电离辐射诱导神经干细胞衰老的模型研究

Establishment of ionizing radiation-induced neural stem cell aging model

关键词:  神经干细胞  电离辐射  衰老  模型  小鼠

Key words:  neural stem cell  ionizing radiation  aging  model  mice

摘要:

目的 研究神经干细胞在不同强度电离辐射(ionizing radiation, IR)处理下的衰老表型变化,建立电离辐射诱导神经干细胞衰老的方法。方法 对成年鼠来源的神经干细胞进行不同剂量的连续多次电离辐射,细胞分为照射组(0Gy组),低剂量连续辐照组(0.5Gy组),中等剂量的隔代辐照组(2Gy组),高剂量的每隔3代辐照组(8Gy组),分别在短期处理和长期处理后,收集细胞样本,提取RNA,检测神经干细胞衰老相关基因(p16、Bmi1)的表达,并用Ki67免疫荧光染色检测细胞的增殖,用SA-β-Gal检测细胞的衰老状态。结果 在长期处理后,p16在0.5Gy、8Gy组细胞中有20倍的表达升高,在2Gy组细胞中有2倍的表达升高；Bmi1在3组细胞中表达都降低,并随着剂量增加降低幅度增加；Ki67免疫荧光染色结果显示3个处理组的细胞增殖速率都降低,但在长期处理后2Gy组细胞增殖加快；SA-β-Gal检测结果显示,0.5Gy组细胞衰老程度不变,8Gy组衰老加重,2Gy组衰老程度降低。结论对神经干细胞长期高剂量的每隔3代辐照处理可以诱导神经干细胞衰老,作为研究干细胞衰老的模型。

Abstract:

Objective To study the aging phenotype of neural stem cells under different dosage of ionizing radiation(IR)and to establish a cellular aging model. Methods The neural stem cells derived from adult mouse were divided into four groups: control(0 group,untreated),low dosage(0.5Gy group,IR=0.5Gy at every passage),medium dosage(2Gy group,IR=2Gy at every other passage)and high dosage(8Gy group,IR=8Gy at every fourth passage). After short and long term passage,the cell samples were collected. The expression of aging related genes was detected with qRT-PCR,cell proliferation was determined with Ki67 immunostaining and cell senescense was examined with SA-β-Gal staining. Results After long term treatment,compared with 0 group the expression level of p16 mRNA was 20-folder increased in 0.5Gy and 8Gy group while only 2-folder rose in 2Gy group. Bmi1 was down regulated in all 3 groups and the extent of decrease was positively related to increased dosage. The Ki67 immunostaining results showed decreased cell proliferation rate in all 3 groups but more dividing cells in 2Gy group. SA-β-Gal staining reveled more severe senescense state in 8Gy group and unchanged state in 0.5Gy group while slightly improved condition of 2Gy group compared to 0 group. Conclusion Ionizing radiation at dosage of 8Gy at every fourth passage can induce cell aging of neural stem cells from young adult mouse, which may be used as a cellular aging model for further research.