MACROMOLECULE MACROMOLECULE INTERACTION IN DRUG DISTRIBUTION .2. EFFECT OF ALPHA-GLOBULIN ON SATURABLE UPTAKE OF FRACTIONATED [H-3] HEPARIN BY RAT PARENCHYMAL HEPATOCYTES IN PRIMARY CULTURE

The uptake of fractionated [H-3]heparin was investigated in rat parenchymal hepatocytes in primary culture. The initial uptake of fractionated [H-3]heparin was found to be saturable with the maximum uptake velocity (V(max)) of 10. 1 +/- 1.46 pmol/min/mg protein and the Michaelis constant (K(m)) of 284 +/- 47.9 nm. The effect of alpha-globulin, the major protein binding to fractionated [H-3]heparin, on the saturable uptake profile of fractionated [H-3]heparin was also investigated. The uptake clearance was reduced, depending on the concentration of fractionated [H-3]heparin, by the addition of 1 mg/ml alpha-globulin. We assumed that fractionated H-3-heparin bound to alpha-globulin was not available for uptake and that the reduction in the uptake clearance was solely attributable to die saturable binding of fractionated [H-3]heparin to alpha-globulin. The uptake clearance versus concentration profile was analyzed to obtain the dissociation constant (K(d)) of 31.8 nm and the capacity (n) of 0.047 for the binding of fractionated [H-3]heparin to alpha-globulin. The saturable binding of fractionated [H-3]heparin to alpha-globulin was supported by in vitro binding experiments using gel chromatography, in which bound fractionated [H-3]heparin decreased with the concentration of fractionated [H-3]heparin in the presence of alpha-globulin. In conclusion, the present study demonstrated the saturable uptake of fractionated [H-3]heparin by rat parenchymal hepatocytes and the saturable binding of fractionated [H-3]heparin to alpha-globulin. The saturable uptake may suggest the involvement of a specific transport system such as receptor-mediated endocytosis.