{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,13]],"date-time":"2026-06-13T01:51:21Z","timestamp":1781315481851,"version":"3.54.1"},"reference-count":54,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,1,21]],"date-time":"2025-01-21T00:00:00Z","timestamp":1737417600000},"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. Oncol."],"abstract":"Introduction<\/jats:title>The role of miRNAs in regulating variable molecular functions has been sought by scientists for its promising utility in regulating the immune response and, hence, in treating various diseases. In hepatocellular carcinoma (HCC) specifically, a reduction in the number and efficiency of circulating and intrahepatic natural killer (NK) cells has been reported. Our project aims to investigate the role of miR-216a-3p<\/jats:italic> in the regulation of NK cell cytotoxicity, especially since it plays a tumor suppressor role in the context of HCC.<\/jats:p><\/jats:sec>Methods<\/jats:title>To achieve our aim, we isolated NK cells from the whole blood of 86 patients with HCC and 23 healthy controls. We assessed the expression profile of miR-216a-3p<\/jats:italic> in NK cells of patients and controls. Furthermore, we induced the expression of miR-216a-3p<\/jats:italic> in NK cells isolated from healthy controls, followed by measuring the release of interferon-gamma (IFN-\u03b3), tumor necrosis factor-alpha (TNF-\u03b1), perforins (PRF) and granzyme B (GrB) using ELISA as well as NK cells cytolytic activity against Huh7 cells using lactate dehydrogenase (LDH) cytotoxicity assay. After that, we performed an in silico<\/jats:italic> analysis to understand the mechanistic regulation imposed by miR-216a-3p<\/jats:italic> on NK cells to study its impact on one of its potential downstream targets.<\/jats:p><\/jats:sec>Results<\/jats:title>Our results have indicated that miR-216a-3p<\/jats:italic> has higher expression in NK cells of patients with HCC, and simulating this elevated expression pattern via forcing miR-216a-3p<\/jats:italic> expression in normal NK cells has negatively impacted the release of TNF- \u03b1, IFN- \u03b3, GrB, and PRF. Consequently, a decrease in cell cytolysis was observed. Our in silico<\/jats:italic> analysis revealed that the predicted downstream targets of miR-216a-3p<\/jats:italic> are enriched in the FOXO-signaling pathway. Among those targets is FOXO-1, which has been reported to play a role in NK cell maturation. Thus, we evaluated FOXO-1 expression upon mimicking miR-216a-3p<\/jats:italic> in control NK cells that showed significant downregulation of FOXO-1 on both RNA and protein levels.<\/jats:p><\/jats:sec>Conclusion<\/jats:title>In conclusion, we report miR-216-3p<\/jats:italic> as a negative regulator of NK cell cytotoxicity.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fonc.2024.1523068","type":"journal-article","created":{"date-parts":[[2025,1,21]],"date-time":"2025-01-21T08:39:00Z","timestamp":1737448740000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":5,"title":["MiR-216a-3p inhibits the cytotoxicity of primary natural killer cells"],"prefix":"10.3389","volume":"14","author":[{"given":"Rowan","family":"Abdelbary","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manon","family":"Ragheb","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shereen A.","family":"El Sobky","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nagwa","family":"El-Badri","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nourhan","family":"Aboud","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed","family":"Tawheed","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Asmaa","family":"Gomaa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mona","family":"Zidan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ramy K.","family":"Aziz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Abd Elrahman","family":"Abouzid","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Radwa