Dataset Information

Effect of Tensile Frequency on the Osteogenic Differentiation of Periodontal Ligament Stem Cells.

ABSTRACT:

Purpose

The role of periodontal ligament stem cells (PDLSCs) in mediating osteogenesis involved in orthodontic tooth movement (OTM) is well established. However, various relevant in vitro studies vary in the frequency of tension. The effect of tensile frequency on the mechanotransduction of PDLSCs is not clear. The current study aimed to determine the effect of different tensile frequencies on the osteogenic differentiation of PDLSCs and to identify important mechano-sensitivity genes.

Methods

Human PDLSCs were isolated, identified, and subjected to cyclic equibiaxial tensile strain of 12% at different frequencies of 0.1 Hz, 0.5 Hz, 0.7 Hz, or static cultures. Osteogenic differentiation of PDLSCs was assessed by using Western blotting. High-throughput sequencing was used to identify differential mRNA expression. Short time-series expression miner (STEM) was utilized to describe the frequency patterns of the mRNAs. The functions and enriched pathways were identified, and the hub genes were identified and validated.

Results

We found that the osteoblastic differentiation capacity of PDLSCs increased with tensile frequency in the range of 0.1-0.7 Hz. Eight frequency-tendency gene expression profiles were identified to be statistically significant. Tensile frequency-specific expressed genes, such as SALL1 and EYA1, which decreased with the increase in tensile frequency, were found.

Conclusion

The osteoblastic differentiation of PDLSCs under mechanical tensile force is frequency dependent. EYA1 and SALL1 were identified as potential important tensile frequency-sensitive genes, which may contribute to the cyclic tension-induced osteogenic differentiation of PDLSCs in a frequency-dependent manner.

SUBMITTER: Wang W

PROVIDER: S-EPMC9259061 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:Long non-coding RNAs (lncRNAs) have been widely known to have an appreciable effect in physiology and pathology. In tooth regeneration, periodontal ligament stem cells (PDLSCs) are regarded as a key effector, whereas, how lncRNA acts in the osteogenic differentiation of PDLSCs have not been completely understood. This study aims to find out the relationship between lncRNA DANCR and the proliferation and osteogenic differentiation of PDLSCs. METHODS:Microarray was used to observe the different expression of lncRNAs in differentiated and undifferentiated PDLSCs. And then osteogenic-related lncRNA, DANCR was screened out. Its effects on proliferation and osteogenic differentiation was explored by constructing an overexpression and inhibition model. qRT-PCR was used to detect the mRNA expression of osteogenesis related genes. MTT assay was performed to assess the effects of DANCR on cell growth curve. To quantify the effects of DANCR on osteogenic differentiation of PDLSCs, ALP staining and alizarin red was performed in basic culture medium and osteogenic medium. Data were statistically processed. RESULTS:Compared with the undifferentiated PDLSCs, the alizarin red staining level was higher in differentiated PDLSCs. And the expressions of osteogenic differentiation marker genes Runt-related transcription factor 2 (Runx2), osteocalcin (OCN) and bone morphogenetic protein (BMP-2) were significantly increased in the differentiated PDLSCs. Furthermore, we noticed that comparing with control groups, the expression of lncRNA DANCR decreases markedly in osteogenically induced PDLSCs. DANCR promoted proliferation of PDLSCs, as evidenced by cell viability. Further investigation has proven that the downregulation of DANCR shows in the calcium sediment forming, alkaline phosphatase (ALP) activation and some osteogenic-related gene markers' upregulation including Runx2, OCN and BMP-2, which finally results in the osteogenic differentiation of PDLSCs following the transfection and induction. Conversely, DANCR upregulation was shown to repress the osteogenic differentiation potential of PDLSCs. CONCLUSIONS:The osteogenic differentiation of PDLSCs has proven to related to the down regulation of lncRNA DANCR. And this paper throws light on the effects of DANCR in the process of PDLSCs' osteogenic differentiation.

Project description:BackgroundAn increasing number of studies have shown that dysregulated miR-589-3p is associated with multiple diseases. However, the role of miR-589-3p in osteogenic differentiation of periodontal ligament stem cells (PDLSCs) remains unknown. This study aimed to explore the biological function and potential molecular mechanism of miR-589-3p in osteogenic differentiation of PDLSCs.MethodsGSE159508 was downloaded from Gene Expression Omibus (GEO, http://www.ncbi.nlm.nih.gov/geo/ ). Differentially expressed miRNAs between osteogenic induction PDLSCs versus non-induction PDLSCs were obtained by R software. miR-589-3p mimic and miR-589-3p inhibitor and corresponding negative control were obtained and to identify the role of miR-589-3p in osteogenic differentiation of PDLSCs. ALP staining and ARS were used to evaluate ALP activity and mineralization, respectively. The targeted binding relationship between miR-589-3p and ATF1 was predicted and verified by target prediction analysis and dual-luciferase assay. Furthermore, the functional mechanism based on miR-589-3p and ATF1 in osteogenic differentiation of PDLSCs was further investigated through rescue experiments.ResultsAccording to the cut-off criteria with log 2 FC > 1.0 and P < 0.05, 514 differentially expressed miRNAs were identified between osteogenic induction and non-induction PDLSCs, including 309 upregulated miRNAs and 205 downregulated miRNAs. Compared with control PDLSCs, miR-589-3p expression level was notably increased in PDLSCs that underwent osteogenic induction. The overexpression of miR-589-3p promoted the cell viability of PDLSCs, while the low expression of miR-589-3p had the opposite effect. The dual luciferase reporter assay verified that ATF1 was proved to be a direct target of miR-589-3p in PDLSCs. And overexpressed miR-589-3p reduced the expression of ATF1. Overexpression of miR-589-3p enhanced the osteogenic capacity of PDLSCs, as demonstrated by increases in ALP activity, matrix mineralization, and RUNX2, OCN and OSX expression. In addition, the rescue experiments confirmed that overexpressed ATF1 restored the effects of overexpressed miR-589-3p on cell proliferation and osteogenic differentiation of PDLSCs.ConclusionmiR-589-3p could down-regulate the expression of ATF1, thereby promote the proliferation and osteogenic differentiation of PDLSCs. This finding may provide a new therapeutic target for molecular therapy of periodontitis.