Potential Use of Brazilian Green Propolis Extracts as New Photosensitizers for Antimicrobial Photodynamic Therapy against Cariogenic Microorganisms.

The synergic effect of Streptococcus mutans and Candida albicans increases dental caries severity. Antimicrobial photodynamic therapy (aPDT) is a non-invasive treatment for antimicrobial aspects. However, the current photosensitizers (PS) have many downsides for dental applications. This study aimed to evaluate the efficiency of two different Brazilian green propolis (BGP-AF and BGP-AG) as PS for aPDT against these microorganisms. A single-species biofilm was irradiated with crude extracts and their fractions and controls. Such extracts showed the best results and were evaluated in dual-species biofilms. Photodegradation, reactive oxygen species (ROS), cytotoxicity, and color stability assays were also investigated. Reductions higher than 3 log10 CFU/mL (p < 0.0001) occurred for crude BGP in single- and dual-species biofilms. Singlet oxygen was produced in BGP (p < 0.0001). BGP-mediated aPDT delayed S. mutans and C. albicans regrowth after 24 h of treatment (p < 0.0001). Both BGP did not change the color of dental materials (p > 0.05). BGP-AF-mediated aPDT showed 72.41% of oral keratinocyte viability (p < 0.0001). BGP extracts may be used in aPDT against S. mutans and C. albicans. Specifically, BGP-AF may represent a promising PS for dental applications.

In Vitro Evaluation of Photodynamic Activity of Plant Extracts from Senna Species against Microorganisms of Medical and Dental Interest.

Background: Bacterial resistance requires new treatments for infections. In this context, antimicrobial photodynamic therapy (aPDT) is an effective and promising option. Objectives: Three plant extracts (Senna splendida, Senna alata, and Senna macranthera) were evaluated as photosensitizers for aPDT. Methods: Cutibacterium acnes (ATCC 6919), Streptococcus mutans (ATCC 35668), Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), and Candida albicans (ATCC 90028) were evaluated. Reactive oxygen species production was also verified. Oral keratinocytes assessed cytotoxicity. LC-DAD-MS analysis identified the chemical components of the evaluated extracts. Results: Most species cultured in the planktonic phase showed total microbial reduction (>6 log10 CFU/mL/p < 0.0001) for all extracts. C. albicans cultured in biofilm showed total microbial reduction (7.68 log10 CFU/mL/p < 0.0001) for aPDT mediated by all extracts. Extracts from S. macranthera and S. alata produced the highest number of reactive oxygen species (p < 0.0001). The S. alata extract had the highest cell viability. The LC-DAD-MS analysis of active extracts showed one naphthopyrone and seven anthraquinones as potential candidates for photoactive compounds. Conclusion: This study showed that aPDT mediated by Senna spp. was efficient in microbial suspension and biofilm of microorganisms of medical and dental interest.

Assessment of synergism between enzyme inhibition of Cu/Zn-SOD and antimicrobial photodynamic therapy in suspension and E. coli biofilm.

BACKGROUND: Antimicrobial Photodynamic Therapy (aPDT) is a treatment based on the interaction between a photosensitizer (PS), oxygen and a light source, resulting in the production of reactive oxygen species (ROS). There are two main types of reactions that can be triggered by this interaction: type I reaction, which can result in the production of hydrogen peroxide, superoxide anion and hydroxyl radical, and type II reaction, which is the Photodynamic Reaction, which results in singlet oxygen production. Antioxidant enzymes (e.g., catalase and superoxide dismutase) are agents that help prevent the damage caused by ROS and, consequently, reduce the effectiveness of aPDT. The aim of this study was to evaluate a possible synergism of the combined inhibition therapy of the enzyme Cu/Zn-Superoxide dismutase (SOD) and the methylene blue- and curcumin-mediated aPDT against Escherichia coli ATCC 25922, in suspension and biofilm. METHODS: Kinetic assay of antimicrobial activity of diethydithiocarbamate (DDC) and Minimum Bactericidal Concentration (MIC) of DDC were performed to evaluate the behavior of the compound on bacterial suspension. Inhibition times of Cu/Zn-SOD, as well as DDC concentration, were evaluated via bacterial susceptibility to combined therapy in suspension and biofilm. RESULTS: DDC did not present MIC at the evaluated concentrations. The inhibition time and Cu/Zn-SOD concentration with the highest bacterial reductions were 30 minutes and 1.2 µg/mL, respectively. Synergism occurred between DDC and MB-mediated aPDT, but not with CUR-mediated aPDT. CONCLUSIONS: The synergism between Cu/Zn-SOD inhibition and aPDT has been confirmed, opening up a new field of study full of possibilities.

