{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,23]],"date-time":"2026-07-23T16:31:40Z","timestamp":1784824300303,"version":"3.55.0"},"reference-count":46,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2022,1,26]],"date-time":"2022-01-26T00:00:00Z","timestamp":1643155200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"TUA grants from the County Council of V\u00e4sterbotten, Sweden","award":["7003193"],"award-info":[{"award-number":["7003193"]}]},{"name":"Fund for medical research, Ume\u00e5 university","award":["xxx"],"award-info":[{"award-number":["xxx"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pathogens"],"abstract":"<jats:p>Carriers of highly leukotoxic genotypes of Aggregatibacter actinomycetemcomitans are at high risk for rapid degradation of tooth-supporting tissues. The leukotoxin (LtxA) expressed by this bacterium induces a rapid pro-inflammatory response in leukocytes that results in cell death. The aim of the present study was to increase the understanding of LtxA-induced leukocyte activation mechanisms and of possible associated osteoclast differentiation. The effect of LtxA on activation of the inflammasome complex was studied in THP-1 wild type and in NLRP3- and ASC knockout cells. Cell-to-cell communication was assessed by fluorescent parachute assays, and THP-1 differentiation into osteoclast-like cells was investigated microscopically. The results showed that LtxA induced inflammatory cell death, which involved activation of the NLRP3 inflammasome and gap junction cell-to-cell communication. THP-1 cells treated with lipopolysaccharide (LPS) and LtxA together differentiated into an osteoclast-like phenotype. Here, LPS prevented LtxA-mediated cell death but failed to induce osteoclast differentiation on its own. However, pit formation was not significantly enhanced by LtxA. We conclude that A. actinomycetemcomitans leukotoxicity mediates activation of the NLRP3 inflammasome and cell-to-cell communication in the induced pro-inflammatory cell death. In addition, LtxA stimulated differentiation towards osteoclasts-like cells in LPS-treated THP-1 cells.<\/jats:p>","DOI":"10.3390\/pathogens11020159","type":"journal-article","created":{"date-parts":[[2022,1,27]],"date-time":"2022-01-27T04:49:51Z","timestamp":1643258991000},"page":"159","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":23,"title":["Aggregatibacter actinomycetemcomitans Leukotoxin Activates the NLRP3 Inflammasome and Cell-to-Cell Communication"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1594-1738","authenticated-orcid":false,"given":"Peyman","family":"Kelk","sequence":"first","affiliation":[{"name":"Department of Integrative Medical Biology, Ume\u00e5 University, 901 85 Ume\u00e5, Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nick Sina","family":"Moghbel","sequence":"additional","affiliation":[{"name":"Department of Odontology, Ume\u00e5 University, 901 87 Ume\u00e5, Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Josefine","family":"Hirschfeld","sequence":"additional","affiliation":[{"name":"Periodontal Research Group, Birmingham Dental School & Hospital, University of Birmingham, Birmingham B5 7EG, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anders","family":"Johansson","sequence":"additional","affiliation":[{"name":"Department of Odontology, Ume\u00e5 University, 901 87 Ume\u00e5, Sweden"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,1,26]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Belibasakis, G.N., Maula, T., Bao, K., Lindholm, M., Bostanci, N., Oscarsson, J., Ihalin, R., and Johansson, J. (2019). Virulence and Pathogenicity Properties of Aggregatibacter actinomycetemcomitans. Pathogens, 8.","DOI":"10.3390\/pathogens8040222"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"604","DOI":"10.1902\/jop.1989.60.11.604","article-title":"Prevalence of periodontal breakdown in adolescents and presence of Actinobacillus actinomycetemcomitans in subjects with attachment loss","volume":"60","author":"Abbas","year":"1989","journal-title":"J. Periodontol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"94","DOI":"10.1177\/0022034515608163","article-title":"Evolutionary Divergence of Aggregatibacter actinomycetemcomitans","volume":"95","author":"Kittichotirat","year":"2016","journal-title":"J. Dent. Res."