Influence of a 3-month training program on muscular damage and neutrophil function in male university freshman judoists.

We studied the effects of a high intensity and high frequency 3-month training program on muscle damage and neutrophil function in male judoists. The study included 15 male judoists who started intensive judo training program after a 6-month break. Creatine kinase (CK), neutrophil counts and reactive oxygen species (ROS) production capability as well as phagocytic activity (PA) of neutrophils were measured at 2 stages; entering university (pre-training) and after 3-month training (post-training). At both points, we investigated parameters three times: just before, immediately after and 24 h after a 2-h practice session. Practice-mediated change in CK was lower at post-training than at pre-training. Neutrophil count significantly increased after 2-h practice but recovered 24 h later whereas it showed no subsequent and further increased at 24 h post-practice. Although neutrophil ROS production capability and PA both decreased (breakdown) after practice session, ROS production capability increased and PA decreased (well-adapted) at the post-training. Long-term training strengthened muscular function and improved neutrophil reaction against practice-mediated stress.

Relationship between periodontitis and hepatic condition in Japanese women.

The liver is an important organ closely associated with lipid and glucose metabolism. This study was performed to clarify the relationship between periodontitis and hepatic condition in apparently healthy Japanese women. A cross-sectional study was performed on 172 apparently healthy, dentulous Japanese women (20-59 years old) who attended a health promotion program at Fukuoka Health Promotion Center. After multivariate adjustment for age, smoking history and oral hygiene, which were known risk factors for periodontitis, the incidence of periodontitis (deepest probing depth > or =4 mm) in females was significantly increased with elevated serum levels of aspartate aminotransferase (AST, p < 0.01), alanine aminotransferase (ALT, p < 0.01) and cholinesterase (p < 0.001), and an AST-to-ALT ratio of less than one (p = 0.02). Further adjustment for either body mass index (BMI) or percent body fat did not attenuate these relationships. These results suggest that hepatic steatosis is associated with periodontitis in Japanese women.

3-Chloro-DL-alanine resistance by L-methionine-alpha-deamino-gamma-mercaptomethane-lyase activity.

The antibacterial agent 3-chloro-DL-alanine (3CA) is an inhibitor of peptidoglycan synthesis. Fusobacterium nucleatum and Porphyromonas gingivalis, the bacteria responsible for oral malodor, are shown to be resistant to 1 mM 3CA, whereas Streptococcus mutans and Escherichia coli are sensitive to this antibacterial agent at the same concentration. We isolated the 3CA resistance gene from F. nucleatum and showed that the gene encodes an L-methionine-alpha-deamino-gamma-mercaptomethane-lyase that catalyzes the alpha,gamma-elimination of L-methionine to produce methyl mercaptan. The enzyme also exhibits 3CA chloride-lyase (deaminating) activity. This antibacterial agent is expected to be useful for specific selection of malodorous oral bacteria producing high amounts of methyl mercaptan.

A novel mechanism for glucose side-chain formation in rhamnose-glucose polysaccharide synthesis.

We have cloned two genes (rgpH and rgpI) that encode proteins for the formation of the glucose side-chains of the Streptococcus mutans rhamnose-glucose polysaccharide (RGP), which consists of a rhamnan backbone with glucose side-chains. The roles of rgpH and rgpI were evaluated in a rhamnan-synthesizing Escherichia coli. An E. coli strain that harbored rgpH reacted with antiserum directed against complete RGP, whereas the E. coli strain that carried rgpI did not react with this antiserum. Although E. coli:rgpH reacted strongly with rhamnan-specific antiserum, co-transformation of this strain with rgpI increased the number of glucose side-chains and decreased immunoreactivity with the rhamnan-specific antiserum significantly. These results suggest that two genes are involved in side-chain formation during S. mutans RGP synthesis in E. coli: one gene encodes a glucosyltransferase, and the other gene probably controls the frequency of branching. This is the first report to identify a gene that is involved in regulation of branching frequency in polysaccharide synthesis.

Correlation between oral malodor and periodontal bacteria.

Volatile sulfur compounds (VSCs), including hydrogen sulfide, methyl mercaptan, and dimethyl sulfide, are primarily responsible for oral malodor. Recently, the mgl gene encoding L-methionine-alpha-deamino-gamma-mercaptomethane-lyase, which produces methyl mercaptan, was cloned from Porphyromonas gingivalis. This article discusses the mechanism and pathogenic role of the formation of VSCs by oral bacteria.

Cloning and characterization of the L-cysteine desulfhydrase gene of Fusobacterium nucleatum.

Hydrogen sulfide and methyl mercaptan are the two major compounds associated with oral malodor. These compounds are highly toxic, and are thought to play an important role in periodontal disease. Fusobacterium nucleatum, an oral bacterium, produces large amounts of hydrogen sulfide from L-cysteine by the enzymatic action of L-cysteine desulfhydrase. We cloned and sequenced the cdl gene encoding L-cysteine desulfhydrase from F. nucleatum ATCC 10953, and revealed that the structural cdl gene consists of 921 bp and encodes a 33.4-kDa protein. The cloned gene was inserted into an expression vector, pDEST17, and expressed in Escherichia coli as a fused protein. The purified enzyme was tested for substrate specificity using various SH-containing compounds. Only L-cysteine served as a substrate for L-cysteine desulfhydrase to produce hydrogen sulfide.

Enhanced production of vascular endothelial growth factor by human monocytic cells stimulated with endotoxin through transcription factor SP-1.

The effect of endotoxin on the regulation of vascular endothelial growth factor (VEGF) mRNA expression in human monocytic (THP-1) cells was examined. Endotoxic lipopolysaccharide (LPS) from Escherichia coli and synthetic E. coli-type lipid A (LA-15-PP) enhanced VEGF mRNA expression. LPS-induced VEGF mRNA accumulation was regulated, at least in part, at the transcriptional level. Enhancement of VEGF gene expression by LPS was shown by gel shift analysis and use of transcription factor inhibitors to be mediated via the activation of SP-1.

Association between alveolar bone loss and elevated serum C-reactive protein in Japanese men.

BACKGROUND: Moderate elevation of C-reactive protein (CRP) is thought to predict type 2 diabetes and cardiovascular disease (CVD), both of which are associated with periodontitis. Recent studies indicate that periodontal disease is associated with moderate elevation of CRP; however, the relationship between alveolar bone loss (ABL) and CRP elevation is unclear. METHODS: A total of 179 Japanese men aged 50 to 54 years old, with at least 10 teeth, were examined as part of a comprehensive health examination before retirement from the Japan Self-Defense Force. ABL was measured at proximal sites of posterior teeth on a panoramic x-ray film. The relationship between the mean ratio of ABL to root length and serum CRP level and other variables was analyzed. RESULTS: ABL was significantly correlated with serum CRP level (P = 0.008), alkaline phosphatase (P = 0.008), high-density lipoprotein (HDL) cholesterol (P = 0.04, inversely), white blood cell count (P < 0.001), erythrocyte sedimentation rate (P = 0.002), age (P = 0.03), and smoking history (P < 0.001). In a multiple logistic regression model adjusted for age, smoking history, systolic blood pressure, body-mass index, triglyceride, and HDL cholesterol, subjects in the highest tertile of ABL had an increased risk for CRP elevation > or = 1.3 mg/l (odds ratio [OR] = 8.20; 95% confidence interval [CI], 1.6 to 40.7; P = 0.01) when compared to the lowest tertile of ABL. CONCLUSION: ABL around posterior teeth was associated with elevated CRP in Japanese men, suggesting an association between periodontal disease and increased risk of type 2 diabetes and CVD.

Relationship between electrocardiographic abnormalities and periodontal disease: the Hisayama Study.

