Treponema denticola Promotes OSCC Development via the TGF-ß Signaling Pathway.

Numerous studies have demonstrated an association between periodontitis and oral squamous cell carcinoma (OSCC), and periodontal pathogens such as Treponema denticola are implicated in the pathogenesis of OSCC. Previous studies have mainly focused on T. denticola surface proteins-for example, chymotrypsin-like proteinase, which was detected in the majority of orodigestive tumor tissues.T. denticola may influence the development of OSCC. Nevertheless, the potential direct regulatory mechanism of T. denticola in OSCC is still unclear. Therefore, this study aimed to explore the direct effect of T. denticola on OSCC cell proliferation and elucidate potential mechanisms of T. denticola in contributing to cell proliferation. A series of in vitro experiments (e.g., CCK-8, EdU, flow cytometry) were performed to explore the effect of T. denticola on cell proliferation, cell cycle, and apoptosis. Mice experiments were performed to explore the effect of T. denticola on tumor growth. Whole mRNA transcriptome sequencing and quantitative real-time polymerase chain reaction were performed to explore the intracellular signaling pathway. Our study found that T. denticola could invade Cal-27 cells and directly promote cell proliferation, regulate the cell cycle, and inhibit apoptosis. T. denticola could also promote the growth of OSCC tumors in mice, and it upregulated Ki67 expression. Regarding the mechanism, T. denticola could promote the development of OSCC by activating the TGF-ß pathway. In conclusion, T. denticola could promote OSCC cell proliferation directly, and the mechanism was associated with intracellular TGF-ß pathway activation.

Oral Treponema denticola Infection Induces Aß1-40 and Aß1-42 Accumulation in the Hippocampus of C57BL/6 Mice.

Accumulation of amyloid-ß (Aß) in the brain is a central component of pathology in Alzheimer's disease. A growing volume of evidence demonstrates close associations between periodontal pathogens including Porphyromonas gingivalis (P. gingivalis) and Treponema denticola (T. denticola) and AD. However, the effect and mechanisms of T. denticola on accumulation of Aß remain to be unclear. In this study, we demonstrated that T. denticola was able to enter the brain and act directly on nerve cells resulting in intra- and extracellular Aß1-40 and Aß1-42 accumulation in the hippocampus of C57BL/6 mice by selectively activating both ß-secretase and γ-secretase. Furthermore, both KMI1303, an inhibitor of ß-secretase, as well as DAPT, an inhibitor of γ- secretase, were found to be able to inhibit the effect of T. denticola on Aß accumulation in N2a neuronal cells. Overall, it is concluded that T. denticola increases the expression of Aß1-42 and Aß1-40 by its regulation on beta-site amyloid precursor protein cleaving enzyme-1 and presenilin 1.

Characterization of the Treponema denticola Virulence Factor Dentilisin.

Treponema denticola is a potent periodontal pathogen that forms a red complex with Porphyromonas gingivalis and Tannerella forsythia. It has many virulence factors, yet there are only a few reports detailing these factors. Among them, dentilisin is a well-documented surface protease. Dentilisin is reported to be involved in nutrient uptake, bacterial coaggregation, complement activation, evasion of the host immune system, inhibition of the hemostasis system, and cell invasion as a result of its action, in addition to its original proteolysis function. Therefore, characterization of dentilisin, and clarifying the relationship between T. denticola and the onset of periodontal disease will be important to better understanding this disease. In this chapter, we explain the methods for analysis of dentilisin activity and pathogenicity.

Oral infectious bacteria in dental plaque and saliva as risk factors in patients with esophageal cancer.

