Methylene Blue-mediated Photodynamic Therapy Research Articles

Hypericin Mediated Photodynamic Therapy (HY-PDT) versus Methylene Blue Mediated Photodynamic Therapy (MB-PDT) in Treatment of Common Warts: A Comparative Study

Abstract Background Many therapeutic modalities are available for the treatment of warts, but no single therapy is universally effective. Warts can resolve, recur, shrink, or grow despite therapy. Aim of the Study to compare between the efficacy and safety of topical hypericin (HY)-mediated photodynamic therapy (PDT) and methylene blue (MB)-mediated PDT in the treatment of common warts. Patients and Methods The study included 40 patients subdivided into two groups (20 patients each). Group one (MB group) patients were treated with topical MB/IPL PDT. Group two (Hypericin group) patients were treated with topical HY/IPL PDT. Patients were assessed using digital photography and dermoscopic examinations. Results MB-mediated PDT group showed that there was significant difference regarding the decrease of size of warts before versus after MB-mediated PDT. HY- mediated PDT group showed that there was a significant difference regarding the decrease of size and number of warts before versus after HY-mediated PDT. There was better response with HY-mediated PDT compared to MB-mediated PDT without statistical significance. Conclusion Both HY and MB-mediated PDT are effective in clearing common warts. Hypericin is a promising photosensitizer alternative that has a potential efficacy for clearing wart by PDT.

Evaluation of cytotoxicity impacts of combined methylene blue-mediated photodynamic therapy and intracanal antibiotic medicaments on dental stem cells

Root canal therapy is a predominant method for treatment of dental pulp and periapical diseases. Conventional methods such as mechanical instrumentations, chemical irrigation and intracanal medicaments pose a huge limitation to root canal disinfection as they kill bacteria and dental stem cells simultaneously. Therefore, much attention has been focused on finding more efficacious antibacterial methods that has no or negligible cytotoxicity for dental stem cells. Herein, we hypothesized that combining antibacterial medicaments with Antimicrobial photodynamic therapy (aPDT) and methylene blue (MB) as a photosensitizer would be effective in reducing death of dental pulp stem cells (DPSCs). To examine this, DPSCs were isolated from third molar teeth through enzymatic digestion. Isolated cells were cultured in αMEM and when reached adequate confluency, were used for further analysis. Cytotoxicity effect of different groups of MB, DAP, MB, LED and their combination on DPSCs was analyzed using MTT assay. DPSCs membrane integrity as a marker of live cells was assessed through measuring lipid peroxidation and lactate dehydrogenase (LDH) release into extracellular space. Results showed that the combination of LED, MB and TAP or aPDT, MB and DAP was more effective in reducing DPSCs death rate compared to TAP and DAP administration alone. Moreover, Malondialdehyde (MDA) and LDH levels were found to be decreased in cells exposed to combination treatment in comparison with single TAP or DAP therapy. Our study shows the promising perspectives of employing combined aPDT, MB and antibiotic medicaments for reduction of dental stem cell death.

Conditioning of PEEK Implant Abutment Surfaces Using Photodynamic Therapy, Nd:YAG Laser, and Conventional Methods to Evaluate Shear Bond Strength.

Objective: This study was done to evaluate shear bond strength (SBS) and modes of failures after different surface treatment modalities on polyetheretherketone (PEEK) implant abutments. Materials and methods: Seventy-five PEEK implant abutment specimens were randomly distributed into five groups based on surface treatment methods: Group I: No treatment, Group II: methylene blue-mediated photodynamic therapy (MB-PDT), Group III: neodymium-doped yttrium aluminum garnet (Nd:YAG) laser, Group IV: Sulfuric acid (H2SO4), and Group V: Sandblasting (Sb). The measurements for SBS and failure modes for PEEK implant abutment specimens were evaluated via Universal Testing Machine and Stereomicroscope, respectively. Two-way analysis of variance and Tukey's post-hoc test (p > 0.05) were used for the statistical analysis. Moreover, the Shapiro-Wilk normality test was also performed for normality. Results: The SBS mean values and standard deviations in megapascals (MPa) for PEEK abutment specimens with different surface pretreatment groups are shown in Table 1 and Fig. 1. The control group had the lowest SBS (9.67 ± 2.1 MPa), while the highest SBS was observed in Group II (MB-PDT; 17.21 ± 1.32 MPa). SBS values for Group IV (H2SO4; 15.83 ± 0.63 MPa) and Group III (Nd:YAG laser; 16.91 ± 2.10 MPa) were similar to Group II (MB-PDT; p > 0.05). The SBS values for the sandblasted specimens (13.90 ± 1.87 MPa) were not significantly different from the control group (p > 0.05). Conclusions: The PEEK implant abutment surfaces treated with MB-PDT and Nd:YAG laser expressed significantly improved SBS.

