Extracellular Vesicles in Periodontal Medicine : The Candidates Linking Oral Health to General Health

The term, periodontal medicine is used to describe the multitude of systemic diseases which are regarded to link periodontal disease. The concept of periodontal medicine has been widely accepted today, however, the molecular mechanisms which periodontal diseases impact general health in whole body are not elucidated in detail. Extracellular vesicles (EVs) and outer membrane vesicles (OMVs) are the nano-sized particles released from mammalian cells and bacterial cells resectively, which influence the health and various disease by transporting biological factors to the neighbor and distant cells. In this review, we will discuss whether EVs and OMVs produced in periodontal diseases could be implicated in periodontal medicine

コツガ サイボウ ノ ブンカ ニオケル PKR ノ ヤクワリ ニツイテ

In this study, it was demonstrated that double-stranded RNA-dependent protein kinase (PKR) is required for the differentiation of osteoblasts. PKR was expressed on the surface of trabecula in rat tibiae. Dominant-negative mutant PKR cDNA, in which the amino acid lysine at 296 was replaced with arginine and which did not have catalytic activity, was transfected into mouse osteoblastic MC3T3-E1 cells; thereby establishing cells that stably expressed the PKR mutant gene (PKR-K/R). PKR-K/R mutant cells exhibited up-regulated cell growth and had low alkaline phosphatase (ALP) activity. PKR-K/R mutant cells were not able to form bone nodules in vitro. The expression of STAT1α protein increased in PKR-K/R mutant cells, in which runt-related gene 2 (Runx2)-mediated transcription decreased compared with the levels in control cells. When the expression of STAT1α protein in PKR mutant cells was suppressed in the presence of STAT1α siRNA, the activity of Runx2-mediated transcription recovered to the control level. These results indicate that PKR is a stimulator of Runx2 transcription and is a negative modulator of STAT1α expression. These findings also suggest that PKR plays important roles in the differentiation and calcification of osteoblasts by modulating STAT1α and/or Runx2 expression

Involvement of Fusobacterium Species in Oral Cancer Progression : A Literature Review Including Other Types of Cancer

Chronic inflammation caused by infections has been suggested to be one of the most important cause of cancers. It has recently been shown that there is correlation between intestinal bacteria and cancer development including metastasis. As over 700 bacterial species exist in an oral cavity, it has been concerning that bacterial infection may cause oral cancer. However, the role of bacteria regarding tumorigenesis of oral cancer remains unclear. Several papers have shown that Fusobacterium species deriving the oral cavities, especially, play a crucial role for the development of colorectal and esophageal cancer. F. nucleatum is a well-known oral bacterium involved in formation of typical dental plaque on human teeth and causing periodontal diseases. The greatest characteristic of F. nucleatum is its ability to adhere to various bacteria and host cells. Interestingly, F. nucleatum is frequently detected in oral cancer tissues. Moreover, detection of F. nucleatum is correlated with the clinical stage of oral cancer. Although the detailed mechanism is still unclear, Fusobacterium species have been suggested to be associated with cell adhesion, tumorigenesis, epithelial-to-mesenchymal transition, inflammasomes, cell cycle, etc. in oral cancer. In this review, we introduce the reports focused on the association of Fusobacterium species with cancer development and progression including oral, esophageal, and colon cancers

Conversion from epithelial to partial-EMT phenotype by Fusobacterium nucleatum infection promotes invasion of oral cancer cells

The ability of cancer cells to undergo partial-epithelial mesenchymal transition (p-EMT), rather than complete EMT, poses a higher metastatic risk. Although Fusobacterium nucleatum mainly inhabits in oral cavity, attention has been focused on the F. nucleatum involvement in colorectal cancer development. Here we examined the p-EMT regulation by F. nucleatum in oral squamous cell carcinoma (OSCC) cells. We cultured OSCC cells with epithelial, p-EMT or EMT phenotype with live or heat-inactivated F. nucleatum. Expression of the genes involved in epithelial differentiation, p-EMT and EMT were examined in OSCC cells after co-culture with F. nucleatum by qPCR. Cell growth and invasion of OSCC cells were also examined. Both live and heat-inactivated F. nucleatum upregulated the expression of p-EMT-related genes in OSCC cells with epithelial phenotype, but not with p-EMT or EMT phenotype. Moreover, F. nucleatum promoted invasion of OSCC cells with epithelial phenotype. Co-culture with other strains of bacteria other than Porphyromonas gingivalis did not alter p-EMT-related genes in OSCC cells with epithelial phenotype. F. nucleatum infection may convert epithelial to p-EMT phenotype via altering gene expression in OSCC. Oral hygiene managements against F. nucleatum infection may contribute to reduce the risk for an increase in metastatic ability of OSCC

