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. 2023 Jan;18(1):105-111.
doi: 10.1016/j.jds.2022.07.003. Epub 2022 Jul 15.

Intermittent compressive force regulates dentin matrix protein 1 expression in human periodontal ligament stem cells

Jeeranan Manokawinchoke  1   2 , Suconta Chareonvit  2 , Vorapat Trachoo  3 , Phoonsuk Limraksasin  1   2 , Hiroshi Egusa  4   5 , Thanaphum Osathanon  1   2
Affiliations

Intermittent compressive force regulates dentin matrix protein 1 expression in human periodontal ligament stem cells

Jeeranan Manokawinchoke et al. J Dent Sci. 2023 Jan.

Abstract

Background/purpose: Mechanical force differentially regulates periodontal ligament functions depending on types, magnitudes, and duration of stimulation. Intermittent compressive force (ICF) promotes an in vitro mineralization in human periodontal ligament cells. The present study investigated the effect of ICF on dentin matrix protein-1 (DMP1) expression in human periodontal ligament stem cells (hPDLSCs).

Materials and methods: Cells were treated with ICF in a serum-free culture medium for 24 h The mRNA and protein expression were examined using real-time polymerase chain reaction, immunofluorescence staining and Western blot analysis, respectively.

Results: The exposure to ICF in a serum-free condition significantly induced DMP1 expression in both mRNA and protein levels. The effect of ICF-induced DMP1 expression was inhibited by pretreatment with cycloheximide, indicating the requirement of the intermediated molecule(s). Pretreatment with transforming growth factor β (TGF-β) receptor inhibitor (SB431542) or neutralized antibody against TGF-β1 prior to ICF application abolished the effect of ICF-induced DMP1 expression. Further, recombinant TGF-β1 treatment stimulated DMP1 expression.

Conclusion: The present study illustrated that ICF induces DMP1 expression in hPDLSCs via the regulation of TGF-β signaling pathway.

Keywords: Compressive force; Dentin matrix protein; Periodontal ligament; Transforming growth factor.

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Figures

Figure 1
Figure 1
Characterization of hPDLSCs. hPDLSC morphology was shown (A). The percentage of mesenchymal stem cell surface marker expression, CD44, CD73, CD90, and CD105, and hematopoietic cell marker expression, CD45 in hPDLSCs was examined using flow cytometry analysis (B). Co-expression of CD44, CD73, CD90, and CD105 in hPDLSCs was illustrated (C–F). hPDLSCs were maintained in an osteogenic induction medium and an in vitro mineralization was determined on day 14 and 21 using alizarin red s staining (G).
Figure 2
Figure 2
ICF induced DMP1 expression in hPDLSCs. The experimental scheme was illustrated (A). hPDLSCs were exposed to ICF for 2, 4, 8, 24, and 48 h in serum-free condition. hPDLSC morphology was demonstrated using a phase-contrast microscope (B). DMP1 mRNA (C) and protein (D, E) expression were investigated by qPCR, immunofluorescence staining, and Western blot analysis, respectively. Bars indicated a significant difference between conditions (∗P < 0.05).
Figure 3
Figure 3
TGF-β pathway regulated ICF-induced DMP1 expression in hPDLSCs. The experimental scheme was illustrated (A). hPDLSCs were treated with the cycloheximide (CHX) (B), or SB431542 (a TGF-β inhibitor) (C), or neutralizing TGF-β1 antibody (D) 30 min prior to ICF application. After cells were exposed to ICF for 24 h, the DMP1 mRNA expression was examined using qPCR. Bars indicated a significant difference between conditions (∗P < 0.05, ∗∗P < 0.01).
Figure 4
Figure 4
Recombinant TGF-β1 protein promoted DMP1 expression in hPDLSCs. The experimental scheme was illustrated (A). hPDLSCs were exposed to recombinant human TGF-β1 protein at 1 and 10 ng/mL in serum-free culture condition for 24 h and then examined the DMP1 mRNA expression (B). Bars indicated a significant difference between conditions (∗P < 0.05).
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