Evaluation of Daylight Methylene Blue Photodynamic Therapy in Multiple Recurrent Molluscum Contagiosum

Abstract Background Molluscum contagiosum is a common viral infection caused by DNA Poxvirus. Daylight-photodynamic therapy (DL-PDT) is defined as a photochemical reaction used to selectively destroy tissue. Objectives To evaluate the efficacy and safety of daylight methylene blue photodynamic therapy in treatment of multiple recurrent molluscum contagiosum. Subject and methods This is a single blinded controlled prospective interventional clinical study including 20 patients with molluscum contagiosum diagnosed clinically and by dermoscopy. Patients subjected to daylight methylene blue-mediated photodynamic therapy. Topical MB liposomal loaded 10% cream was applied over the lesions area with occlusion and left for 15 minutes. Two leisons were left as control in each studied patient. Patients were instructed to stay outdoors exposing their lesions and the adjacent area to daylight continuously for at least 90 minutes (if it was a sunny day) or 120 minutes (if it was an overcast or partially sunny day) between 11:00 am to 02:00 pm. Treatment sessions were done weekly for 10 weeks or till complete clearance. Follow up of the patients for 6 months after end of study to detect recurrence or appearance of new lesions. Results Daylight MB Mediated PDT response in a total of 20 (100%) studied patients showed Excellent response to treatment shown as complete clearance in 65%( no.=13) of patients, very good response to treatment in 15.0%( no.=3), good response to treatment in 10.0%( no.=2) and poor response to treatment in 10.0%( no.=2). The total number of lesions was 313. Response of the lesions in studied patents to MB-PDT was monitored by dermoscopic follow up of lesions. Lesions showed complete response in 66.5% (no. = 208) of lesions, partial response in 25.2% (no. = 79) of lesions and no response in 8.3% (no. = 26) of lesions. No serious side effects and also MB-DL/PDT is a painless treatment. No relapse was detected at the site of cleared MC or at a near site during the follow-up period of 6 months following the procedure. Conclusion Daylight-PDT using MB is effective in clearing MC lesions, also is safe, easy to carry out, economic, acceptable cosmetic results with no recurrence, convenient especially for children as it is a painless treatment.

The effect of photodynamic therapy on the salivary flow rate, IgA concentration and C-reactive protein levels in active smokers: a case-control study.

This study aimed to evaluate the effect of photodynamic therapy (PDT) on the salivary flow rate, secretory immunoglobulin A, and C-reactive protein levels in active smokers. The present study is a prospective case-control study. Twenty active smokers were allocated to two groups randomly of ten participants each: the experimental group was irradiated while the control was exposed to sham irradiation by turning off the equipment. In the experimental group, methylene blue mediated PDT was applied both intra- and extra-orally over the major and minor salivary glands using a diode laser. 780 nm wavelength and 4 J/cm2 of energy were used to irradiate the 10 points of major salivary glands (6 for parotid and 2 for submandibular glands and 2 for sublingual glands). On the other hand, 660 nm was used to apply 10 J/cm2 of energy over the minor salivary glands at numerous points. The samples of the stimulated and unstimulated saliva were collected from both groups to assess the SFR. ELISA method was used to assess the level of salivary IgA levels, statistical analysis was done using a one-way ANOVA, and a p-value of <0.05 was considered significant. The results showed a significant increment in salivary and secretory immunoglobulin A levels of subjects undergone photodynamic therapy. C-reactive protein levels were significantly decreased in subjects exposed to irradiation. The present study concludes that photodynamic therapy significantly improves the salivary flow rate, secretory Immunoglobulin A, and oral health quality of life in smokers. The inflammatory salivary marker C-reactive protein, which is usually raised in smokers, is also reduced.

