Drinking Hydrogen-Rich Water Has Additive Effects on Non-Surgical Periodontal Treatment of Improving Periodontitis: A Pilot Study.

Oxidative stress is involved in the pathogenesis of periodontitis. A reduction of oxidative stress by drinking hydrogen-rich water (HW) might be beneficial to periodontal health. In this pilot study, we compared the effects of non-surgical periodontal treatment with or without drinking HW on periodontitis. Thirteen patients (3 women, 10 men) with periodontitis were divided into two groups: The control group (n = 6) or the HW group (n = 7). In the HW group, participants consumed HW 4-5 times/day for eight weeks. At two to four weeks, all participants received non-surgical periodontal treatment. Oral examinations were performed at baseline, two, four and eight weeks, and serum was obtained at these time points to evaluate oxidative stress. At baseline, there were no significant differences in periodontal status between the control and HW groups. The HW group showed greater improvements in probing pocket depth and clinical attachment level than the control group at two, four and eight weeks (p < 0.05). The HW group also exhibited an increased serum level of total antioxidant capacity at four weeks, compared to baseline (p < 0.05). Drinking HW enhanced the effects of non-surgical periodontal treatment, thus improving periodontitis.

Occlusal disharmony accelerates the initiation of atherosclerosis in apoE knockout rats.

BACKGROUND: Psychosocial stress is one of the risk factors for atherosclerosis. As occlusal disharmony induces psychological stress, we hypothesized that psychological stress by occlusal disharmony accelerates atherosclerosis. The aim of this study was to investigate the effects of occlusal disharmony on the initiation of atherosclerosis in apolipoprotein E (apoE) knockout rats. METHODS: Fourteen male apoE-knockout rats (age; 8 weeks) (Sprague-Dawley strain background) were divided into two groups of seven rats: the occlusal disharmony group and the no treatment (control) group. In the occlusal disharmony group, the maxillary molar cusps were cut off for the 8-week experimental period. RESULTS: In the occlusal disharmony group, the percentages of the area of total aortic lumen occupied by plaques and lipid were significantly higher than those in the control group (p < 0.05, t-test). The occlusal disharmony group also showed significantly higher serum levels of very low-density lipoprotein-cholesterol (VLDL) and low-density lipoprotein-cholesterol (LDL), plasma levels of corticosterone (1.9, 1.3 and 1.3 times, respectively), higher aortic protein expression levels of vascular cell adhesion molecule-1 (VCAM1) and intercellular adhesion molecule-1 (ICAM1) (1.5 and 1.4 times, respectively), and higher aortic gene expression of levels of VCAM1 and Toll-like receptor 4 (TLR4) (1.9 and 4.3 times, respectively), as compared to the control group (p < 0.05). However, there were no significant differences in serum levels of oxidized LDL, reactive oxygen metabolites and C-reactive protein between the two groups. CONCLUSION: In apoE knockout rats, occlusal disharmony may induce VCAM1, ICAM1 and TLR4 expression and accelerate the initiation of atherosclerosis.

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-1ß 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.

Anti-ageing effects of dentifrices containing anti-oxidative, anti-inflammatory, and anti-bacterial agents (Tomarina®) on gingival collagen degradation in rats.

OBJECTIVE: Previous studies have demonstrated the relationship between ageing and oxidative stress. In this study, we examined the effects of topical application of a dentifrice containing anti-oxidative, anti-inflammatory, and anti-bacterial agents (Tomarina®) to the gingival surface on gingival collagen degradation in rats. DESIGN: Fischer 344 male rats (4 or 8 months old) were divided into two groups: experimental group and control group. Tomarina® (the experimental group) or control dentifrice (the control group) was applied 5 days per week for 2 months. RESULTS: In the control group, gingival collagen density decreased with ageing. In the experimental group, the collagen density did not change with ageing, and was greater than that in the control group at 10 months of age (p < 0.0083). In addition, the control group showed an increase in serum oxidative stress with ageing. The experimental group also showed increased serum oxidative stress, but the value was lower than the control group at 10 months of age (p < 0.0083). Furthermore, low expressions of protein oxidative damage in the periodontal tissue were observed in the experimental group, compared to the control group at 6 months and 10 months. CONCLUSION: These findings indicate that Tomarina® might suppress the effects of ageing on gingival collagen degradation, by decreasing oxidative stress in the rat model.

