p38 mitogen-activated protein kinase protects human retinal pigment epithelial cells exposed to oxidative stress.
OBJECTIVE
To investigate the role of p38 mitogen-activated protein kinase (MAPK) in human retinal pigment epithelial (RPE) cells exposed to acute oxidative stress.


METHODS
Experimental study.


METHODS
Oxidative stress was induced by the chemical oxidant tert-butyl hydroperoxide (t-BOOH) in the human RPE cell line ARPE-19. Cell viability was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The phosphorylation of p38 MAPK was measured by Western blot analysis. Small interfering (si)RNA and plasmid DNA of a constitutive active mutant of a MAPK kinase (MKK6E) were transfected to knock down and activate p38 MAPK in ARPE-19 cells, respectively.


RESULTS
t-BOOH induced ARPE-19 cell death in a time- and dose-dependent manner. t-BOOH increased phosphorylation of p38 MAPK. Both inhibition of p38 MAPK with a selective inhibitor SB203580 and knockdown of p38? MAPK with siRNA enhanced t-BOOH-induced ARPE-19 cell death (by 29% and 20%, respectively). Overexpression of MKK6E, a kinase upstream of p38 MAPK, increased phosphorylation of p38 MAPK and attenuated t-BOOH-induced ARPE-19 cell death by 16%. Preconditioning with a low dose of t-BOOH decreased the ARPE-19 cell death induced by a higher dose of t-BOOH by 13%. This protective effect was absent when ARPE-19 cells were pretreated with SB203580 for 1 hour before preconditioning.


CONCLUSIONS
Under the conditions tested, activation of p38 MAPK protects ARPE-19 cells against oxidant-induced death. Because RPE cells play a key role in retinal homeostasis, this finding may have important implications for retinal disease.
Pocrnich
Cady E
CE
Department of Ophthalmology, University of Western Ontario, London, Ontario, Canada.
Liu
Hong
H
Feng
Mary
M
Peng
Tianqing
T
Feng
Qingping
Q
Hutnik
Cindy M L
CM
D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection / Q000187:drug effects; D004247:DNA; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D018384:Oxidative Stress / Q000187:drug effects*; D010766:Phosphorylation; D010957:Plasmids / Q000235:genetics; D034741:RNA, Small Interfering / Q000494:pharmacology; D055213:Retinal Pigment Epithelium / Q000187:drug effects* / Q000201:enzymology* / Q000473:pathology; D013997:Time Factors; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D020122:tert-Butylhydroperoxide / Q000633:toxicity*
D004791:Enzyme Inhibitors; D034741:RNA, Small Interfering; D004247:DNA; D020122:tert-Butylhydroperoxide; D048051:p38 Mitogen-Activated Protein Kinases