Nef synergizes with IFNβ in iNOS production.

<p>(<b>A</b>) BV-2 cells were treated for 6 h with myr⁺Nef_SF2 (100 ng/ml), IFNβ (200 IU/ml) or a combination of both. iNOS mRNA was measured by real time RT-PCR as reported in the materials and methods section. (<b>B</b>) Cells were incubated for 24 h with the indicated dose of myr⁺Nef_SF2 with or without IFNβ (50 IU/ml). Cells were also incubated with heat-inactivated recNef_SF2 (inactiv. recNef, 500 ng/ml) or, as control, with LPS or pre-heated LPS (500 ng/ml each). Total cellular lysates were analyzed by Western Blot for iNOS expression. β-Tubulin expression was used as an internal loading control. (<b>C</b>) Cells were treated like in (<b>B</b>) and NO₂^- content in supernatants was measured using the Griess colorimetric assay. Empty diamond: IFNβ; filled circles: myr⁺Nef_SF2; gray empty circles: heat-inactivated myr⁺Nef_SF2; filled squares: recNef_SF2 plus IFNβ; gray empty squares: heat-inactivated myr⁺Nef_SF2 plus IFNβ.