局部辐照大鼠血清对成骨细胞增殖分化的影响

Effects of serum of local radiated rat on the proliferation and differentiation of osteoblast

SD大鼠胫骨部位30 Gy照射2周和12周后处死,取血离心得血清;酶消化法分离培养新生SD大鼠头盖骨成骨细胞。采用MTT和PNPP法观察不同血清对成骨细胞增殖和分化的影响;用流式细胞仪观察成骨细胞周期变化;用Realtime PCR方法观察成骨细胞CyclinD、CyclinE、p21和p53的表达。结果显示:辐照2周和12周的大鼠血清抑制成骨细胞增殖,对成骨细胞ALP的表达无明显影响;辐照2周和12周的大鼠血清增加G1期成骨细胞比例,减少S期和G2期细胞比例;辐照2周大鼠血清抑制成骨细胞CyclinD和CyclinE的表达;12周大鼠血清促进成骨细胞p21和p53的表达,抑制成骨细胞CyclinE的表达。局部辐照大鼠血清通过抑制成骨细胞CyclinD和CyclinE表达,上调p21和p53的表达持续阻滞细胞于G1期抑制成骨细胞增殖。

The tibia of rat was irradiated at the dose of 30 Gy, and then the rats were killed at 2 and 12 weeks after radiation to collect the serum; the osteoblast was cultured from cranium of fetal rat with enzymatic digestion method, and then the proliferation and differentiation of osteoblasts were detected by MTT and PNPP after 72 h, and the cell cycle of osteoblasts were detected by flow cytometry, meanwhile, the mRNA expression of cyclinD, cyclinE, p21 and p53 of the osteoblast were detected after 24 h. The results showed that the serum of the radiated rats inhibits the proliferation of osteoblast and increases the number of osteoblast in G1 phase, while decreases the number of osteoblast in S phase and G2 phase. Serum from the radiated rats 2 weeks later down-regulates the mRNA expression of CyclinD and CyclinE, meanwhile the serum from the radiated rats 12 weeks later up-regulates the mRNA expression of p21 and p53 and down-regulate the mRNA expression of CyclinE. The results suggest that the serum of the radiated rat inhibits the proliferation of osteoblast by down-regulating the expression of CyclinD and CyclinE and up-regulating the expression of p21 and p53 which retards osteoblasts in G1 phase.