ENHANCED TNF-ALPHA PRODUCTION BY MONOCYTIC-LIKE CELLS EXPOSED TO DENGUE VIRUS-ANTIGENS

The studies indicating the importance of TNF alpha in dengue virus inf ection have led us to determine whether monocyte-like cells produce TN F alpha after exposure to dengue virus. The supernatant fluids of mosq uito cells (AP61) infected with dengue virus (DV) type 1 and DV type 3 were harvested 7 days post-infection and clarified. DV inactivation w as performed in the presence of betapropiolactone that preserves antig enicity of viruses. We used the monocytic-like cell line THP-1 that is a model system of TNF alpha production. Polymyxin B (50 mu g/ml) was added to block untoward effects resulting from possible LPS contaminat ion of media or cultures. THP-1 cells were primed with a phorbol ester (PMA) for 24 h, then they were cultured for 4 and 24 h in the presenc e of inactivated culture supernatant of dengue infected AP61 cells or control preparations. The concentrations of TNF alpha in the culture s upernatants were measured by using an immunoenzymatic assay. PMA-treat ed THP-1 cells rapidly secreted TNF alpha in response to inactivated c ulture supernatant of. DV-infected cells. We found high levels of TNF alpha with cells exposed to DV1 and DV3 preparations compared with con trols (mean values; 465 and 829 vs. 70 pg/ml, respectively, at 24 h po st exposure, n = 4). We obtained a substantial inhibition of the enhan cing activity of DV1 and DV3 infected supernatants in the presence of dengue hyperimmune mouse ascitic fluids. Our results demonstrate that exposure of monocytes/macrophages to DV particles or virus proteins de rived from DV may be responsible for the enhanced production of TNF al pha in DV-infected patients.