RESUMO
Objective To observe whether the bone marrow stromal cells (BMSCs) which carry BMP-2/Tet-on genes and are microcapsulated by alginate polylysine alginate (APA) and seeded onto calcium phosphate cement (CPC) scaffolds can express bone morphogenetic proteins-2(BMP-2) and secrete collagen proteins.Methods The BMSCs from rats were prepared,transfected with adenoviruses carrying BMP-2/Tet-on genes,and then microcapsulated by APA.Eight porous CPC scaffolds were prepared and mounted onto a 12-hole plate.The experiment was conducted in 3 groups,with 4 parallel specimens in each group.Microcapsulated BMSCs with the target gene were seeded onto the CPC scaffolds in the experimental group; the transfected BMSCs were seeded into the plate holes as positive controls; APA microcapsules without cells were seeded onto the CPC scaffolds as blank controls.The culture medium in each group was taken for ELISA examination on 3,6,9 and 12 days to evaluate the BMP-2 concentrations respectively.The samples in the experimental group were fixed,sliced and dyed to observe secretion of collagen.Results On 3,6,9 and 12 days after culture,the BMP-2 concentrations were 4713.98 ± 178.50 μg/mL,3288.85 ± 194.38 μg/mL,1292.25 ± 300.11 μg/mL and 337.19 ± 84.49 μg/mL respectively in the experimental group; 4663.87 ±242.99 μg/mL,3250.67 ±293.72 μg/mL,1276.74 ± 157.10 μg/mL and 293.65 ±92.48 μg/mL respectively in the positive controls; 105.14 ± 10.93 μg/mL,91.42 ± 18.00 μg/mL,89.63 ± 12.99 μg/mLand 108.72 ± 23.90 μg/mL respectively in the blank controls.There were significant differences between the 3 groups at each same time point (P ＜ 0.05).The blank controls were significantly different from both the experimental group and the positive controls (P ＜ 0.05),but there were no significant differences between the experimental group and the positive controls (P ＞ 0.05).Collagen fibrils were observed by optic microscopy in the sections of the experimental group.Conclusion As microcapsulated BMSCs can survive in CPC scaffolds and express a significant amount of BMP-2 and secrete collagen proteins as expected,they can be used for further experiments in vivo.