Effects of apoE deficiency and occlusal disharmony on amyloid-beta production and spatial memory in rats.

Amyloid-ß (Aß) plays a causative role in Alzheimer's disease. Apolipoprotein E (apoE) is involved in Aß accumulation, whereas occlusal disharmony increases Aß production in the rat hippocampus. The purpose of the present study was to investigate the effects of apoE deficiency and occlusal disharmony on Aß production and spatial memory. Wild-type (WT) (n = 12) and apoE-deficient [ApoE(-/-)] (n = 12) rats (Sprague-Dawley; 8 weeks old) were used. These rats were randomly divided into four groups of six rats each: two control (C) groups: WT (C-WT) and ApoE [C-ApoE(-/-)], and two occlusal disharmony (D) groups: WT (D-WT) and ApoE [D-ApoE(-/-)]. The C group received no treatment for 8 weeks. In the D group, the maxillary molar cusps were cut off for 8 weeks. The spatial memory of rats was assessed according to their behavioral performance in a radial arm maze. In both genotypes of rats, significant differences in the reference memory, Aß42 production, ß-secretase expression and plasma corticosterone levels were observed between the C and D groups (P < 0.0125). The levels of Aß42 and glucocorticoid receptor in the C-ApoE(-/-) group were also significantly higher than those in the C-WT group (P < 0.0125). However, no significant differences in these parameters were found between the two genotypes with occlusal disharmony. In conclusion, occlusal disharmony induces cognitive dysfunction and Aß accumulation in the rat hippocampus, and the effects of occlusal disharmony on Aß accumulation and cognitive dysfunction were larger than those of apoE deficiency.

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

BACKGROUND: Dyslipidemia 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. METHODS: Levels 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. RESULTS: Apolipoprotein 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). CONCLUSION: These findings underscore the important role for TLR2 and TLR4 in mediating the osteoclast differentiation on alveolar bone response to dyslipidemia.

Preventive effects of trehalose on osteoclast differentiation in rat periodontitis model.

AIM: Trehalose, which is a disaccharide formed by a 1,1 linkage of two glucose molecules, was suggested to have a suppressive effect on bone resorption. In this study, we examined the effects of topical application of trehalose on osteoclast differentiation in a rat periodontitis model. MATERIAL AND METHODS: Rats were divided into four groups. One group received no treatment. In the other groups, experimental periodontitis was induced by ligature placement. These rats with experimental periodontitis received topical application of pure water (vehicle group), 30 mg/ml trehalose solution (30 mg/ml trehalose group) or 60 mg/ml trehalose solution (60 mg/ml trehalose group) to the gingival sulcus respectively. RESULTS: The vehicle group showed higher numbers of polymorphonuclear leucocytes, receptor activator of nuclear factor kappa B ligand (RANKL)-positive cells and osteoclasts compared with the no treatment group respectively. Trehalose-applied groups exhibited lower numbers of these cells compared with the vehicle group. Gene expressions of tumour necrosis factor-α, RANKL and toll-like receptor 4 were suppressed by trehalose. In addition, protein expressions of RANKL inducing pathway were less activated by trehalose. CONCLUSION: Topical application of trehalose could suppress osteoclast differentiation by inactivation of RANKL inducing pathway in the rat periodontitis model.

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.

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.

Stage of hepatocellular carcinoma is associated with periodontitis.

