Study On The Mechanism Of Bone Marrow Mesenchymal Stem Cells Expressing Purinergic Receptor P2Y6 To Promote Osteogenic Differentiation Induced By Titanium Dioxide Nanotubes In Vitro

Background:The micromorphology of titanium dioxide(TiO₂)nanotubes can improve the wear resistance,corrosion resistance and mechanical properties of titanium(Ti).At the same time,it can induce bone marrow mesenchymal stem cells(BMSCs)to differentiate into osteoblasts and promote the formation of new bone tissue,but the exact mechanism has not been fully elucidated.Purinergic receptor P2Y6 can be expressed in BMSCs and participates in the regulation of bone metabolism,but it is not clear whether P2Y6 is involved in the osteogenic differentiation of BMSCs induced by TiO₂ nanotubes.Methods:TiO₂ nanotubes were prepared on the surface of titanium specimens using the anodizing method.The surface morphology of the nanotubes was observed by a scanning electron microscope(SEM).The culture environment of SD rat BMSCs in vitro was constructed,and the expression levels of P2Y6 and osteogenic differentiation markers in BMSCs on the surface of TiO₂ nanotubes were detected by quantitative reverse transcriptase polymerase chain reaction(q RT-PCR)and Western blotting(WB).After that,the specific inhibitors and agonists of P2Y6 were used to detect the expression levels of osteogenic differentiation markers and key sites in PKC?-ERK1/2pathway,and to explore the effects of purine receptor P2Y6 and PKC?-ERK1/2pathway on osteogenic differentiation of BMSCs induced by TiO₂ nanotubes.Results:The average inner diameter of the TiO₂ nanotubes increased with an increase in voltage(voltage range of 30–90V),and the expression of P2Y6 in BMSCs could be upregulated by TiO₂ nanotubes in osteogenic culture.Inhibition of P2Y6 expression partially inhibited the osteogenic effect of TiO₂ nanotubes and downregulated the activity of the PKC?-ERK1/2 pathway.When combined with P2Y6 agonists,the bone-promoting effect of TiO₂ nanotubes was more obvious.Conclusions:TiO₂ nanotubes promoted the P2Y6 expression of BMSCs during osteogenic differentiation and promoted osteogenesis by activating the PKC?-ERK1/2pathway.The combined application of TiO₂ nanotubes and P2Y6 agonists may be an effective new strategy to improve the osseointegration of titanium implants.