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"501","DOI":"10.1128\/iai.62.2.501-508.1994","article-title":"Regulation of Actinobacillus actinomycetemcomitans leukotoxin expression: Analysis of the promoter regions of leukotoxic and minimally leukotoxic strains","volume":"62","author":"Brogan","year":"1994","journal-title":"Infect. Immun."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"237","DOI":"10.1016\/S0140-6736(08)60135-X","article-title":"Risk of aggressive periodontitis in adolescent carriers of the JP2 clone of Aggregatibacter (Actinobacillus) actinomycetemcomitans in Morocco: A prospective longitudinal cohort study","volume":"371","author":"Haubek","year":"2008","journal-title":"Lancet"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1111\/jcpe.12209","article-title":"Progression of attachment loss is strongly associated with presence of the JP2 genotype of Aggregatibacter actinomycetemcomitans: A prospective cohort study of a young adolescent population","volume":"41","author":"Kwamin","year":"2014","journal-title":"J. Clin. Periodontol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"e12967","DOI":"10.1111\/cmi.12967","article-title":"Aggregatibacter actinomycetemcomitans leukotoxin causes activation of lymphocyte function-associated antigen 1","volume":"21","author":"Nygren","year":"2019","journal-title":"Cell. Microbiol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"707096","DOI":"10.3389\/fimmu.2021.707096","article-title":"The Hunger Games: Aggregatibacter actinomycetemcomitans Exploits Human Neutrophils As an Epinephrine Source for Survival","volume":"12","author":"Ozuna","year":"2021","journal-title":"Front. Immunol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"18086","DOI":"10.1038\/s41598-021-97545-4","article-title":"Leukotoxin (LtxA\/Leukothera) induces ATP expulsion via pannexin-1 channels and subsequent cell death in malignant lymphocytes","volume":"11","author":"Prince","year":"2021","journal-title":"Sci. Rep."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"4448","DOI":"10.1128\/IAI.71.8.4448-4455.2003","article-title":"Caspase 1 involvement in human monocyte lysis induced by Actinobacillus actinomycetemcomitans leukotoxin","volume":"71","author":"Kelk","year":"2003","journal-title":"Infect. Immun."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1260","DOI":"10.1161\/CIRCRESAHA.120.315937","article-title":"Interleukin-1 and the Inflammasome as Therapeutic Targets in Cardiovascular Disease","volume":"126","author":"Abbate","year":"2020","journal-title":"Circ. Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"e126","DOI":"10.1038\/cddis.2011.6","article-title":"Cellular and molecular response of human macrophages exposed to Aggregatibacter actinomycetemcomitans leukotoxin","volume":"2","author":"Kelk","year":"2011","journal-title":"Cell Death Dis."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2324","DOI":"10.1182\/blood-2008-03-146720","article-title":"Differential requirement for the activation of the inflammasome for processing and release of IL-1beta in monocytes and macrophages","volume":"113","author":"Netea","year":"2009","journal-title":"Blood"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"242","DOI":"10.1016\/j.job.2020.07.003","article-title":"Role of interleukin-1 and inflammasomes in oral disease","volume":"62","author":"Murakami","year":"2020","journal-title":"J. Oral Biosci."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1111\/prd.12084","article-title":"The inflammasome and danger molecule signaling: At the crossroads of inflammation and pathogen persistence in the oral cavity","volume":"69","author":"Yilmaz","year":"2015","journal-title":"Periodontology 2000"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Sordi, M.B., Magini, R.d.S., Panahipour, L., and Gruber, R. (2022). Pyroptosis-Mediated Periodontal Disease. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23010372"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1016\/j.jdsr.2021.09.005","article-title":"Mechanism of alveolar bone destruction in periodontitis\u2014Periodontal bacteria and inflammation","volume":"57","author":"Usui","year":"2021","journal-title":"Jpn. Dent. Sci. Rev."