Efficacy of indocyanine green and methylene blue mediated-photodynamic therapy on peri-implant outcomes among diabetics with peri-implant mucositis

BackgroundThis study aimed to assess the efficacy of indocyanine green (ICG)-mediated versus methylene blue (MB)-mediated photodynamic therapy (PDT) as an adjunct to conventional mechanical debridement (MD) on the peri‑implant clinical, radiographic, microbiological, and immunological outcomes among diabetics with peri‑implant mucositis (pi-M). MethodsFor this 3-month follow-up study, diabetics having pi-M were randomly divided into 3 groups: group-I (n = 20) subjects received only MD; group-II (n = 20) participants received ICG-mediated adjunct PDT; and group-III (n = 20) subjects received MB-mediated adjunct PDT. Peri-implant clinical (i.e., plaque index [PI], bleeding on probing [BOP], probing depth [PD]), radiographic (crestal bone loss [CBL]), microbiological (Fusobacterium nucleatum [F. nucleatum], Tannerella forsythia [T. forsythia], Prevotella intermedia [P. intermedia], Porphyromonas gingivalis [P. gingivalis], Aggregatibacter actinomycetemcomitans [A. actinomycetemcomitans]), and immunological (interleukin [IL]-6, IL-1β, tumor necrosis factor-alpha [TNF-α]) outcomes were assessed at baseline and 3-month follow-up. ResultsMean changes between baseline and 3-month follow-up in peri‑implant clinico-radiographic parameters were significantly different between control (PI: 12.42±21.80%; BOP: 12.10±19.30%; PD: 0.45±0.41 mm; CBL: 1.10±1.02 mm) and test groups (ICG-mediated PDT [PI: 26.55±25.80%; BOP: 28.77±29.24%; PD: 0.84±0.62 mm; CBL: 1.98±1.85 mm] and MB-mediated PDT [PI: 27.24±26.15%; BOP: 27.71±28.16%; PD: 0.85±0.63 mm; CBL: 1.95±1.80 mm]), however comparable differences were observed in peri‑implant PI, BOP, PD, and CBL between group-II and group-III participants (p>0.05). The proportions of T. forsythia were significantly reduced in group-II (4.78 × 104 colony-forming unit per milliliter [CFU/mL]) and group-III (4.76 × 104 CFU/mL) as compared to group-I (-4.40 × 103 CFU/mL) at 3-month follow-up (p = 0.02). No statistically significant differences were observed between the study groups regarding the proportions of the other assessed target bacterial species. For IL-6 (group-I: 210±108; group-II: 298±165; group-III: 277±121 pg/mL; p = 0.03), IL-1β (group-I: 101±95; group-II: 84±98; group-III: 86±74 pg/mL; p = 0.02), and TNF-α (group-I: 336±121; group-II: 385±210; group-III: 366±198 pg/mL; p = 0.03) peri‑implant sulcular fluid [PISF] levels, all three study groups demonstrated statistically significant reduction at 3-month follow-up. ConclusionsICG-mediated and MB-mediated adjunctive PDT showed statistically significant improvements in peri‑implant clinical, radiographic, microbiological, and immunological parameters as compared to conventional MD alone at 3-month follow-up among diabetics with pi-M. However, comparable outcomes were demonstrated by ICG-mediated and MB-mediated adjunctive PDT regarding the assessed peri‑implant parameters.

Methylene blue-mediated photodynamic therapy for the treatment of oral leukoplakia

Methylene blue-mediated photodynamic therapy for the treatment of oral leukoplakia

Methylene blue-mediated photodynamic therapy in the treatment of oral microbiota. A Systematic Review