Current applications of drug delivery nanosystems associated with antimicrobial photodynamic therapy for oral infections.

The oral cavity is colonized by several species of microorganisms that can cause dental caries, periodontal diseases, candidiasis, endodontic infections, and, among other diseases related to the dental field. Conventional treatment consists of mechanical removal associated with systemic administration of antimicrobials, which can cause various side effects and microbial resistance. In this context, alternative therapies have been developed, including Antimicrobial Photodynamic Therapy (aPDT). For the improvement of therapy, the implementation of nanotechnology is very important to optimize the delivery system of the dyes or photosensitizers on biological targets. Besides, this combination provides a non-invasive treatment, better solubility and bioavailability, delivery to the target site, controlled release and protection against external and physical-chemical factors, low side effects, and, unlikely resistant species. Although, there are numerous researches on aPDT and nanotechnology, few review articles based on the combination of these three aspects: nanosystems, aPDT and oral infections are available. For this reason, this article aims to discuss the advances and advantages of this combination. Therefore, this article was divided into different types of nanosystems (organic and inorganic nanoparticles) associated with aPDT bringing a description of it is definitions, properties, and, applications in oral infections.

Conjugation of superparamagnetic iron oxide nanoparticles and curcumin photosensitizer to assist in photodynamic therapy.

In this work, we describe the synthesis and characterization of the SPIONP-CUR conjugate between curcumin (CUR) and superparamagnetic iron oxide nanoparticles (SPIONPs), in addition to its application in photodynamic therapy (PDT) using a protocol free of organic solvents as a dispersant. The SPIONP-CUR conjugate was characterized by X-ray diffraction, transmission electron microscopy, zeta potential measurements, Fourier transform infrared spectroscopy, thermogravimetry, magnetometry and magnetic hyperthermia assays. The SPIONP-CUR conjugation occurred by bonding between the keto-enol moiety of CUR and the iron atoms present on the surfaces of the SPIONPs. The conjugate showed heating power under an alternating magnetic field (AMF) and photodynamic action when irradiated with blue LED light. In experiments using PDT against Staphylococcus aureus in the planktonic phase, it was demonstrated that with application of blue light at 3.12 J cm-2, the conjugate (dispersed in water) caused a total reduction of the bacterial load. In the absence of light, the reduction was insignificant, even after 24 h of contact with the bacteria.

Influence of light intensity and irradiation mode on methylene blue, chlorin-e6 and curcumin-mediated photodynamic therapy against Enterococcus faecalis.

This study aimed to evaluate the effect of the continuous irradiation with low intensity (continuous mode) and fractioned irradiation with high intensity (fractionated mode), keeping the same dose of light by using Light Emitting Diode (LEDs) with wavelength emission centered at 450 and at 660 nm, using methylene blue (MB), chlorin-e6 (Ce6) and curcumin (CUR) as photosensitizers (PSs) against planktonic phase of E. faecalis. Cell viability was assessed by counting colonies forming per mL (CFU/mL), and the quantification of reactive species was performed by fluorescence with the photodegradation rate evaluated by measurements of absorbance of PSs at different times. The results revealed that MB-mediated PDT was efficient to achieve total microbial load reduction in both irradiation modes, but in fractional mode it was possible to use a lower light dose. Using Ce6, a total bacterial reduction was observed when fractional light was used, but at the same light dose, there was no reduction in the continuous irradiation mode. CUR-mediated PDT under continuous irradiation mode promoted the total microbial load reduction. However, for fractional mode, a higher concentration of CUR was required to completely reduce E. faecalis cell viability. Our results suggest that the biological response to PDT is variable depending on the irradiation mode and on the photosensitizer. Therefore, these studies indicate that the irradiation mode, intensity and the specific PSs should be taken into account for the development of clinical protocols for PDT.

Antimicrobial photodynamic therapy against metronidazole-resistant dental plaque bactéria.

