大鼠液压冲击脑损伤半胱氨酰天冬氨酸特异性蛋白酶-12动态表达变化和意义  

Time-course expression and role of cysteinyl aspartate-specific protease-12 following fluid percussion injury in rats

作　　者：李秀山[1] 孙博宇[2] 孙国柱[3] 张记民 王惊涛 Li Xiushan;Sun Boyu;Sun Guozhu;Zhang Jimin;Wang Jingtao(Department of Neurosurgery,Affiliated Hospital of Hebei University of Engineering,Handan 056002,China;Basic Medical College of Hebei Medical University,Shijiazhuang 050017,China;Department of Neurosurgery,the 2nd Hospital of Hebei Medical University,Shijiazhuang 050000,China)

机构地区：[1]河北工程大学附属医院神经外科,邯郸056002 [2]河北医科大学基础医学院,石家庄050017 [3]河北医科大学第二医院神经外科,石家庄050000

出　　处：《中华实验外科杂志》2019年第10期1806-1809,共4页Chinese Journal of Experimental Surgery

基　　金：河北省自然科学基金(H2014206383);河北省引进留学人员资助项目(CY201710);河北省医学科学重点课题(20170088,20190127).

摘　　要：目的探讨大鼠颅脑液压冲击伤脑损伤半胱氨酰天冬氨酸特异性蛋白酶(Caspase)-12蛋白及其mRNA的表达变化及其意义.方法成年雄性SD大鼠48只应用Dixon法制作颅脑损伤模型,造模后6、12、24h、3d用免疫组织化学方法、蛋白质印迹法(Western blot)以及实时定量聚合酶链反应(Real-time PCR)分别测定Caspase-12蛋白及其mRNA的表达,应用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记测定法(TUNEL)测定神经细胞凋亡变化.应用SPSS 22.0统计软件分析,计量资料以均值±标准差(Mean±SD)表示,组间比较差异性分析采用Student's t检验.结果与假手术组(Sham组)比较,模型组(TBI组)自伤后6h,损伤灶周脑组织Caspase-12蛋白开始增加[积分吸光度(IA)0.165±0.021],24 h达高峰(0.489±0.034),3d开始下降(0.258±0.027),差异有统计学意义(￡=2.293、4.594、6.488、3.605,P<0.05).Western blot进一步印证了Caspase-12蛋白变化趋势.Caspase-12 mRNA于伤后6h开始增加(0.626±0.054),12 h达高峰(0.867±0.048),持续至3d后下降(0.764±0.077),差异有统计学意义(t=2.326、4.185、3.607、2.215,P<0.05);TUNEL发现神经细胞凋亡指数(AI)自6h开始增高(22.372 ±3.304),24h达到峰值(55.222 ±4.123),3d时出现下降(28.226±3.273),差异有统计学意义(t=3.093、6.224、6.984、4.133,P<0.05).结论内质网应激的特异性关键蛋白Caspase-12表达上调在液压冲击脑损伤的神经细胞凋亡中起重要作用.Objective To investigate the expression of cysteinyl aspartate-specific protease (Caspase)-12 protein and its mRNA following fluid percussion injury in rats. Methods The experimental models were established in 48 rats according to Dixon’s method. Neuronal apoptosis, the expression of Caspase-12 protein and its mRNA were measured with terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL), immunohistochemistry, Western blotting and real-time quantitative polymerase chain reaction (Real-time PCR) at 6, 12, 24 h, 3 d after operation respectively. Results Compared with animals of Sham group, the Caspase-12 immuno-reactive expression significantly increased in TBI group from 6 h [integral optical density (IA) 0.165±0.021], reached its peak at 24 h (IA 0.489±0.034), and then began to drop at 3th day (IA 0.258±0.027)(t=2.293, 4.594, 6.488, 3.605, P<0.05). Western blotting further confirmed the trend of Caspase-12 protein expression. Furthermore, compared with Sham group, Caspase-12 mRNA significantly increased from 6 h (0.626±0.054), reached its maximum at 12 h (0.867±0.048), and began to drop at 3th day (0.764±0.077)(t=2.326, 4.185, 3.607, 2.215, P<0.05). TUNEL showed neuronal apoptotic index (AI) significantly increased from 6 h (22.372±3.304), reached its maximum at 24 h (55.222±4.123), and began to drop at 3th day (28.226±3.273)(t=3.093, 6.224, 6.984, 4.133, P<0.05). Conclusion Caspase-12 expression upregulation plays an important role in neuronal apoptosis following fluid percussion injury in rats.

关 键 词：颅脑液压冲击伤 半胱氨酰天冬氨酸特异性蛋白酶-12 神经细胞凋亡 大鼠 

分 类 号：R651.15[医药卫生—外科学]