BACKGROUND: High levels of periodontopathic bacteria as well as Streptococcus anginosus were detected in cancer tissue from patients with esophageal cancer. An association between oral infectious bacteria and esophageal cancer has been reported. METHODS: Characteristics of the oral microbiota and periodontal conditions were studied as clinicopathologic factors in patients with esophageal cancer. The study included 61 patients with esophageal cancer and 62 matched individuals without any cancers. Samples of subgingival dental plaque and unstimulated saliva were collected to evaluate the prevalence and abundance of the following oral bacteria using a real-time polymerase chain reaction assay: Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, Treponema denticola, and S. anginosus. RESULTS: In the cancer group, the prevalence of all bacteria, with the exception of F. nucleatum, in dental plaque; the prevalence of A. actinomycetemcomitans in saliva; the abundance of all bacteria, with the exception of F. nucleatum and P. intermedia, in dental plaque; and the abundance of A. actinomycetemcomitans and S. anginosus in saliva were significantly higher. Furthermore, a logistic regression analysis suggested that the prevalence of T. forsythia and S. anginosus in dental plaque and of A. actinomycetemcomitans in saliva, as well as a drinking habit, were associated with a high risk of esophageal cancer, with a high odds ratio. CONCLUSIONS: The current findings have potential implications for the early diagnosis of esophageal cancer.

Oral spirochetes: Pathogenic mechanisms in periodontal disease.

Periodontitis is an infectious inflammatory disease resulting from infection of biofilm forming bacteria. Several bacterial factors regulate inflammatory response and cause to tissue damage and loss of connection between gingival and tooth. Since bacterial virulence factors and also host immune responses have role, understanding of periodontal disease is complex, in overall we can say that in this disease epithelium is deleted by bacteria. Oral spirochetes are related to periodontitis, among them, Treponema denticola, have been associated with periodontal diseases such as early-onset periodontitis, necrotizing ulcerative gingivitis, and acute pericoronitis. This review will analyse mechanisms of pathogenesis of spirochetes in periodontitis. Microorganisms cause destruction of gingival tissue by two mechanisms. In one, damage results from the direct action of bacterial enzymes and cytotoxic products of bacterial metabolism. In the other, only bacterial components have role, and tissue destruction is the inevitable side effect of a subverted and exaggerated host inflammatory response to plaque antigens.

Polymicrobial periodontal disease triggers a wide radius of effect and unique virome.

Periodontal disease is a microbially-mediated inflammatory disease of tooth-supporting tissues that leads to bone and tissue loss around teeth. Although bacterially-mediated mechanisms of alveolar bone destruction have been widely studied, the effects of a polymicrobial infection on the periodontal ligament and microbiome/virome have not been well explored. Therefore, the current investigation introduced a new mouse model of periodontal disease to examine the effects of a polymicrobial infection on periodontal ligament (PDL) properties, changes in bone loss, the host immune response, and the microbiome/virome using shotgun sequencing. Periodontal pathogens, namely Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Fusobacterium nucleatum were used as the polymicrobial oral inoculum in BALB/cByJ mice. The polymicrobial infection triggered significant alveolar bone loss, a heightened antibody response, an elevated cytokine immune response, a significant shift in viral diversity and virome composition, and a widening of the PDL space; the latter two findings have not been previously reported in periodontal disease models. Changes in the PDL space were present at sites far away from the site of insult, indicating that the polymicrobial radius of effect extends beyond the bone loss areas and site of initial infection and wider than previously appreciated. Associations were found between bone loss, specific viral and bacterial species, immune genes, and PDL space changes. These findings may have significant implications for the pathogenesis of periodontal disease and biomechanical properties of the periodontium. This new polymicrobial mouse model of periodontal disease in a common mouse strain is useful for evaluating the features of periodontal disease.

Glutathione catabolism by Treponema denticola impacts its pathogenic potential.

Treponema denticola is a spirochete that is etiologic for periodontal diseases. This bacterium is one of two periodontal pathogens that have been shown to have a complete three step enzymatic pathway (GTSP) that catabolizes glutathione to H2S. This pathway may contribute to the tissue pathology seen in periodontitis since diseased periodontal pockets have lower glutathione levels than healthy sites with a concomitant increase in H2S concentration. In order to be able to demonstrate that glutathione catabolism by the GTSP is critical for the pathogenic potential of T. denticola, allelic replacement mutagenesis was used to make a deletion mutant (Δggt) in the gene encoding the first enzyme in the GTSP. The mutant cannot produce H2S from glutathione since it lacks gamma-glutamyltransferase (GGT) activity. The hemolytic and hemoxidation activities of wild type T. denticola plus glutathione are reduced to background levels with the Δggt mutant and the mutant has lost the ability to grow aerobically when incubated with glutathione. The Δggt bacteria with glutathione cause less cell death in human gingival fibroblasts (hGFs) in vitro than do wild type T. denticola and the levels of hGF death correlate with the amounts of H2S produced. Importantly, the mutant spirochetes plus glutathione make significantly smaller lesions than wild type bacteria plus glutathione in a mouse back lesion model that assesses soft tissue destruction, a major symptom of periodontal diseases. Our results are the first to prove that T. denticola thiol-compound catabolism by its gamma-glutamyltransferase can play a significant role in the in the types of host tissue damage seen in periodontitis.