BACKGROUND: Recent studies have suggested a relationship between periodontitis and cardiovascular disease (CVD). This study investigated the relationship between periodontitis and electrocardiographic (ECG) abnormalities, which are known predictors of CVD. METHODS: We examined the periodontal status of 1,111 residents of Hisayama Town, Fukuoka, Japan. Nine hundred fifty-seven (957) subjects (374 males, 583 females) with > or = 10 teeth and without a medical history of CVD were included in the analysis. Probing depth (PD) and clinical attachment level (CAL) were measured on two randomly selected quadrants, one maxillary and one mandibular. A 12-lead ECG was recorded using a standard electrocardiograph. ECG abnormalities included left ventricular hypertrophy (Minnesota code 3-1) and ST depression (4-1, 2, 3). The relation of periodontal condition and ECG abnormalities was assessed with logistic regression analysis. RESULTS: Univariate analysis revealed that mean probing depth, mean attachment loss, number of teeth, and plaque index were significantly associated with ECG abnormalities, as well as with known risk factors of CVD. In multivariate analysis, the subjects with deep pockets (mean probing depth > or = 2 mm) had an increased risk for ECG abnormalities (odds ratio [OR] = 1.6; 95% confidence interval [CI] = 1.01 to 2.50) compared to the subjects with mean PD < 2 mm. Subjects with severe attachment loss (mean CAL > or = 2.5 mm) had also significant risk for ECG abnormalities (OR = 1.7; 95% CI = 1.07 to 2.67) compared to those whose mean CAL was < 2.5 mm. CONCLUSION: This study clearly shows the relationship between periodontitis and ECG abnormalities, which are important predictors of CVD.

Pulmonary hyperinflation and respiratory distress following solvent aspiration in a patient with asthma: expectoration of bronchial casts and clinical improvement with high-frequency chest wall oscillation.

An 18-year-old student with a history of asthma accidentally inhaled organic solvent during a class, with immediate cough and dyspnea that worsened over several hours. He presented in severe respiratory distress, with hypoxemia and marked pulmonary hyperinflation. Administration of inhaled bronchodilator was ineffective because of agitation, and the patient could not be positioned for chest physiotherapy to treat presumed widespread mucus plugging. High-frequency chest wall oscillation (HFCWO) in the sitting position initially caused increased distress but was subsequently tolerated when noninvasive positive-pressure ventilation (NPPV) via nasal mask was initiated. Almost immediately, the patient began expectorating bronchial mucus casts, with concomitant clinical improvement. Endotracheal intubation was avoided, and with aggressive pharmacologic treatment for acute severe asthma and continuation of intermittent HFCWO-NPPV, the patient made a full recovery over the next several days. This case suggests that the combination of HFCWO and NPPV may be helpful in the presence of mucus plugging as a complication of acute inhalation injury or acute severe asthma.

Monitoring ammonia to assess halitosis.

OBJECTIVE: This study examined the applicability of ammonia monitoring for assessing halitosis. STUDY DESIGN: The actual degree of halitosis was determined by using an organoleptic test in 61 subjects aged 28 +/- 10 years (mean +/- SD). Levels of volatile sulfur compounds and ammonia were determined by using gas chromatography and ammonia monitoring, respectively. Levels of ammonia and methyl mercaptan produced by bacteria in dental plaque and tongue-coating samples obtained from 25 subjects were quantified. In addition, changes in ammonia levels were measured before and after removing tongue coating or dental plaque. RESULTS: There was no significant correlation between the organoleptic score and the ammonia level measured with ammonia monitoring, whereas there was a significant correlation between ammonia level and the total level of volatile sulfur compounds measured with gas chromatography. Significant correlations were also observed between ammonia level and levels of methyl mercaptan produced by bacteria in dental plaque and tongue coating. Furthermore, the ammonia level decreased after the removal of tongue coating and dental plaque. CONCLUSION: These results indicate that measuring ammonia levels is useful for assessing halitosis, specifically for halitosis arising from a lack of oral hygiene.

Risk factors for tooth loss in the institutionalised elderly; a six-year cohort study.

OBJECTIVE: The aim of this study was to examine risk factors for tooth loss and edentulism in institutionalised elderly people. DESIGN: Six-year prospective cohort study. SETTING: Twenty nine of the 30 institutions for elderly people in Kitakyushu, Japan. SUBJECTS: Four hundred and eighteen of the 1,096 subjects who were dentate at the baseline examination. OUTCOME MEASURES: Factors that influenced tooth loss and edentulism. RESULTS: In both sexes, older subjects had fewer remaining teeth at baseline. Males lost more teeth than females, and edentulism was also more likely in males than in females. Stepwise multiple linear regression analysis showed that the number of teeth, number of decayed teeth, periodontal pocket depth, and plaque index were significant predictors for the number of teeth lost. The incidence of edentulism was higher in males than in females. Stepwise multiple logistic regression analysis revealed that poorer mental health status, fewer teeth, having deep periodontal pocket, and poorer oral hygiene status were independently associated with the six-year incidence of edentulism. CONCLUSIONS: Tooth loss in institutionalised elderly people in Japan is related to both poor oral health status and poor mental health status.

