Angiogenesis, the sprouting of new capillary blood vessels from existing vasculature, is a complex biological process of critical importance to the treatment of numerous pathologies and the success of tissue engineering. Cell-based strategies to promote therapeutic vascularization have shown pre-clinical and clinical success, particularly for the treatment of critical limb ischemia, and typically involve the delivery of a single mesenchymal cell type to ischemic sites to stimulate angiogenesis. By contrast, strategies to vascularize engineered tissues typically involve two cell types, with endothelial cells (or their progenitors) combined with some sort of supporting stromal cell type and delivered via a hydrogel-based extracellular matrix (ECM). However, the choices of stromal cells and ECM have varied widely across studies. In the prior funding period, we used a combination of 3D cell culture models in vitro and subcutaneous implants in vivo to discover that the formation of nascent vasculature is regulated by both the biophysical properties of the ECM and the identity of the supporting stromal cells. Our data suggest that stromal cells of different origins differentially control ECM proteolysis during angiogenic sprouting, and that controlling the rate o ECM breakdown is critical to yield stable, functional vessels. In this competing renewal application, we propose to mechanistically investigate how these two critical instructive elements of the local microenvironment (i.e., the stromal cells and the ECM) influence the quantity, functional quality, and stability of new vasculature.
Aim 1 will quantify the impact of stromal cell identity on the rates of ECM proteolysis and capillary morphogenesis in a fibrin-based 3D co-culture model, using active microrheology to monitor local ECM mechanics as a function of time.
Aim 2 will use an engineered biomaterial platform with tunable degradative properties to determine the impact of ECM proteolytic susceptibility on the quantity and functional qualities of capillary networks.
Aim 3 will investigate how stromal cell identity and ECM proteolytic susceptibility affect the quantity and quality of neovasculature in a clinically relevant ischemic model. Successful completion of these studies will enhance current understanding of the role of the microenvironment in capillary morphogenesis, and ultimately guide the judicious selection of the ideal combination of cells and ECM for the treatment of ischemic conditions.

Public Health Relevance

Capillary morphogenesis is critical to many normal (e.g., wound healing) and pathologic (e.g., tumor progression) conditions, as well as emerging therapeutic approaches to treat ischemic conditions. In broad terms, this application will determine how two keys features of the microenvironment (e.g., the extracellular matrix and supporting stromal cells) regulate the quantity, functional quality, and stability of new capillary blood vessels. Results from the proposed studies will help bring cell-based strategies for revascularization therapies closer to a translational endpoint, and enhance ongoing efforts to vascularize engineered tissues.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Biomaterials and Biointerfaces Study Section (BMBI)
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Lundberg, Martha
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University of Michigan Ann Arbor
Biomedical Engineering
Schools of Engineering
Ann Arbor
United States
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Janson, Isaac A; Putnam, Andrew J (2015) Extracellular matrix elasticity and topography: material-based cues that affect cell function via conserved mechanisms. J Biomed Mater Res A 103:1246-58
Putnam, Andrew J (2014) The Instructive Role of the Vasculature in Stem Cell Niches. Biomater Sci 2:1562-1573
Ceccarelli, Jacob; Putnam, Andrew J (2014) Sculpting the blank slate: how fibrin's support of vascularization can inspire biomaterial design. Acta Biomater 10:1515-23
Vigen, Marina; Ceccarelli, Jacob; Putnam, Andrew J (2014) Protease-sensitive PEG hydrogels regulate vascularization in vitro and in vivo. Macromol Biosci 14:1368-79
Carrion, Bita; Janson, Isaac A; Kong, Yen P et al. (2014) A safe and efficient method to retrieve mesenchymal stem cells from three-dimensional fibrin gels. Tissue Eng Part C Methods 20:252-63
Rao, Rameshwar R; Ceccarelli, Jacob; Vigen, Marina L et al. (2014) Effects of hydroxyapatite on endothelial network formation in collagen/fibrin composite hydrogels in vitro and in vivo. Acta Biomater 10:3091-7
Grainger, Stephanie J; Carrion, Bita; Ceccarelli, Jacob et al. (2013) Stromal cell identity influences the in vivo functionality of engineered capillary networks formed by co-delivery of endothelial cells and stromal cells. Tissue Eng Part A 19:1209-22
Singh, Rahul K; Seliktar, Dror; Putnam, Andrew J (2013) Capillary morphogenesis in PEG-collagen hydrogels. Biomaterials 34:9331-40
Rao, Rameshwar R; Peterson, Alexis W; Ceccarelli, Jacob et al. (2012) Matrix composition regulates three-dimensional network formation by endothelial cells and mesenchymal stem cells in collagen/fibrin materials. Angiogenesis 15:253-64
Kniazeva, Ekaterina; Weidling, John W; Singh, Rahul et al. (2012) Quantification of local matrix deformations and mechanical properties during capillary morphogenesis in 3D. Integr Biol (Camb) 4:431-9

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