Ayman","family":"Salah","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohamed","family":"El-Kassas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Imam","family":"Waked","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed","family":"Moustafa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Injie Omar","family":"Fawzy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nada","family":"El-Ekiaby","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed Ihab","family":"Abdelaziz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2025,1,21]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2021.655697","article-title":"Hepatocellular carcinoma immune landscape and the potential of immunotherapies","volume":"12","author":"Giraud","year":"2021","journal-title":"Front Immunol"},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.1182\/blood-2015-12-629055","article-title":"Exploitation of natural killer cells for the treatment of acute leukemia","volume":"127","author":"Handgretinger","year":"2016","journal-title":"Blood"},{"key":"B3","doi-asserted-by":"publisher","DOI":"10.1080\/2162402X.2015.1119354","article-title":"Dual targeting of glioblastoma with chimeric antigen receptor-engineered natural killer cells overcomes heterogeneity of target antigen expression and enhances antitumor activity and survival","volume":"5","author":"Gen\u00dfler","year":"2016","journal-title":"Oncoimmunology"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1038\/nri3174","article-title":"Targeting natural killer cells and natural killer T cells in cancer","volume":"12","author":"Vivier","year":"2012","journal-title":"Nat Rev Immunol"},{"key":"B5","article-title":"Transfusion medicine and hemotherapy: offizielles Organ der Deutschen Gesellschaft fur\u00f6 Transfusionsmedizin und Immunham\u00f6atologie","author":"Karger","year":"2003"},{"key":"B6","doi-asserted-by":"publisher","first-page":"6837241","DOI":"10.1155\/2016\/6837241","article-title":"Dual effects of cellular immunotherapy in inhibition of virus replication and prolongation of survival in HCV-positive hepatocellular carcinoma patients","volume":"2016","author":"Qian","year":"2016","journal-title":"J Immunol Res"},{"key":"B7","doi-asserted-by":"publisher","first-page":"77","DOI":"10.1016\/j.ygyno.2024.04.001","article-title":"Adoptive T cell therapy for ovarian cancer","volume":"186","author":"Gitto","year":"2024","journal-title":"Gynecol Oncol"},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.1080\/2162402X.2016.1250050","article-title":"Targeting Ewing sarcoma with activated and GD2-specific chimeric antigen receptor-engineered human NK cells induces upregulation of immune-inhibitory HLA-G","volume":"6","author":"Kailayangiri","year":"2017","journal-title":"Oncoimmunology.>"},{"key":"B9","doi-asserted-by":"publisher","DOI":"10.1038\/ni1582","article-title":"Functions of natural killer cells","volume":"9","author":"Vivier","year":"2008","journal-title":"Nat Immunol"},{"key":"B10","doi-asserted-by":"publisher","DOI":"10.1002\/ijc.2910160204","article-title":"Natural cytotoxic reactivity of mouse lymphoid cells against syngeneic acid allogeneic tumors. I. Distribution of reactivity and specificity","volume":"16","author":"Herberman","year":"1975","journal-title":"Int J Cancer"},{"key":"B11","doi-asserted-by":"publisher","DOI":"10.1002\/eji.1830050208","article-title":"Natural\u201d killer cells in the mouse. I. Cytotoxic cells with specificity for mouse Moloney leukemia cells. Specificity and distribution according to genotype","volume":"5","author":"Kiessling","year":"1975","journal-title":"Eur J Immunol"},{"key":"B12","doi-asserted-by":"publisher","DOI":"10.1038\/icb.2010.78","article-title":"Activating and inhibitory receptors of natural killer cells","volume":"89","author":"Pegram","year":"2011","journal-title":"Immunol Cell Biol"},{"key":"B13","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2013.00459\/full","article-title":"Natural killer cells modulation in hematological Malignancies","volume":"4","author":"Baier","year":"2013","journal-title":"Front Immunol"},{"key":"B14","doi-asserted-by":"publisher","DOI":"10.1016\/j.clim.2008.08.012","article-title":"Functional