Effect of OPN on the proliferation of A549.

<p>(A and B) A549 were transfected with 20 nM human <i>OPN</i> siRNA or control siRNA. Twenty-four h after transfection, A549 were incubated for 48 h in the presence of 1 µg/mL bleomycin or PBS. (A) <i>OPN</i> mRNA was assessed by qRT-PCR. (B) OPN concentrations in cell culture media were determined by ELISA. (C and D) A549 were transfected with 20 nM human <i>OPN</i> siRNA or control siRNA. Twenty-four h after transfection, A549 were exposed to 1 µg/mL bleomycin for 48 h and then incubated with EdU for an additional 15 h. A549 were stained with Alexa Fluor 488 azide and HCS NuclearMask Blue stain. The cells were visualized by pseudocoloring with the Floid Cell Imaging Station. (C) Nuclei (i, ii) show representative photomicrographs of all cells. EdU (iii, iv) show representative photomicrographs of proliferating cells. Merge (v, vi) show representative photomicrographs of merged images. Bar: 100 µm. (D) Twelve fields per well were recorded and the number of positively-staining cells was counted. The percentages of proliferating A549 that were transfected with either human <i>OPN</i> siRNA or control siRNA are shown. Data are presented as mean ± S.E. *, P<0.05; **, P<0.01.