Extracellular 2′,3′-Cyclic Adenosine Monophosphate Is a Potent Inhibitor of Preglomerular Vascular Smooth Muscle Cell and Mesangial Cell Growth

Abstract

Recently we discovered that intact kidneys release into the extracellular compartment 2′,3′-cAMP (a positional isomer of 3′,5′-cAMP with unknown pharmacology) and metabolize 2′,3′-cAMP to 2′-AMP, 3′-AMP, and adenosine. Because adenosine inhibits growth of vascular smooth muscle cells and mesangial cells, we tested the hypothesis that extracellular 2′,3′-cAMP attenuates growth of preglomerular vascular smooth muscle and mesangial cells via production of adenosine. For comparison, all of the experiments were performed with both 2′,3′-cAMP and 3′,5′-cAMP. In study 1, 2′,3′-cAMP, 3′,5′-cAMP, 5′-AMP, 3′-AMP, or 2′-AMP was incubated with cells and purines measured in the medium by mass spectrometry. Both preglomerular vascular smooth muscle and mesangial cells metabolized 3′,5′-cAMP to 5′-AMP and adenosine; 5′-AMP to adenosine; 2′,3′-cAMP to 2′-AMP, 3′-AMP, and adenosine; and 2′-AMP and 3′-AMP to adenosine. 3-Isobutyl-1-methylxanthine (phosphodiesterase inhibitor) and 1,3-dipropyl-8-p-sulfophenylxanthine (ecto-phosphodiesterase inhibitor) blocked conversion of 3′,5′-cAMP to 5′-AMP and adenosine, and α,β-methylene-adenosine-5′-diphosphate (CD73 inhibitor) blocked conversion of 5′-AMP to adenosine. These enzyme inhibitors had little effect on metabolism of 2′,3′-cAMP, 2′-AMP, or 3′-AMP. For study 2, 2′,3′-cAMP and 3′,5′-cAMP profoundly inhibited proliferation (thymidine incorporation and cell number) of both cell types, with 2′,3′-cAMP more potent than 3′,5′-cAMP. Antagonism of A 2B receptors (MRS-1754), but not A 1 (1,3-dipropyl-8-cyclopentylxanthine), A 2A (SCH-58261), or A 3 (VUF-5574) receptors, attenuated the growth inhibitory effects of 2′,3′-cAMP and 3′,5′-cAMP. Extracellular 2′,3′-cAMP inhibits growth of preglomerular vascular smooth muscle and mesangial cells more profoundly than does 3′,5′-cAMP. Although both cAMPs inhibit growth in part via conversion to adenosine followed by A 2B receptor activation, their metabolism is mediated by different enzymes.