{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,2]],"date-time":"2026-10-02T05:48:13Z","timestamp":1790920093124,"version":"4.1.0"},"reference-count":35,"publisher":"Public Library of Science (PLoS)","issue":"9","license":[{"start":{"date-parts":[[2025,9,8]],"date-time":"2025-09-08T00:00:00Z","timestamp":1757289600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Higher Education and Science Committee, The RA Ministry of Education, Science, Culture and Sports","award":["SCS 21AG-1F072"],"award-info":[{"award-number":["SCS 21AG-1F072"]}]},{"name":"Higher Education and Science Committee, The RA Ministry of Education, Science, Culture and Sports","award":["SCS 24AA-1F007"],"award-info":[{"award-number":["SCS 24AA-1F007"]}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    The short lifespan of polymorphonuclear neutrophils (PMNs)\n                    <jats:italic>\n                      <jats:italic>in vitro<\/jats:italic>\n                    <\/jats:italic>\n                    poses challenges, as their limited viability restricts functional assays and experimental manipulations. The HL-60 cell line serves as a valuable model for neutrophil-like differentiation, yet the functional relevance of ATRA- and DMSO-induced differentiation remains incompletely understood. In the present study, we aimed to characterize the differentiation potential of all-trans retinoic acid (ATRA) and dimethyl sulfoxide (DMSO) on HL-60 cells and compare their functionality with primary PMNs. Besides that, we performed profound immunophenotypes of the cells with multicolor cytometry, and evaluated their antitumor capabilities. Our findings indicate that both differentiation conditions yield cells resembling immature neutrophils, exhibiting promyelocyte-like morphology, lacking key maturity markers. However, ATRA-differentiated cells exhibit a more mature phenotype, with higher expression of\n                    <jats:italic>C\/EBP\u03b1<\/jats:italic>\n                    and reduced proliferation rates, indicating advanced differentiation. Functionally, ATRA-dHL-60 cells displayed limited immune responses, showing minimal phagocytic activity, low ROS production, and a reduced response to LPS. In contrast, DMSO-dHL-60 cells, despite their less mature phenotype, showed enhanced NET formation, and tumor-promoting potential. Additionally, DMSO-dHL-60 cells demonstrated superior adhesion and migration abilities, likely due to increased expression of CD18 and CD31. Overall, different differentiation conditions shape the functional specialization of HL-60 cells, with ATRA promoting a more neutrophil-like maturation and moderate activation, while DMSO results in a more immature phenotype with enhanced NET formation. These distinct properties suggest that ATRA-dHL-60 cells may better model neutrophils in chronic inflammation, whereas DMSO-dHL-60 cells could be more suitable for studying NETosis-driven autoimmune, thrombotic disorders and cancer.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0331783","type":"journal-article","created":{"date-parts":[[2025,9,8]],"date-time":"2025-09-08T17:27:12Z","timestamp":1757352432000},"page":"e0331783","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":10,"title":["Unraveling the functional landscape of ATRA- and DMSO-differentiated HL-60 cells"],"prefix":"10.1371","volume":"20","author":[{"given":"Susanna","family":"Ghonyan","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Poghosyan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anush","family":"Martirosyan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sona","family":"Margaryan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aida","family":"Avetisyan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zaruhi","family":"Khachatryan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7541-5743","authenticated-orcid":true,"given":"Gayane","family":"Manukyan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2025,9,8]]},"reference":[{"key":"pone.0331783.ref001","doi-asserted-by":"crossref","DOI":"10.3389\/fphys.2018.00113","article-title":"Neutrophil: A Cell with Many Roles in Inflammation or Several Cell Types?","volume":"9","author":"C Rosales","year":"2018","journal-title":"Front Physiol"},{"issue":"1","key":"pone.0331783.ref002","doi-asserted-by":"crossref","first-page":"235","DOI":"10.1038\/s41392-024-01933-x","article-title":"Neutrophil extracellular traps in homeostasis and disease","volume":"9","author":"H Wang","year":"2024","journal-title":"Signal Transduct Target Ther"},{"issue":"4","key":"pone.0331783.ref003","doi-asserted-by":"crossref","first-page":"539","DOI":"10.1189\/jlb.3VMR1014-468R","article-title":"Neutrophil homeostasis and inflammation: novel paradigms from studying periodontitis","volume":"98","author":"G Hajishengallis","year":"2015","journal-title":"J Leukoc