SPRC Suppresses Experimental Periodontitis by Modulating Th17/Treg Imbalance

Object: The aims of the study were to explore the protective effects of S-propargyl-cysteine (SPRC) on periodontitis and to determine the underlying mechanisms.Methods: A rat periodontitis model was constructed by injecting LPS and SPRC (0, 25, and 50 mg/kg/d) was administered intraperitoneally. H2S and CSE level were detected. The alveolar bone level was evaluated by micro-CT, HE staining and methylene blue staining analysis. Inflammation-related factors, Treg and Th17 cells were detected by immunohistochemistry, RT-PCR, immunofluorescence, Western blot and flow cytometry. Phosphorylation levels of ERK1/2 and CREB were analysed.Results: The administration of SPRC significantly increased the expression of CSE in the gingival tissue and the concentration of endogenous H2S in the peripheral blood. Simultaneously, SPRC significantly inhibited the resorption of alveolar bone based on the H&E staining, micro-CT and methylene blue staining analysis. Compared with the periodontitis group, the levels of IL-17A, IL-10 were downregulated and IL-6,TGF-β1 were upregulated in the SPRC groups. In the SPRC group, the percentage of TH17 cells and the expression of ROR-γt were downregulated, while the percentage of Tregs and the expression of Foxp3 were upregulated accompanied with inhibition of phosphorylation ERK1/2 and CREB.Conclusion: SPRC can prevent the progression of periodontitis by regulating the Th17/Treg balance by inhibition of the ERK/CREB signalling pathway.

Pleckstrin Levels Are Increased in Patients with Chronic Periodontitis and Regulated via the MAP Kinase-p38α Signaling Pathway in Gingival Fibroblasts

Chronic periodontitis (CP) is a bacteria-driven inflammatory disease characterized by the breakdown of gingival tissue, the periodontal ligament, and alveolar bone, leading ultimately to tooth loss. We previously reported the pleckstrin gene (PLEK) to be highly upregulated in gingival tissue of patients with CP and the only gene concurrently upregulated in other inflammatory diseases including rheumatoid arthritis and cardiovascular diseases. Using saliva from 169 individuals diagnosed with CP and healthy controls, we investigated whether pleckstrin could serve as a novel biomarker of periodontitis. Additionally, we explored signal pathways involved in the regulation of PLEK using human gingival fibroblasts (HGFs). Pleckstrin levels were significantly higher (p < 0.001) in the saliva samples of patients with CP compared to controls and closely associated with CP severity. Immunohistochemical analysis revealed the expression of pleckstrin in inflammatory cells and gingival fibroblasts of CP patients. To explore the signal pathways involved in pleckstrin regulation, we stimulated HGFs with either interleukin-1β (IL-1β) or lipopolysaccharides (LPS) alone, or in combination with inhibitors targeting c-Jun N-terminal kinase, tyrosine kinase, protein kinase C, or p38 MAP kinase. Results showed that IL-1β and LPS significantly increased PLEK mRNA and pleckstrin protein levels. VX-745, the p38 MAP kinase inhibitor significantly decreased IL-1β- and LPS-induced pleckstrin levels at both the mRNA and the protein level. Together, these findings show that pleckstrin could serve as a salivary biomarker for the chronic inflammatory disease periodontitis and a regulator of inflammation via the p38 MAP kinase pathway.

Gene expression of inflammatory immune factors and clinical parameters in diabetes and nondiabetes patients with periodontal disease

