Synergistic induction of nitric oxide synthase type II: in vitro effect of leptin and interferon-gamma in human chondrocytes and ATDC5 chondrogenic cells.

Overview

OBJECTIVE: To study, in vitro, the effect of leptin (OB), alone or in combination with interferon-gamma (IFNgamma), on inducible nitric oxide synthase (iNOS) and NO production in human primary chondrocytes and in mouse embryonic chondrogenic ATDC5 cells. METHODS: Leptin receptor expression and iNOS messenger RNA expression were evaluated by reverse transcriptase-polymerase chain reaction. Then, iNOS activity was indirectly studied by measuring nitrite accumulation, using the Griess colorimetric reaction, in culture medium of human primary chondrocytes and ATDC5 cells. RESULTS: ATDC5 mouse embryonic cells expressed functional OB receptor. Alone, neither OB nor IFNgamma produced nitrite accumulation in culture medium. However, costimulation with OB and IFNgamma resulted in dose-dependent up-regulation of the expression of iNOS and NO production in human primary chondrocytes and ATDC5 cells. Production of NO was blunted by the iNOS-specific inhibitors L-N(G)-nitroarginine methyl ester and aminoguanidine. In addition, the janus-activated kinase 2 (JAK2)-specific inhibitor Tyrphostin AG 490 completely blocked OB + IFNgamma-driven up-regulation of iNOS and NO production. CONCLUSION: Our data show for the first time a putative proinflammatory role of OB via iNOS induction and NO production. This occurs via activation of JAK2.