title: Antisense inhibition of silica-induced tumor necrosis factor in alveolar macrophages.
abstract: Tumor necrosis factor-alpha (TNFalpha) has been shown to play an important role in the pathogenesis of silicotic fibrosis. In this study, antisense oligonucleotides targeted to TNFalpha mRNA were used to inhibit silica-induced TNFalpha gene expression in alveolar macrophages. To achieve macrophage-specific oligonucleotide delivery, a molecular conjugate consisting of mannosylated polylysine that exploits endocytosis via the macrophage mannose receptor was used. Complexes were formed between the mannosylated polylysine and oligonucleotides and added to the cells in the presence of silica. Enzyme-linked immunoadsorbent assay showed that the complex consisting of the conjugate and antisense oligomer effectively inhibited TNFalpha production, whereas the oligomer alone had much less effect. Reverse transcriptase-polymerase chain reaction analysis revealed that the reduction in TNFalpha secretion was associated with specific ablation of targeted TNFalpha mRNA. The conjugate alone or conjugate complexed with inverted or sense sequence oligonucleotide had no effect. The promoting effect of the conjugate on antisense activity was shown to be due to enhanced cellular uptake of the oligomer via mannose receptor-mediated endocytosis. Cells lacking mannose receptors showed no susceptibility to the conjugate treatment. These results indicate that effective and selective inhibition of macrophage TNFalpha expression can be achieved using the antisense mannosylated polylysine system.
authors.affiliation: Schools of Pharmacy and Medicine, West Virginia University, Health Sciences Center, Morgantown, West Virginia 26506, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Rojanasakul
authors.forename: D N
authors.initials: DN
authors.lastname: Weissman
authors.forename: X
authors.initials: X
authors.lastname: Shi
authors.forename: V
authors.initials: V
authors.lastname: Castranova
authors.forename: J K
authors.initials: JK
authors.lastname: Ma
authors.forename: W
authors.initials: W
authors.lastname: Liang
mesh_terms: D000818:Animals; D002460:Cell Line; D016676:Macrophages, Alveolar / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008358:Mannose / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D011107:Polylysine / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012822:Silicon Dioxide / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000096:biosynthesis
chemical_list: D016376:Oligonucleotides, Antisense; D014409:Tumor Necrosis Factor-alpha; D011107:Polylysine; D012822:Silicon Dioxide; D008358:Mannose