The mechanism of inhibition of DNA synthesis in articular chondrocytes from young and old rats by nitric oxide.

The mechanism by which nitric oxide inhibits the incorporation of [3H]thymidine into rat articular chondrocytes (AC) in culture was studied. First-passage articular chondrocytes, isolated by collagenase digestion of cartilage fragments from humeral and femoral heads of 1- and 18-month-old rats, were used in all experiments. NO-generating compounds, isosorbide dinitrate or sodium nitoprusside, inhibited the incorporation of [3H]thymidine and the release of prostaglandin E2 (PGE2) and stimulated cyclic guanosine monophosphate (cGMP) production by rat AC monolayers in a concentration-dependent manner. The cells from old rats were much less sensitive to NO donors and also produced less PGE2 and cGMP. Blocking the production of endogenous NO with NG-monomethyl-L-arginine (L-NMA), an inhibitor of NO synthase, stimulated DNA synthesis. cGMP was found to be a key mediator of the inhibition of DNA synthesis by NO donors in rat AC. 6-Anilino-5,8-quinolinedione (LY83583), an inhibitor of NO-dependent cGMP release, stimulated [3H]-thymidine incorporation, whereas the cGMP analog, 8- bromo-cGMP, inhibited L-NMA-induced or LY83583-induced stimulation of [3H]thymidine incorporation. NO donors blocked the stimulation of DNA synthesis induced by L-NMA and only marginally blocked that of LY83583. Indomethacin had no effect on the inhibition of DNA synthesis by NO or 8-bromo-cGMP. These results show that NO donors induce inhibition of DNA synthesis probably by elevating cGMP. The relative insensitivity of senescent cells to NO donors may be due, at least in part, to their decreased capacity to produce cGMP.