Self-extinguishing relay waves enable homeostatic control of human neutrophil swarming

Abstract

Neutrophils exhibit self-amplified swarming to sites of injury and infection. How swarming is controlled to ensure the proper level of neutrophil recruitment is unknown. Using an ex vivo model of infection, we find that human neutrophils use active relay to generate multiple pulsatile waves of swarming signals. Unlike classic active relay systems such as action potentials, neutrophil swarming relay waves are self-extinguishing, limiting the spatial range of cell recruitment. We identify an NADPH-oxidase-based negative feedback loop that is needed for this self-extinguishing behavior. Through this circuit, neutrophils adjust the number and size of swarming waves for homeostatic levels of cell recruitment over a wide range of initial cell densities. We link a broken homeostat to neutrophil over-recruitment in the context of human chronic granulomatous disease.

Arp2/3 complex and the pentose phosphate pathway regulate late phases of neutrophil swarming

“… 36 Previous work has shown that the activity of NADPH oxidase and ROS signaling are dispensable for initiating neutrophil swarming. 13 , 16 , 18 Based on the use of neutrophils from NADPH oxidase-deficient chronic granulomatous disease (CGD) patients in these studies, ROS production appears to emerge as a control mechanism to avoid over-recruitment of neutrophils at late stages of swarming. However, none of these studies have pinpointed the sources of ROS production in populations of swarming neutrophils.…”

“…Several studies have used these controllable swarming devices for the functional characterization of human neutrophils from the blood of patients with different disease backgrounds. 9 , 16 , 17 , 18 , 19 Furthermore, technical adaptations now also allow the study of neutrophil swarming in response to patterned arrays of living microbes, in particular different species of living fungi. 18 , 20 , 21 , 22 Thus, recent developments of swarming-on-a-chip platforms have brought forward versatile applications for the study of primary human and mouse neutrophils, but also neutrophil-like cell lines.…”

“… 23 Several biological findings were derived from such studies, including the analysis of the neutrophil swarm secretome, 9 the involvement of exosomes in swarming, 24 the interaction of swarming neutrophils with monocytes, 25 and the functional mapping of molecular pathways involved in the neutrophil swarming response in vitro. 11 , 16 , 18 , 21 , 26 Although several of these studies performed live-cell imaging to capture neutrophil swarming dynamics, the use of bright-field and widefield fluorescence microscopy at low resolution prevented the visualization of more detailed biological information, e.g., subcellular components inside individual cells. By using fluorescent calcium sensors, Khazen et al.…”

“…These elegant studies showed the power and great potential of applying fluorescence microscopy techniques to studies on in vitro swarming. 16 , 27 Despite all these technical developments, the analysis of crowding neutrophils during the late stages of swarming has been similarly problematic in swarming-on-a-chip platforms as in native tissues. Experimental in vitro setups were also limited in spatiotemporally resolving individual cells in dense neutrophil clusters.…”

A LTB4/CD11b self-amplifying loop drives pyogranuloma formation in chronic granulomatous disease

“… 21 , 43 , 44 The subsequent recruitment of macrophages leads to swarm resolution, making this a transient process 44 , 45 with the LTB 4 amplification loop ultimately self-limited by NADPH oxidase activity. 46 LTB 4 signaling appears to be heightened in CGD, where loss of NADPH oxidase activity fails to inhibit the production of LTB 4 46 and cell aggregation results in the heightened transcellular production of LTB 4 . 47 LTB 4 is of particular importance in innate responses to fungal pathogens generally 47 , 48 , 49 , 50 and, in CGD mice, has been shown to be the key chemoattractant for early neutrophil recruitment following zymosan installation and the initiator of bone marrow neutrophil aggregation.…”