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1.
J Vis Exp ; (207)2024 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-38856211

RESUMO

Changes in calcium concentration in cells are rapidly monitored in a high-throughput fashion with the use of intracellular, fluorescent, calcium-binding dyes and imaging instruments that can measure fluorescent emissions from up to 1,536 wells simultaneously. However, these instruments are much more expensive and can be challenging to maintain relative to widely available plate readers that scan wells individually. Described here is an optimized plate reader assay for use with an endothelial cell line (EA.hy926) to measure the protease-activated receptor (PAR)-driven activation of Gαq signaling and subsequent calcium mobilization using the calcium-binding dye Fluo-4. This assay has been used to characterize a range of PAR ligands, including the allosteric PAR1-targeting anti-inflammatory "parmodulin" ligands identified in the Dockendorff lab. This protocol obviates the need for an automated liquid handler and permits the medium-throughput screening of PAR ligands in 96-well plates and should be applicable to the study of other receptors that initiate calcium mobilization.


Assuntos
Cálcio , Humanos , Cálcio/metabolismo , Cálcio/análise , Xantenos/química , Compostos de Anilina/química , Linhagem Celular , Corantes Fluorescentes/química , Ligantes , Receptor PAR-1/metabolismo , Células Endoteliais/metabolismo , Sinalização do Cálcio/fisiologia
2.
Eur J Med Chem ; 272: 116474, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38735149

RESUMO

Small molecule photosensitizers for combined in vivo tailored cancer diagnostics and photodynamic/photothermal therapy are desperately needed. Monoamine oxidase A (MAO-A)-activated therapeutic and diagnostic compounds provide great selectivity because MAO-A can be employed as a biomarker for associated Tumors. In order to screen photosensitizers with photodynamic therapeutic potential, we have created a range of near-infrared fluorescent molecules in this work by combining dihydroxanthene parent with various heterocyclic fluorescent dyes. The NIR fluorescent diagnostic probe, DHMQ, was created by combining the screened fluorescent dye matrices with the propylamino group, which is the recognition moiety of MAO-A, based on the oxidative deamination mechanism of the enzyme. This probe has a low toxicity level and can identify MAO-A precisely. It has the ability to use fluorescence imaging on mice and cells to track MAO-A activity in real-time. It has strong phototoxicity and can produce singlet oxygen when exposed to laser light. The temperature used in photothermal imaging can get up to 50 °C, which can harm tumor cells permanently and have a positive phototherapeutic impact on tumors grown from SH-SY5Y xenograft mice. The concept of using MAO-A effectively in diseases is expanded by the MAO-A-activated diagnostic-integrated photosensitizers, which offer a new platform for in vivo cancer diagnostics and targeted anticancer treatment.


Assuntos
Monoaminoxidase , Fotoquimioterapia , Fármacos Fotossensibilizantes , Terapia Fototérmica , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/síntese química , Animais , Humanos , Monoaminoxidase/metabolismo , Camundongos , Xantenos/química , Xantenos/farmacologia , Xantenos/síntese química , Estrutura Molecular , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Relação Estrutura-Atividade , Corantes Fluorescentes/química , Corantes Fluorescentes/síntese química , Corantes Fluorescentes/farmacologia , Proliferação de Células/efeitos dos fármacos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Camundongos Nus
3.
Analyst ; 149(12): 3372-3379, 2024 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-38712551

RESUMO

A series of novel near-infrared (NIR) xanthene-chalcone fluorophores were constructed through a modular synthesis with the electron-donating xanthene moiety and the electron-withdrawing chalcone moiety. These fluorophores are convenient for fluorescence imaging in living cells, benefiting from their NIR emissions (650-710 nm), large Stokes shifts (>100 nm), moderate quantum yields and low cytotoxicity. The substituted hydroxyl group of the xanthene-chalcone fluorophore HCA-E facilitates the development of multifunctional fluorescent probes. As an example, a highly sensitive and selective probe N-HCA-E for glutathione (GSH) detection was developed based on the fluorophore HCA-E. A 4-nitrobenzenesulfonyl (4-Ns) group was introduced to cage the hydroxyl group of HCA-E, which was used as a selective recognition site for the thiol of GSH and an effective fluorescence quencher. Probe N-HCA-E revealed NIR "turn-on" fluorescence (709 nm) for endogenous and exogenous GSH detection in lysosomes with a large Stokes shift (129 nm) and high anti-interference ability.


Assuntos
Corantes Fluorescentes , Glutationa , Imagem Óptica , Xantenos , Corantes Fluorescentes/química , Corantes Fluorescentes/toxicidade , Corantes Fluorescentes/síntese química , Xantenos/química , Humanos , Glutationa/química , Imagem Óptica/métodos , Chalconas/química , Células HeLa , Lisossomos/química , Lisossomos/metabolismo , Raios Infravermelhos , Chalcona/química
4.
J Dent ; 145: 104997, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38621525

