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1.
Sci Rep ; 11(1): 21723, 2021 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-34741051

RESUMO

Coronavirus with intact infectivity attached to PPE surfaces pose significant threat to the spread of COVID-19. We tested the hypothesis that an electroceutical fabric, generating weak potential difference of 0.5 V, disrupts the infectivity of coronavirus upon contact by destabilizing the electrokinetic properties of the virion. Porcine respiratory coronavirus AR310 particles (105) were placed in direct contact with the fabric for 1 or 5 min. Following one minute of contact, zeta potential of the porcine coronavirus was significantly lowered indicating destabilization of its electrokinetic properties. Size-distribution plot showed appearance of aggregation of the virus. Testing of the cytopathic effects of the virus showed eradication of infectivity as quantitatively assessed by PI-calcein and MTT cell viability tests. This work provides the rationale to consider the studied electroceutical fabric, or other materials with comparable property, as material of choice for the development of PPE in the fight against COVID-19.


Assuntos
COVID-19/prevenção & controle , COVID-19/transmissão , Eletroquímica/métodos , Têxteis , Animais , Anti-Infecciosos , Líquidos Corporais , Linhagem Celular , Sobrevivência Celular , Fluoresceínas , Humanos , Peróxido de Hidrogênio , Cinética , Nanopartículas , Propídio , SARS-CoV-2 , Suínos , Temperatura , Sais de Tetrazólio , Tiazóis , Vírion , Cicatrização
2.
Sensors (Basel) ; 21(21)2021 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-34770479

RESUMO

Ischemic stroke is one of the leading causes of death among the aged population in the world. Experimental stroke models with rodents play a fundamental role in the investigation of the mechanism and impairment of cerebral ischemia. For its celerity and veracity, the 2,3,5-triphenyltetrazolium chloride (TTC) staining of rat brains has been extensively adopted to visualize the infarction, which is subsequently photographed for further processing. Two important tasks are to segment the brain regions and to compute the midline that separates the brain. This paper investigates automatic brain extraction and hemisphere segmentation algorithms in camera-based TTC-stained rat images. For rat brain extraction, a saliency region detection scheme on a superpixel image is exploited to extract the brain regions from the raw complicated image. Subsequently, the initial brain slices are refined using a parametric deformable model associated with color image transformation. For rat hemisphere segmentation, open curve evolution guided by the gradient vector flow in a medial subimage is developed to compute the midline. A wide variety of TTC-stained rat brain images captured by a smartphone were produced and utilized to evaluate the proposed segmentation frameworks. Experimental results on the segmentation of rat brains and cerebral hemispheres indicated that the developed schemes achieved high accuracy with average Dice scores of 92.33% and 97.15%, respectively. The established segmentation algorithms are believed to be potential and beneficial to facilitate experimental stroke study with TTC-stained rat brain images.


Assuntos
Isquemia Encefálica , Cérebro , Acidente Vascular Cerebral , Algoritmos , Animais , Encéfalo/diagnóstico por imagem , Isquemia Encefálica/diagnóstico por imagem , Processamento de Imagem Assistida por Computador , Ratos , Acidente Vascular Cerebral/diagnóstico por imagem , Sais de Tetrazólio
3.
Int J Mol Sci ; 22(18)2021 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-34576274

RESUMO

Biotransformation of four bioactive phenolic constituents from licorice, namely licoisoflavanone (1), glycyrrhisoflavone (2), echinatin (3), and isobavachalcone (4), was performed by the selected fungal strain Aspergillus niger KCCM 60332, leading to the isolation of seventeen metabolites (5-21). Structures of the isolated compounds were determined on the basis of extensive spectroscopic methods, twelve of which (5-7, 10-17 and 19) have been previously undescribed. A series of reactions including hydroxylation, hydrogenation, epoxidation, hydrolysis, reduction, cyclization, and alkylation was observed in the biotransformation process. All compounds were tested for their cytotoxic activities against three different human cancer cell lines including A375P, MCF-7, and HT-29. Compounds 1 and 12 exhibited most considerable cytotoxic activities against all the cell lines investigated, while compounds 2 and 4 were moderately cytotoxic. These findings will contribute to expanding the chemical diversity of phenolic compounds, and compounds 1 and 12 may serve as leads for the development of potential cancer chemopreventive agents.


