Glycometabolic reprogramming in cementoblasts: A vital target for enhancing cell mineralization.

Cementum, a constituent part of periodontal tissues, has important adaptive and reparative functions. It serves to attach the tooth to alveolar bone and acts as a barrier delimit epithelial growth and bacteria evasion. A dynamic and highly responsive cementum is essential for maintaining occlusal relationships and the integrity of the root surface. It is a thin layer of mineralized tissue mainly produced by cementoblasts. Cementoblasts are osteoblast-like cells essential for the restoration of periodontal tissues. In recent years, glucose metabolism has been found to be critical in bone remodeling and osteoblast differentiation. However, the glucose metabolism of cementoblasts remains incompletely understood. First, immunohistochemistry staining and in vivo tracing with 18 F-fluorodeoxyglucose (18 F-FDG) revealed significantly higher glucose metabolism in cementum formation. To test the bioenergetic pathways of cementoblast differentiation, we compared the bioenergetic profiles of mineralized and unmineralized cementoblasts. As a result, we observed a significant increase in the consumption of glucose and production of lactate, coupled with the higher expression of glycolysis-related genes. However, the expression of oxidative phosphorylation-related genes was downregulated. The verified results were consistent with the RNA sequencing results. Likewise, targeted energy metabolomics shows that the levels of glycolytic metabolites were significantly higher in the mineralized cementoblasts. Seahorse assays identified an increase in glycolytic flux and reduced oxygen consumption during cementoblast mineralization. Apart from that, we also found that lactate dehydrogenase A (LDHA), a key glycolysis enzyme, positively regulates the mineralization of cementoblasts. In summary, cementoblasts mainly utilized glycolysis rather than oxidative phosphorylation during the mineralization process.

Mitochondrial Dysfunction in Periodontitis and Associated Systemic Diseases: Implications for Pathomechanisms and Therapeutic Strategies.

Periodontitis is a chronic infectious disorder damaging periodontal tissues, including the gingiva, periodontal ligament, cementum, and alveolar bone. It arises from the complex interplay between pathogenic oral bacteria and host immune response. Contrary to the previous view of "energy factories", mitochondria have recently been recognized as semi-autonomous organelles that fine-tune cell survival, death, metabolism, and other functions. Under physiological conditions, periodontal tissue cells participate in dynamic processes, including differentiation, mineralization, and regeneration. These fundamental activities depend on properly functioning mitochondria, which play a crucial role through bioenergetics, dynamics, mitophagy, and quality control. However, during the initiation and progression of periodontitis, mitochondrial quality control is compromised due to a range of challenges, such as bacterial-host interactions, inflammation, and oxidative stress. Currently, mounting evidence suggests that mitochondria dysfunction serves as a common pathological mechanism linking periodontitis with systemic conditions like type II diabetes, obesity, and cardiovascular diseases. Therefore, targeting mitochondria to intervene in periodontitis and multiple associated systemic diseases holds great therapeutic potential. This review provides advanced insights into the interplay between mitochondria, periodontitis, and associated systemic diseases. Moreover, we emphasize the significance of diverse therapeutic modulators and signaling pathways that regulate mitochondrial function in periodontal and systemic cells.

Irisin attenuates P. gingivalis-suppressed osteogenic/cementogenic differentiation of periodontal ligament cells via p38 signaling pathway.

Regeneration of periodontal hard tissues damaged by Porphyromonas gingivalis (P. gingivalis) is essential for tooth stability and dental health. Irisin, a myokine secreted by skeletal muscle, is involved in different biological processes, such as myogenesis, adipogenesis, neurogenesis and osteogenesis. However, whether irisin regulates the osteogenic/cementogenic differentiation of human periodontal ligament cell (hPDLCs), especially under P. gingivalis-triggered inflammation, remains unknown. In this study, we verified the suppression role of P. gingivalis in the osteogenic/cementogenic differentiation of hPDLCs. Also, compared with the control cells, hPDLCs with irisin stimulation showed higher expression of osteogenic-/cementogenic-related markers, ALP activity and mineralization ability, as measured by RT-qPCR, western blotting, ALP staining and Alizarin red staining, respectively. Moreover, the osteogenic/cementogenic differentiation-facilitating role of irisin was also demonstrated under P. gingivalis-elicited inflammation, which implied a rescue function of irisin in P. gingivalis-suppressed hPDLC differentiation. Finally, the underlying mechanism involved in the process was explored. We observed that the p38 signaling pathway was activated during irisin-accelerated hPDLC differentiation. Furthermore, hPDLC differentiation was weakened after the p38 inhibitor was applied. In summary, we found that irisin can facilitate the osteogenic/cementogenic differentiation of hPDLCs partially through the p38 signaling pathway, which may provide evidence for the regeneration of P. gingivalis-destroyed periodontal hard tissues.

