Kenta Kasuyama

Involvement of toll-like receptor 2 and 4 in association between dyslipidemia and osteoclast differentiation in apolipoprotein E deficient rat periodontium

BackgroundDyslipidemia increases circulating levels of oxidized low-density lipoprotein (OxLDL) and this may induce alveolar bone loss through toll-like receptor (TLR) 2 and 4. The purpose of this study was to investigate the effects of dyslipidemia on osteoclast differentiation associated with TLR2 and TLR4 in periodontal tissues using a rat dyslipidemia (apolipoprotein E deficient) model.MethodsLevels of plasma OxLDL, and the cholesterol and phospholipid profiles in plasma lipoproteins were compared between apolipoprotein E-deficient rats (16-week-old males) and wild-type (control) rats. In the periodontal tissue, we evaluated the changes in TLR2, TLR4, receptor activator of nuclear factor kappa B ligand (RANKL) and tartrate resistant acid phosphatase (TRAP) expression.ResultsApolipoprotein E-deficient rats showed higher plasma levels of OxLDL than control rats (p<0.05), with higher plasma levels of total cholesterol (p<0.05) and LDL-cholesterol (p<0.05) and lower plasma levels of high-density lipoprotein cholesterol (p<0.05). Their periodontal tissue also exhibited a higher ratio of RANKL-positive cells and a higher number of TRAP-positive osteoclasts than control rats (p<0.05). Furthermore, periodontal gene expression of TLR2, TLR4 and RANKL was higher in apolipoprotein E-deficient rats than in control rats (p<0.05).ConclusionThese findings underscore the important role for TLR2 and TLR4 in mediating the osteoclast differentiation on alveolar bone response to dyslipidemia.

Effects of Obesity on Gingival Oxidative Stress in a Rat Model

BACKGROUND Studies indicate a correlation between obesity and periodontitis. Oxidative stress is involved in the progression of periodontitis. The purpose of this study was to investigate the effects of obesity on gingival oxidative stress in a rat periodontitis model. METHODS The obese Zucker rats (n = 14) and their lean littermates (n = 14) were each divided into two groups of seven rats. In one of each group, periodontitis was induced by ligature for 4 weeks, whereas the other group was left unligated. The level of 8-hydroxydeoxyguanosine and the ratio of reduced/oxidized glutathione were determined to examine gingival oxidative stress. The serum level of reactive oxygen metabolites and the gingival gene-expression pattern related to oxidative/metabolic stress, inflammation, and cell behavior were also evaluated. RESULTS The obese rats weighed more than the lean rats at 4 weeks. Compared to lean rats, obese rats had enhanced gingival 8-hydroxydeoxyguanosine levels and a decreased ratio of reduced/oxidized glutathione in the gingival tissue, with increasing serum reactive oxygen metabolites. However, there were no significant differences in the degree of alveolar bone loss between lean and obese rats, except for teeth with and without ligatures in both rats. In addition, the periodontal lesion in obese rats showed higher 8-hydroxydeoxyguanosine levels and polymorphonuclear leukocyte infiltration than the inflamed ones in lean rats, with downregulation of multiple cytochrome P450 gene expression. CONCLUSIONS Obesity induced gingival oxidative stress with increasing serum reactive oxygen metabolites in rats. In the periodontal lesion, gene expressions related to a capacity for xenobiotic detoxification were downregulated in the obese model.