Objective: The objective of the present review was to relate the scientific production presented in the last 5 years on the effect of photodynamic therapy using the methylene blue photosensitizer on the microorganisms present in the oral biofilm. Methods: systematic review looking for research in English of primary studies such as randomized clinical trials and without randomization, individual case-control, and qualitative descriptive studies, using the PubMed/MEDLINE and Scopus databases as references, from 2017 to 2021. Results: 15 articles were analyzed, being 67% national publications and 33% international studies. Considering the samples described in the studies, 73% were in vitro, 7% were in vivo and 20% were clinical samples. As for the light source, 66% used laser, 20% used LED and 14% the association of laser and LED devices. The power density used ranged from 0.0026 W/cm2 to 1415 W/cm2. The energy density ranged from 3 J/cm2 to 640 J/cm2. The power ranged from 10 mW to 110 mW. The irradiation time ranged from 24 to 294 seconds. In most publications, 87% used the wavelength of 660 nm. As for the photosensitizer, the most used concentration, in 33% of the studies, was 0.01%, with the pre-irradiation time most used being 300 seconds in 40% of the studies. Conclusion: Photodynamic Methylene Blue Therapy is being accepted as effective in therapies of various oral curative conditions and has been shown to be a useful and effective therapy, as well as a complementary approach to control the development/growth of oral biofilm in clinical situations.

Enhancing effect of chitosan on methylene blue-mediated photodynamic therapy against C. albicans: A study in planktonic growth, biofilms, and persister cells

Chitosan (CS) is a natural polymer extracted from the exoskeleton of crustaceans. Due to its cationic structure, CS has been studied as a possible enhancer of antimicrobial photodynamic therapy (aPDT). The objective was to evaluate the association of CS with methylene blue (MB)-mediated aPDT on Candida albicans, investigating its effects on planktonic growth, biofilms, and cells persistent to fluconazole. The ability of CS to interfere with MB absorption by Candida cells was also evaluated. For the assays, planktonic cells of C. albicans were cultivated for 24 h, and the biofilms were formed for 48 h. For the induction of persister cells, C. albicans was cultivated with high concentration of fluconazole for 48 h. Treatments were performed with MB, CS or MB+CS, followed by irradiation with LED (660 nm ). As results, aPDT with MB (300 µm) reduced the planktonic cells by 1.6 log10 CFU, while the MB+CS association led to a reduction of 4.8 log10 CFU. For aPDT in biofilms, there was a microbial reduction of 2.9 log10 CFU for the treatment with MB (600 µm) and 5.3 log10 CFU for MB+CS. In relation to persister cells, the fungal reductions were 0.4 log10 CFU for MB and 1.5 log10 CFU for MB+CS. In the absorption assays, the penetration of MB into Candida cells was increased in the presence of CS. It was concluded that CS enhanced the antimicrobial activity of aPDT in planktonic growth, biofilms, and persister cells of C. albicans, probably by facilitating the penetration of MB into fungal cells.

Depth distributions of bacteria for the Pseudomonas aeruginosa-infected burn wounds treated by methylene blue-mediated photodynamic therapy in rats: effects of additives to photosensitizer.

.SignificancePseudomonas (P.) aeruginosa, a common cause of infection in burns, acquires antibiotic resistance easily and forms biofilms efficiently. Thus, it is difficult to control P. aeruginosa infection in burn wounds, which causes lethal septicemia. Antimicrobial photodynamic therapy (aPDT) is attractive as a new strategy to treat burn wound infections with drug-resistant bacteria.AimWe examined the efficacy of methylene blue (MB)-mediated aPDT with various additives in a tissue depth-resolved manner to find conditions that minimize the bacterial invasion.ApproachWe applied MB-mediated aPDT with LED array illumination to an extensive, full-thickness burn infected with P. aeruginosa in rats for three consecutive days (days 0, 1, and 2). On day 2, the depth distributions of bacteria were assessed based on the histological analysis using Gram staining. We examined how the addition of ethylenediaminetetraacetic acid (EDTA), ethanol, and dimethyl sulfoxide (DMSO) affected the efficacy of aPDT.ResultsPure MB-mediated aPDT significantly reduced the numbers of bacteria with biofilms on the wound surface and in the epidermis compared with those for the control tissue (saline only). However, there were many bacteria in the deeper region of the tissue. In contrast, MB/EDTA/ethanol/DMSO-mediated aPDT minimized the numbers of bacteria in the broad depth region of the tissue. Still, a limited number of bacteria was observed in the subcutaneous tissue.ConclusionsThe depthwise analysis of bacteria demonstrated the efficacy of the MB-mediated aPDT with the addition of EDTA, ethanol, and DMSO in controlling burn wound infections. However, further improvement of the therapy is needed to suppress bacterial migration into the deep tissue completely.

