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目的探讨钙反应性反式激活因子(CREST)在肌萎缩侧索硬化症(ALS)转基因鼠脊髓及脑干组织中的表达变化及相关性作用。方法分别取出生后95 d、108 d、122 d(早、中、晚期)野生型与SOD1-G93A转基因鼠,通过RT-PCR、蛋白免疫印迹技术检测脊髓组织中CREST mRNA及蛋白水平表达变化,通过免疫荧光双标记技术检测脊髓及脑干组织中CREST的表达变化及与神经元及星形胶质细胞的共表达情况。结果 (1)与野生型鼠相比较,SOD1-G93A转基因鼠脊髓组织中CREST蛋白水平下调(P<0.05),在mRNA水平变化并不明显。(2)免疫荧光双标检测显示,CREST主要表达于细胞核,与神经元有共表达,与星形胶质细胞不存在共表达;与野生型鼠比较,SOD1-G93A转基因鼠脊髓和脑干组织中CREST表达下调(P<0.05)。结论 CREST的差异表达可能与ALS运动神经元的功能障碍有关。
Objective To investigate the expression and correlative role of calcium reactive transactivator (CREST) in the spinal cord and brainstem tissue of amyotrophic lateral sclerosis (ALS) transgenic mice. Methods The wild-type and SOD1-G93A transgenic mice at 95 d, 108 d and 122 d after birth were harvested. The expression of CREST mRNA and protein in spinal cord tissues was detected by RT-PCR and Western blot, respectively. Immunofluorescence double labeling technique was used to detect the expression of CREST in spinal cord and brainstem tissue and its co-expression with neurons and astrocytes. Results (1) Compared with the wild type mice, the level of CREST protein in the spinal cord of SOD1-G93A transgenic mice was down-regulated (P <0.05), but not obvious in mRNA level. (2) Immunofluorescence double-labeled assay showed that CREST was mainly expressed in the nucleus, co-expressed with neurons and co-expressed with astrocytes. Compared with wild-type mice, the expression of CREST in spinal cord and brainstem tissue of SOD1-G93A transgenic mice CREST expression was down-regulated (P <0.05). Conclusion The differential expression of CREST may be related to the dysfunction of ALS motor neurons.