Neutrophil Methods and Protocols

Dorward, David A., Rossi, Adriano G., Dransfield, Ian, Lucas, Christopher D., David A. Dorward, Adriano G. Rossi, Ian Dransfield, Christopher D. Lucas

Timely neutrophil apoptosis and cell clearance by surrounding phagocytes are essential components of the resolution phase of acute inflammation. Programmed cell death by apoptosis occurs with maintenance of an intact cell membrane in order to prevent the release of histotoxic intracellular products such as proteases and reactive oxidant species into the extracellular surroundings as occurs during necrosis. Macrophage phagocytosis results in attenuation of toll-like receptor-driven proinflammatory mediator production further promoting inflammation resolution. Failures in this cascade of events can result in tissue damage, chronic inflammation and disease. By studying human neutrophil apoptosis and phagocytic clearance in vitro, it is possible to delineate key control mechanisms in the regulation of these processes and therefore also identify potential therapeutic targets. Apoptotic signalling pathways are well described in the literature using a variety of laboratory techniques. In this paper, we outline the key in vitro assays used to assess neutrophil apoptosis, activation of key components of the apoptotic machinery, and phagocytic clearance of these cells.