###### Abstract  
Rapid new microvascular network induction was critical for bone regeneration, which required the spatiotemporal delivery of growth factors and transplantation of endothelial cells. In this study, the linear poly(d,l-lactic-\(co\)-glycolic acid)-\(b\)-methoxy poly(ethylene glycol) (PLGA-mPEG) block copolymer microspheres were prepared for simultaneously delivering vascular endothelial growth factor (VEGF) and monocyte chemotactic protein-1 (MCP-1). Then, vascular endothelial cells (VECs) with growth factor loaded microspheres were composited into a star-shaped PLGA-mPEG block copolymer solution. After this, composite hydrogel (microspheres ratio: 5 wt%) was formed by increasing the temperature to 37 \({}^{\circ}\)C. The release profiles of VEGF and MCP-1 from composite hydrogels in 30 days were investigated to confirm the different simultaneous delivery systems. The VECs exhibited a good proliferation in the composite hydrogels, which proved that the composite hydrogels had a good cytocompatibility. Furthermore, in vivo animal experiments showed that the vessel density and the mean vessel diameters increased over weeks after the composite hydrogels were implanted into the necrosis site of the rabbit femoral head. The above results suggested that the VECs-laden hydrogel composited with the dual-growth factor simultaneous release system has the potential to enhance angiogenesis in bone tissue engineering. composite hydrogel; simultaneous release; growth factor; angiogenesis; bone tissue engineering +
Footnote †: journal: Applied  
including angiogenesis and osteogenesis [1; 2; 3; 4]. Thereinto, the ingrowth of new blood vessels was critical for stimulating osteogenesis, and the formed capillary network also served as a template for new bone formation [5]. The lack of microvasculature formation remains a key roadblock to apply engineered composites to clinical applications up to now [6]. Several reports proved that many soluble growth factors could regulate vasculogenesis and angiogenesis, mainly including the vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), insulin-like growth factor (IGF), hepatocyte growth factor (HGF), platelet-derived growth factor-BB (PDGF-BB), monocyte chemoattractant protein-1 (MCP-1), etc. [7; 8; 9; 10; 11]. VEGF was one of the most important growth factors across all stages of neovascularization, mainly through initiating the sprouting of existing blood vessels by mitogenic and chemotactic effects, and facilitating the recruitment of circulating endothelial progenitor cells [9; 10]. Furthermore, VEGF was also important for maintaining the survival of blood vessels, the data indicated that the withdrawal of VEGF caused vascular collapse and regression [9; 10; 11]. As a matter of fact, a sustaining release of VEGF at least for a period of four weeks could improve the maturing and stabilizing of newly formed blood vessels [12]. On the other hand, the VEGF could not only stimulate the proliferation, migration, and survival of VECs, but also enhanced the alkaline phosphatase activity, mineralized tissue production, and adhered molecule, which played an important role in fracture healing [13; 14]. Because of its short half-life and enzymatic degradation of the growth factors, some side effects were unavoidable by using very high doses. Thus, an effective delivery system was significant. On the other hand, new bone formation and angiogenesis were intimately connected during tissue regeneration [5]. Many studies confirmed that bone tissue regeneration could be ameliorated via the spatiotemporal release of VEGF and bone growth factors, for instance, bone morphogenetic protein-2 (BMP-2) [15; 16; 17]. MCP-1, known as chemokine (C-C motif) ligand 2 (CCL2), was an arteriogenic factor that could induce a remarkable host vascularization response at an early stage, and it was clear that the dual delivery of MCP-1 and VEGF induced the mural cell recruitment and improved survival of endothelial cells (ECs) [4].