Bactericidal activity and recovery effect of hydroxyl radicals generated by ultraviolet irradiation and silver ion application on an infected titanium surface.

This study investigated the bactericidal effect, the underlying mechanisms of treatment, and recovery of biocompatibility of the infected titanium surface using a combination treatment of silver ion application and ultraviolet-A (UV-A) light irradiation. Streptococcus mutans and Aggregatibacter actinomycetemcomitans were used in suspension and as a biofilm on a titanium surface to test for the bactericidal effect. The bactericidal effect of the combination treatment was significantly higher than that of silver ion application or UV-A light irradiation alone. The bactericidal effect of the combination treatment was attributable to hydroxyl radicals, which generated from the bacterial cell wall and whose yield increased with the silver concentration. To assess the biocompatibility, proliferation and calcification of MC3T3E1 cells were evaluated on the treated titanium surface. The treated titanium screws were implanted into rat tibias and the removal torques were measured 28 days post-surgery. The titanium surface that underwent the combination treatment exhibited recovery of biocompatibility by allowing cellular proliferation or calcification at levels observed in the non-infected titanium surfaces. The removal torque 28 days after surgery was also comparable to the control values. This approach is a novel treatment option for peri-implantitis.

Oral mucosal irritation potential of antimicrobial chemotherapy involving hydrogen peroxide photolysis with high-power laser irradiation for the treatment of periodontitis.

In the present study, we assessed the oral mucosal irritation potential of antimicrobial chemotherapy involving hydrogen peroxide (H2O2) photolysis with a 405-nm laser device at an output power of ≥100 mW in hamsters. Twenty-four cheek pouches from 12 male Syrian hamsters received 7-min treatment with pure water (PW), 3% H2O2, laser irradiation of PW at 100 mW, laser irradiation of 3% H2O2 at 100 mW, laser irradiation of PW at 200 mW, or laser irradiation of 3% H2O2 at 200 mW (n = 4 each). The diameter of the irradiation area was set at 3 mm; accordingly, the calculated irradiances (optical power densities) of the 100- and 200-mW laser lights were approximately 1400 and 2800 mW/cm2, respectively. In addition, 12 cheek pouches from six animals received laser irradiation of 3% H2O2 at 100 mW for 1, 3, or 5 min (n = 4 each). Each treatment was repeated three times at 1-h intervals. Macroscopic and histological changes were evaluated 24 h after the last treatment. In addition, in vitro bactericidal activity of the treatment against periodontal pathogens was evaluated. We found that 405-nm laser irradiation of 3% H2O2 caused moderate to severe oral mucosal irritation when performed at powers of 100 and 200 mW for ≥3 min, while the same treatment performed at 100 mW for 1 min resulted in mild irritation. Moreover, 1-min H2O2 photolysis at 100 mW caused a >4-log decrease in viable bacterial counts. These findings suggest that 1-min H2O2 photolysis, which can effectively kill periodontal pathogens, may be acceptable when a 405-nm laser device is used at 100 mW. However, use of the laser at a lower power would be preferable for the prevention of unnecessary oral mucosal irritation.

Potential adverse effects of antimicrobial chemotherapy based on ultraviolet-A irradiation of polyphenols against the oral mucosa in hamsters and wounded skin in rats.

Antimicrobial chemotherapy based on ultraviolet-A (UVA) irradiation of polyphenol solution has been proposed as an adjunctive treatment for dental caries. However, the safety of this treatment has not been thoroughly evaluated. Therefore, the aim of the present study was to assess the influence of this treatment on the oral mucosa in hamsters and wounded skin in rats. An oral mucosal irritation test was performed in hamsters. The cheek pouch was everted and treated with UVA irradiation (wavelength: 365 nm) of pure water, 1 mg/mL of caffeic acid, or 1 mg/mL of grape seed extract using a light-emitting diode at an irradiance of 275 mW/cm2. Each treatment was performed for 2 or 5 min and repeated three times. Macroscopic and histological evaluations were performed 24 h after the last treatment. We also examined the effects of the treatment on the healing process of skin wounds in rats. Full-thickness skin wounds created on the back of the rats were treated for 2 min as described above, but only once. The wound area was then assessed daily for 9 days. The results demonstrated that the treatment induced oral mucosal irritation depending on the irradiation time; however, it did not influence the wound healing process. The oral mucosal irritation potential of three treatment sessions performed for 2 and 5 min was minimal and mild to moderate, respectively, according to histological analysis. These findings suggest that the duration of treatment based on UVA irradiation of polyphenols in the oral cavity should be as short as possible, considering the clinical efficacy of the antimicrobial effects and the irritation potential.

