A quantitative assessment of non specific pinocytosis by human endothelial cells surviving in vitro

Human umbilical vein endothelial cells have been assayed in vitro, 24 hrs. after plating, for non specific pinocytic activity. The culture conditions were designed to minimize the exogenous stimulations of pinocytosis, such as those possibly coming from mitotic induction and chemical and contact-dependent signaling. Two different markers were used: Lucifer Yellow CH (LY), and three different preparations of horseradish peroxidase, a multiple form (type II) composed of five different isoenzymes, and two purified acidic (type VIII) and basic (type IX) isoenzymes. The uptake of LY appears to depend on both fluid-phase incorporation and non specific adsorption to the cell surface, and it shows a linear monophasic dependence on time and a linear diphasic dependence on concentration. This probe is actively chased from the cells to an extent proportional to the amount incorporated. Therefore, the endocytic index obtained from the LY incorporation data is not a reliable estimate of extracellular fluid incorporation. The three different forms of HRP share an incorporation pattern linearly dependent on both time and concentration, consistent with the classical interpretation of a simple fluid-phase mechanism of intracellular uptake; however, the rates of uptake and chase activity of the pure isoenzymes are clearly different from that of the multiple form. The observed differences are related to possible local variations in the physicochemical properties of the cell surface, which may restrict the cell surface area suitable for fluid-phase uptake of differently charged macromolecular probes.