Effects Of Strontium-containing Hydroxyapatite On Proliferation, Differentiation And Mineralization Of Osteoblasts In Vitro

Objective: The purpose of this in vitro study was to compare strontium-containing hydroxyapatite (Sr-HA)bioceramic with hydroxyapatite (HA)bioceramic on the effects upon the proliferation,differentiation and mineralization of osteoblast cells, thereby exploring the mechanism of Sr-HA on bone formation,thus to provide theoretical guidance to further investigation, development and improvement of biological materials.Methods:Primary osteoblasts were cultured from SD rats' calvaria by enzyme digestion method , and then identified, purified and passaged. Four types of bioceramic powders extracts(pure HA,1%Sr-HA,5% Sr-HA,10% Sr-HA) were prepared, and the strontium ion content of each type was detected. thereafter, these 4 types of extracts were co-cultured with the third passage of osteoblast cells respectively. The effects of extracts upon cell proliferation were measured by MTT ,the effects of extracts upon osteoblast alkaline phosphatase activity were measured by Alkaline phosphatase activity test kit, the specific transcription factors :core-binding factors(cbfa1)were measured by RT-PCR,and the effects of extracts upon cell mineralization were measured by Alizarin Red staining.Results: Determination of ALP activity revealed: on the 3rd day of cell cultivation, the ALP activity of each group had no significant difference (P> 0.05);With the extension of time, the ALP activity of each group has increased;on the 14th as well as 21st days of cell cultivation,it was found that Sr-HA was superior to HA in the secretion of ALP,and the difference was extremely significant (P <0.01); 5%Sr-HA group was shown to have the most significantly promoting effect on ALP activity of osteoblasts,however, compared with the 10%Sr-HA group, there was no significant difference (P> 0.05). Analysis on cbfa1 mRNA expression through RT-PCR showed:Sr-HA was superior to HA in the expression of cbfa1 mRNA,and the difference was extremely significant (P <0.01); 5%Sr-HA group was shown to have the most significantly promoting effect on the expression of cbfa1 mRNA,however, compared with the 10%Sr-HA group, there was no significant difference (P> 0.05). Calculation of mineralization nodules in vitro indicated : on the 14th day as well as 21st days of cell cultivation ,it was found that Sr-HA was superior to HA in the mineralization of osteoblast cells, and the difference was extremely significant (P <0.01),especially the 5% Sr-HA group (P <0.01). Results of cell proliferation showed:there was no significant difference in cell proliferation for each group(P>0.05).Conclusion: our results suggest that compared with HA bioceramic ,Sr-HA bioceramic is superior in the expression of cbfa1 mRNA and secretion of alkaline phosphatase,which can accelerate the differentiation and mineralization of osteoblasts in vitro,thus to activate bone formation. It has been proved to be a kind of good repair material with better biological characteristics for internal fixating and bone filling ,and the prospect of its clinical application is broad.