中文摘要：

目的：探讨白细胞介素1β（IL-1β）刺激不同时间对大鼠脑微血管内皮细胞（rCMEC）的环氧合酶（COX）活性及其mRNA表达和PGE2释放的影响。方法：建立rCMEC培养，进行Ⅷ因子相关抗原鉴定。细胞长至融合状态后加入IL-1β（30μg／L），分别刺激0．5、1、2、4、8、12、24h，ELISA测定细胞内COX-1、COX-2的活性及细胞外液PGE2含量，荧光实时定量PCR检测COX-1、COX-2的mRNA表达量，扩增产物进行熔解曲线图及琼脂糖电泳分析，并与未刺激的rCMEC作比较。结果：①Ⅷ因子抗体免疫组化染色可见90％以上的培养细胞呈阳性，确认为rCMEC。②IL-1β激4h时细胞培养液PGE2含量已明显高于未刺激组（P〈0．05）；12h时PGE2含量达到最大值（P〈0．01）；24h时PGE2含量有所回降，但与未刺激组比较仍有显著差异（P〈0．05）。③IL-1β刺激不同时间rCMEC内COX-1活性与未刺激组相比无统计学差异（P〉0．05）；COX-2活性在第8h时已明显高于未刺激组（P〈0．05），12h活性达峰值（P〈0．01），24h活性有所回降，但仍具显著差异（P〈0．05）。④IL-1β刺激不同时间COX-1mRNA表达与未刺激组比较无明显差异（P〉0．05）；未刺激组在本实验条件下未检测到COX-2mRNA表达，IL-1β激1h时可见COX-2mRNA表达，4h时COX-2mRNA表达至峰值，而后开始回降，第12h时已未见表达。熔解曲线图显示无非特异性扩增；琼脂糖电泳可见扩增基因与目的基因长度相符，结果与荧光定量PCR一致。结论：IL-1β用下，rCMEC释放的PGE2持续增加并于12h达峰值，这一过程主要与COX-2mRNA表达激活及COX-2活性增加有关。

英文摘要：

AIM： To study the cyclooxygenase （COX） activity and its mRNA expression, and PGE2 release from rats cerebral microvascular endothelial cells （rCEMC） stimulated by IL - 1β （30 μg/L） at different times. METHODS： rCMEC were cultured, and identified by immunohistochemistry for yon Willebrand factor （Ⅷ factor, a marker for all endothelial cells） in cytoplasm of the cells. After rCEMC grew to confluency, they were stimulated with IL - 1β for 0. 5, 1, 2, 4, 8, 12 and 24 h, respectively. Activity of COX - 1 and COX -2 in rCEMC and production of PGE2 in the conditioned media were detected by ELISA. COX - 1 and COX - 2 mRNA expressions were measured by real - time quantity PCR. The amplification product was tested by melting curve and identified by electrophoretic gel. RESULTS： ① Positive immunostaining for Ⅷ factor was present diffusely in the cytoplasm in more than 90% rCMEC. ② Compared to the cells without IL - 1β stimulation, the production of PGE2 increased significantly （P 〈 0. 05） at 4 h after rCEMC were incubated with IL - 1β and reached the top level at 12 h （P 〈0. 01 ）, then declined thereafter at 24 h （P 〈0. 05）. ③ There was no significant difference on COX - 1 activity between IL - 1β group and non - IL - 1β group. COX - 2 activity increased significantly compared with those in non - IL - 1β （ P 〈 0. 05 ） at 8 h after rCEMC were incubated with IL - 1β and reached the top level at 12 h （P〈0. 01） , then declined thereafter at 24 h （P〈0. 05）. ④ There was no significant difference on COX - 1 mRNA expression between IL - 1β group and non - IL - 1β group. COX - 2 mRNA was induced and became detectable at 1 h, and reached the top level at 4 h, then declined thereafter at 8 h and became undetectable by 12 h and 24 h after incubation with IL - 1β. The melting curve showed there was no nonspecific amplification and electrophoretic gel showed the lengths of amplification products accorded with the predicted lengths. CONCLUSION： While rCEMC