-
Biochimica Et Biophysica Acta.... Mar 2023Photodynamic therapy (PDT) is a process in which a photosensitizer (PS) is exposed to specific wavelengths and generates reactive oxygen species (ROS) which act within...
Photodynamic therapy (PDT) is a process in which a photosensitizer (PS) is exposed to specific wavelengths and generates reactive oxygen species (ROS) which act within nanometers. The low invasive nature and directed cytotoxicity of this approach render it attractive to the treatment of different conditions, including the ones that affect the central nervous system (CNS). The effect of PDT on healthy neurons is one main concern over its use in the CNS, since neuronal-like cells were shown to be particularly sensitive to certain PSs. Among available PSs, 1,9-dimethyl-methylene blue (DMMB) stands out as being resistant to reduction to its inactive leuco form and by being able to produce high levels of singlet‑oxygen. In this study, we aimed to investigate DMMB photodamage mechanisms in the hippocampal cell line HT22. Our results demonstrate that DMMB-PDT decrease in cell viability was linked with an increase in cell death and overall ROS production. Besides, it resulted in a significant increase in mitochondrial ROS production and decreased mitochondria membrane potential. Furthermore, DMMB-PDT significantly increased the presence of acidic autolysosomes, which was accompanied by an increase in ATG1 and ATG8 homologue GaBarap1 expression, and decreased DRAM1 expression. Taken together our results indicated that mitochondrial and autophagic dysfunction underlie DMMB-PDT cytotoxicity in neuronal cells.
Topics: Photochemotherapy; Methylene Blue; Reactive Oxygen Species; Mitochondria
PubMed: 36608805
DOI: 10.1016/j.bbamcr.2022.119429 -
Scientific Reports Jun 2023Assessing the in vitro toxicity of compounds on cell cultures is an important step during the screening of candidate molecules for diverse applications. Among the...
Assessing the in vitro toxicity of compounds on cell cultures is an important step during the screening of candidate molecules for diverse applications. Among the strategies employed to determine cytotoxicity, MTT, neutral red, and resazurin are commonly used. Methylene blue (MB), a phenothiazinium salt, has several uses, such as dye, redox indicator, and even as treatment for human disease and health conditions, such as malaria and methemoglobinemia. However, MB has only been sparsely used as a cellular toxicity indicator. As a viability indicator, MB is mostly applied to fixed cultures at high concentrations, especially when compared to MTT or neutral red. Here we show that MB and its related compounds new methylene blue (NMB), toluidine blue O (TBO), and dimethylmethylene blue (DMMB) can be used as cytotoxicity indicators in live (non-fixed) cells treated for 72 h with DMSO and cisplatin. We compared dye uptake between phenothiazinium dyes and neutral red by analyzing supernatant and cell content via visible spectra scanning and microscopy. All dyes showed a similar ability to assess cell toxicity compared to either MTT or neutral red. Our method represents a cost-effective alternative to in vitro cytotoxicity assays using cisplatin or DMSO, indicating the potential of phenothiazinium dyes for the screening of candidate drugs and other applications.
Topics: Humans; Coloring Agents; Phenothiazines; Cisplatin; Neutral Red; Dimethyl Sulfoxide; Methylene Blue
PubMed: 37353536
DOI: 10.1038/s41598-023-36721-0 -
Pharmacological Research Aug 2020Phenothiazines inhibit major antioxidant defense mechanisms in trypanosomatids and exhibit potent cytotoxic effects in vitro. However, the relevance of these drugs in...
Phenothiazines inhibit major antioxidant defense mechanisms in trypanosomatids and exhibit potent cytotoxic effects in vitro. However, the relevance of these drugs in the treatment of Trypanosoma cruzi-induced acute myocarditis is poorly explored, especially in combination with reference trypanocidal drugs. Thus, we compared the antiparasitic and cardioprotective potential of thioridazine (TDZ) and benznidazole (Bz) administered in monotherapy and combined in a murine model of T. cruzi-induced acute myocarditis. Female mice were randomized into six groups: (i) uninfected untreated, (ii) infected untreated, or infected treated with (iii) Bz (100 mg/kg), (iv) TDZ (80 mg/kg), (v) Bz (100 mg/kg) + TDZ (80 mg/kg), or (vi) Bz (50 mg/kg) + TDZ (80 mg/kg). Infected animals were inoculated with 2000 T. cruzi trypomastigotes and treated by gavage for 20 days. Animals that received TDZ alone presented the highest levels of parasitemia, parasitic load and anti-T. cruzi immunoglobulin G titers; cardiac upregulation of N-acetyl-β-D-glucosaminidase activity, nitric oxide, malondialdehyde and cytokines (IFN-γ, TNF-α, IL-10 and IL-17); as well as microstructural damage compared to the other groups (p < 0.05). These parameters were reduced in groups receiving Bz monotherapy compared to the other groups (p < 0.05). The combination of TDZ and Bz attenuated the response to treatment, worsening parasitological control, oxidative heart damage and myocarditis compared to the group treated with Bz alone (p < 0.05). Our results indicate that when administered alone, TDZ potentiated the pathological outcomes in animals infected with T. cruzi. Moreover, TDZ attenuated the antiparasitic effect of Bz when administered together, impairing parasitological control, potentiating inflammation, molecular oxidation and pathological microstructural remodeling of the heart. Thus, our findings indicate that TDZ acts as a pharmacological risk factor and Bz-based monotherapy remains a better cardioprotective drug against Trypanosoma cruzi-induced acute myocarditis.
