Characterization of Specific Growth Hormone Binding Sites in Mouse Fibroblasts*

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Abstract
The demonstration that mouse embryo mesenchymal cells produce sotmatomedin-like immunoreactivity suggests that fibroblasts may be a target tissue for GH [growth hormone] action. The aim of this study was to investigate the interaction of GH with mouse fibroblasts. Specific binding sites for GH in mouse fibroblasts (BALB/c and Swiss 3T3) were characterized. Binding of [125I]iodo-human GH (hGH) was rapid, reversible, and time and temperature dependent. Maximal binding was achieved within 2 h at 30.degree. C and was rapidly dissociable at this temperature with or without excess unlabeled hormone. Specific binding of [125I]iodo-hGH was similar over the pH range 6.6-8.2. Half-maximal inhibition of specific binding was obtained with 10 ng/ml hGH. A linear relationship between specific binding and cell number was found and negligible degradation of [125I]iodo-hGH occurred during the binding studies. Somatogenic hormones from various species, including mouse GH, rat GH, porcine GH, and bovine GH competed for binding with [125I]iodo-hGH. Lactogenic hormones did not displace [125I]iodo-hGH at low concentrations. Scatchard analysis revealed curvilinear plots suggesting that [125I]-iodo-hGH was binding to 2 sites. The affinity constants and capacities of these binding sites on BALB/c 3T3 cells were 7.46 .times. 109 M-1 and 4000 sites per cell and 0.26 .times. 109 M-1 and 67,000 sites/cell. Using [125I]iodo-bovine GH and [125I]iodo-human placental lactogen as labeled ligands 2 distinct binding sites were found with affinity constants of 6.1 .times. 109 M-1 and 1.1 .times. 109 M-1, respectively. These data are consistent with the presence of cell surface somatogenic and lactogenic receptos in mouse fiboblasts and suggest GH receptors may be present on many cell types not previously considered to be target tissues for GH action. The mouse fibroblast GH receptor may provide a useful model for somatogenic receptors, particularly if it is coupled to somatomedin production as appears to be the case with human fibroblasts.