impairment in circulating and intrahepatic NK cells and relative mechanism in hepatocellular carcinoma patients","volume":"129","author":"Cai","year":"2008","journal-title":"Clin Immunol Orlando Fla"},{"key":"B15","first-page":"17","article-title":"Diminished absolute counts of CD56dim and CD56bright natural killer cells in peripheral blood from Egyptian patients with hepatocellular carcinoma","volume":"16","author":"Fathy","year":"2009","journal-title":"Egypt J Immunol"},{"key":"B16","doi-asserted-by":"publisher","DOI":"10.1002\/hep.26192","article-title":"Monocyte\/macrophage-elicited natural killer cell dysfunction in hepatocellular carcinoma is mediated by CD48\/2B4 interactions","volume":"57","author":"Wu","year":"2013","journal-title":"Hepatol Baltim Md"},{"key":"B17","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2023.1284669\/full","article-title":"Characterizing hepatocellular carcinoma stem markers and their corresponding susceptibility to NK-cell based immunotherapy","volume":"14","author":"Chiang","year":"2023","journal-title":"Front Immunol"},{"key":"B18","doi-asserted-by":"publisher","first-page":"799","DOI":"10.1002\/hep.23054","article-title":"Myeloid derived suppressor cells inhibit natural killer cells in patients with hepatocellular carcinoma via the NKp30 receptor","volume":"50","author":"Hoechst","year":"2009","journal-title":"Hepatol Baltim Md"},{"key":"B19","doi-asserted-by":"publisher","DOI":"10.1016\/j.jhep.2005.05.026","article-title":"Impairment of natural killer cell and dendritic cell functions by the soluble form of MHC class I-related chain A in advanced human hepatocellular carcinomas","volume":"43","author":"Jinushi","year":"2005","journal-title":"J Hepatol"},{"key":"B20","doi-asserted-by":"publisher","first-page":"77","DOI":"10.1016\/j.chom.2008.05.013","article-title":"Position-dependent function for a tandem microRNA miR-122-binding site located in the hepatitis C virus RNA genome","volume":"4","author":"Jopling","year":"2008","journal-title":"Cell Host Microbe"},{"key":"B21","doi-asserted-by":"publisher","DOI":"10.1080\/08977194.2016.1200571","article-title":"Contradicting interplay between insulin-like growth factor-1 and miR-486-5p in primary NK cells and hepatoma cell lines with a contemporary inhibitory impact on HCC tumor progression","volume":"34","author":"Youness","year":"2016","journal-title":"Growth Factors Chur Switz"},{"key":"B22","doi-asserted-by":"publisher","first-page":"52","DOI":"10.1016\/j.imlet.2015.10.013","article-title":"Contradicting roles of miR-182 in both NK cells and their host target hepatocytes in HCV","volume":"169","author":"El Sobky","year":"2016","journal-title":"Immunol Lett"},{"key":"B23","doi-asserted-by":"publisher","first-page":"76","DOI":"10.1080\/08977194.2017.1354859","article-title":"MiR-615-5p depresses natural killer cells cytotoxicity through repressing IGF-1R in hepatocellular carcinoma patients","volume":"35","author":"Rahmoon","year":"2017","journal-title":"Growth Factors Chur Switz"},{"key":"B24","doi-asserted-by":"publisher","DOI":"10.1016\/j.humimm.2016.04.020","article-title":"Enhancing NK cell cytotoxicity by miR-182 in hepatocellular carcinoma","volume":"77","author":"Abdelrahman","year":"2016","journal-title":"Hum Immunol"},{"key":"B25","doi-asserted-by":"publisher","first-page":"326","DOI":"10.3390\/ijms17030326","article-title":"Natural killer cells\u2014An epigenetic perspective of development and regulation","volume":"17","author":"Schenk","year":"2016","journal-title":"Int J Mol Sci"},{"key":"B26","doi-asserted-by":"publisher","DOI":"10.1016\/j.clim.2016.03.003","article-title":"Transcriptional and post-transcriptional regulation of NK cell development and function","volume":"177","author":"Leong","year":"2017","journal-title":"Clin Immunol Orlando Fla"},{"key":"B27","doi-asserted-by":"publisher","DOI":"10.1039\/d4na00488d","article-title":"Revolutionizing cancer therapy: nanoformulation of miRNA-34 \u2013 enhancing delivery and efficacy for various cancer immunotherapies: a