Genetic causal association between rheumatoid arthritis and periodontitis: a bidirectional two-sample Mendelian randomization analysis.

BACKGROUND: A link between rheumatoid arthritis (RA) and periodontitis (PD) has been established. However, their causal relationship remains unclear and the effect of different serotypes of RA on the PD development has not been investigated. This study aims to elucidate the causal association between PD and different serotypes of RA using Mendelian randomization (MR). METHODS: A bidirectional two-sample MR analysis was performed using available large-scale genome-wide association studies statistics. The inverse-variance weighted (IVW) or multiplicative random-effects IVW was used to determine causality, depending on the heterogeneity of instrumental variables. Additional sensitivity analyses were also performed. RESULTS: The forward MR analysis identified that seropositive RA (odds ratio (OR), 1.26; 95% confidence interval (CI), 1.07-1.44; p = 0.0018), but not seronegative RA (OR, 1.01; 95% CI, 0.95-1.06; p = 0.9098), was associated with an increased risk of PD. The reverse MR analysis did not show any significant causal effect of PD on RA, independent of the serotypes. The sensitivity tests (p > 0.05) confirmed the robustness and accuracy of these findings. CONCLUSION: This study revealed that there was a genetic causal effect of seropositive RA on PD, suggesting that this subtype of RA patients may require specific clinical attention to prevent the development of PD.

The Oral Microbiome and Its Role in Systemic Autoimmune Diseases: A Systematic Review of Big Data Analysis.

Introduction: Despite decades of research, systemic autoimmune diseases (SADs) continue to be a major global health concern and the etiology of these diseases is still not clear. To date, with the development of high-throughput techniques, increasing evidence indicated a key role of oral microbiome in the pathogenesis of SADs, and the alterations of oral microbiome may contribute to the disease emergence or evolution. This review is to present the latest knowledge on the relationship between the oral microbiome and SADs, focusing on the multiomics data generated from a large set of samples. Methodology: By searching the PubMed and Embase databases, studies that investigated the oral microbiome of SADs, including systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and Sjögren's syndrome (SS), were systematically reviewed according to the PRISMA guidelines. Results: One thousand and thirty-eight studies were found, and 25 studies were included: three referred to SLE, 12 referred to RA, nine referred to SS, and one to both SLE and SS. The 16S rRNA sequencing was the most frequent technique used. HOMD was the most common database aligned to and QIIME was the most popular pipeline for downstream analysis. Alterations in bacterial composition and population have been found in the oral samples of patients with SAD compared with the healthy controls. Results regarding candidate pathogens were not always in accordance, but Selenomonas and Veillonella were found significantly increased in three SADs, and Streptococcus was significantly decreased in the SADs compared with controls. Conclusion: A large amount of sequencing data was collected from patients with SAD and controls in this systematic review. Oral microbial dysbiosis had been identified in these SADs, although the dysbiosis features were different among studies. There was a lack of standardized study methodology for each study from the inclusion criteria, sample type, sequencing platform, and referred database to downstream analysis pipeline and cutoff. Besides the genomics, transcriptomics, proteomics, and metabolomics technology should be used to investigate the oral microbiome of patients with SADs and also the at-risk individuals of disease development, which may provide us with a better understanding of the etiology of SADs and promote the development of the novel therapies.

Role of molybdenum in material immunomodulation and periodontal wound healing: Targeting immunometabolism and mitochondrial function for macrophage modulation.

Recently, strategies that can target the underlying mechanisms of phenotype change to modulate the macrophage immune response from the standpoint of biological science have attracted increasing attention in the field of biomaterials. In this study, we printed a molybdenum-containing bioactive glass ceramic (Mo-BGC) scaffold as an immunomodulatory material. In a clinically relevant critical-size periodontal defect model, the defect-matched scaffold featured robust immunomodulatory activity, enabling long-term stable macrophage modulation and leading to enhanced regeneration of multiple periodontal tissues in canines. Further studies demonstrated that the regeneration-enhancing function of Mo-BGC scaffold was macrophage-dependent by using canines with host macrophage depletion. To investigate the role of Mo in material immunomodulation, in vitro investigations were performed and revealed that Mo-BGC powder extract, similar to MoO42--containing medium, induced M2 polarization by enhancing the mitochondrial function of macrophages and promoted a cell metabolic shift from glycolysis toward mitochondrial oxidative phosphorylation. Our findings demonstrate for the first time an immunomodulatory role of a Mo-containing material in the dynamic cascade of wound healing. By targeting the immunometabolism and mitochondrial function of macrophages, Mo-mediated immunomodulation provides new avenues for future material design in the field of tissue engineering and regenerative medicine.