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"821","DOI":"10.1016\/j.cell.2010.01.040","article-title":"The inflammasomes","volume":"140","author":"Schroder","year":"2010","journal-title":"Cell"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"110","DOI":"10.1016\/j.it.2011.01.003","article-title":"Inflammasome activation and IL-1\u03b2 and IL-18 processing during infection","volume":"32","author":"Netea","year":"2011","journal-title":"Trends Immunol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"124","DOI":"10.1016\/j.cyto.2012.03.016","article-title":"Aggregatibacter actinomycetemcomitans targets NLRP3 and NLRP6 inflammasome expression in human mononuclear leukocytes","volume":"59","author":"Belibasakis","year":"2012","journal-title":"Cytokine"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"752546","DOI":"10.3389\/fendo.2021.752546","article-title":"NLRP3 Inflammasome: A New Target for Prevention and Control of Osteoporosis?","volume":"12","author":"Jiang","year":"2021","journal-title":"Front. Endocrinol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"691013","DOI":"10.3389\/fimmu.2021.691013","article-title":"Inflammasomes in Alveolar Bone Loss","volume":"12","author":"Li","year":"2021","journal-title":"Front. Immunol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"929","DOI":"10.1080\/17435390.2020.1777476","article-title":"Multi-walled carbon nanotubes induce IL-1\u03b2 secretion by activating hemichannels-mediated ATP release in THP-1 macrophages","volume":"14","author":"Fan","year":"2020","journal-title":"Nanotoxicology"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"528","DOI":"10.1002\/jcb.20866","article-title":"Regulation of osteoclastogenesis by gap junction communication","volume":"99","author":"Matemba","year":"2006","journal-title":"J. Cell. Biochem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"2497","DOI":"10.1111\/j.1582-4934.2008.00275.x","article-title":"Gap junctional communication in human osteoclasts in vitro and in vivo","volume":"12","author":"Schilling","year":"2008","journal-title":"J. Cell. Mol. Med."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"8380","DOI":"10.3892\/mmr.2017.7625","article-title":"High-dose PMA with RANKL and MCSF induces THP-1 cell differentiation into human functional osteoclasts in vitro","volume":"16","author":"Li","year":"2017","journal-title":"Mol. Med. Rep."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"453","DOI":"10.1128\/IAI.73.1.453-458.2005","article-title":"Abundant secretion of bioactive interleukin-1beta by human macrophages induced by Actinobacillus actinomycetemcomitans leukotoxin","volume":"73","author":"Kelk","year":"2005","journal-title":"Infect. Immun."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1016\/bs.mie.2019.10.005","article-title":"Methods for macrophage differentiation and in vitro generation of human tumor associated-like macrophages","volume":"632","author":"Heideveld","year":"2020","journal-title":"Methods Enzymol."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Wang, W., Hu, D., Feng, Y., Wu, C., Song, Y., Liu, W., Li, A., Wang, Y., Chen, K., and Tian, M. (2020). Paxillin mediates ATP-induced activation of P2X7 receptor and NLRP3 inflammasome. BMC Biol., 18.","DOI":"10.1186\/s12915-020-00918-w"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Nice, J.B., Balashova, N.V., Kachlany, S.C., Koufos, E., Krueger, E., Lally, E.T., and Brown, A.C. (2018). Aggregatibacter actinomycetemcomitans Leukotoxin Is Delivered to Host Cells in an LFA-1-Indepdendent Manner When Associated with Outer Membrane Vesicles. Toxins, 10.","DOI":"10.3390\/toxins10100414"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Peng, L., Jiang, J., Chen, T., Xu, D., Hou, F., Huang, Q., Peng, Y., Ye, C., Hu, D.L., and Fang, R. (2021). Toxic Shock Syndrome Toxin 1 Induces Immune Response via the Activation of NLRP3 Inflammasome. Toxins, 13.","DOI":"10.3390\/toxins13010068"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"e12973","DOI":"10.1111\/cpr.12973","article-title":"NLRP3 regulates alveolar bone loss in ligature-induced periodontitis by promoting osteoclastic differentiation","volume":"54","author":"Chen","year":"2021","journal-title":"Cell Prolif."