Antibodies to rat soluble IL-6 receptor stimulate B9 hybridoma cell proliferation

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. (C) 1997 Federation of European Biochemical Societies.