Topics: Animals; Antiprotozoal Agents; Chagas Cardiomyopathy; Chagas Disease; Drug Therapy, Combination; Female; Mice; Myocarditis; Nitroimidazoles; Phenothiazines; Trypanocidal Agents; Trypanosoma cruzi
PubMed: 32416214
DOI: 10.1016/j.phrs.2020.104907 -
Molecular Informatics Oct 2023Phenothiazine derivatives can unselectively inhibit the trypanothione-dependent antioxidant system enzyme trypanothione reductase (TR). A virtual screening of 2163...
Phenothiazine derivatives can unselectively inhibit the trypanothione-dependent antioxidant system enzyme trypanothione reductase (TR). A virtual screening of 2163 phenothiazine derivatives from the ZINC15 and PubChem databases docked on the active site of T. cruzi TR showed that 285 compounds have higher affinity than the natural ligand trypanothione disulfide. 244 compounds showed higher affinity toward the parasite's enzyme than to its human homolog glutathione reductase. Protein-ligand interaction profiling predicted that the main interactions for the top scored compounds were with residues important for trypanothione disulfide binding: Phe396, Pro398, Leu399, His461, Glu466, and Glu467, particularly His461, which participates in catalysis. Two compounds with the desired profiles, ZINC1033681 (Zn_C687) and ZINC10213096 (Zn_C216), decreased parasite growth by 20 % and 50 %, respectively. They behaved as mixed-type inhibitors of recombinant TR, with Ki values of 59 and 47 μM, respectively. This study provides a further understanding of the potential of phenothiazine derivatives as TR inhibitors.
Topics: Humans; Molecular Docking Simulation; Molecular Dynamics Simulation; Trypanosoma cruzi; Ligands; Phenothiazines; Disulfides
PubMed: 37490403
DOI: 10.1002/minf.202300069 -
Bioorganic Chemistry Oct 2020In the incessant search for innovative cancer control strategies, this study was devoted to the design, synthesis and pharmacological evaluation of dual inhibitors of...
In the incessant search for innovative cancer control strategies, this study was devoted to the design, synthesis and pharmacological evaluation of dual inhibitors of farnesyltransferase and tubulin polymerization (FTI/MTIs). A series of indolizine-phenothiazine hybrids 16 (amides) and 17 (ketones) has been obtained in a 4-step procedure. The combination of the two heterocycles provided potent tubulin polymerization inhibitors with similar efficiency as the reference phenstatin and (-)-desoxypodophyllotoxin. Ketones 17 were also able to inhibit human farnesyltransferase (FTase) in vitro. Interestingly, three molecules 17c, 17d and 17f were very effective against both considered biological targets. Next, nine indolizine-phenothiazine hybrids 16c, 16f, 17a-f and 22b were evaluated for their cell growth inhibition potential on the NCI-60 cancer cell lines panel. Ketones 17a-f were the most active and displayed promising cellular activities. Not only they arrested the cell growth of almost all tested cancer cells, but they displayed cytotoxicity potential with GI values in the low nanomolar range. The most sensitive cell lines upon treatment with indolizine-phenothiazine hybrids were NCI-H522 (lung cancer), COLO-205 and HT29 (colon cancer), SF-539 (human glioblastoma), OVCAR-3 (ovarian cancer), A498 (renal cancer) and especially MDA-MB-435 (melanoma). Demonstrating the preclinical effectiveness of these dual inhibitors can be crucial. A single dual molecule could induce a synergy of antitumor activity, while increasing the effectiveness and reducing the toxicity of the classical combo treatments currently used in chemotherapy.