The antimicrobial photodynamic therapy (aPDT) has stood out as an alternative and promising method of disinfection and has been exploited for the treatment of oral bacteria. In this study, we evaluate in vitro the action of aPDT, mediated by methylene blue, chlorin-e6, and curcumin against clinical subgingival plaques that were resistant to metronidazole. The sensitivity profile of the samples to metronidazole was analyzed by the agar dilution method. Cell viability in the planktonic and biofilm phase was assessed by CFU / mL. The composition of the biofilm was evaluated by the checkboard DNA-DNA Hibrydization technique. Photosensitizers internalization was qualitatively assessed by confocal fluorescence microscopy (CLSM). The aPDT mediated by the three photosensitizers tested was able to reduce the totality of the planktonic microbial load and partially reduce the biofilm samples. The analysis performed by CLSM showed that the photosensitizers used in the application of aPDT were able to permeate the interior of the biofilm. The aPDT has been shown to be useful in a supportive and effective approach to the treatment of periodontal disease.

Effect of Photodynamic Therapy on Microorganisms Responsible for Dental Caries: A Systematic Review and Meta-Analysis.

The aim of this study was to perform a systematic review of the literature followed by a meta-analysis about the efficacy of photodynamic therapy (PDT) on the microorganisms responsible for dental caries. The research question and the keywords were constructed according to the PICO strategy. The article search was done in Embase, Lilacs, Scielo, Medline, Scopus, Cochrane Library, Web of Science, Science Direct, and Pubmed databases. Randomized clinical trials and in vitro studies were selected in the review. The study was conducted according the PRISMA guideline for systematic review. A total of 34 articles were included in the qualitative analysis and four articles were divided into two subgroups to perform the meta-analysis. Few studies have achieved an effective microbial reduction in microorganisms associated with the pathogenesis of dental caries. The results highlight that there is no consensus about the study protocols for PDT against cariogenic microorganisms, although the results showed the PDT could be a good alternative for the treatment of dental caries.

Antimicrobial Photodynamic therapy enhanced by the peptide aurein 1.2.

In the past few years, the World Health Organization has been warning that the post-antibiotic era is an increasingly real threat. The rising and disseminated resistance to antibiotics made mandatory the search for new drugs and/or alternative therapies that are able to eliminate resistant microorganisms and impair the development of new forms of resistance. In this context, antimicrobial photodynamic therapy (aPDT) and helical cationic antimicrobial peptides (AMP) are highlighted for the treatment of localized infections. This study aimed to combine the AMP aurein 1.2 to aPDT using Enterococcus faecalis as a model strain. Our results demonstrate that the combination of aPDT with aurein 1.2 proved to be a feasible alternative capable of completely eliminating E. faecalis employing low concentrations of both PS and AMP, in comparison with the individual therapies. Aurein 1.2 is capable of enhancing the aPDT activity whenever mediated by methylene blue or chlorin-e6, but not by curcumin, revealing a PS-dependent mechanism. The combined treatment was also effective against different strains; noteworthy, it completely eliminated a vancomycin-resistant strain of Enterococcus faecium. Our results suggest that this combined protocol must be exploited for clinical applications in localized infections as an alternative to antibiotics.

Susceptibility of Enterococcus faecalis and Propionibacterium acnes to antimicrobial photodynamic therapy.

Bacterial resistance to available antibiotics nowadays is a global threat leading researchers around the world to study new treatment modalities for infections. Antimicrobial photodynamic therapy (aPDT) has been considered an effective and promising therapeutic alternative in this scenario. Briefly, this therapy is based on the activation of a non-toxic photosensitizing agent, known as photosensitizer (PS), by light at a specific wavelength generating cytotoxic singlet oxygen and free radicals. Virtually all studies related to aPDT involve a huge screening to identify ideal PS concentration and light dose combinations, a laborious and time-consuming process that is hardly disclosed in the literature. Herein, we describe an antimicrobial Photodynamic Therapy (aPDT) study against Enterococcus faecalis and Propionibacterium acnes employing methylene blue, chlorin-e6 or curcumin as PS. Similarities and discrepancies between the two bacterial species were pointed out in an attempt to speed up and facilitate futures studies against those clinical relevant strains. Susceptibility tests were performed by the broth microdilution method. Our results demonstrate that aPDT mediated by the three above-mentioned PS was effective in eliminating both gram-positive bacteria, although P. acnes showed remarkably higher susceptibility to aPDT when compared to E. faecalis. PS uptake assays revealed that P. acnes is 80 times more efficient than E. faecalis in internalizing all three PS molecules. Our results evidence that the cell wall structure is not a limiting feature when predicting bacterial susceptibility to aPDT treatment.

