Functional effects of NAD(P)H oxidase p22(phox) C242T mutation in human leukocytes and association with thrombotic cerebral infarction.
BACKGROUND
Previous study showed that polymorphism of the NAD(P)H oxidase p22(phox) gene is associated with atherosclerosis, although others could not confirm such association. We investigated the association between p22(phox) C242T polymorphism and thrombotic cerebral infarction and the role of this polymorphism on superoxide-production activity in human neutrophils and promyelocytic HL-60 cells as a model system.


METHODS
PCR-RFLP analysis revealed that genotype and allele frequencies of C242T polymorphism in 120 patients with thrombotic cerebral infarction and 177 control subjects. The superoxide-production activity in neutrophils was determined by cytochrome c reduction assay. To clarify the role of p22(phox) C242T polymorphism on NAD(P)H oxidase activity, we used transgenic HL-60 cells transfected with expression plasmids carrying p22(phox) cDNAs with or without C242T polymorphism.


RESULTS
Genotype and allele frequencies of C242T polymorphism in patients and control subjects were not significantly different. The superoxide-production activity in neutrophils with T allele was higher than in neutrophils without T allele. Moreover, expression analysis showed that superoxide-production activity in p22(phox) C242T-expressing HL-60 cells were significantly higher than in p22(phox)-expressing HL-60 cells.


CONCLUSIONS
We conclude that C242T of p22(phox) gene is not involved in thrombotic cerebral infarction but more likely in increased NAD(P)H oxidase activity in phagocytes.
Shimo-Nakanishi
Yumi
Y
Department of Neurology, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Hasebe
Takeshi
T
Suzuki
Asuka
A
Mochizuki
Hideki
H
Nomiyama
Takashi
T
Tanaka
Yasushi
Y
Nagaoka
Isao
I
Mizuno
Yoshikuni
Y
Urabe
Takao
T
D000328:Adult; D019943:Amino Acid Substitution; D015152:Blotting, Northern; D002544:Cerebral Infarction / Q000209:etiology / Q000235:genetics*; D045304:Cytochromes c / Q000378:metabolism; D005260:Female; D005787:Gene Frequency; D005838:Genotype; D018922:HL-60 Cells; D006801:Humans; D015151:Immunoblotting; D020767:Intracranial Thrombosis / Q000150:complications / Q000201:enzymology / Q000235:genetics*; D007962:Leukocytes / Q000378:metabolism*; D008297:Male; D026901:Membrane Transport Proteins / Q000235:genetics*; D008875:Middle Aged; D009154:Mutation; D009252:NADPH Dehydrogenase / Q000235:genetics*; D019255:NADPH Oxidases / Q000235:genetics*; D009504:Neutrophils / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length; D013481:Superoxides / Q000378:metabolism; D014162:Transfection
D026901:Membrane Transport Proteins; D010750:Phosphoproteins; D013481:Superoxides; D045304:Cytochromes c; D019255:NADPH Oxidases; C086986:CYBA protein, human; D009252:NADPH Dehydrogenase