Biol"},{"issue":"1","key":"pone.0331783.ref004","doi-asserted-by":"crossref","first-page":"55","DOI":"10.1093\/rb\/rbw041","article-title":"An overview of the role of neutrophils in innate immunity, inflammation and host-biomaterial integration","volume":"4","author":"GS Selders","year":"2017","journal-title":"Regen Biomater"},{"issue":"1","key":"pone.0331783.ref005","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.immuni.2016.12.012","article-title":"Neutrophils in Homeostasis, Immunity, and Cancer","volume":"46","author":"J\u00c1 Nicol\u00e1s-\u00c1vila","year":"2017","journal-title":"Immunity"},{"key":"pone.0331783.ref006","article-title":"How Neutrophils Shape Adaptive Immune Responses","volume":"6","author":"PHC Leliefeld","year":"2015","journal-title":"Front Immunol"},{"key":"pone.0331783.ref007","doi-asserted-by":"crossref","first-page":"2456","DOI":"10.3389\/fimmu.2018.02456","article-title":"Understanding the Multifaceted Role of Neutrophils in Cancer and Autoimmune Diseases","volume":"9","author":"X Wang","year":"2018","journal-title":"Front Immunol"},{"issue":"18","key":"pone.0331783.ref008","doi-asserted-by":"crossref","first-page":"2173","DOI":"10.1182\/blood-2016-01-688887","article-title":"Neutrophil heterogeneity: implications for homeostasis and pathogenesis","volume":"127","author":"C Silvestre-Roig","year":"2016","journal-title":"Blood"},{"issue":"1","key":"pone.0331783.ref009","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1038\/s41392-024-02050-5","article-title":"Tissue-resident immune cells: from defining characteristics to roles in diseases","volume":"10","author":"J Li","year":"2025","journal-title":"Signal Transduct Target Ther"},{"key":"pone.0331783.ref010","doi-asserted-by":"crossref","first-page":"141","DOI":"10.2147\/JIR.S284941","article-title":"Studying Neutrophil Function in vitro: Cell Models and Environmental Factors","author":"M Blanter","year":"2021","journal-title":"JIR"},{"issue":"5635","key":"pone.0331783.ref011","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1038\/270347a0","article-title":"Continuous growth and differentiation of human myeloid leukaemic cells in suspension culture","volume":"270","author":"SJ Collins","year":"1977","journal-title":"Nature"},{"key":"pone.0331783.ref012","doi-asserted-by":"crossref","first-page":"30","DOI":"10.1186\/1471-2121-5-30","article-title":"Cytoskeletal influences on nuclear shape in granulocytic HL-60 cells","volume":"5","author":"AL Olins","year":"2004","journal-title":"BMC Cell Biol"},{"issue":"4","key":"pone.0331783.ref013","doi-asserted-by":"crossref","first-page":"413","DOI":"10.1111\/imcb.12015","article-title":"The influence of agents differentiating HL-60 cells toward granulocyte-like cells on their ability to release neutrophil extracellular traps","volume":"96","author":"A Manda-Handzlik","year":"2018","journal-title":"Immunol Cell Biol"},{"issue":"13","key":"pone.0331783.ref014","first-page":"3561","article-title":"Loss of amplified c-myc genes in the spontaneously differentiated HL-60 cells","volume":"54","author":"N Shimizu","year":"1994","journal-title":"Cancer Res"},{"issue":"2","key":"pone.0331783.ref015","doi-asserted-by":"crossref","first-page":"260","DOI":"10.1006\/bcmd.2002.0513","article-title":"Surface Antigen Changes during Normal Neutrophilic Development: A Critical Review","volume":"28","author":"MT Elghetany","year":"2002","journal-title":"Blood Cells Mol Dis"},{"issue":"2","key":"pone.0331783.ref016","doi-asserted-by":"crossref","DOI":"10.1016\/j.immuni.2018.02.002","article-title":"Developmental Analysis of Bone Marrow Neutrophils Reveals Populations Specialized in Expansion, Trafficking, and Effector Functions","volume":"48","author":"M Evrard","year":"2018","journal-title":"Immunity"},{"key":"pone.0331783.ref017","doi-asserted-by":"crossref","DOI":"10.7554\/eLife.84752","article-title":"CD31 signaling promotes the detachment at the uropod of extravasating neutrophils allowing their migration to sites of inflammation","volume":"12","author":"F Andreata","year":"2023","journal-title":"ELife"},{"issue":"7","key":"pone.0331783.ref018","doi-asserted-by":"crossref","first-page":"517","DOI":"10.1038\/nrm1155","article-title":"Calcium signalling: dynamics, homeostasis and remodelling","volume":"4","author":"MJ Berridge","year":"2003","journal-title":"Nat Rev Mol Cell Biol"},{"key":"pone.0331783.ref019","doi-asserted-by":"crossref","DOI":"10.3389\/fimmu.2021.602963","article-title":"Neutrophils: Need for Standardized Nomenclature","volume":"12","author":"E McKenna","year":"2021","journal-title":"Front Immunol"},{"issue":"2","key":"pone.0331783.ref020","doi-asserted-by":"crossref","first-page":"483","DOI":"10.1182\/blood.V100.2.483","article-title":"Granulocyte inducer C\/EBPalpha inactivates the myeloid master regulator PU.1: possible role in lineage commitment decisions","volume":"100","author":"VA