Calcitriol inhibits osteoclastogenesis in an inflammatory environment by changing the proportion and function of T helper cell subsets (Th2/Th17).

OBJECTIVES: Previously, we found that by regulating T helper (Th) cell polarization, calcitriol intervention inhibited lipopolysaccharide (LPS)-induced alveolar bone loss in an animal periodontitis model, but the underlying cellular events remain unknown. MATERIALS AND METHODS: In this study, mouse Th cells were incubated in an inflammatory environment in the presence of dendritic cells (DCs) and LPS. Then, the potential of the Th cells to undergo Th2/Th17 polarization, the RANKL expression of the polarized Th cells and the subsequent influences of the polarized Th cells on RAW264.7 cell osteoclastogenesis in response to calcitriol administration were assessed. Finally, the effects of calcitriol on antigen presentation by DCs during these cellular events were evaluated. RESULTS: In response to calcitriol administration, Th cells in an inflammatory environment exhibited an enhanced potential for Th2 polarization along with a decreased potential for Th17 polarization. In addition, RANKL expression in Th17-polarized cells was largely inhibited. Furthermore, inflammation-induced osteoclastogenesis in RAW264.7 cells was suppressed following coculture with calcitriol-treated Th cells. During these cellular events, increased expression of Th2 promoters (such as OX-40L and CCL17) and decreased expression of Th17 promoters (such as IL-23 and IL-6) were found in DCs. CONCLUSIONS: Calcitriol can inhibit osteoclastogenesis in an inflammatory environment by changing the proportion and function of Th cell subsets. Our findings suggest that calcitriol may be an effective therapeutic agent for treating periodontitis.

Impacts of non-impacted third molar removal on the periodontal condition of adjacent second molars.

OBJECTIVE: The aim of this study was to determine how the removal of non-impacted third molars (N-M3s) affects the periodontal status of neighboring second molars (M2s). SUBJECTS AND METHODS: The periodontal condition of M2s for which the neighboring N-M3s were removed (more than 6 months previously) and those with intact N-M3s was analyzed in a cross-sectional observation study. In an additional case series, periodontal changes in M2s in response to adjacent N-M3 removal were observed during a 6-month follow-up period. RESULTS: A total of 457 patients with 1,301 M2s were enrolled in this cross-sectional observational study. Compared to M2s with neighboring N-M3s, M2s without neighboring N-M3s (teeth removed more than 6 months previously) exhibited a 0.27-mm reduction in the average pocket depth (PD) (p < .001) and a 0.38-fold reduced risk of at least one probing site with PD ≥5 mm (PD5+) (p < .001). Subsequently, a 41-case follow-up study showed that 6 months after neighboring N-M3 extraction, the PD of the M2s decreased by 0.31 mm (p < .001), while the incidence of PD5+ decreased by 21.9% when compared to the parameters detected before tooth extraction (p = .004). CONCLUSIONS: Removing N-M3s was associated with an improved periodontal condition in neighboring M2s.

Surface modification via plasmid-mediated pLAMA3-CM gene transfection promotes the attachment of gingival epithelial cells to titanium sheets in vitro and improves biological sealing at the transmucosal sites of titanium implants in vivo.