Using 3D Medical Modeling to Evaluate the Accuracy of Single-Photon Emission Computed Tomography (SPECT) Bone Scintigraphy in Diagnosing Condylar Hyperplasia.

PURPOSE: To evaluate the accuracy of single-photon emission computed tomography (SPECT) in diagnosing unilateral condylar hyperplasia (UCH) and to describe the condylar growth of patients with UCH. MATERIALS AND METHODS: Using a retrospective study design, patients with UCH who had undergone SPECT and cone-beam computed tomography (CBCT) examinations at the same time were included in the study. We used 3D medical models based on CBCT data as the gold standard. The SPECT results were compared with the model data, and the sensitivity and specificity were calculated. To further describe the condylar growth activity, statistical analysis was performed, and the P value was set at 0.05. RESULTS: The sample was composed of 75 patients. The sensitivity of SPECT was 55.3%, the specificity was 48.6%, and the area under the receiver operating characteristic curve was 0.53. There was no significant difference in sex between patients with and without active growth. CONCLUSION: The sensitivity and specificity of SPECT are poor, and SPECT alone is not suitable for evaluating the active stage of condylar growth. 3D medical modeling has good prospects for application in the diagnosis of condylar hyperplasia.

Platr3/NUDT21/NF-κB Axis Mediates P. gingivalis-Suppressed Cementoblast Mineralization.

Porphyromonas gingivalis (P. gingivalis) is one of the major pathogens causing periodontitis and apical periodontitis (AP). Long noncoding RNA (lncRNA) can regulate cellular mineralization and inflammatory diseases. The aim of this study was to investigate the role and mechanism of lncRNA in P. gingivalis-stimulated cementoblast mineralization. In vivo, C57BL/6 mice were divided into the healthy, the AP, and AP + P. gingivalis groups (n = six mice per group). Micro computed tomography, immunohistochemistry staining, and fluorescence in situ hybridization were used to observe periapical tissue. In vitro, cementoblasts were treated with osteogenic medium or P. gingivalis. Pluripotency associated transcript 3 (Platr3), interleukin 1 beta (IL1B), and osteogenic markers were analyzed by quantitative real-time polymerase chain reaction and western blot. RNA pull-down and RNA immunoprecipitation assays were used to detect proteins that bind to Platr3. RNA sequencing was performed in Platr3-silenced cementoblasts. In vivo, P. gingivalis promoted periapical tissue destruction and IL1B expression, but inhibited Platr3 expression. In vitro, P. gingivalis facilitated IL1B expression (P < 0.001), whereas suppressed the expression of Platr3 (P < 0.001) and osteogenic markers (P < 0.01 or 0.001). In contrast, Platr3 overexpression alleviated the repressive effect of P. gingivalis on cementoblast mineralization (P < 0.01 or 0.001). Furthermore, Platr3 bound to nudix hydrolase 21 (NUDT21) and regulated the nuclear factor-κB (NF-κB) signaling pathway. Knocking down NUDT21 suppressed osteogenic marker expression and activated the above signaling pathway. Collectively, the results elucidated that Platr3 mediated P. gingivalis-suppressed cementoblast mineralization through the NF-κB signaling pathway by binding to NUDT21.

M2 macrophages with inflammation tropism facilitate cementoblast mineralization.

BACKGROUND: Cementum regeneration was regarded as the critical goal for periodontal regeneration, and M2 macrophage-based therapy was expected to be a promising strategy. However, little is known about the effects of M2 macrophages on cementoblast mineralization and tropism, especially under inflammation. Here we investigated for the first time the crosstalk between M2 macrophages and Porphyromonas gingivalis (Pg)-stimulated cementoblasts. METHODS: M2 macrophages were induced with interleukin (IL)-4, and identified. CC-chemokine ligand 2 (CCL2) expression and secretion of inflammatory cementoblasts were detected by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), western blotting (WB), immunohistochemistry for apical periodontitis (AP) mice, and by enzyme-linked immunosorbent assay. Crystal violet staining was used to observe macrophage migration. Conditional medium (CM) and transwell coculture methods were applied to evaluate the effects of M2 macrophages on cementum mineralization with or without Pg, and to explore the mechanism. Mineralization-related markers and pathway-related proteins were measured by RT-qPCR and WB. RESULTS: M2 macrophages were identified successfully. We found an increase of CCL2 in cementoblasts and their supernatant. Also, higher CCL2 in cementoblasts was observed in the AP model. Superior recruitment of M2 macrophages to supernatant from Pg-stimulated cementoblasts or CCL2-containing medium was verified. Moreover, CM2 and Trans-M2 showed better mineralization-accelerating and rescuing effects when compared to their controls, and application of p38 inhibitor partially blocked the promotion. CONCLUSIONS: Our study demonstrated the inflammation-targeting and mineralization-promoting effects of M2 macrophages on cementoblasts, which may provide evidence for M2 macrophage-based cementum regeneration.