Reddy","year":"2002","journal-title":"Blood"},{"issue":"3","key":"pone.0331783.ref021","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1002\/JLB.1RU1219-504RR","article-title":"Transcriptional regulation of neutrophil differentiation and function during inflammation","volume":"107","author":"Z Ai","year":"2020","journal-title":"J Leukoc Biol"},{"issue":"1","key":"pone.0331783.ref022","doi-asserted-by":"crossref","DOI":"10.1038\/s42003-022-03888-7","article-title":"PU.1-c-Jun interaction is crucial for PU.1 function in myeloid development","volume":"5","author":"X Zhao","year":"2022","journal-title":"Commun Biol"},{"issue":"18","key":"pone.0331783.ref023","doi-asserted-by":"crossref","first-page":"14502","DOI":"10.1074\/jbc.M111.329003","article-title":"The role of CD38 in Fc\u03b3 receptor (Fc\u03b3R)-mediated phagocytosis in murine macrophages","volume":"287","author":"J Kang","year":"2012","journal-title":"J Biol Chem"},{"issue":"11","key":"pone.0331783.ref024","doi-asserted-by":"crossref","first-page":"4091","DOI":"10.1128\/IAI.00340-13","article-title":"CD38 controls the innate immune response against Listeria monocytogenes","volume":"81","author":"T Lischke","year":"2013","journal-title":"Infect Immun"},{"issue":"3","key":"pone.0331783.ref025","doi-asserted-by":"crossref","first-page":"953","DOI":"10.1182\/blood-2010-06-290171","article-title":"Myeloperoxidase is required for neutrophil extracellular trap formation: implications for innate immunity","volume":"117","author":"KD Metzler","year":"2011","journal-title":"Blood"},{"key":"pone.0331783.ref026","doi-asserted-by":"crossref","first-page":"1024","DOI":"10.1016\/j.freeradbiomed.2015.10.398","article-title":"Neutrophil NET formation is regulated from the inside by myeloperoxidase-processed reactive oxygen species","volume":"89","author":"H Bj\u00f6rnsdottir","year":"2015","journal-title":"Free Radic Biol Med"},{"issue":"1","key":"pone.0331783.ref027","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1186\/s13045-021-01187-y","article-title":"Neutrophils in cancer carcinogenesis and metastasis","volume":"14","author":"S Xiong","year":"2021","journal-title":"J Hematol Oncol"},{"key":"pone.0331783.ref028","doi-asserted-by":"crossref","DOI":"10.3389\/fimmu.2020.01749","article-title":"The Emerging Role of Neutrophil Extracellular Traps (NETs) in Tumor Progression and Metastasis","volume":"11","author":"MT Masucci","year":"2020","journal-title":"Front Immunol"},{"issue":"7","key":"pone.0331783.ref029","doi-asserted-by":"crossref","first-page":"565","DOI":"10.1016\/j.it.2019.04.012","article-title":"Neutrophil Diversity in Health and Disease","volume":"40","author":"C Silvestre-Roig","year":"2019","journal-title":"Trends Immunol"},{"key":"pone.0331783.ref030","doi-asserted-by":"crossref","first-page":"117324","DOI":"10.1016\/j.biopha.2024.117324","article-title":"Phenotypes and functions of \u201caged\u201d neutrophils in cardiovascular diseases","volume":"179","author":"S Ling","year":"2024","journal-title":"Biomed Pharmacother"},{"issue":"7570","key":"pone.0331783.ref031","doi-asserted-by":"crossref","first-page":"528","DOI":"10.1038\/nature15367","article-title":"Neutrophil ageing is regulated by the microbiome","volume":"525","author":"D Zhang","year":"2015","journal-title":"Nature"},{"issue":"1","key":"pone.0331783.ref032","doi-asserted-by":"crossref","first-page":"5894","DOI":"10.1038\/s41467-023-41484-3","article-title":"CREB1-driven CXCR4hi neutrophils promote skin inflammation in mouse models and human patients","volume":"14","author":"J Chen","year":"2023","journal-title":"Nat Commun"},{"issue":"3","key":"pone.0331783.ref033","article-title":"Chronic stress increases metastasis via neutrophil-mediated changes to the microenvironment","volume":"42","author":"X-Y He","year":"2024","journal-title":"Cancer Cell"},{"issue":"2","key":"pone.0331783.ref034","article-title":"The Gut Microbiome Regulates Psychological-Stress-Induced Inflammation","volume":"53","author":"C Xu","year":"2020","journal-title":"Immunity"},{"issue":"10","key":"pone.0331783.ref035","doi-asserted-by":"crossref","first-page":"739","DOI":"10.1038\/nrc3581","article-title":"History of myeloid-derived suppressor cells","volume":"13","author":"JE Talmadge","year":"2013","journal-title":"Nat Rev Cancer"}],"container-title":["PLOS One"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0331783","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,9,8]],"date-time":"2025-09-08T17:27:18Z","timestamp":1757352438000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0331783"}},"subtitle":[],"editor":[{"given":"Ajit","family":"Prakash","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2025,9,8]]},"references-count":35,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2025,9,8]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0331783","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,9,8]]}}}