Proanthocyanidin-Rich Grape Seed Extract Modulates Intestinal Microbiota in Ovariectomized Mice.

Grape-seed extract (GSE) is rich in proanthocyanidins (polymers of flavan-3-ols). GSE is well known to have various beneficial effects to health. The objective of this study was to examine the effect of dietary GSE on the intestinal microbiota in ovariectomized (OVX) mice as a model of menopause. Phylum-level analyses using 16S rRNA-targeted group-specific polymerase-chain reaction primers in fecal samples collected 8 weeks postoperatively from OVX mice revealed that the proportion of Firmicutes and Bacteroidetes populations became imbalanced as compared with that in sham-operated control mice. That is, the ratio of Firmicutes:Bacteroidetes populations in the OVX group were increased significantly. When OVX animals were given dietary GSE, the imbalanced proportion of Firmicutes and Bacteroidetes populations was normalized to that seen in control mice. In addition, the body weight of OVX animals measured at 6 weeks postoperatively was significantly higher than that in sham-operated control animals. Dietary GSE also prevented OVX animals from increasing body weight. Thus, we postulated that GSE can improve imbalanced populations of intestinal microbiota, leading to prevention of obesity under conditions of not only menopause but morbidity. PRACTICAL APPLICATION: The GSE has a great potential to be a functional food to improve dysbiosis in post-menopausal women.

Adjunctive antimicrobial chemotherapy based on hydrogen peroxide photolysis for non-surgical treatment of moderate to severe periodontitis: a randomized controlled trial.

Treatment of severe periodontitis with non-surgical therapy remains challenging in dentistry. The present study aimed to evaluate the clinical efficacy of hydrogen peroxide (H2O2) photolysis-based antimicrobial chemotherapy adjunctively performed with root debridement (RD) for moderate to severe periodontitis. A randomized controlled trial was conducted that included 53 patients with 142 test teeth. The test teeth were randomly assigned to one of three treatment groups: Group 1, RD + H2O2 photolysis; Group 2, RD followed by administration of a local drug delivery system (minocycline chloride gel); or Group 3, RD alone. Clinical and microbiological examination were performed for up to 12 weeks following treatment. Probing pocket depth (PPD) and bleeding on probing (BoP) were improved after each treatment session. At 12 weeks, Group 1 had achieved significantly lower PPDs than the other groups, though there were no significant differences in BoP between Group 1 and the other groups. Counts of Porphyromonas gingivalis, a known periodontal pathogen, in Group 1 were significantly lower than those in Group 3, and were comparable to those in Group 2. Therefore, it is suggested that H2O2 photolysis treatment can be used as a novel adjunctive antimicrobial chemotherapy for non-surgical periodontal treatment.

Bactericidal Action of Photodynamic Antimicrobial Chemotherapy (PACT) with Photosensitizers Used as Plaque-Disclosing Agents against Experimental Biofilm.

Our previous study revealed that the photo-irradiation of rose bengal, erythrosine, and phloxine, xanthene photosensitizers, used as dental plaque disclosing agents, could exert bactericidal action on planktonic Streptococcus mutans via the singlet oxygen. In the present study, the photo-irradiation induced bactericidal activity of the three xanthene compounds against the experimental biofilm of S. mutans was investigated in combination with acid electrolyzed water (AcEW) and alkaline electrolyzed water (AlEW). As a result, only the photo-irradiated rose bengal in AlEW showed prominent bactericidal activity with a >3-log reduction of the viable bacterial count. Since our previous study showed that the affinity of rose bengal to bacterial cells was superior to that of erythrosine and phloxine, it was speculated that AlEW damaged the extracellular matrix of the experimental biofilm, which would let the rose bengal easily be bound to the bacterial cells. From these results, it is strongly suggested that rose bengal is a suitable photosensitizer for use as a plaque disclosing agent in photodynamic antimicrobial chemotherapy to treat dental plaque.

Bactericidal Activity and Mechanism of Photoirradiated Polyphenols against Gram-Positive and -Negative Bacteria.

