Pyroptosis, an inflammatory caspase-dependent programmed cell death, plays a vital role in maintaining tissue homeostasis and activating inflammatory responses. Orthodontic tooth movement (OTM) is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament (PDL) progenitor cells. However, whether and how force induces PDL progenitor cell pyroptosis, thereby influencing OTM and alveolar bone remodeling remains unknown. In this study, we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process. Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively. Using Caspase-1^-/- mice, we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1. Moreover, mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro, which influenced osteoclastogenesis. Mechanistically, transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells. Overall, this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli, indicating a promising approach to accelerate OTM by targeting Caspase-1.
Animals ; Humans ; Mice ; Rats ; Bone Remodeling/physiology* ; Caspase 1 ; Periodontal Ligament ; Pyroptosis ; Tooth Movement Techniques

OBJECTIVES: To investigate the effect of recombinant human fibroblast growth factor 21 (rhFGF21) on the proliferation and mineralization of cementoblasts and its mechanism. METHODS: Hematoxylin eosin, immunohistochemical staining, and immunofluorescence were used to detect the expression and distribution of fibroblast growth factor 21 (FGF21) in rat periodontal tissues and cementoblasts (OCCM-30), separately. Cell Counting Kit-8 was used to detect the proliferation of OCCM-30 under treatment with rhFGF21. Alkaline phosphatase staining and Alizarin Red staining were used to detect the mineralization state of OCCM-30 after 3 and 7 days of mineralization induction. The transcription and protein expression of the osteogenic-related genes Runx2 and Osterix were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot analysis. The expression levels of genes of transforming growth factor β (TGFβ)/bone morphogenetic protein (BMP) signaling pathway in OCCM-30 were detected through PCR array analysis. RESULTS: FGF21 was expressed in rat periodontal tissues and OCCM-30. Although rhFGF21 had no significant effect on the proliferation of OCCM-30, treatment with 50 ng/mL rhFGF21 could promote the mineralization of OCCM-30 cells after 7 days of mineralization induction. The transcriptional levels of Runx2 and Osterix increased significantly at 3 days of mineralization induction and decreased at 5 days of mineralization induction. Western blot analysis showed that the protein expression levels of Runx2 and Osterix increased during mineralization induction. rhFGF21 up-regulated Bmpr1b protein expression in cells. CONCLUSIONS rhFGF21 can promote the mineralization ability of OCCM-30. This effect is related to the activation of the TGFβ/BMP signaling pathway.
Humans ; Rats ; Animals ; Dental Cementum ; Core Binding Factor Alpha 1 Subunit/metabolism* ; Cell Differentiation ; Bone Morphogenetic Proteins/metabolism* ; Transforming Growth Factor beta/pharmacology*

OBJECTIVES: This study aimed to investigate how naringenin (Nar) affected the anti-inflammatory, vascula-rization, and osteogenesis differentiation of human periodontal ligament stem cells (hPDLSCs) stimulated by lipopolysaccharide (LPS) and to preliminarily explore the underlying mechanism. METHODS: Cell-counting kit-8 (CCK8), cell scratch test, and Transwell assay were used to investigate the proliferation and migratory capabilities of hPDLSCs. Alkaline phosphatase (ALP) staining, alizarin red staining, lumen-formation assay, enzyme-linked immunosorbent assay, quantitative timed polymerase chain reaction, and Western blot were used to measure the expression of osteopontin (OPN), Runt-related transcription factor 2 (RUNX2), vascular endothlial growth factor (VEGF), basic fibroblast growth factor (bFGF), von Willebrand factor (vWF), tumor necrosis factor-α (TNF-α), and interleukin (IL)-6. RESULTS: We observed that 10 μmol/L Nar could attenuate the inflammatory response of hPDLSCs stimulated by 10 μg/mL LPS and promoted their proliferation, migration, and vascularization differentiation. Furthermore, 0.1 μmol/L Nar could effectively restore the osteogenic differentiation of inflammatory hPDLSCs. The effects of Nar's anti-inflammatory and promotion of osteogenic differentiation significantly decreased and inflammatory vascularization differentiation increased after adding AMD3100 (a specific CXCR4 inhibitor). CONCLUSIONS Nar demonstrated the ability to promote the anti-inflammatory, vascularization, and osteogenic effects of hPDLSCs stimulated by LPS, and the ability was associated with the stromal cell-derived factor/C-X-C motif chemokine receptor 4 signaling axis.
Humans ; Anti-Inflammatory Agents/pharmacology* ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Chemokine CXCL12 ; Lipopolysaccharides/pharmacology* ; Osteogenesis ; Periodontal Ligament/metabolism* ; Receptors, Chemokine/metabolism* ; Stem Cells ; Interleukin-8/metabolism*

