p38 MAP kinase mediates gp120-induced iDC chemotaxis downstream of Pyk2. (A) iDCs were pretreated with SB203589 (10 μM) for 1 hour at 37°C and stimulated with gp120 (10 nM) at 37°C for various periods of time. The cells were lysed and analyzed by Western blotting with anti–phospho-p38 antibodies (left panel). The same blots were reprobed with anti-p38 antibodies. iDCs were pretreated with vehicle or tyrphostin A9 (5 μM) for 1 hour at 37°C. The cells were then stimulated with HIV-1 gp120 for 15 minutes, lysed, and analyzed by Western blotting with anti–phospho-p38 antibody (right panel). The same blot was reprobed with anti-p38 antibody. The bar graph at the bottom of the panels represents the phosphorylation index as obtained by densitometry. *P < .05 versus the unstimulated control. Data represent mean ± SD of 3 independent experiments. (B) iDCs were pretreated with PD98059 (10 μM) for 1 hour at 37°C and stimulated with gp120 (10 nM) at 37°C for various periods of time. The cells were lysed and analyzed by Western blotting with anti–p-ERK1/2 antibodies. The same blots were reprobed with anti–ERK1/2 antibodies. Data show 1 representative experiment of 3 independent experiments. (C) iDCs were pretreated with vehicle or various concentrations of the p38 MAP kinase inhibitors, SB203580 (top left panel) and SB220025 (top right panel) as well as the ERK kinase inhibitor (PD98059; bottom panel) for 1 hour at 37°C. The migration of these cells in response to M-tropic gp120 (10 nM) was determined after 3 hours of incubation. *P < .05 versus the vehicle control.