肿瘤坏死因子-α和白细胞介素-1通过一条共同途径上调膜结合型和可溶性E-选择素。

TNF-alpha and IL-1 upregulate membrane-bound and soluble E-selectin through a common pathway.

作者信息

Wyble C W, Hynes K L, Kuchibhotla J, Marcus B C, Hallahan D, Gewertz B L

机构信息

Department of Surgery, University of Chicago, MC 5029, 5841 S. Maryland Ave., Chicago, Illinois 60637, USA.

出版信息

J Surg Res. 1997 Dec;73(2):107-12. doi: 10.1006/jsre.1997.5207.

DOI:10.1006/jsre.1997.5207

PMID:9441802

Abstract

BACKGROUND

Endothelial cell adhesion molecules such as E-selectin promote the capture of neutrophils (PMN) in the microcirculation and initiate the inflammatory response. In contrast, when "shed" into the microcirculation, soluble E-selectin can bind PMN in the blood stream, reducing the number available for adhesion to injured tissue. These experiments were designed to better characterize the molecular response to cytokines and the balance between cell surface (bound) and soluble (unbound) E-selectin.

METHODS

Cultured human umbilical veins, exposed to human recombinant TNF-alpha or IL-1 (10 pg/ml), were analyzed for E-selectin mRNA induction (Northern blot), E-selectin cell surface expression (flow cytometry), and sE-selectin release (ELISA). Transcriptional regulation was analyzed via Raf kinase dominant negative gene transfection.

RESULTS

E-selectin mRNA expression was markedly increased at 2 h and sustained through 8 h. No further induction was noted at 12 h. Upregulation of cell surface E-selectin was noted (mean fluorescence) as early as 2 h for TNF-alpha (baseline, 12.28 +/- 1.32; TNF-alpha, 23.03 +/- 1.81) or 4 h for IL-1 (baseline, 12.28 +/- 1.32; IL-1, 70.00 +/- 3.04) with maximum expression at 6 h (TNF-alpha, 118.8+/-15; IL-1, 94.11 +/- 9. 34). Expression returned to baseline levels by 24 h. Soluble E-selectin (ng/ml) assays demonstrated later increases beginning at 12 h (TNF-alpha, 0.313 +/- 0.077; IL-1, 0.159 +/- 0.075) and continuing through 24 h (TNF-alpha, 0.340 +/- 0.062; IL-1, 0.157 +/- 0.030). Transfection of endothelial cells with Raf kinase 301 dominant negative gene resulted in proportionate decreases in the peak expression in both surface (bound) E-selectin (TNF-alpha, 51. 7%; IL-1, 29.6%) and sE-selectin (TNF-alpha, 49.2%; IL-1, 34.5%).

CONCLUSION

The temporal sequence of late decreases in cell surface E-selectin accompanied by increases in soluble E-selectin indicates that the source of E-selectin in the microcirculation is shed receptors rather than synthesis of a different type of receptor. Enhancement of such "shedding" may decrease PMN adhesion to injured tissue and have therapeutic potential.

摘要

背景

内皮细胞黏附分子如E选择素可促进中性粒细胞（PMN）在微循环中的捕获并引发炎症反应。相反，当“脱落”进入微循环时，可溶性E选择素可与血流中的PMN结合，减少可黏附于受损组织的PMN数量。这些实验旨在更好地表征细胞因子的分子反应以及细胞表面（结合型）和可溶性（未结合型）E选择素之间的平衡。

方法

将培养的人脐静脉暴露于重组人肿瘤坏死因子-α或白细胞介素-1（10 pg/ml），分析E选择素mRNA诱导情况（Northern印迹法）、E选择素细胞表面表达情况（流式细胞术）以及可溶性E选择素释放情况（酶联免疫吸附测定法）。通过Raf激酶显性负性基因转染分析转录调控。

结果

E选择素mRNA表达在2小时时显著增加，并持续至8小时。12小时时未观察到进一步诱导。细胞表面E选择素上调（平均荧光强度）在肿瘤坏死因子-α作用下最早于2小时出现（基线值，12.28±1.32；肿瘤坏死因子-α，23.03±1.81），在白细胞介素-1作用下最早于4小时出现（基线值，12.28±1.32；白细胞介素-1，70.00±3.04），在6小时时达到最大表达（肿瘤坏死因子-α，118.8±15；白细胞介素-1，94.11±9.34）。到24小时时表达恢复至基线水平。可溶性E选择素（ng/ml）检测显示，从12小时开始出现后期增加（肿瘤坏死因子-α，0.313±0.077；白细胞介素-1，0.159±0.075），并持续至24小时（肿瘤坏死因子-α，0.340±0.062；白细胞介素-1，0.157±0.030）。用Raf激酶301显性负性基因转染内皮细胞导致表面（结合型）E选择素（肿瘤坏死因子-α，51.7%；白细胞介素-1，29.6%）和可溶性E选择素（肿瘤坏死因子-α，49.2%；白细胞介素-1，34.5%）的峰值表达成比例降低。