外伤性视神经损伤后大鼠视网膜中细胞因子的变化和来源
The Changes and Sources of Cytokines in the Retina of Rats after Traumatic Optic Nerve Injury
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摘要: 目的 利用大鼠视神经夹持损伤模型,观察视网膜中小胶质细胞和星形胶质细胞在不同时间点的激活水平,炎性介质和神经营养因子的时空表达变化情况,探讨这2种胶质细胞在大鼠视神经夹持损伤后在视网膜中可能的免疫调控机制.方法 建立SD大鼠视神经夹持模型,分别在建模后1 d、3 d、5d和7 d处死模型大鼠并取材.Western blot检测分析各时间点视网膜IL-1β和TNF-α及iNOS的表达变化情况;免疫荧光组织染色法标记检测各时间点视网膜小胶质细胞和星形胶质细胞的激活情况并检测相应组织中IL-1β、TNF-α、iNOS以及Nestin的表达及分布情况.结果 夹持组与对照组相比Western blot法检测发现IL-1β蛋白在损伤后3 d表达量最高,差异与对照组相比有统计学意义(P<0.05),5 d后表达下降,差异与对照组相比无统计学意义(P>0.05).TNF-α在损伤后各时间点表达均升高(P<0.05),损伤后3d表达量达到峰值,与对照组相比差异有显著统计学意义(P<0.01).iNOS水平在损伤后1d显著上升(P<0.05),且表达量最高,在损伤后3 d出现下降,差异与对照组相比无统计学意义(P>0.05).免疫组织荧光化学染色法显示手术组视网膜中小胶质细胞及星形胶质细胞出现了活化.在小胶质细胞中和星形胶质细胞中检测到了IL-1β,iNOS的表达,但在所有时间点都未能观察到星形胶质细胞中TNF-α的表达.在损伤后7d,在星形细胞中观察到了Nestin表达.结论 视神经夹持损伤后,视网膜内活化的星形胶质细胞和小胶质细胞是IL-1β、iNOS的主要来源.星形胶质细胞在该模型中随着时间的推移可能展现出神经保护作用.Abstract: Objective To explore the immunoregulatory mechanisms of glial cells in retina after optic nerve injury in rats by using rats with traumatic optic nerve injury as TON model to detect the activation of microglia,astrocyte,and observe the temporal and spatial expression of inflammatory mediators and neurotrophic factors in retina at different time points after TON. Methods We established an animal model of rat optic nerve traumatic and draw materials from the rats at 1d, 3d, 5d and 7d after injury.Western-blot was used to dectect the expression levels of IL-1β, TNF-α and iNOS at all time points after injury. Immunofluorescence staining was used to detect microglia, astrocytes and the specific lesions expression and distribution of IL-1β,TNF-α,iNOS,Nestin inthe retina. Results Compared with the control group, the expression of IL-1β protein was peaked at 3 days after injury, and the difference was statistically significant(P <0.05), the expression declined at 5 days, the difference was not statistically significant(P>0.05). The expression of TNF-α increased at all time points after injury(P<0.05), and the expression level of TNF-α reached the peak at 3 days after injury. Compared with the control group, the difference was a statistical significant(P <0.01). The level of iNOS was significantly up-regulated at 1 day after injury, and the expression level of iNOS was significantly higher than the control group,and the difference was statistically significant(P<0.05), the expression declined at 3 days, the difference was not statistically significant(P>0.05). Immunohistochemical staining showed that after the injury,we can observe the activation of microglia and astrocytes. IL-1β and iNOS were expressed in activated microglia and astrocytes.But we were unable to observe the expression of TNF-α in astrocytes at all time points. Nestin expression was observed in astrocytes at 7 days after injury. Conclusions The activated astrocytes and microglia in retina are the main sources of IL-1β and iNOS after traumatic optic nerve injury. The activated astrocytes may have neuroprotective effects in the model.
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Key words:
- Optic nerve injury /
- Retina /
- Microglia /
- Astrocyte /
- Activation /
- IL-1β /
- TNF-α /
- iNOS /
- Nestin
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