Nitric oxide production by the vacuolar-type (H+)-ATPase inhibitors bafilomycin A1 and concanamycin A and its possible role in apoptosis in RAW 264.7 cells.
In the mouse leukemic monocyte cell line RAW 264.7, the vacuolar-type (H(+))-ATPase (V-ATPase) inhibitors bafilomycin A1 and concanamycin A induced nitric oxide (NO) production through the expression of inducible nitric-oxide synthase mRNA and its protein and decreased cell growth and survival as determined by 3-(4,5-dimethyl(thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Bafilomycin A1 and concanamycin A activated nuclear factor (NF)-kappaB and activator protein-1 and decreased the level of IkappaB-alpha and increased that of phosphorylated c-Jun N-terminal kinase (JNK). NO production induced by these V-ATPase inhibitors was suppressed by the NF-kappaB inhibitor Bay 11-7082 [(E)3-[(4-methylphenyl)sulfonyl])-2-propenenitrile] and the JNK inhibitor SP600125 [anthra[1,9-cd]pyrazol-6(2H)-one] in parallel with the partial alleviation of the V-ATPase inhibitor-induced decrease in MTT response. The Na(+),K(+)-ATPase inhibitor dibucaine and the F-ATPase inhibitor oligomycin did not induce NO production at which concentrations the MTT response was decreased. The NO donor S-nitroso-N-acetyl-dl-penicillamine further lowered the V-ATPase inhibitor-induced decrease in the MTT response, and the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, sodium salt (carboxy-PTIO) alleviated it partially. Mitochondrial depolarization, an index of apoptosis, was induced by bafilomycin A1 and concanamycin A. On treatment with the nitric-oxide synthase inhibitor N(G)-monomethyl-l-arginine acetate, the disruption of mitochondrial membrane potential induced by bafilomycin A1 and concanamycin A was alleviated partially in parallel with the decrease in NO production. Carboxy-PTIO also alleviated it partially. Our findings suggest that the V-ATPase inhibitors bafilomycin A1 and concanamycin A similarly induce NO production and the newly produced NO participates partially in the V-ATPase inhibitor-induced apoptosis in RAW 264.7 cells.
Hong
Jangja
J
Laboratory of Pathophysiological Biochemistry, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba Aramaki, Aoba-ku, Sendai, Miyagi 980-8578, Japan.
Nakano
Yasuhiro
Y
Yokomakura
Aya
A
Ishihara
Kenji
K
Kim
Soonjoo
S
Kang
Young-Sook
YS
Ohuchi
Kazuo
K
D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D001565:Benzoates / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003992:Dibucaine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007093:Imidazoles / Q000494:pharmacology; D018942:Macrolides / Q000494:pharmacology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II / Q000032:analysis / Q000235:genetics; D009570:Nitriles / Q000494:pharmacology; D009840:Oligomycins / Q000494:pharmacology; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D010766:Phosphorylation; D012333:RNA, Messenger / Q000032:analysis; D013450:Sulfones / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000037:antagonists & inhibitors*; D019323:omega-N-Methylarginine / Q000494:pharmacology
C434003:3-(4-methylphenylsulfonyl)-2-propenenitrile; D000873:Anthracenes; D001565:Benzoates; D004791:Enzyme Inhibitors; D007093:Imidazoles; D018942:Macrolides; D016328:NF-kappa B; D009570:Nitriles; D009840:Oligomycins; D012333:RNA, Messenger; C110881:S-nitro-N-acetylpenicillamine; D013450:Sulfones; D018808:Transcription Factor AP-1; C079393:1,3-dihydroxy-4,4,5,5-tetramethyl-2-(4-carboxyphenyl)tetrahydroimidazole; C432165:pyrazolanthrone; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D019284:Thapsigargin; C036978:concanamycin A; C040929:bafilomycin A1; D052247:Nitric Oxide Synthase Type II; D025262:Vacuolar Proton-Translocating ATPases; D010396:Penicillamine; D003992:Dibucaine