title: Specific inhibition of macrophage TNF-alpha expression by in vivo ribozyme treatment.
abstract: The overproduction of the cytokine TNF-alpha is associated with inflammatory and autoimmune diseases. We have developed a means to block TNF-alpha production with ribozymes directed against TNF-alpha mRNA to selectively inhibit its production in vitro and in vivo. Following cationic lipid-mediated delivery to peritoneal murine macrophages in culture, anti-TNF-alpha ribozymes were more effective inhibitors of TNF-alpha secretion than catalytically inactive ribozyme controls. Inhibition of TNF-alpha secretion was proportional to the concentration of ribozyme administered, with an IC50 of approximately 10 nM. After i.p. injection of cationic lipid/ribozyme complexes, elicited macrophages accumulated approximately 6% of the administered ribozyme. The catalytically active ribozyme suppressed LPS-stimulated TNF-alpha secretion by approximately 50% relative to an inactive ribozyme control without inhibiting secretion of another proinflammatory cytokine produced by macrophages, IL-1alpha. Ribozyme-specific TNF-alpha mRNA degradation products were found among the mRNA extracted from macrophages following in vivo ribozyme treatment and ex vivo stimulation. Thus, catalytic ribozymes can accumulate in appropriate target cells in vivo; once in the target cell, ribozymes can be potent inhibitors of specific gene expression.
authors.affiliation: Department of Cell Biology, School of Medicine, University of California, Davis 95616, USA.
authors.forename: K O
authors.initials: KO
authors.lastname: Kisich
authors.forename: R W
authors.initials: RW
authors.lastname: Malone
authors.forename: P A
authors.initials: PA
authors.lastname: Feldstein
authors.forename: K L
authors.initials: KL
authors.lastname: Erickson
mesh_terms: D000818:Animals; D001202:Ascitic Fluid / Q000166:cytology / Q000276:immunology; D001483:Base Sequence; D002412:Cations; D002478:Cells, Cultured; D005260:Female; D006868:Hydrolysis; D007274:Injections, Intraperitoneal; D007700:Kinetics; D008055:Lipids / Q000008:administration & dosage / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D010537:Peritoneum / Q000166:cytology / Q000201:enzymology; D010714:Phosphatidylethanolamines / Q000008:administration & dosage / Q000494:pharmacology; D000644:Quaternary Ammonium Compounds / Q000008:administration & dosage / Q000494:pharmacology; D016337:RNA, Catalytic / Q000008:administration & dosage* / Q000378:metabolism / Q000494:pharmacology; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000378:metabolism; D013096:Spermine / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
chemical_list: C098223:2,3-dioleyloxy-N-(2-(sperminecarboxamido)ethyl)-N,N-dimethyl-1-propanaminium trifluoroacetate; D002412:Cations; D008055:Lipids; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D016337:RNA, Catalytic; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; C054005:N-(1-(2,3-dioleyloxy)propyl)-N,N,N-trimethylammonium; C087276:(3-dimyristyloxypropyl)(dimethyl)(hydroxyethyl)ammonium; C094877:dioleoyl phosphatidylethanolamine; D013096:Spermine