Methylene blue-mediated Photodynamic Therapy in human retinoblastoma cell lines

Retinoblastoma is a malignant tumor of the retinal precursor cells and one of the rarest types of pediatric tumor, often occurring in the earliest years of life. Symptoms are conditioned by tumor size and location; one of the most recurrent symptoms is a white reflex in the pupillary area, called leukocoria or cat's eye reflex. In the present work, we studied the in vitro effectiveness of Photodynamic treatment (Pdt) in two types of human retinoblastoma, Y79 and WERI-Rb cell lines, using methylene blue (MB), a photosensitizer (PS) from the phenothiazine group. The two cell lines were incubated with varying concentrations of MB (3, 7, 10, 15, 20, 25, 30, 40, and 50μM), in the absence of light (dark cytotoxicity) and, in the presence of 664nm laser light (phototoxicity) with fluences of 1, 1.5, 3, 5, 7, 10, and 15J/cm2. The Y79 cell line showed higher cellular uptake values for MB than the WERI-Rb cell line. After three hours of incubation, the Y79 and WERI-Rb took up 48% and 34% of the total photosensitizer present in the medium, respectively. Using MTT assay, the results showed that the Y79 cell line was more affected by the photo treatment as demonstrated by the combination of MB concentration and light doses compared with WERI-Rb cell line. The results were correlated with the more pronounced singlet oxygen emission observed in Y79 cells. While MB does show efficacy for eradication of retinoblastoma in vitro, only studies in appropriate animal models will reveal whether the selectivity of photokilling at tolerable drug and light doses is sufficient to suggest clinical trials.

The effectiveness of topical forms of dexamethasone in the treatment of oral lichen planus- A systematic review.

The goal of this systematic review was to assess the efficacy of dexamethasone compared to other treatments in oral lichen planus (OLP). The literature search used the following inclusion criteria: randomized controlled trials (RCT) comparing dexamethasone and other treatment strategies in patients with OLP. The outcome measures included relief of symptoms, decrement of erosive area size, and changes in quality of life. A computer and manual search was performed in Pubmed, Web of Science, and Cochrane Library up to January 31, 2021. The risk of bias was measured with the Revised Cochrane risk-of-bias tool for randomized trials. Eight trials with 131 study participants and 132 controls were identified. The following interventions were compared dexamethasone mouthwash, and 5% methylene blue-mediated photodynamic therapy, low-level laser therapy, amlexanox, clobetasol mouthwash, ketoconazole with amitriptyline, and thalidomide 1% paste. The therapeutic outcomes were more advantageous for dexamethasone in comparison with photodynamic therapy (PDT) (2 RCT) and low-level laser therapy (LLLT). Comparable effects were observed for dexamethasone, amlexanox, thalidomide, and PDT (1 RCT). Clobetasol showed more effective action than dexamethasone. Given the small sample sizes, heterogeneity and the few studies included, there is limited evidence to support the selection of treatment for OLP.

Methylene blue-mediated photodynamic therapy may be superior to 5% amorolfine nail lacquer for non-dermatophyte onychomycosis.

Methylene blue-mediated photodynamic therapy as an antimicrobial has been reported to improve onychomycosis. To compare the short-term efficacy of methylene blue-mediated photodynamic therapy (MB-PDT) and 5% amorolfine nail lacquer (AMO) for toenail onychomycosis using higher intensity and shorter total treatment period than previously reported. Twenty-seven toenails with onychomycosis were randomized to receive either six biweekly sessions of MB-PDT or AMO for twelve weeks. Dermoscopic photography was used for onychomycosis severity index assessment under a dermoscopic inspection (d-OSI) at baseline, weeks 6, 10, 14 and 22 as well as microscopic and microbiological tests. Adverse events were recorded. All subjects completed the study. Causative organisms found were exclusively non-dermatophytes including Fusarium spp., Asperillus spp.,and yeasts. Fifteen toenails received MB-PDT, whilst 12 received AMO. D-OSI showed greater improvement in MB-PDT than in AMO groups at weeks 6, 10, 14 as well as 22, with median changes of -2, -3, -4 (P=.055). and-3 respectively in the MB-PDT group. The AMO group displayed the median d-OSI change of 0 throughout the study period. Mycological cure rate at 22weeks in MB-PDT and AMO group was 73.3% and 66.67% (P>.05). Clinical cure rate at 22weeks in MB-PDT (26.7%) was higher than AMO (16.7%), (P>.05). All patients only felt comfortably warm during the MB-PDT treatment. No major adverse events were found in both groups. MB-PDT appeared to be more efficacious for non-dermatophyte onychomycosis than AMO particularly in a limited period and moderately severe onychomycosis.

