Abstract 14370: Direct Conversion of Adult Fibroblasts to Functional Endothelial Cells by Defined Factors

Endothelial dysfunction contributes significantly to the development of ischemic vascular and heart diseases. Cell based therapies to augment endothelial cells (EC) and vascular function as a whole hold great therapeutic promise. Herein, we report the novel approach to generate functional endothelial cells directly from adult fibroblasts. Methods & Results: 11 genes, known key regulators of EC development, were selected by literature review. Skin fibroblasts (SFBs) were prepared from 8 week old male Tie2-GFP mice and SFBs infected with lentiviruses allowing for simultaneous overexpression of all 11 factors. Tie2-GFP + cells (0.9%), representing positive Tie2 expression, were detected by FACS. Further studies revealed 5 key genes (5F) which were required for efficient reprogramming of SFBs to Tie2-GFP + cells (4%). Moreover, following 5F infection, GFP + cells exhibited endothelial-like cobblestone morphology and expressed molecular makers of EC including VE cadherin, CD31, ICAM2, and Tie2. These induced endotheial cells (iECs) were capable of BS1 lectin binding, acLDL uptake and capillary formation on Matrigel. VE cadherin and Tie2 promoters of iECs were demethylated. mRNA profiling showed highly enriched endothelial genes in iECs. Further, iECs were found to cluster with bona fide ECs isolated from mouse lungs and an MS1 EC line, but were distinct from original SFB. 5F cocktail was able to reprogram tail tip FBs with efficiency of 7.2% suggesting the universal effects of our iEC 5F. Importantly, our novel reprogramming strategy did not involve activation of pluripotency, as neither Oct4 nor nanog expression was induced with 5F overexpression. In a mouse hindlimb ischemia model, iEC implantation enhanced limb perfusion, and increased capillary density. Conclusions: We demonstrate the first direct reprograming of adult fibroblasts to functional EC and provide a novel therapeutic modality for regenerative medicine in ischemic vascular and heart diseases.