COPPER UPTAKE BY CULTURED TROPHOBLAST CELLS ISOLATED FROM HUMAN TERM PLACENTA

This paper has examined copper uptake from CuHis(2) complexes by cytot rophoblast cells isolated from term human placenta Uptake is time-depe ndent, reaching equilibrium after about 90 min, and saturable, with a calculated apparent K-m of 0.174 +/- 0.061 mu M and V-max, measured ov er 30 min, of 0.721 +/- 0.092 pmol/min/mu g DNA. To determine whether ATP was required for uptake, cells were incubated with inhibitors of g lycolysis (iodoacetate) and the TCA cycle (sodium azide and cyanide). Iodoacetate and sodium azide had no effect on uptake, but cyanide decr eased the initial rate of uptake. This effect was due to copper bindin g to the inhibitor and decreasing the effective substrate concentratio n rather than inhibition of uptake through ATP depletion. Ouabain and monensin had no effect, showing that neither the Na+ gradient nor endo cytosis were involved in uptake. The monovalent ion chelator, bathocup roine sulphonate, had no effect on uptake but buthionine sulfoximine, an inhibitor of glutathione synthesis, did decrease both the rate of u ptake and equilibrium copper levels, suggesting that copper may bind t o glutathione within the cell. The data show that copper is taken up b y a passive carrier-mediated transporter and, following uptake, binds to glutathione within the cell.