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Increased ADMA/protein methylation modulates nitric oxide generation in hypoxic human lung microvascular endothelial cells
Author(s) -
Han Zhaosheng,
Pope Arthur J.,
Cardounel Arturo J.,
Nelin Leif D,
Chicione Louis G.
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.1024.6
Subject(s) - asymmetric dimethylarginine , enos , nitric oxide , hypoxia (environmental) , medicine , endocrinology , western blot , chemistry , downregulation and upregulation , nitric oxide synthase , methylation , nitric oxide synthase type iii , protein methylation , endogeny , arginine , biology , biochemistry , oxygen , gene , amino acid , organic chemistry , methyltransferase
Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) synthase (NOS) and is elevated in serum of animals exposed to chronic hypoxia. We tested the hypothesis that exposing endothelial cells (EC) to hypoxia would increase ADMA levels and thereby decrease NO production. Human lung microvascular EC were exposed to normoxia or hypoxia (1% O 2 ) for 48 hours, NO generation and ADMA were measured using EPR and HPLC, respectively. eNOS, protein arginine methyltransferases2 (PRMT2), dimethylarginine dimethylaminohydrolases I and II (DDAH) were assayed by Western blot. Assessment of protein methylation was performed by immunoprecipatation/immunoblotting. We found that NO production was reduced by 30% following 48h of hypoxia compared to normoxia. The protein levels of eNOS were similar between normoxic and hypoxic EC. Furthermore, we found that ADMA, methylated protein, and PRMT2 were increased by 112%, 105%, and 113%, respectively in hypoxic compared to normoxic EC. DDAH1 protein levels were decreased by 50% in hypoxia, while DDAH2 protein was undetectable. Thus, hypoxia caused an increase in PRMT2 and reduced DDAH1 leading to an increase in ADMA levels and an associated decrease in NO production. Therefore, we conclude that the increased ADMA/protein methylation due to enhanced PRMT2 and diminished DDAH protein may play an important role in hypoxia induced reduction of NO production.