Production of IL-8 by cultured endothelial cells in response to Borrelia burgdorferi occurs independently of secreted IL-1 and TNFa and is required for subsequent transendothelial migration of neutrophils

Previous studies have shown that Borrelia burgdorferi (Bb), the spirochetal agent of Lyme disease, promotes inflammation by stimulating endothelial cells to upregulate adhesion molecules for leukocytes and to produce a soluble agent that is chemotactic for neutrophils. We determined that IL-8 was the chemotactic agent for neutrophils present in conditioned media from cultured human umbilical vein endothelial cells (HUVEC) stimulated with Bb. As few as one spirochete per HUVEC stimulated production of IL-8 within 8 h of co-incubation. When ten spirochetes per HUVEC were added, IL-8 was detected after 4 h of co-culture. Production of IL-8 continued in a linear fashion for at least 24 h. Neutralizing antibodies against IL-8 reduced migration of neutrophils across spirochete-stimulated HUVEC monolayers by 93%. In contrast, pretreatment of neutrophils with antagonists of platelet activating factor did not inhibit migration. Increases in production of IL-8 and expression of the adhesion molecule E-selectin by HUVEC in respone to Bb were not inhibited by IL-1 receptor antagonist or a neutralizing monoclonal antibody directed against tumor necrosis factor alpha, used either alone or in combination. These results suggest that activation of HUVEC by Bb is not mediated through the autocrine action of secreted IL-1 or tumor necrosis factor alpha. Rather, it appears that Bb must stimulate endothelium either by a direct signaling mechanism or by induction of a novel host-derived cytokine.