Fig. 3.

Effect of puerarin on VSMCs proliferation. VSMCs viability was assessed by CCK-8 assay. a Cells were stimulated with different concentrations of PM2.5 (0, 25, 50, 100, 200, 400 mg/L) for 24 h. b Cells were stimulated with 200 mg/L PM2.5 for the different time points (0, 3, 6, 12, 24, 48 h). c Cells were stimulated with different concentrations of puerarin (0, 5, 10, 25, 50, 100 μM) for 24 h. d Cells were pre-treated with puerarin at different concentrations (0, 12.5, 25, 50 μM) or p38 MAPK inhibitor SB203580 (20 μM) for 1 h and followed by the addition of 200 mg/L PM2.5 for 24 h. VSMCs proliferation was measured according to the BrdU immunofluorescence. e Confocal images. a, the untreated cells; b, cells were stimulated with 200 mg/L PM2.5 for 24 h; c, cells were pre-treated with 12.5 μM puerarin for 1 h and followed by the addition of 200 mg/L PM2.5 for 24 h; d, cells were pre-treated with 25 μM puerarin for 1 h and followed by the addition of 200 mg/L PM2.5 for 24 h; e, cells were pre-treated with 50 μM puerarin for 1 h and followed by the addition of 200 mg/L PM2.5 for 24 h; f, cells were pre-treated with 50 μM puerarin and p38 MAPK inhibitor SB203580 (20 μM) for 1 h and followed by the addition of 200 mg/L PM2.5 for 24 h. Bars = 100 μm. f Graphs of BrdU positive ratios. The results are presented as mean ± SEM. n = 3. Compared to the untreated cells, ^*P < 0.05, ^**P < 0.01; compared to 200 mg/L PM2.5 group, ^#P < 0.05, ^##P < 0.01; compared to the co-incubation of puerarin with SB203580 group, ^▲P < 0.05, ^▲▲P < 0.01