OBJECTIVES: This work aimed to investigate the molecular mechanism of cyclic tensile stress (CTS) stimulating autophagy in human periodontal ligament cells (hPDLCs). METHODS: hPDLCs were isolated and cultured from normal periodontal tissues. hPDLCs were loaded with tensile stress by force four-point bending extender to simulate the autophagy of hPDLCs induced by orthodontic force du-ring orthodontic tooth movement. XMU-MP-1 was used to inhibit the Hippo signaling pathway to explore the role of the Hippo-YAP signaling pathway in activating hPDLC autophagy by tensile stress. The expression levels of autophagy-related genes (Beclin-1, LC3, and p62) in hPDLCs were detected by real-time quantitative polymerase chain reaction. Western blot was used to detect the expression levels of autophagy-related proteins (Beclin-1, LC3-Ⅱ/LC3-Ⅰ, and p62) and Hippo-YAP pathway proteins (active-YAP and p-YAP) in hPDLCs. Immunofluorescence was used to locate autophagy-related proteins (LC3-Ⅱand p62) and Hippo-YAP pathway proteins (active-YAP) of hPDLCs. RESULTS: CTS-activated autophagy in hPDLCs and expression of autophagy-related proteins initially increased and then decreased; it began to increase at 30 min, peaked at 3 h, and decreased (P<0.05). CTS increased the expression of active-YAP protein and decreased the expression of p-YAP protein (P<0.05). When XMU-MP-1 inhibited the Hippo-YAP signaling pathway (P<0.05), active-YAP protein was promoted to enter the nucleus and autophagy expression was enhanced (P<0.05). CONCLUSIONS The Hippo-YAP signaling pathway is involved in the regulation of autophagy activation in hPDLCs under CTS.
Humans ; Hippo Signaling Pathway ; Periodontal Ligament/metabolism* ; Beclin-1/metabolism* ; Cells, Cultured ; Autophagy

OBJECTIVES: This study aimed to clarify the effects of Foxp3 silencing on the expression of inflammatory cytokines in human periodontal ligament cells (hPDLFs) in an inflammatory environment and on cell proliferation and invasiveness, as well as to explore the role of Foxp3 gene in the development of periodontitis. METHODS: An small interfering RNA (siRNA) construct specific for Foxp3 was transfected into hPDLFs. Foxp3 silencing efficiency was verified by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, and the siRNA with the optimum silencing effect of Foxp3 gene was screened. Using lipopolysaccharide to simulate an inflammatory environment in vitro, CCK-8 detected the effect of silencing Foxp3 on hPDLFs proliferation under inflammatory conditions. Wound-healing experiments and transwell assays were conducted to detect the effect of silencing Foxp3 on hPDLF migration under inflammatory conditions. The expression of the inflammatory cytokines interleukin (IL)-6 and IL-8 was detected by RT-PCR and Western blotting under inflammatory conditions. RESULTS: After siRNA transfection, RT-PCR and Western blotting analyses showed that the expression of Foxp3 mRNA in the Foxp3-si3 group decreased significantly (t=21.03, P<0.000 1), and the protein expression of Foxp3 also decreased significantly (t=12.8, P<0.001). In the inflammatory environment, Foxp3 gene silencing had no significant effect on hPDLFs proliferation (P>0.05), and Foxp3 gene silencing promoted hPDLFs migration (P<0.05). Moreover, the expression of IL-6 and IL-8 increased (P<0.05). CONCLUSIONS In an inflammatory environment, Foxp3 gene silencing promoted hPDLFs migration but had no significant effect on hPDLFs proliferation. The expression of inflammatory factors expressed in hPDLFs increased after Foxp3 gene silencing, indicating that Foxp3 gene inhibited inflammation in periodontitis.
Humans ; Cell Proliferation/genetics* ; Cells, Cultured ; Cytokines/metabolism* ; Fibroblasts/metabolism* ; Forkhead Transcription Factors/metabolism* ; Gene Silencing ; Interleukin-6/metabolism* ; Interleukin-8/metabolism* ; Periodontal Ligament/metabolism* ; Periodontitis/metabolism* ; RNA, Small Interfering/metabolism* ; Transcription Factors/metabolism*