A pleiotropic role of FlaG in regulating the cell morphogenesis and flagellar homeostasis at the cell poles of Treponema denticola.

FlaG homologue has been found in several bacteria including spirochetes; however, its function is poorly characterised. In this report, we investigated the role of TDE1473, a putative FlaG, in the spirochete Treponema denticola, a keystone pathogen of periodontitis. TDE1473 resides in a large gene operon that is controlled by a σ70 -like promoter and encodes a putative FlaG protein of 123 amino acids. TDE1473 can be detected in the periplasmic flagella (PFs) of T. denticola, suggesting that it is a flagella-associated protein. Consistently, in vitro studies demonstrate that the recombinant TDE1473 interacts with the PFs in a dose-dependent manner and that such an interaction requires FlaA, a flagellar filament sheath protein. Deletion of TDE1473 leads to long and less motile mutant cells. Cryo-electron tomography analysis reveal that the wild-type cells have 2-3 PFs with nearly homogenous lengths (ranging from 3 to 6 µm), whereas the mutant cells have less intact PFs with disparate lengths (ranging from 0.1 to 9 µm). The phenotype of T. denticola TDE1473 mutant reported here is different from its counterparts in other bacteria, which provides insight into further understanding the role of FlaG in the regulation of bacterial cell morphogenesis and flagellation.

Bacterial species associated with persistent apical periodontitis exert differential effects on osteogenic differentiation.

AIM: To determine if bacteria associated with persistent apical periodontitis induce species-specific pro-inflammatory cytokine responses in macrophages, and the effects of this species-specific microenvironment on osteogenic differentiation. METHODOLOGY: Macrophages were exposed to Enterococcus faecalis, Streptococcus oralis, Streptococcus mitis, Fusobacterium nucleatum, Treponema denticola or Tannerella forsythia, and levels of TNF-α and IL-1ß elicited were determined by immunoassay. Following treatment of MG-63 pre-osteoblasts with conditioned media from bacteria-exposed macrophages, osteogenic differentiation and viability of osteoblasts were analyzed by Alizarin Red Staining and MTS assay, respectively. Statistical analysis was carried out by one-way anova with the Tukey post-hoc test. Differences were considered to be significant if P < 0.05. RESULTS: Macrophages exposed to Gram-positive bacteria did not produce significant amounts of cytokines. F. nucleatum-challenged macrophages produced up to four-fold more TNF-α and IL-1ß compared to T. denticola or T. forsythia. Only conditioned media from macrophages treated with Gram-negative bacteria decreased mineralization and viability of osteoblasts. CONCLUSIONS: Gram-positive bacteria did not impact osteogenic differentiation and appeared innocuous. Gram-negative bacteria, in particular F. nucleatum elicited an enhanced pro-inflammatory response in macrophages, inhibited osteogenic differentiation and reduced cell viability. The findings suggest that the presence of this organism could potentially increase the severity of persistent apical periodontitis.

The Role of Treponema denticola Motility in Synergistic Biofilm Formation With Porphyromonas gingivalis.

