Surface mediated in situ differentiation of mesenchymal stem cells on gene-functionalized titanium films fabricated by layer-by-layer technique.

In this work, multilayered and gene-functionalized titanium films composed of chitosan (Chi) and plasmid DNA (pEGFP-hBMP2, pGB) were employed to investigate the surface mediated in situ differentiation of mesenchymal stem cells (MSCs). The Chi/pGB multilayered structures were fabricated by layer-by-layer (LbL) assembly technique and degraded to release plasmid DNA complexes depending on bilayer numbers over 7 days. Therefore, the differentiation behaviors of MSCs cultured onto Chi/pGB multilayered titanium films surface were investigated. Chi/pGB LbL-modified titanium films show significant higher (p<0.01) transfection efficiency than those of other groups transfected by lipofectamine 2000 regarding the expression of green fluorescent protein (GFP). Reverse transcription-polymerase chain reaction (RT-PCR) assay revealed that MSCs adhered onto Chi/pGB LbL-modified titanium films could still express hBMP2 mRNA over 7 days culture. Compared with control groups, MSCs cultured onto Chi/pGB LbL-modified titanium films display significantly higher (p<0.01 or p<0.05) production levels of alkaline phosphatase (ALP) and osteocalcin over 7 days and 14 days culture, respectively. These results demonstrate that Chi/pGB LbL-modified titanium films are beneficial for sustained in situ inducing osteoprogenitor cells to differentiate into mature osteoblasts over long time. The approach presented here has potential applications in the development of gene-stimulating biomaterials and implant technology.