Effects of periodontitis on aortic insulin resistance in an obese rat model

The combination of obesity and its associated risk factors, such as insulin resistance and inflammation, results in the development of atherosclerosis. However, the effects of periodontitis on atherosclerosis in an obese body remain unclear. The aim of the study was to investigate the effects of ligature-induced periodontitis in Zucker fatty rats on initiation of atherosclerosis by evaluating aortic insulin resistance. Zucker fatty rats (n=24) were divided into two groups. In the periodontitis group, periodontitis was ligature-induced for 4 weeks, whereas the control group was left unligated. After the 4-week experimental period, descending aorta was used for measuring the levels of lipid deposits, immunohistochemical analysis, and evaluation of gene expression. Levels of serum C-reactive protein (CRP), tumor necrosis factor-α (TNF-α), and insulin were also measured. Rats in the periodontitis group had significantly enhanced lipid deposits in the aorta, but not in the control group. Expression of suppressor of cytokine signaling 3, vascular cell adhesion molecule 1, reactive oxygen species, nitrotyrosine, and endothelin-1 in the periodontitis group was more intense than that in the control group. Significantly decreased levels of phosphatidylinositol 3-kinase (Pi3k) catalytic β-polypeptide (Pi3kcb), Pi3kp85, and insulin receptor substrate 1 and 2 were observed in the periodontitis group. Levels of serum CRP and TNF-α were significantly increased in the periodontitis group. Under insulin-stimulated conditions, aorta in the periodontitis group altered the Akt phosphorylation. Periodontitis in obesity induced the initial stage of atherosclerosis and disturbed aortic insulin signaling.

Combined effects of hydrogen sulfide and lipopolysaccharide on osteoclast differentiation in rats.

BACKGROUND Lipopolysaccharide (LPS) stimulates osteoclast differentiation through toll-like receptors (TLRs) 2 and 4, and hydrogen sulfide (H(2)S) induces osteoclast differentiation. If H(2)S activates TLRs, H(2)S may enhance the effects of LPS on osteoclast differentiation. The purpose of the present study is to examine the combined effects of sodium hydrogen sulfide (NaHS, an H(2)S donor drug) and LPS on osteoclast differentiation and TLR expression in rat periodontal tissue. METHODS Twenty-eight male Wistar rats (8 weeks old) were divided into four groups (n = 7 per group): a control (no treatment) group and three experimental groups (NaHS group, LPS group, and a combination [NaHS + LPS] group). At 1 day after topical application of NaHS and/or Porphyromonas gingivalis LPS into the gingival sulcus of first molars, the number of tartrate-resistant acid phosphate (TRAP)-positive osteoclasts in the periodontal tissue was counted. Expression of TLR2 and TLR4 mRNAs and proteins in the gingival was also assessed. RESULTS The number of TRAP-positive osteoclasts was significantly higher in the combination group than in any other group (P <0.01). The combination group had 11.0-fold higher TLR4 mRNA levels than the control group. TLR4 protein levels were also higher in the combination group than in the NaHS or LPS group. However, the TLR2 mRNA and protein levels were not significantly different in the combination group and the LPS group. CONCLUSION In rat periodontal tissue, NaHS and LPS had an additive effect on osteoclast differentiation through activation of the TLR4 pathway but not the TLR2 pathway.

Effects of exercise training on gingival oxidative stress in obese rats

OBJECTIVE The purpose of the present study was to investigate the effects of exercise training on serum reactive oxygen species (ROS) level and gingival oxidative stress in obese rats fed a high-fat diet. DESIGN Rats were divided into three groups (n = 14/group): one control group (fed a regular diet) and two experimental groups (fed a high-fat diet with and without exercise training [treadmill: 5 days/week]). The rats were sacrificed at 4 or 8 weeks. The level of serum reactive oxidative metabolites (ROM) was measured as an indicator of circulating ROS. The level of 8-hydroxydeoxyguanosine (8-OHdG) and reduced-form glutathione (GSH)/oxidised-form glutathione (GSSG) ratio were determined to evaluate gingival oxidative stress. RESULTS The obese rats fed a high-fat diet without exercise training showed higher serum ROM levels [Carratelli Units (CARR U)] (mean ± SD; 413 ± 64) than the control (333 ± 12) at 4 weeks (p = 0.023). Such a condition resulted in higher 8-OHdG levels (ng/mg mtDNA) (0.97 ± 0.18) (p < 0.05) and a lower GSH/GSSG ratio (17.0 ± 3.1) (p < 0.05) in gingival tissues, compared to the control (0.55 ± 0.13 for 8-OHdG and 23.6 ± 5.8 for GSH/GSSG ratio) at 8 weeks. In addition, the obese rats fed a high-fat diet with exercise training showed lower serum ROM (623 ± 103) (p < 0.001) and gingival 8-OHdG levels (0.69 ± 0.17) (p = 0.012) than those without exercise training (1105 ± 95 for ROM and 0.55 ± 0.13 for 8-OHdG) at 8 weeks. CONCLUSIONS Obesity prevention by exercise training may effectively suppress gingival oxidative stress by decreasing serum ROS in rats.

