CHARACTERIZATION OF A TUMOR NECROSIS FACTOR-ALPHA (TNF-ALPHA) INHIBITOR - EVIDENCE OF IMMUNOLOGICAL CROSS-REACTIVITY WITH THE TNF RECEPTOR

Previous studies have shown that urine of febrile patients contains a tumor necrosis factor α inhibiting activity (TNF-α Inh) when tested in a cytotoxicity assay using the tumor necrosis factor α (TNF-α)-susceptible cell line L929. In the present study, we investigated the relationship between the TNF-α Inh and a potential soluble form of the receptor, as the former has been shown to block TNF-α activities by binding to the ligand. We demonstrate that human TNF-α is affected to a greater extent than is murine TNF-α. This species specificity of the inhibitor correlates with the binding studies of TNF receptor interactions already reported. We raised a polyclonal antibody to TNF-α Inh that neutralizes its activity and does not recognize TNF-α. Solubilized cross-linked 125I-labeled TNF-α receptor complex could be inununoprecipitated by using either anti-TNF-α or anti-TNF-α Inh antibody, suggesting immunological cross-reactivity between the receptor and the inhibitor. By using fluorescein isothiocyanate-coupled TNF-α, it was possible to visualize by fluorescence-activated cell sorter analysis the TNF-α receptor on phytohemagglutinin/interleukin 2-activated T cells. A similar increase of immunofluorescence intensity of the activated T cells was observed by using anti-TNF-α Inh antibody revealed with a fluorescein isothiocyanate-coupled goat anti-rabbit IgG1 conjugate, suggesting that the TNF-α Inh is also expressed as a membrane protein. Taken together, our results suggest that the TNF-α Inh originally described might be a soluble form of the TNF receptor itself.