Physical characterization of cyclosporine binding sites in lymphocytes.

The binding of cyclosporine to human peripheral blood lymphocytes (PBLs) was studied by measuring the fluorescence emission spectrum and lifetime of the fluorescent and immunosuppressive cyclosporine derivative dansyl-cyclosporine (DCs). The emission maximum and fluorescence lifetime of DCs were characterized in several solvents. The fluorescence emission maximum and lifetime of DCs increased at a high dielectric constant. The fluorescence lifetime decay curve of DCs was a monoexponential function in all solvents tested. Fluorescence micrographs of lipid vesicles and erythrocytes labeled with DCs exhibit uniform staining patterns, whereas PBLs show heterogeneous DCs labeling. DCs exhibits a relatively low emission maximum (490 nm) in erythrocyte membranes. Such an emission maximum is characteristic of a hydrophobic environment. DCs in PBLs also has a low emission maximum (484 nm). The lifetime of DCs in PBLs required two exponential terms to properly fit the lifetime decay curve and could not be attributed to light scattering. One short component (4.7 +/- 1.0 ns) and a second long component (18.5 +/- 1.0 ns) were resolved from the DCs fluorescence decay curves. Time-resolved anisotropy of DCs in PBLs revealed that the labeled drug was present in an anisotropic environment, consistent with at least some DCs being bound to a membrane. These fluorescence studies suggest that DCs interacts with multiple and/or heterogeneous sites in peripheral blood lymphocytes.