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Glutamine is essential for nitric oxide synthesis by murine macrophages.
J Surg Res. 1999 Oct; 86(2):213-9.JS

Abstract

BACKGROUND

Recent studies suggest an interaction between l-arginine (Arg) and l-glutamine (Gln) in the control of nitric oxide (NO) synthesis. Endotoxemia enhances Gln demand and NO production. This study was initiated to investigate the effects of altered Gln availability on the capacity of macrophages to produce NO and the interaction of Gln with l-citrulline (Cit) and Arg in the regulation of endotoxin-stimulated NO synthesis.

METHODS

Cultures of RAW 264.7 macrophages in MEM containing Gln (0 to 100 mM) or Arg (0 or 0.6 mM) and supplemented or not with Cit (0.31 to 10 mM) were exposed to Escherichia coli lipopolysaccharide (LPS) at 0.001 and 1 microg/ml. After 24-h incubation, supernatants were evaluated for nitrite concentrations by Greiss reaction as a measure of NO synthesis.

RESULTS

LPS stimulated nitrite synthesis in a dose-dependent fashion. Macrophages cultured in Gln-free medium containing Arg (0.6 mM) did not produce NO when stimulated with LPS. In contrast, in the presence of Arg and 0.001 microg/ml LPS, adding as little as 0.31 mM Gln resulted in a 23-fold increase in NO production (from 0.13 +/- 0. 02 to 2.92 +/- 0.06 nmol/ml) (P < 0.0001). Furthermore, a dose-dependent increase in LPS-stimulated nitrite release was observed with increasing amounts of Gln to as much as 1 mM. LPS-stimulated macrophages cultured in Arg-free medium containing Gln (0.31-10 mM) did not produce significant amounts of nitrite. However, in the absence of Arg, increasing extracellular Gln levels to 100 mM in the culture medium resulted in nitrite synthesis (2.39 +/- 0.11 nmol/ml). Detectable levels of nitrite (2.84 +/- 0.21 nmol/ml) were also documented when stimulated macrophages were incubated in culture medium lacking Arg but containing Cit (0.31 mM) and Gln (2 mM). Increasing Cit levels (0.63 to 10 mM) significantly augmented nitrite release (P < 0.05). Once again, no detectable levels of nitrite were observed when macrophages were cultured in Gln-free medium, even when Arg and Cit were present.

CONCLUSION

These results suggest that Gln is an essential amino acid for NO synthesis by macrophages and raise the strong possibility that Gln acts with nitric oxide synthase to catalyze the conversion of Arg to NO. The consumption of Gln during sepsis may represent NO production.

Authors+Show Affiliations

Department of Surgery, Tulane University School of Medicine, New Orleans, Louisiana, 70112, USA.No affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

10534426

Citation

Bellows, C F., and B M. Jaffe. "Glutamine Is Essential for Nitric Oxide Synthesis By Murine Macrophages." The Journal of Surgical Research, vol. 86, no. 2, 1999, pp. 213-9.
Bellows CF, Jaffe BM. Glutamine is essential for nitric oxide synthesis by murine macrophages. J Surg Res. 1999;86(2):213-9.
Bellows, C. F., & Jaffe, B. M. (1999). Glutamine is essential for nitric oxide synthesis by murine macrophages. The Journal of Surgical Research, 86(2), 213-9.
Bellows CF, Jaffe BM. Glutamine Is Essential for Nitric Oxide Synthesis By Murine Macrophages. J Surg Res. 1999;86(2):213-9. PubMed PMID: 10534426.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Glutamine is essential for nitric oxide synthesis by murine macrophages. AU - Bellows,C F, AU - Jaffe,B M, PY - 1999/10/27/pubmed PY - 1999/10/27/medline PY - 1999/10/27/entrez SP - 213 EP - 9 JF - The Journal of surgical research JO - J Surg Res VL - 86 IS - 2 N2 - BACKGROUND: Recent studies suggest an interaction between l-arginine (Arg) and l-glutamine (Gln) in the control of nitric oxide (NO) synthesis. Endotoxemia enhances Gln demand and NO production. This study was initiated to investigate the effects of altered Gln availability on the capacity of macrophages to produce NO and the interaction of Gln with l-citrulline (Cit) and Arg in the regulation of endotoxin-stimulated NO synthesis. METHODS: Cultures of RAW 264.7 macrophages in MEM containing Gln (0 to 100 mM) or Arg (0 or 0.6 mM) and supplemented or not with Cit (0.31 to 10 mM) were exposed to Escherichia coli lipopolysaccharide (LPS) at 0.001 and 1 microg/ml. After 24-h incubation, supernatants were evaluated for nitrite concentrations by Greiss reaction as a measure of NO synthesis. RESULTS: LPS stimulated nitrite synthesis in a dose-dependent fashion. Macrophages cultured in Gln-free medium containing Arg (0.6 mM) did not produce NO when stimulated with LPS. In contrast, in the presence of Arg and 0.001 microg/ml LPS, adding as little as 0.31 mM Gln resulted in a 23-fold increase in NO production (from 0.13 +/- 0. 02 to 2.92 +/- 0.06 nmol/ml) (P < 0.0001). Furthermore, a dose-dependent increase in LPS-stimulated nitrite release was observed with increasing amounts of Gln to as much as 1 mM. LPS-stimulated macrophages cultured in Arg-free medium containing Gln (0.31-10 mM) did not produce significant amounts of nitrite. However, in the absence of Arg, increasing extracellular Gln levels to 100 mM in the culture medium resulted in nitrite synthesis (2.39 +/- 0.11 nmol/ml). Detectable levels of nitrite (2.84 +/- 0.21 nmol/ml) were also documented when stimulated macrophages were incubated in culture medium lacking Arg but containing Cit (0.31 mM) and Gln (2 mM). Increasing Cit levels (0.63 to 10 mM) significantly augmented nitrite release (P < 0.05). Once again, no detectable levels of nitrite were observed when macrophages were cultured in Gln-free medium, even when Arg and Cit were present. CONCLUSION: These results suggest that Gln is an essential amino acid for NO synthesis by macrophages and raise the strong possibility that Gln acts with nitric oxide synthase to catalyze the conversion of Arg to NO. The consumption of Gln during sepsis may represent NO production. SN - 0022-4804 UR - https://www.unboundmedicine.com/medline/citation/10534426/Glutamine_is_essential_for_nitric_oxide_synthesis_by_murine_macrophages_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-4804(99)95713-X DB - PRIME DP - Unbound Medicine ER -