Abstract
Carbon nanotubes (CNTs) have performed outstandingly in the field of biomedicine due to their unique properties including electrical conductivity, nanostructures of hollow graphite cylinders, and mechanical strength. Based on these, we used multi-carbon nanotubes as biomaterials in directing neural differentiation and cell sheet preparation. The first part is that we demonstrated carboxylated multiwalled CNTs (MWCNTs) can induce and maintain neural differentiation of human bone marrow mesenchymal stem cells (hBMMSCs) without any exogenous differentiating factors, as evidenced by the protein expression. The low cytotoxicity of carboxylated MWCNTs was also shown by a proliferation assay. Quantitative real-time polymerase chain reaction (Q-PCR) data revealed that neural-associated genes, including growth and transcription factors, were promoted while bone-associated genes were inhibited when the cells were cultured on carboxylated MWCNTs. These up-regulated neural growth factors can also adsorb onto carboxylated MWCNTs. The data suggest that carboxylated MWCNTs play dual roles: promoting hBMMSC neural differentiation, including up-regulating the neural growth factors; and trapping these neural growth factors to create a suitable environment for long-term neural differentiation. The second part is fabrication of poly(N-isopropylacrylamide) (PNIPAAM) hydrogels interpenetrated with multiwalled CNTs (MWCNTs) as substrates for cell sheet preparation. The results demonstrate that PNIPAAM hydrogels with interpenetrating MWCNTs still exhibit thermosensitive behavior. It is also found that epithelial Madin-Darby canine kidney (MDCK) cells can only attach and proliferate on MWCNT-interpenetrated PNIPAAM hydrogels. Furthermore, the PNIPAAM hydrogels with MWCNTs possess higher elastic moduli and hydrophobicities than those without MWCNTs, suggesting these two characteristics are necessary for the cells to attach to the hydrogel surfaces. Moreover, cell sheets can only be harvested from PNIPAAM hydrogels with MWCNTs because of their high ratio of cell attachment. Thus, this simple method provides sufficient mechanical strength to PNIPAAM hydrogels so that anchorage-dependent cells can be cultivated and provides a superior system for preparing cell sheets.