Vascularization remains a large obstacle for creating a functional pancreas-tissue equivalent for transplantation. In this study, a pre-vascularized pancreatic decellularized scaffold was prepared through endothelializing with endothelial progenitor cells (EPCs) in a bioreactor, and the ability to regenerate new blood vessels was detected in vivo. Initially, pancreases of Sprague-Dawley (SD) rats were perfused with 1% Triton X-100 and 0.1% ammonium hydroxide to remove the cellular components while the intact vascular network was preserved. Then, the decellularized scaffold was reseed with EPCs, which were primarily characterized by dual staining for dil-labeled acetylated low-density lipoprotein (Dil-acLDL) and fluorescein isothiocyanate labeled ulex europaeus agglutinin 1 (FITC-UEA-1), to reconstruct the vascular network. Thus, a scaffold covered with EPCs in the vessel structure was created. After that, the scaffold was transplanted into the rat in vivo to observe the anastomosis with the host vascular network. The results showed that EPCs can be located around the blood vessel wall, and re-endothelialized scaffold connected with the host through new blood vessel formation earlier than the control group (p < 0.05). These findings all indicated that the pancreatic decellularized scaffold endothelialized with EPCs may be further applied to solve the problem of blood supply and support the function of insulin-secreting cells after in vivo transplantation.