Assuntos
Biotransformação , Glycyrrhiza/química , Fenol/química , Anticarcinógenos/farmacologia , Antineoplásicos/química , Aspergillus niger/metabolismo , Linhagem Celular Tumoral , Fermentação , Fungos/metabolismo , Células HT29 , Humanos , Hidrólise , Concentração Inibidora 50 , Células MCF-7 , Fenóis , Extratos Vegetais , Raízes de Plantas/efeitos dos fármacos , Pós , Rizoma/metabolismo , Espectrofotometria , Sais de Tetrazólio/farmacologia , Tiazóis/farmacologia
4.
Cancer Sci ; 112(10): 4037-4049, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34309966

RESUMO

Immunotherapy with immune-checkpoint therapy has recently been used to treat oral squamous cell carcinomas (OSCCs). However, improvements in current immunotherapy are expected because response rates are limited. Transforming growth factor-ß (TGF-ß) creates an immunosuppressive tumor microenvironment (TME) by inducing the production of regulatory T-cells (Tregs) and cancer-associated fibroblasts and inhibiting the function of cytotoxic T-lymphocytes (CTLs) and natural killer cells. TGF-ß may be an important target in the development of novel cancer immunotherapies. In this study, we investigated the suppressive effect of TGF-ß on CTL function in vitro using OSCC cell lines and their specific CTLs. Moreover, TGFB1 mRNA expression and T-cell infiltration in 25 OSCC tissues were examined by in situ hybridization and multifluorescence immunohistochemistry. We found that TGF-ß suppressed the function of antigen-specific CTLs in the priming and effector phases in vitro. Additionally, TGF-ß inhibitor effectively restored the CTL function, and TGFB1 mRNA was primarily expressed in the tumor invasive front. Interestingly, we found a significant negative correlation between TGFB1 mRNA expression and the CD8+ T-cell/Treg ratio and between TGFB1 mRNA expression and the Ki-67 expression in CD8+ T-cells, indicating that TGF-ß also suppressed the function of CTLs in situ. Our findings suggest that the regulation of TGF-ß function restores the immunosuppressive TME to active status and is important for developing new immunotherapeutic strategies, such as a combination of immune-checkpoint inhibitors and TGF-ß inhibitors, for OSCCs.


Assuntos
Inibidores de Checkpoint Imunológico/uso terapêutico , Imunoterapia Adotiva/métodos , Neoplasias Bucais/terapia , Carcinoma de Células Escamosas de Cabeça e Pescoço/terapia , Linfócitos T Citotóxicos/efeitos dos fármacos , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Microambiente Tumoral/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Fibroblastos Associados a Câncer/citologia , Fibroblastos Associados a Câncer/imunologia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Humanos , Interferon gama/análise , Interferon gama/metabolismo , Antígeno Ki-67/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Linfócitos do Interstício Tumoral/citologia , Linfócitos do Interstício Tumoral/imunologia , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/metabolismo , RNA Mensageiro/metabolismo , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Sais de Tetrazólio/farmacologia , Fator de Crescimento Transformador beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/imunologia , Fator de Crescimento Transformador beta1/análise , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
5.
Biomed Res Int ; 2021: 5514669, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34136566

RESUMO

Pyruvate kinase (PK), a key enzyme that determines glycolytic activity, has been known to support the metabolic phenotype of tumor cells, and specific pyruvate kinase isoform M2 (PKM2) has been reported to fulfill divergent biosynthetic and energetic requirements of cancerous cells. PKM2 is overexpressed in several cancer types and is an emerging drug target for cancer during recent years. Therefore, this study was carried out to identify PKM2 inhibitors from natural products for cancer treatment. Based on the objectives of this study, firstly, plant extract library was established. In order to purify protein for the establishment of enzymatic assay system, pET-28a-HmPKM2 plasmid was transformed to E. coli BL21 (DE3) cells for protein expression and purification. After the validation of enzymatic assay system, plant extract library was screened for the identification of inhibitors of PKM2 protein. Out of 51 plant extracts screened, four extracts Mangifera indica (leaf, seed, and bark) and Bombex ceiba bark extracts were found to be inhibitors of PKM2. In the current study, M. indica (leaf, seed, and bark) extracts were further evaluated dose dependently against PKM2. These extracts showed different degrees of concentration-dependent inhibition against PKM2 at 90-360 µg/ml concentrations. We have also investigated the anticancer potential of these extracts against MDA-MB231 cells and generated dose-response curves for the evaluation of IC50 values. M. indica (bark and seed) extracts significantly halted the growth of MDA-MB231 cells with IC50 values of 108 µg/ml and 33 µg/ml, respectively. Literature-based phytochemical analysis of M. indica was carried out, and M. indica-derived 94 compounds were docked against three binding sites of PKM2 for the identification of PKM2 inhibitors. The results of in silico based screening have unveiled various PKM2 modulators; however, further studies are recommended to validate their PKM2 inhibitory potential via in vitro biochemical assay. The results of this study provide novel findings for possible mechanism of action of M. indica (bark and seed) extracts against TNBC via PKM2 inhibition suggesting that M. indica might be of therapeutic interest for the treatment of TNBC.


