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Recombinant arginine deiminase reduces inducible nitric oxide synthase iNOS‐mediated neurotoxicity in a coculture of neurons and microglia
Author(s) -
Yu HaoHsin,
Wu FeLin Lin,
Lin ShanErh,
Shen LiJiuan
Publication year - 2008
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.21740
Subject(s) - microglia , nitric oxide , nitric oxide synthase , neuroprotection , viability assay , arginine , citrulline , lipopolysaccharide , chemistry , immunostaining , microbiology and biotechnology , biochemistry , inflammation , biology , pharmacology , immunology , in vitro , endocrinology , immunohistochemistry , amino acid
Modulation of nitric oxide (NO) production is considered a promising approach to therapy of diseases involving excessive inducible nitric oxide synthase (iNOS) expression, such as certain neuronal diseases. Recombinant arginine deiminase (rADI, EC3.5.3.6) catalyzes the conversion of L ‐arginine ( L ‐arg), the sole substrate of NOS for NO production, to L ‐citrulline ( L ‐cit) and ammonia. To understand the effect of the depletion of L ‐arg by rADI on NO concentration and neuroprotection, a direct coculture of neuron SHSY5Y cells and microglia BV2 cells treated with lipopolysaccharide (LPS) and interferon‐γ (IFN‐γ) was used as a model of iNOS induction. The results showed that rADI preserved cell viability (4‐fold higher compared with the cells treated with LPS/IFN‐γ only) by the MTT assay, corresponding with the results of neuronal viability by neuron‐specific immunostaining assay. NO production (mean ± SD) decreased from 67.0 ± 1.3 to 19.5 ± 5.5 μM after a 2‐day treatment of rADI by the Griess assay; meanwhile, induction of iNOS protein expression by rADI was observed. In addition, rADI substantially preserved the neuronal function of dopamine uptake in the coculture. The replenishment of L ‐arg in the coculture eliminated the neuroprotective and NO‐suppressive effects of rADI in the coculture, indicating that L ‐arg played a crucial role in the effects of rADI. These results highlight the important role of L ‐arg in the neuron‐microglia coculture in excessive induction of iNOS. Regulation of L ‐arg by ADI demonstrated that rADI has a potentially therapeutic role in iNOS‐related neuronal diseases. © 2008 Wiley‐Liss, Inc.