Involvement of p38 MAPK in the up-regulation of tissue factor on endothelial cells by antiphospholipid antibodies.
Involvement of p38 MAPK in the up-regulation of tissue factor on endothelial cells by antiphospholipid antibodies.
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p38 MAPK 参与抗磷脂抗体对内皮细胞组织因子的上调。
DOI：
10.1002/art.21009
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发表时间：
2005
期刊：
影响因子：
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通讯作者：
Pierangeli,SilviaS
中科院分区：
文献类型：
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作者：
Vega-Ostertag,Mariano;Casper,Katherine;Swerlick,Robert;Ferrara,Dardo;Harris,ENigel;Pierangeli,SilviaS
目的研究抗磷脂抗体（aPL）上调内皮细胞（ECs）组织因子（TF）的细胞内机制，研究aPL对内皮细胞（HUVECs）组织因子（TF）的转录、表达和功能、白细胞介素- 6 （IL - 6）和IL - 8的表达、诱导型一氧化氮合酶（iNOS）的诱导以及p38 MAPK磷酸化的影响。方法将培养的HUVECs用抗磷脂综合征（APS）患者的IgG aPL或正常人血清的IgG作为对照。以肉豆蔻酸酯佛波酯（PMA）和细菌脂多糖（LPS）为阳性对照。采用酶联免疫吸附试验（ELISA）检测HUVECs表面的TF表达。用细胞裂解物显色法测定TF活性，用实时定量聚合酶链反应测定TF信使RNA （mRNA）。Western blotting检测p38 MAPK磷酸化水平和iNOS诱导水平，ELISA检测IL - 6和IL - 8水平。结果与对照组相比，spma、LPS和aPL显著提高了TF的表达。这种上调被SB203580 （p38 MAPK的特异性抑制剂）和MG132 （NF - κB的特异性抑制剂）显著抑制。IgG aPL显著提高了TF活性，SB203580也抑制了这种作用。与aPL一起孵育的HUVECs使TF mRNA增加2-15倍；这些影响已被SB203580废除。IgG aPL诱导p38 MAPK显著磷酸化，并以时间依赖性的方式在HUVECs上产生iNOS。IgG - aPL也能诱导HUVECs上IL - 6和IL - 8的表达增加。我们的数据表明，aPL诱导了TF转录、功能和表达的显著增加，IL - 6和IL - 8的上调，以及HUVECs上iNOS的表达，这些过程涉及p38 MAPK的磷酸化和NF - κB的激活。了解aPL介导的EC激活的细胞内事件可能有助于设计针对APS血栓形成的新的靶向治疗方法。
ObjectiveTo study the intracellular mechanism involved in the up‐regulation of tissue factor (TF) on endothelial cells (ECs) by antiphospholipid antibodies (aPL), we examined the effects of aPL on the transcription, expression, and function of TF, the expression of interleukin‐6 (IL‐6) and IL‐8, the induction of inducible nitric oxide synthase (iNOS), and the phosphorylation of p38 MAPK on human umbilical vein ECs (HUVECs).MethodsCultured HUVECs were treated with IgG aPL (from patients with antiphospholipid syndrome [APS]) or with control IgG (from normal human serum). Phorbol myristate acetate (PMA) and bacterial lipopolysaccharide (LPS) were used as positive controls. TF expression was determined on the surface of HUVECs using an enzyme‐linked immunosorbent assay (ELISA). TF activity was determined with the use of a chromogenic assay in cell lysates, and TF messenger RNA (mRNA) was determined by real‐time quantitative polymerase chain reaction. Phosphorylation of p38 MAPK and induction of iNOS were determined by Western blotting, and levels of IL‐6 and IL‐8 were determined by ELISA.ResultsPMA, LPS, and aPL significantly increased the expression of TF compared with controls. This up‐regulation was significantly inhibited by SB203580 (a specific inhibitor of p38 MAPK) and by MG132 (a specific inhibitor of NF‐κB). TF activity was significantly increased by treatment with IgG aPL and this effect was also inhibited by SB203580. Incubation of HUVECs with aPL increased TF mRNA 2–15‐fold; these effects were abrogated by SB203580. IgG aPL induced significant phosphorylation of p38 MAPK and produced iNOS on HUVECs in a time‐dependent manner. Treatment with IgG aPL also induced increased expression of IL‐6 and IL‐8 on HUVECs.ConclusionOur data show that aPL induces significant increases in TF transcription, function, and expression, in IL‐6 and IL‐8 up‐regulation, and in iNOS expression on HUVECs and that these processes involve phosphorylation of p38 MAPK and activation of NF‐κB. Understanding intracellular events in aPL‐mediated EC activation may help in designing new targeted therapies for thrombosis in APS.