Role of Protein Phosphatase 2A in Osteoblast Differentiation and Function

The reversible phosphorylation of proteins plays hugely important roles in a variety of cellular processes, such as differentiation, proliferation, and apoptosis. These processes are strictly controlled by protein kinases (phosphorylation) and phosphatases (de-phosphorylation). Here we provide a brief history of the study of protein phosphorylation, including a summary of different types of protein kinases and phosphatases. One of the most physiologically important serine/threonine phosphatases is PP2A. This review provides a description of the phenotypes of various PP2A transgenic mice and further focuses on the known functions of PP2A in bone formation, including its role in osteoblast differentiation and function. A reduction in PP2A promotes bone formation and osteoblast differentiation through the regulation of bone-related transcription factors such as Osterix. Interestingly, downregulation of PP2A also stimulates adipocyte differentiation from undifferentiated mesenchymal cells under the appropriate adipogenic differentiation conditions. In osteoblasts, PP2A is also involved in the ability to control osteoclastogenesis as well as in the proliferation and metastasis of osteosarcoma cells. Thus, PP2A is considered to be a comprehensive factor in controlling the differentiation and function of cells derived from mesenchymal cells such as osteoblasts and adipocytes

Cyclosporine A and FK506 as Potent Inhibitors of Streptococcus intermedius Intermedilysin-Induced NFAT-1 Activation

Cyclosporine A (CsA) and tacrolimus (FK506), a member of calcineurin inhibitors, inhibit inflammation process as part of immune response. Nuclear activated T cells subfamily NFAT1 is a trascription factor responsible for the regulation of immune response genes. Streptococcus intermedius, an oral commensal bacterium, has been shown to strongly associate with liver abscess.  The S. intermedius strains produce intermedilysin (ILY), which is responsible for the bacterial virulence. Cyclosporine A and FK506 have been widely used to control NFAT activation in most of cell types, however the ability of CsA and FK506 to inhibit ILY-induced NFAT1 activation remains to be investigated. The aim of this study was to investigate the effect of CsA and FK506 on NFAT1 activation caused by ILY. Human cholangiocellular cell line HuCCT1 was stimulated with various concentrations of ILY. The cell and nuclear morphological change was observed by microscopy analysis. The NFAT1 nuclear translocation that indicates its activation was detected by immunocytochemistry. The inhibitory effect of CsA and FK506 was tested after 30 min application before ILY treatment by using immunofluorescence microscope. The results showed cell and nuclear shrinkage in ILY-treated cells. The NFAT1 was translocated to the nuclei in HuCCT1 cells, and observed in dose dependent manner.  Cyclosporine A and FK506 inhibited ILY-induced NFAT1 nuclear translocation.  In conclusion, CsA and FK506 may act as potent inflammation control agents in S. intermedius ILY-infected cells.Keywords: Cyclosporine A, FK506, NFAT1, intermedilysi

Porphyromonas gingivalis attenuates the insulin-induced phosphorylation and translocation of forkhead box protein O1 in human hepatocytes

