Lactobacillus rhamnosus inhibits osteoclast differentiation by suppressing the TLR2/NF-κB pathway.

OBJECTIVE: This study aimed to investigate the role of ultrasonicated Lactobacillus rhamnosus extract in osteoclast differentiation and its underlying mechanism, providing new strategies for the treatment of periodontitis. MATERIALS AND METHODS: Osteoclasts were induced using macrophage colony-stimulating factor and receptor activator for nuclear factor-κB ligand. Lactobacillus rhamnosus extracts were obtained via ultrasonic crushing and ultracentrifugation. The effects of the LGG extract on osteoclast differentiation were evaluated, and the related signaling pathways were examined using western blotting. A mouse periodontitis model was established, and Lactobacillus rhamnosus extract was injected into the gingival sulcus to evaluate the inhibitory effect of Lactobacillus rhamnosus extract on alveolar bone resorption. RESULTS: At 50 µg/mL, Lactobacillus rhamnosus extract inhibited osteoclast differentiation with no effect on apoptosis and proliferation. This phenomenon was achieved by deactivating the NF-κB/c-Fos/NFATc1 signaling pathway through toll-like receptor 2. The in vivo results showed that the local injection of Lactobacillus rhamnosus extract suppressed osteoclast differentiation and alveolar bone resorption. CONCLUSION: The ultrasonicated extract of Lactobacillus rhamnosus inhibited osteoclast differentiation by suppressing the activation of the NF-κB/c-Fos/NFATc1 pathway. Furthermore, it inhibited the destruction of the alveolar bone, providing a new strategy for the use of probiotics in the treatment of periodontitis.

Clinical and biochemical effect of laser as an adjunct to non-surgical treatment of chronic periodontitis.

OBJECTIVES: To evaluate the clinical and biochemical efficacy of laser therapy as an adjunct to non-surgical treatment in chronic periodontitis. METHODS: A systematic search was performed through the PubMed, EMBASE, and Cochrane Library for eligible articles published as of May 2, 2020, supplemented by information search in the System for Information on Programme Literature in Europe and a manual literature search. Only randomized controlled trials (RCTs) used to compare the adjunctive use of laser and non-surgical treatment alone with an observation period of at least 6 months were included. RESULTS: Sixteen RCTs with a total of 525 subjects were included. Meta-analysis suggested that the additional use of laser to scaling and root planing (SRP) showed significant superiority over SRP alone among most of clinical parameters involved. Regarding the GCF, although volume in the laser group was lower at week 4 and 12, no significant difference was found regarding the cytokines level. Subgroup analysis revealed that the combined therapy produced no significant difference in PD, CAL and PI at most time points for studies in respect to smokers. No treatment-related adverse events had been reported in the included studies. CONCLUSIONS: Pooled analysis suggested that laser-assisted non-surgical treatment improved clinical outcome to SRP alone in the management of non-smoking chronic periodontitis patients.

DNA methylation alterations and their potential influence on macrophage in periodontitis.

OBJECTIVES: To explore how various methylation mechanisms function and affect macrophages in periodontitis, with an aim of getting a comprehensive understanding of pathogenesis of the disease. SUBJECT: Alterations in DNA methylation are associated with different periodontitis susceptible factors and disrupt immunity homeostasis. The host's immune response to stimulus plays a vital role in the progression of periodontitis. Macrophages are key immune cells of immune system. They act as critical regulators in maintaining issue homeostasis with their nature of high plasticity. The altered methylation status of genes may cause abnormal expression of proteins in the progress of periodontitis, thus, exert potential influence on macrophages. RESULTS: Certain genes are selectively activated or silenced due to the changes in the methylation status, which causes the alteration of the expression level of cytokines/chemokines, signal molecules, extracellular matrix molecules, leads to the change in local microenvironment, affects activation states of immune cells including macrophages, thus influences the host immune response during periodontitis.. This results in differential susceptibility and therapeutic outcome. CONCLUSION: DNA methylation alteration may cause aberrant expression level of genes associated with periodontal diseases, thus results in deregulation of macrophages, which supports the prospect of using DNA methylation-related parameter as a new biomarker for the diagnosis and treatment of periodontitis.

Porphyromonas gingivalis aggravates colitis via a gut microbiota-linoleic acid metabolism-Th17/Treg cell balance axis.

