激光共聚焦图像系统与免疫荧光双标记对脊髓缺血再灌注损伤中ICAM-1表达定量分析

目的 采用激光共聚焦显微镜与免疫荧光双标记进一步探讨实验脊髓缺血再灌注损伤ICAM-1的表达变化规律。方法 采用Zinin法建立脊髓缺血再灌注损伤模型。以荧光探针Fluo23PAM负载后，通过激光扫描共聚焦显微镜（InsightplusIQ Meridian，usA）检测脊髓微血管内皮细胞表面ICAM-lmRNA荧光强度。结果 在激光扫描共聚焦显微镜下，可以清晰地观察ICAM-1微量表达于脊髓微血管内皮细胞表面，而单纯缺血不引起ICAM-1表达。再灌注后损伤区ICAM-1表达先后发生了改变；再灌注4h，ICAM-1mRNA表达量明显升高，再灌注12h其在单位微血管面积上的荧光强度约比单纯缺血组增加了1／2。提示脊髓损伤后微血管内皮细胞初始ICAM-1表达增加是由再灌注损伤激发，其后延迟递增性规律性表达增强，说明ICAM-1参与了脊髓再灌注损伤的炎症病理过程。结论 应用激光扫描共聚焦图像分析与免疫荧光双重标记技术能对黏附分子进行精确的定位定量研究。ICAM-1可做为评判脊髓损伤程度和脊髓I/R后炎症反应程度的监测指标。

Quantitative Analysis of ICAM-1 in the Ischemia/Reperfusion of the Spinal Cord Injury Using ConfocaI Laser Scanning Microscopy And Immunofluorescence Technique

Objective To investigate the expression change and its regularity of ICAM-1 after spinal cord ischemia/reperfusion injury.Methods To set up spinal cord ischemia/reperfusion model. The expression of vascular endotheliocytes ICAM-1mRNA in spinal cord injury was detected through the adoption of LSCM and immunofluorescence technique. Results It can distinct observed that ICAM- 1 have expressed on vascular endothelial cell. The expression of intercellular adhesion molecule was not increased in the normal group and the ischemia group. But the expression of cytokines and adhesion molecules in ischemia area after reperfusion were changed successively. The expression of ICAM -lmRNA was increased after reperfusion for 4 hours, which was about twice as much as that of the comparative groups.It peaked after reperfusion for 12 hours. After reperfusion for twelve hours its fluorescent intensity on microvascular area per unit was increased about one second mere than that of the ischemia group. Condusion Using ConfocaI Laser Scanning Microscopy And immunofluorescence technique can undertake location and its quantitative study of spinal cord injury. ICAM- lcan become a judge target monitoring target of the spinal cord injury.