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The aim of this study was to investigate the osteogenic effect of low\u2010intensity pulsed US on human periodontal ligament cells and to determine whether bone morphogenetic protein (BMP)\u2010Smad signaling was involved.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>Human periodontal ligament cells were exposed to low\u2010intensity pulsed US at a frequency of 1.5 MHz and intensity of 90 mW\/cm<jats:sup>2<\/jats:sup> for 20 min\/d. Osteogenic differentiation was determined by assaying alkaline phosphatase (ALP) and calcium deposition. Expression of BMP\u20102, BMP\u20106, and BMP\u20109 was detected by real\u2010time polymerase chain reaction analysis. Phosphorylated Smad was detected by western blotting; Smad in the cells was labeled by an immunofluorescent antibody and observed by laser\u2010scanning confocal microscopy.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>The optical density of ALP stimulated by US at 1.5 MHz and 90 mW\/cm<jats:sup>2<\/jats:sup> for 20 min\/d was significantly higher than in other groups (<jats:italic>P<\/jats:italic> &lt; .01); therefore, this dosage was considered optimal for promoting osteogenic differentiation. After 13 days of US exposure, ALP increased gradually after 5 days, peaked at 11 days, and decreased at 13 days, with a significant difference compared with the control group (<jats:italic>P<\/jats:italic> &lt; .05). Osteocalcin production increased from 9 to 13 days and peaked at 15 days, with a significant difference compared with the control group (<jats:italic>P<\/jats:italic> &lt; .05). BMP\u20102 and BMP\u20106 increased dynamically after exposure for 13 days. BMP\u20102 increased 6.07\u2010fold at 3 days, 6.39\u2010fold at 11 days, and 5.97\u2010fold at 13 days. BMP\u20106 expression increased 6.82\u2010fold at 1 day and 51.5\u2010fold at 3 days and decreased thereafter. BMP\u20109 was not expressed. Phospho\u2010Smad1\/5\/8 expression was significantly increased after exposure (<jats:italic>P<\/jats:italic>&lt; .05) and transferred from the cytoplasm into the nuclei.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>Low\u2010intensity pulsed US effectively induced osteogenic differentiation of human periodontal ligament cells, and the BMP\u2010Smad signaling pathway was involved in the mechanism.<\/jats:p><\/jats:sec>","DOI":"10.7863\/ultra.33.5.865","type":"journal-article","created":{"date-parts":[[2014,4,24]],"date-time":"2014-04-24T20:44:17Z","timestamp":1398372257000},"page":"865-873","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":34,"title":["Low\u2010Intensity Pulsed Ultrasound Induces Osteogenic Differentiation of Human Periodontal Ligament Cells Through Activation of Bone Morphogenetic Protein\u2013Smad 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