Specific Growth Hormone Receptors on Human Peripheral Mononuclear Cells: Reexpression, Identification, and Characterization

Abstract

Although specific GH receptors have been demonstrated in various tissues of a number of species, the presence of GH receptors on human peripheral mononuclear cells (PMC) is controversial. Binding of human GH (hGH) to its receptor as the hypothesized initial step of hormone action was consequently studied using mononuclear cells from peripheral venous blood of normal subjects. Specific binding of [¹²⁵I]hGH was rapid, reversible, and time and temperature dependent. Specific GH binding to PMC was maximal after 8–24 h of preincubation. Binding of hormone was maximal at 37 C after incubation of cells for 2 h. Dissociation of GH was maximal at 37 C after the addition of 6 M NaCl. A linear relationship between specific GH binding and cell number was found. Saturation of GH binding to 10⁶ PMC was obtained with 25 ng iodinated hormones. Halfmaximal inhibition of GH binding occurred at 12–25 ng unlabeled hGH/tube. Hypothalamic and pituitary hormones as well as insulin did not interfere with specific hGH binding to PMC. Scatchard analysis of ]¹²⁵I]hGH binding to PMC revealed a receptor with a mean affinity constant of 1.5 ± 0.2 (±SD) ×lO⁹/ M−1 (n = 72) and a maximal binding capacity of 7.1 ± 2.0 × 10−¹¹M/10⁶ cells. The concentrations of calcium, sodium, and magnesium ions in the incubation medium strongly influenced GH binding, whereas pH or potassium concentration did not. As interassay variation of the binding assay was low (14% for total binding; 6% for specific hGH binding), this direct approach to study tissue receptors for hGH in a human in vitro test was reproducible and should encourage the investigation of receptor regulation as well as the study of binding in human disease.