AIM: Periodontitis induces overproduction of reactive oxygen species (ROS). This state increases circulating ROS levels and may affect hepatocellular carcinoma (HCC). The Japan Integrated Stage (JIS) score is a novel staging system for HCC. The objective of the present study was to compare JIS scores in HCC patients with and without periodontitis. MATERIAL AND METHODS: We recruited 64 HCC patients comprising 31 chronic periodontitis subjects (HCC + P) and 33 periodontally healthy controls (HCC + H). Their JIS scores were recorded. Serum levels of reactive oxygen metabolites (ROM) from HCC + P, HCC + H and healthy age- and gender-matched subjects with healthy gingiva (control, n = 15) were also assessed for circulating ROS levels. RESULTS: The HCC + P and HCC + H groups had similar body mass index, habitual drinking and tobacco exposure data. The HCC + P group showed higher JIS scores than the HCC + H group (p = 0.027). Both the HCC + P and HCC + H groups had higher serum levels of ROM than controls (p < 0.001), while serum levels of ROM in the HCC + P group were a further 25.8% higher than those in the HCC + H group (p < 0.001). CONCLUSION: HCC patients with periodontitis had higher JIS score and circulating ROS level than HCC patients without periodontitis.

Hydrogen-rich water attenuates experimental periodontitis in a rat model.

AIM: Reactive oxygen species (ROS) contribute to the development of periodontitis. As molecular hydrogen can act as a scavenger of ROS, we examined the effects of treatment with hydrogen-rich water on a rat model of periodontitis. MATERIAL & METHODS: A ligature was placed around the maxillary molars for 4 weeks to induce periodontitis, and the animals were given drinking water with or without hydrogen-rich water. RESULTS: The rats with periodontitis which were treated with pure water showed a time-dependent increase in serum ROS level. Compared with the rats without periodontitis, the periodontitis-induced rats which were given pure water also showed polymorphonuclear leucocyte infiltration and alveolar bone loss at 4 weeks. Hydrogen-rich water intake inhibited an increase in serum ROS level and lowered expression of 8-hydroxydeoxyguanosine and nitrotyrosine in the periodontal tissue at 4 weeks. Such conditions prevented polymorphonuclear leucocyte infiltration and osteoclast differentiation following periodontitis progression. Furthermore, inflammatory signalling pathways, such as mitogen-activated protein kinases, were less activated in periodontal lesions from hydrogen-rich water-treated rats as compared with pure water-treated rats. CONCLUSION: Consuming hydrogen-rich water might be beneficial in suppressing periodontitis progression by decreasing gingival oxidative stress.

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

BACKGROUND: The 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. METHODS: A 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. RESULTS: In 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. CONCLUSION: The B/B Checker® is useful for objective evaluation of malodor in oral, exhaled and nasal air and for screening subjects with halitosis. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01139073.

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.

Occlusal disharmony induces BDNF level in rat submandibular gland.

OBJECTIVES: brain-derived neurotrophic factor (BDNF), which is produced in rat submandibular gland, is one of the most abundant neurotrophins in the central nervous system. It is generally accepted that occlusal disharmony causes stress. The purpose of the present study was to investigate whether occlusal disharmony-induced chronic stress affects BDNF levels and morphology in rat submandibular gland. DESIGN: eight wks old male Wistar rats (n=21) were randomly divided into three groups of 7 rats. In a control (C) group, the rats received no treatment for 8 wks. In a molar cusp-less (OD) group, maxillary molar cusps were cut off with a dental turbine at baseline and kept for 8 wks. In a molar cusp-less + recovered cusp (OR) group, maxillary molar cusps were cut off and then were recovered after 4 wks using resin material. After the experimental period, expression of BDNF mRNA and protein as well as histological findings were evaluated in the submandibular glands. The comparisons between the groups were made using the Mann-Whitney U test with Bonferroni correction. RESULTS: the OD group showed a significant increase in submandibular gland BDNF mRNA and protein expression after 8 wks, and plasma adrenocorticotropic hormone and corticosterone levels increased in a time-dependent manner. There were no significant differences in BDNF expression in the submandibular glands and in levels of plasma adrenocorticotropic hormone and corticosterone between the OR and C groups. CONCLUSIONS: these results indicate that psychological stress induced by occlusal disharmony reversibly induces BDNF expression in the rat submandibular gland.

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-alpha (TNF-alpha), 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 beta-polypeptide (Pi3kcb), Pi3kp85, and insulin receptor substrate 1 and 2 were observed in the periodontitis group. Levels of serum CRP and TNF-alpha 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.

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.