Semaphorin 3F signaling actively retains neutrophils at sites of inflammation
Tracie Plant, … , Moira K.B. Whyte, Sarah R. Walmsley
Tracie Plant, … , Moira K.B. Whyte, Sarah R. Walmsley
Published March 19, 2020
Citation Information: J Clin Invest. 2020;130(6):3221-3237. https://doi.org/10.1172/JCI130834.
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Research Article Inflammation Pulmonology

Semaphorin 3F signaling actively retains neutrophils at sites of inflammation

Abstract

Neutrophilic inflammation is central to disease pathogenesis, for example, in chronic obstructive pulmonary disease, yet the mechanisms that retain neutrophils within tissues remain poorly understood. With emerging evidence that axon guidance factors can regulate myeloid recruitment and that neutrophils can regulate expression of a class 3 semaphorin, SEMA3F, we investigated the role of SEMA3F in inflammatory cell retention within inflamed tissues. We observed that neutrophils upregulate SEMA3F in response to proinflammatory mediators and following neutrophil recruitment to the inflamed lung. In both zebrafish tail injury and murine acute lung injury models of neutrophilic inflammation, overexpression of SEMA3F delayed inflammation resolution with slower neutrophil migratory speeds and retention of neutrophils within the tissues. Conversely, constitutive loss of sema3f accelerated egress of neutrophils from the tail injury site in fish, whereas neutrophil-specific deletion of Sema3f in mice resulted in more rapid neutrophil transit through the airways, and significantly reduced time to resolution of the neutrophilic response. Study of filamentous-actin (F-actin) subsequently showed that SEMA3F-mediated retention is associated with F-actin disassembly. In conclusion, SEMA3F signaling actively regulates neutrophil retention within the injured tissues with consequences for neutrophil clearance and inflammation resolution.

Authors

Tracie Plant, Suttida Eamsamarng, Manuel A. Sanchez-Garcia, Leila Reyes, Stephen A. Renshaw, Patricia Coelho, Ananda S. Mirchandani, Jessie-May Morgan, Felix E. Ellett, Tyler Morrison, Duncan Humphries, Emily R. Watts, Fiona Murphy, Ximena L. Raffo-Iraolagoitia, Ailiang Zhang, Jenna L. Cash, Catherine Loynes, Philip M. Elks, Freek Van Eeden, Leo M. Carlin, Andrew J.W. Furley, Moira K.B. Whyte, Sarah R. Walmsley

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Figure 2

Knockdown of semaphorin 3F by MO injection or by TALEN-induced mutation accelerates resolution of neutrophilic inflammation in zebrafish.

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Knockdown of semaphorin 3F by MO injection or by TALEN-induced mutation ...
(A–D) sema3fa and/or sema3fb MO (1 nL of 0.5 mM) was injected into 1-cell-stage zebrafish mpx:GFP embryos, with 1 nL of 0.5 mM control MO used as a negative control. Tail fin transection was performed at 2 dpf, and neutrophils were counted at 6 hpi and 24 hpi. (A) Neutrophil counts at the 6 hpi time point with (B) overlaid fluorescence and bright-field photomicrographs (recruitment). (C) Neutrophil counts at the 24 hpi time point with (D) overlaid fluorescence and bright-field photomicrographs (resolution). Scale bars: 60 μm (B and D). Data are mean ± SEM, with individual data points (n = 30) from 3 independent experiments. (E–G) sema3fa- or sema3fb-mutated F1 fish were incrossed and compared with mpx:GFP fish. Tail fin transection was performed at 2 dpf, and (E) neutrophils were counted at 6 hpi. Data are mean ± SEM, with individual data points (n = 30) from 4 independent experiments. (F) Whole-body total neutrophil numbers were counted at 3 dpf. Data are mean ± SEM, with individual data points (n = 60) from 4 independent experiments. (G) Tail fin transection was performed at 2 dpf and neutrophils were counted at 24 hpi. Data are mean ± SEM, with individual data points (n = 5–60) from 4 independent experiments. (H) sema3fa and or sema3fb MOs (1 nL of 0.5 mM) were injected into 1-cell-stage zebrafish mpx:kaede embryos, and tail fin transection was performed at 2 dpf. Neutrophils at 6 hpi were recruited to the wound and photoconverted, and red fluorescence neutrophils were tracked for 3.5 hours. Data are from 3 independent experiments (n = 9). Statistical analysis was by 1-way ANOVA and Bonferroni’s post hoc test. ***P < 0.001.