Inhibition of p38-MAPK potentiates cisplatin-induced apoptosis via GSH depletion and increases intracellular drug accumulation in growth-arrested kidney tubular epithelial cells.
We were interested in analyzing the regulation by mitogen-activated protein kinases (MAPKs) of cisplatin-provoked toxicity in epithelial renal tubule cell lines, when assayed under culture conditions (cell confluence plus serum deprivation), which mimic the characteristics of a nonproliferating epithelium. Under these restrictive growth conditions, cisplatin induced apoptosis with lower efficacy than in exponentially growing cells, and decreased p38-MAPK phosphorylation in NRK-52E and other (LLC-PK1, MDCK, HK2) cell lines. Moreover, cisplatin-provoked apoptosis was potentiated by cotreatment with p38-MAPK-specific inhibitors (SB203580, SB220025) or transfection with a kinase-negative mutant of MKK6, whereas c-Jun NH2-terminal kinase or extracellular signal-regulated kinase/MAPK and ERK Kinase inhibitors were ineffective. By contrast, when applied to exponentially growing cells, cisplatin stimulated p38-MAPK phosphorylation and apoptosis, was attenuated by kinase inhibitors. Treatment of confluent/serum-deprived cells with cisplatin caused mitochondrial transmembrane potential disruption and activated the mitochondrial apoptotic pathway, as indicated by the decrease in Bcl-X(L) expression, increase in Bax expression and cytochrome c release, and these effects were potentiated by cotreatment with SB203580. Treatment of confluent/serum-deprived cells with cisplatin plus SB203580 decreased the intracellular reduced glutathione (GSH) content, and increased intracellular cisplatin accumulation as well as cisplatin binding to DNA. Cotreatment with the GSH-depleting agent D,L-buthionine-R,S-sulfoximine also potentiated cisplatin-provoked apoptosis. In summary, p38-MAPK inhibition potentiates cisplatin-provoked apoptosis in growth-arrested epithelial renal tubule cells, a result that may be explained at least in part by GSH depletion and drug transport alteration.
Rodríguez-García
Maria Elena
ME
Departamento de Bioquímica y Biología Molecular I, Universidad Complutense de Madrid, Madrid 28040, Spain.
Quiroga
Adoración G
AG
Castro
José
J
Ortiz
Alberto
A
Aller
Patricio
P
Mata
Felicísima
F
D000818:Animals; D000970:Antineoplastic Agents / Q000493:pharmacokinetics / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002460:Cell Line; D002945:Cisplatin / Q000493:pharmacokinetics / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism*; D007684:Kidney Tubules / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism
D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D048051:p38 Mitogen-Activated Protein Kinases; D005978:Glutathione; D002945:Cisplatin