Weber C, Negrescu E, Erl W, Pietsch A, Frankenberger M, Ziegler-Heitbrock H W, Siess W, Weber P C
Institute for Cardiovascular Diseases, Ludwig Maximilians University, Munich, Germany.
J Immunol. 1995 Jul 1;155(1):445-51.
We studied the role of tyrosine phosphorylation in the induction of vascular cell adhesion molecule 1 (VCAM-1), endothelial leukocyte adhesion molecule 1 (ELAM-1), and intercellular adhesion molecule 1 (ICAM-1) in HUVEC. Induction of VCAM-1 and ELAM-1 surface expression by TNF was dose-dependently reduced by pretreatment with the protein tyrosine kinase inhibitors herbimycin A (HMA, IC50 300 nM) or genistein (IC50 30 microM). Only genistein attenuated ICAM-1 induction. Genistein or HMA did not affect adhesion molecule up-regulation by PMA. U937 monocyte adhesion to TNF-stimulated HUVEC was markedly inhibited by a combination of anti-VCAM-1 and anti-ELAM-1 mAb, as well as by HMA or genistein, probably due to suppression of VCAM-1 and ELAM-1 up-regulation. HMA appeared to prevent VCAM-1 transcription, since it reduced induction of VCAM-1 mRNA by TNF. Gelshift analysis demonstrated inhibition of TNF-induced nuclear factor-kappa B (NF-kappa B) mobilization by HMA. TNF rapidly enhanced tyrosine phosphorylation of a protein migrating with an apparent molecular mass of 35 kDa. HMA and genistein suppressed constitutive tyrosine phosphorylation of all detectable proteins and prevented TNF-induced tyrosine phosphorylation of the 35 kDa protein with an IC50 and dose range, similar to inhibition of VCAM-1 and ELAM-1 induction. Our data suggest that specific phosphorylation following protein tyrosine kinase activation may be required for NF-kappa B mobilization and induction of VCAM-1 and ELAM-1 by TNF.
我们研究了酪氨酸磷酸化在人脐静脉内皮细胞（HUVEC）中诱导血管细胞黏附分子1（VCAM-1）、内皮白细胞黏附分子1（ELAM-1）和细胞间黏附分子1（ICAM-1）表达过程中的作用。用蛋白酪氨酸激酶抑制剂除莠霉素A（HMA，IC50为300 nM）或染料木黄酮（IC50为30 μM）预处理后，肿瘤坏死因子（TNF）诱导的VCAM-1和ELAM-1表面表达呈剂量依赖性降低。只有染料木黄酮减弱了ICAM-1的诱导。染料木黄酮或HMA不影响佛波酯（PMA）引起的黏附分子上调。抗VCAM-1和抗ELAM-1单克隆抗体（mAb）的组合，以及HMA或染料木黄酮，均可显著抑制U937单核细胞与TNF刺激的HUVEC的黏附，这可能是由于抑制了VCAM-1和ELAM-1的上调。HMA似乎可阻止VCAM-1转录，因为它降低了TNF诱导的VCAM-1 mRNA的表达。凝胶迁移分析表明，HMA可抑制TNF诱导的核因子κB（NF-κB）的激活。TNF可迅速增强一种表观分子量为35 kDa的蛋白质的酪氨酸磷酸化。HMA和染料木黄酮可抑制所有可检测蛋白质的组成型酪氨酸磷酸化，并阻止TNF诱导的35 kDa蛋白质的酪氨酸磷酸化，其IC50和剂量范围与抑制VCAM-1和ELAM-1诱导相似。我们的数据表明，蛋白酪氨酸激酶激活后的特异性磷酸化可能是NF-κB激活以及TNF诱导VCAM-1和ELAM-1表达所必需的。