Hydrogen-rich water prevents lipid deposition in the descending aorta in a rat periodontitis model

OBJECTIVE Periodontitis has been causally linked to atherosclerosis, which is mediated by the oxidative stress. As hydrogen-rich water (HW) scavenges reactive oxygen species (ROS), we hypothesized that HW could prevent lipid deposition induced by periodontitis in the aorta. The aim of this study was to investigate the effects of HW on the initiation of atherosclerosis in a rat periodontitis model. DESIGN Eighteen 8-wk-old male Wistar rats were divided into three groups of six rats; the periodontitis group, periodontitis+HW group and the no treatment (control) group. In the periodontitis and periodontitis+HW groups, periodontitis was induced using a ligature for 4 wk, while the periodontitis+HW group was given water containing 800-1000 μg/L hydrogen during the 4-wk experimental period. RESULTS In the periodontitis group, lipid deposition in the descending aorta was observed. The periodontitis group also showed significant higher serum levels for ROS and oxidised low-density lipoprotein-cholesterol (ox-LDL) (1.7 and 1.4 times, respectively), and higher aortic expression levels of nitrotyrosine and hexanoyl-lysine (HEL) (7.9 and 16.0 times, respectively), as compared to the control group (p<0.05). In the periodontitis+HW group, lipid deposition was lower. Lower serum levels of ROS and ox-LDL (0.46 and 0.82 times, respectively) and lower aortic levels of nitrotyrosine and HEL (0.27 and 0.19 times, respectively) were observed in the periodontitis+HW group than in the periodontitis group (p<0.05). CONCLUSIONS HW intake may prevent lipid deposition in the rat aorta induced by periodontitis by decreasing serum ox-LDL levels and aortic oxidative stress.

Anti-aging Effects of Co-enzyme Q10 on Periodontal Tissues

Oxidative stress is associated with age-related reactions. The anti-oxidative effects of a reduced form of co-enzyme Q10 (rCoQ10) suppress oxidative stress, which may contribute to the prevention of age-related inflammatory reactions. We examined the effects of topically applied rCoQ10 on periodontal inflammatory reactions in a rat aging model. Male Fischer 344 rats, 2 (n = 6) and 4 mos (n = 18) of age, were used. All of the two-month-old rats and 6 of the four-month-old rats were sacrificed and 12 remaining four-month-old rats received topically applied ointment with or without 1% rCoQ10 on the gingival surface until they reached 6 mos of age. The rats showed an age-dependent increase in circulating oxidative stress. RCoQ10 decreased oxidative DNA damage and tartrate-resistant acid-phosphatase-positive osteoclasts in the periodontal tissue at 6 mos of age as compared with the control. The same conditions lowered gene expression of caspase-1 and interleukin-1β in the periodontal tissue. Furthermore, Nod-like receptor protein 3 inflammasomes were less activated in periodontal tissues from rCoQ10-treated rats as compared with the control rats. Our results suggest that rCoQ10 suppresses age-related inflammatory reactions and osteoclast differentiation by inhibiting oxidative stress.

Effects of hydrogen-rich water on aging periodontal tissues in rats

Oxidative damage is involved in age-related inflammatory reactions. The anti-oxidative effects of hydrogen-rich water suppress oxidative damage, which may aid in inhibiting age-related inflammatory reactions. We investigated the effects of drinking hydrogen-rich water on aging periodontal tissues in healthy rats. Four-month-old male Fischer 344 rats (n = 12) were divided into two groups: the experimental group (hydrogen-rich water treatment) and the control group (distilled water treatment). The rats consumed hydrogen-rich water or distilled water until 16 months of age. The experimental group exhibited lower periodontal oxidative damage at 16 months of age than the control group. Although protein expression of interleukin-1β did not differ, gene expression of Nod-like receptor protein 3 inflammasomes was activated in periodontal tissues from the experimental group as compared with the control group. Drinking hydrogen-rich water is proposed to have anti-aging effects on periodontal oxidative damage, but not on inflammatory reactions in healthy rats.