Assuntos
Proteínas de Transporte/antagonistas & inibidores , Mangifera/metabolismo , Proteínas de Membrana/antagonistas & inibidores , Extratos Vegetais/farmacologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Cristalografia por Raios X , Inibidores Enzimáticos/farmacologia , Feminino , Humanos , Concentração Inibidora 50 , Cinética , Casca de Planta/metabolismo , Folhas de Planta/metabolismo , Plasmídeos/metabolismo , Sementes/metabolismo , Sais de Tetrazólio , Tiazóis , Hormônios Tireóideos , Neoplasias de Mama Triplo Negativas/enzimologia
6.
PLoS One ; 16(6): e0252961, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34138928

RESUMO

N. gonorrhoeae is one of the most pressing antibiotic resistant threats of our time and low-cost diagnostics that can easily identify antibiotic resistance are desperately needed. However, N. gonorrhoeae responds so uniquely to growth conditions that it cannot be assumed gonorrhea will respond to common microbiological methods used for other pathogenic organisms. In this paper, we explore visual colorimetric indicators of N. gonorrhoeae growth that can be seen without a microscope or spectrophotometer. We evaluate growth media, pH indicators, resazurin-based dyes, and tetrazolium-based dyes for their use in simple colorimetric system. Overall, we identified Graver Wade media as the best at supporting robust gonococcal growth while also providing the least background when analyzing results of colorimetric tests. XTT, a tetrazolium-based dye, proved to show to brightest color change over time and not negatively impact the natural growth of N. gonorrhoeae. However, other dyes including PrestoBlue, MTT, and NBT are less expensive than XTT and work well when added after bacterial growth has already occurred. By identifying the specific use cases of these dyes, this research lays the groundwork for future development of a color-based antibiotic susceptibility low-cost test for N. gonorrhoeae.


Assuntos
Técnicas Bacteriológicas/métodos , Gonorreia/diagnóstico , Neisseria gonorrhoeae/crescimento & desenvolvimento , Colorimetria , Meios de Cultura/química , Diagnóstico Precoce , Humanos , Concentração de Íons de Hidrogênio , Neisseria gonorrhoeae/isolamento & purificação , Oxazinas/química , Sensibilidade e Especificidade , Sais de Tetrazólio/química , Xantenos/química
7.
Molecules ; 26(11)2021 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-34071298

RESUMO

Chemotherapeutic agents, which contain the Michael acceptor, are potent anticancer molecules by promoting intracellular reactive oxygen species (ROS) generation. In this study, we synthesized a panel of PL (piperlongumine) analogs with chlorine attaching at C2 and an electron-withdrawing/electron-donating group attaching to the aromatic ring. The results displayed that the strong electrophilicity group at the C2-C3 double bond of PL analogs plays an important role in the cytotoxicity whereas the electric effect of substituents, which attached to the aromatic ring, partly contributed to the anticancer activity. Moreover, the protein containing sulfydryl or seleno, such as TrxR, could be irreversibly inhibited by the C2-C3 double bond of PL analogs, and boost intracellular ROS generation. Then, the ROS accumulation could disrupt the redox balance, induce lipid peroxidation, lead to the loss of MMP (Mitochondrial Membrane Potential), and ultimately result in cell cycle arrest and A549 cell line death. In conclusion, PL analogs could induce in vitro cancer apoptosis through the inhibition of TrxR and ROS accumulation.


Assuntos
Apoptose , Dioxolanos/química , Espécies Reativas de Oxigênio , Células A549 , Antineoplásicos/farmacologia , Ciclo Celular , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células , Cloro/química , Elétrons , Humanos , Peroxidação de Lipídeos , Potencial da Membrana Mitocondrial , Oxirredução , Sais de Tetrazólio/química , Tiazóis/química , Tiorredoxina Dissulfeto Redutase/metabolismo
8.
Molecules ; 26(11)2021 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-34072474

