The Effects of Air Cold Atmospheric Plasma on Cellular Early Attachment, Proliferation and Migration on Pure Titanium Surfaces

Cold plasma has been studied for several fields of medicine, for example, pathogen inactivation and implant functionalization. Previous studies have provided evidence that plasma treatment promotes the adhesion of osteoblasts and is becoming a popular method for modifying the characteristics of substrate surfaces. However, its clinical application is limited as the effects of plasma treatment cannot be maintained for a long time. Air cold atmospheric plasma (CAP) is cost-efficient and convenient for clinical application compared to other work gas atmospheric plasma. In this study, the behavior of MC3T3-E1 cells on titanium discs was analyzed after treatment with air CAP. The characteristics of the titanium surfaces before and after the air CAP treatment were analyzed by a field emission scanning electron microscope (SEM), atomic force microscope (AFM), X-ray photoemission spectroscopy (XPS) and contact angle measurements. The morphologies of cells attached to the titanium surfaces were observed by SEM and fluorescence microscope at 2, 8 and 24 h after seeding. The gene expression levels of integrin beta 1, alpha 2 and alpha 5 were examined by RT-PCR after incubating 2, 8 and 24 h, respectively. The abilities of cell proliferation and migration were studied by MTT assay and migration assay, respectively. The roughness of titanium surfaces with and without air CAP were 30.6 +/- 9.00 nm and 27.66 +/- 3.68 nm, respectively. There was no obvious difference (p > 0.05). The air CAP made the titanium surfaces more hydrophilic. The MC3T3-E1 cells adhered to the untreated surface were fusiform, whereas the cells covered a larger surface area on the air CAP-treated surface at 2 h. The gene expression levels of integrin beta 1, alpha 2, and alpha in cells on the surfaces treated by air CAP were upregulated at 2 and 8 h compared to those untreated. And the air CAP-treated titanium surfaces enhanced cell proliferation and migration with more developed cellular network. In conclusion, air CAP treatment is a potential surface modifying method that can enhance the initial cellular attachment, proliferation and migration. Since the effects of plasma treatment cannot be maintained for a long time, it is expected that implants treated by air CAP immediately before implantation could improve the successful rate of implants.