The effect of X-ray on microglial Ml and M2 polarization

Abstract

Objective To investigate the effect of X-ray on the polarization of mouse microglia BV-2 cells.

Methods BV-2 cells at the logarithmic growth stage were randomly divided into the Sham irradiation group and 10 Gy irradiation group. The latter group was given a single X-ray irradiation at a dose of 1.28 Gy/min for 7 min 49 s. The activation rate of BV-2 cells was observed and analyzed under a microscope at 1, 3, 6, 24 h and 48 h after irradiation.The changes of cell morphology were observed by HE staining and immunofluorescence staining; The levels of Ml-type activation markers (TNF-α and IL-1β) and M2-type activation marker TGF-β1 in the supernatant of BV-2 cells were detected by ELISA. The levels of polarization-related proteins of Ml-type (CD86 and iNOS) and M2-type (CD206) in BV-2 cells were detected by Western blotting.

Results Morphological results showed that BV-2 cells became larger, and their protrude became coarse and shorter, showing “amoeba” like changes after 10 Gy X-ray irradiation. Compared with the Sham group, the activation rate of BV-2 cells was significantly increased at 3 h, and reached the peak at 6 h, and began to recover at 48 h after irradiation. ELISA results showed an obvious increase in the level of TNF-α and TGF-β1 48 h after irradiation.The level of IL-1β showed a transient decrease at 3~6 h, increased at 24 h, and reached the peak 48 h after irradiation. Western blotting results showed that CD86 protein level did not change significantly at each time points after irradiation, and iNOS protein level increased significantly at 1, 6,24 h and 48 h after irradiation. A fluctuating change in CD206 protein level was found after irradiation.

Conclusion 10 Gy X-ray irradiation can induce the activation of BV-2 cells in vitro, and the polarization type changes with the time after irradiation.

Abstract

摘要： 目的 探讨 X 射线对小鼠小胶质 BV-2 细胞极化的影响。 方法 取对数生长期 BV-2 细胞随机分为假照射组 (Sham) 和 10 Gy 照射组, 后者给予 X 射线单次照射, 剂量为 1.28 Gy/min, 照射时间 7 min 49 s。于照射后 1、3、6、24 h 和 48 h, 光镜下观察并分析 BV-2 细胞活化率; HE 染色及 Iba-1 免疫荧光染色观察 BV-2 细胞形态; ELISA 检测 BV-2 细胞培养液上清中 Ml 型活化标志物 TNF-α、IL-1β 和 M2 型活化标志物 TGF-β1 水平变化; Western blotting 检 测 BV-2 细胞 M1 型 (CD86、iNOS) 和 M2 型 (CD206) 极化相关蛋白水平变化。 结果 形态学结果显示, 10 Gy X 射线 照射后细胞出现胞体变大、突起变粗变短,呈“阿米巴”样改变。与 Sham 组相比, 10 Gy 照射后 3 h BV-2 细胞活化率即 明显升高, 并在照后 6 h 达到高峰, 48 h 开始恢复; ELISA 结果显示, TNF-α 和 TGF-β1 水平在照后 48 h 时均明显升 高; IL-1β 水平在照后 3~6 h 出现一过性降低趋势, 在照后 24 h 开始增高, 至照后 48 h 达到高峰; Western blotting 结果 显示, CD86 蛋白水平在照后各时间点无显著变化, iNOS 蛋白表达水平在照后 1、6、24 h 和 48 h 显著增高; CD206 在 照后 3 h 出现先降低后升高交替波动性变化。 结论 10 Gy X 射线单次照射可诱导离体培养的 BV-2 细胞激活, 其极 化类型随照后时间的不同而变化。