Reassessment of minimal media reveals differences in growth among Porphyromonas gingivalis standard strains

ObjectivesPorphyromonas gingivalis is one of the etiologic agents of chronic periodontitis. Our previous study showed that the use of minimal media for P. gingivalis allowed to isolate novel inhibitors of P. gingivalis growth. However, growth of P. gingivalis in minimal media was not always reproducible.MethodsTo explain this phenomenon, we analyzed the growth of seven wild-type ATCC 33277 strains and two wild-type W83 strains in 10 minimal media and three complex media.ResultsAll nine strains grew in LF (Lactalbumin-Ferric chloride), GC (bovine γ–immunoglobulin G-Calcium chloride), and newly developed mC (milk-Casein) minimal media. Therefore, LF, GC, and mC could be used as minimal media for P. gingivalis. In contrast, other six minimal media containing bovine serum albumin (BSA) supported the growth of several less strains; among these, two media also showed lack of reproducibility in growth among ATCC 33277 strains. On the other hand, four ATCC 33277 strains grew similarly in all 13 media, but two W83 and other three ATCC 33277 strains grew differently in at least one medium.ConclusionsThese results suggest that the lack of reproducibility of P. gingivalis growth on minimal media is caused by the presence of BSA, and by differences among the standard strains of P. gingivalis.     

An in vitro study of the effects of Phellodendron bark extract and berberine chloride on periodontal pathogenic bacteria in the oral microbiome

ObjectivesPeriodontal disease is triggered by oral microbiome dysbiosis. Thus, to prevent its onset, it is important to maintain relative abundance of periodontal pathogenic bacteria in the oral microbiome at a low level. While Phellodendron bark extract (PBE) and its active ingredient, berberine, exert antibacterial effects on periodontal pathogenic bacteria, such as Porphyromonas gingivalis, their effects on the oral microbiome as a whole remain unknown. Therefore, we aimed to clarify the potential of PBE and berberine chloride (BC) in regulating the relative abundance of periodontal pathogenic bacteria in the oral microbiome.MethodsSaliva was collected from 20 participants. Each participant's saliva was combined separately with P. gingivalis suspension and either PBE or BC in a modified basal medium. The samples were then incubated under anaerobic conditions for 24 h. After cultivation, we determined the total bacterial concentration using quantitative polymerase chain reaction analysis and the bacterial composition using 16 S ribosomal RNA gene sequencing.ResultsThe total bacterial concentration was reduced because of treatment with PBE and BC. Bacterial 16 S ribosomal RNA gene sequencing confirmed that treatment with PBE and BC significantly reduced the relative abundance of periodontal pathogenic bacteria, including red and orange complex bacteria.ConclusionsOur findings suggest that PBE and BC reduce the relative abundance of periodontal pathogenic bacteria in the oral microbiome. Thus, PBE and BC can aid in preventing periodontal disease, given their ability to regulate the oral microbiome composition and their anti-inflammatory effects.     

Bioactive composition and antibacterial activity of three herbal extracts (lemongrass,sage, and guava leaf) against oral bacteria: An in vitro study

ObjectiveThe objective of this study was to estimate the antibacterial activity of three different herbal extracts against oral bacteria and their bioactive composition.MethodsUsing the disk diffusion technique, the antibacterial activities of three different extracts (lemongrass, sage, and guava leaf) were evaluated against oral bacteria (Streptococcus mutans, Staphylococcus aureus, and Enterococcus faecalis). Additionally, the bioactive components of the herbal extracts were assessed by employing the gas chromatography-mass spectrometry technique.ResultsThe sage, lemongrass, and guava leaf extracts suppressed the proliferation of all three tested bacterial strains at different rates. The phytochemical analysis revealed that sage extract possessed the highest content of antioxidants, phenols, and flavonoid compounds. The gas chromatography-mass spectrometry analysis of the tested plants revealed the presence of vital bioactive compounds.ConclusionsLemongrass, sage, and guava leaf extracts have potent antibacterial activities, are rich in bioactive compounds, and could be utilized as natural remedies for the prevention of oral diseases.     

Genetic background influences the capacity for medial edge epithelium disintegration and phenotype of cleft palate in TGFβ3 knockout mice

ObjectivesCleft palate is a frequent congenital craniofacial malformation of unknown etiology. Transforming growth factor (TGF) β3 is required for palatal shelf fusion. Although TGFβ3 knockout (KO) mice are widely used mouse models for cleft palate, cleft palate phenotypes differ among these mice. This study aimed to determine the effects of genetic background on the cleft palate phenotype in mice.MethodsWe produced TGFβ3 KO congenic mouse strains with five different genetic backgrounds. The phenotypes of the congenic strains were determined by visual examination. The capacity for disintegration of the medial edge epithelium (MEE) and basement membrane (BM) of palatal shelves of all five mouse strains was analyzed by using immunofluorescence staining after single palatal shelf suspension culture. The relationship between phenotype and disappearance of the MEE and BM was analyzed.ResultsAlthough the five congenic strains carried the same defective Tgfb3 gene, the fetal palate phenotypes differed among strains. The loss of the MEE cells and BM also differed with the genetic background, and the degree of such loss correlated with the cleft palate phenotype.ConclusionsThe cleft palate phenotype in mice is influenced by the genetic background, which governs the capacity for MEE and BM disintegration.     

Pathophysiological association between periodontal disease and Alzheimer's disease: Importance of periodontal health in the elderly

BackgroundAs the global aging population is rapidly advancing, recognizing the full potential of periodontal disease (PD) in the onset or progression of Alzheimer's disease (AD) is important for reducing geriatric morbidity. This review explores the possible role of PD in the pathogenesis of AD, as the pathological mechanisms underlying AD are the most well-studied among all types of dementia. The investigation was conducted using the electronic academic databases PubMed and ScienceDirect, employing a combination of keywords “periodontal disease,” “periodontitis,” “Alzheimer's disease,” “dementia,” and “Porphyromonas gingivalis.” After applying the selection and eligibility criteria and removing overlaps, from an initial search finding of 5933 studies, 11 were finally included for qualitative analysis.HighlightThe inflammatory reaction induced by oral pathogenic bacteria related to PD, through complex pathways, may exacerbate inflammation in the central nervous system, thereby contributing to the progression of AD.ConclusionMaintenance of adequate oral hygiene in patients diagnosed with AD is significant because they suffer from a gradual loss of manual dexterity as the disease advances. Additionally, the evidence presents the potential of systemic inflammation from PD-induced pathogenic bacteria, illustrating the grave cyclical progression of AD.     

Adipose-derived mesenchymal stem cells promote salivary duct regeneration via a paracrine effect

ObjectivesThe self-regeneration of exocrine tissues, including salivary glands, is limited and their regeneration mechanism has not yet been fully elucidated. Here we identify the role of adipose-derived mesenchymal stem cells (AMSCs) in salivary gland regeneration.MethodsAMSCs expressing mesenchymal stem cell markers were applied to a submandibular gland injury model and the mechanism of salivary gland repair and regeneration was analyzed.ResultsTransplanted green fluorescent protein (GFP)-labeled AMSCs grew tightly together and promoted ductal regeneration in the regenerative nodule, with slight infiltration of nonspecific immune cells. A comprehensive gene analysis through RNA-sequencing revealed increased expression of bone morphogenetic protein (BMP), transforming growth factor (TGF), and Wnt in AMSC-transplanted regenerative nodules. The factors released from AMSCs scavenge hydrogen peroxidase-induced reactive oxygen species (ROS) through Wnt promoter activity in vitro. Furthermore, AMSC-conditioned medium recovered the growth of the hydrogen peroxidase-damaged primordium of the submandibular gland culture ex vivo.ConclusionsThese results suggest that AMSC-released factors scavenge ROS and maintain salivary gland repair and regeneration via paracrine effects. Thus, AMSCs could be a practical and applicable tool for use in salivary gland regeneration.     