RESUMO

Ficus deltoidea var. deltoidea is used as traditional medicine for diabetes, inflammation, and nociception. However, the antimutagenic potential and cytoprotective effects of this plant remain unknown. In this study, the mutagenic and antimutagenic activities of F. deltoidea aqueous extract (FDD) on both Salmonella typhimurium TA 98 and TA 100 strains were assessed using Salmonella mutagenicity assay (Ames test). Then, the cytoprotective potential of FDD on menadione-induced oxidative stress was determined in a V79 mouse lung fibroblast cell line. The ferric-reducing antioxidant power (FRAP) assay was conducted to evaluate FDD antioxidant capacity. Results showed that FDD (up to 50 mg/mL) did not exhibit a mutagenic effect on either TA 98 or TA 100 strains. Notably, FDD decreased the revertant colony count induced by 2-aminoanthracene in both strains in the presence of metabolic activation (p < 0.05). Additionally, pretreatment of FDD (50 and 100 µg/mL) demonstrated remarkable protection against menadione-induced oxidative stress in V79 cells significantly by decreasing superoxide anion level (p < 0.05). FDD at all concentrations tested (12.5-100 µg/mL) exhibited antioxidant power, suggesting the cytoprotective effect of FDD could be partly attributed to its antioxidant properties. This report highlights that F. deltoidea may provide a chemopreventive effect on mutagenic and oxidative stress inducers.


Assuntos
Antimutagênicos/química , Antioxidantes/química , Ficus/metabolismo , Extratos Vegetais/química , Animais , Ânions , Linhagem Celular , Cricetulus , Diabetes Mellitus , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Glutationa , Camundongos , Mutagênese/efeitos dos fármacos , Testes de Mutagenicidade , Mutagênicos , Estresse Oxidativo , Salmonella typhimurium/efeitos dos fármacos , Sais de Tetrazólio/química , Tiazóis/química , Vitamina K 3/química , Água
9.
BMC Vet Res ; 17(1): 198, 2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34034733

RESUMO

BACKGROUND: Betulin, a natural pentacyclic triterpene with the lupane structure that is present in significant amounts in the outer bark of birch, is known for its broad array of biological and pharmacological properties. Betulin has attracted attention as a potential, natural-origin antimicrobial substance. The literature describes it as selectively toxic to neoplastic cells but safe for normal cells. The research aim was to evaluate the basal cytotoxicity of betulin towards fish (BF-2) and murine (NIH/3T3) fibroblasts. We used four colorimetric tests that provide a preliminary evaluation of possible mechanisms of the cytotoxicity of a compound to assess the degree of the toxicity of betulin after 24, 48 and 72 h of incubation with cells: the MTT assay (mitochondrial activity assessment), the NRU assay (lysosomal membrane integrity assessment), the LDH assay (cellular membrane integrity assessment) and the SRB assay (total cellular protein content determination). RESULTS: The results revealed an exceptionally high sensitivity of mitochondria to the effect of betulin, with the other endpoints being less sensitive. Although murine fibroblasts were more vulnerable to the toxic effect of betulin than fish fibroblasts, the betulin CC50 values for both cell lines were comparable with analogous IC50 values determined by other researchers in studies involving cancerous cells. CONCLUSIONS: The results indicate the need to verify the claim about the selective toxicity of betulin towards malignant cells and to conduct safety/toxicity tests before any potential therapeutic use of betulin in veterinary medicine.


Assuntos
Antineoplásicos Fitogênicos/toxicidade , Fibroblastos/efeitos dos fármacos , Triterpenos/toxicidade , Células 3T3 , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Linhagem Celular , Citotoxinas/toxicidade , Dimetil Sulfóxido/toxicidade , Peixes , L-Lactato Desidrogenase/metabolismo , Camundongos , Vermelho Neutro/metabolismo , Solubilidade , Sais de Tetrazólio/metabolismo , Tiazóis/metabolismo , Triterpenos/química , Triterpenos/farmacologia
10.
Biomater Sci ; 9(9): 3300-3305, 2021 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-33870966

RESUMO

Tetrazolium-based assays such as the MTT assay have been commonly employed in evaluating biocompatibility. Here, we show that PDA (or its precursor dopamine (DA)) spontaneously reduces MTT and produces exaggerated cytocompatibility inferences. The extent of interference depends on the method of DA polymerization. We observed that the trypan blue exclusion assay allowed more accurate determination of cell viability in the presence of DA- and PDA-based nanomaterials.


