Effects of low-intensity pulsed ultrasound on the differentiation of C2C12 cells.
Low-intensity pulsed ultrasound (LIPUS) is known to accelerate bone regeneration, but the precise cellular mechanism is still unclear. The purpose of this study was to determine the effect of LIPUS on the differentiation of pluripotent mesenchymal cell line C2C12. The cells were cultured in differentiation medium with or without the addition of LIPUS stimulation. The ultrasound signal consisted of 1.5 MHz at an intensity of 70 mW/cm2 for 20 min for all cultures. To verify the cell lineage after LIPUS stimulation, mRNA expression of cellular phenotype-specific markers characterizing osteoblasts (Runx2, Msx2, Dlx5, AJ18), chondroblasts (Sox9), myoblasts (MyoD), and adipocytes (C/EBP, PPARgamma) was studied using real-time polymerase chain reaction analysis. The protein expression of Runx2 and activated phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (p38 MAPK) were performed using Western blotting. The mRNA expression of Runx2, Msx2, Dlx5, AJ18, and Sox9 was increased markedly by the LIPUS stimulation, whereas the expression of MyoD, C/EBP, and PPARgamma was drastically decreased. In the Western blot analysis, LIPUS stimulation increased Runx2 protein expression and phosphorylation of ERK1/2 and p38 MAPK. Our study demonstrated that LIPUS stimulation converts the differentiation pathway of C2C12 cells into the osteoblast and/or chondroblast lineage via activated phosphorylation of ERK1/2 and p38 MAPK.
Ikeda
Kyoko
K
Department of Periodontology, Nihon University School of Dentistry, Tokyo, Japan.
Takayama
Tadahiro
T
Suzuki
Naoto
N
Shimada
Koichi
K
Otsuka
Kichibee
K
Ito
Koichi
K
D000818:Animals; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics / Q000528:radiation effects; D002460:Cell Line; D019070:Cell Lineage; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D050659:Core Binding Factor Alpha 1 Subunit / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000528:radiation effects; D008648:Mesoderm / Q000166:cytology* / Q000378:metabolism / Q000528:radiation effects*; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000032:analysis / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000032:analysis / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000032:analysis / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D010766:Phosphorylation; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D014465:Ultrasonics; D048051:p38 Mitogen-Activated Protein Kinases / Q000032:analysis / Q000378:metabolism
D015415:Biomarkers; D050659:Core Binding Factor Alpha 1 Subunit; D012333:RNA, Messenger; C493716:Runx2 protein, mouse; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases