylation and a master regulator of SMC differentiation. Our central hypothesis will be further tested by addressing the following specific aims.
Aim 1 will determine the impact of loss of SMC-H3K4me2 on SMC participation to arteriogenesis. The functional consequences of SMC-H3K4me2 removal on SMC phenotype, their ability to invest capillaries, as well as tissue reperfusion will be evaluated.
Aim 2 will test the hypothesis that H3K4me2 plays a key role in promoting SMC differentiation by serving as a docking site for TET2 on the SMC marker genes. We expect the completion of these studies will lead to the identification of novel mechanisms controlling smooth muscle cell identity, plasticity, and lineage memory and their participation to beneficial microvascular arterialization and maturation. These results could serve from the development of new strategies for enhancing therapeutic vascularization in patients with PAD.

Public Health Relevance

Peripheral Artery Disease is an occlusive disease of non-cardiac and non-intracranial arteries affecting over 8.5 million patients in the US and leads to morbid and debilitating complications including lower extremity ischemic ulceration, gangrene, and eventually amputation due to a defect in tissue vascularization. Since recent attempts to promote vascularization by therapeutic endothelial cell-mediated angiogenesis have failed, there is a need to define mechanisms controlling and promoting the participation of smooth muscle cell in microvascular remodeling, since these cells play a key role in maturation and muscularization of immature capillaries, process leading to better tissue vascularization and perfusion. The studies of this proposal will identify the functional role of epigenetic programming, major regulatory process of cell gene expression, in controlling smooth muscle cells phenotype and participation in microvascular remodeling and may lead to novel therapeutic approaches to improve lower extremity vascularization in patients with Periphery Artery Disease.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Vascular Cell and Molecular Biology Study Section (VCMB)
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Reid, Diane M
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University of Pittsburgh
Internal Medicine/Medicine
Schools of Medicine
United States
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