ABSTRACT Cell therapy combined with biomimetic polymer engineering offers a promising strategy for cardiovascular regeneration. Compartmentalizing single cells in hydrogels creates three‐dimensional (3D) micro‐niches that enhance bio‐responsiveness and the therapeutic efficacy of cell therapy. A non‐microfluidic strategy for single‐cell encapsulation using a stimuli‐responsive amphiphilic copolymer, gelatin‐poly(glycerol sebacate)‐methacrylate (GPM) is developed. When incubated with cells at room temperature, GPM self‐assembled into single‐cell vesicles via ligand–receptor–mediated interactions, creating a nanoscale matrix around each cell with near 100% encapsulation efficiency. The presence of bioactive moieties, dynamic mechanical properties, and matrix metalloproteinase mediated degradability enabled reciprocal, cell‐matrix interactions. Consequently, 3D cultures of GPM vesicles encapsulating single endothelial cell (GPM/EC vesicles) promoted deposition of thread‐like extracellular matrix (ECM), which facilitated cell migration and served as a guidance scaffold for vessel‐like network formation. For enhanced in vivo distribution and immunoprotection, GPM/reprogrammed EC vesicles were assembled into injectable microspheres, termed mGPM‐rEC. Upon injection into a mouse hindlimb ischemia model, mGPM‐rEC demonstrated robust cell survival and intramuscular migration with the deposition of dense ECM. This ECM functioned as a reservoir for growth factors and signaling molecules, recruiting host vasculature to promote neovascularization and providing structural and mechanical support for implanted cells to form de novo blood vessels.