Different susceptibility of spores and hyphae of Trichophyton rubrum to methylene blue mediated photodynamic treatment in vitro.

In recent years, methylene blue mediated-photodynamic therapy (MB-PDT) has proved to be an effective inhibitor to a variety of microorganisms, including Trichophyton rubrum, the most common dermatophyte worldwide. However, previous studies mainly focused on the spore form of Trubrum, but rarely on its hyphal form, although the latter is the main pathogenic form of Trubrum in vivo. To investigate the inhibitory effect of MB-PDT on Trubrum in different growth phases in vitro. The suspensions of spores and hyphae obtained from Trubrum (ATCC28188) were prepared, respectively, incubated with MB solution (0.15-40μg/mL) and irradiated with 635nm red light. Varied light energy and MB concentration were used. The specimen in the absence of light exposure or/and MB served as controls. MIC determination, colony counts and MTT assay were employed to evaluate the antifungal effect of MB-PDT. The MICs of MB-PDT for hyphae and spores of T. rubrum were 6.300±1.072μg/mL and 1.984±1.072μg/mL, respectively, at a fixed light dose of 60J/cm2 . CFU counts gave the minimum critical combinations of MB concentration and light dose to achieve 100% inhibitory rate. For hyphae, they were 5μg/mL+100J/cm2 or 10μg/mL+60J/cm2 . For spores, they were 1.25μg/mL+40J/cm2 or 5μg/mL+20J/cm2 . The outcomes of MTT assay were consistent with those of CFU counts, but less accurate. MB-PDT is a potent inhibitor to both spores and hyphae of T. rubrum in vitro, and the spores are more sensitive to it. Its antifungal efficacy is positively correlated with the concentration of MB and light dose.

Evaluation of Topical Photo-Enhancer Assisted Intense Pulsed Light versus Methylene Blue Assisted Intense Pulsed Light for Treatment of Seborrheic Keratosis

Abstract Background seborrheic keratosis is one of the most common benign epidermal tumors that affects both sexes equally, and usually arises in individuals older than 50 years. It presents as sharply demarcated, slightly raised brownish patches or plaques, usually on sun-exposed surfaces of the skin. The clinical presentation can be quite variable and includes clinical variants, such as stucco keratosis and dermatosis papulosa nigra. Aim of the Work to evaluate the efficacy and safety of topical carbon gel photo-enhancer assisted intense pulsed light versus methylene blue assisted intense pulsed light photodynamic therapy for treatment of Seborrheic Keratosis. Patients and Methods this is a comparative prospective study that included thirty patients of skin types III to V Fitzpatrick class having multiple Seborrheic Keratosis lesions with no sex or age predilection. They were randomly recruited from Dermatology out-patient clinic of Ain –Shams University Hospitals, during period from June 2017 till March 2018. The study was approved by Research Ethical Committee of Ain Shams University and fulfilled all the ethical aspects required for human research. Results this denoted that the methylene blue-mediated photodynamic therapy (MB-PDT) (liposomal-loaded methylene blue gel 10%).and topical carbon gel photo-enhancer assisted intense pulsed light). could be effective in treating patients with Sks. Our findings denote the efficacy of them in curing multiple Sk lesions. The efficacy of methylene blue-mediated photodynamic therapy (MB-PDT) (liposomal-loaded methylene blue gel 10%). Conclusion that Intense Light Pulse using liposomal-loaded MB and Topical carbon suspension-assisted IPL treatment could be a good option for patients with indistinct pigmented or thick melanogenesic lesions. Adverse reactions to this treatment were minimal and the results acceptable, though appropriate lesions need to be chosen care- fully.