Effects of topical application of inorganic polyphosphate on tissue remodeling in rat inflamed gingiva

BACKGROUND AND OBJECTIVE Inorganic polyphosphate [poly(P)] is a biopolymer found in almost all cells and tissues, and which promotes tissue remodeling. However, there is limited information on how poly(P) affects the connective tissue in inflamed gingiva. This study examined the effects of topical application of poly(P) on gingival connective tissue and its remodeling in a rat periodontitis model. MATERIAL AND METHODS Male Wistar rats (n = 36, 8 wk of age) were used in this 6-wk study. The rats were divided into six groups of six rats each. The control group received no treatment. In the other groups, periodontitis was ligature-induced for 4 wk. After 4 wk, the rats with periodontitis were further divided into five groups, and were left untreated (periodontitis group) or subjected to topical application of oral rinses containing 0, 0.1, 1 or 5% poly(P) for 2 wk. RESULTS The periodontitis and 0% poly(P) groups showed a higher density of polymorphonuclear leukocytes and a lower density of collagen in gingival tissue than the control group (p < 0.05). In contrast, groups treated with more than 1% poly(P) exhibited a lower density of polymorphonuclear leukocytes (p < 0.05) and a higher density of collagen than the periodontitis and 0% poly(P) groups (p < 0.05). A higher expression of fibroblast growth factor-2 was observed in the gingiva of rats treated with 1% poly(P) than in those treated with 0% poly(P) (p < 0.05). CONCLUSION Topical application of poly(P) may induce connective tissue remodeling, contributing to improvement of inflamed gingiva in rats.

A new portable monitor for measuring odorous compounds in oral, exhaled and nasal air

BackgroundThe B/B Checker®, a new portable device for detecting odorous compounds in oral, exhaled, and nasal air, is now available. As a single unit, this device is capable of detecting several kinds of gases mixed with volatile sulfur compounds (VSC) in addition to other odorous gasses. The purpose of the present study was to evaluate the effectiveness of the B/B Checker® for detecting the malodor level of oral, exhaled, and nasal air.MethodsA total of 30 healthy, non-smoking volunteers (16 males and 14 females) participated in this study. The malodor levels in oral, exhaled, and nasal air were measured using the B/B Checker® and by organoleptic test (OT) scores. The VSCs in each air were also measured by gas chromatography (GC). Associations among B/B Checker® measurements, OT scores and VSC levels were analyzed using Spearman correlation coefficients. In order to determine the appropriate B/B Checker® level for screening subjects with malodor, sensitivity and specificity were calculated using OT scores as an identifier for diagnosing oral malodor.ResultsIn oral and nasal air, the total VSC levels measured by GC significantly correlated to that measured by the B/B Checker®. Significant correlation was observed between the results of OT scores and the B/B Checker® measurements in oral (r = 0.892, p < 0.001), exhaled (r = 0.748, p < 0.001) and nasal air (r = 0.534, p < 0.001). The correlation between the OT scores and VSC levels was significant only for oral air (r = 0.790, p < 0.001) and nasal air (r = 0.431, p = 0.002); not for exhaled air (r = 0.310, p = 0.096). When the screening level of the B/B Checker® was set to 50.0 for oral air, the sensitivity and specificity were 1.00 and 0.90, respectively. On the other hand, the screening level of the B/B Checker® was set to 60.0 for exhaled air, the sensitivity and specificity were 0.82 and 1.00, respectively.ConclusionThe B/B Checker® is useful for objective evaluation of malodor in oral, exhaled and nasal air and for screening subjects with halitosis.Trial registrationClinicalTrials.gov: NCT01139073