Tumor Necrosis Factor-α Pathway Plays a Critical Role in Regulating Interferon-γ Induced Protein-10 Production in Initial Allogeneic Human Monocyte–Endothelial Cell Interactions

Abstract

T-cell infiltration of allografts is a major pathologic component defining acute rejection episodes (ARE). We have shown that monocytes interact with allogeneic endothelial cells (ECs) for costimulation to achieve T-cell allorecognition. However, the production of T-cell interferon-γ induced protein-10 (IP-10) and regulation of this chemokine during the initial monocyte–EC interaction are unclear. We hypothesized that the tumor necrosis factor (TNF)-α pathway plays a key role to regulate IP-10 production during the initial monocyte–EC interaction. Cytokine-activated ECs were analyzed for IP-10 production and adhesion molecule expression. Established, monocyte–EC cocultures were analyzed using real-time polymerase chain reaction and a chemokine assay for IP-10 and activation factors. Anti–TNF-α antibody was used to neutralize TNF-α release during monocyte–EC interactions. TNF-α–activated ECs upregulated CD62E and CD54 as determined by flow cytometry, releasing high levels of IP-10 and interleukin (IL)-6. Interferon-γ–stimulated ECs also produced high levels of IP-10 and IL-6. Monocyte–EC interactions demonstrated upregulation of gene transcripts for TNF-α, IL-6, and IP-10. The cytokine/chemokine assay detected high levels of TNF-α, IL-6, and IP-10 in coculture supernates in a time-dependent manner. Anti–TNF-α antibody dramatically reduced IP-10 production by monocyte–ECs interactions. However, anti–TNF-α antibody did not prevent the release of IL-6 by monocytes in EC cocultures. Our results showed that ECs activated by TNF-α are an important source of IP-10. The monocyte–EC interaction produces high levels of IP-10. The TNF-α pathway plays a key role to regulate IP-10 production during monocyte-EC interactions. We thus proposed that the initial monocyte–EC interaction with increased expression of IP-10 may play a critical role to initiate and augment T-cell–mediated ARE.