Synergistic antimicrobial effect of photodynamic therapy and ciprofloxacin.

The occurrence of a variety of pathogens resistant to current antibiotics remains the major problem in medical care, especially when bacterial infections are established as biofilms. In this study, we propose the use of photodynamic therapy (PDT) as a monotherapy and associated with antibiotic as an alternative treatment. The aim of this study was to analyze the effects of PDT mediated by methylene blue (MB) on Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922) in both biofilm and planktonic phases. Several concentrations of MB and light doses were tested. The bactericidal effects of PDT as a monotherapy did not increase with the concentration of photosensitizer, but were light dose-dependent. In addition, bacteria in biofilms were less affected than cells in the planktonic phase. Although not concentration-dependent, the disruption effect of PDT on biofilms was clearly illustrated by scanning electron microscopy (SEM). We also carried out experiments that evaluated the synergistic effect of photodynamic therapy and the antibiotic ciprofloxacin. The best results were obtained after combination treatment of photodynamic therapy followed by ciprofloxacin on biofilms, which increased bacterial reduction on biofilms, resulting in a 5.4 log reduction for S. aureus biofilm and approximately 7 log for E. coli biofilm.

Effects of photodynamic therapy with blue light and curcumin as mouth rinse for oral disinfection: a randomized controlled trial.

OBJECTIVE: The purpose of this study was to evaluate the effects of the antimicrobial photodynamic therapy (a-PDT) with blue light and curcumin on oral disinfection during the 2 h after treatment. BACKGROUND DATA: a-PDT is a technique that can potentially affect the viability of bacterial cells, with selective action targeting only areas with photosensitizer accumulation. MATERIALS AND METHODS: A randomized controlled trial was undertaken. Twenty-seven adults were randomly divided into three groups: (1) the PDT group, which was treated with the drug, curcumin, and blue light (n=9); (2) the light group, which was treated only with the blue light, and no drug (n=9) and; (3) the curcumin group, which was treated only with the drug, curcumin, and no light (n=9). The irradiation parameters were: blue light-emitting diode (LED) illumination (455±30 nm), 400 mW of average optical power, 5 min of application, illumination area of 0.6 cm(2), 600 mW/cm(2) of intensity, and 200 J/cm(2) of fluence. A curcumin concentration of 30 mg/L was used. The saliva samples were collected for bacterial counts at baseline and after the experimental phases (immediately after treatment, and 1 and 2 h after treatment). Serial dilutions were performed, and the resulting samples were cultured on blood agar plates in microaerophilic conditions. The number of colony-forming units (CFU) was determined. RESULTS: The PDT group showed a significant reduction of CFU immediately after treatment (post-treatment) with PDT (5.71±0.48, p=0.001), and 1 h (5.14±0.92, p=0.001) and 2 h (5.35±0.76, p=0.001) after treatment, compared with pretreatment (6.61±0.82). There were no significant changes for the light group. The curcumin group showed a significant increase of CFU 1 h after treatment (6.77±0.40, p=0.02) compared with pretreatment (5.57±0.91) falling to baseline values at 2 h after treatment (5.58±0.70). CONCLUSIONS: The PDT group showed significant difference in microbial reduction (p<0.05) compared with both the light and curcumin groups until 2 h post-treatment. The new blue LED device for PDT using curcumin may be used for reduction of salivary microorganisms, leading to overall disinfection of the mouth (e.g., mucosa, tongue, and saliva), but new protocols should be explored.

A 12-month follow-up of hypopigmentation after laser hair removal.

INTRODUCTION: Laser hair removal is becoming an increasingly popular alternative to traditional methods such as shaving, waxing, among other methods. Semiconductor diode lasers are considered the most efficient light sources available and are especially well suited for clinical applications including hair reduction. The effectiveness of laser hair reduction depends on many variables, including the skin type of the patient. MATERIAL AND METHODS: A patient with Fitzpatrick Skin Type IV was submitted to laser hair removal of the arms with a high-power diode laser system with long pulses with a wavelength of 800 nm, a fluence of 40 J/cm(2) and a pulse width of 20 ms. A 12-month follow-up assessment was performed and included photography and questionnaire. RESULTS: Hypopigmentation was observed after a single laser hair removal section. After 6 months with the area totally covered, a gradual suntan with a sun screen lotion with an SPF of 15 was prescribed by the dermatologist. After 12 months of the initial treatment, a complete recovery of the hypopigmentation was achieved. CONCLUSION: Although a safe procedure, lasers for hair removal may be associated with adverse side effects including undesired pigment alterations. Before starting a laser hair removal treatment, patients seeking the eradication of hair should be informed that temporary, and possibly permanent, pigmentary changes may occur.