Chronic periodontitis has a polymicrobial biofilm etiology and interactions between key oral bacterial species, such as Porphyromonas gingivalis and Treponema denticola contribute to disease progression. P. gingivalis and T. denticola are co-localized in subgingival plaque and have been previously shown to exhibit strong synergy in growth, biofilm formation and virulence in an animal model of disease. The motility of T. denticola, although not considered as a classic virulence factor, may be involved in synergistic biofilm development between P. gingivalis and T. denticola. We determined the role of T. denticola motility in polymicrobial biofilm development using an optimized transformation protocol to produce two T. denticola mutants targeting the motility machinery. These deletion mutants were non-motile and lacked the gene encoding the flagellar hook protein of the periplasmic flagella (ΔflgE) or a component of the stator motor that drives the flagella (ΔmotB). The specificity of these gene deletions was determined by whole genome sequencing. Quantitative proteomic analyses of mutant strains revealed that the specific inactivation of the motility-associated gene, motB, had effects beyond motility. There were 64 and 326 proteins that changed in abundance in the ΔflgE and ΔmotB mutants, respectively. In the ΔflgE mutant, motility-associated proteins showed the most significant change in abundance confirming the phenotype change for the mutant was related to motility. However, the inactivation of motB as well as stopping motility also upregulated cellular stress responses in the mutant indicating pleiotropic effects of the mutation. T. denticola wild-type and P. gingivalis displayed synergistic biofilm development with a 2-fold higher biomass of the dual-species biofilms than the sum of the monospecies biofilms. Inactivation of T. denticola flgE and motB reduced this synergy. A 5-fold reduction in dual-species biofilm biomass was found with the motility-specific ΔflgE mutant suggesting that T. denticola periplasmic flagella are essential in synergistic biofilm formation with P. gingivalis.

Treponema denticola Induces Epithelial Barrier Dysfunction in Polarized Epithelial Cells.

Treponema denticola, an anaerobic spirochete found mainly in the oral cavity, is associated with periodontal disease and has a variety of virulence factors. Although in vitro studies have shown that T. denticola is able to penetrate epithelial cell monolayers, its effect on the epithelial barrier junction is not known. Human gingival epithelial cells are closely associated with adjacent membranes, forming barriers in the presence of tight junction proteins, including zonula occludens-1 (ZO-1), claudin-1, and occludin. Tight junction proteins are also expressed by Madin-Darby canine kidney (MDCK) cells in culture. In this study, the MDCK cell profile was investigated following infection with T. denticola (ATCC 35405) wild-type, as well as with its dentilisin-deficient mutant, K1. Basolateral exposure of MDCK cell monolayers to T. denticola at a multiplicity of infection (MOI) of 104 resulted in a decrease in transepithelial electrical resistance (TER). Transepithelial electrical resistance in MDCK cell monolayers also decreased following apical exposure to T. denticola (MOI=104), although this took longer with basolateral exposure. The effect on the TER was time-dependent and required the presence of live bacteria. Meanwhile, MDCK cell viability showed a decrease with either basolateral or apical exposure. Immunofluorescence analysis demonstrated decreases in the amounts of immunoreactive ZO-1 and claudin-1 in association with disruption of cell-cell junctions in MDCK cells exposed apically or basolaterally to T. denticola. Western blot analysis demonstrated degradation of ZO-1 and claudin-1 in culture lysates derived from T. denticola-exposed MDCK cells, suggesting a bacteria-induced protease capable of cleaving these tight junction proteins.

Litsea japonica Leaf Extract Suppresses Proinflammatory Cytokine Production in Periodontal Ligament Fibroblasts Stimulated with Oral Pathogenic Bacteria or Interleukin-1ß.

Periodontal disease, a chronic disease caused by bacterial infection, eventually progresses to severe inflammation and bone loss. Regulating excessive inflammation of inflamed periodontal tissues is critical in treating periodontal diseases. The periodontal ligament (PDL) is primarily a connective tissue attachment between the root and alveolar bone. PDL fibroblasts (PDLFs) produce pro-inflammatory cytokines in response to bacterial infection, which could further adversely affect the tissue and cause bone loss. In this study, we determined the ability of Litsea japonica leaf extract (LJLE) to inhibit pro-inflammatory cytokine production in PDLFs in response to various stimulants. First, we found that LJLE treatment reduced lipopolysaccharide (LPS)-induced pro-inflammatory cytokine (interleukin-6 and interleukin-8) mRNA and protein expression in PDLFs without cytotoxicity. Next, we observed the anti-inflammatory effect of LJLE in PDLFs after infection with various oral bacteria, including Fusobacterium nucleatum, Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia. These anti-inflammatory effects of LJLE were dose-dependent, and the extract was effective following both pretreatment and posttreatment. Moreover, we found that LJLE suppressed the effect of interleukin-1 beta-induced pro-inflammatory cytokine production in PDLFs. Taken together, these results indicate that LJLE has anti-inflammatory activity that could be exploited to prevent and treat human periodontitis by controlling inflammation.

