Figure 2. Inhibition of Tat-induced TNFα production by ibudilast is not reversed by blockade of adenosine A_2A receptor.

A, BV-2 cells (1.5×10⁵) were left untreated (NT) or were treated with Tat (100 nM) for 8 h with or without pre-treatment for 30 min. with either anti-Tat or control IgG antibodies (8 µg/mL) or adenosine A_2A receptor agonist, CGS21680 (CGS, 1 µM) or antagonist, ZM241385 (ZM, 100 nM), as indicated. TNFα release was measured by ELISA. The Tat-treated samples were set to 100% and all other samples were compared to this value (the TNFα concentration for this sample was 3475 pg/µg total protein content). Results are shown as mean ± SEM of values derived from four replicates from a single representative experiment; two total experiments were performed. Statistical significance (p<0.001) is indicated, as compared to Tat-treated cells (***) or as compared to Tat+CGS-treated cells (###). B, BV-2 cells (1.2×10⁵) were left untreated (NT) or were treated with Tat (100 nM) for 8 h with or without pre-treatment for 30 min. with ibudilast (Ib, 50 µM) or vehicle (Veh) alone or together with increasing concentrations of the adenosine A_2A receptor antagonist, ZM241385, as indicated. TNFα release was measured by ELISA. The Tat+Veh-treated samples were set to 100% and all other samples were compared to this value (the TNFα concentration for this sample was 696.2 pg/mL). Results are shown as mean ± SEM of values derived from three replicates from a single representative experiment; two total experiments were performed. Statistical significance is indicated, as compared to Tat+Veh-treated cells (p<0.05, *) or as compared to Tat+ZM241385-treated cells (p<0.001, ###).