Breathing intolerance index: a new indicator for ventilator use.

OBJECTIVES: The purpose was to study the efficacy of a proposed breathing intolerance index for justifying ventilator use by patients with pulmonary or chest wall diseases and to compare with values obtained from healthy controls. DESIGN: A total of 42 patients with lung/chest wall disease, including 11 ventilator users and 25 age-matched controls, were studied. The breathing intolerance index was defined as (Ti/Ttot) x (Vt/VC), where Ti = inspiratory time of one breath (in seconds), Ttot = total time of one breath (in seconds), Vt = tidal volume (in milliliters) at rest, and VC = vital capacity (in milliliters). A digital spirometer with custom computer software was used. RESULTS: The examinations were completed uneventfully. The mean breathing intolerance index values of the 25 healthy volunteers, the 31 nonventilator user patients, and the 11 users of nocturnal noninvasive positive-pressure ventilation were 0.050 +/- 0.009 (mean +/- standard deviation), 0.087 +/- 0.022, and 0.186 +/- 0.038, respectively. The breathing intolerance indices of the ventilator users were significantly greater (P < 0.0001) than those of the other nonventilator user groups, and there was no overlap in values. CONCLUSIONS: Ventilator users have a significantly higher breathing intolerance index than nonventilator users. The index may be useful for justifying ventilator use.

L-Methionine-gamma-lyase, as a target to inhibit malodorous bacterial growth by trifluoromethionine.

Methyl mercaptan is a major component responsible for oral malodor. This compound arises from the bacterial metabolism of methionine. Here we show that the growth of Porphyromonas gingivalis, a periodontal microorganism that produces large amounts of methyl mercaptan, was strongly inhibited by l-trifluoromethionine (TFM), a fluorinated derivative of methionine. In contrast, TFM had no effect on the growth of bacteria which do not produce methyl mercaptan. In addition, the survival rate of P. gingivalis-infected mice was remarkably increased by the co-injection of TFM. These results suggest that TFM is a promising antibacterial agent specific to the malodorous oral bacteria.

Functional domain of bovine milk lactoferrin which inhibits the adherence of Streptococcus mutans cells to a salivary film.

The bovine lactoferrin molecule and relatively long lactoferrin fragments containing residues 473 to 538 strongly inhibited adherence of Streptococcus mutans to saliva-coated hydroxyapatite beads. Each cysteine residue in Lf411 (residues 473 to 538) was replaced by a serine residue, and the mutants Lf411-C481S and Lf411-C532S strongly inhibited S. mutans adherence. These results suggest that the functional domain of lactoferrin that binds to a salivary film lies in residues 473 to 538 and that the region might be concealed by disulfide bond formation between Cys481 and Cys532 in the Lf411 fragment.

Immunization against dental caries.

Dental caries is one of the most common infectious diseases. Of the oral bacteria, mutans streptococci, such as Streptococcus mutans and S. sobrinus, are considered to be causative agents of dental caries in humans. There have been numerous studies of the immunology of mutans streptococci. To control dental caries, dental caries vaccines have been produced using various cell-surface antigens of these organisms. Progress in recombinant DNA technology and peptide synthesis has been applied to the development of recombinant and synthetic peptide vaccines to control dental caries. Significant protective effects against dental caries have been shown in experimental animals, such as mice, rats and monkeys, which have been subcutaneously, orally, or intranasally immunized with these antigens. Only a few studies, however, have examined the efficacy of dental caries vaccines in humans. Recently, local passive immunization using murine monoclonal antibodies, transgenic plant antibodies, egg-yolk antibodies, and bovine milk antibodies to antigens of mutans streptococci have been used to control the colonization of the organisms and the induction of dental caries in human. Such immunization procedures may be a safer approach for controlling human dental caries than active immunization.

Tumor necrosis factor alpha enhances Actinobacillus actinomycetemcomitans leukotoxin-induced HL-60 cell apoptosis by stimulating lymphocyte function-associated antigen 1 expression.

