MONONUCLEAR CELL-LINE THP-1 INTERNALIZES BACTERICIDAL PERMEABILITY-INCREASING PROTEIN BY A NON-RECEPTOR-MEDIATED MECHANISM CONSISTENT WITH PINOCYTOSIS

Background: Bactericidal/permeability-increasing protein (BPI) binds l ipopolysaccharide and neutralizes its toxic effects in vitro and in en dotoxemic animals. Our recent work identified physiologically signific ant interactions between BPI, lipopolysaccharide, and mononuclear cell s. Objective: To determine whether the interaction between BPI and mon onuclear cells is receptor mediated. Design: Labeled BPI was incubated with THP-1 cells in the presence of up to 100-fold excess of unlabele d BPI. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and W estern blotting were performed to evaluate competitive binding and tot al uptake of BPI. Crosslinking was performed to determine whether BPI binds to a single protein entity. Acid washing experiments and flow cy tometric analysis were performed to determine whether BPI remains on t he cellular surface. Finally, flow cytometry analysis was used to dete rmine whether BPI incubation with THP-1 cells affects the surface expr ession of the lipopolysaccharide-binding protein-lipopolysaccharide re ceptor CD14. Results: Labeled BPI uptake was not inhibited by the pres ence of 100-fold excess of unlabeled BPI at 37 degrees C or 4 degrees C in the presence of azide. Uptake was not saturable under either cond ition with incubation concentrations up to 10 mu g/mL. Cross-linking d id not show BPI bound to a single entity. Acid washing and flow cytome try experiments disclosed rapid internalization of BPI. Finally, BPI u ptake by THP-1 cells had no effect on the surface expression of CD14. Conclusions: Bactericidal/permeability-increasing protein is rapidly i nternalized by mononuclear cells in a nonspecific fashion not saturabl e at very high doses, which is consistent with pinocytosis. This proce ss may represent a disposal mechanism for lipopolysaccharide in closed -space infections and may be partially responsible for the rapid clear ance of BPI from the peripheral circulation.