Interspecies metabolite transfer fuels the methionine metabolism of Fusobacterium nucleatum to stimulate volatile methyl mercaptan production

Hara Takeshi, Sakanaka Akito, Lamont Richard J., et al. Interspecies metabolite transfer fuels the methionine metabolism of Fusobacterium nucleatum to stimulate volatile methyl mercaptan production. mSystems 2, 118 (2024); https://doi.org/10.1128/msystems.00764-23.The major oral odor compound methyl mercaptan (CH3SH) is strongly associated with halitosis and periodontitis. CH3SH production stems from the metabolism of polymicrobial communities in periodontal pockets and on the tongue dorsum. However, understanding of CH3SH-producing oral bacteria and their interactions is limited. This study aimed to investigate CH3SH production by major oral bacteria and the impact of interspecies interactions on its generation. Using a newly constructed large-volume anaerobic noncontact coculture system, Fusobacterium nucleatum was found to be a potent producer of CH3SH, with that production stimulated by metabolic interactions with Streptococcus gordonii, an early dental plaque colonizer. Furthermore, analysis of extracellular amino acids using an S. gordonii arginine-ornithine antiporter (ArcD) mutant demonstrated that ornithine excreted from S. gordonii is a key contributor to increased CH3SH production by F. nucleatum. Further study with 13C, 15N-methionine, as well as gene expression analysis, revealed that ornithine secreted by S. gordonii increased the demand for methionine through accelerated polyamine synthesis by F. nucleatum, leading to elevated methionine pathway activity and CH3SH production. Collectively, these findings suggest that interaction between S. gordonii and F. nucleatum plays a key role in CH3SH production, providing a new insight into the mechanism of CH3SH generation in oral microbial communities. A better understanding of the underlying interactions among oral bacteria involved in CH3SH generation can lead to the development of more appropriate prophylactic approaches to treat halitosis and periodontitis. An intervention approach like selectively disrupting this interspecies network could also offer a powerful therapeutic strategy

Cigarette smoke extract impairs gingival epithelial barrier function

We previously showed that junctional adhesion molecule 1 (JAM1) and coxsackievirus and adenovirus receptor (CXADR), tight junction-associated proteins, have important roles to maintain epithelial barrier function in gingival tissues. Smoking is considered to be a significant risk factor for periodontal disease. The present study was conducted to examine the effects of cigarette smoke extract (CSE) on JAM1 and CXADR in human gingival epithelial cells. CSE was found to cause translocation of JAM1 from the cellular surface to EGFR-positive endosomes, whereas CXADR did not. Using a three-dimensional multilayered gingival epithelial tissue model, CSE administration was found to increase permeability to lipopolysaccharide and peptidoglycan, whereas overexpression of JAM1 in the tissue model prevented penetration by those substrates. Furthermore, vitamin C increased JAM1 expression, and inhibited penetration of LPS and PGN induced by CSE. These findings strongly suggest that CSE disrupts gingival barrier function via dislocation of JAM1, thus allowing bacterial virulence factors to penetrate into subepithelial tissues. Furthermore, they indicate that vitamin C increases JAM1 expression and prevents disruption of gingival barrier function by CSE.Yamaga S., Tanigaki K., Nakamura E., et al. Cigarette smoke extract impairs gingival epithelial barrier function. Scientific Reports 13, 9228 (2023); https://doi.org/10.1038/s41598-023-36366-z

Saliva and Plasma Reflect Metabolism Altered by Diabetes and Periodontitis

Periodontitis is an inflammatory disorder caused by disintegration of the balance between the periodontal microbiome and host response. While growing evidence suggests links between periodontitis and various metabolic disorders including type 2 diabetes (T2D), non-alcoholic liver disease, and cardiovascular disease (CVD), which often coexist in individuals with abdominal obesity, factors linking periodontal inflammation to common metabolic alterations remain to be fully elucidated. More detailed characterization of metabolomic profiles associated with multiple oral and cardiometabolic traits may provide better understanding of the complexity of oral-systemic crosstalk and its underlying mechanism. We performed comprehensive profiling of plasma and salivary metabolomes using untargeted gas chromatography/mass spectrometry to investigate multivariate covariation with clinical markers of oral and systemic health in 31 T2D patients with metabolic comorbidities and 30 control subjects. Orthogonal partial least squares (OPLS) results enabled more accurate characterization of associations among 11 oral and 25 systemic clinical outcomes, and 143 salivary and 78 plasma metabolites. In particular, metabolites that reflect cardiometabolic changes were identified in both plasma and saliva, with plasma and salivary ratios of (mannose + allose):1,5-anhydroglucitol achieving areas under the curve of 0.99 and 0.92, respectively, for T2D diagnosis. Additionally, OPLS analysis of periodontal inflamed surface area (PISA) as the numerical response variable revealed shared and unique responses of metabolomic and clinical markers to PISA between healthy and T2D groups. When combined with linear regression models, we found a significant correlation between PISA and multiple metabolites in both groups, including threonate, cadaverine and hydrocinnamate in saliva, as well as lactate and pentadecanoic acid in plasma, of which plasma lactate showed a predominant trend in the healthy group. Unique metabolites associated with PISA in the T2D group included plasma phosphate and salivary malate, while those in the healthy group included plasma gluconate and salivary adenosine. Remarkably, higher PISA was correlated with altered hepatic lipid metabolism in both groups, including higher levels of triglycerides, aspartate aminotransferase and alanine aminotransferase, leading to increased risk of cardiometabolic disease based on a score summarizing levels of CVD-related biomarkers. These findings revealed the potential utility of saliva for evaluating the risk of metabolic disorders without need for a blood test, and provide evidence that disrupted liver lipid metabolism may underlie the link between periodontitis and cardiometabolic disease.Sakanaka A., Kuboniwa M., Katakami N., et al. Saliva and Plasma Reflect Metabolism Altered by Diabetes and Periodontitis. Frontiers in Molecular Biosciences, 8, , 742002. https://doi.org/https://doi.org/10.3389/fmolb.2021.742002

Salivary metabolic signatures of carotid atherosclerosis in patients with type 2 diabetes hospitalized for treatment

Atherosclerosis is a life-threatening disease associated with morbidity and mortality in patients with type 2 diabetes (T2D). This study aimed to characterize a salivary signature of atherosclerosis based on evaluation of carotid intima-media thickness (IMT) to develop a non-invasive predictive tool for diagnosis and disease follow-up. Metabolites in saliva and plasma samples collected at admission and after treatment from 25 T2D patients hospitalized for 2 weeks to undergo medical treatment for diabetes were comprehensively profiled using metabolomic profiling with gas chromatography-mass spectrometry. Orthogonal partial least squares analysis, used to explore the relationships of IMT with clinical markers and plasma and salivary metabolites, showed that the top predictors for IMT included salivary allantoin and 1,5-anhydroglucitol (1,5-AG) at both the baseline examination at admission and after treatment. Furthermore, though treatment induced alterations in salivary levels of allantoin and 1,5-AG, it did not modify the association between IMT and these metabolites (pinteraction > 0.05), and models with these metabolites combined yielded satisfactory diagnostic accuracy for the high IMT group even after treatment (area under curve = 0.819). Collectively, this salivary metabolite combination may be useful for non-invasive identification of T2D patients with a higher atherosclerotic burden in clinical settings.Sakanaka A, Katakami N, Furuno M, Nishizawa H, Omori K, Taya N, Ishikawa A, Mayumi S, Inoue M, Tanaka Isomura E, Amano A, Shimomura I, Fukusaki E and Kuboniwa M (2022) Salivary metabolic signatures of carotid atherosclerosis in patients with type 2 diabetes hospitalized for treatment. Front. Mol. Biosci. 9:1074285. doi: 10.3389/fmolb.2022.107428

Periodontal tissue susceptibility to glycaemic control in type 2 diabetes

Inoue M., Sakanaka A., Katakami N., et al. Periodontal tissue susceptibility to glycaemic control in type 2 diabetes. Diabetes, Obesity and Metabolism 26, 4684 (2024); https://doi.org/10.1111/dom.15835.Aim: To assess the direct effect of intensive glycaemic control on periodontal tissues in patients with diabetes mellitus. Materials and Methods: Twenty-nine patients with type 2 diabetes were enrolled and hospitalized to receive a 2-week intensive glycaemic control regimen. We observed and analysed the systemic and oral disease indicators before and after treatment and clarified the indicators related to periodontal inflammation. Results: A significant reduction in glycaemic and periodontal parameters, including glycated albumin levels and periodontal inflamed surface area (PISA), was observed after treatment. The changes in PISA per tooth, indicative of periodontal healing, exhibited a bimodal distribution; the patients were divided into two groups on this basis. Correlations were observed between the changes in PISA per tooth and fasting plasma glucose, acetoacetic acid, and beta-hydroxybutyrate levels in the PISA-improved group. Significantly lower levels of C-peptide, coefficient of variation of R-R interval, and ankle-brachial pressure index were observed before treatment in the PISA non-improved group. Conclusions: Glycaemic control treatment can effectively improve periodontitis in patients with type 2 diabetes, even in the absence of specific periodontal treatments. However, the periodontal responsiveness to glycaemic control treatment depends on the systemic condition of the patient