Assuntos
Dopamina , Polímeros , Sobrevivência Celular , Indóis/toxicidade , Polímeros/toxicidade , Sais de Tetrazólio
11.
Environ Sci Pollut Res Int ; 28(33): 45390-45401, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33866499

RESUMO

Microbial toxicity of test substances in tetrazolium assay is often quantified while referring to their IC50 values. However, the implication of such an estimate is very limited and can differ across studies depending on prevailing test conditions. In this work, a factorial design-based end-point microbial toxicity assay was performed, which suggests a significant interaction (P= 0.041) between inoculum and tetrazolium dose on formazan production. Subsequently, a dynamic model framework was utilized to capture the nonlinearities in biomass, substrate, formazan profiles and to project the toxicant inhibition parameter as a robust alternative to IC50 value. Microbial growth, glucose uptake and formazan production in the presence or absence of toxicant (Cu2+) from designed batch experiments were used for sequential estimation of model parameters, and their confidence intervals. A logistic growth model with multiplicative inhibition terms for formazan content and toxicant concentration fits the experimental data reasonably well (R2>0.96). Dynamic relative sensitivity analysis revealed that both microbial growth and formazan production profiles were sensitive to toxicant inhibition parameter. The modelling framework not only provides a better insight into the underlying toxic effect but also offers a stable toxicity index for the test substances that can be extended to design a versatile, robust in vitro assay system.


Assuntos
Bioensaio , Testes de Toxicidade , Formazans , Sais de Tetrazólio
12.
Cells ; 10(5)2021 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-33925786

RESUMO

Inflammation-mediated skeletal muscle wasting occurs in patients with sepsis and cancer cachexia. Both conditions severely affect patient morbidity and mortality. Lithium chloride has previously been shown to enhance myogenesis and prevent certain forms of muscular dystrophy. However, to our knowledge, the effect of lithium chloride treatment on sepsis-induced muscle atrophy and cancer cachexia has not yet been investigated. In this study, we aimed to examine the effects of lithium chloride using in vitro and in vivo models of cancer cachexia and sepsis. Lithium chloride prevented wasting in myotubes cultured with cancer cell-conditioned media, maintained the expression of the muscle fiber contractile protein, myosin heavy chain 2, and inhibited the upregulation of the E3 ubiquitin ligase, Atrogin-1. In addition, it inhibited the upregulation of inflammation-associated cytokines in macrophages treated with lipopolysaccharide. In the animal model of sepsis, lithium chloride treatment improved body weight, increased muscle mass, preserved the survival of larger fibers, and decreased the expression of muscle-wasting effector genes. In a model of cancer cachexia, lithium chloride increased muscle mass, enhanced muscle strength, and increased fiber cross-sectional area, with no significant effect on tumor mass. These results indicate that lithium chloride exerts therapeutic effects on inflammation-mediated skeletal muscle wasting, such as sepsis-induced muscle atrophy and cancer cachexia.


Assuntos
Caquexia/prevenção & controle , Cloreto de Lítio/farmacologia , Músculo Esquelético/patologia , Atrofia Muscular/patologia , Neoplasias/tratamento farmacológico , Sepse/tratamento farmacológico , Sepse/prevenção & controle , Animais , Peso Corporal , Diferenciação Celular , Proliferação de Células , Meios de Cultivo Condicionados , Glicogênio Sintase Quinase 3 beta/biossíntese , Inflamação , Lipopolissacarídeos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Contração Muscular , Fibras Musculares Esqueléticas/efeitos dos fármacos , Proteínas Musculares/biossíntese , Músculo Esquelético/metabolismo , Neoplasias/complicações , Células RAW 264.7 , RNA Interferente Pequeno/metabolismo , Proteínas Ligases SKP Culina F-Box/biossíntese , Sais de Tetrazólio/farmacologia , Tiazóis/farmacologia
13.
Carbohydr Polym ; 257: 117617, 2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33541645

RESUMO

Current work focuses on the synthesis and characterization of novel pH-sensitive biocompatible gelatin/carboxymethyl cellulose based hydrogels by free radical polymerization technique cross-linked with glutaraldehyde. Effect of pH, polymer ratio and variable crosslinking concentrations on dynamic swelling, equilibrium swelling, porosity, sol-gel analysis and in vitro release pattern was investigated. Hydrogel structure was confirmed by FTIR, XRD, and DSC. Moreover scanning electron microscopy confirmed the porous structure of gel network. Various structure property relationships like average molecular weight between crosslinks (Mc), solvent interaction parameters, volume fraction of polymer (V2,s) and diffusion coefficient (D) that affect the release behaviour were determined. Results showed that maximum swelling and highest release of drug occurred at pH 1.2. Porosity and gel fraction increased by increasing polymer load. The invivo absorption and pharmacokinetics evaluation in rabbit's models revealed the controlled nature of hydrogels. MTT assay confirmed the biocompatible nature of blank hydrogels against Vero cell lines and cytotoxic potential against HeLa cell lines. The preliminary safety evaluation and oral tolerability revealed that the hydrogel solution is safe up to 4000 mg/kg body weight without causing any hematological or histopathological changes in rabbits.


