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. 2022 Sep 21;13:5529. doi: 10.1038/s41467-022-33230-y

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — a, b Hematologic analysis of WT and Wash-cKO (Washc1^Δhaemo) mice. a White blood cells (WBC) counts. Mean ± SEM, n = 9 independent mice; ns, not significant. Two-tailed Mann-Whitney test. b Neutrophil counts in WT and Wash-cKO mice. Mean ± SEM, n = 9. *p = 0.0229. Two-tailed Mann-Whitney test. c Flow cytometry analysis of neutrophils (Ly6G+) in bone marrows from WT and Wash-cKO neutrophils. n = 3. Mean ± SEM. ****p < 0.0001. Two-tailed Student’s t-test. d CyTOF identifies similar leukocyte characteristics in WT and Wash-cKO mice. Bone marrow cells were stained with a panel of 43 surface markers and analyzed by CyTOF. Live CD45 + cells were selected for viSNE and FLOWSOM analysis. e, f Analysis of neutrophil lineage in wild-type and Wash-cKO mice. e Flow cytometry analysis of WT and Wash-cKO bone marrow neutrophil lineage maturation status. n = 4. f CyTOF analysis of wild-type and Wash-cKO bone marrows identifying distinctive neutrophil progenitor and precursor phenotypes. Bone marrow cells were stained with a panel of 43 surface markers and analyzed by CyTOF. Neutrophil lineage clusters from d were selected for viSNE and FLOWSOM analysis. n = 3. g Wash-cKO mature neutrophils show a similar developmental phenotype as in the WT mice. viSNE maps show neutrophil lineage clusters’ expression patterns of each CD marker as spectrum-colored dots. Blue = low expression. Red = high expression. n = 3. h, i Wash-cKO mice present neutrophil-mediated systemic inflammation. h Plasma levels of MMP-9 (Gelatinase granules) or MPO (myeloperoxidase, azurophilic granules) quantified by ELISA. Each symbol represents an individual mouse. n = 9. Mean ± SEM. ns, not significant. ****p < 0.0001. Two-tailed unpaired Student’s t-test. i Analysis of neutrophil infiltration into kidneys of wild-type and Wash-cKO mice measured as tissue-associated myeloperoxidase by ELISA. Each symbol represents an individual mouse. n = 6. Mean ± SEM, ****p < 0.0001. Two-tailed unpaired Student’s t-test. ns, not significant. Source data are provided as a Source Data file.