MMPs-responsive release of DNA from electrospun nanofibrous matrix for local gene therapy: In vitro and in vivo evaluation

摘要

DNA-incorporated nanofibrous matrix was fabricated to control release of DNA in response to high concentration of MMPs such as diabetic ulcers. Nanofibrous matrix was electrospun employing amine groups-functionalized block copolymers. Linear polyethyleneimine was chemically conjugated to surface-exposed amine groups of the nanofibrous matrix by a MMP-cleavable peptide linkage. After confirming conjugation of LPEI by a survey scan of X-ray photoelectron spectroscopy, DNA was electrostatically incorporated on the matrix at various charge ratios of DNA to the immobilized LPEI. Release profiles of DNA and LPEI were monitored in the presence of MMP-2 to examine MMP-responsiveness of the matrix. In vitro transfection efficiencies were examined by employing the released fraction of DNA and LPEI. Animal study was performed to examine in vivo transfection efficiency of DNA-incorporated LPEI nanofibrous matrix in comparison to unmodified controls and naked DNA. Surface-exposed amine groups of the matrix at 0.387. nmol/mg were reacted with a MMP-responsive peptide and subsequently employed for LPEI immobilization. DNA and LPEI were fast released out in the presence of MMP-2 and the released DNA and the release LPEI were confirmed to form electrostatic complex according the charge ratios by dynamic light scattering and gel retardation assay. Transfection efficiency of the release fractions was significantly dependent on the charge ratios of the incorporated DNA and the matrix rather than the amount of the released DNA in the released fractions. Animal experiments employing diabetic animals with dorsal ulcers revealed that the LPEI-immobilized nanofibrous matrix was superior to naked DNA or DNA-incorporated matrix without LPEI in terms of in vivo transfection efficiencies. Thus, controlled release of DNA in response to MMPs can be potentially applied to local gene delivery system for treating diabetic ulcers.