Mechanical properties of rat bone marrow and circulating neutrophils and their responses to inflammatory mediators

Abstract

Abstract Neutrophils are continuously released from the bone marrow (BM), and this release is accelerated during inflammation. This study compared the mechanical properties of mature neutrophils within the BM and the circulating blood, as well as the role of microtubule rearrangement in the release of neutrophils from the BM in rats. Neutrophils isolated from the BM were stiffer than neutrophils in the circulating blood, using magnetic twisting cytometry. BM neutrophils also contained more F-actin within the submembrane region than circulating neutrophils when examined using confocal microscopy, suggesting that mature quiescent neutrophils within the BM are stiffer than circulating neutrophils because of increased formation of F-actin beneath the plasma membrane. Complement protein 5 fragments or formylmethionyl-leucylphenylalanine (fMLP) induced a stiffening response within 2 minutes that was greater in circulating than in BM neutrophils. This stiffening required F-actin formation within the submembrane region but not microtubule rearrangement in both circulating and BM neutrophils. fMLP-induced shape changes were more pronounced in circulating than in BM neutrophils, which showed fewer and smaller pseudopods and fewer membrane irregularities. In vivo, fMLP induced neutropenia, sequestration of neutrophils within the pulmonary capillaries, and release of neutrophils from the BM. Studies using colchicine demonstrated that rearrangement of microtubules was not required for any of these processes but was required for normal trafficking of neutrophils through the pulmonary capillaries.