Cytotoxic effect and apoptosis pathways activated by methylene blue-mediated photodynamic therapy in fibroblasts.

Antimicrobial photodynamic therapy (aPDT) has been used as an adjuvant treatment of oral infections as a minimal intervention clinical approach. Its antimicrobial efficacy was demonstrated in several studies; however, there is a lack of evidence on its cytotoxic effect on mouse fibroblasts (NIH/3T3). The aim of this study was to evaluate the cytotoxicity and apoptotic pathways of methylene blue-mediated aPDT on mouse fibroblasts. Cells were treated with 0.1 or 1.0 mg.L-1 methylene blue (MB), and 0.075 or 7.5 J.cm-² LED at 630 nm. Cell viability was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and crystal violet (CV) assays, while cDNA expression for Bax, Bad, Bcl-2, VDAC-1, cytochrome C and Fas-L was assessed by qRT-PCR (1, 3, 6 and 24 h). The differences between groups were detected by Kruskal-Wallis and post-hoc Dunn's tests for MTT and CV assays, and by ANOVA and post-hoc Tukey test for qPCR (P < 0.05). The combination of 1.0 mg.L-1 MB and 7.5 J.cm-² LED significantly reduced the cellular viability, whereas MB and LED alone were innocuous to fibroblasts. MB-mediated aPDT increased the expression of cytochrome C and Fas-L after 3 h, and Bax/Bcl-2, Bad/Bcl-2, and VDAC-1 after 6 h from treatment. Based on these results, MB-mediated aPDT induced cytotoxicity on mouse fibroblasts, with consequent activation of Bcl-2 apoptosis signaling pathways. Further studies are needed to determine the adequate parameters of aPDT to inactivate microorganisms without damaging fibroblasts.

Methylene Blue-Mediated Photodynamic Therapy Induces Macrophage Apoptosis via ROS and Reduces Bone Resorption in Periodontitis.

Aim To investigate whether methylene blue-mediated photodynamic therapy (MB-PDT) can affect the “fate” of macrophages in vitro or in periodontitis tissues and to explore the potential mechanism. Methods For in vitro treatments, THP-1 macrophages were divided into three experimental groups: C/control, no treatment; MB, methylene blue treatment; and MB-PDT, MB and laser irradiation treatment. Then, apoptosis and apoptosis-related proteins were detected in each group. For in vivo treatments, periodontitis was ligature-induced in the first molars of the bilateral maxilla in 12 Sprague Dawley (SD) rats. After six weeks, the ligatures were removed and all the induced molars underwent scaling and root planning (SRP). Then, the rats were divided into three groups according to the following treatments: SRP, saline solution; MB, phenothiazinium dye; and MB-PDT, MB and laser irradiation. Apoptotic macrophages, inflammation levels, and alveolar bone resorption in the periodontal tissues of rats were analyzed in each group. Results In vitro, flow cytometry analysis demonstrated that 10 μM MB and 40 J/cm2 laser irradiation maximized the apoptosis rate (34.74%) in macrophages. Fluorescence probe and Western blot analyses showed that MB-PDT induced macrophage apoptosis via reactive oxygen species (ROS) and the mitochondrial-dependent apoptotic pathway. Conversely, the addition of exogenous antioxidant glutathione (GSH) and the pan-caspase inhibitor Z-VAD-FMK markedly reduced the apoptotic response in macrophages. In vivo, immunohistochemistry, histology, radiographic, and molecular biology experiments revealed fewer infiltrated macrophages, less bone loss, and lower IL-1β and TNF-α levels in the MB-PDT group than in the SRP and MB groups (P < 0.05). Immunohistochemistry analysis also detected apoptotic macrophages in the MB-PDT group. Conclusion MB-PDT could induce macrophage apoptosis in vitro and in rats with periodontitis. This may be another way for MB-PDT to relieve periodontitis in addition to its antimicrobial effect. Meanwhile, MB-PDT induced apoptosis in THP-1 macrophages via the mitochondrial caspase pathway.

Antimicrobial efficacy of methylene blue-mediated photodynamic therapy on titanium alloy surfaces in vitro.

