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Myeloid cells are elevated in the peripheral blood and contribute to peritonitis, but their circulating dynamics are not clear. In vivo flow cytometry (IVFC) is a noninvasive technique for monitoring the dynamics of circulating cells in live animals. It has been extensively used to detect circulating tumor cells, but rarely for monitoring immune cells. Here, we describe a method adapting an intravital microscope for IVFC so that we can monitor LysM-EGFP-labeled circulating myeloid cells in a tumor necrosis factor (TNF) \u03b1-induced peritonitis mouse model. Using this IVFC method, we quantified the blood flow velocity and cell concentration in circulation. We observed a significant increase in LysM-EGFP+ cells in circulation after TNF\u03b1 intraperitoneal (i.p.) injection, which reached a plateau in ~20 min. Conventional cytometry analysis showed that most LysM-EGFP+ cells were neutrophils. Increasing blood neutrophils were accompanied by neutrophil recruitment to the peritoneal cavity and neutrophil emigration from the bone marrow. We then monitored neutrophil CD64 expression in vivo and found a significant increase in TNF\u03b1-induced peritonitis. We also found that CD18 blockade doubled the circulating neutrophil number in TNF\u03b1-induced peritonitis, suggesting that CD18 is critical for neutrophil recruitment in peritonitis. Overall, we demonstrate that IVFC techniques are useful for studying the circulating dynamics of immune cells during inflammatory diseases.<\/jats:p>","DOI":"10.3390\/biom14080886","type":"journal-article","created":{"date-parts":[[2024,7,23]],"date-time":"2024-07-23T14:26:50Z","timestamp":1721744810000},"page":"886","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Monitoring Circulating Myeloid Cells in Peritonitis with an In Vivo Imaging Flow Cytometer"],"prefix":"10.3390","volume":"14","author":[{"given":"Sunitha","family":"Pulikkot","sequence":"first","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Souvik","family":"Paul","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555, USA"},{"name":"Department of Pathology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alexxus","family":"Hall","sequence":"additional","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0775-8722","authenticated-orcid":false,"given":"Brianna","family":"Gardner","sequence":"additional","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wei","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liang","family":"Hu","sequence":"additional","affiliation":[{"name":"Academy of Integrative Medicine, Shanghai University of Traditional Chinese Medicine, 1200 Cai Lun Road, Shanghai 201203, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anthony T.","family":"Vella","sequence":"additional","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7618-107X","authenticated-orcid":false,"given":"Yunfeng","family":"Chen","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555, USA"},{"name":"Department of Pathology, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5385-9626","authenticated-orcid":false,"given":"Zhichao","family":"Fan","sequence":"additional","affiliation":[{"name":"Department of Immunology, School of Medicine, UConn Health, 263 Farmington Ave., Farmington, CT 06030, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,7,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"21313","DOI":"10.1038\/s41598-022-25176-4","article-title":"NLRP3 inflammasome activation in neutrophils directs early inflammatory response in murine peritonitis","volume":"12","author":"Fukui","year":"2022","journal-title":"Sci. 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