In vivo osteogenic response to different ratios of BMP-2 and VEGF released from a biodegradable porous system Antonio Herna ´ ndez, 1,2 Ricardo Reyes, 1,3 Esther S anchez, 1,3 Marı´aRodrı´guez-E ´ vora, 1,3 Araceli Delgado, 1,3 Carmen E ´ vora 1,3 1 Department of Chemical Engineering and Pharmaceutical Technology, University of La Laguna, 38200 La Laguna, Spain 2 Traumatology Service, Hospit en Rambla Ltd., Santa Cruz de Tenerife, Spain 3 Institute of Biomedical Technology, University of La Laguna, Spain Received 5 October 2011; revised 20 February 2012; accepted 12 March 2012 Published online 24 April 2012 in Wiley Online Library (wileyonlinelibrary.com). DOI: 10.1002/jbm.a.34183 Abstract: Bone regeneration and vascularization with porous PLGA scaffolds loaded with VEGF (0.35 and 1.75 lg) and BMP-2 (3.5 and 17.5 lg), incorporated in PLGA microspheres, or the combination of either dose of BMP-2 with the low dose of VEGF were investigated in an intramedullary femur defect in rabbits. The system was designed to control growth factor (GF) release and maintain the GFs localized within the defect. An incomplete release was observed in vitro whereas in vivo VEGF and BMP-2 were totally delivered during 3 and 4 weeks, respectively. A weak synergistic effect of the dual delivery of VEGF and BMP-2 (high dose) was found by 4 weeks. However, the absence of an apparent synergistic long-term effect (12 weeks) of the combination over BMP-2 alone suggests that more work has to be done to optimize VEGF dose, sequential presentation, and the ratio of the two GFs to obtain a beneficial bone repair response. V C 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 100A: 2382–2391, 2012. Key Words: VEGF, BMP-2, PLGA, bone regeneration, delivery system How to cite this article: Herna ´ ndez A, Reyes R, S anchez E, Rodrı ´guez-E ´ vora M, Delgado A, E ´ vora C. 2012. In vivo osteogenic response to different ratios of BMP-2 and VEGF released from a biodegradable porous system. J Biomed Mater Res Part A 2012:100A:2382–2391. INTRODUCTION Bone regeneration is a complex process that requires the coordinated activity of multiple cell types, involving a con- certed regulatory system of growth factors (GFs) and cyto- kines. Synergistic contribution and crosstalk between GF signaling pathways was suggested to be crucial for the over- all outcome in osteogenesis. 1–3 Prominent amongst osteo- genic factors are the bone morphogenetic proteins (BMPs), which play an important role in many processes associated with embryogenesis and in adult bone repair. 4–7 Bone is a highly vascularized tissue reliant on the close spatial and temporal association between blood vessels and bone cells to maintain skeletal integrity. Angiogenesis thus plays a piv- otal role in skeletal development and bone fracture repair. 8 A variety of molecules are implicated in tissue vasculariza- tion. Vascular endothelial growth factor (VEGF) is regarded as the key molecule for neo-angiogenesis and also plays a significant role in skeletal growth and repair. 9,10 Thus, it seems to be a crucial molecule to address bone repair. How- ever, according to the complexity of osseous regeneration, it is improbable that a single factor, no matter how potent it is, could induce bone healing without the contribution of other GFs, cytokines, and cells. Therefore, combinations of growth factors have been intensely investigated in bone regeneration studies. 7 What does seem clear to date is that BMPs have the potential to significantly improve management of critical bone defects, on the one hand, and that VEGF plays an important role during the initial phase of fracture healing 10,11 and bone regeneration, on the other hand. 8 Despite some authors report a synergistic effect of dual delivery of VEGF and BMP-2 on bone formation and suggest interplay between these GFs in bone regeneration, inconsis- tent results can be found in the recent literature. The stud- ies by Peng et al., 12 Kakudo et al., 13 and Kanczler et al. 14 seem to suggest that the combination of BMP-2 and VEGF provides bone tissue engineering constructs with increased osteogenic potential over BMP-2 or VEGF alone. However, the results obtained by Young et al., 15 Patel et al., 16 and Kempen et al. 17 suggest an unclear long-term effect of the combined delivery of VEGF and BMP-2 versus BMP-2 alone. These contradictions may be result of various discrepan- cies as for example in doses and routes of GF administration as well as the ratio of the combinations and the delivery kinetics. Although GFs are clearly implicated in harnessing and controlling cell activity in tissue regeneration, the Correspondence to: C. E ´ vora; e-mail: cevora@ull.es Contract grant sponsor: Ministry of Science and Technology; contract grant number: MAT2011-23819 2382 V C 2012 WILEY PERIODICALS, INC.