Periodontitis is closely related to inflammatory bowel disease (IBD). An excessive and non-self-limiting immune response to the dysbiotic microbiome characterizes the two. However, the underlying mechanisms that overlap still need to be clarified. We demonstrate that the critical periodontal pathogen Porphyromonas gingivalis (Pg) aggravates intestinal inflammation and Th17/Treg cell imbalance in a gut microbiota-dependent manner. Specifically, metagenomic and metabolomic analyses shows that oral administration of Pg increases levels of the Bacteroides phylum but decreases levels of the Firmicutes, Verrucomicrobia, and Actinobacteria phyla. Nevertheless, it suppresses the linoleic acid (LA) pathway in the gut microbiota, which was the target metabolite that determines the degree of inflammation and functions as an aryl hydrocarbon receptor (AHR) ligand to suppress Th17 differentiation while promoting Treg cell differentiation via the phosphorylation of Stat1 at Ser727. Therapeutically restoring LA levels in colitis mice challenged with Pg exerts anti-colitis effects by decreasing the Th17/Treg cell ratio in an AHR-dependent manner. Our study suggests that Pg aggravates colitis via a gut microbiota-LA metabolism-Th17/Treg cell balance axis, providing a potential therapeutically modifiable target for IBD patients with periodontitis.

Mesenchymal stem cell-derived apoptotic bodies alleviate alveolar bone destruction by regulating osteoclast differentiation and function.

Periodontitis is caused by overactive osteoclast activity that results in the loss of periodontal supporting tissue and mesenchymal stem cells (MSCs) are essential for periodontal regeneration. However, the hypoxic periodontal microenvironment during periodontitis induces the apoptosis of MSCs. Apoptotic bodies (ABs) are the major product of apoptotic cells and have been attracting increased attention as potential mediators for periodontitis treatment, thus we investigated the effects of ABs derived from MSCs on periodontitis. MSCs were derived from bone marrows of mice and were cultured under hypoxic conditions for 72 h, after which ABs were isolated from the culture supernatant using a multi-filtration system. The results demonstrate that ABs derived from MSCs inhibited osteoclast differentiation and alveolar bone resorption. miRNA array analysis showed that miR-223-3p is highly enriched in those ABs and is critical for their therapeutic effects. Targetscan and luciferase activity results confirmed that Itgb1 is targeted by miR-223-3p, which interferes with the function of osteoclasts. Additionally, DC-STAMP is a key regulator that mediates membrane infusion. ABs and pre-osteoclasts expressed high levels of DC-STAMP on their membranes, which mediates the engulfment of ABs by pre-osteoclasts. ABs with knock-down of DC-STAMP failed to be engulfed by pre-osteoclasts. Collectively, MSC-derived ABs are targeted to be engulfed by pre-osteoclasts via DC-STAMP, which rescued alveolar bone loss by transferring miR-223-3p to osteoclasts, which in turn led to the attenuation of their differentiation and bone resorption. These results suggest that MSC-derived ABs are promising therapeutic agents for the treatment of periodontitis.

Force-Induced Nitric Oxide Promotes Osteogenic Activity during Orthodontic Tooth Movement in Mice.

Objectives: The aim of this study was to investigate the effect of nitric oxide (NO) on orthodontic tooth movement and the regulatory effect on bone formation. Design: A mouse orthodontic tooth movement model was established to measure the level of releasing NO. Besides, orthodontic tooth movement distance and the bone formation in the tension side of the orthodontic tooth were also analyzed. In vitro, human periodontal ligament stem cells (hPDLSCs) were cultured under tensile force stimulation. The production of NO and the expression level of nitric oxide synthase (NOS) were detected after mechanical stimulation. Furthermore, the downstream cellular signaling pathway regulated by NO was also explored. Results: The generation of NO steadily increased throughout the orthodontic tooth movement in mice. Orthodontic tooth movement was decreased in the NOS inhibitor group while it was accelerated in the NO precursor group. Force-induced NO promoted the osteogenic differentiation of human hPDLSCs under tensile force stimulation. And force-induced NO in hPDLSCs regulated the PI3K/Akt/ß-catenin signal pathway. Conclusion: NO is involved in the regulation of orthodontic tooth movement and promotes bone formation on the tension side of the orthodontic tooth. The PI3K/Akt/ß-catenin pathway is one of the downstream cell signal transduction pathways of NO in the orthodontic process.

