不同比例共培养内皮细胞和成骨细胞的成骨及成血管效应*☆

背景：近年来研究显示内皮细胞和成骨细胞共培养修复骨组织的过程中存在相互促进的作用。目的：观察不同比例下内皮细胞和成骨细胞共培养对其成血管和成骨作用的影响。方法：分离培养人脐静脉内皮细胞和成骨细胞MG63，内皮细胞和成骨细胞按照1∶0，8∶1，4∶1，1∶1，1∶4，1∶8，      0∶1在24孔板中进行共培养，内皮细胞和成骨细胞分别单纯单层培养作对照。在第7，14，21天时观察细胞生长状态、血管内皮生长因子表达水平及碱性磷酸酶活性。结果与结论：各组细胞在21 d中生长良好，第14天1∶8组可见成骨细胞开始集落式生长，第21天1∶4组钙化染色结节要明显强于1∶8组，而单纯成骨细胞组并未出现钙化结节。1∶4组碱性磷酸酶含量在3个时间点均明显增高(P < 0.05)且第7天时血管内皮生长因子开始增高，直到第21天时达到高峰。说明内皮细胞和成骨细胞共培养比例为4∶1时，成血管作用较强，而比例为1∶4时成骨作用更强。 

Osteogenic and angiogenic effects of endothelial cells and osteoblasts co-cultured at different ratios

BACKGROUND: Recent studies have shown that co-cultured endothelial cells and osteoblasts are mutually promotive in bone tissues repair.OBJECTIVE: To observe the effects of co-culture of endothelial cells and osteoblasts at different ratios on vasculogenesis and bone formation. METHODS: Human umbilical vein endothelial cells and osteoblasts MG63 were isolated and cultured. Endothelial cells were co-cultured with osteoblasts at ratios of 1:0, 8:1, 4:1, 1:1, 1:4, 1:8, 0:1 in a 24-well plate, and monolayers of endothelial cells and osteoblasts were cultured alone respectively as controls. At 7, 14, 21 days after co-culture, cells growth condition, vascular endothelia growth factor level and alkaline phosphatase activity were observed and detected in each group.RESULTS AND CONCLUSION: Cells grew well in each group in 21 days. At day 14, in the 1:8 group, osteoblasts began to cluster together. At day 21, the 1:4 group showed significantly more calcification nodules formed by Alizarin red staining than the 1:8 group, and no calcification nodules were seen in osteoblasts cultured alone. Alkaline phosphatase activity in the 1:4 group was significantly increased at the three time points (P < 0.05). The level of vascular endothelia growth factor in the 1:4 group began to increase at day 7 and reached its peak at day 21. These findings suggest that angiogenic effect is more effective when endothelial cells are co-cultured with osteoblasts at the ratio of 4:1, and osteogenic effect is more effective at the ratio of 1:4.