Effects Of Domestic Porous Tantalum Composite SD Rats Bone Marrow Mesenchymal Stem Cells On Osteoblast Differentiation And Biological Activity

Objective To investigate the effect of domestic porous tantalum combined with bone morphogenetic protein-2(BMP-2)on the differentiation of SD rat bone marrow mesenchymal stem cells(BMSCs)into osteoblasts and the expression of related osteogenic factors under osteogenic induction.Methods 1 BMSCs of SD rats growing for 1 month were cultured to the 3rd generation and induced to osteoblast direction,chondroblast direction and adipoblast direction respectively.Alizarin red,alixin blue and oil red O staining were performed to identify after 14 days of continuous induction.2 BMSCs were compounded with porous tantalum and their proliferation was detected by cell counting.3 14 days after osteogenesis induction,BMSCs-porous tantalum composite was stained by phalloidin to detect its composite growth.4 experimental grouping: group a(blank control group): BMSCs alone plus complete osteoblast conditioned medium culture;Group b(porous tantalum group): BMSCs were inoculated with tantalum scaffold material and cultured in complete osteoblast conditioned medium.Group C is(BMP-2 group): BMSCs are cultured with BMP-2-containing complete osteogenic conditioned medium,while Group D is(porous tantalum-BMP-2 group): BMSCs are cultured with BMP-2-containing complete osteogenic conditioned medium after being inoculated with tantalum scaffold material.The expression of COL-?,OCN,OPN mRNA and protein after BMSCs differentiated into osteoblasts were detected by ELISA,real-time fluorescence quantitative PCR and Western-blot after 7,14 and 21 days of induction culture.Results 1 The cultured BMSCs were extracted and cultured to the third generation for testing.After BMSCs were continuously induced toward osteoblasts,the cell morphology changed.After 3 days of induction,the cell arrangement had certain directivity and showed vortex-like dense growth.Identification of BMSCs: alizarin red staining was positive 14 days after induction of bmscs into osteoblasts;Alixinblue staining was positive after induction toward chondrocytes.Oil red O staining was positive after induction toward adipocytes.3 CCK-8 method showed no significant difference in cell proliferation between the porous tantalum group and the blank control group(P>0.05).As time went on,the difference was statistically significant(P<0.05)between the groups at different time points.It was confirmed that porous tantalum had no effect on BMSCs growth.4 Fluorescence microscope observation of domestic porous tantalum-BMSCs complex stained by phalloidin showed that BMSCs adhered to the surface of porous tantalum and grew inside,secreting many cell products at the same time.5 ELISA;6 qRT-PCR;7 Western-blot results showed that the expressions of OCN,OPN,col-I in porous tantalum-BMP-2 group and BMP-2 group were higher than those in blank control group and porous tantalum group(P<0.05)at the initial stage of osteoblast induction.However,compared with the expression of porous tantalum group,the expression of porous tantalum-BMP-2 group and BMP-2 group had no significant difference(P>0.05).with the extension of induction time,the expression of OCN and OPN in porous tantalum group,BMP-2 group,porous tantalum-BMP-2 group was higher than that in the blank control group(P<0.05).The expression of col-I in BMP-2 group and porous tantalumBMP-2 group was higher than that in porous tantalum group and blank control group(P<0.05),while the expression of col-I in porous tantalum group and blank control group was not significantly different from that in BMP-2 group and porous tantalum-BMP-2 group(P>0.05).Conclusion Domestic porous tantalum scaffold combined with BMP-2 can increase the proliferation and differentiation ability of BMSCs to osteoblasts,and promote the high expression of osteoblast factors COL-?,OCN,OPN genes and protein levels during the differentiation process of BMSCs to osteoblasts,thus realizing the rapid mineralization of osteoblasts and the stable connection of bone tissue and scaffold materials.Figure16;Table16;Reference 69...