The Effects of Antimicrobial Peptide Nal-P-113 on Inhibiting Periodontal Pathogens and Improving Periodontal Status.

Periodontal disease consists of chronic gingival inflammation characterized by both degradation of the periodontal connective tissue and alveolar bone loss. Drug therapy is used as an auxiliary treatment method in severe chronic periodontitis, aggressive periodontitis, and periodontitis-associated systemic disease. Nal-P-113, a modified antimicrobial peptide, specifically replaces the histidine residues of P-113 with the bulky amino acid ß-naphthylalanine, and our previous studies have verified that this novel peptide is not toxic to the human body within a certain concentration range. The objective of the present study was to evaluate the effect of Nal-P-113 on periodontal pathogens and periodontal status in clinical studies. In a split-mouth clinical trial, the pocket depth and bleeding index values tended to decrease in the experimental group compared with those in the control group. SEM results verified that Nal-P-113 restrained the maturation of plaque. Based on real-time polymerase chain reaction, the levels of Fusobacterium nucleatum, Streptococcus gordonii, Treponema denticola, and Porphyromonas gingivalis in subgingival plaque were decreased when the subjects were given Nal-P-113. Bacterial growth curve analysis and a biofilm susceptibility assay verified that Nal-P-113 at a concentration of 20 µg/mL restrained the growth of S. gordonii, F. nucleatum, and P. gingivalis and biofilm formation. Therefore, Nal-P-113 effectively reduces periodontal pathogens and ameliorates periodontal status.

Treponema denticola chymotrypsin-like proteinase is present in early-stage mobile tongue squamous cell carcinoma and related to the clinicopathological features.

BACKGROUND: Certain periodontopathogenic bacteria have been linked to cancers. Treponema denticola (Td) is associated with severe periodontitis. Chymotrypsin-like proteinase (CTLP), a major virulence factor of Td, can degrade various host proteins and peptides, and modulate inflammatory responses. However, the role of Td in the tongue carcinogenesis remains unknown. This study aimed to investigate the presence of Td-CTLP in early-stage mobile tongue squamous cell carcinoma (MTSCC) and its relation to clinical and pathological characteristics. METHODS: The immunopositivity of Td-CTLP was assessed in samples obtained from 60 patients with MTSCC and associated with their clinicopathological data. Additionally, Td-CTLP expression was compared with immunoexpression of matrix metalloproteinases (MMP-8 and MMP-9), toll-like receptors (TLR-2, TLR-4, TLR-7 and TLR-9), c-Myc, Ki-67, Bmi-1 and Snail. RESULTS: Treponema denticola-chymotrypsin-like proteinase was present in 95% of MTSCC tumours of which many (40.4%) showed high immunopositivity. Td-CTLP positivity was significantly associated with invasion depth, tumour diameter and the expression of TLR-7, TLR-9 and c-Myc. High Td-CTLP immunopositivity in younger patients (≤ 60 years old) predicted early relapse. CONCLUSION: Our data indicate that Td and its CTLP are present in early-stage MTSCC carcinoma and may contribute to carcinogenesis, and therefore provide novel perspectives into intervention and therapeutic measures of MTSCC.

A Novel Rat Model of Polymicrobial Peri-Implantitis: A Preliminary Study.

