{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T15:25:36Z","timestamp":1783524336904,"version":"3.55.0"},"reference-count":35,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2020,7,30]],"date-time":"2020-07-30T00:00:00Z","timestamp":1596067200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["81874018"],"award-info":[{"award-number":["81874018"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Orthopaedic Surgery"],"published-print":{"date-parts":[[2020,8]]},"abstract":"<jats:sec><jats:title>Objective<\/jats:title><jats:p>Iron plays a significant role in multiple biological processes. The purpose of this study was to measure whether iron mediated osteoclast differentiation through regulation of triggering receptor expressed in myeloid cells\u20102 (Trem\u20102) expression and the PI3K\/Akt signaling pathway.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>The effects of six different concentrations of ferric ammonium citrate (FAC) (100, 80, 40, 20, 10 and 0 \u03bcmol\/L) on RAW 264.7 cells proliferation were assessed by Cell Counting Kit\u20108 (CCK\u20108) gassay. Tartrate resistant acid phosphatase (TRAP) assay was performed to detect the effects of FAC on osteoclast formation. The expression of osteoclast differentiation\u2010related (TRAP, NFATc\u20101, and c\u2010Fos) and Trem\u20102 mRNA and proteins was analyzed by reverse transcription\u2010polymerase chain reaction and western blot, respectively. Si\u2010Trem\u20102 was constructed and transfected to RAW264.7 to measure the effects of Trem\u20102 on FAC\u2010mediated osteoclast formation. TRAP assay and osteoclast differentiation\u2010related gene analyses were further performed to identify the role of Trem\u20102 in osteoclastogenesis. The Search Tool for the Retrieval of Interacting Genes (STRING) was used to explore the target genes of Trem\u20102. Trem\u20102\u2010related gene ontology and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway were used for further in\u2010depth analysis. PI3K\/Akt pathway\u2010related proteins were detected by immunofluorescence and western blot.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>In groups with FAC concentration of 10 (102.5\u2009\u00b1\u20093.1), 20 (100.5\u2009\u00b1\u20091.5), and 40 \u03bcmol\/L (98.7\u2009\u00b1\u20093.1), compared with the control group (100.1\u2009\u00b1\u20092.2), cell viability was not significantly different from the control (<jats:italic>P<\/jats:italic>\u2009&gt;\u20090.05). When the concentration of FAC exceeded 80 \u03bcmol\/L, cell viability was significantly decreased (87.5\u2009\u00b1\u20092.8 <jats:italic>vs<\/jats:italic> 100.1\u2009\u00b1\u20092.2, <jats:italic>P<\/jats:italic>\u2009&lt;\u20090.05). FAC promotes Trem\u20102 expression and osteoclast differentiation in a dose\u2010response manner (<jats:italic>P<\/jats:italic>\u2009&lt;\u20090.05). The number of osteoclast\u2010like cells was found to be reduced following transfection with the siRNA of Trem\u20102 (42\u2009\u00b1\u20093 <jats:italic>vs<\/jats:italic> 30\u2009\u00b1\u20095, <jats:italic>P<\/jats:italic>\u2009&lt;\u20090.05). We observed that most of Trem\u20102 target genes are primarily involved in response to organic substance, regulation of reactive oxygen species metabolic process, and regulation of protein phosphorylation. The STRING database revealed that Trem\u20102 directly target two gene nodes (Pik3ca and Pik3r1), which are key transcriptional cofactors of the PI3K\/Akt signaling pathway. KEGG pathways include the \u201cPI3K\u2010Akt signaling pathway,\u201d the \u201cthyroid hormone signaling pathway\u201d, \u201cprostate cancer,\u201d the \u201clongevity regulating pathway,\u201d and \u201cinsulin resistance.\u201d Expression of p\u2010PI3K and p\u2010Akt protein, measured by immunofluorescence and western blotting, was markedly increased in the FAC groups. Trem\u20102 siRNA caused partial reduction of these two proteins (p\u2010PI3K and p\u2010Akt) compared to the FAC alone group.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusion<\/jats:title><jats:p>The FAC promoted osteoclast differentiation through the Trem\u20102\u2010mediated PI3K\/Akt signaling pathway. However, its regulation osteoclastogenesis should be verified through further <jats:italic>in vivo<\/jats:italic> studies.<\/jats:p><\/jats:sec>","DOI":"10.1111\/os.12750","type":"journal-article","created":{"date-parts":[[2020,7,30]],"date-time":"2020-07-30T07:35:24Z","timestamp":1596094524000},"page":"1304-1312","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["Ferric Ion Induction of Triggering Receptor Expressed in Myeloid Cells\u20102 Expression and <scp>PI3K<\/scp>\/Akt Signaling Pathway in Preosteoclast Cells to Promote Osteoclast Differentiation"],"prefix":"10.1111","volume":"12","author":[{"given":"Lu\u2010lin","family":"Liu","sequence":"first","affiliation":[{"name":"Department of Orthopaedics The Second Affiliated Hospital of Soochow University  Suzhou China"},{"name":"Department of Orthopaedics The First Affiliated Hospital of Gannan Medical University  Ganzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zi\u2010hou","family":"Cao","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The Second Affiliated Hospital of Soochow University  Suzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chun\u2010lei","family":"He","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The First Affiliated Hospital of Gannan Medical University  Ganzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yan\u2010chun","family":"Zhong","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The First Affiliated Hospital of Gannan Medical University  Ganzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wu\u2010yang","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The First Affiliated Hospital of Gannan Medical University  Ganzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peng","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The Second Affiliated Hospital of Soochow University  Suzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fan","family":"Yang","sequence":"additional","affiliation":[{"name":"Osteoporosis Institute of Soochow University  Suzhou China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4255-850X","authenticated-orcid":false,"given":"You\u2010jia","family":"Xu","sequence":"additional","affiliation":[{"name":"Department of Orthopaedics The Second Affiliated Hospital of Soochow University  Suzhou China"},{"name":"Osteoporosis Institute of Soochow University  Suzhou 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