Interaction of nitric oxide and cyclic guanosine 3′,5′-monophosphate in erythropoietin production

Abstract

The present study was designed to investigate whether in vivo and in vitro erythropoietin (EPO) production is modulated by nitric oxide (NO) and cyclic guanosine 3′,5′-monophosphate (cGMP). Serum levels of EPO in exhypoxic polycythemic mice were significantly increased after injections of 200 μg/kg sodium nitroprusside for 4 d. One injection of NG-nitro-L-arginine methyl ester (L-NAME) produced a significant dose-related decrease in serum levels of EPO in exhypoxic polycythemic mice in response to hypoxia. When EPO producing Hep3B cells were incubated in 1 % O2 for 30 min, cGMP levels in the Hep3B cells were significantly elevated, compared with cells incubated in 20% O2. The elevation of cGMP by hypoxia was inhibited by L-NAME (100 μM). Sodium nitropruside (10 and 100 μM) and NO (2 μM) also significantly increased cGMP levels in Hep3B cells. L-NAME, LY 83583 (6-Anilino-5,8-quinolinedione, a soluble guanylate cyclase inhibitor), and Rp-8-Bromo-cGMPS (Rp-8-Bromo-guanosine 3′,5′-cyclic monophosphothioate, a cGMP-dependent protein kinase inhibitor) significantly inhibited the hypoxia-induced increase in medium levels of EPO in Hep3B cells. 8-Bromo-cGMPS produced a dose-dependent decrease in EPO messenger RNA levels in Hep3B cells in response to hypoxia. 8-Bromo-cGMP (10-3M) produced significant increases in medium levels of EPO in Hep3B cell cultures incubated under normoxic conditions, which was enhanced by the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (0.2 mM). These results suggest that NO and cGMP may interact in modulating hypoxic stimulation of EPO production.