{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T15:39:03Z","timestamp":1792078743572,"version":"4.4.0"},"reference-count":37,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,10,2]],"date-time":"2025-10-02T00:00:00Z","timestamp":1759363200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:sec><jats:title>Introduction<\/jats:title><jats:p>Neutrophils use Fc gamma receptors (Fc\u03b3Rs) to recognize IgG-opsonized pathogens, triggering antimicrobial functions including phagocytosis, ROS production, and cytokine release. CD16b, the most abundant Fc\u03b3R on neutrophils, plays a key role in initiating these responses, while CD32a is another abundant Fc\u03b3R on neutrophils that contributes to modulating immune functions. CD16b lacks an intracellular domain and its signaling mechanisms remain unclear. The prevalence of the CD16b-deficient phenotype on donor neutrophils is estimated at &amp;lt;1% of the global population, which complicates its study. To address this, we employed CRISPR\/Cas9 to generate HL-60-derived neutrophil-like cells deficient for CD16b or CD32a, that facilitate investigation of their respective roles in neutrophil biology.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>We disrupted the <jats:italic>FCGR3B<\/jats:italic> or <jats:italic>FCGR2A<\/jats:italic> genes using CRISPR\/Cas9 in the HL-60 cell line and differentiated clones into neutrophil-like cells using 1.3% DMSO. Functional assays were performed, including phagocytosis, ROS production, SYK phosphorylation, and cytokine responses.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results and discussion<\/jats:title><jats:p>Both CD16b<jats:sup>-\/-<\/jats:sup> and CD32a<jats:sup>-\/-<\/jats:sup> HL-60-derived clones maintained neutrophilic differentiation and phagocytic capacity but displayed impaired Fc\u03b3R-mediated ROS production and SYK phosphorylation, with more pronounced defects in CD16b<jats:sup>-\/-<\/jats:sup> cells. Cytokine production was altered in both lines, with CD16b<jats:sup>-\/-<\/jats:sup> cells producing less IL-6 and IL-1\u03b2, and CD32a<jats:sup>-\/-<\/jats:sup> cells producing less TNF-\u03b1 and IL-10. This model provides new insights into the distinct roles of CD16b and CD32a in neutrophil activation and immune responses.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fimmu.2025.1633609","type":"journal-article","created":{"date-parts":[[2025,10,2]],"date-time":"2025-10-02T05:31:50Z","timestamp":1759383110000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":3,"title":["Development of CRISPR\/Cas9-mediated CD16b-\/- and CD32a-\/- promyelocytic cell lines to study Fc\u03b3R signaling in human neutrophils"],"prefix":"10.3389","volume":"16","author":[{"given":"Jos\u00e9 Antonio","family":"Cruz-C\u00e1rdenas","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alejandra","family":"L\u00f3pez-Arredondo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jorge Andr\u00e9s","family":"C\u00e1zares-Preciado","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mabel","family":"Rodr\u00edguez-Gonzalez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Laura A.","family":"Palomares","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marion E. 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