Ribozyme modulation of lipopolysaccharide-induced tumor necrosis factor-alpha production by peritoneal cells in vitro and in vivo.
We have utilized synthetic ribozymes to modulate the lipopolysaccharide (LPS)-induced production of tumor necrosis factor-alpha (TNF-alpha) by peritoneal cells. Two hammerhead ribozymes (mRz1 and mRz2) were prepared by transcription in vitro and their activities in vitro and in vivo were investigated. Both ribozymes cleaved their RNA target with an apparent turnover number (kcat) of 2 min(-1), and inhibited TNF-alpha gene expression in vitro by 50% and 70%, respectively. When mRz1 and mRz2, entrapped in liposomes, were delivered into mice by intraperitoneal injection, they inhibited LPS-induced TNF-alpha gene expression in vivo with mRz2 being the most effective. This enhanced activity could result from the facilitation of catalysis by cellular endogenous proteins, since they specifically bind to mRz2 as compared to mRz1. Furthermore, a significant mRz2 activity can be recovered from peritoneal cells 2 days post-administration in vivo. The anti-TNF-alpha ribozyme treatment in vivo resulted in a more significant reduction of LPS-induced IFN-gamma protein secretion compared to IL-10. In contrast to this pleiotropic effect, the anti-TNF-alpha ribozyme treatment did not affect the heterogenous expression of Fas ligand by peritoneal cells, indicating the specificity of the treatment. Taken together, the present data indicate that the biological effects of TNF-alpha can be modulated by ribozymes. In addition, the data suggest that ribozymes can be administered in a drug-like manner, and therefore indicate their potential in clinical applications.
Sioud
M
M
Institute of Immunology and Rheumatology, National Hospital, Oslo, Norway.
D000818:Animals; D000918:Antibody Specificity; D001483:Base Sequence; D003593:Cytoplasm / Q000378:metabolism; D005786:Gene Expression Regulation / Q000276:immunology; D007274:Injections, Intraperitoneal; D007371:Interferon-gamma / Q000235:genetics; D016753:Interleukin-10 / Q000235:genetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D017737:Macrophages, Peritoneal / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016337:RNA, Catalytic / Q000008:administration & dosage / Q000302:isolation & purification / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology
D008070:Lipopolysaccharides; D016337:RNA, Catalytic; D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10; D007371:Interferon-gamma