Topics: Antineoplastic Agents; Binding Sites; Cell Line, Tumor; Cell Proliferation; Drug Design; Drug Screening Assays, Antitumor; Enzyme Inhibitors; Farnesyltranstransferase; Humans; Indolizines; Molecular Docking Simulation; Molecular Structure; Phenothiazines; Protein Binding; Structure-Activity Relationship; Tubulin; Tubulin Modulators
PubMed: 32891861
DOI: 10.1016/j.bioorg.2020.104184 -
Journal of Fluorescence Nov 2023Hypochlorous acid (HClO), a highly reactive oxygen species, has important effects on human health. High selectivity and sensitivity remain challenges of fluorescent...
Hypochlorous acid (HClO), a highly reactive oxygen species, has important effects on human health. High selectivity and sensitivity remain challenges of fluorescent probes for detection of ClO with a large Stokes shift. This work designed and synthesized a novel phenothiazine-based fluorescent probe TF which can detect ClO by colorimetric and fluorescent dual signals. TF displayed turn-on fluorescence effect toward ClO with high selectivity (≥ 28-folds) and sensitivity (LOD = 0.472 μM), fast response time (< 1 min) and large Stokes shift (150 nm) in PBS (pH = 7.4, 40% DMSO). Meanwhile, TF can visualize ClO on the mung bean sprouts model and apply as testing strips for portable and rapid detecting ClO by the naked eyes. A phenothiazine-based fluorescent probe with large Stokes shift was synthesized and its responding rapidly ability to detect ClO was studied.
Topics: Humans; Fluorescent Dyes; Hypochlorous Acid; Phenothiazines; Colorimetry
PubMed: 37129794
DOI: 10.1007/s10895-023-03215-1 -
Photodiagnosis and Photodynamic Therapy Dec 2019Orthodontics involves diagnosis and treatment of dental and skeletal malocclusions. Orthodontic apparatus may repair these malocclusions but may also impair oral hygiene...
BACKGROUND
Orthodontics involves diagnosis and treatment of dental and skeletal malocclusions. Orthodontic apparatus may repair these malocclusions but may also impair oral hygiene making patients prone to develop both periodontal diseases and caries. Antimicrobial agents may be used to prevent this.To avoid increased antimicrobial resistance to available drugs, A-PDT (Antimicrobial Photodynamic Therapy) appears as a viable alternative.
OBJECTIVE
This work aimed to evaluate the efficacy of A-PDT on reducing the number of colony forming units (CFU) through the use of phenothiazine compound (methylene blue+ toluidine blue) as a photosensitizer, associated with red LED (λ640±5ηm) irradiation in orthodontic patients.
METHODOLOGY
Twenty-one patients consented to participate in the study. Three biofilm collections were performed around the brackets and gums of the inferior central incisors; first before any intervention (Control); second after 5min of pre-irradiation and the last one immediately after AmPDT. Subsequently, a microbiological routine for microorganism growth period were performed and CFU counting after a 24h done.
RESULTS
The data showed that the AmPDT was able to reduce CFU count around 90% when compared to Control group (p=0.007) and also between the A-PDT and Photosensitizer groups (p=0.010). However, there were no differences between the Control and Photosensitizer groups.
CONCLUSION
A-PDT associated with the use of phenothiazine compounds and red LED was able to significantly reduce the number of CFUs in orthodontic patients.
Topics: Antiprotozoal Agents; Biofilms; Colony Count, Microbial; Cross-Over Studies; Dental Caries; Humans; Methylene Blue; Orthodontic Brackets; Periodontal Diseases; Phenothiazines; Photochemotherapy; Photosensitizing Agents; Tolonium Chloride
PubMed: 31394297
DOI: 10.1016/j.pdpdt.2019.08.002 -
Photodermatology, Photoimmunology &... Jul 2022Photodynamic therapy is a two-step procedure, involving the use of photosensitizing agents followed by selective illumination of the target lesion with visible light....
BACKGROUND
Photodynamic therapy is a two-step procedure, involving the use of photosensitizing agents followed by selective illumination of the target lesion with visible light. Photodynamic therapy has been described recently as a promising strategy for treatment of leishmaniasis. This study aims to evaluate the in vitro phototoxic, morphological, and apoptotic effect of methylene blue, toluidine blue, chloro-aluminum phthalocyanine, and pheophorbide a-mediated photodynamic therapy on the viability of Leishmania tropica promastigotes.