review","volume":"6","author":"Paula Fawzy M","year":"2024","journal-title":"Nanoscale Adv"},{"key":"B28","doi-asserted-by":"publisher","DOI":"10.1007\/s13277-015-4506-8","article-title":"miR-615-5p prevents proliferation and migration through negatively regulating serine hydromethyltransferase 2 (SHMT2) in hepatocellular carcinoma","volume":"37","author":"Wu","year":"2016","journal-title":"Tumour Biol J Int Soc Oncodevelopmental Biol Med"},{"key":"B29","doi-asserted-by":"publisher","DOI":"10.1016\/j.febslet.2012.06.054","article-title":"miR-615-5p is restrictedly expressed in cirrhotic and cancerous liver tissues and its overexpression alleviates the tumorigenic effects in hepatocellular carcinoma","volume":"586","author":"El Tayebi","year":"2012","journal-title":"FEBS Lett"},{"key":"B30","doi-asserted-by":"publisher","first-page":"19","DOI":"10.1186\/s12935-023-02865-2","article-title":"Molecular mechanisms of microRNA-216a during tumor progression","volume":"23","author":"Hamidi","year":"2023","journal-title":"Cancer Cell Int"},{"key":"B31","doi-asserted-by":"publisher","first-page":"6874","DOI":"10.1111\/jcmm.v25.14","article-title":"Establishment and experimental validation of an immune miRNA signature for assessing prognosis and immune landscape of patients with colorectal cancer","volume":"25","author":"Liu","year":"2021","journal-title":"J Cell Mol Med"},{"key":"B32","unstructured":"OncomiR: Database search"},{"key":"B33","doi-asserted-by":"publisher","DOI":"10.1002\/hep.26369","article-title":"MicroRNA-216a\/217-induced epithelial-mesenchymal transition targets PTEN and SMAD7 to promote drug resistance and recurrence of liver cancer","volume":"58","author":"Xia","year":"2013","journal-title":"Hepatology.>"},{"key":"B34","doi-asserted-by":"publisher","DOI":"10.1002\/jcb.v119.2","article-title":"MiR-216a-3p inhibits colorectal cancer cell proliferation through direct targeting COX-2 and ALOX5","volume":"119","author":"Wang","year":"2018","journal-title":"J Cell Biochem"},{"key":"B35","doi-asserted-by":"publisher","DOI":"10.3727\/096504017X15031557924150","article-title":"miR-216a-3p inhibits the proliferation, migration, and invasion of human gastric cancer cells via targeting RUNX1 and activating the NF-\u03baB signaling pathway","volume":"26","author":"Wu","year":"2018","journal-title":"Oncol Res"},{"key":"B36","doi-asserted-by":"publisher","first-page":"13636","DOI":"10.3390\/ijms222413636","article-title":"Multi-omics reveal the immunological role and the theragnostic value of miR-216a\/GDF15 axis in human colon adenocarcinoma","volume":"22","author":"Tung","year":"2021","journal-title":"Int J Mol Sci"},{"key":"B37","doi-asserted-by":"publisher","DOI":"10.18632\/aging.103670","article-title":"MicroRNA-216a-3p promotes sorafenib sensitivity in hepatocellular carcinoma by downregulating MAPK14 expression","volume":"12","author":"Wan","year":"2020","journal-title":"Aging"},{"key":"B38","doi-asserted-by":"publisher","first-page":"103288","DOI":"10.1016\/j.xpro.2024.103288","article-title":"Protocol for isolating high-purity natural killer cells from peripheral blood of septic patients using magnetic cell separation","volume":"5","author":"Coulibaly","year":"2024","journal-title":"STAR Protoc"},{"key":"B39","doi-asserted-by":"publisher","DOI":"10.1046\/j.1365-2567.1997.00174.x","article-title":"Secretion of cytokines by natural killer cells primed with interleukin-2 and stimulated with different lipoproteins","volume":"90","author":"De Sanctis","year":"1997","journal-title":"Immunology"},{"key":"B40","doi-asserted-by":"publisher","first-page":"3036","DOI":"10.1038\/s41598-019-39211-4","article-title":"Efficient cell penetration and delivery of peptide nucleic acids by an argininocalix[4]arene","volume":"9","author":"Gasparello","year":"2019","journal-title":"Sci Rep"},{"key":"B41","doi-asserted-by":"publisher","first-page":"18","DOI":"10.1186\/s13059-019-1629-z","article-title":"Prediction of functional microRNA targets by integrative modeling of microRNA binding and target expression