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Nitric oxide synthesis in neutrophil precursor cells and effect of nitric oxide modulators on their maturation
Author(s) -
Kumar Sachin,
Gupta Kanishka,
Barthwal Manoj Kumar,
Dikshit Madhu
Publication year - 2008
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.22.1_supplement.749.5
Subject(s) - propidium iodide , chemistry , nitric oxide , percoll , apoptosis , myeloperoxidase , bone marrow , microbiology and biotechnology , biochemistry , in vitro , immunology , biology , programmed cell death , inflammation , organic chemistry
Neutrophil maturation in bone marrow and their release in circulation have been shown to be modulated by nitric oxide (NO). Present study was undertaken to further assess the role of NO in neutrophil maturation in vitro. Neutrophil precursor cells were isolated from Sprague Dawley rat (200–250 g) bone marrow by Percoll density gradient in myeloblasts/promyelocytes rich band 3, myelocytes/metamyelocytes rich band 2 and band cells/segmented Neutrophils rich band 1, purity was confirmed by Giemsa staining/CD markers, NADPH oxidase and myeloperoxidase activity. Intracellular NO level (assessed by DAF‐2DA), total nitrite (measured by Griess reagent) content, NOS activity (using 3 H‐L‐arginine) and nNOS protein (Western blotting) were highest in band 1 and lowest in band 3. We also investigated effect of NO donor (SNP 1μM‐1mM) and NOS inhibitor (L‐NAME 1mM) on cell cycle by using Propidium iodide and BrdU staining during their culture for 24 hrs. SNP treated cells exhibited an increase in apoptosis, while viable cells showed a marginal increase in the S phase in band 2 and band 3. NOS inhibitor (L‐NAME) however increased the survival of cells without any modulation of cell cycle. Results obtained indicate that NO content augment with neutrophil maturation and seems to play an important role in the maturation of neutrophils. The work was supported by CSIR and DBT India.