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Effects of stimulation of adenosine A2A receptors on lipopolysaccharide-induced production of reactive oxygen species by equine neutrophils

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  • 1 Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 2 Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 3 Departments of Physiology and Pharmacology, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 4 Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 5 Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 6 Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602
  • | 7 Department of Pharmacology, School of Medicine, Creighton University, Omaha, NE 68178

Abstract

Objective—To assess the anti-inflammatory effects of an adenosine analogue on lipopolysaccharide (LPS)-stimulated equine neutrophils.

Sample Population—Neutrophils obtained from 10 healthy horses.

Procedures—An adenosine analogue (5′-N-ethylcarboxamidoadenosine [NECA]) was tested for its ability to inhibit production of reactive oxygen species (ROS) in LPS-stimulated equine neutrophils. Selective adenosine receptor antagonists were used to identify the receptor subtype responsible for effects. To assess the mechanism of action of NECA, cAMP concentrations were measured, and effects of dibutyryl cAMP (a stable analogue of cAMP) and rolipram (a type 4 phosphodiesterase inhibitor) were investigated.

Results—NECA elicited concentration-dependent inhibition of ROS production that was inhibited by ZM241385, a selective adenosine A2A receptor antagonist; this effect of NECA was not affected by the adenosine A2B receptor antagonist MRS1706. Also, ZM241385 blocked NECA-induced increases in cAMP concentrations, whereas MRS1706 did not alter this effect of NECA. Rolipram potentiated NECA-induced inhibition of ROS production, and dibutyryl cAMP also inhibited ROS production.

Conclusions and Clinical Relevance—Activation of adenosine A2A receptors inhibited ROS production by LPS-stimulated equine neutrophils in a cAMP-dependent manner. These results suggest that stable adenosine A2A receptor agonists may be developed as suitable anti-inflammatory drugs in horses.

Abstract

Objective—To assess the anti-inflammatory effects of an adenosine analogue on lipopolysaccharide (LPS)-stimulated equine neutrophils.

Sample Population—Neutrophils obtained from 10 healthy horses.

Procedures—An adenosine analogue (5′-N-ethylcarboxamidoadenosine [NECA]) was tested for its ability to inhibit production of reactive oxygen species (ROS) in LPS-stimulated equine neutrophils. Selective adenosine receptor antagonists were used to identify the receptor subtype responsible for effects. To assess the mechanism of action of NECA, cAMP concentrations were measured, and effects of dibutyryl cAMP (a stable analogue of cAMP) and rolipram (a type 4 phosphodiesterase inhibitor) were investigated.

Results—NECA elicited concentration-dependent inhibition of ROS production that was inhibited by ZM241385, a selective adenosine A2A receptor antagonist; this effect of NECA was not affected by the adenosine A2B receptor antagonist MRS1706. Also, ZM241385 blocked NECA-induced increases in cAMP concentrations, whereas MRS1706 did not alter this effect of NECA. Rolipram potentiated NECA-induced inhibition of ROS production, and dibutyryl cAMP also inhibited ROS production.

Conclusions and Clinical Relevance—Activation of adenosine A2A receptors inhibited ROS production by LPS-stimulated equine neutrophils in a cAMP-dependent manner. These results suggest that stable adenosine A2A receptor agonists may be developed as suitable anti-inflammatory drugs in horses.

Contributor Notes

Supported by the White Fox Farm Research Fund, Pfizer Animal Health, and a grant from the USDA Small Business Innovation Research (No. 2003-33610-13038).

The authors thank Natalie Norton, Dr. Monica Figueiredo, and Dr. Shozo Okano for technical assistance.

Address correspondence to Dr. Moore.