Cellular and molecular mechanisms of bromate-induced cytotoxicity in human and rat kidney cells.
The mechanisms of bromate (BrO(3)(-))-induced toxicity in Normal Rat Kidney (NRK) and human embryonic kidney 293 (HEK293) cells were investigated. BrO(3)(-) (added as KBrO(3)) induced concentration-dependent decreases in 3-(4, dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) staining after 48 h. BrO(3)(-)-induced necrosis based on tandem increases in annexin V and PI staining. Cell cycle analysis demonstrated that BrO(3)(-) also induced G2/M arrest and nuclear fragmentation, prior to alterations in MTT staining or annexin V and PI staining. Immunoblot analysis demonstrated that the G2/M arrest correlated to induction of phosphorylated (p)-p53, p21, cyclin B1 and p-cdc2. Further, BrO(3)(-) induced time-dependent increases in the activity of the mitogen activated protein kinases p38 and ERK1/2. Treatment of cells with the p38 inhibitor SB202190, but not the ERK1/2 inhibitor PD98059, partially reversed BrO(3)(-)-induced G2/M arrest and decreased BrO(3)(-)-induced p-p53, p21 and cyclin B1 expression. In addition, BrO(3)(-) treatment induced reactive oxygen species (ROS) based on increases in CM-H(2)DCFDA fluorescence. The antioxidant ascorbic acid inhibited BrO(3)(-)-induced p38 activation, G2/M arrest, p-p53, p21 and cyclin B1 expression; however, ascorbic acid had no effect on BrO(3)(-)-induced formation of 8-OHdG, a marker of DNA oxidative damage, whose increases preceded cell death by 24h. These data suggest that ROS mediated MAPK activation is involved in the molecular mechanisms of BrO(3)(-)-induced cell cycle arrest, which occurs independently of 8-OH-dG production. The similar mode of action in both NRK and HEK293 cells suggests that the mechanisms of BrO(3)(-)-induced renal cell death are model-independent.
Zhang
Xiaoling
X
Department of Pharmaceutical and Biomedical Sciences, 336 Pharmacy South, College of Pharmacy, University of Georgia, Athens, GA 30602, USA.
De Silva
Dilhara
D
Sun
Bin
B
Fisher
Jeffery
J
Bull
Richard J
RJ
Cotruvo
Joseph A
JA
Cummings
Brian S
BS
D000818:Animals; D001959:Bromates / Q000633:toxicity*; D002453:Cell Cycle / Q000187:drug effects* / Q000502:physiology; D002460:Cell Line; D006801:Humans; D007668:Kidney / Q000187:drug effects* / Q000473:pathology* / Q000502:physiology; D051381:Rats
D001959:Bromates; C019536:potassium bromate