BACKGROUND: Peri-implantitis is a complex polymicrobial biofilm-induced inflammatory osteolytic gingival infection that results in orofacial implant failures. To the best knowledge of the authors, there are no preclinical in vivo studies in implant dentistry that have investigated the inflammatory response to known microbial biofilms observed in humans. The aim of this study is to develop a novel peri-implant rat model using an established model of polymicrobial periodontitis. METHODS: Wistar rats were used for the study of experimental peri-implantitis. One month after extraction of maxillary first molars, a titanium mini-implant was inserted. Two months after implant healing, implants were uncovered, and abutment fixing was done using cyanoacrylate to prevent abutment loosening. Rats were separated into two groups (group A: polymicrobial-infected and group B: sham-infected). One week after healing of abutments, rats were infected with Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia for 12 weeks. Bacterial colonization, bone resorption, and implant inflammation were evaluated by polymerase chain reaction (PCR), microcomputed tomography, and histology, respectively. RESULTS: Three rats with four implants in the infection group and two rats with three implants in the sham-infection group were analyzed. PCR analysis revealed presence of bacterial genomic DNA, and infection elicited significant immunoglobulin (Ig)G and IgM antibody responses, indicating bacterial colonization/infection around implants. Infection induced an enhanced mean distance from implant platform to the first bone-to-implant contact, extensive peri-implantitis with advanced bone resorption, and extensive inflammation with granulation tissue and polymorphonuclear leukocytes. CONCLUSIONS: To the best knowledge of the authors, this is the first study to develop a novel rat model of polymicrobial peri-implantitis. With modifications to improve implant retention it could offer significant advantages for studies of initiation and progression of peri-implantitis.

High-risk periodontal pathogens contribute to the pathogenesis of atherosclerosis.

Periodontal disease (PD) is generated by microorganisms. These microbes can enter the general circulation causing a bacteraemia. The result can be adverse systemic effects, which could promote conditions such as cardiovascular disease. Level A evidence supports that PD is independently associated with arterial disease. PD is a common chronic condition affecting the majority of Americans 30 years of age and older. Atherosclerosis remains the largest cause of death and disability. Studies indicate that the adverse cardiovascular effects from PD are due to a few putative or high-risk bacteria: Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola or Fusobacterium nucleatum There are three accepted essential elements in the pathogenesis of atherosclerosis: lipoprotein serum concentration, endothelial permeability and binding of lipoproteins in the arterial intima. There is scientific evidence that PD caused by the high-risk pathogens can influence the pathogenesis triad in an adverse manner. With this appreciation, it is reasonable to state PD, due to high-risk pathogens, is a contributory cause of atherosclerosis. Distinguishing this type of PD as causal provides a significant opportunity to reduce arterial disease.

Treponema denticola, Porphyromonas gingivalis, and Tannerella forsythia induce cell death and release of endogenous danger signals.

OBJECTIVE: The aim of this study was to analyze whether periodontopathogens induced inflammatory cell death and the release of diverse endogenous danger molecules in THP-1-derived macrophages. METHODS: The macrophages were treated with Treponema denticola, Porphyromonas gingivalis, and Tannerella forsythia. Activation of caspase-1 and caspase-4 was detected by Western blotting. Cell death of bacteria-stimulated macrophages was examined using a lactate dehydrogenase (LDH) assay and propidium iodide (PI)/annexin V (AV) staining. Levels of endogenous danger signals, including adenosine triphosphate (ATP), uric acid, heat shock protein 60 (HSP60), high-mobility group box protein 1 (HMGB1), and fibronectin in the culture supernatants were determined using an ATP bioluminescence assay kit, a uric acid assay kit, and Western blotting, respectively. RESULTS: T. denticola, P. gingivalis, and T. forsythia induced activation of caspase-1 and caspase-4. The LDH assay and PI/AV staining showed that all three pathogens induced pyroptotic cell death. All three bacteria induced release of ATP, which is an important ligand for inflammasome activation; the increase in ATP ultimately leads to caspase-1 activation. T. denticola induced release of HSP60 and fibronectin, while T. forsythia induced release of HMGB1 in addition to HSP60 and fibronectin. None of the endogenous molecules except for fibronectin were detected in P. gingivalis-infected cells, possibly due to degradation of these factors by the proteolytic activity of the bacteria. Interestingly, P. gingivalis induced uric acid release. CONCLUSION: Inflammatory cell death and endogenous danger molecules released from cells infected with periodontopathogens may play critical roles in the pathogenesis and progression of periodontitis by augmenting immune and inflammatory responses.

