Cyclic adenosine monophosphate-stimulating agents induce ecto-5′-nucleotidase activity and inhibit DNA synthesis in rat cultured mesangial cells

Abstract

The ecto-5′-nucleotidase activity of rat glomerular mesangial cells increases after exposure to prostaglandin E 2 (PGE 2) via cAMP stimulation (Savic et al., 1990, Immunology 70, 321). Therefore we examined whether other cAMP-stimulating agents had a similar effect. Forskolin (1 μ m), PGE 2 (10 μ m), and isoproterenol (10 μ m), three products stimulating rat mesangial cell adenylate cyclase activity, enhanced cAMP accumulation within 5 min and 5′-nucleotidase activity after a lag time of at least 24 h. 3-Isobutyl-1-methylxanthine (IBMX) and Ro 20–1724, two drugs inhibiting cAMP degradation, also stimulated cAMP accumulation and 5′-nucleotidase activity. The effects of these agents on 5′-nucleotidase activity were additive with those of the three products stimulating adenylate cyclase activity, except for Ro 20–1724 and forskolin which acted synergistically. Cycloheximide, a blocker of protein synthesis, suppressed the cAMP-dependent increase of 5′-nucleotidase activity. Because ecto-5′-nucleotidase activity is a marker of cell differentiation, the effect of the same cAMP-stimulating agents on cell proliferation was also studied. Forskolin, PGE 2, and isoproterenol inhibited [ 3H]thymidine incorporation into rat mesangial cells in a dose-dependent manner. The same effect was obtained with IBMX (100 μ m) and Ro 20–1724 (50 μ m). Stimulation of 5′-nucleotidase activity and inhibition of [ 3H]thymidine incorporation occurred over the same range of concentrations for the various agonists tested. Taken together, these results indicate that expression of ecto-5′-nucleotidase in rat mesangial cells is induced by cAMP whatever the reason for its accumulation. The simultaneous inhibition of DNA synthesis may occur independently or be associated with the stimulation of 5′-nucleotidase expression.