Porphyromonas gingivalis and Lactobacillus rhamnosus GG regulate the Th17/Treg balance in colitis via TLR4 and TLR2.

OBJECTIVES: CD4+ T cells are the key to many immune-inflammatory diseases mediated by microbial disorders, especially inflammatory bowel disease (IBD). The purpose of this study was to explore how pathogenic and probiotic bacteria directly affect the T helper (Th)17 and T regulatory (Treg) cell balance among CD4+ T cells to regulate inflammation. METHODS: Porphyromonas gingivalis (Pg; ATCC 33277) and Lactobacillus rhamnosus GG (LGG; CICC 6141) were selected as representative pathogenic and probiotic bacteria, respectively. Bacterial extracts were obtained via ultrasonication and ultracentrifugation. Flow cytometry, RT-qPCR, ELISAs, immunofluorescence and a Quantibody cytokine array were used. The dextran sodium sulphate (DSS)-induced colitis model was selected for verification. RESULTS: The Pg ultrasonicate induced the apoptosis of CD4+ T cells and upregulated the expression of the Th17-associated transcription factor RoRγt and the production of the proinflammatory cytokines IL-17 and IL-6, but downregulated the expression of the essential Treg transcription factor Foxp3 and the production of the anti-inflammatory factors TGF-ß and IL-10 via the TLR4 pathway. However, LGG extract maintained Th17/Treg homeostasis by decreasing the IL-17+ Th17 proportion and increasing the CD25+ Foxp3+ Treg proportion via the TLR2 pathway. In vivo, Pg-stimulated CD4+ T cells aggravated DSS-induced colitis by increasing the Th17/Treg ratio in the colon and lamina propria lymphocytes (LPLs), and Pg + LGG-stimulated CD4+ T cells relieved colitis by decreasing the Th17/Treg ratio via the JAK-STAT signalling pathway. CONCLUSIONS: Our findings suggest that pathogenic Pg and probiotic LGG can directly regulate the Th17/Treg balance via different TLRs.

Topical Host-Modulating Therapy for Periodontal Regeneration: A Systematic Review and Meta-Analysis.

Background: Over the past decades, locally delivered host-modulating pharmacological agents have been extensively investigated to treat periodontitis. Although a small category of agents has been tested in clinical trials, most of them were reported in the animal experiments. This systematic review evaluates the efficacy of local application of currently available host-modulating agents, with or without mechanical removal of plaque, in animal models with periodontitis or periodontal defect. Methods: PubMed and Embase were searched on February 11, 2019. The inclusion criteria were as follows: (1) experiments were performed in healthy animals (all ages, sexes, and species) with periodontitis, and (2) outcome data for the local application of host-modulating approaches were presented for bone quantity, bone loss, and attachment loss and compared with a vehicle control group. Study characteristics and outcome data were extracted and internal validity was assessed. The efficacy of host-modulating agents was analyzed in a meta-analysis. A standardized mean difference (SMD) and its 95% confidence intervals for each individual comparison were calculated to estimate the overall effect. Subgroup analyses were conducted on animal species and different types of agents. Results: Forty-eight articles were included in the review, of which 42 were included in meta-analysis on bone quantity, bone loss, and attachment loss. The results showed that host-modulating therapy significantly increased bone formation (SMD: 2.200 [1.560-2.840], n = 24), and decreased bone loss (SMD: -1.659 [-1.969 to -1.348], n = 51) and attachment loss (SMD: -1.572 [-2.211 to -0.933], n = 17). No significant subgroup effects were identified, indicating that the effects are similar across species and drug types. Conclusions: The current scientific evidence supports the use of local host-modulation therapy in the treatment of periodontitis in experimental animals. Our findings seem robust for various animal models and designs included in this review, which increases our confidence of the translational value of the results to the clinical situation. Impact Statement Over the past decades, locally delivered host-modulating pharmacological agents have been extensively investigated for periodontal regeneration. Although a small category of agents has been tested in clinical trials, most of them were reported in the animal experiments. Considering the increasing amount of preclinical evidence and the need for future clinical trials, a systematic review and meta-analysis of all preclinical data was performed to determine the translational value of host-modulating drugs for human application. The results showed that host-modulating therapy significantly increased bone formation and decreased bone and attachment loss. This study contributes to researchers working on the periodontal translational research.