In the orthodontics process, intervention and sliding of an orthodontic bracket during the orthodontic process can arise large response of the labio-cheek soft tissue. Soft tissue damage and ulcers frequently happen at the early stage of orthodontic treatment. In the field of orthodontic medicine, qualitative analysis is always carried out through statistics of clinical cases, while quantitative explanation of bio-mechanical mechanism is lacking. For this purpose, finite element analysis of a three-dimensional labio-cheek-bracket-tooth model is conducted to quantify the bracket-induced mechanical response of the labio-cheek soft tissue, which involves complex coupling of contact nonlinearity, material nonlinearity and geometric nonlinearity. Firstly, based on the biological composition characteristics of labio-cheek, a second-order Ogden model is optimally selected to describe the adipose-like material of the labio-cheek soft tissue. Secondly, according to the characteristics of oral activity, a two-stage simulation model of bracket intervention and orthogonal sliding is established, and the key contact parameters are optimally set. Finally, the two-level analysis method of overall model and submodel is used to achieve efficient solution of high-precision strains in submodels based on the displacement boundary obtained from the overall model calculation. Calculation results with four typical tooth morphologies during orthodontic treatment show that: ① the maximum strain of soft tissue is distributed along the sharp edges of the bracket, consistent with the clinically observed profile of soft tissue deformation; ② the maximum strain of soft tissue is reduced as the teeth align, consistent with the clinical manifestation of common damage and ulcers at the beginning of orthodontic treatment and reduced patient discomfort at the end of treatment. The method in this paper can provide reference for relevant quantitative analysis studies in the field of orthodontic medical treatment at home and abroad, and further benefit to the product development analysis of new orthodontic devices.
Humans ; Periodontal Ligament/physiology* ; Orthodontic Wires ; Cheek ; Ulcer ; Tooth ; Finite Element Analysis

Periodontitis is a complex chronic inflammatory disease. The invasion of pathogens induces the inflammatory microenvironment in periodontitis. Cell behavior changes in response to changes in the microenvironment, which in turn alters the local inflammatory microenvironment of the periodontium through factors secreted by cells. It has been confirmed that periodontal ligament stem cells (PDLSCs) are vital in the development of periodontal disease. Moreover, PDLSCs are the most effective cell type to be used for periodontium regeneration. This review focuses on changes in PDLSCs, their basic biological behavior, osteogenic differentiation, and drug effects caused by the inflammatory microenvironment, to provide a better understanding of the influence of these factors on periodontal tissue homeostasis. In addition, we discuss the underlying mechanism in detail behind the reciprocal responses of PDLSCs that affect the microenvironment.
Humans ; Periodontal Ligament ; Osteogenesis ; Stem Cells ; Periodontitis/metabolism* ; Cell Differentiation/physiology* ; Cells, Cultured

OBJECTIVE: To analyze the cement flow in the abutment margin-crown platform switching structure by using the three-dimensional finite element analysis, in order to prove that whether the abutment margin-crown platform switching structure can reduce the inflow depth of cement in the implantation adhesive retention. METHODS: By using ANSYS 19.0 software, two models were created, including the one with regular margin and crown (Model one, the traditional group), and the other one with abutment margin-crown platform switching structure (Model two, the platform switching group). Both abutments of the two models were wrapped by gingiva, and the depth of the abutment margins was 1.5 mm submucosal. Two-way fluid structure coupling calculations were produced in two models by using ANSYS 19.0 software. In the two models, the same amount of cement were put between the inner side of the crowns and the abutments. The process of cementing the crown to the abutment was simulated when the crown was 0.6 mm above the abutment. The crown was falling at a constant speed in the whole process spending 0.1 s. Then we observed the cement flow outside the crowns at the time of 0.025 s, 0.05 s, 0.075 s, 0.1 s, and measured the depth of cement over the margins at the time of 0.1 s. RESULTS: At the time of 0 s, 0.025 s, 0.05 s, the cements in the two models were all above the abutment margins. At the time of 0.075 s, in Model one, the gingiva was squeezed by the cement and became deformed, and then a gap was formed between the gingiva and the abutment into which the cement started to flow. In Model two, because of the narrow neck of the crown, the cement flowed out from the gingival as it was pressed by the upward counterforce from the gingival and the abutment margin. At the time of 0.1 s, in Model one, the cement continued to flow deep inside with the gravity force and pressure, and the depth of the cement over the margin was 1 mm. In Model two, the cement continued to flow out from the gingival at the time of 0.075 s, and the depth of the cement over the margin was 0 mm. CONCLUSION When the abutment was wrapped by the gingiva, the inflow depth of cement in the implantation adhesive retention can be reduced in the abutment margin-crown platform switching structure.
Finite Element Analysis ; Cementation/methods* ; Gingiva ; Crowns ; Dental Abutments ; Dental Cements ; Dental Stress Analysis

