肝素对脂多糖诱导内皮细胞通透性增高的保护作用  

Protective effects of heparin against increased permeability of endothelial cells induced by lipopolysaccharide

作　　者：毛怡然[1] 肇冬梅[1] 马晓春[1] 

机构地区：[1]中国医科大学附属第一医院重症医学科,辽宁沈阳110001

出　　处：《中国危重病急救医学》2012年第5期278-282,共5页Chinese Critical Care Medicine

基　　金：辽宁省沈阳市科技项目（FIO-205-1-01）

摘　　要：目的研究肝素对脂多糖（LPS）诱导人脐静脉内皮细胞（HUVECs）单层通透性和细胞骨架形态的影响。方法HUVECs细胞株传代培养后随机分为空白对照组、LPS组、肝素组、LPS＋肝素组，n=8。用四甲基偶氮唑盐（MTT）比色法测定内皮细胞活性；用Transwell小室法测定单层内皮细胞通透性；用免疫荧光染色法测定内皮细胞纤维状肌动蛋白fF-actin）形态。结果与空白对照组比较，LPS10zg／L及100mg／L对细胞活性起到明显的抑制作用（0．695±0．015、0．476±0．030比0．860±0．053，P〈O．05和P〈O．01），肝素100kU／L对内皮细胞活性产生抑制作用（0．675±0．030比0．840±0．023，P〈0．05）。LPS10mg／L可增加单层内皮细胞通透性，于4h时达峰值，与空白对照组比较差异有统计学意义（5．882±0．101比4．489±O．015，P〈0．05），并能增加细胞骨架应力纤维的形成；而LPS＋肝素刺激2-12h时单层内皮细胞的通透性较LPS组明显降低（2h：4．382±0．053比5．084±0．129，4h：4．528±0．044比5．882±0．101，6h：4．38l±0．089比5．479±0．125，12h：4．447±0．054比4．719±0．080，均P〈O．05），并能减少内皮细胞骨架的重排及应力纤维的形成。结论肝素可以减轻LIPS诱导单层内皮细胞通透性增高及细胞骨架重排，对内皮细胞屏障功能起到保护作用。Objective To evaluate the effects of heparin on the changes in permeability and cytoskeleton of cultured human umbilical vascular endothelial cells （HUVECs） induced by fipopolysaecharide （LPS）. Methods HUVECs were cultured in vitro and randomly assigned to blank control group, LPS group, heparin group, and LPS ＋ heparin group, n =8. Cell viability was determined by methyl thiazolyl tetrazolium （MTF） method. Endothelial permeability was measured with the Transwell chamber models. F-actin of cytoskeletons was assayed with iluorescein isothiocyanate （FITC）-phalloidine. Results Compared with the blank control group, 10 mg/L and 100 mg/L of LPS significantly inhibited cell viability （0.695±0.015, 0.476±0.030 vs. 0.860±0.053, P〈0.05 and P〈O.01 ）. Heparin 100 kU/L also inhibited cell viability （0.675±0.030 vs. 0.840±0.023, P〈0.05）. Increase in permeability of cndothelium was induced by 10 mg/L of LPS, and it peaked at 4 hours, there was significant difference compared with blank control group （5.882±0.101 vs. 4.489±0.015, P〈0.05）. At the same time, LPS led to the reorganization of F-actin eytoskeleton and the formation of stress fibers. LPS ＋ heparin decreased the increase in permeability of endothelium at 2-12hours （2hours： 4.382±0.053 vs. 5.084±0.129, 4hours： 4.528±0.044vs. 5.882±0.101, 6 hours： 4.381±0.089 vs. 5.479±0.125, 12 hours： 4.447±0.054 vs. 4.719±0.080, all P〈O.05）, and attenuated the reorganization of F-actin and the formation of stress fibers. Conclusion Heparin attenuates LPS-induced increase in Dermeabilitv of HUVECs and alterations in F-actin organization, thus orotects endothelial barrier.

关 键 词：脓毒症 肝素 内皮细胞通透性 细胞骨架 内皮细胞纤维状肌动蛋白 

分 类 号：R459.7[医药卫生—急诊医学]