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Induction of nitric oxide synthase in rat peritoneal neutrophils and its inhibition by dexamethasone
T B McCall1, R M Palmer, S Moncada
1Wellcome Research Laboratories, Beckenham, Kent, Great Britain.
Abstract:
Rat peritoneal polymorphonuclear leukocytes (PMN) elicited with oyster glycogen contain a Ca(2+)-independent nitric oxide (NO) synthase which is induced in vivo in a time-dependent manner. When washed PMN containing low levels of enzyme activity were cultured ex vivo further expression of NO synthase was observed. This was inhibited by cycloheximide indicating that de novo synthesis of the enzyme occurred during the ex vivo incubation. Enzyme activity was enhanced by interferon (IFN)-gamma, but not by tumor necrosis factor (TNF)-alpha when added ex vivo. However, IFN-gamma and TNF-alpha synergized to increase further the expression of NO synthase. Treatment of rats with dexamethasone inhibited the induction of NO synthase in elicited PMN. This treatment reduced the accumulation of PMN by approximately 30%, without affecting cell viability. Dexamethasone also inhibited the induction of the NO synthase ex vivo in a concentration-dependent manner. Furthermore, the enhanced enzyme activity following treatment of PMN with cytokines was also inhibited by dexamethasone. Once induced, dexamethasone did not affect enzyme activity. These data indicate that PMN elicited in the rat peritoneum with oyster glycogen express an NO synthase in vivo and ex vivo. The induction of the enzyme can be further stimulated ex vivo with IFN-gamma and TNF-alpha and inhibited by dexamethasone. The inhibition of the induction of NO synthase in the PMN by dexamethasone may contribute to the anti-inflammatory activity of this and other glucocorticoids.
Insights
Rat peritoneal cells produce nitric oxide (NO) synthase, an enzyme inducible in vivo and ex vivo. Interferon-gamma and tumor necrosis factor-alpha enhance its expression, while dexamethasone inhibits NO synthase induction, suggesting anti-inflammatory effects.
Area of Science:
- Immunology
- Molecular Biology
- Biochemistry
Background:
- Rat peritoneal polymorphonuclear leukocytes (PMN) possess Ca(2+)-independent nitric oxide (NO) synthase.
- This enzyme is induced in vivo in response to oyster glycogen.
- PMN cultured ex vivo show further NO synthase expression, indicating de novo synthesis.
Purpose of the Study:
- To investigate the regulation of NO synthase induction in rat peritoneal PMN.
- To explore the effects of cytokines (IFN-gamma, TNF-alpha) and dexamethasone on NO synthase expression.
- To determine the potential anti-inflammatory role of dexamethasone-mediated inhibition of NO synthase.
Main Methods:
- Elicitation of rat peritoneal PMN with oyster glycogen.
- Culture of PMN ex vivo with or without cytokines and dexamethasone.
- Measurement of NO synthase activity.
- Assessment of PMN accumulation and viability.
Main Results:
- NO synthase induction occurred both in vivo and ex vivo in elicited PMN.
- Interferon-gamma and tumor necrosis factor-alpha synergistically enhanced NO synthase expression ex vivo.
- Dexamethasone inhibited NO synthase induction in a concentration-dependent manner, both in vivo and ex vivo, and reduced PMN accumulation.
- Dexamethasone did not affect already induced enzyme activity.
Conclusions:
- Rat peritoneal PMN express an inducible NO synthase.
- Cytokines like IFN-gamma and TNF-alpha stimulate NO synthase induction, while dexamethasone inhibits it.
- Dexamethasone's inhibition of NO synthase induction in PMN may contribute to its anti-inflammatory properties.