Although titanium implants have been applied in dental clinics to replace lost teeth and to restore masticatory function for decades, strategies to design the surface of the transmucosal sites of implants to achieve ideal and predictable biological sealing following implantation remain to be optimized. In this study, we hypothesized that gingival epithelial cell (GEC) adhesion and new tissue attachment to titanium sheets/implants could be promoted by the release of plasmid pLAMA3-CM (encoding a motif of the C-terminal globular domain of LAMA3) from a titanium surface. To test this hypothesis, a chitosan/collagen (Chi/Col) coating was immobilized on the surfaces of titanium substrates with nanotube topography (NT-Ti) through cathodic electrophoretic deposition; it was found that pLAMA3-CM could be released from the coating in a highly sustained manner. After culturing on titanium with nanotube topography coated by Chi/Col with the plasmid pLAMA3-CM (Chi/Col/pLAMA3-CM-Ti), human GECs (hGECs) were found to effectively uptake the incorporated plasmids, which resulted in improved attachment, as evidenced by morphological and immunofluorescence analyses. In addition, Chi/Col/pLAMA3-CM-Ti induced better biological sealing at transmucosal sites following immediate implantation into Sprague-Dawley rats. Our findings indicate that the modification of titanium implants by plasmid-mediated pLAMA3-CM gene transfection points to a practical strategy for optimizing biological sealing around the transmucosal sites of implants.

Calcitriol suppresses lipopolysaccharide-induced alveolar bone damage in rats by regulating T helper cell subset polarization.

BACKGROUND: Although the immunomodulatory properties of calcitriol in bone metabolism have been documented for decades, its therapeutic role in the management of periodontitis remains largely unexplored. In this study, we hypothesized that calcitriol suppresses lipopolysaccharide (LPS)-induced alveolar bone loss by regulating T helper (Th) cell subset polarization. METHODS: To test this hypothesis, we determined the effect of calcitriol intervention on the development of LPS-induced periodontitis in rats in terms of bone loss (micro-CT analysis), local inflammatory infiltration levels, the number of osteoclasts (hematoxylin and eosin staining) and the level of osteoclastogenesis (tartrate-resistant acid phosphatase method). Furthermore, immunohistochemistry was used to assess the expression levels of the receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) as well as the cytokine levels of interferon-γ (IFN-γ), interleukin-4 (IL-4), IL-17, and IL-10 throughout the LPS-injected region. Finally, the polarization potential of Th cells in peripheral blood was analyzed using flow cytometry. RESULTS: Calcitriol intervention decreased alveolar bone loss in response to LPS injection and inflammatory cell infiltration. Analysis of osteoclast number and RANKL and OPG expression showed that bone resorption activity was largely suppressed in response to calcitriol administration, along with decreased IL-17 levels but increased IL-4 and IL-10 levels in periodontal tissues (the LPS-injected region). Similarly, the percentages of Th2 and Treg cells in peripheral blood increased, but the percentages of Th1 and Th17 cells decreased in rats receiving calcitriol. CONCLUSION: Our findings suggest that calcitriol can be used to inhibit bone loss in experimental periodontitis, likely via the regulation of local and systemic Th cell polarization.

The relationship between T-helper cell polarization and the RANKL/OPG ratio in gingival tissues from chronic periodontitis patients.

This study aimed to investigate the relationship between inflammation-related T-helper cell polarization and the receptor activator for nuclear factor-κB ligand (RANKL)/osteoprotegerin (OPG) ratio, which is associated with bone resorption or remodeling of chronic periodontitis patients. Gingival crevicular fluid (GCF) and gingival tissues were obtained from periodontally healthy individuals (PH group) and chronic periodontitis patients (CP group). The GCF levels of IFN-γ, IL-4, IL-17, and IL-10 linked to T-helper cell polarization toward the Th1, Th2, Th17, and Treg phenotypes, respectively, were determined by ELISA. The expression levels of these cytokines and the polarized T-helper cells in gingival tissues were assessed through immunohistochemical and immunofluorescence assays. In addition, the RANKL and OPG expression levels in gingival tissues were detected by immunohistochemical assays, and linear regression analysis was used to identify the potential relationship between T-helper cell polarization and the RANKL/OPG ratio. In total, 22 individuals and 35 patients were enrolled in the present study. In both GCF and gingival tissues, increased levels of IL-17 and the decreased levels of IL-4 and IL-10 were observed in the CP group. When polarized T-helper cells were identified in gingival tissues, more Th1 and Th17 cells were found in the CP group, whereas more Th2 and Treg cells were found in the PH group. Although there was no significant difference in OPG expression between the two groups, the RANKL/OPG ratio in the CP group was higher than that in the PH group. The linear regression analysis showed that the presence of more Th1 and Th17 cells correlated with a higher RANKL/OPG ratio, whereas the presence of more Th2 cells correlated with a lower RANKL/OPG ratio. Th1 and Th17 cells are positively correlated and Th2 cells are negatively correlated with the RANKL/OPG ratio. Our data suggest that T-helper cell polarization is closely linked to the RANKL/OPG ratio in gingival tissues from chronic periodontitis patients.