Assuntos
Carboximetilcelulose Sódica/química , Sistemas de Liberação de Medicamentos , Fluoruracila/química , Gelatina/química , Hidrogéis/química , Animais , Disponibilidade Biológica , Varredura Diferencial de Calorimetria , Bovinos , Chlorocebus aethiops , Reagentes para Ligações Cruzadas/química , Liberação Controlada de Fármacos , Células HeLa , Humanos , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Microscopia Eletrônica de Varredura , Nanopartículas/química , Polimerização , Polímeros/química , Porosidade , Coelhos , Espectroscopia de Infravermelho com Transformada de Fourier , Sais de Tetrazólio/química , Tiazóis/química , Células Vero , Difração de Raios X
14.
Phys Chem Chem Phys ; 23(6): 3761-3770, 2021 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-33538741

RESUMO

Tetrazolium salts (TZs) are pervasively utilized as precursors in the dye industry, colorimetric probes in enzyme assays and for exploring nanomaterial toxicity, but its own toxicity is not investigated enough so far. Using femtosecond transient absorption spectroscopy, nanosecond pulse radiolysis (ns-PRL), western blotting and UV-vis absorption spectroscopy, here we characterized a neutral tetrazolinyl radical (with the same maximum absorption at 420 nm and different lifetimes of 5.0 and 9.0 µs for two selected TZs), the key intermediate of TZs reduction, and noticed TZs-formazan production under UV light irradiation accompanied by 41% increase in the cross-linking of lysozyme (Lyso, model protein) compared to TZs-free sample, which uncovered the photoenhanced oxidation of TZs towards Lyso. The ns-PRL in a reductive atmosphere simulated the electron/proton donors of amino acid residues in Lyso upon photoexcitation and revealed the reduction mechanism of TZs, as that first followed one-electron-transfer and then probably proton-coupled electron transfer. This is the first time to report on the photoenhanced oxidation mechanism of TZs, which would provide new insights into the applications of TZs in cell biology, "click" chemistry and nanotoxicology.


Assuntos
Aminoácidos/química , Muramidase/química , Sais de Tetrazólio/química , Aminoácidos/efeitos da radiação , Animais , Galinhas , Radicais Livres/química , Muramidase/efeitos da radiação , Oxirredução , Sais de Tetrazólio/efeitos da radiação , Raios Ultravioleta
15.
Acta Trop ; 217: 105864, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33607062

RESUMO

Accurate measurement of glucose-6-phosphate dehydrogenase (G6PD) activity is critical for malaria treatment as misclassification of G6PD deficiency could cause serious harm to patients. G6PD activity should be assessed in blood samples on the day of collection. Otherwise, specimens should be stored under suitable conditions to prevent loss of G6PD activity. Here, we assessed stability and integrity of G6PD testing in samples from normal controls, heterozygous females, and G6PD deficient individuals using water-soluble tetrazolium salts (WST-8) assay. Specimens were stored as ethylenediaminetetraacetic acid (EDTA) whole blood and dried blood spots (DBS) at various temperatures (37 °C, room temperature, 4 °C and -20 °C) and under different humidity conditions (with and without desiccant). G6PD normal samples were stable for up to 1 year when stored at -20 °C under controlled conditions, with 85% and 91% G6PD activity in EDTA whole blood and DBS in the presence of desiccant, respectively. Specimens from heterozygous females showed greater G6PD activity when stored as DBS, with 85% enzyme activity after 1 year of storage at -20 °C under controlled conditions in the presence of desiccant. G6PD deficient samples rapidly lost enzyme activity in all storage conditions tested. However, the reduction in G6PD enzyme activity in G6PD deficient samples did not interfere with G6PD classification. Samples stored under suitable conditions for G6PD testing will allow accurate measurement of enzyme activity, prevent misclassification of G6PD deficiency and enable safe and effective use of antimalarial drugs such as primaquine and tafenoquine.


