DOWN-REGULATION OF TRANSFORMING GROWTH-FACTOR-BETA GENE-EXPRESSION BYANTISENSE OLIGODEOXYNUCLEOTIDES INCREASES RECOMBINANT INTERFERON-GAMMA-INDUCED NITRIC-OXIDE SYNTHESIS IN MURINE PERITONEAL-MACROPHAGES

Increasing evidence indicates that the production of nitric oxide (NO) by inducible NO synthase (iNOS) is tightly regulated. Transforming gr owth factor-beta (TGF-beta) is a family of multifunctional peptides se creted during macrophage activation, but several lines of evidence sug gest that TGF-beta is selectively suppressive for macrophage NO produc tion. We therefore reasoned that a strategy employing oligodeoxynucleo tides (ODN) complementary to TGF-beta mRNA (antisense ODN) might incre ase NO production in interferon-gamma (IFN-gamma) treated murine perit oneal macrophages. To evaluate this concept, we tested the effects of antisense ODN targeted to TGF-beta mRNA (25-mer ODN complementary to T CF-beta mRNA sequences) by introducing them into the medium of culture d macrophages. Phosphorothioation of ODN was employed to retard their degradation. Antisense ODN had no effect on NO production by itself, w hereas IFN-gamma alone had a modest effect. When antisense ODN were us ed in combination with IFN-gamma, there was a marked cooperative induc tion of NO production. These effects of antisense ODN were associated with decreased TGF-beta expression in activated macrophages. However, sense ODN had no effect. Adding anti-TGF-beta antibodies to the IFN-ga mma-treated macrophages mimicked the positive effect of antisense ODN on NO production. In addition, the effects of either antisense ODN or anti-TGF-beta antibodies were blocked by adding exogenous TGF-beta in cultured macrophages. These results indicate that the generation of TG F-beta by activated macrophages provides a self-regulating mechanism b y which the temporal and perhaps spatial production of NO, a reactive and potentially toxic mediator, can be finely regulated.