Elastase Controls the Binding of the Vitamin D-Binding Protein (Gc-Globulin) to Neutrophils: A Potential Role in the Regulation of C5a Co-Chemotactic Activity

摘要

The vitamin D-binding protein (DBP) binds to the plasma membranes of numerous cell types and mediates a diverse array I] cellular functions. DBP bound to the surface of leukocytes serves as a co-chemotactic factor for CSa, significantly enhancing th chemotactic activity of pM concentrations of CSa. This study investigated the regulation of DBP binding to neutrophils as : possible key step in the process of chemotaxis enhancement to CSa. Using radioiodinated DBP as a probe, neutrophils release. 70% of previously bound DBP into the extracellular media during a 60-min incubation at 37°C. This was suppressed by serilll protease inhibitors (PMSF, Pefabloc SC), but not by metallo- or thiol-protease inhibitors. DBP shed from neutrophils had Dl detectable alteration in its m.w., suggesting that a serine protease probably cleaves the DBP binding site, releasing DBP in al unaltered form. Cells treated with PMSF accumulate DBP vs time with over 90% of the protein localized to the plasma membrane Purified neutrophil plasma membranes were used to screen a panel of protease inhibitors for their ability to suppress sheddinl of the DBP binding site. Only inhibitors to neutrophil elastase prevented the loss of membrane DBP-binding capacity. Moreover treatment of intact neutrophils with elastase inhibitors prevented the generation of CSa co-chemotactic activity from DBP. Thesl results indicate that steady state binding of DBP is essential for co-chemotactic activity, and further suggest that neutrophi elastase may playa critical role in the CSa co-chemotactic mechanism.