Porphyromonas gingivalis induces penetration of lipopolysaccharide and peptidoglycan through the gingival epithelium via degradation of junctional adhesion molecule 1.

Porphyromonas gingivalis is a major pathogen in severe and chronic manifestations of periodontal disease, which is one of the most common infections of humans. A central feature of P. gingivalis pathogenicity is dysregulation of innate immunity at the gingival epithelial interface; however, the molecular basis underlying P. gingivalis-dependent abrogation of epithelial barrier function remains unknown. Gingival epithelial cells express junctional adhesion molecule (JAM1), a tight junction-associated protein, and JAM1 homodimers regulate epithelial barrier function. Here we show that Arg-specific or Lys-specific cysteine proteases (gingipains) secreted by P. gingivalis can specifically degrade JAM1 at K134 and R234 in gingival epithelial cells, resulting in permeability of the gingival epithelium to 40 kDa dextran, lipopolysaccharide (LPS), and proteoglycan (PGN). A P. gingivalis strain lacking gingipains was impaired in degradation of JAM1. Knockdown of JAM1 in monolayer cells and a three-dimensional multilayered tissue model also increased permeability to LPS, PGN, and gingipains. Inversely, overexpression of JAM1 in epithelial cells prevented penetration by these agents following P. gingivalis infection. Our findings strongly suggest that P. gingivalis gingipains disrupt barrier function of stratified squamous epithelium via degradation of JAM1, allowing bacterial virulence factors to penetrate into subepithelial tissues

Characterization of Binding of Streptococcus oralis Glyceraldehyde-3-Phosphate Dehydrogenase to Porphyromonas gingivalis Major Fimbriae

Binding of Streptococcus oralis glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to Porphyromonas gingivalis fimbriae was characterized via a biomolecular interaction analysis system. The interaction was specific, and the association constant value was 4.34 × 10(7) M(−1), suggesting that S. oralis GAPDH functions as a dominant receptor for P. gingivalis and contributes to P. gingivalis colonization

Porphyromonas gingivalis Induces Receptor Activator of NF-κB Ligand Expression in Osteoblasts through the Activator Protein 1 Pathway

Porphyromonas gingivalis, an important periodontal pathogen, is closely associated with inflammatory alveolar bone resorption, and several components of the organism such as lipopolysaccharides have been reported to stimulate production of cytokines that promote inflammatory bone destruction. We investigated the effect of infection with viable P. gingivalis on cytokine production by osteoblasts. Reverse transcription-PCR and real-time PCR analyses revealed that infection with P. gingivalis induced receptor activator of nuclear factor κB (NF-κB) ligand (RANKL) mRNA expression in mouse primary osteoblasts. Production of interleukin-6 was also stimulated; however, osteoprotegerin was not. SB20350 (an inhibitor of p38 mitogen-activated protein kinase), PD98059 (an inhibitor of classic mitogen-activated protein kinase kinase, MEK1/2), wortmannin (an inhibitor of phosphatidylinositol 3 kinase), and carbobenzoxyl-leucinyl-leucinyl-leucinal (an inhibitor of NF-κB) did not prevent the RANKL expression induced by P. gingivalis. Degradation of inhibitor of NF-κB-alpha was not detectable; however, curcumin, an inhibitor of activator protein 1 (AP-1), prevented the RANKL production induced by P. gingivalis infection. Western blot analysis revealed that phosphorylation of c-Jun, a component of AP-1, occurred in the infected cells, and an analysis of c-Fos binding to an oligonucleotide containing an AP-1 consensus site also demonstrated AP-1 activation in infected osteoblasts. Infection with P. gingivalis KDP136, an isogenic deficient mutant of arginine- and lysine-specific cysteine proteinases, did not stimulate RANKL production. These results suggest that P. gingivalis infection induces RANKL expression in osteoblasts through AP-1 signaling pathways and cysteine proteases of the organism are involved in RANKL production