Pyrogallol, ROS generator inhibits As4.1 juxtaglomerular cells via cell cycle arrest of G2 phase and apoptosis

Pyrogallol as a catechin compound has been employed as an O-2(center dot-) generator and often used to investigate the role of ROS in the biological system. Here, we investigated the in vitro effect of pyrogallol on cell growth, cell cycle and apoptosis in As4.1 juxtaglomerular cells. Dose-dependent inhibition of cell growth was observed with IC50 of about 60 I-LM for 48 It using MTT assay. Pyrogallol (100 mu M) did not alter intracellular H2O2 level and catalase activity, but increased the intracellular O-2(center dot-) level and decreased SOD activity in As4.1 cells. DNA flow cytometric analysis indicated that 50 and 100 mu M pyrogallol significantly increased G2 phase cells as compared with those of pyrogall ol -untreated cells. Also, pyrogallol induced apoptosis as evidenced by flow cytometric detection of sub-GI DNA content, annexin V binding assay and DAPI staining. This apoptosis process was accompanied with the loss of mitochondrial transmembrane potential (Delta psi(m)), Bcl-2 decrease, caspase-3 activation and PARP cleavage. Pan caspase inhibitor (Z-VAD) could significantly rescue As4.1 cells from pyrogallol-induced cell death. But, the inhibitors of caspase-3, caspase-8, and caspase-9 did not prevent apoptotic events in pyrogallol-treated As4.1 cells. Taken together, we have demonstrated that an ROS inducer, pyrogallol inhibits the growth of As4.1 JG cells via cell cycle arrest and apoptosis, and suggest that the compound exhibits an anti-proliferative efficacy on these cells. (C) 2007 Published by Elsevier Ireland Ltd.