Fig. 5.

Effects of isoproterenol (ISO) on phosphorylation of S6 in HMB45-positive lung LAM cells. (A and B) Lung LAM cells after a 1-h incubation with DMSO or 200 nM sirolimus (Siro), followed by a 1-h incubation with 1 μM ISO, were fixed and stained with indicated antibodies. After incubation with DMSO or 200 nM Siro for 3 d, with or without 1 μM ISO (C) or with ISO for 1 h each day for 3 d (D), cells were fixed, reacted with rabbit anti–phospho-S6 polyclonal antibodies and mouse HMB45 monoclonal antibodies, and then prepared for confocal immunofluorescence microscopy. (Scale bar: 10 μm.) Mean fluorescence intensity of phospho-S6 in outlined area of each cell (>50 in each population) was measured in pixels by ImageJ software (NIH). *P < 0.01 vs. DMSO; **P < 0.05 vs. Siro. Similar results were obtained from three independent experiments.