应用激光共聚焦扫描显微镜技术检测缺氧人肺微血管内皮细胞内钙离子浓度动态变化

目的：探讨应用激光共聚焦扫描显微镜（LSCM）技术检测缺氧状态的人肺微血管内皮细胞（HPMVEC）内钙离子（Ca2＋）浓度动态变化的价值。方法：HPMVEC常规培养,按观察时间点不同分为5个缺氧培养组（1h hyp组、2h hyp组、4h hyp组、6h hyp组和8h hyp组）以及1个对照组（0h con组）共6个组,每组设8个复孔,应用LSCM技术测定缺氧后HPMVEC内Ca2＋浓度水平及随时间推移的变化。结果：LSCM技术显示HPMVEC内Ca2＋的荧光强度1h hyp组与0h con组比较、2h hyp组与1h hyp组比较、4h hyp组与2h hyp组比较、6h hyp组与4h hyp组比较、8h hyp组与6h hyp组比较有显著差异（P〈0.05）。线性回归分析结果显示Ca2＋荧光强度与缺氧时间成正相关（r=0.969,P〈0.01）。结论：HPMVEC内Ca2＋浓度随缺氧时间增长而增高;LSCM在动态检测缺氧状态下HPMVEC内的Ca2＋浓度变化中具有明显优势。

Detection of Dynamic Changes of Intracellular Calcium in Hypoxia Human Pulmonary Microvascular Endothelial Cells by Laser Scanning Confocal Microscope

Objective：To investigate the dynamic changes of intracellular calcium concentration（i） by laser scanning confocal microscope（LSCM） in hypoxia human pulmonary microvascular endothelial cells（HPMVEC）.Methods：HPMVEC conventional culture,divided into six groups：one control group（Group 0h con） and five hypoxia groups,respectively hypoxia for 1h,2h,4h,6h and 8h（1h hyp group,2h hyp group,4h hyp group,6h hyp group and 8h hyp group）,each group has 8 wells.The changes of the intracellular calcium concentration（i） were determined after hypoxia by LSCM.Results：The concentration of the intracellular calcium was significantly increased in the 1h hyp group compared to the 0h hyp group（P0.05）,and in the 2h hyp group compared to the 1h hyp group（P0.05）,and in the 4h hyp group compared to the 2h hyp group（P0.05）,and in the 6h hyp group compared to the 4h hyp group（P0.05）,and in the 8h hyp group compared to the 6h hyp group（P0.05）.Linear regression showed：Fluorescence intensity of the intracellular calcium concentration positively correlated with hypoxia（r=0.969,p0.01）.Conclusions：The intracellular calcium concentration of hypoxia HPMVEC were increasing with the time.It is an obvious advantages that LSCM dynamicly detects the change of intracellular calcium concentration of the HPMVEC.