Influences of TiO2 nanotubes and chemical stimulation on osteogenetic differentiation of MSCs

Abstract:

Objective：To investigate the osteogenic differentiation of bone marrow mesenchymal stem cells（MSCs） derived from Sprague Dawley rats on TiO2 nanotubes under different inducing conditions. Methods：Isolated and purified MSCs were cultured on the surface structures of plane titalium and TiO2 nanotubes anodized of 30，70，100 nm diameter. Four osteogenic differentiation methods were ap－proached：regular culture medium（control group），regular medium with L-Ascorbic acid and β-glycerophosphate，regular medium with L-ascorbic acid，β-glycerophosphate and 1α，25-dihydroxyvitamin D3 and regular medium with L-ascorbic acid，β-glycerophosphate and dexamethasone. Alkaline phosphatase（ALP） activities of all groups were analyzed on 3，7，14 d after the culture，respectively. Calcium deposition was detected on 21 d after the culture. Real-time PCR was applied to quantitate the expression of osteoblast related gene Runx2 and OSX on 14 d after the culture. Results：MSCs cultured on the same surface structure demonstrated higher ALP activity（F=338.542，P=0.000），calcium deposition（F=417.012，P=0.000） as well as Runx2（F=14.419，P=0.000） and OSX（F=42.011，P=0.000） gene expression in the L-ascorbic acid + β-glycerophosphate + dexamethasone group than in the others，with statistically significant differences. Within the same culture condition，ALP activity（F=53.170，P=0.000），calcium deposition（F=264.268，P=0.000） and Runx2（F=3.196，P=0.037） and OSX（F=5.895，P=0.003） gene expression were much higher on the surface structure of 30 nm diameter TiO2 nanotubes，with statistically significant differences. Culture condition and material structure had interactions（ALP activity（F=6.322，P=0.000），calcium deposition（F=33.330，P=0.000） and gene expression of OSX（F=2.825，P=0.015）），in which L-ascorbic acid + β-glycerophosphate + dexamethasone culture and 30 nm diameter TiO2 nanotubes was the best combination. Conclusion：The osteogenic differentiation of MSCs can be influenced by both the structure of material surface and chemical stimulation. On the same surface structure，MSCs in the L-ascorbic acid + β-glycerophosphate + dexamethasone culture group has greater potential of osteogenic differentiation than the others. Under the same chemical stimulation，MSCs on 30 nm diameter TiO2 nanotubes are more apt to differentiating towards osteoblasts than the way on other surface structures. The combination condition of L-ascorbic acid + β-glycerophosphate + dexamethasone culture and 30 nm diameter TiO2 nanotubes is most conductive to the os－teogenic differentiation of MSCs.