METHODS
Parasites were treated with methylene blue, toluidine blue, chloro-aluminum phthalocyanine, and pheophorbide a or/and methylene blue, toluidine blue, chloro-aluminum phthalocyanine, and pheophorbide a-mediated photodynamic therapy, and cell proliferation, morphological changes, and apoptosis were evaluated by XTT, giemsa staining, DAPI staining, and DNA fragmentation, respectively.
RESULTS
Parasite viability was significantly different in between the groups treated with methylene blue, toluidine blue, and pheophorbide a, with or without irradiation. chloro-aluminum phthalocyanine treatment did not lead to any alterations in cell viability in Leishmania tropica promastigotes with or without irradiation. DAPI staining results indicated that apoptotic bodies and nucleus fragmentation started to be visible in methylene blue, chloro-aluminum phthalocyanine, and pheophorbide a-mediated photodynamic therapy groups. DNA ladder pattern which is used to define apoptosis was observed in irradiated methylene blue, chloro-aluminum phthalocyanine, and pheophorbide a groups.
CONCLUSIONS
The results revealed that apoptosis-induced cell death was observed in Leishmania tropica promastigotes after the application of photosensitizers in combination with light irradiation.
Topics: Humans; Leishmania tropica; Methylene Blue; Photochemotherapy; Photosensitizing Agents; Tolonium Chloride
PubMed: 34897808
DOI: 10.1111/phpp.12758 -
Journal of Photochemistry and... Sep 2021Retinoblastoma 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....
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 50 μM), in the absence of light (dark cytotoxicity) and, in the presence of 664 nm laser light (phototoxicity) with fluences of 1, 1.5, 3, 5, 7, 10, and 15 J/cm. 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.
Topics: Cell Line, Tumor; Cell Proliferation; Cell Survival; Humans; Lasers; Methylene Blue; Photochemotherapy; Photosensitizing Agents; Retinoblastoma; Singlet Oxygen
PubMed: 34304071
DOI: 10.1016/j.jphotobiol.2021.112260 -
Journal of Veterinary Emergency and... Nov 2021To determine the effects of time after sampling on CO-oximetry measurements of equine blood samples and the effects of adding ascorbic acid (AscAc) and methylene blue...
OBJECTIVES
To determine the effects of time after sampling on CO-oximetry measurements of equine blood samples and the effects of adding ascorbic acid (AscAc) and methylene blue (MetBlue) to samples with methemoglobinemia.
DESIGN
Experimental study.
SETTING
University teaching hospital.
ANIMALS
Thirty healthy adult horses assigned to 5 groups.
INTERVENTIONS
Repeated CO-oximetry determinations were performed on venous (n = 6) and arterial blood samples (n = 7) stored at 0°C for 48 hours. Methemoglobinemia was induced in vitro in 17 additional blood samples. Six were used as untreated controls, 6 had AscAc added, and 5 had MetBlue added. Total hemoglobin, oxyhemoglobin, carboxyhemoglobin, methemoglobin (MetHb), and oxygen saturation of hemoglobin (SO ) were measured.
MEASUREMENTS AND MAIN RESULTS
Oxyhemoglobin and SO increased from 69.8% ± 10.2% and 90% ± 3% to 82.8% ± 7.9% and 99% ± 3%, respectively, after 8 hours in venous blood (mean ± SD, P < 0.001). There was an effect of treatment (P = 0.032) and of time (interaction P = 0.003) on MetHb% in methemoglobinemic samples. The difference in absolute MetHb% from time 0 was as follows: 7.0% (interquartile range [IQR] = 21.2), -0.2% (IQR = 3.5), and -4.4% (IQR = 5.2) at 48 hours in control, AscAc, and MetBlue groups, respectively (P < 0.05). There was no effect of time on MetHb% in the AscAc group (23% [IQR = 52.6] at time 0 to 23.2% [IQR = 56.9] after 48 h).
CONCLUSIONS
Storage of blood in ice water to determine O Hb and SO using a CO-oximeter should not exceed 4 hours. Measurement of MetHb% could be delayed by up to 48 hours if AscAc is added to the sample. MetBlue significantly decreased MetHb% over time. The limitations of this study include the fact that the antioxidant effects of AscAc and MetBlue were evaluated in vitro and not in vivo. Further studies are needed to evaluate different storage temperatures and syringe types.
Topics: Animals; Antioxidants; Ascorbic Acid; Horses; Methylene Blue; Oximetry; Oxygen Saturation
PubMed: 34427385
DOI: 10.1111/vec.13089