Vaday G G, Hershkoviz R, Rahat M A, Lahat N, Cahalon L, Lider O
Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.
J Leukoc Biol. 2000 Nov;68(5):737-47.
Tumor necrosis factor alpha (TNF-alpha) is a proinflammatory cytokine implicated in the stimulation of matrix metalloproteinase (MMP) production by several cell types. Our previous studies demonstrated that TNF-alpha avidly binds fibronectin (FN) and laminin, major adhesive glycoproteins of extracellular matrix (ECM) and basement membranes. These findings suggested that TNF-alpha complexing to insoluble ECM components may serve to concentrate its activities to distinct inflamed sites. Herein, we explored the bioactivity and possible function of ECM-bound TNF-alpha by examining its effects on MMP-9 secretion by monocytes. Immunofluorescent staining indicated that LPS-activated monocytes deposited newly synthesized TNF-alpha into ECM-FN. FN-bound TNF-alpha (FN/TNF-alpha) significantly up-regulated MMP-9 expression and secretion by the human monocytic cell line MonoMac-6 and peripheral blood monocytes. Such secretion could be inhibited by antibodies that block TNF-alpha activity and binding to its receptors TNF RI (p55) and TNF RII (p75). Cheniotaxis through ECM gels in the presence of soluble or bound TNF-alpha was inhibited by a hydroxamic acid inhibitor of MMPs (GM6001). It is interesting that, although the adhesion of MonoMac-6 cells to FN/TNF-alpha required functional activated beta1 integrins, FN/TNF-alpha-induced MMP-9 secretion was independent of binding to beta1 integrins, since MMP-9 secretion was unaffected by: (1) neutralizing nAb to alpha4, alpha5, and beta1 subunits, which blocked cell adhesion; (2) a mAb that stimulated beta1 integrin-mediated adhesion; and (3) binding TNF-alpha to the 30-kDa amino-terminal fragment of FN, which lacks the major cell adhesive binding sites. Thus, in addition to their cell-adhesive roles, ECM glycoproteins, such as FN, may play a pivotal role in presenting proinflammatory cytokines to leukocytes within the actual inflamed tissue, thereby affecting their capacities to secrete ECM-degrading enzymes. These TNF-alpha-ECM interactions may serve to limit the cytokine's availability and bioactivity to target areas of inflammation.
肿瘤坏死因子α（TNF-α）是一种促炎细胞因子，与多种细胞类型刺激基质金属蛋白酶（MMP）的产生有关。我们之前的研究表明，TNF-α能与纤连蛋白（FN）和层粘连蛋白紧密结合，这两种蛋白是细胞外基质（ECM）和基底膜的主要黏附糖蛋白。这些发现表明，TNF-α与不溶性ECM成分的复合可能有助于将其活性集中于不同的炎症部位。在此，我们通过研究其对单核细胞分泌MMP-9的影响，探讨了与ECM结合的TNF-α的生物活性和可能的功能。免疫荧光染色表明，LPS激活的单核细胞将新合成的TNF-α沉积到ECM-FN中。与FN结合的TNF-α（FN/TNF-α）显著上调了人单核细胞系MonoMac-6和外周血单核细胞中MMP-9的表达和分泌。这种分泌可被阻断TNF-α活性及其与受体TNF RI（p55）和TNF RII（p75）结合的抗体所抑制。在存在可溶性或结合型TNF-α的情况下，通过ECM凝胶的趋化作用被MMPs的一种异羟肟酸抑制剂（GM6001）所抑制。有趣的是，尽管MonoMac-6细胞与FN/TNF-α的黏附需要功能性激活的β1整合素，但FN/TNF-α诱导的MMP-9分泌与β1整合素的结合无关，因为MMP-9的分泌不受以下因素影响：（1）针对α4、α5和β1亚基的中和性单克隆抗体，这些抗体可阻断细胞黏附；（2）一种刺激β1整合素介导黏附的单克隆抗体；（3）TNF-α与FN的30 kDa氨基末端片段结合，该片段缺乏主要的细胞黏附结合位点。因此，除了其细胞黏附作用外，ECM糖蛋白，如FN，可能在向实际炎症组织中的白细胞呈递促炎细胞因子方面发挥关键作用，从而影响它们分泌ECM降解酶的能力。这些TNF-α-ECM相互作用可能有助于限制细胞因子在炎症靶区域的可用性和生物活性。