Low-level laser therapy in pediatric Bell's palsy: case report in a three-year-old child.

OBJECTIVES: The objective of this study was to apply low-level laser therapy (LLLT) to accelerate the recovery process of a child patient with Bell's palsy (BP). DESIGN: This was a prospective study. SUBJECT: The subject was a three-year-old boy with a sudden onset of facial asymmetry due to an unknown cause. MATERIALS AND METHODS: The low-level laser source used was a gallium aluminum arsenide semiconductor diode laser device (660 nm and 780 nm). No steroids or other medications were given to the child. The laser beam with a 0.04-cm(2) spot area, and an aperture with approximately 1-mm diameter, was applied in a continuous emission mode in direct contact with the facial area. The duration of a laser session was between 15 and 30 minutes, depending on the chosen points and the area being treated. Light was applied 10 seconds per point on a maximum number of 80 points, when the entire affected (right) side of the face was irradiated, based on the small laser beam spot size. According to the acupuncture literature, this treatment could also be carried out using 10-20 Chinese acupuncture points, located unilaterally on the face. In this case study, more points were used because the entire affected side of the face (a large area) was irradiated instead of using acupuncture points. OUTCOME MEASURES: The House-Brackmann grading system was used to monitor the evolution of facial nerve motor function. Photographs were taken after every session, always using the same camera and the same magnitude. The three-year-old boy recovered completely from BP after 11 sessions of LLLT. There were 4 sessions a week for the first 2 weeks, and the total treatment time was 3 weeks. RESULTS: The result of this study was the improvement of facial movement and facial symmetry, with complete reestablishment to normality. CONCLUSIONS: LLLT may be an alternative to speed up facial normality in pediatric BP.

Overall-mouth disinfection by photodynamic therapy using curcumin.

BACKGROUND DATA: Photodynamic therapy is a technique that involves the activation of photosensitizers by light in the presence of tissue oxygen, resulting in the production of reactive radicals capable of inducing cell death. OBJECTIVE: The present study assessed the overall susceptibility of pathogens of salivary flora to photodynamic therapy after sensitization with curcumin and exposure to blue light at 450 nm. METHODS: A randomized trial was executed with 13 adult volunteers. Three different groups were analyzed: L-D- (no light, no drug; control group), L-D+ (treated only with the drug; curcumin group) and L+D+ (treated with drug and light; photodynamic therapy group). Non-stimulated saliva samples were collected for bacterial counts at baseline and after the experimental phase, and adverse events experienced were recorded. Serial dilutions were performed, and the resulting samples were cultured on blood agar plates in microaerophilic conditions. The number of colony-forming units was then determined. RESULTS: There was a considerable difference between the two experimental groups with regard to effectiveness of bacterial reduction. In the L-D+ group, the bacterial decline was considerably smaller (9%) than in the L+D+ group, with a 68% decrease in bacteria. A statistically significant reduction in the bacterial population was observed only in the photodynamic therapy group (p<0.05). CONCLUSIONS: Photodynamic therapy was found to be effective in the reduction of salivary microorganisms. No significant reduction was found for the group in which only curcumin was used, proving the absence of dark toxicity of the drug. This work shows that overall disinfection of the mouth can be done with a simple procedure involving photodynamic action.

Temperature variation at soft periodontal and rat bone tissues during a medium-power diode laser exposure.

OBJECTIVE: The aim of this study was to evaluate temperature variation induced by a diode laser in periodontal repair. BACKGROUND DATA: A diode laser operating with medium power can constitute an alternative device for bacterial reduction at periodontal pockets. This procedure will probably avoid any kind of undesirable thermal damage to the irradiated tissues. METHODS: The temperature variation induced by a 810-nm diode laser was investigated in an in vitro study, varying the soft tissue thickness, and in an in vivo study for soft periodontal and bone tissues. The laser powers used were 600 mW, 800 mW, 1.0 W, and 1.2 W, and the light was delivered by a 300-microm fiber. RESULTS: The laser parameters and irradiation time used did not induce a temperature variation high enough to cause thermal irreversible damage to the periodontal tissues investigated. CONCLUSIONS: This study contributes to the establishment of thermally safe working parameters of a diode medium power laser.