data","volume":"20","author":"Liu","year":"2019","journal-title":"Genome Biol"},{"key":"B42","unstructured":""},{"key":"B43","doi-asserted-by":"publisher","first-page":"11023","DOI":"10.1038\/ncomms11023","article-title":"FoxO1-mediated autophagy is required for NK cell development and innate immunity","volume":"7","author":"Wang","year":"2016","journal-title":"Nat Commun"},{"key":"B44","doi-asserted-by":"publisher","DOI":"10.3892\/or.2018.6727","article-title":"Upregulation of microRNA\u2212300 induces the proliferation of liver cancer by downregulating transcription factor FOXO1","volume":"40","author":"Chang","year":"2018","journal-title":"Oncol Rep"},{"key":"B45","doi-asserted-by":"publisher","DOI":"10.18632\/oncotarget.13786","article-title":"FOXO1 inhibits the invasion and metastasis of hepatocellular carcinoma by reversing ZEB2-induced epithelial-mesenchymal transition","volume":"8","author":"Dong","year":"2017","journal-title":"Oncotarget"},{"key":"B46","doi-asserted-by":"publisher","DOI":"10.1002\/mc.22286","article-title":"OncomiR miR-96 and miR-182 promote cell proliferation and invasion through targeting ephrinA5 in hepatocellular carcinoma","volume":"55","author":"Wang","year":"2016","journal-title":"Mol Carcinog"},{"key":"B47","doi-asserted-by":"publisher","first-page":"128","DOI":"10.3892\/mmr.2023.13015","article-title":"hsa\u2212miR\u2212216a\u22123p regulates cell proliferation in oral cancer via the Wnt3a\/\u03b2\u2212catenin pathway","volume":"27","author":"Wang","year":"2023","journal-title":"Mol Med Rep"},{"key":"B48","doi-asserted-by":"publisher","DOI":"10.1002\/jcp.v235.12","article-title":"MiR-216a-3p suppresses the proliferation and invasion of cervical cancer through downregulation of ACTL6A-mediated YAP signaling","volume":"235","author":"Zhao","year":"2020","journal-title":"J Cell Physiol"},{"key":"B49","doi-asserted-by":"publisher","DOI":"10.26355\/eurrev_201811_16410","article-title":"MiR-216a-3p promotes differentiation of BMMSCs into ACE II cells via Wnt\/\u03b2-catenin pathway","volume":"22","author":"Song","year":"2018","journal-title":"Eur Rev Med Pharmacol Sci"},{"key":"B50","doi-asserted-by":"publisher","DOI":"10.1002\/JLB.MR0718-269R","article-title":"Mechanisms of natural killer cell-mediated cellular cytotoxicity","volume":"105","author":"Prager","year":"2019","journal-title":"J Leukoc Biol"},{"key":"B51","doi-asserted-by":"publisher","DOI":"10.1038\/cddis.2017.364","article-title":"MicroRNA-519a-3p mediates apoptosis resistance in breast cancer cells and their escape from recognition by natural killer cells","volume":"8","author":"Breunig","year":"2017","journal-title":"Cell Death Dis"},{"key":"B52","doi-asserted-by":"publisher","first-page":"222","DOI":"10.1186\/1471-2148-9-222","article-title":"FoxO gene family evolution in vertebrates","volume":"9","author":"Wang","year":"2009","journal-title":"BMC Evol Biol"},{"key":"B53","doi-asserted-by":"publisher","DOI":"10.1038\/ni.1689","article-title":"Foxo1 links homing and survival of naive T cells by regulating L-selectin, CCR7 and interleukin 7 receptor","volume":"10","author":"Kerdiles","year":"2009","journal-title":"Nat Immunol"},{"key":"B54","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2015.02.006","article-title":"Transcription factor Foxo1 is a negative regulator of natural killer cell maturation and function","volume":"42","author":"Deng","year":"2015","journal-title":"Immunity"}],"container-title":["Frontiers in Oncology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fonc.2024.1523068\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,1,21]],"date-time":"2025-01-21T08:39:03Z","timestamp":1737448743000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fonc.2024.1523068\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,1,21]]},"references-count":54,"alternative-id":["10.3389\/fonc.2024.1523068"],"URL":"https:\/\/doi.org\/10.3389\/fonc.2024.1523068","relation":{},"ISSN":["2234-943X"],"issn-type":[{"value":"2234-943X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,1,21]]},"article-number":"1523068"}}