OBJECTIVE: The angiogenic potential of endothelial progenitor cells (EPCs) may be limited by the absence of their natural biologic foundation, namely smooth muscle pericytes. We hypothesized that joint delivery of EPCs and smooth muscle cells (SMCs) in a novel, totally bone marrow-derived cell sheet will mimic the native architecture of a mature blood vessel and act as an angiogenic construct to limit post infarction ventricular remodeling. METHODS: Primary EPCs and mesenchymal stem cells were isolated from bone marrow of Wistar rats. Mesenchymal stem cells were transdifferentiated into SMCs by culture on fibronectin-coated culture dishes. Confluent SMCs topped with confluent EPCs were detached from an Upcell dish to create a SMC-EPC bi-level cell sheet. A rodent model of ischemic cardiomyopathy was then created by ligating the left anterior descending artery. Rats were randomized into 3 groups: cell sheet transplantation (n = 9), no treatment (n = 12), or sham surgery control (n = 7). RESULTS: Four weeks postinfarction, mature vessel density tended to increase in cell sheet-treated animals compared with controls. Cell sheet therapy significantly attenuated the extent of cardiac fibrosis compared with that of the untreated group (untreated vs cell sheet, 198 degrees [interquartile range (IQR), 151-246 degrees] vs 103 degrees [IQR, 92-113 degrees], P = .04). Furthermore, EPC-SMC cell sheet transplantation attenuated myocardial dysfunction, as evidenced by an increase in left ventricular ejection fraction (untreated vs cell sheet vs sham, 33.5% [IQR, 27.8%-35.7%] vs 45.9% [IQR, 43.6%-48.4%] vs 59.3% [IQR, 58.8%-63.5%], P = .001) and decreases in left ventricular dimensions. CONCLUSIONS: The bone marrow-derived, spatially arranged SMC-EPC bi-level cell sheet is a novel, multilineage cellular therapy obtained from a translationally practical source. Interactions between SMCs and EPCs augment mature neovascularization, limit adverse remodeling, and improve ventricular function after myocardial infarction.