Hemagglutinating properties of a Streptococcus gordonii strain expressing sialic acid-binding adhesin homolog with low binding site similarity to that of strain DL1

ObjectivesThe Hsa adhesin of Streptococcus gordonii strain DL1 was previously identified as a hemagglutinin that binds specifically to sialoglycoconjugates. We recently found that among oral streptococcal species, S. gordonii strains most frequently express Hsa homologs on the bacterial cell surface. However, the effect of amino acid sequence diversity of nonrepetitive region 2 (NR2), a putative binding site of Hsa, on antigenicity and hemagglutinating (HA) properties is unclear due to difficulties in DNA sequencing the NR2 coding region. The aim of this study was to elucidate the similarity of the low NR2 antigenicity Hsa homolog of strain NDU1118 to that of strain DL1 and the association of the homolog with HA properties of the strain.MethodsThe hsa homolog of NDU1118 was sequenced using a long-read next-generation sequencer, and the Hsa homolog was assessed by alignment analysis of the deduced amino acid sequences. The hsa mutant of NDU1118 was generated by insertion of the erythromycin resistance gene. The HA properties of the wild type and the hsa mutant were assessed with human erythrocytes.ResultsThe NR2 amino acid sequence of the NDU1118 Hsa homolog was almost identical to that of the S. gordonii M99 Hsa homolog, also known as GspB, and less similar to that of DL1 Hsa. The hsa mutation of NDU1118 induced reduction of HA activity in untreated erythrocytes, but surprisingly increased lactose-inhibitable HA activity in neuraminidase-treated erythrocytes.ConclusionsThe results suggest the existence of an adhesin other than the Hsa homolog on the cell surface of NDU1118.     

Role of Streptococcus intermedius phosphoglucosamine mutase in bacterial growth,cell morphology,and resistance to polymorphonuclear leukocyte killing

ObjectivesStreptococcus intermedius is a member of the anginosus group of streptococci, an oral commensal bacterium found in infected root canals, and the causative agent of deep-seated abscesses. This organism has slow clearance when phagocytosed within neutrophils. Here, we investigated the role of its phosphoglucosamine mutase (GlmM), an enzyme associated with peptidoglycan synthesis, in bacterial growth, cell morphology, and resistance to polymorphonuclear leukocyte killing.MethodsThe glmM-deletion (ΔglmM) mutant and the plasmid-borne complementation (ΔglmM/glmM) strain of S. intermedius were generated. The wild type, the ΔglmM mutant, and the ΔglmM/glmM strain were phagocytosed with human polymorphonuclear leukocytes (PMNs), and bacterial viability in PMNs was determined by LIVE/DEAD staining. Additionally, bacterial growth and cell morphology were also compared.ResultsThe survival rate of the ΔglmM mutant was significant lower than that of the wild type. Although the difference in the survival rate of the ΔglmM/glmM strain compared to that of the wild type or the ΔglmM mutant was not significant, the rate appeared to be restored to the middle level. Compared to the wild type and the ΔglmM/glmM strain, the ΔglmM mutant showed reduced growth potential, a significant increase in the number of bacterial chains, and heterogeneous bacteria.ConclusionsGlmM is one of the factors responsible for the stable resistance of S. intermedius to clearance by PMNs.     

Effect of vitamin E on periodontitis: Evidence and proposed mechanisms of action

BackgroundPeriodontitis is a noncommunicable inflammatory disease of the soft tissue and bone surrounding the teeth in the jaw, which affects susceptible individuals with poor oral hygiene. A growing interest has been seen in the use of dietary supplements and natural products for the treatment and prevention of periodontitis. Vitamin E consists of two major groups, namely tocopherols and tocotrienols, which are botanical lipophilic compounds with excellent anti-inflammatory and antioxidant properties.HighlightThis review aimed to summarize the preclinical and clinical findings on the effects of vitamin E on periodontitis. The current literature suggests that vitamin E could improve the periodontal status by correcting redox status imbalance, reducing inflammatory responses, and promoting wound healing, thus highlighting the potential of vitamin E in the management of periodontitis.ConclusionDirect evidence for the use of vitamin E supplementation or treatment of periodontitis in humans is still limited. More well-designed and controlled studies are required to ascertain its effectiveness.     

