X线辐照对小鼠成骨前体细胞MC3T3-E1增殖及RANTES表达的影响

目的探讨X线辐照对体外培养小鼠成骨前体细胞MC3T3-E1增殖及其分泌蛋白的影响,为放射性骨损伤的防治提供研究基础。方法取对数生长期的小鼠成骨前体细胞MC3T3-E1,按辐照剂量分成0 Gy对照组、2 Gy组、4 Gy组、8 Gy组,使用6 MV直线加速器分别予以相应剂量的X线辐照,观察辐照后细胞形态的改变,CCK-8法检测细胞增殖情况,检测辐照后7 d细胞内碱性磷酸酶(alkaline phosphatase,ALP)活性;收集各组细胞上清,采用高通量蛋白质芯片技术检测分泌蛋白的表达情况,筛选出放射性损伤特异性的差异蛋白,并通过酶联免疫吸附实验进行验证。结果辐照后MC3T3-E1细胞胞体肿大,细胞核变大。与对照组相比,从3 d开始4 Gy和8 Gy组细胞增殖速度降低,差异有统计学意义(P<0.05),并且增殖降低呈现辐照剂量依赖性。辐照后培养7 d,各辐照组细胞内ALP活性均比对照组低,差异均有统计学意义(P<0.05)。与对照组相比,辐照后细胞上清共发现36个差异表达蛋白,其中受激活调节正常T细胞表达和分泌因子(regulated upon activation,normal T cell expressed and secreted factor,RANTES)含量明显增加,呈剂量依赖性。结论一定剂量的辐照可抑制小鼠成骨前体细胞MC3T3-E1的增殖;RANTES可能是细胞受放射线辐照损伤后引发的重要信号因子。

Effects of X-ray irradiation on proliferation and RANTES expression of the mouse osteogenic precursor cell line MC3T3-E1

Objective To investigate the effects of X-ray irradiation on the proliferation and protein secretion in vitro in a cultured mouse osteoblast precursor cell line(MC3T3-E1) and to provide a basis for the prevention and treatment of radiation-induced bone injury. Methods Mouse osteoblast precursor cells(MC3T3-E1) in the logarithmic growth phase were divided into a 0 Gy control group, 2 Gy group, 4 Gy group and 8 Gy group according to the irradiation dose. The corresponding dose of X-rays was applied, and the changes in cell morphology after irradiation were observed. Cell proliferation was detected by the CCK-8 method, and alkaline phosphorus was detected at 7 days after irradiation. The alkaline phosphatase(ALP) activity and expression of secreted proteins were detected by high-throughput protein chip technology. The differential expression of proteins due to radiation damage was screened and verified by an ELISA. Results After irradiation, the soma and nucleus of MC3T3-E1 cells were enlarged. Compared to the control group, the cell proliferation rates of the 4 Gy and 8 Gy groups were significantly decreased starting at 3 days in culture(P < 0.05), and the cell proliferation decreased in a dose-dependent manner. After 7 days of culture, the ALP activity in the cells of each irradiated group was lower than that of the control group, and the difference was statistically significant(P < 0.05). Compared to the control group, 36 differentially expressed proteins were found in the supernatant after irradiation. Among them, the content of regulated upon activation, normal T cell expression and secreted factor(RANTES)was significantly increased in a dose-dependent manner. Conclusion A certain dose of irradiation can inhibit the proliferation of mouse osteoblast precursor cells(MC3T3-E1), and RANTES may be an important signaling factor induced by radiation damage.