硫氧还蛋白TXN对视神经轴突损伤的保护作用

发布时间：2019-05-16 15:19

【摘要】：目的研究硫氧还蛋白(thioredoxin,TXN)在视神经轴突保护中的作用,为视神经损伤提供有效的治疗方案。方法制作大鼠视神经夹伤模型,以正常大鼠作为正常对照组。按夹伤后处死时间不同,选取伤后1周、2周、4周三个时间点,提取实验组和正常对照组中大鼠视网膜的mRNA和蛋白,通过RT-PCR和Western blot实验方法检测TXN表达情况;用曲古抑菌素A(TSA)刺激RGC-5细胞,观察在TSA刺激后,RGC-5细胞的轴突形成情况,并检测TXN表达量的变化。构建TXN的真核表达质粒,转染进RGC-5细胞,在TNF-α刺激条件下,观察RGC-5细胞的轴突形成情况以及轴突长度的改变。结果与正常大鼠比较,视神经夹伤模型大鼠中TXN的mRNA以及蛋白水平明显下降。在伤后1周,TXN mRNA表达量仅下降了32%,而在伤后2周、4周则分别下降了55%以及70%,说明TXN表达水平的降低是随着夹伤后时间的延长而增加的。用TSA诱导RGC-5细胞24 h,观察到RGC-5细胞形态发生明显改变,约65%的细胞分化产生轴突,而对照组(DMSO处理)中细胞没有形成轴突,同时Western blot结果显示,相对于DMSO处理,TSA刺激后TXN的表达明显升高。在TNF-α刺激下,RGC-5细胞的轴突形成比率从(65±5)%减少至(30±4)%,轴突长度从(100±6)μm下降至(74±6)μm。而在过表达TXN情况下,TNF-α刺激RGC-5细胞,轴突形成比率为(52±7)%,轴突的长度为(92±8)μm,较未刺激细胞仅有较少的降低。结论在轴突发生损伤后,TXN的表达量显著下降。而在轴突形成过程中,TXN的表达量明显上升并且过表达TXN能很好地减轻外界刺激诱导的轴突损伤。TXN可能参与了轴突形成并且能有效地防止轴突的损伤。
[Abstract]:Objective to study the role of thioredoxin (thioredoxin,TXN) in the protection of optic nerve axons and to provide an effective treatment for optic nerve injury. Methods the rat model of optic nerve clip injury was established, and the normal rats were used as the normal control group. According to the time of death after clamping injury, mRNA and protein were extracted at three time points 1 week, 2 weeks and 4 weeks after injury, and the expression of TXN was detected by RT-PCR and Western blot assay. RGC-5 cells were stimulated by trigutin A (TSA). The axonal formation of RGC-5 cells was observed after TSA stimulation, and the expression of TXN was detected. The eukaryotic expression plasmid of TXN was constructed and transformed into RGC-5 cells. The axonal formation and axonal length of RGC-5 cells were observed under the condition of TNF- 伪 stimulation. Results compared with normal rats, the mRNA and protein levels of TXN in optic nerve clip injury model rats were significantly decreased. The expression of, TXN mRNA decreased by only 32% at 1 week after injury, 55% and 70% at 2 weeks and 4 weeks after injury, respectively, indicating that the decrease of TXN expression increased with the prolongation of clamping time. RGC-5 cells were induced by TSA for 24 h. The morphology of RGC-5 cells was significantly changed, about 65% of the cells differentiated to produce axons, while the cells in the control group (DMSO treatment) did not form axons. At the same time, Western blot results showed that compared with DMSO treatment, The expression of TXN increased significantly after TSA stimulation. Under the stimulation of TNF- 伪, the axonal formation ratio of RGC-5 cells decreased from (65 卤5)% to (30 卤4)%, and the axon length decreased from (100 卤6) 渭 m to (74 卤6) 渭 m. Under the condition of overexpression of TXN, the axonal formation rate and axon length of RGC-5 cells stimulated by TNF- 伪 were (52 卤7)% and (92 卤8) 渭 m, respectively, which were only lower than those of unstimulated cells. Conclusion the expression of TXN decreased significantly after axonal injury. In the process of axonal formation, the expression of TXN increased obviously and overexpression of TXN could alleviate the axonal injury induced by external stimulation. TXN may be involved in axonal formation and can effectively prevent axonal injury.
【作者单位】：武汉市中心医院眼科;
【分类号】：R774.6

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