The effect of antisense oligodeoxynucleotides on nitric oxide secretion from macrophage-like cells.

ABSTRACT

Nitric oxide (NO) plays an important role in cellular signaling and host defense, and it also contributes to the deleterious effects of immune response. Until recently, the lack of specific inhibitors of various forms of nitric oxide synthase (NOS) hampered a stringent evaluation of the role played by inducible NOS (iNOS) in cell damage. The present study investigated the use of antisense oligodeoxynucleotides (AS-ODNs) to selectively inhibit the expression of iNOS. AS-ODNs (1-10 microM) inhibited, in a time-dependent and dose-dependent manner, iNOS activity in RAW 264.7 murine macrophages. Maximal inhibitory effect was >90%, and control ODNs had little or no effect on NO production. Treatment with AS-ODNs decreased iNOS protein and mRNA level in studied cell, and control ODNs again were ineffective. The decreased levels of the target mRNA in AS-ODN-treated samples suggest that the AS-ODNs used act as substrates for ribonuclease (RNase) H. Lipofection enhanced the effect of AS-ODNs on iNOS activity. However, this potentiation appears to be different from the antisense effect, in which the AS-ODNs studied were involved. Liposaccharide/interferon-gamma (LPS/IFN-gamma) induced iNOS, and increased NO production impaired the viability of macrophages. Treatment of RAW 264.7 cells with 10 microM AS-ODNs prevented the NO-induced lethal cell damage.