目的 探讨负载siRNA胶原/生物活性玻璃复合材料的成骨作用.方法 冷冻干燥法分别制备胶原/生物玻璃、负载阴性对照siRNA胶原/生物玻璃和负载noggin胶原/生物玻璃3组复合材料.用CCK8试验检测空白组上述3组不同支架材料浸提液对细胞增殖的影响,ALP活性检测、q-PCR检测、茜素红染色评估3组不同支架对MC3T3细胞矿化的影响.结果 材料浸提液共培养MC3T3细胞3、5 d后,胶原/生物玻璃复合材料、负载阴性对照siRNA胶原/生物玻璃复合材料和负载noggin胶原/生物玻璃支架组细胞增殖数目均明显高于空白组,差异具有统计学意义(P<0.05).培养14 d后,负载siRNA胶原/生物玻璃支架组ALP活性高于单纯支架组,差异具有统计学意义(P<0.05).培养14 d后,负载siRNA胶原/生物玻璃支架组ALP、Runx2、BSP表达量高于单纯支架组,差异具有统计学意义(P<0.05).茜素红染色结果表明负载siRNA胶原/生物玻璃支架组较其余组有更多矿化结节形成,差异具有统计学意义(P<0.05).结论 负载siRNA胶原/生物玻璃复合材料支架具有良好的生物相容性,胶原/生物玻璃复合材料和siRNA noggin可协同增效促进成骨.%Objective To investigate osteogenic effect of collagen/bioglass composites loaded with a small interfering RNA (siRNA) targeting noggin. Methods The collagen/bioglass composites loaded with the negative control siRNA or noggin siRNA were prepared by freeze-drying method. CCK8 test was used to evaluate the proliferation of MC3T3 cells exposed to the aqueous extracts of collagen/bioglass composites and the siRNA-loaded collagen/bioglass composites. ALP activity assay, quantitative real-time PCR and Alizarin Red staining were used to assess the effect of the 3 composites on mineralization in MC3T3 cells. Results MC3T3 cells cultured for 3 and 5 days in the presence of the extracts of the 3 composites all showed significantly more active proliferation than the blank control cells (P<0.05). Compared with the cells seeded on the scaffold without siRNA, MC3T3 cells seeded on collagen/bioglass scaffold loaded with noggin siRNA showed a significantly higher ALP activity at 14 days after seeding (P<0.05) with significantly increased expression of ALP, Runx2 and BSP mRNAs (P<0.05). Alizarin Red staining showed that the cells seeded on the noggin siRNA-loading collagen/bioglass scaffold contained significantly more mineralized nodules than the other cells (P<0.05). Conclusion The collagen/bioglass composites loaded with noggin siRNA have a good biocompatibility, and the collagen/bioglass composites and noggin siRNA show a synergistic effect in promoting osteogenesis.
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