Detection of human growth hormone receptors on IM-9 cells and peripheral blood mononuclear cell subsets by flow cytometry: Correlation with growth hormone-binding protein levels

We have developed a method using flow cytometry to identify fluorescein- conjugated GH receptors (GHR) on IM-9 lymphocytes and circulating peripheral blood mononuclear cell subsets. Binding to IM-9 cells and peripheral blood mononuclear cells was concentration dependent and could be competitively blocked by the addition of unlabeled human GH, but not by the addition of rat or bovine GH or human insulin or PRL. Using two-color flow cytometric analysis, fluorescein-conjugated human GHR were readily detected on more than 90% of B lymphocytes and monocytes, but only variably on T lymphocytes. B Lymphocytes and monocytes had approximately 5000 GHR/cell. Using two color flow cytometry, we identified GHR on circulating B lymphocytes in subjects with GH deficiency (n = 9), precocious puberty (n = 6), and Turner syndrome (n = 5) and in seven subjects with miscellaneous disorders, including familial short stature, bone dysplasia, Crohn disease, congenital adrenal hyperplasia, and acromegaly. The percentage of B lymphocytes expressing GHR in subjects with GH deficiency (mean ± SD, 95 ± 9%), precocious puberty (91 ± 15%), and Turner syndrome (84 ± 15%) was not different from that in normal volunteers (90 ± 12%; n = 14). In 10 subjects, serum GH-binding protein levels were assayed simultaneously with B lymphocyte GHR. GH-binding protein was normal in all (mean, 1255 pmol/L; range, 773-1809). There was a good correlation between GHR expression on B lymphocytes and GH-binding protein levels (r = 0.75; P = 0.01). We postulate that GHR found on circulating B lymphocytes may contribute to the pool of receptors identified in serum as GH-binding proteins. Two color flow cytometry appears to be an effective method for the detection of GHR on circulating peripheral blood mononuclear cell subsets. The evaluation of GHR on circulating B lymphocytes may prove to be a useful means of evaluating GH-GHR interactions in subjects with growth disorders.