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In vivo cell-mediated immune reactions are characterized by mixtures of monocytes and T cells. The purpose of this study was to investigate the role of monocytes on T-cell migration and induction of endothelial adhesion molecules. The in vitro model consisted of adding peripheral blood mononuclear cells (PBMC), T cells or mixtures of monocytes and T cells, to endothelial cells on a porous membrane and using flow cytometry to distinguish between the monocyte and lymphocyte components. PBMC and PBMC supernatants were highly potent at upregulating intercellular adhesion molecule-1 (ICAM-1) and inducing expression of vascular cell adhesion molecule-1 (VCAM-1) and E-selectin. Induction by supernatants was inhibited by antibodies to tumour necrosis factor-α (TNF-α) and interleukin-1 (IL)-1β. Using monocyte-enriched populations, as few as one monocyte to 100 endothelial cells was sufficient to upregulate adhesion molecules. Fixed monocytes also induced adhesion molecules and expressed surface-bound cytokines. In contrast, highly purified unstimulated T cells were not found to induce adhesion molecules at 4, 6, 24 or 48 hr of coculture. Purified T cells showed low-level migration through resting (VCAM-1 negative) endothelium, which was approximately doubled by addition of small numbers of monocytes or TNF-α. In conclusion, monocytes, via cell surface or released cytokines play an essential role in allowing large-scale recruitment of T cells to inflammatory sites in vivo. [ABSTRACT FROM AUTHOR]

Objective: Decay-accelerating factor (DAF) is a widely expressed, multifunctional cell surface protein involved in complement regulation and cell signaling. Previous studies have demonstrated that endothelial cell (EC) DAF is up-regulated by tumor necrosis factor alpha and inhibits complement binding. Because vascular endothelial growth factor (VEGF) is cytoprotective to endothelium and is expressed at sites of chronic inflammation, we hypothesized that VEGF may induce DAF expression during inflammatory angiogenesis., Methods: Human umbilical vein and dermal microvascular EC were isolated using routine procedures, and the regulation and function of DAF, as well as other complement-regulatory proteins (membrane cofactor protein and CD59), were analyzed following stimulation with VEGF., Results: Incubation of large- or small-vessel EC with VEGF led to increased expression of DAF, with maximal expression after 48-72 hours of stimulation. This effect depended on the activation of protein kinase C (PKC) and required increased steady-state messenger RNA levels and de novo protein synthesis. Although VEGF-induced EC proliferation was inhibited by both p38 and p42/44 mitogen-activated protein kinase (MAPK) antagonists, DAF up-regulation in response to VEGF was only sensitive to inhibition of p38 MAPK. VEGF-stimulated EC showed a 60% reduction in C3 deposition following complement activation, and this resulted in a marked reduction in complement-mediated EC lysis. These protective effects were abolished by anti-DAF monoclonal antibody 1H4., Conclusion: This study confirms the importance of PKC for the regulation of DAF expression by EC and reveals VEGF to be a physiologic agonist for this pathway. The up-regulation of DAF expression by VEGF may represent an important mechanism for the protection of EC from complement-mediated injury during angiogenesis in inflammatory rheumatic diseases.