Objective: Porphyromonas gingivalis (P. gingivalis) is a pathogen involved in periodontal disease. Recently, periodontal disease has been demonstrated to increase the risk of developing diabetes mellitus, although the molecular mechanism is not fully understood. Forkhead box protein O1 (FoxO1) is a transcriptional factor that regulates gluconeogenesis in the liver. Gluconeogenesis is a key process in the induction of diabetes mellitus; however, little is known regarding the relationship between periodontal disease and gluconeogenesis. In this study, to investigate whether periodontal disease influences hepatic gluconeogenesis, we examined the effects of P. gingivalis on the phosphorylation and translocation of FoxO1 in insulin-induced human hepatocytes. Design: The human hepatocyte HepG2 was treated with insulin and Akt and FoxO1 phosphorylation was detected by western blot analysis. The localization of phosphorylated FoxO1 was detected by immunocytochemistry and western blot analysis. HepG2 cells were treated with SNAP26b-tagged P. gingivalis (SNAP-P. g.) before insulin stimulation, and then the changes in Akt and FoxO1 were determined by western blot analysis and immunocytochemistry. Results: Insulin (100 nM) induced FoxO1 phosphorylation 60 min after treatment in HepG2 cells. Phosphorylated FoxO1 translocated to the cytoplasm. SNAP-P.g. internalized into HepG2 cells and decreased Akt and FoxO1 phosphorylation induced by insulin. The effect of insulin on FoxO1 translocation was also attenuated by SNAP-P.g. Conclusions: Our study shows that P. gingivalis decreases the phosphorylation and translocation of FoxO induced by insulin in HepG2 cells. Our results suggest that periodontal disease may increase hepatic gluconeogenesis by reducing the effects of insulin on FoxO1

Porphyromonas gingivalis Outer Membrane Vesicles Stimulate Gingival Epithelial Cells to Induce Pro-Inflammatory Cytokines via the MAPK and STING Pathways

Porphyromonas gingivalis (Pg) is a keystone pathogen associated with chronic periodontitis and produces outer membrane vesicles (OMVs) that contain lipopolysaccharide (LPS), gingipains, and pathogen-derived DNA and RNA. Pg-OMVs are involved in the pathogenesis of periodontitis. Pg-OMV-activated pathways that induce the production of the pro-inflammatory cytokines, interleukin (IL)-6, and IL-8 in the human gingival epithelial cell line, OBA-9, were investigated. The role of mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB in levels of Pg-OMV-induced pro-inflammatory cytokines was investigated using Western blot analysis and specific pathway inhibitors. Pg-OMVs induced IL-6 and IL-8 production via the extracellular signal-regulated kinase (Erk) 1/2, c-Jun N-terminal kinase (JNK), p38 MAPK, and NF-κB signaling pathways in OBA-9 cells. In addition, the stimulator of interferon genes (STING), an essential innate immune signaling molecule, was triggered by a cytosolic pathogen DNA. Pg-OMV-induced IL-6 and IL-8 mRNA expression and production were significantly suppressed by STING-specific small interfering RNA. Taken together, these results demonstrated that Pg-OMV-activated Erk1/2, JNK, p38 MAPK, STING, and NF-κB signaling pathways resulting in increased IL-6 and IL-8 expression in human gingival epithelial cells. These results suggest that Pg-OMVs may play important roles in periodontitis exacerbation by stimulating various pathways

Outer membrane vesicles of Porphyromonas gingivalis: Novel communication tool and strategy

Extracellular vesicles (EVs) have been recognized as a universal method of cellular communications and are reportedly produced in bacteria, archaea, and eukaryotes. Bacterial EVs are often called "Outer Membrane Vesicles" (OMVs) as they were the result of a controlled blebbing of the outer membrane of gram-negative bacteria such as Porphyromonas gingivalis (P. gingivalis). Bacterial EVs are natural messengers, implicated in intra-and inter-species cell-to-cell communication among microorganism populations present in microbiota. Bacteria can incorporate their pathogens into OMVs; the content of OMVs differs, depending on the type of bacteria. The production of distinct types of OMVs can be mediated by different factors and routes. A recent study highlighted OMVs ability to carry crucial molecules implicated in immune modulation, and, nowadays, they are considered as a way to communicate and transfer messages from the bacteria to the host and vice versa. This review article focuses on the current understanding of OMVs produced from major oral bacteria, P. gingivalis: generation, characteristics, and contents as well as the involvement in signal transduction of host cells and systemic diseases. Our recent study regarding the action of P. gingivalis OMVs in the living body is also summarized