title: Antibodies to rat soluble IL-6 receptor stimulate B9 hybridoma cell proliferation.
abstract: Interleukin-6 mediates its pleiotropic effects by interacting with its membrane bound receptor (gp80) or the soluble counterpart gp54, resulting in activation of a complex that includes the transducer protein gp130. We have generated a polyclonal antibody against the rat soluble IL-6 receptor (anti-rat sIL-6R) in rabbits. By Western blot analysis we show that purified anti-rat sIL-6R IgG antibody reacts specifically with recombinant rat sIL-6R generated from E. coli, baculovirus or adenovirus expression systems. Anti-rat sIL-6R inhibited IL-6-induced acute phase protein synthesis in rat (H35) but not human (HepG2) hepatoma cells, and did not affect stimulation of those cells by Oncostatin-M. Conversely, on the mouse hybridoma B9 cell line, IgG anti-rat sIL-6R showed a dose-dependent stimulation of proliferation. Fab fragments of this antibody did not stimulate, but abrogated IL-6-mediated hepatoma cell stimulation and B9 cell proliferation. Gel shift analysis of STAT nuclear factors showed activation of STAT DNA binding in nuclei of B9 cells treated with IgG anti-rat sIL-6R, whereas in H35, NIH-3T3 and M1 cells, only IL-6 could trigger a similar STAT activation. Our data suggest that mechanisms of IL-6 receptor activation and signalling in mouse B9 hybridoma cells show subtle but important differences from other IL-6-responsive cells.
authors.affiliation: Department of Pathology, Health Sciences Center, McMaster University, Hamilton, Ont., Canada.
authors.forename: V
authors.initials: V
authors.lastname: Thibault
authors.forename: C D
authors.initials: CD
authors.lastname: Richards
authors.forename: F
authors.initials: F
authors.lastname: Botelho
authors.forename: J
authors.initials: J
authors.lastname: Gauldie
mesh_terms: D000209:Acute-Phase Proteins / Q000096:biosynthesis; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D015703:Antigens, CD / Q000276:immunology / Q000502:physiology*; D002455:Cell Division; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000378:metabolism; D006801:Humans; D006825:Hybridomas; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D015850:Interleukin-6 / Q000494:pharmacology*; D053683:Oncostatin M; D010455:Peptides / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D018123:Receptors, Interleukin / Q000276:immunology / Q000502:physiology*; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D015398:Signal Transduction; D012995:Solubility; D015534:Trans-Activators / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D000209:Acute-Phase Proteins; D000906:Antibodies; D015703:Antigens, CD; D015853:Cysteine Proteinase Inhibitors; D004268:DNA-Binding Proteins; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D015850:Interleukin-6; C508351:OSM protein, human; C508352:Osm protein, mouse; D010455:Peptides; D018123:Receptors, Interleukin; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C494088:Stat3 protein, rat; D015534:Trans-Activators; D053683:Oncostatin M