Fibronectin mediates cell adhesion, migration and initiates extracellular matrix assembly. Therefore, it plays critical roles on wound healing... Show moreFibronectin mediates cell adhesion, migration and initiates extracellular matrix assembly. Therefore, it plays critical roles on wound healing processes. The purpose of the study is to elucidate the effects of the combination of two functional domains, III1-2 and III9-10 subunits, on the fibronectin matrix assembly. In the present study, NIH 3T3 cells were cultured on the functionalized polyurethane (0.5 μM III9-10 and the mixture with 0.5 μM III9-10 and 0.5 μM III1-2). Functionalization enabled the formation of the covalent linkages between the coverslip polyurethane and the recombinant proteins. Surface characterization was examined by ELISA. NIH3T3 cells were plated on the functionalized coverslips and harvested after 3, 6, 12, and 24 hours culture. Total fibronectin and extracellular matrix fibronectin which is insoluble in deoxycholic acid (DOC) were analyzed through western blotting. The quantification results showed that the mixture groups tended to have higher total fibronectin than the III9-10 alone groups at most time point (3, 6, and 12 hours). Especially obvious differences of DOC-insoluble fibronectin between the mixture group and III9-10 alone group were found in the early stage (3 hour) of the fibronectin matrix assembly. Immunostaining images also showed that fiber-like FN had been generated after three hours culture in the mixture group but FN still looked faint in III9-10 group. Longer and larger FN fibrils were identified in the mixture groups than in III9-10 groups at all time points. In conclusion, III1-2 enhances fibronectin matrix assembly when present with III9-10 through increase the rate of the formation of DOC-insoluble fibronectin in 24 hours culture. M.S. in Biology, July 2011 Show less
