Effect of porous tantalum on promoting the osteogenic differentiation of bone marrow mesenchymal stem cells in vitro through the MAPK/ERK signal pathway

Background: As an ideal new graft material, porous tantalum (pTa) has excellent mechanical properties and corrosion resistance and has received increased attention in the biomedical field because of its excellent cytocompatibility and ability to induce bone formation. However, the molecular mechanism of its potential to promote osteogenesis remains unclear, and very few reports have been published on this topic. Methods: In this study, we first produced porous Ti6Al4V (pTi6Al4V) and pTa with the same pore size by three-dimensional printing combined with chemical vapour deposition. The number of adhesions between pTa and pTi6Al4V and bone marrow mesenchymal stem cells (BMSCs) after 1 day of culture was detected by the live/dead cell staining method. The proliferation activity of the two groups was determined after culture for 1, 3, 5 and 7 days by the cell counting kit-8 method. In addition, the osteogenic activity, mRNA expression levels of osteogenic genes alkaline phosphatase (ALP), osterix (OSX), collagen-I (Col-I), osteonectin (OSN) and osteocalcin (OCN) and protein expression levels of the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signalling pathway marker p-ERK of the two groups cultured for 7, 14 and 21 days were determined by the ALP activity assay, real-time quantitative polymerase chain reaction (Q-PCR) and Western blotting, respectively. Subsequently, the two groups were treated with the MAPK/ERK-specific inhibitor U0126, and then, the mRNA expression levels of osteogenic genes and protein expression levels of p-ERK in the cultures were determined by Q-PCR and Western blotting, respectively. Results: The live/dead cell staining and cell counting kit-8 assays showed that the adhesion and proliferation activities of BMSCs on pTa were significantly better than those on pTi6Al4V. In addition, the ALP activity assay and Q-PCR showed that pTa harboured osteogenic activity and that the osteogenic genes ALP, OSX, Col-I, OSN and OCN were highly expressed, and by Western blotting, the expression of p-ERK protein in the pTa group was also significantly higher than that in the pTi6Al4V group. Subsequently, using the MAPK/ERK-specific inhibitor U0126, Western blotting showed that the expression of p-ERK protein was significantly inhibited and that there was no difference between the two groups. Furthermore, Q-PCR showed that osteogenic gene expression and ALP expression levels were significantly increased in the pTa group, and there were no differences in the OSX, Col-I, OSN and OCN mRNA expression levels between the two groups. Conclusion: Overall, our research found that compared with the widely used titanium alloy materials, our pTa can promote the adhesion and proliferation of BMSCs, and the molecular mechanism of pTa may occur via activation of the MAPK/ERK signalling pathway to regulate the high expression of OSX, Col I, OSN and OCN osteogenic genes and promote the osteogenic differentiation of BMSCs in vitro. The translational potential of this article: Our self-developed pTa material produced by three-dimensional printing combined with the chemical vapour deposition method not only retains excellent biological activity and osteoinductive ability of the original tantalum metal but also saves considerably on material costs to achieve mass production of personalised orthopaedic implants with pTa as a stent and to accelerate the wide application of pTa implants in clinical practice, which have certain profound significance. (C) 2019 The Authors. Published by Elsevier (Singapore) Pte Ltd on behalf of Chinese Speaking Orthopaedic Society.