大鼠脊髓损伤模型的改良及伤后再生基因-2蛋白的表达变化

Modification of the rats model of spinal cord injury and expression of regenerating gene-2 protein after spinal cord injury in rats

目的:制备改良的大鼠脊髓损伤(SCI)动物模型,并探讨再生基因(Reg)-2蛋白在SCI后的表达规律。方法:采用36只SD大鼠。参考Allen法,使用自制打击装置致大鼠T13段脊髓中度损伤。以行为联合评分法(CBS)评定模型的可靠性。免疫印迹法和免疫组织化学(SABC)法对正常对照组、伤后第1天、第2天、第3天、第5天和第7天的大鼠脊髓组织中的Reg-2蛋白进行检测。结果:SCI后大鼠一般情况符合临床损伤特点,且稳定性强;各损伤组大鼠神经功能联合评分均呈明显下降趋势,与正常对照组比较差异具有显著性意义(P<0.05);在正常对照组大鼠脊髓神经元内有微量的Reg-2蛋白表达(阳性细胞数为17.3±2.6,Reg-2相对表达量为0.038±0.007)。SCI后1天,大鼠脊髓内Reg-2表达的免疫阳性细胞随着损伤时间的推移逐渐增多,至伤后第7天仍呈高水平表达(阳性细胞数为90.0±3.6,相对表达量为0.694±0.018),各组间比较差异具有显著性意义(P<0.05)。伤后3天内,Reg-2免疫阳性细胞以后角神经元为主,而伤后7天以前角神经元和胶质细胞为主。结论:本实验装置制作的大鼠SCI模型稳定、可靠;SCI后Reg-2蛋白表达开始升高,对受损神经起保护和修复作用。

Objective： To design and produce a kind of modified rat model of spinal cord injury（SCI）, and explore the ex- pression pattern and effects of regenerating gene （Reg） -2 protein after SCI in rats. Method： Thirty-six SD rats were subjected to moderate SCI by modified Allen＇s crush method with a self-de- signed experimental device at T13 level of spinal cord. Combined behavioral score（CBS） was used to assess the reliability of model. Western-blot and immunohistochemical techniques （SABC） were used to detect the expression of Reg-2 in rats of different groups （normal control, 1, 2, 3, 5 and 7d post-injury）. Result： After operation the general health status of rat model was accorded with the clinical characteristics of SCI in human, and was with high-stability. The CBS in model groups declined with the time of injury, and com- pared with the normal control the difference was statistically significant （P〈 0.05）. In normal control group, the number of Reg-2-positive cells and relative expression of Reg-2 in spinal cord were 17.3 ± 2.6 and 0.038 ± 0.007. At ld post-injury the level of Reg-2 elevated and reached a high level at 7d post-injury （the number of Reg-2-positive cells and relative expression of Reg-2 were 90.0 ± 3.6 and 0.694 ± 0.018）, the difference compared with model groups were statistically significant （P 〈 0.05）. In the early phase of injury （in 3d post-injury）, neuronsin dorsal horn of spinal cord were accounted for the most part of Reg-2-positive cells. But the main Reg-2-posi- tive cells were neurons of spinal anterior horn and gliocytes in the later phase of injury. Conclusion： The modified rat model prepared with our self-designed experimental device was with high-stability and higher reliability, and the experimental results suggested that SCI could stimulate the Reg-2 expression, which might contribute to maintenance of nerve cell survival and repair of damaged neural tissues after SCI.