Characterization and evaluation of ascorbic acid-induced cell sheet formation in human periodontal ligament stem cells: An in vitro study

ObjectivesPeriodontal ligament-derived stem cells (PDLSCs) are regarded as a viable option for periodontal regeneration using cell sheet technology. The objective of the present in vitro study was to characterize human PDLSCs based on their phenotypic and biological properties and to evaluate the ascorbic acid (AA or vitamin C)-induced cell sheet by analyzing the molecular markers.MethodsPDLSCs were established from premolars, and their morphology, viability, proliferation, phenotypic marker expression, and ability to differentiate into osteocytes and adipocytes were analyzed. PDLSCs were then induced to form cell sheets using 100 μM AA, and gene expression was examined by real-time polymerase chain reaction.ResultsPDLSCs showed fibroblastic morphology with >95% viability. The cells were highly proliferative and positive for surface antigens CD29, CD73, and CD90 but negative for CD34 and CD45. They were capable of differentiating into osteocytes and adipocytes. Induction with 100 μM AA transformed PDLSCs into two-to three-layered cell sheets. There was no significant upregulation in ALP and RUNX2 expression in the AA-induced cell sheet. However, the expression levels of late osteoblast differentiation marker (bone gamma-carboxy glutamate protein); cementogenic markers (cementum attachment protein and CP23), and genes encoding extracellular matrix (ECM) proteins [collagen type 1 alpha 1 and integrin beta 1) were higher in AA-induced cell sheets by PDLSCs.ConclusionsThe stimulating effect of AA on cell sheet formation by PDLSCs was confirmed by the expression of typical markers involved in osteogenesis/cementogenesis and ECM secretion, which makes this procedure a prospective option for periodontal tissue regeneration applications.     

A Ser252Trp substitution in mouse FGFR2 results in hyperplasia of embryonic salivary gland parenchyma

ObjectivesMutations in the fibroblast growth factor receptor 2 (FGFR2) gene are responsible for several severe forms of craniosynostotic disorders, such as Apert and Crouzon syndromes. Patients with craniosynostotic disorders caused by a mutation in Fgfr2 present with several clinical symptoms, including hypersalivation. Here we used a transgenic mouse model of Apert syndrome (Fgfr2^+/S252W mice) to evaluate the morphology of the submandibular glands at embryonic day 15.5 (E15.5), the time point reported to mark the start of lumen formation.MethodsFgfr2^+/S252W mice were generated by crossing ACTB-Cre^+/+ and Fgfr2^+/Neo-S252W mice. After measuring body weight, the submandibular glands were collected at E15.5. H&E staining, immunostaining, and RT-qPCR were performed to investigate the development of the submandibular gland.ResultsThe number of ducts and acini in Fgfr2^+/S252W mice was significantly higher than in control littermates; however, lumen formation was not affected. The mRNA expression of Fgf1, Fgfr1, Mmp2, Bmp4, Bmp7, Dusp6, and Etv5 in Fgfr2^+/S252W mice was significantly higher compared to control littermates. Immunoreactivity for FGF3, FGF1, BMP4, and F4/80 was detected in the parenchyma of Fgfr2^+/S252W mice. The area of apoptotic cells stained with TUNEL in Fgfr2^+/S252W mice was significantly larger than that of the control littermates.ConclusionsThese results suggested that increased FGFR1 signaling and apoptosis in the submandibular glands of Fgfr2^+/S252W mice occurred at E15.5, leading to parenchymal hyperplasia. This study demonstrated that a Ser252Trp substitution in mouse FGFR2 resulted in hyperplasia of the submandibular gland parenchyma during development.     