The bactericidal effect of various types of photoirradiated polyphenols against Gram-positive and -negative bacteria was evaluated in relation to the mode of action. Gram-positive bacteria (Enterococcus faecalis, Staphylococcus aureus, and Streptococcus mutans) and Gram-negative bacteria (Aggregatibacter actinomycetemcomitans, Escherichia coli, and Pseudomonas aeruginosa) suspended in a 1 mg/mL polyphenol aqueous solution (caffeic acid, gallic acid, chlorogenic acid, epigallocatechin, epigallocatechin gallate, and proanthocyanidin) were exposed to LED light (wavelength, 400 nm; irradiance, 260 mW/cm(2)) for 5 or 10 min. Caffeic acid and chlorogenic acid exerted the highest bactericidal activity followed by gallic acid and proanthocyanidin against both Gram-positive and -negative bacteria. It was also demonstrated that the disinfection treatment induced oxidative damage of bacterial DNA, which suggests that polyphenols are incorporated into bacterial cells. The present study suggests that blue light irradiation of polyphenols could be a novel disinfection treatment.

Bactericidal action of photogenerated singlet oxygen from photosensitizers used in plaque disclosing agents.

BACKGROUND: Photodynamic therapy (PDT) has been suggested as an efficient clinical approach for the treatment of dental plaque in the field of dental care. In PDT, once the photosensitizer is irradiated with light of a specific wavelength, it transfers the excitation energy to molecular oxygen, which gives rise to singlet oxygen. METHODOLOGY/PRINCIPAL FINDINGS: Since plaque disclosing agents usually contain photosensitizers such as rose bengal, erythrosine, and phloxine, they could be used for PTD upon photoactivation. The aim of the present study is to compare the ability of these three photosensitizers to produce singlet oxygen in relation to their bactericidal activity. The generation rates of singlet oxygen determined by applying an electron spin resonance technique were in the order phloxine > erythrosine ≒ rose bengal. On the other hand, rose bengal showed the highest bactericidal activity against Streptococcus mutans, a major causative pathogen of caries, followed by erythrosine and phloxine, both of which showed activity similar to each other. One of the reasons for the discrepancy between the singlet oxygen generating ability and bactericidal activity was the incorporation efficiency of the photosensitizers into the bacterial cells. The incorporation rate of rose bengal was the highest among the three photosensitizers examined in the present study, likely leading to the highest bactericidal activity. Meanwhile, the addition of L-histidine, a singlet oxygen quencher, cancelled the bactericidal activity of any of the three photoactivated photosensitizers, proving that singlet oxygen was responsible for the bactericidal action. CONCLUSIONS: It is strongly suggested that rose bengal is a suitable photosensitizer for the plaque disclosing agents as compared to the other two photosensitizers, phloxine and erythrosine, when used for PDT.

Reevaluation of analytical methods for photogenerated singlet oxygen.

The aim of the present study is to compare different analytical methods for singlet oxygen and to discuss an appropriate way to evaluate the yield of singlet oxygen photogenerated from photosensitizers. Singlet oxygen photogenerated from rose bengal was evaluated by electron spin resonance analysis using sterically hindered amines, spectrophotometric analysis of 1,3-diphenylisobenzofuran oxidation, and analysis of fluorescent probe (Singlet Oxygen Sensor Green®). All of the analytical methods could evaluate the relative yield of singlet oxygen. The sensitivity of the analytical methods was 1,3-diphenylisobenzofuran < electron spin resonance < Singlet Oxygen Sensor Green®. However, Singlet Oxygen Sensor Green® could be used only when the concentration of rose bengal was very low (<1 µM). In addition, since the absorption spectra of 1,3-diphenylisobenzofuran is considerably changed by irradiation of 405 nm laser, photosensitizers which are excited by light with a wavelength of around 400 nm such as hematoporphyrin cannot be used in the 1,3-diphenylisobenzofuran oxidation method. On the other hand, electron spin resonance analysis using a sterically hindered amine, especially 2,2,6,6-tetramethyl-4-piperidinol and 2,2,5,5-tetramethyl-3-pyrroline-3-carboxamide, had proper sensitivity and wide detectable range for the yield of photogenerated singlet oxygen. Therefore, in photodynamic therapy, it is suggested that the relative yield of singlet oxygen generated by various photosensitizers can be evaluated properly by electron spin resonance analysis.