Identification of an α2-Macroglobulin Receptor in Human Mammary Epithelial Cells

Abstract

Several cases of zinc (Zn) deficiency in human infants caused by abnormally low concentrations of Zn in breast milk were recently reported, the underlying mechanism of which is not known. Alpha2-macroglobulin (α2-M), a major Zn-binding ligand in serum, presents a potential vehicle for mammary Zn uptake. This study was conducted to determine if an α2-M receptor is present in human mammary epithelial cells, where it may be involved in the endocytosis of α2-M into the mammary gland. Normal human mammary epithelial cells were grown to confluency in serum-free medium. For all binding and uptake studies, α2-M, preactivated with methylamine and labeled with 125I, was added to cells for varied lengths of time to determine saturation over time and at varied concentrations to determine saturation over increasing concentration of ligand. Nonspecific and competitive binding were measured by addition of a 100-fold molar excess of unlabeled α2-M and serum albumin or lactoferrin, respectively. Binding at 4°C was specific for α2-M and approached saturation kinetics at 56 nmol/L. Scatchard plot analysis of the binding data demonstrated more than one binding site: a high affinity, saturable binding site and a low affinity, nonsaturable binding site. Uptake of α2-M at 37°C was rapid and continuous over increasing concentrations of α2-M, and internalized α2-M was rapidly degraded. Results from this study present evidence for receptor-mediated uptake of α2-M in human mammary epithelial cells, which in turn, provides a potential mechanism for Zn acquisition by the cell.