摘要
目的探讨乌司他丁（UTI）对肿瘤坏死因子-α（TNF-α）诱导的血管内皮细胞高通透性的影响。方法建立人脐静脉内皮细胞系EA．hy926细胞TNF-α诱导炎症模型，将其分为正常组、TNF-α组、uTI组及TNF-α＋不同浓度的UTI组（T＋U组），分别采用四甲基偶氮唑盐（MTT）比色法检测EA．hy926细胞活性，EVOM法测单层细胞电阻。RT-PCR法、免疫细胞化学法检测血管内皮钙黏蛋白（VE-cadherin）的表达。结果与正常组相比，TNF-α作用下EA．hy926单层细胞电阻值明显降低（67．200±8．937vs33．600±8．771，P=0．010），通透性增加，UTI在1-100U／mL范围内以剂量依赖性方式改善TNF-α所致的EA．hy926细胞高通透性（40．133±7．484vs33．600±8．771，P=0．382；49．232±3．162vs．33．600±8．771，P=0．044；63．700±8．515vs．33．600±8．771，P=0．013）。TNF-α作用下EA．hy926细胞VE-cadherinmRNA的表达较正常组显著降低（1．089±0．018vs0．835±0．021，P=0．000），UTI可抑制TNF-α引起的VE-cadherin低表达，其中UTI浓度为10U／mL和100U／mL时VE-cadherin mRNA的表达较TNF-α．组差异具有统计学意义（0．976±0．014vs．0．835±0．021，P=0．001；1．115±0．015vs．0．835±0．021，P=0．000），UTI的上述作用在1-100U／mL浓度范围内呈现出剂量依赖性。免疫细胞化学结果显示，与正常组相比，TNF-α作用下VE-cadherin的表达明显降低（0．061±0．013vs0．093±0．014，P=0．049），而UTI可改善TNF-α导致的VE-cadherin表达降低（0．032±0．004vs．0．061±0．013，P=0．016）。结论UTI可抑制TNF-α引起的EA．hy926细胞通透性增加，且在一定范围内呈现出剂量依赖性。
Objective To investigate the influence of Ulinastatin （UTI） on the hyper-permeability of vascular endothelial cells induced by tumor necrosis factor alpha （TNF-α）. Methods Inflammation model was induced by TNF-α in human umbilical vein endothelial cell line （ EA. hy926 ）. The experiment was designed into 4 groups： normal group, TNF-ct group, UTI group and TNF-α with UTI （U ＋ T） group. Methyl thiazolyl tetrazolium （MTT） method and epithelial vohameter （EVOM） method were used to measure cell viability [ absorbanee （A） value ] and transepithelial electrical resistance （TER） of EA. hy926 cells respectively. The expression of VE-eadherin was measured by reverse transcription polymerase chain reaction （ RT-PCR ） and immunocytochemistry. Results Compared with normal group, the TER of EA. hy926 cells induced by TNF- αwas significantly decreased （67. 200 ± 8. 937 vs. 33. 600 ± 8. 771, P = 0. 010）. The permeability in EA. hy926cells increased obviously. The hyper- permeability of EA. hy926 cells induced by TNF- αcould be alleviated by UTI at the concentrations of 1 - 100 U/mL in a dose-dependent manner （40. 133 ± 7.484 vs. 33. 600± 8.771, P=0.382;49.232±3.162 vs. 33. 600±8.771, P=0.044; 63. 700 ±8.515 vs. 33. 600±8.771, P= 0. 013）. The expression of VE-cadherin mRNA reduced significantly in the TNF-α group （1. 089 ±0. 018 vs. 0. 835 ±0. 021, P = 0. 000） compared with normal group. This effect of TNF-αcould be attenuated by UTI. When EA. hy926 cells exposed to UTI at 10 U/mL and 100 U/mL, a significant increase of the expression of VE-cadherin mRNA was observed （0. 976 ±0. 014 vs. 0. 835 ±0. 021, P =0. 001 ; 1. 115 ±0. 015 vs. 0. 835 ±0. 021, P = 0. 000）. And the inhibition of UTI manifested a dose-dependent manner （ 1 - 100 U/mL）. The results of the immunoeytochemistry showed that the expression of VE-cadherin in TNF-α group was decreased significantly （0. 061± 0. 013 vs. 0. 093 ±0. 014, P = 0.049） compared with normal group. And the low- expression of VE-cadherin could be alleviated by UTI （ 0. 032 ± 0. 004 vs. 0. 061 ± 0. 013, P = 0. 016 ）. Conclusion The high permeability of EA. hy926 cells induced by TNF-αcould be inhibited by UTI at the concentrations of 1 - 100 U/mL in a dose-dependent manner.
出处
《中华急诊医学杂志》
CAS
CSCD
北大核心
2016年第3期320-324,共5页
Chinese Journal of Emergency Medicine
基金
国家自然科学基金（81071531
81372102）
重庆市自然科学基金（CSTC2009BB5066）
天普研究基金项目（UF201314）
