The healing of full-thickness burns treated by using plasmid DNA encoding VEGF-165 activated collagen–chitosan dermal equivalents

Abstract

Repair of deep burn by use of the dermal equivalent relies strongly on the angiogenesis and thereby the regeneration of dermis. To enhance the dermal regeneration, in this study plasmid DNA encoding vascular endothelial growth factor-165 (VEGF-165)/N,N,N-trimethyl chitosan chloride (TMC) complexes were loaded into a bilayer porous collagen–chitosan/silicone membrane dermal equivalents (BDEs), which were applied for treatment of full-thickness burn wounds. The DNA released from the collagen–chitosan scaffold could remain its supercoiled structure but its degree was decayed along with the prolongation of incubation time. The released DNA could transfect HEK293 cells in vitro with decayed efficiency too. Human umbilical vein endothelial cells (HUVECs) in vitro cultured in the scaffold loaded with TMC/pDNA-VEGF complexes expressed a significantly higher level of VEGF and showed higher viability than those cultured in the controls, i.e. blank scaffold, and scaffolds loaded with naked pDNA-VEGF and TMC/pDNA-eGFP, respectively. The four different BDEs were then transplanted in porcine full-thickness burn wounds. Results showed that the TMC/pDNA-VEGF group had a significantly higher number of newly-formed and mature blood vessels, and fastest regeneration of the dermis. RT-qPCR and western blotting found that the experimental group also had the highest expression of VEGF, CD31 and α-SMA in both mRNA and protein levels. Furthermore, ultra-thin skin grafting was performed on the regenerated dermis 14 days later, leading to complete repair of the burn wounds with normal histology. Moreover, the tensile strength of the repaired tissue increased along with the time prolongation of post grafting, resulting in a value of approximately 70% of the normal skin at 105 days.