Release of thrombomodulin from endothelial cells by concerted action of TNF-alpha and neutrophils: in vivo and in vitro studies.
Release of thrombomodulin from endothelial cells by concerted action of TNF-alpha and neutrophils: in vivo and in vitro studies.
复制标题
通过 TNF-α 和中性粒细胞的协同作用从内皮细胞释放血栓调节蛋白：体内和体外研究。
作者：
M. Boehme;You;U. Raeth;A. Bierhaus;R. Ziegler;Wolfgang Stremmel;Peter P. Nawroth
炎症细胞因子通过抑制血栓调节素（TM）的转录和翻译或内化并随后降解来降低内皮细胞表面血栓调节素（TM）的表达。然而，血清TM水平升高是在系统性或局部炎性细胞因子水平升高相关的疾病中发现的。为了直接研究肿瘤坏死因子- α (tnf - α)在体内的作用，我们测定了系统性重组人（rh） tnf - α治疗后血清TM的过程。采用酶联免疫吸附试验（ELISA）检测TM水平。全身rhtnf - α治疗导致血清TM显著升高。通过小鼠模型，我们研究了血清TM的增加是否与体内内皮表面TM表达的减少有关。注射tnf - α后4小时，小鼠移植的甲基a肉瘤的血管免疫组化染色显示TM免疫反应性丧失。为了研究tnf - α介导的TM释放机制，我们将培养的内皮细胞与中性粒细胞和tnf - α孵育。单独与tnf - α孵育不导致体外TM的增加。然而，当用tnf - α预处理的内皮细胞暴露于中性粒细胞时，TM被释放到培养上清中。这与内皮细胞损伤的形态学证据有关。因此，细胞因子刺激的内皮细胞和中性粒细胞的协同作用导致rhtnf - α治疗后培养的内皮细胞释放TM。这可能解释了尽管tnf - α对TM转录和翻译具有诱导内化和直接抑制作用，但在与全身或局部炎症细胞因子水平升高相关的疾病中观察到的血清TM水平升高。
Inflammatory cytokines decrease the expression of thrombomodulin (TM) on the endothelial cell surface by suppression of TM transcription and translation or internalization with subsequent degradation. Nevertheless, elevated serum TM levels are found in diseases associated with systemical or locally increased levels of inflammatory cytokines. To study directly the in vivo effects of tumour necrosis factor-alpha (TNF-alpha) we determined the course of serum TM after systemic recombinant human (rh)TNF-alpha therapy. The TM levels were determined by enzyme-linked immunosorbent assay (ELISA). Systemic rhTNF-alpha therapy resulted in a marked and significant increase of serum TM. Using a mouse model we studied whether increased serum TM is associated with a decreased expression of TM on the endothelial surface in vivo. The immunohistochemical staining of the vasculature of meth-A sarcoma transplanted in mice showed a loss of TM immunoreactivity 4 hr after intravenous TNF-alpha application. To study the mechanism of TNF-alpha mediated release of TM, cultured endothelial cells were incubated with neutrophils and TNF-alpha. Incubation with TNF-alpha alone did not lead to an increase of TM in vitro. However TM was released into the culture supernatant when endothelial cells pretreated with TNF-alpha were exposed to neutrophils. This was associated with morphological evidence of endothelial cell damage. Therefore, the concerted action of cytokine-stimulated endothelial cells and neutrophils results in release of TM from cultured endothelial cells after rhTNF-alpha therapy. This might explain the increased serum TM levels observed in diseases associated with increased systemic or local levels of inflammatory cytokines despite the induced internalization and the direct inhibitory effects of TNF-alpha on TM transcription and translation.