Objective: This study evaluated clinical, glucose, and immunological parameters in patients with type 2 diabetes mellitus (DM) compared to those without systemic alterations (NDM), both with generalized chronic periodontitis. Methodology: Twenty-one patients were selected with indications of tooth loss. Surgeries were performed using the Widman flap modified to obtain a gingival collar at 1 mm from the gingival margin. Before the surgical procedure, the following clinical parameters were evaluated: pocket probing depth (PPD), gingival recession (GR), clinical attachment level (CAL), and bleeding on probing (BOP). Fasting glucose levels (FGL) and glycosylated hemoglobin Hba1c (HbA1c) were also assessed. During the surgery, gingival tissue samples were collected and frozen for later laboratorial analysis. The samples were processed to obtain mRNA, cDNA and determine the gene expression of the immune parameters IL-1-β, IL-6, TNF-α, IL-10 and NF-kB by real-time polymerase chain reaction (RT-PCR). Data were analyzed statistically considering p<0.05. Results: The clinical and glucose parameters BOP, FGL, and HbA1c were statistically higher in the DM group. RNAm levels of IL-1β, TNF-α, and NF-kB were higher in the DM group (p<0.05). Conclusion: The presence of diabetes and hyperglycemic status increase the levels of pro-inflammatory immune factors and severity of the periodontal disease.

Gingival overgrowth and associated factors in a population of Nigerian hypertensives

Background: Gingival overgrowth may be idiopathic or secondary. Drug Induced Gingival Overgrowth (DIGO) occurs within 3 months of treatment and is more prevalent in younger age group with predilection for the anterior gingival tissue and usually not associated with attachment loss or tooth mobility unless there is an existing periodontal disease. Methodology: 170 hypertensive patients were recruited for the study; 85 calcium channel blocker (CCB) and 85 non-CCB users. Interviewer-administered questionnaires was used to obtain socio-demographic information as well as medical and drug history. GO was assessed using New Clinical Index for DIGO and data was analyzed with SPSS version 21 (Armonk, NY: IBM Corp). Continuous and nominal variables were described with means, standard deviations and frequencies. Statistical significance was set at P < 0.05. Results: Amlodipine was the most commonly used CCB. The prevalence of DIGO in CCB and non-CCB was the same (49.5%). Gingival enlargement was found equally among both sexes in the CCB and non-CCB groups. A third of the participants with GO were 70 years and above while those without were majorly in the fifth and sixth decade of life. Two-third of those with DIGO had fair oral hygiene status, two-fifth had gingival bleeding and three-fifth had mild gingival inflammation. Those without DIGO in both groups had a slight female predominance and majorly good oral hygiene. Associated factors with DIGO were female sex, 60-69 age group, 10mg drug dosage, been on medication less than 10 years, mild gingival inflammation and generalized gingivitis. Conclusion: There was no difference in the prevalence of DIGO between BBC and non-BBC users. However, there was mild gingival inflammation in all participants with DIGO and amlodipine users were three times more at risk of developing DIGO than nifedipine users. Thus, it is imperative to advise the hypertensives on the importance of maintaining adequate oral hygiene measures and incorporate periodontal care in their management so as to ameliorate the side effects of their medication.

Clinical application of concentrate growth factors combined with bone substitute in Alveolar ridge preservation of anterior teeth

Objective: To investigate the clinical effect of concentrated growth factors (CGF) combined with Bio-oss bone powder on Alveolar ridge preservation (ARP) during implantology. Methods: A total of 38 patients were selected and randomly divided into 2 groups, with 19 cases in each group. The extraction sockets were filled with Bio-oss bone powder with or without CGF. VAS pain score was recorded within1 week and Landry wound healing index (LWHI) was recorded at 1, 2 and 3 weeks after operation. CBCT was taken 3 and 6 months after operation to measure and compare the changes of vertical height, width and gray value of alveolar bone at extraction site. The changes of alveolar bone contour were observed clinically and compared between the two groups. Results: The VAS score of CGF group was lower than control group on the 1st and 3rd day after operation (P < 0.05). The LWHI of CGF group was higher than control group 1 week after operation (P < 0.05). The absorption of the labial and palatal plates height and the width in the CGF group was significantly less than the control group at 3 months (P<0.05). The gray value of alveolar bone in CGF group was significantly higher than control group at 3 months (P < 0.05). There was no significant difference in new bone contour between the two groups (P > 0.05). 94.7% cases in CGF group did not undergo bone grafting, which was significantly higher than control group (78.9%). Conclusions: The use of CGF combined with Bio-oss bone powder can help to reduce postoperative pain at the early stage of healing, form sufficient keratinized gingival tissue, effectively maintain the height and width of alveolar bone in the three-dimensional direction and provide good conditions for implant repair in the future.

