Apoptosis of Bovine Neutrophils Following Diapedesis Through a Monolayer of Endothelial and Mammary Epithelial Cells

Abstract

In a two-chamber system, isolated blood polymorphonuclear neutrophil leukocytes (PMN) were allowed to migrate (5h, 37°C) in response to bovine complement component C5a across calfskin and rat-tail type I collagen-coated micropore membranes, arterial endothelial, or mammary epithelial cell monolayer on calfskin and rat-tail collagen-coated membranes, respectively. Migration through calfskin collagen-coated membranes resulted in 14.5%±3.4% apoptotic PMN, which was significantly higher than 6.6%±1.2% apoptotic nonmigrated C5a-treated PMN. The addition of an endothelial or epithelial cell monolayer to collagen-coated membranes prevented apoptosis of migrated PMN. After removing the membranes, nonmigrated (untreated and C5a treated) and migrated PMN were incubated for an additional 20h. At this time point, 69.1%±4.5% and 47%±4.5% of PMN that have migrated through a calfskin-coated membrane and an endothelial monolayer, respectively, were apoptotic, compared with 28.2%±3.0% and 21.1%±4.5% apoptotic untreated and C5a-treated PMN, respectively; 46.9%±4.8% of PMN that have migrated through rat-tail-coated membranes were apoptotic compared with 14.7%±2.3% and 9.3%±1.2% apoptotic untreated and C5a-treated PMN, respectively. Migration across rat-tail collagen-coated membranes with a monolayer of epithelial cells did not affect apoptosis of migrated PMN, even after 20h of incubation. In conclusion, migration of PMN across collagen-coated membranes (either calfskin or rat-tail collagen) induced an apoptotic response, which was downregulated by a monolayer of endothelial cells and was negated by an epithelial cell monolayer.