General Control Nonrepressed Protein 5 Modulates Odontogenic Differentiation Through NF-κB Pathway in Tumor Necrosis Factor-α-Mediated Impaired Human Dental Pulp Stem Cells.

Dental pulp stem cells (DPSCs) from pulpitis patients showed defective osteogenic differentiation. However, as the most well-studied histone acetyltransferase, the impaired general control nonrepressed protein 5 (GCN5) plays essential roles in various developmental processes. The aim of this study was to investigate the effect of GCN5 on DPSCs odontogenic differentiation. The healthy dental pulp tissues were obtained from the extracted impacted third molar of patients with the informed consent. DPSCs were treated with a high concentration of tumor necrosis factor-alpha (TNF-α) (100 ng/mL) and odontogenic differentiation-related gene and GCN5 protein level by Western blot analysis. Proliferation of the DPSCs was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Immunofluorescence staining detected GCN5 and NF-κB signaling for p-p65. The mechanism of GCN5 regulating odontogenic differentiation of DPSCs was determined by small interfering RNA analysis. Our data suggested that TNF-α can significantly reduce mineralization and the expression of dentin matrix acidic phosphoprotein 1 and dentin sialophosphoprotein at higher concentration (100 ng/mL). Meanwhile, it showed that the inflammation in microenvironment resulted in a downregulation of GCN5 expression and GCN5 knockdown caused decreased odontogenic differentiation of DPSCs was also found. In addition, the knockdown of GCN5 increased the expression of phosphorylation of p65, thus activating NF-κB pathway of DPSCs. Meanwhile, NF-κB pathway inhibitor pyrrolidinedithiocarbamic acid reversed the siGCN5 decreased odontogenic differentiation of DPSCs. Altogether, our findings indicated that in inflammatory microenvironments GCN5 plays a protective role in pulpitis impaired odontogenic differentiation of DPSCs by activating NF-κB pathway, which may provide a potential approach to dentin regeneration.

MicroRNA-125a-3p participates in odontoblastic differentiation of dental pulp stem cells by targeting Fyn.

Fyn is a member of the protein tyrosine kinase family and its overexpression is associated with various types of inflammation. MicroRNAs can regulate the expression of target genes and play an important role in varied physiological and pathological processes. Based on the important role of Fyn and microRNA-125a-3p (miR-125a-3p) in inflammation, and combined with the bioinformatics studies, we performed in this study and chose miR-125a-3p as the focus of our research. During the progression of inflammation, we found that the expression of miR-125a-3p was decreased while the expression of Fyn was up-regulated. Fyn formed a complex with Neuropilin-1, which inhibited odontoblastic differentiation and expanded inflammatory responses through nuclear factor-κB signal pathways in dental pulp stem cells (DPSCs). These findings suggested that miR-125a-3p plays an important role in odontoblastic differentiation of DPSCs by targeting Fyn, implying its therapeutic potential in dental caries.

Dysfunction of MiR-148a-NRP1 Functional Axis Suppresses Osteogenic Differentiation of Periodontal Ligament Stem Cells Under Inflammatory Microenvironment.

Periodontitis is a chronic inflammatory disease that can lead to the loss of periodontal bone tissue. The osteogenic potential of periodontal ligament stem cells (PDLSCs) is significantly decreased in periodontitis microenvironment. However, the mechanism is still unclear. We used Porphyromonas gingivalis lipopolysaccharide (LPS) as a stimulator of PDLSCs to mimic the periodontal inflammatory environment. The mineralization capability was restrained in LPS-stimulated PDLSCs, and the level of miR-148a increased, while the level of Neuropilin 1 (NRP1) decreased. Downregulation of miR-148a could reverse the osteogenesis deficiency of PDLSCs under LPS treatment. In addition, the expression of miR-148a in PDLSCs was negatively correlated with the expression of NRP1. Furthermore, overexpression of NRP1 upregulated the osteogenesis ability of LPS-stimulated PDLSCs, while inhibition of NRP1 eliminated the stimulative effect of miR-148a inhibitor on osteogenic differentiation. These data illustrated that the inflammatory environment mimicked by LPS inhibits osteogenesis by upregulation of miR-148a and subsequent downregulation of NRP1. We also found, compared to healthy periodontal tissues, miR-148a level increased, while NRP1 level decreased in periodontitis tissues. These two phenomena also exist in PDLSCs that come from the upper two types of tissues. To summarize, the decline of osteogenic potential of PDLSCs under inflammatory condition of periodontitis is related to miR-148a/NRP1 functional axis. This study may provide a novel strategy in the molecular aspect for the therapy of periodontitis.

Comparison of immunomodulatory properties of exosomes derived from bone marrow mesenchymal stem cells and dental pulp stem cells.

Substantial discoveries suggested that exosomes released from multiple sources of stem cells can affect the biological functions of target cells. In present period, the immunosuppressive properties of exosomes derived from bone marrow mesenchymal stem cells (BMMSCs-E) have been extensively recognized, but few studies have been reported about exosomes secreted from dental pulp stem cells (DPSCs-E) in the field of medical immunity. Hence, the aim of this study is to compare the immunomodulatory capacity of BMMSCs-E and DPSCs-E. Peripheral blood mononuclear cells (PBMCs) were co-cultured with them respectively and the proportion of regulatory T cells (Treg) was detected to increase. Subsequently, we stimulated CD4+T cells with BMMSCs-E and DPSCs-E to observe their effects on the polarizations, chemokines secretion, apoptosis, and proliferation of CD4+T cells. We found that DPSCs-E inhibited the differentiation of CD4+T cells into T helper 17 cells (Th17) and reduced the secretions of pro-inflammatory factors IL-17 and TNF-α, while promoted the polarization of CD4+T cells into Treg and increased the release of anti-inflammatory factors IL-10 and TGF-ß. What's more, these capabilities of DPSCs-E were stronger than those of BMMSCs-E. In addition, DPSCs-E were more effective in inducing apoptosis of CD4+T cells compared with BMMSCs-E, and DPSCs-E inhibited the proliferation of CD4+T cells, which is similar to BMMSCs-E. We draw a conclusion that DPSCs-E have stronger immune-modulating activities than BMMSCs-E, and may be a new therapeutic tool for the treatment of immunological diseases.

Exosomal microRNA-155-5p from PDLSCs regulated Th17/Treg balance by targeting sirtuin-1 in chronic periodontitis.

The mechanism of local inflammation and systemic injury in chronic periodontitis is complicated, in which and exosomes play an important role. In our study, we found that T helper cell 17 (Th17)/regulatory T cell (Treg) balance is destabilized in the peripheral blood of patients with periodontitis, with upregulated Th17 or downregulated Treg, respectively. Porphyromonas gingivalis lipopolysaccharide (LPS) was used to simulate the inflammatory microenvironment of chronic periodontitis. The exosomes were extracted from periodontal ligament stem cells (PDLSCs) in LPS-induced periodontitis environment, which inversely effected on CD4+ T cells under normal and inflammatory conditions. Furthermore, compared with exosomes from normal PDLSCs, lower expression of microRNA-155-5p (miR-155-5p) and higher expression of Sirtuin-1 (SIRT1) were observed in exosomes from LPS-stimulated PDLSCs. Exosomes from PDLSCs alleviated inflammatory microenvironment through Th17/Treg/miR-155-5p/SIRT1 regulatory network. This study aimed to find the "switching" factors that affected the further deterioration of periodontitis to maximally control the multiple downstream damage signal factors to further understand periodontitis and find new targets for its treatment.