CKIP-1 mediates P. gingivalis-suppressed osteogenic/cementogenic differentiation of periodontal ligament cells partially via p38 signaling pathway.

Objectives: Casein kinase 2 interacting protein-1 (CKIP-1) is a versatile player involved in various biological processes. However, whether CKIP-1 mediates the osteogenic/cementogenic differentiation of periodontal ligament cells (PDLCs) under Porphyromonas gingivalis (Pg) stimulation remains unknown. Material and Methods: The effect of Pg on PDLC differentiation was first verified. CKIP-1 expression in Pg-infected PDLCs or in PDL of apical periodontitis (AP) mice was detected. The changes of CKIP-1 during PDLC differentiation was also determined. PDLC differentiation capacity in CKIP-1 knockout (KO) mice and CKIP-1-silenced PDLCs with or without Pg stimulation were further studied. Inhibitor was finally applied to verify the involvement of p38 signaling pathway in PDLC differentiation. Results: The suppression effect of Pg on PDLC differentiation was demonstrated. CKIP-1 increased in the PDL of AP mice and Pg-induced PDLCs, and decreased gradually during PDLC differentiation. Increased OSX and RUNX2 expression in PDL were observed in CKIP-1 KO mice. Also, CKIP-1 silencing facilitated and rescued Pg-inhibited PDLC differentiation. Inhibitor for p38 signaling pathway blocked CKIP-1 silencing-facilitated PDLC differentiation. Conclusions: CKIP-1 mediated the osteogenic/cementogenic differentiation of PDLCs partially through p38 signaling pathway, which may provide evidence for the regeneration of periodontal hard tissues damaged by Pg.

CKIP-1 Promotes P. gingivalis-Induced Inflammation of Periodontal Soft Tissues by Inhibiting Autophagy.

As a chronic inflammatory disease, periodontitis involves many biological processes including autophagy. At the same time, casein kinase 2 interacting protein-1 (CKIP-1) was reported to play a role in regulation of inflammation. But whether CKIP-1 and autophagy interact in periodontitis remains unclear. In this paper, our research team verified the levels of CKIP-1 expression and autophagy increase in the periodontal tissues of a ligature-induced periodontitis mouse model. And this result was also confirmed in Porphyromonas gingivalis (Pg)-induced human gingival fibroblasts (HGF) and human periodontal ligament cells (PDLC). We also showed the autophagy level in periodontal tissues is higher in Ckip-1 knockout (KO) mice than wild type (WT). At the same time, CKIP-1 knockdown lentivirus was used in PDLC and HGF, and it was found that silencing CKIP-1 significantly activated autophagy. Unfortunately, the regulatory role of autophagy in periodontitis is still unclear. Then, the autophagy agonist Rapamycin and inhibitor 3-MA were used in a periodontitis mouse model to investigate periodontal tissue destruction. We found the inflammation in periodontal tissue was reduced when autophagy activated. All these conclusions have been verified both in vivo and in vitro experiments. Finally, our research proved that silencing CKIP-1 reduces the expression of inflammatory cytokines in Pg-induced PDLC and HGF by regulating autophagy. Overall, a new role for CKIP-1 in regulating periodontal tissue inflammation was demonstrated in our study, and it is possible to treat periodontitis by targeting the CKIP-1 gene.

Tet methylcytosine dioxygenase 1 modulates Porphyromonas gingivalis-triggered pyroptosis by regulating glycolysis in cementoblasts.

Porphyromonas gingivalis is involved in the pathogenesis of multiple polymicrobial biofilm-induced inflammatory diseases, including apical periodontitis, and it triggers pyroptosis accompanied by robust inflammatory responses. Tet methylcytosine dioxygenase 1 (TET1), an epigenetic modifier enzyme, has been is correlated with inflammation, though an association of TET1 and P. gingivalis-related pyroptosis in cementoblasts and the molecular mechanisms has not been shown. Our study here demonstrated that P. gingivalis downregulated Tet1 expression and elicited CASP11- and GSDMD-dependent pyroptosis. Additionally, Tet1 mRNA silencing in cementoblasts appeared to result in a more severe pyroptotic phenotype, where levels of CASP11 and GSDMD cleavage, lactate dehydrogenase release, and IL-1ß and IL-18 production were significantly increased. Moreover, Tet1 overexpression resulted in blockade of pyroptosis activation accompanied by inflammation moderation. Further analyses revealed that TET1 modulated glycolysis, confirmed by the application of the specific inhibitor 2-deoxy-d-glucose (2-DG). The pyroptosis phenotype enhanced by Tet1 silencing was moderated by 2-DG upon P. gingivalis invasion. Taken together, these data show the effects and underlying mechanisms of TET1 on pyroptosis and inflammatory phenotype induced by P. gingivalis in cementoblasts, and provides insight into the involvement of P. gingivalis in apical periodontitis and, possibly, other inflammatory diseases.