Macrophage polarization in human gingival tissue in response to periodontal disease.

OBJECTIVE: Although accumulating evidence indicates that macrophages are central players in the destructive and reparative phases of periodontal disease, their polarization states at different stages of periodontal inflammation remain unclear. METHODS: We collected gingival biopsies from patients with chronic periodontitis (P group), gingivitis (G group), or periodontally healthy individuals (H group). Polarized macrophages were identified through immunofluorescence. M1- and M2-related cytokines were detected by immunohistochemistry. RESULTS: Compared with the H group, the P group had more M1 cells (higher M1/M2 ratio) and significantly higher TNF-α, IFN-γ, IL-6, and IL-12 levels. Although the G group also exhibited higher TNF-α and IL-12 levels than the H group, they had similar M1/M2 ratios. The M1/M2 ratio and IFN-γ and IL-6 levels were significantly higher in the P than the G group. Among M2-related cytokines, IL-4 levels were significantly higher in the G than the H group. The M1/M2 ratio was positively correlated with clinical probing depth (PD), and both were positively correlated with IFN-γ and IL-6. PD was negatively correlated with IL-4. CONCLUSION: Macrophage polarization in gingival tissue may be responsible for the development and progression of inflammation-induced tissue destruction, and modulating macrophage function may be a potential strategy for periodontal disease management.

Human platelet lysate supports the formation of robust human periodontal ligament cell sheets.

The use of stem cell-derived sheets has become increasingly common in a wide variety of biomedical applications. Although substantial evidence has demonstrated that human platelet lysate (PL) can be used for therapeutic cell expansion, either as a substitute for or as a supplement to xenogeneic fetal bovine serum (FBS), its impact on cell sheet production remains largely unexplored. In this study, we manufactured periodontal ligament stem cell (PDLSC) sheets in vitro by incubating PDLSCs in sheet-induction media supplemented with various ratios of PL and FBS, i.e. 10% PL without FBS, 7.5% PL + 2.5% FBS, 5% PL + 5% FBS, 2.5% PL + 7.5% FBS or 10% FBS without PL. Cultures with the addition of all the designed supplements led to successful cell sheet production. In addition, all the resultant cellular materials exhibited similar expression profiles of matrix-related genes and proteins, such as collagen I, fibronectin and integrin ß1. Interestingly, the cell components within sheets generated by media containing both PL and FBS exhibited improved osteogenic potential. Following in vivo transplantation, all sheets supported significant new bone formation. Our data suggest that robust PDLSC sheets can be produced by applying PL as either an alternative or an adjuvant to FBS. Further examination of the relevant influences of human PL that benefit cell behaviour and matrix production will pave the way towards optimized and standardized conditions for cell sheet production.

Heparin improves BMSC cell therapy: Anticoagulant treatment by heparin improves the safety and therapeutic effect of bone marrow-derived mesenchymal stem cell cytotherapy.

Systemic infusion of bone marrow-derived mesenchymal stem cells (BMSCs) has become a promising strategy for disease treatment and tissue regeneration. Strategies to enhance the efficiency of BMSC cell therapy are crucial to promote its clinical application. Here, we aimed to improve BMSC cell therapy by inhibiting the BMSC-induced coagulation reaction. Intravenous injection of gradient BMSCs into mice showed that BMSCs were not fully compatible with blood. Large doses of BMSCs induced a series of symptoms of respiratory failure and heart failure. Histological and homeostasis analysis confirmed that large doses of BMSCs induced disseminated intravascular thrombosis, exhaustion of platelets and coagulation factors, and prolonged prothrombin time (PT) and activated partial thromboplastin time (APTT). Similar to mouse BMSCs, goat and human BMSCs also induced coagulation reactions in vitro and in vivo. The coagulation was induced mostly by tissue factor, the overexpression of which enhanced the procoagulant activity of BMSCs during in vitro culture. Notably, clinical doses of BMSCs in cell therapy also induced mild and reversible coagulation, which increased BMSC lung embolism and clearance. Anticoagulation treatment by heparin (400 U/kg) prevented BMSC-induced coagulation and the acute adverse effects of large-dose BMSCs infusion efficiently. Importantly, heparin treatment led to decreased BMSC lung embolism and enhanced migration and maintenance of BMSCs to target organs in cell therapy. Based on an experimental colitis model, we confirmed that heparin treatment enhanced the effect of BMSC therapy efficiently to reduce mortality, prevent weight loss, suppress inflammation reaction and alleviate tissue injury. In conclusion, BMSCs possess procoagulant activity that could induce disseminated coagulation and thrombosis in recipients. Anticoagulation treatment by heparin is a practical strategy to improve both the safety and therapeutic effect of BMSC therapy.

