Production of insulin-like growth factor binding protein (IGFBP), IGFBP-3 protease, and expression of IGF-I receptors by cells of the sheep immune system.
Single-cell suspensions of sheep thymus cells were cultured in serum-free medium with or without polyclonal activators (phytohaemagglutinin or concanavalin A) and the resultant conditioned medium was assayed for insulin-like growth factor binding protein (IGFBP) activity by binding of [125I]IGF-I, using charcoal to separate free from bound. All cultures produced IGFBP but mitogen stimulation significantly increased IGFBP concentrations, indicating production by lymphoid cells. Conditioned medium also degraded recombinant human [125I]IGFBP-3, suggesting IGFBP-3 protease production within the thymus. This degradation was inhibited by several protease inhibitors (phenylmethylsulphonyl fluoride, aprotinin, N-alpha-p-tosyl-L-lysine chloromethyl ketone), suggesting the presence of a serine protease. Cell surface [125I]IGF-I binding was demonstrated on cells from thymus, mesenteric lymph node, peripheral blood mononuclear cells and platelets. The [125I]IGF-I binding to platelets could be inhibited by unlabelled peptides, with relative potencies IGF-I > IGF-II >> insulin. Scatchard analysis of IGF-I competitive binding revealed a Kd of 266 pmol/l and approximately 40 receptor sites per cell. The high-affinity binding of IGF-I and competition by insulin suggested that the [125I]IGF-I binding was to an IGF-I receptor rather than to a membrane-associated IGFBP, to which insulin does not bind. These data provide further support for the role of the IGF-IGFBP axis in the immune system, particularly in relation to the thymus.
Tonner
E
E
Hannah Research Institute, Ayr, UK.
Beattie
J
J
Flint
D J
DJ
D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis*; D002478:Cells, Cultured; D010450:Endopeptidases / Q000096:biosynthesis*; D007107:Immune System / Q000166:cytology / Q000378:metabolism*; D018969:Insulin-Like Growth Factor Binding Proteins; D007334:Insulin-Like Growth Factor I / Q000378:metabolism*; D017451:Receptors, Somatomedin / Q000378:metabolism*; D012756:Sheep; D013002:Somatomedins / Q000378:metabolism; D013950:Thymus Gland / Q000166:cytology / Q000378:metabolism
D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D017451:Receptors, Somatomedin; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D010450:Endopeptidases; C080473:insulin-like growth factor binding protein-3 protease