Interferon-gamma enhances superoxide production in human mesangial cells via the JAK-STAT pathway.
Immune reactive cytokines, such as interferon (IFN)-gamma, have multiple effects in glomerulonephritis. Superoxide anions (O(2)(-)), which are associated with the progression of glomerulonephritis, are mainly generated by nicotinamide adenine dinucleotide phosphate (reduced form) NAD(P)H oxidases. We determined the effects of IFN-gamma on O(2)(-) production, phosphorylation of signal transducer and activator of transcription (STAT)-1alpha, and the mRNA and protein expressions of p22phox and Nox1, components of NAD(P)H oxidases, in human mesangial cells (HMCs). Significant increases in O(2)(-) production with IFN-gamma were completely abolished by the flavin-containing enzyme inhibitor, diphenyleneiodonium (10 micromol/l), and the Janus-activated kinase (JAK)2 inhibitor, AG490 (100 micromol/l). Phosphorylated STAT-1alpha was detected after 5 min of IFN-gamma stimulation using Western blot analysis, and binding to the gamma-activating site was observed from 30 min to 4 h, thereafter by electrophoretic mobility shift assay (EMSA). Super-shift analysis in EMSA revealed that the main transcription factor was STAT-1alpha. IFN-gamma significantly increased the expression of p22phox mRNA and protein, although expression was inhibited by AG490. These data suggest that IFN-gamma stimulates O(2)(-) production in HMCs via the JAK-STAT pathway and NAD(P)H oxidase.
Moriwaki
K
K
Department of CardioRenal and Cerebrovascular Medicine, Faculty of Medicine, Kagawa University, Kagawa, Japan.
Kiyomoto
H
H
Hitomi
H
H
Ihara
G
G
Kaifu
K
K
Matsubara
K
K
Hara
T
T
Kondo
N
N
Ohmori
K
K
Nishiyama
A
A
Fukui
T
T
Kohno
M
M
D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D053613:Janus Kinase 1; D053614:Janus Kinase 2; D050527:Mesangial Cells / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D009249:NADP / Q000235:genetics / Q000378:metabolism; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D050791:STAT Transcription Factors / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D013481:Superoxides / Q000378:metabolism*; D020032:Tyrphostins / Q000494:pharmacology
D004791:Enzyme Inhibitors; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D050791:STAT Transcription Factors; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; D013481:Superoxides; D009249:NADP; D007371:Interferon-gamma; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C402799:NOX1 protein, human; C086986:CYBA protein, human; D011505:Protein-Tyrosine Kinases; C507904:JAK1 protein, human; C507924:JAK2 protein, human; D053613:Janus Kinase 1; D053614:Janus Kinase 2; D010100:Oxygen