Effects of short-term inflammatory and/or hypoxic pretreatments on periodontal ligament stem cells: in vitro and in vivo studies.

In this study, we extensively screened the in vitro and in vivo effects of PDLSCs following short-term inflammatory and/or hypoxic pretreatments. We found that the 24-h hypoxic pretreatment of PDLSCs significantly enhanced cell migration and improved cell surface CXCR4 expression. In addition, hypoxia-pretreated PDLSCs exhibited improved cell colony formation and proliferation. Cells that were dually stimulated also formed more colonies compared to untreated cells but their proliferation did not increase. Importantly, the hypoxic pretreatment of PDLSCs enhanced cell differentiation as determined by elevated RUNX-2 and ALP protein expression. In this context, the inflammatory stimulus impaired cell OCN protein expression, while dual stimuli led to decreased RUNX-2 and OCN mRNA levels. Although preconditioning PDLSCs with inflammatory and/or hypoxic pretreatments resulted in no differences in the production of matrix proteins, hypoxic pretreatment led to the generation of thicker cell sheets; the inflammatory stimulus weakened the ability of cells to form sheets. All the resultant cell sheets exhibited clear bone regeneration following ectopic transplantation as well as in periodontal defect models; the amount of new bone formed by hypoxia-preconditioned cells was significantly greater than that formed by inflammatory stimulus- or dual-stimuli-treated cells or by nonpreconditioned cells. The regeneration of new cementum and periodontal ligaments was only identified in the hypoxia-stimulus and no-stimulus cell groups. Our findings suggest that PDLSCs that undergo short-term hypoxic pretreatment show improved cellular behavior in vitro and enhanced regenerative potential in vivo. The preconditioning of PDLSCs via combined treatments or an inflammatory stimulus requires further investigation.

Age-related decline in the matrix contents and functional properties of human periodontal ligament stem cell sheets.

In this study, periodontal ligament (PDL) stem cells (PDLSCs) derived from different-aged donors were used to evaluate the effect of aging on cell sheet formation. The activity of PDLSCs was first determined based on their colony-forming ability, surface markers, proliferative/differentiative potentials, senescence-associated ß-galactosidase (SA-ßG) staining, and expression of pluripotency-associated transcription factors. The ability of these cells to form sheets, based on their extracellular matrix (ECM) contents and their functional properties necessary for osteogenic differentiation, was evaluated to predict the age-related changes in the regenerative capacity of the cell sheets in their further application. It was found that human PDLSCs could be isolated from the PDL tissue of different-aged subjects. However, the ability of the PDLSCs to proliferate and to undergo osteogenic differentiation and their expression of pluripotency-associated transcription factors displayed age-related decreases. In addition, these cells exhibited an age-related increase in SA-ßG expression. Aged cells showed an impaired ability to form functional cell sheets, as determined by morphological observations and Ki-67 immunohistochemistry staining. Based on the production of ECM proteins, such as fibronectin, integrin ß1, and collagen type I; alkaline phosphatase (ALP) activity; and the expression of osteogenic genes, such as ALP, Runt-related transcription factor 2, and osteocalcin, cell sheets formed by PDLSCs derived from older donors demonstrated a less potent osteogenic capacity compared to those formed by PDLSCs from younger donors. Our data suggest that the age-associated decline in the matrix contents and osteogenic properties of PDLSC sheets should be taken into account in cell sheet engineering research and clinical periodontal regenerative therapy.