Assuntos
Coleta de Amostras Sanguíneas/métodos , Deficiência de Glucosefosfato Desidrogenase/sangue , Deficiência de Glucosefosfato Desidrogenase/diagnóstico , Programas de Rastreamento/métodos , Sais de Tetrazólio , Feminino , Deficiência de Glucosefosfato Desidrogenase/genética , Heterozigoto , Humanos , Masculino , Temperatura
16.
Biosci Biotechnol Biochem ; 85(3): 739-742, 2021 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-33624771

RESUMO

The applicability of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay to an industrially valuable filamentous cyanobacterium Arthrospira platensis was examined. When it was applied to A. platensis NIES-39, as few as 10 viable trichomes were quantitatively detected. However, depending on the experimental conditions, it also generated artifactual viability signals. The results should help clarify the scope and limits of the MTT assay in viability analysis.


Assuntos
Colorimetria/métodos , Spirulina/química , Spirulina/metabolismo , Bioensaio , Sais de Tetrazólio , Tiazóis
17.
Biomolecules ; 11(2)2021 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-33573343

RESUMO

Green synthesis of metal nanoparticles using plant extracts as capping and reducing agents for the biomedical applications has received considerable attention. Moreover, emergence and spread of multidrug resistance among bacterial pathogens has become a major health concern and lookout for novel alternative effective drugs has gained momentum. In current study, we synthesized gold nanoparticles using the seed extract of Trachyspermum ammi (TA-AuNPs), assessed its efficacy against drug resistant biofilms of Listeria monocytogenes and Serratia marcescens, and evaluated its anticancer potential against HepG2 cancer cell lines. Microwave-assisted green synthesis of gold nanoparticles was carried out and characterization was done using UV-vis spectroscopy, X-ray diffraction (XRD), transmission electron microscopy (TEM), and dynamic light scattering (DLS). Most nanoparticles were observed as spherical and spheroidal with few anisotropies with an average crystalline size of 16.63 nm. Synthesized TA-AuNPs demonstrated significant biofilm inhibitory activity against L. monocytogenes (73%) as well as S. marcescens (81%). Exopolysaccharide (EPS), motility, and CSH, key elements that facilitate the formation and maintenance of biofilm were also inhibited significantly at the tested sub-minimum inhibitory concentrations (sub-MICs). Further, TA-AuNPs effectively obliterated preformed mature biofilms of S. marcescens and L. monocytogenes by 64% and 58%, respectively. Induction of intracellular ROS production in TA-AuNPs treated bacterial cells could be the plausible mechanism for the reduced biofilm formation in test pathogens. Administration of TA-AuNPs resulted in the arrest of cellular proliferation in a concentration-dependent manner. TA-AuNPs decrease the intracellular GSH in HepG2 cancer cell lines, cells become more prone to ROS generation, hence induce apoptosis. Thus, this work proposes a new eco-friendly and rapid approach for fabricating NPs which can be exploited for multifarious biomedical applications.


Assuntos
Antineoplásicos/farmacologia , Apiaceae/metabolismo , Ouro/química , Nanopartículas Metálicas/química , Espécies Reativas de Oxigênio , Sementes/metabolismo , Anisotropia , Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Sobrevivência Celular , Glutationa Transferase/metabolismo , Química Verde , Células Hep G2 , Humanos , Luz , Peroxidação de Lipídeos , Listeria monocytogenes/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Transmissão , Micro-Ondas , Extratos Vegetais/farmacologia , Polissacarídeos Bacterianos/química , Espalhamento de Radiação , Serratia marcescens/efeitos dos fármacos , Sais de Tetrazólio/química , Tiazóis/química , Difração de Raios X
18.
Carbohydr Polym ; 258: 117659, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33593545

RESUMO

Chemically modified biopolymers derived nanomaterials have shown great potential in drug delivery and live-cell imaging. We have developed two materials, doxorubicin-loaded chitosan-gold nanoparticles and beads, both embedded with functionalized silk fibroin. Nanoparticles with size 8 ± 3 nm were synthesized using chitosan as reducing and stabilizing agent. Beads with 900-1000 µm size were formulated by the ionic gelation technique. Both the materials were coated with functionalized silk fibroin for targeted and sustained drug release properties. The coated materials showed retarded drug release compared to the uncoated ones. The cytotoxicity was assessed in HeLa cell lines, which demonstrated a maximum dose-dependent decrease in cell viability for the cells treated with folate conjugated silk fibroin coated nanoparticles. The live-cell imaging of the nanoparticles unveiled the increased cellular uptake of the coated materials by seven folds than the uncoated ones. Thus, functionalized silk coated materials can be effective drug delivery tools for targeted and sustained drug release.