Fusobacterium nucleatum (F. nucleatum) is an early pathogenic colonizer in periodontitis, but the host response to infection with this pathogen remains unclear. In this study, we built an F. nucleatum infectious model with human periodontal ligament stem cells (PDLSCs) and showed that F. nucleatum could inhibit proliferation, and facilitate apoptosis, ferroptosis, and inflammatory cytokine production in a dose-dependent manner. The F. nucleatum adhesin FadA acted as a proinflammatory virulence factor and increased the expression of interleukin(IL)-1β, IL-6 and IL-8. Further study showed that FadA could bind with PEBP1 to activate the Raf1-MAPK and IKK-NF-κB signaling pathways. Time-course RNA-sequencing analyses showed the cascade of gene activation process in PDLSCs with increasing durations of F. nucleatum infection. NFκB1 and NFκB2 upregulated after 3 h of F. nucleatum-infection, and the inflammatory-related genes in the NF-κB signaling pathway were serially elevated with time. Using computational drug repositioning analysis, we predicted and validated that two potential drugs (piperlongumine and fisetin) could attenuate the negative effects of F. nucleatum-infection. Collectively, this study unveils the potential pathogenic mechanisms of F. nucleatum and the host inflammatory response at the early stage of F. nucleatum infection.
Humans ; Fusobacterium nucleatum/metabolism* ; NF-kappa B/metabolism* ; Periodontal Ligament/metabolism* ; Signal Transduction ; Fusobacterium Infections/pathology* ; Stem Cells/metabolism*

In dentistry, orthodontic root resorption is a long-lasting issue with no effective treatment strategy, and its mechanisms, especially those related to senescent cells, remain largely unknown. Here, we used an orthodontic intrusion tooth movement model with an L-loop in rats to demonstrate that mechanical stress-induced senescent cells aggravate apical root resorption, which was prevented by administering senolytics (a dasatinib and quercetin cocktail). Our results indicated that cementoblasts and periodontal ligament cells underwent cellular senescence (p21⁺ or p16⁺) and strongly expressed receptor activator of nuclear factor-kappa B (RANKL) from day three, subsequently inducing tartrate-resistant acid phosphatase (TRAP)-positive odontoclasts and provoking apical root resorption. More p21⁺ senescent cells expressed RANKL than p16⁺ senescent cells. We observed only minor changes in the number of RANKL⁺ non-senescent cells, whereas RANKL⁺ senescent cells markedly increased from day seven. Intriguingly, we also found cathepsin K⁺p21⁺p16⁺ cells in the root resorption fossa, suggesting senescent odontoclasts. Oral administration of dasatinib and quercetin markedly reduced these senescent cells and TRAP⁺ cells, eventually alleviating root resorption. Altogether, these results unveil those aberrant stimuli in orthodontic intrusive tooth movement induced RANKL⁺ early senescent cells, which have a pivotal role in odontoclastogenesis and subsequent root resorption. These findings offer a new therapeutic target to prevent root resorption during orthodontic tooth movement.
Rats ; Animals ; Root Resorption/prevention & control* ; Senotherapeutics ; Stress, Mechanical ; Dasatinib/pharmacology* ; Quercetin/pharmacology* ; Osteoclasts ; Tooth Movement Techniques ; Periodontal Ligament ; RANK Ligand