Spatiotemporal release of VEGF from biodegradable polylactic-co-glycolic acid microspheres induces angiogenesis in chick chorionic allantoic membrane assay

The file attached to this record is the author's final peer reviewed version. The Publisher's final version can be found by following the DOI link.While vascular endothelial growth factor (VEGF) is an acknowledged potent pro-angiogenic agent there is a need to deliver it at an appropriate concentration for several days to achieve angiogenesis. The aim of this study was to produce microspheres of biodegradable polylactic-co-glycolic acid (PLGA) tailored to achieve sustained release of VEGF at an appropriate concentration over seven days, avoiding excessive unregulated release of VEGF that has been associated with the formation of leaky blood vessels. Several formulations were examined to produce microspheres loaded with both human serum albumin (HSA) and VEGF to achieve release of VEGF between 3 and 10 ng per ml for seven days to match the therapeutic window desired for angiogenesis. In vitro experiments showed an increase in endothelial cell proliferation in response to microspheres bearing VEGF. Similarly, when microspheres containing VEGF were added to the chorionic membrane of fertilised chicken eggs, there was an increase in the development of blood vessels over seven days in response, which was significant for microspheres bearing VEGF and HSA, but not VEGF alone. There was an increase in both blood vessel density and branching – both signs of proangiogenic activity. Further, there was clearly migration of cells to the VEGF loaded microspheres. In summary, we describe the development of an injectable delivery vehicle to achieve spatiotemporal release of physiologically relevant levels of VEGF for several days and demonstrate the angiogenic response to this. We propose that such a treatment vehicle would be suitable for the treatment of ischemic tissue or wounds

Developing A Synthetic Composite Membrane For Cleft Palate Repair

An oronasal fistula is a passage between the oral and nasal cavity. Currently, surgical procedures use mucosal flaps or collagen grafts to make a barrier between oral and nasal cavities. Our aim was to develop a cell-free synthetic repair material for closure of nasal fistulas. We surface functionalized electrospun polyurethane (PU) and poly-L-lactic acid (PLLA) and composite polymer (PU-PLLA) membranes with acrylic acid through plasma polymerization. Membranes were treated in a layer-by-layer approach to develop highly charged electrostatic layer that could bind heparin as a pro-angiogenic glycosaminoglycan. The properties were evaluated through physical, chemical, and mechanical characterization techniques. Cytotoxicity was tested with MC3T3 pre-osteoblast cell lines for 3, 7, and 14 days, and vasculogenesis was assessed by implantation into the chorio-allantoic membrane in chick embryos for 7 days. In vivo biocompatibility was assessed by subcutaneous implantation in rats for 1, 3, and 6 weeks. The membranes consisted of random fibers of PLLA-PU with fiber diameters of 0.47 and 0.12 μm, respectively. Significantly higher cell proliferation and migration of MC3T3 cells at 3, 7, and 14 days were shown on plasma-coated membranes compared with uncoated membranes. Further, it was found that plasma-coated membranes were more angiogenic than controls. In vivo implantation of membranes in rats did not reveal any gross toxicity to the materials, and wound healing was comparable with the native tissue repair (sham group). We therefore present a plasma-functionalized electrospun composite polymer membrane for use in the treatment of fistulas. These membranes are flexible, non-cytotoxic, and angiogenic, and we hope it should lead to permanent closure of oronasal fistula

Developing a layer-by-layer coating to make materials angiogenic for urological applications

Tissue engineered constructs often fail due to poor integration with the patient’s tissues. Specifically, they fail to be vascularised, leading to the death and loss of the implanted tissues. Thus, there is a need to produce angiogenic materials to improve tissue integration. We describe the development of a layer-by-layer (LBL) approach to coat electrospun scaffolds to help promote angiogenesis into these biomaterials once implanted. Initially, Electrospun poly-L-lactic acid (PLLA) was coated comparing two different techniques – one using alternative layers of PolyEthyleneImine (PEI) and PolyAcrylic Acid (PAC) and one with alternative layers of PEI and heparin (HEP) for a total of seven layers in both cases. Both scaffolds were then coated with heparin as the final layer. The scaffold coated with alternate PEI and PAC showed a clear ability to bind the most heparin. This scaffold was then studied further for its ability to bind VEGF which was confirmed using an ELISA. The scaffold coated with 7 alternate layers of PEI and PAC and heparin was then implanted in a chick chorionic allantoic membrane (CAM) assay. After a period of 7 days in the CAM, the coated scaffold showed strong angiogenic activity. In contrast, the uncoated scaffolds did not promote angiogenesis. Sub sequentially, this approach was used to coat two other different materials, electrospun polyurethane and a commercially available polypropylene mesh. They were also implanted into the chick CAM. We conclude that this approach to functionalising scaffolds is effective within a clinically relevant time period (7 days in an in vivo angiogenic model) and suggest this will be useful for improving integration of scaffolds once implanted