Assuntos
Antineoplásicos/farmacologia , Quitosana/química , Ouro/química , Nanopartículas Metálicas/química , Microesferas , Seda/metabolismo , Sobrevivência Celular , Preparações de Ação Retardada/química , Doxorrubicina/farmacocinética , Portadores de Fármacos , Sistemas de Liberação de Medicamentos , Liberação Controlada de Fármacos , Fibroínas/química , Fluoresceína , Ácido Fólico/química , Química Verde , Células HeLa , Humanos , Hidrogéis/química , Microscopia de Fluorescência , Peso Molecular , Tamanho da Partícula , Espectroscopia de Infravermelho com Transformada de Fourier , Propriedades de Superfície , Sais de Tetrazólio/farmacologia , Tiazóis/farmacologia
19.
J Biosci Bioeng ; 131(4): 396-404, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33386278

RESUMO

2,3,5-Triphenyl tetrazolium chloride (TTC) staining is a method to distinguish the mitochondrial activity of cells based on the color: colorless TTC turns red when under reducing conditions. Although the assay reflects the mitochondrial activity of cells, which enzyme(s) in the electron transport system contribute to TTC reduction has been unclear. TTC staining assays using gene disruptants related to the electron transport system in Saccharomyces cerevisiae revealed those disruptants related to electron transport from each electron donor to ubiquinone (red colonies) and disruptants that were related to ubiquinol-cytochrome c oxidoreductase and cytochrome c oxidase (white colonies). In addition, when the enzyme activities of ubiquinol-cytochrome c oxidoreductase and cytochrome c oxidase were measured using TTC as the electron acceptor, only ubiquinol-cytochrome c oxidoreductase showed TTC reduction activity, and the activity was enhanced by potassium cyanide, an inhibitor of cytochrome c oxidase. These results indicated that ubiquinol-cytochrome c oxidoreductase is involved in TTC reduction in S. cerevisiae. The fermentation profiles of BY4741UΔcor1 and BY4741UΔcox4, which exhibited no TTC staining activity, were almost identical to that of the parental strain BY4741U. However, cell growth and ethanol and succinate production of the ura3-mutated strain BY4741, which also exhibited no TTC staining activity, was altered compared to those of BY4741U, indicating that the fermentation profile varies among strains that show no TTC staining activity. The relationship between uracil metabolism and TTC staining activity was also determined based on metabolome analysis.


Assuntos
Fermentação , Saccharomyces cerevisiae/metabolismo , Sais de Tetrazólio/química , Transporte de Elétrons , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Coloração e Rotulagem , Ubiquinona/análogos & derivados , Ubiquinona/metabolismo
20.
Mol Cell Biochem ; 476(3): 1455-1465, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33389496

RESUMO

Melanoma ranks second in aggressive tumors, and the occurrence of metastasis in melanoma results in a persistent drop in the survival rate of patients. Therefore, it is very necessary to find a novel therapeutic method for treating melanoma. It has been reported that lncRNA XIST could promote the tumorigenesis of melanoma. However, the mechanism by which lncRNA XIST regulates the progression of melanoma remains unclear. The proliferation of A375 cells was measured by clonal formation. Cell viability was detected by MTT assay. Flow cytometry was performed to detect cell apoptosis and cycle. The level of GINS2, miR-23a-3p, and lncRNA XIST was investigated by qRT-PCR. Protein level was detected by Western blot, and the correctness of prediction results was confirmed by Dual luciferase. In present study, GINS2 and lncRNA XIST were overexpressed in melanoma, while miR-23a-3p was downregulated. Silencing of GINS2 or overexpression of miR-23a-3p reversed cell growth and promoted apoptosis in A375 cells. Mechanically, miR-23a-3p directly targeted GINS2, and XIST regulated GINS2 level though mediated miR-23a-3p. Moreover, XIST exerted its function on cell proliferation, cell viability, and promoted the cell apoptosis of A375 cells though miR-23a-3p/GINS2 axis. LncRNA XIST significantly promoted the tumorigenesis of melanoma via sponging miR-23a-3p and indirectly targeting GINS2, which can be a potential new target for treating melanoma.


Assuntos
Apoptose , Proteínas Cromossômicas não Histona/biossíntese , Regulação Neoplásica da Expressão Gênica , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Regulação para Baixo , Regulação da Expressão Gênica , Inativação Gênica , Células HEK293 , Humanos , Melanócitos/metabolismo , Melanoma/metabolismo , MicroRNAs/genética , Transdução de Sinais , Sais de Tetrazólio/farmacologia , Tiazóis/farmacologia
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