Binding of cyclosporine by human lymphocytes and phospholipid vesicles.

Abstract

The purpose of this investigation was to identify and characterize a possible plasma membrane receptor for cyclosporine (CsA) on human lymphoid cells. Binding of 3H-CsA by normal human lymphocytes and purified lymphoid subpopulations was examined using ligand concentrations ranging from 5 X 10(-7) to 10(-10) M. Specificity was determined using a 500-fold excess of unlabeled drug. No differences were observed in the uptake of 3H-CsA by B cells or T cells using Cartesian or semilogarithmic analyses. Scatchard analysis of the specific binding of CsA by normal peripheral blood lymphocytes yielded two dissociation constants: a high affinity site with a KD of 2 to 6 X 10(-9) M and a low affinity site with a KD of about 10(-7) M CsA. Scatchard analysis of specific CsA uptake by purified splenic T cells and B cells showed both populations to exhibit a low affinity site (KD = 3 to 6 X 10(-7) M and 5 to 8 X 10(-7) M, respectively), whereas only B cells bore a high affinity site (KD = 2 X 10(-9) M). Analysis of specific CsA uptake by cultured human kidney cells and phospholipid vesicles (1:1 molar ratio of phosphatidylcholine:cholesterol) showed both these targets to display a single, low affinity binding site (KD = 1 X 10(-7) M and 2 X 10(-8) M, respectively). The relevance of the low affinity site to CsA-mediated in vitro immunosuppression was suggested by the need for 10(-7) M CsA to achieve 50% suppression of lymphoproliferation during mixed lymphocyte culture. Taken together, these data argue against the existence of a specific CsA receptor on the surface of human T cells. Although a role for the high affinity B cell receptor in some types of CsA-induced immunomodulation cannot be excluded, the present study suggests that immunosuppression is achieved by the partitioning of the hydrophobic CsA molecule into the membrane lipid bilayer, thereby perturbing homeostatic control of membrane function.