Bacterial elimination using antimicrobial photodynamic therapy (aPDT) has been considered an alternative therapeutic modality in peri-implantitis treatment. The present in vitro study evaluated the dose-dependent and pH-dependent bactericidal effects of methylene blue (MB)-mediated aPDT at eliminating Gram-negative (P. gingivalis and A. actinomycetemcomitans) and Gram-positive (S. mutans) bacteria on sandblasting, large-grit and acid-etching (SLA)-pretreated titanium alloy. The effects of different MB concentrations (50, 100, and 200 μg/mL), the pH of the MB (4, 7, and 10), and irradiation time (0, 30, and 60 s) on the bacterial viability and residual lipopolysaccharide (LPS) levels were examined. The variations in the pH of the MB solution after aPDT for 60 s on the uncontaminated and contaminated specimens were also detected. The experimental results indicated that MB-mediated PDT could effectively kill the majority of bacteria on the titanium alloy surfaces of biofilm-contaminated implants compared with the MB alone. Of note, aPDT exhibited better antibacterial efficacy with increase in the MB concentration and irradiation time. While treated in an acidic solution on the biofilm-contaminated specimens, aPDT caused the pH to increase. By contrast, the initially high alkaline pH decreased to a value of about pH 8.5 after aPDT. Intriguingly, the neutral pH had minor changes, independent of the MB concentration and bacterial species. As expected, aPDT with higher MB concentration at higher pH environment significantly lowered the LPS concentration of A. actinomycetemcomitans and P. gingivalis. On the basis of the data, the aPDT with 200 μg/mL MB at pH 10 for 60 s of irradiation time might be an effectively treatment to eliminate bacteria and LPS adherent to titanium surface, however, the use of the multispecies biofilm model and the evaluation of in vitro osteogenesis needed to be further evaluated.

Effects of methylene blue-mediated photodynamic therapy on a mouse model of squamous cell carcinoma and normal skin.

Photodynamic therapy is used to treat a variety of cancers and skin diseases by inducing apoptosis, necrosis, immune system activation, and/or vascular damage. Here, we describe the effects of a single photodynamic therapy session using methylene blue on a mouse model of squamous cell carcinoma and normal skin. The photodynamic therapy protocol comprised application of a 1% methylene blue solution, followed by irradiation with a diode laser for 15 min at 74 mW/cm2, for a total dose of 100 J/cm2. Morphological changes, cell proliferation, apoptosis, collagen quantity, immune system activity, and blood vessel number were analyzed 24 h and 15 days after photodynamic therapy. In the squamous cell carcinoma group, photodynamic therapy reduced tumor size and cell proliferation and raised cytokine levels. In normal skin, it decreased cell proliferation and collagen quantity and increased apoptosis and blood vessel numbers. The effects of photodynamic therapy were greater on normal skin than squamous cell carcinoma tissues. The reduced epithelial thickness and keratinization of the former are factors that contribute to the efficacy of this treatment. Adjustments to the treatment protocol are necessary to potentiate the effects for squamous cell carcinoma therapy.

Assessment of Photodynamic Therapy and Miconazole in the Management of Denture Stomatitis

Objectives: Candida-induced denture stomatitis can be treated with different approaches; one of the most effective methods is the Photodynamic therapy (PDT). The current study aimed to assess the collective effects of PDT and low-level laser therapy (LLLT) in comparison to the topical application of miconazole gel. Materials and Methods: Thirty complete denture wearers with a history of Candida-induced denture stomatitis were haphazardly divided into two groups 15 patient each. Group I: The patients were subjected to a session of methylene blue-mediated PDT followed by to 2 sessions of (LLLT) 2 times per week for 15 days, while those of Group II: were subjected to miconazole gel four times per day for fifteen days. Results: The clinical and cytolological findings revealed that 50% of the patients in Group I have been cured from denture stomatitis while in Group II the curing rate was 85%. After two weeks following treatment stoppage, 30% of Group I patients showed reappearance of symptoms while the ration of recurrence was 15% in Group II patients. Conclusion: Photodynamic therapy associated with low level laser therapy (LLLT) and Miconazole gel can be considered effective methods in the management of Candida-induced denture stomatitis with higher curing rate and lower recurrence ratio for Miconazole.