Application of Hyaluronic Acid as a Biopolymer Material in Reconstruction of Interdental Papilla in Rats

Applying hyaluronic acid, a biopolymer material, in the treatment of interdental papilla reconstruction has become a trend. The main objective of this research is to investigate the histologic effect of hyaluronic acid on interdental papilla over time. Deficient interdental papilla models were surgically created in sixty-two Sprague-Dawley (SD) rats and were randomly treated with the injection of hyaluronic acid (HA group) or phosphate-buffered saline (sham control group) or left untreated (control group). After 2, 4, and 8 weeks, the rats were sacrificed in batches to observe the histological changes. A fluorochrome label was used to monitor bone formation in 8 weeks. Immunohistochemical analyses were performed to evaluate the expression of potentially relevant cytokines, vascular endothelial growth factor (VEGF), bone morphogenetic protein-2 (BMP-2), and Wnt-induced secreted protein 1 (WISP1) in the gingival tissue in 8 weeks. A preliminary study of HA degradation after 24 weeks was performed in two rats. Following the HA injection, no inflammation or granulomatous foreign body reaction was observed. HA was able to promote collagen fiber and alveolar bone regular formation in the reconstruction site. HA also enhanced VEGF, BMP-2, and WISP-1 expression in gingival tissue (p＜0.05). After 24 weeks, there was no HA filler observed in the interdental papilla. In conclusion, our study suggested that HA is an effective way to reconstruct deficient interdental papilla.

Prolyl-hydroxylase inhibitor-induced regeneration of alveolar bone and soft tissue in a mouse model of ligature-induced periodontitis

Bone injuries and fractures reliably heal through a process of regeneration with restoration to original structure and function when the gap between adjacent sides of a fracture site is small. However, when there is significant volumetric loss of bone, bone regeneration usually does not occur. In the present studies, we explore a particular case of volumetric bone loss in a mouse model of human periodontal disease (PD) in which alveolar bone surrounding teeth is permanently lost and not replaced. This model employs the placement a ligature around the upper second molar which leads to inflammation and bone breakdown and faithfully replicates the bacterially-induced inflammatory etiology of human PD to induce bone degeneration. After 10 days, the ligature is removed and the mice are treated with a timed-release formulation of a small molecule inhibitor of prolylhydroxylases (PHDi; 1,4-DPCA) previously shown to induce epimorphic regeneration of soft tissue in non-regenerating mice. This PHDi induces high expression of HIF-1α and the regenerative response is completely blocked by siHIF1a. Here, we observe that timed-release 1,4-DPCA rapidly and completely restores bone and soft tissue with normal anatomic fidelity and with increased stem cell markers due to stem cell migration into the site and/or de-differentiation of local tissue, PDL cell proliferation, and increased vascularization. In-vitro studies using gingival tissue show that 1,4-DPCA indeed induces de-differentiation and the expression of stem cell markers but does not exclude the role of migrating stem cells.

Proposed protocol for treatment of severe periodontitis without platelet transfusion in patients with aplastic anemia: a case report

Background Aplastic anemia is an intractable disease characterized by pancytopenia, susceptibility to infection, and difficulty in achieving hemostasis. In patients with severe periodontal disease and aplastic anemia, spontaneous bleeding from the gingival tissue due to thrombocytopenia and during brushing is common, which may further exacerbate dental issues. Comprehensive periodontal treatment for patients with aplastic anemia is highly challenging and requires collaboration with a hematologist. Here, we discuss the case of a patient with aplastic anemia and severe periodontitis who was successfully treated in collaboration with our hematology department. Case presentation A 36-year-old Japanese woman with chief complaints of spontaneous gingival bleeding, pain, and increasing tooth mobility consulted our department. She had developed pancytopenia at age 11 years and was later diagnosed with aplastic anemia, making her susceptible to infection due to leukopenia. The results of the initial periodontal examination led to a diagnosis of severe generalized periodontitis (generalized stage IV grade C periodontitis) caused by leukopenia and poor oral hygiene. We adopted a comprehensive treatment plan, including invasive dental procedures. The patient exhibited no postoperative bleeding due to aplastic anemia-induced thrombocytopenia and experienced a good outcome. Conclusions Both physicians and dentists should be aware that immunocompromised patients with aplastic anemia are at risk of developing severe periodontitis with severe alveolar bone resorption if the condition is combined with poor oral hygiene. Even in the presence of aplastic anemia, patients with severe periodontitis can undergo comprehensive dental treatment, including dental extraction and periodontal surgery, if bleeding and susceptibility to infection are controlled. This requires the cooperation of the patient and hematologists and can ultimately contribute to improving the patient’s quality of life.

