A Thermoresponsive Chitosan−NIPAAm/Vinyl Laurate Copolymer Vector for Gene Transfection

Abstract

A carboxyl-terminated N-isopropylacrylamide/vinyl laurate (VL) copolymer was prepared and coupled with chitosan (molecular weight = 2000) to produce a chitosan-NIPAAm/VL copolymer (PNVLCS) vector. The aqueous solution of PNVLCS displayed an obvious thermoresponsive behavior with a lower critical solution temperature (LCST) about 26 degrees C. The transmission electron microscopy (TEM) showed that the size of PNVLCS/DNA complexes varied with charge ratios (+/-), and the smaller nanoparticles were formed at higher charge ratios. DLS revealed that the size of complex particles was dependent on temperature. The results of temperature-variable circular dichroism (CD), UV, and electrophoresis retardation indicated that at lower charge ratios, DNA in the complexes assume a B conformation, whereas increasing charge ratios caused B --> C type conformation transformation; the dissociation-formation of PNVLCS/DNA complexes could be tuned by varying temperature: at 37 degrees C, the collapse of PNIPAAm in PNVLCS was favorable for the formation of compact complexes, shielding more DNA from exposure; at 20 degrees C, the hydrated and extended PNIPAAm chains facilitated the unpacking of DNA from PNVLCS, increasing the exposure of DNA. PNVLCS was used to transfer plasmid-encoding beta-galactosidase into C2C12 cells. The level of gene expression could be controlled by varying incubation temperature. The transfection efficiency of PNVLCS was well improved by temporarily reducing culture temperature to 20 degrees C, whereas naked DNA and Lipofectamine 2000 did not demonstrate the characteristics of thermoresponsive gene transfection.