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Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1985
Several studies using crude liver cell suspensions incubated with labeled transferrin have led to... more Several studies using crude liver cell suspensions incubated with labeled transferrin have led to a conclusion that bepatocytes have transferrin receptors. When a visual probe, which permits evaluation of transferrin binding to individual cells, was used, the binding was unexpectedly found to be limited to endothelial cells in liver cell suspensions. Neither hepatocytes nor Kupffer cells contained transferrin receptors. In the present study, we fractionated liver cell suspensions using metrizamide gradients and centrifugal elutriation to obtain hepatocytes, Kupffer cell and endothelial cell fractions of high purity. Incubation of these fractions with 125Ior SgFe-labeled transferrin led to exclusive binding to endothelial cells but not bepatocytes nor Kupffer cells. Kinetic analysis demonstrated K d of 1.9" 10-7 M, B,,ax of 3.1 pmol/106 cells with receptor numbers estimated as 1.8. 106/cell. The rate of binding was at least 28 ng (0.35 pmoi) protein/106 cells per min, corresponding to 2.1 • 105 molecules/cell per min. At 4°C, the binding reached a steady-state plateau within 5 min. Comparison of our data with those of previous investigators demonstrates a consistency if we consider that crude liver cell suspensions are contaminated with 2-3% endothelial cells. Thus, the previously reported findings may be entirely due to the contamination of crude liver cell suspensions with a small number of endothelial cells.
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1985
Several studies using crude liver cell suspensions incubated with labeled transferrin have led to... more Several studies using crude liver cell suspensions incubated with labeled transferrin have led to a conclusion that bepatocytes have transferrin receptors. When a visual probe, which permits evaluation of transferrin binding to individual cells, was used, the binding was unexpectedly found to be limited to endothelial cells in liver cell suspensions. Neither hepatocytes nor Kupffer cells contained transferrin receptors. In the present study, we fractionated liver cell suspensions using metrizamide gradients and centrifugal elutriation to obtain hepatocytes, Kupffer cell and endothelial cell fractions of high purity. Incubation of these fractions with 125Ior SgFe-labeled transferrin led to exclusive binding to endothelial cells but not bepatocytes nor Kupffer cells. Kinetic analysis demonstrated K d of 1.9" 10-7 M, B,,ax of 3.1 pmol/106 cells with receptor numbers estimated as 1.8. 106/cell. The rate of binding was at least 28 ng (0.35 pmoi) protein/106 cells per min, corresponding to 2.1 • 105 molecules/cell per min. At 4°C, the binding reached a steady-state plateau within 5 min. Comparison of our data with those of previous investigators demonstrates a consistency if we consider that crude liver cell suspensions are contaminated with 2-3% endothelial cells. Thus, the previously reported findings may be entirely due to the contamination of crude liver cell suspensions with a small number of endothelial cells.