Determination of Total Protein and Calcium in Gingival Mesenchymal Stem Cell-Conditioned Medium

Gingival mesenchymal stem cell-conditioned medium (GMSC-CM) is a conditioned medium obtained from cultured gingival mesenchymal stem cells. GMSCs are easily isolated from gingival tissue, whereas gingival tissue is easily obtained by minimally invasive techniques. On the other hand, CM contains proteins, cytokines, chemokines and growth factors that play an important role in osteogenic differentiation.This study aims to determine the total protein and calcium levels in GMSC-CM.GMSCs were grown in culture media with 10% of FBS. CM of GMSC was obtained from the media collection process with a 0.22 µm filter and concentrated with a centrifugal filter to obtain a concentrated GMSC-CM. The concentration of total protein was performed by bicinchoninic acid protein assay on GMSC-CM and concentrated GMSC-CM. Calcium level was performed by atomic absorption spectroscopy method. The results arethat GMSC-CM had a total protein concentration of 2502±0.06 µg/ml, and the concentratedGMSC-CM was 1.912±0.08 μg/ml.The calcium level of GMSC-CM was 0.009% and concentratedGMSC-CM was 0.009%. It can be concluded thatGMSC-CM had a high concentration of total protein and calcium. These parameters were used to develop bioprocesses to enhance the production of GMSC-CM, which will support the implementation of cell-free therapy for tissue regeneration.

Pathogenesis of Enamel-Renal Syndrome Associated Gingival Fibromatosis: A Proteomic Approach

The enamel renal syndrome (ERS) is a rare disorder featured by amelogenesis imperfecta, gingival fibromatosis and nephrocalcinosis. ERS is caused by bi-allelic mutations in the secretory pathway pseudokinase FAM20A. How mutations in FAM20A may modify the gingival connective tissue homeostasis and cause fibromatosis is currently unknown. We here analyzed conditioned media of gingival fibroblasts (GFs) obtained from four unrelated ERS patients carrying distinct mutations and control subjects. Secretomic analysis identified 109 dysregulated proteins whose abundance had increased (69 proteins) or decreased (40 proteins) at least 1.5-fold compared to control GFs. Proteins over-represented were mainly involved in extracellular matrix organization, collagen fibril assembly, and biomineralization whereas those under-represented were extracellular matrix-associated proteins. More specifically, transforming growth factor-beta 2, a member of the TGFβ family involved in both mineralization and fibrosis was strongly increased in samples from GFs of ERS patients and so were various known targets of the TGFβ signaling pathway including Collagens, Matrix metallopeptidase 2 and Fibronectin. For the over-expressed proteins quantitative RT-PCR analysis showed increased transcript levels, suggesting increased synthesis and this was further confirmed at the tissue level. Additional immunohistochemical and western blot analyses showed activation and nuclear localization of the classical TGFβ effector phospho-Smad3 in both ERS gingival tissue and ERS GFs. Exposure of the mutant cells to TGFB1 further upregulated the expression of TGFβ targets suggesting that this pathway could be a central player in the pathogenesis of the ERS gingival fibromatosis.In conclusion our data strongly suggest that TGFβ -induced modifications of the extracellular matrix contribute to the pathogenesis of ERS. To our knowledge this is the first proteomic-based analysis of FAM20A-associated modifications.