Identification of an alpha2-macroglobulin receptor in human mammary epithelial cells

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 (alpha2-M), a major Zn-binding ligand in serum, presents a potential vehicle for mammary Zn uptake. This study was conducted to determine if an alpha2-M receptor is present in human mammary epithelial cells, where it may be involved in the endocytosis of alpha2-M into the mammary gland. Normal human mammary epithelial cells were grown to confluency in serum-free medium. For all binding and uptake studies, alpha2-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 alpha2-M and serum albumin or lactoferrin, respectively. Binding at 4 degreesC was specific for alpha2-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 alpha2-M at 37 degreesC was rapid and continuous over increasing concentrations of alpha2-M, and internalized alpha2-M was rapidly degraded. Results from this study present evidence for receptor-mediated uptake of alpha2-M in human mammary epithelial cells, which in turn, provides a potential mechanism for Zn acquisition by the cell.