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Hydrogen peroxide mediates hypoxic induction of arginase II in human microvascular pulmonary endothelial cells
Author(s) -
Chicoine Louis G,
Nelin Leif D,
Han Zhaosheng
Publication year - 2012
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.26.1_supplement.1130.11
Subject(s) - peroxynitrite , chemistry , catalase , nadph oxidase , reactive oxygen species , hypoxia (environmental) , superoxide , microbiology and biotechnology , arginase , oxidase test , western blot , nitric oxide , biochemistry , oxidative stress , oxygen , enzyme , biology , arginine , organic chemistry , amino acid , gene
Hypoxia induced up‐regulation of arginase II (Arg II) in endothelial cells (ECs) has been recently reported, however, the underlying mechanism(s) is largely unknown. We hypothesized that reactive oxygen species (ROS), in particular H 2 O 2 , may be involved. Human lung microvascular ECs were subjected to hypoxia (1% O 2 , 5% CO 2 , bal N 2 ) for 24–48h and harvested for immune‐blot analysis, which showed a one‐ and three‐fold increase in ECs Arg II expression, respectively. The experiment was repeated in the presence of each of the following: L‐NAME (1mM), N‐Acetyl‐Cysteine (NAC, 1mM), PEG‐SOD (100 μ/ml), PEG‐Catalase (200 μ/ml) and uric acid (UA, 100μM). We observed that Arg II expression was abolished in the presence of NAC and not altered by L‐NAME indicating that ROS played a role in hypoxia induced Arg II up‐regulation. Treatment with PEG‐SOD demonstrated enhanced Arg II expression, while PEG‐Catalase prevented hypoxic up‐regulation of Arg II. UA did not attenuate hypoxia induced up‐regulation of Arg II indicating that H 2 O 2 , but not superoxide (O 2 − ) nor peroxynitrite (ONOO − ), played a role in hypoxia induced Arg II up‐regulation. ECs treated directly with H 2 O 2 (1 and 50 μm) for 48h showed that Arg II expression was increased in a dose‐dependent manner. We investigated the cellular source of H 2 O 2 utilizing NADPH oxidase inhibition with diphenylene iodonium, 50μM, as well as with mitochondrial complex inhibition with rotenone 2μM and with antimycin A 10μM. These inhibitors had no effect on hypoxia‐induced Arg II over‐expression. Finally, xanthine oxidase inhibition with oxypurinol, 20μM, showed marked decreases in hypoxia‐induced Arg II over expression. Taken together, these data support our conclusion that xanthine oxidase derived H 2 O 2 mediate, at least in part, Arg II up‐regulation in human lung microvascular ECs exposed to hypoxia.