The use of gene activated matrix to mediate effective SMAD2 gene silencing against hypertrophic scar

Hypertrophic scar (HS) originates from the over-expression of transforming growth factor β (TGF-β) and downstream SMAD2. With attempts to rectify HS by RNA interference (RNAi) against SMAD2, we report the design of plasmid DNA encoding SMAD2 siRNA (pSUPER-SMAD2), and identify the optimal siRNA sequence toward maximal RNAi efficiency. To realize effective and sustained RNAi, we developed gene activated matrix (GAM) based on porous atelocollagen scaffold and embedded trimethyl chitosan-cysteine (TMCC)/pSUPER-SMAD2 polyplexes for promoting cell growth and gene transfection. The GAM exhibited porosity higher than 80%, pore size of 200-250 μm, desired mechanical strength, and sustained pSUPER-SMAD2 release profiles. Normal skin fibroblasts (NSFs) and hypertrophic scar fibroblasts (HSFs) were allowed to infiltrate and proliferate in GAM; at the meantime they were transfected with TMCC/pSUPER-SMAD2 polyplexes to display remarkably reduced SMAD2 levels that lasted for up to 10 days, consequently inhibiting the over-production of type I and type III collagen. We further unraveled the notably higher transfection levels of GAM in three-dimensional (3D) than in 2D environment, which was attributed to the improved cell-matrix interactions that promote cell proliferation and polyplex internalization. This highly safe and effective GAM may serve as a promising candidate towards HS treatment.