Periodontitis contributes to adipose tissue inflammation through the NF-B, JNK and ERK pathways to promote insulin resistance in a rat model.

This study aimed to investigate the mechanism by which periodontitis affects the inflammatory response and systemic insulin resistance in the white adipose and liver tissues in an obese rat model. The obese model was generated by feeding rats a high fat diet. The periodontitis model was induced by ligatures and injection of "red complex", which consisted of Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia, for two weeks. When compared with rats without periodontitis, fasting glucose levels and homeostasis model assessment index were significantly increased in rats with periodontitis, suggesting that periodontitis promotes the development of insulin resistance in obese rats. Gene and protein expression analysis in white adipose and liver tissue revealed that experimental periodontitis stimulated the expression of inflammatory cytokines, such as tumor necrosis factors-alpha, interleukin-1 beta, toll-like receptor 2 and toll-like receptor 4. Signals associated with inflammation and insulin resistance, including nuclear factor- B, c-Jun amino-terminal kinase and extracellular-signal regulated kinase were significantly activated in the white adipose tissue from obese rats with periodontitis compared to obese rats without periodontitis. Taken together, these findings suggest that periodontitis plays an important role in aggravating the development of local white adipose inflammation and systemic insulin resistance in rat models.

Treponema denticola invasion into human gingival epithelial cells.

Host cell invasion is important for periodontal pathogens in evading host defenses and spreading into deeper areas of the periodontal tissue. Treponema denticola has been implicated in a number of potentially pathogenic processes, including periodontal tissue penetration. Here we tested the ability of T. denticola strains to invade human gingival epithelial cells (HGEC). After 2 h infection, intracellular location of T. denticola cells was confirmed by confocal laser scanning microscopy (CLSM). Results from an antibiotic protection assay following [(3)H]uridine labeling indicated that invasion efficiency reached a maximum at 2 h after infection. Internalized T. denticola cells were still observed in HGEC at 24 h by CLSM. A dentilisin deficient mutant exhibited significantly decreased invasion (p < 0.05) compared with the wild-type strain. In inhibition assays, phenylmethylsulfonyl fluoride and metabolic inhibitors such as methyl-ß-cyclodextrin and staurosporine significantly reduced T. denticola invasion. Under CLSM, T. denticola colocalized with GM-1 ganglioside-containing membrane microdomains in a cholesterol-dependent manner. These results indicated that T. denticola has the ability to invade into and survive within HGECs. Dentilisin activity of T. denticola and lipid rafts on HGEC appear to play important roles in this process.

Interaction of spirochetes with the host fibrinolytic system and potential roles in pathogenesis.

The pathogenic spirochetes Borrelia burgdorferi, B. hermsii, B. recurrentis, Treponema denticola and Leptospira spp. are the etiologic agents of Lyme disease, relapsing fever, periodontitis and leptospirosis, respectively. Lyme borreliosis is a multi-systemic disorder and the most prevalent tick-borne disease in the northern hemisphere. Tick-borne relapsing fever is persistent in endemic areas worldwide, representing a significant burden in some African regions. Periodontal disease, a chronic inflammatory disorder that often leads to tooth loss, is caused by several potential pathogens found in the oral cavity including T. denticola. Leptospirosis is considered the most widespread zoonosis, and the predominant human disease in tropical, undeveloped regions. What these diseases have in common is that they are a significant burden to healthcare costs in the absence of prophylactic measures. This review addresses the interaction of these spirochetes with the fibrinolytic system, plasminogen (Plg) binding to the surface of bacteria and the generation of plasmin (Pla) on their surface. The consequences on host-pathogen interactions when the spirochetes are endowed with this proteolytic activity are discussed on the basis of the results reported in the literature. Spirochetes equipped with Pla activity have been shown to degrade extracellular matrix (ECM) components, in addition to digesting fibrin, facilitating bacterial invasion and dissemination. Pla generation triggers the induction of matrix metalloproteases (MMPs) in a cascade of events that enhances the proteolytic capacity of the spirochetes. These activities in concert with the interference exerted by the Plg/Pla on the complement system - helping the bacteria to evade the immune system - should illuminate our understanding of the mechanisms involved in host infection.