We demonstrated previously that Actinobacillus actinomycetemcomitans leukotoxin (Ltx) is greatly able to induce apoptotic signaling in cells that are positive for lymphocyte function-associated antigen 1 (LFA-1), a cell receptor of Ltx. We investigated in this study whether inflammatory cytokines can regulate apoptosis of human leukemic HL-60 cells induced by Ltx. Of the cytokines tested, tumor necrosis factor alpha (TNF-alpha) significantly enhanced the Ltx-induced cell apoptosis. Northern and Western blotting analyses showed that TNF-alpha enhanced the expression of CD11a in the cells at both the mRNA and protein levels but did not do so for CD18 expression. TNF-alpha also enhanced the binding of Ltx to the cells. We also observed by measuring the mitochondrial transmembrane potential and the generation of superoxide anion that the cytokine enhanced Ltx-induced apoptosis in HL-60 cells. In addition, interleukin-1beta significantly enhanced Ltx-induced cell apoptosis, although the enhancing activity was lower than that of TNF-alpha. These stimulatory effects of both cytokines were also observed for human polymorphonuclear leukocytes. The ability of TNF-alpha to increase cell susceptibility to Ltx could be inhibited by preincubation of the cells with a monoclonal antibody against TNF receptor 1 but not by preincubation of the cells with a monoclonal antibody against anti-TNF receptor 2. Furthermore, the results of an assay of caspase 3 intracellular activity (PhiPhiLuxG1D2) showed that Ltx-induced caspase 3 activation was completely neutralized by CD18 antibody treatment, although significant neutralization was also observed with anti-CD11a antibody. Taken together, the results of the present study indicate that TNF-alpha acts as a potent stimulator of Ltx-induced HL-60 cell apoptosis via TNF receptor 1-mediated upregulation of LFA-1 expression.

Expression and characterization of streptococcal rgp genes required for rhamnan synthesis in Escherichia coli.

Six genes (rgpA through rgpF) that were involved in assembling the rhamnose-glucose polysaccharide (RGP) in Streptococcus mutans were previously identified (Y. Yamashita, Y. Tsukioka, K. Tomihisa, Y. Nakano, and T. Koga, J. Bacteriol. 180:5803-5807, 1998). The group-specific antigens of Lancefield group A, C, and E streptococci and the polysaccharide antigen of Streptococcus sobrinus have the same rhamnan backbone as the RGP of S. mutans. Escherichia coli harboring plasmid pRGP1 containing all six rgp genes did not synthesize complete RGP. However, E. coli carrying a plasmid with all of the rgp genes except for rgpE synthesized the rhamnan backbone of RGP without glucose side chains, suggesting that in addition to rgpE, another gene is required for glucose side-chain formation. Synthesis of the rhamnan backbone in E. coli required the initiation of transfer of N-acetylglucosamine to a lipid carrier and the expression of the rgpC and rgpD genes encoding the putative ABC transporter specific for RGP. The similarities in RGP synthesis between E. coli and S. mutans suggest common pathways for rhamnan synthesis. Therefore, we evaluated the rhamnosyl polymerization process in E. coli by high-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the lipooligosaccharide (LOS). An E. coli transformant harboring rgpA produced the LOS modified by the addition of a single rhamnose residue. Furthermore, the rgpA, rgpB, and rgpF genes of pRGP1 were independently mutated by an internal deletion, and the LOS chemotypes of their transformants were examined. The transformant with an rgpA deletion showed the same LOS profile as E. coli without a plasmid. The transformant with an rgpB deletion showed the same LOS profile as E. coli harboring rgpA alone. The transformant with an rgpF deletion showed the LOS band with the most retarded migration. On the basis of these results, we speculated that RgpA, RgpB, and RgpF, in that order, function in rhamnan polymerization.

Development of a 5' fluorogenic nuclease-based real-time PCR assay for quantitative detection of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis.

A 5' nuclease TaqMan PCR was developed for the quantitative detection of the periodontopathic bacteria Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. The relative numbers of bacteria were measured by the comparative threshold cycle method. This simplified method is a way of obtaining the relative quantities of these organisms from specimens and of monitoring the effect of therapy.

Development of a 5' nuclease-based real-time PCR assay for quantitative detection of cariogenic dental pathogens Streptococcus mutans and Streptococcus sobrinus.

A 5' nuclease TaqMan PCR assay was developed for the quantitative detection of the major cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus. The absolute and relative numbers of bacteria were measured by this method. This assay will be useful for quantifying these organisms in oral specimens and for analyzing biofilm formation.