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"AlQranei, M.S., Senbanjo, L.T., Aljohani, H., Hamza, T., and Chellaiah, M.A. (2021). Lipopolysaccharide- TLR-4 Axis regulates Osteoclastogenesis independent of RANKL\/RANK signaling. BMC Immunol., 22.","DOI":"10.1186\/s12865-021-00409-9"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"232","DOI":"10.1002\/jlb.65.2.232","article-title":"Lipopolysaccharide can block the potential of monocytes to differentiate into dendritic cells","volume":"65","author":"Palucka","year":"1999","journal-title":"J. Leukoc. Biol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1016\/j.jss.2005.12.018","article-title":"Lipopolysaccharide transiently activates THP-1 cell adhesion","volume":"135","author":"Kounalakis","year":"2006","journal-title":"J. Surg. Res."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"546","DOI":"10.1016\/j.bbapap.2012.12.004","article-title":"Monodisperse and LPS-free Aggregatibacter actinomycetemcomitans leukotoxin: Interactions with human \u03b22 integrins and erythrocytes","volume":"1834","author":"Reinholdt","year":"2013","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1876","DOI":"10.1084\/jem.164.6.1876","article-title":"Adhesion-promoting receptors on human macrophages recognize Escherichia coli by binding to lipopolysaccharide","volume":"164","author":"Wright","year":"1986","journal-title":"J. Exp. Med."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3231","DOI":"10.1096\/fj.06-7579com","article-title":"Characterization of two novel LPS-binding sites in leukocyte integrin beta A domain","volume":"21","author":"Wong","year":"2007","journal-title":"FASEB J."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"67","DOI":"10.1046\/j.1365-2249.2003.02189.x","article-title":"Up-regulation of ICAM-1, CD11a\/CD18 and CD11c\/CD18 on human THP-1 monocytes stimulated by Streptococcus suis serotype 2","volume":"133","author":"Segura","year":"2003","journal-title":"Clin. Exp. Immunol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1006\/cimm.1994.1164","article-title":"Regulated expression of integrins and other adhesion molecules during differentiation of monocytes into macrophages","volume":"156","author":"Prieto","year":"1994","journal-title":"Cell. Immunol."},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Kitaura, H., Marahleh, A., Ohori, F., Noguchi, T., Shen, W.R., Qi, J., Nara, Y., Pramusita, A., Kinjo, R., and Mizoguchi, I. (2020). Osteocyte-Related Cytokines Regulate Osteoclast Formation and Bone Resorption. Int. J. Mol. Sci., 2.","DOI":"10.3390\/ijms21145169"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1034\/j.1399-302x.2000.150402.x","article-title":"Inhibition of Actinobacillus actinomycetemcomitans leukotoxicity by bacteria from the subgingival flora","volume":"15","author":"Johansson","year":"2000","journal-title":"Oral Microbiol. Immunol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1125","DOI":"10.1038\/nprot.2008.75","article-title":"Neutral red uptake assay for the estimation of cell viability\/cytotoxicity","volume":"3","author":"Repetto","year":"2008","journal-title":"Nat. Protoc."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"210","DOI":"10.3181\/00379727-90-21985","article-title":"Lactic dehydrogenase activity in blood","volume":"90","author":"Wroblewski","year":"1955","journal-title":"Proc. Soc. Exp. Biol. Med."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1006\/excr.1999.4760","article-title":"Gap-junctional coupling measured by flow cytometry","volume":"255","author":"Czyz","year":"2000","journal-title":"Exp. Cell Res."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"e1836","DOI":"10.21769\/BioProtoc.1836","article-title":"Pit Assay to Measure the Bone Resorptive Activity of Bone Marrow-derived Osteoclasts","volume":"6","author":"Vesprey","year":"2016","journal-title":"Bio-protocol"}],"container-title":["Pathogens"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-0817\/11\/2\/159\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T22:08:01Z","timestamp":1760134081000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-0817\/11\/2\/159"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,1,26]]},"references-count":46,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2022,2]]}},"alternative-id":["pathogens11020159"],"URL":"https:\/\/doi.org\/10.3390\/pathogens11020159","relation":{},"ISSN":["2076-0817"],"issn-type":[{"value":"2076-0817","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,1,26]]}}}