Figure 6.

Rac1 and NADP(H) oxidases are key effectors of renal epithelial growth inhibition in response to TGF-β1. A) A schematic representation of study design for Rac1 involvement in renal epithelial cell cycle arrest. Briefly, subconfluent control shRNA or Rac1 shRNA stably transduced HK-2 epithelial cells at a similar density were serum-deprived for 1 d then incubated with TGF-β for 1 d followed by serum addition (1%) for 3 d to stimulate cell growth. B) Relative epithelial cell counts (mean ± sd) are plotted in setting the cell number in untreated (−) control shRNA cultures as 1. *P < 0.05 as indicated. N.S., not significant. C–F) To investigate the potential involvement of p22^Phox and p47^Phox subunits of the NADP(H) oxidases in TGF-β1–induced epithelial growth inhibition, study designs are adopted similar to above and the schematics (C, E) illustrate the experimental approaches. Semiconfluent and serum-starved control shRNA and p22^Phox or p47^Phox shRNA stably expressing renal epithelial cells at a similar density were incubated with TGF-β for 1 d followed by 3 d of 1% serum stimulation. Relative cell counts (mean ± sd) provided the comparisons of cell growth between control shRNA and p22^Phox shRNA stable transductants (D) or control shRNA and p47^Phox shRNA (F) stably expressing HK-2 cells setting the cell number in untreated control shRNA as 1 in each case. Con, control; NS, not significant; n = 3. *P < 0.05 as indicated.