三维重建技术观察1-磷酸鞘氨醇刺激其受体2在内皮细胞的表达

目的应用激光扫描共聚焦显微镜三维重建技术观察1-磷酸鞘氨醇(S1P)刺激下S1P受体2(S1PR2)的表达变化,并探讨共聚焦三维重建技术与普通单层扫描技术在生物组织功能和形态学研究中的应用价值。方法 S1P(10μmol/L)刺激脐静脉内皮细胞(HUVECs)6 h,Western blot检测S1PR2的表达变化;对细胞进行免疫荧光染色后,采用激光扫描共聚焦显微镜分别进行多层层切扫描和单层扫描,层切扫描之后对图像进行三维重建,比较两种技术的优劣。结果 Western blot结果显示S1P刺激6 h后S1PR2表达显著性增高(P0.05)。荧光图像显示,单层扫描时S1P 6 h组荧光强度与对照组比较差异无统计学意义;而层切之后三维重建图像中,S1P刺激后S1PR2荧光强度显著高于正常对照组和单层扫描所得图像,差异均有统计学意义(P0.05)。结论激光扫描共聚焦显微镜三维重建技术,可以获得三维重构图像,并且具有三维立体及图像信息量更全的特点,更能客观的表现出整个细胞特定分子荧光强度的变化。

Application of laser scanning confocal microscope three-dimensional reconstruction to monitor the changes of sphingosine-l-phosphate receptor 2

Objective This research was designed to observe sphingosine-l-phosphate （S1P）-induced change of S1P receptor 2 （S1PR2） expression and compare the application value of laser scanning confocal three-dimensional reconstruction and ordinary single scanning technology in a morphological study. Methods Human umbilical vein endothelial cells （HUVECs） were stimulated by SIP （10 μmol/L） for 6 h. The expression of S1PR2 was investigated by Western blot. Then S1PR2 was observed by laser scanning confocal three-dimensional reconstruction and ordinary single scanning technology. Results Western blot result revealed that 10 μmol/L S1P significantly increased the expression of S1P2 （P〈0.05）. And there was no significant difference between S1P and control group when S1PR2 was observed by ordinary single scanning technology. But laser scanning confocal three-dimensional reconstruction technology can display the difference obviously. Conclusion Laser scanning confocal three-dimensional reconstruction technology reveals the changes of fluorescence intensity more objectively than ordinary single scanning technology.