Systematic review and meta-analysis of randomized controlled studies comparing oscillating-rotating and other powered toothbrushes

BackgroundThe aim of this study was to systematically review and analyze the difference in efficacy of oscillating-rotating toothbrushes compared with other powered toothbrushes.MethodsThe authors performed a systematic search of the literature according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The authors used the population, intervention, comparison, and outcome format to develop a search strategy to answer the study question. The authors searched PubMed-MEDLINE databases. Inclusion criteria were randomized controlled clinical trials comparing oscillating-rotating (OR) toothbrushes with other powered toothbrushes published from January 1, 2009 through March 1, 2019.ResultsThe authors’ search resulted in 454 articles; 21 articles were downloaded for review, 15 articles were included in the report, and 12 could be used for meta-analysis. All of the studies were randomized controlled clinical trials that assessed plaque removal and gingival inflammation indexes. Results of the meta-analysis of the randomized controlled clinical trials showed that OR toothbrushes had superior, statistically significant outcomes for whole-mouth plaque reduction, assessed using the Rustogi Modified Navy Plaque Index (P < .01), and gingivitis, assessed by using number of bleeding sites (P < .001), but not for the modified gingival index (P > .05) or gingival bleeding index (P > .05).Practical ImplicationsThere is some evidence to suggest that OR powered toothbrushes might remove more plaque and reduce the number of bleedings sites better than other powered toothbrushes, specifically, sonic action toothbrushes.     

The impact of COVID-19 lockdown on maintenance of children's dental health: A questionnaire-based survey

BackgroundThe COVID-19 pandemic has been associated with several changes in maintenance of children's dental health. The aim of this study was to evaluate the extent of these changes.MethodsParents were asked to respond anonymously to a questionnaire regarding alterations in their children's oral habits, such as frequency of eating and drinking, toothbrushing, signs of stress, and receiving oral health care during the lockdown period. The participants were reached either during their visit to the clinics or via the social media groups of the authors.ResultsThere were 308 parents of children aged 1 through 18 years who responded to the questionnaires. The authors found associations between increased frequency of eating and drinking, decreased frequency of toothbrushing, and postponing oral health care. Among the children, 11% experienced more frequent oral signs of stress, such as temporomandibular disorder and aphthous stomatitis, during the lockdown. Although children from all age groups ate and drank more frequently between meals, younger children received a diagnosis of carious lesions more often during the lockdown (P = .015).ConclusionsDuring the lockdown, many children changed their eating, drinking, and toothbrushing habits and, thus, increased their risk of developing caries.Practical ImplicationsDuring pandemic-associated re-care visits or recall visits, it is imperative to conduct a detailed interview regarding changes in oral health habits. In children at high risk, dentists recommended more diagnostic and preventive measures to prevent deterioration of their oral health. Moreover, dentists should put more emphasis on motivational interviewing to help children resume healthier routines after the lockdown.     

Biochemical and X-ray micro-computed tomographic analyses of critical size bone defects grafted with autogenous bone and mercerized bacterial cellulose membranes salified with alendronate

ObjectivesThis study aimed to evaluate the repair of critical-sized bone defects grafted with autogenous bone and mercerized bacterial cellulose membranes (BCm) salified with alendronate (ALN).MethodsForty-eight male Wistar rats underwent surgery to create a 5 mm-diameter bone defect in the calvarium. The removed bone was particularized, regrafted into the defect, and covered by a BCm according to the group: control group (CG), simply mercerized BCm; group 1 (G1), negatively charged BCm (BCm-CM^-) salified with ALN; and group 2 (G2), positively charged BCm (BCm-DEAE⁺) salified with ALN. Serum samples were collected preoperatively and before euthanasia to analyze osteoprotegerin (OPG), parathyroid hormone (PTH), sclerostin (SOST), and fibroblast growth factor 23 (FGF23) levels. The animals were euthanized after 15 or 60 d. Calvaria were analyzed using quantitative microtomography (μCT).ResultsThere was an increased level of PTH in the CG compared to the G2 group, at day 60 (p = 0.019). When analyzing the same group over time, G1 presented an increased FGF23 level on days 15 and 60 (p < 0.05). CG presented an increase in PTH (p = 0.037) at day 60. The μCT analysis detected increased trabecular separation on day 15 in G2 compared to G1 (p = 0.040).ConclusionsSalification of ionized BCm with ALN had no direct effect on bone repair; however, BCm-CM^- increased the levels of FGF23 over time. BCm-DEAE⁺ decreased PTH levels compared to mercerized BCm. BCm-CM-salified with ALN-induced superior bone quality, with respect to trabecular separation, compared to BCm-DEAE⁺.