RESUMO

OBJECTIVE: To assess the effects of arginine, with or without sodium fluoride (NaF; 1,450 ppm), on saliva-derived microcosm biofilms and enamel demineralization. METHODS: Saliva-derived biofilms were grown on bovine enamel blocks in 0.2 % sucrose-containing modified McBain medium, according to six experimental groups: control (McBain 0.2 %); 2.5 % arginine; 8 % arginine; NaF; 2.5 % arginine with NaF; and 8 % arginine with NaF. After 5 days of growth, biofilm viability was assessed by colony-forming units counting, laser scanning confocal microscopy was used to determine biofilm vitality and extracellular polysaccharide (EPS) production, while biofilm metabolism was evaluated using the resazurin assay and lactic acid quantification. Demineralization was evaluated by measuring pH in the culture medium and calcium release. Data were analyzed by Kruskal-Wallis' and Dunn's tests (p < 0.05). RESULTS: 8 % arginine with NaF showed the strongest reduction in total streptococci and total microorganism counts, with no significant difference compared to arginine without NaF. Neither 2.5 % arginine alone nor NaF alone significantly reduced microbial counts compared to the control, although in combination, a reduction in all microbial groups was observed. Similar trends were found for biofilm vitality and EPS, and calcium released to the growth medium. CONCLUSIONS: 8 % Arginine, with or without NaF, exhibited the strongest antimicrobial activity and reduced enamel calcium loss. Also, NaF enhanced the effects of 2.5 % arginine, yielding similar results to 8 % arginine for most parameters analyzed. CLINICAL SIGNIFICANCE: The results provided further evidence on how arginine, with or without NaF, affects oral microcosm biofilms and enamel mineral loss.


Assuntos
Arginina , Biofilmes , Cariostáticos , Esmalte Dentário , Microscopia Confocal , Saliva , Fluoreto de Sódio , Desmineralização do Dente , Biofilmes/efeitos dos fármacos , Arginina/farmacologia , Fluoreto de Sódio/farmacologia , Esmalte Dentário/efeitos dos fármacos , Esmalte Dentário/microbiologia , Bovinos , Animais , Desmineralização do Dente/prevenção & controle , Desmineralização do Dente/microbiologia , Cariostáticos/farmacologia , Saliva/microbiologia , Saliva/metabolismo , Saliva/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Viabilidade Microbiana/efeitos dos fármacos , Cálcio/análise , Cálcio/metabolismo , Streptococcus/efeitos dos fármacos , Xantenos/farmacologia , Contagem de Colônia Microbiana , Oxazinas/farmacologia
5.
Actas Esp Psiquiatr ; 52(2): 83-98, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38622006

RESUMO

BACKGROUND: Vascular dementia (VaD) is a prevalent neurodegenerative disease characterized by cognitive impairment due to cerebrovascular factors, affecting a significant portion of the aging population and highlighting the critical need to understand specific targets and mechanisms for effective prevention and treatment strategies. We aimed to identify pathways and crucial genes involved in the progression of VaD through bioinformatics analysis and subsequently validate these findings. METHODS: We conducted differential expression analysis, Weighted Gene Co-expression Network Analysis (WGCNA), Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and Protein-Protein Interaction (PPI) analysis. We utilized pheochromocytoma 12 (PC12) cells to create an in vitro oxygen-glucose deprivation (OGD) model. We investigated the impact of overexpression and interference of adrenoceptor alpha 1D (ADRA1D) on OGD PC12 cells using TdT-mediated dUTP nick-end labeling (TUNEL), reverse transcription-quantitative polymerase chain reaction (RT-qPCR), western blot (WB), and Fluo-3-pentaacetoxymethyl ester (Fluo-3 AM) analysis. RESULTS: We found 187 differentially expressed genes (DEGs) in the red module that were strongly associated with VaD and were primarily enriched in vasoconstriction, G protein-coupled amine receptor activity, and neuroactive ligand-receptor interaction, mitogen-activated protein kinase (MAPK) signaling pathway, and cell adhesion. Among these pathways, we identified ADRA1D as a gene shared by vasoconstriction, G protein-coupled amine receptor activity, and neuroactive ligand-receptor interaction. The TUNEL assay revealed a significant decrease in PC12 cell apoptosis with ADRA1D overexpression (p < 0.01) and a significant increase in apoptosis upon silencing ADRA1D (p < 0.01). RT-qPCR and WB analysis revealed elevated ADRA1D expression (p < 0.001) and decreased phospholipase C beta (PLCß) and inositol 1,4,5-trisphosphate receptor (IP3R) expression (p < 0.05) with ADRA1D overexpression. Moreover, the Fluo-3 AM assessment indicated significantly lower intracellular Ca2+ levels with ADRA1D overexpression (p < 0.001). Conversely, interference with ADRA1D yielded opposite results. CONCLUSION: Our study provides a new perspective on the pathogenic mechanisms of VaD and potential avenues for therapeutic intervention. The results highlight the role of ADRA1D in modulating cellular responses to OGD and VaD, suggesting its potential as a target for VaD treatment.


Assuntos
Compostos de Anilina , Demência Vascular , Doenças Neurodegenerativas , Xantenos , Animais , Ratos , Humanos , Idoso , Demência Vascular/genética , Ligantes , Aminas , Transdução de Sinais/genética , Proteínas de Ligação ao GTP
6.
Biosensors (Basel) ; 14(4)2024 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-38667149

RESUMO

The resazurin assay, also known as the Alamar Blue assay, stands as a cornerstone technique in cell biology, microbiology, and drug development. It assesses the viability of cells through the conversion of resazurin into highly fluorescent resorufin. The resulting fluorescence intensity provides a reliable estimate of viable cell numbers. Cytotoxicity assays, such as the resazurin-based method, play a crucial role in the screening of potential drug candidates and in the assessment of pharmaceutical and chemical toxicity. In recent years, inconsistencies have arisen in pharmacogenomic studies, often due to poorly optimized laboratory protocols. These inconsistencies hinder progress in understanding how substances affect cell health, leading to unreliable findings. Thus, the need for standardized and rigorously optimized protocols is evident to ensure consistent and accurate results in cytotoxicity studies. This manuscript describes a standardized procedure for optimizing resazurin-based viability assays to improve the reliability of cytotoxicity data. This optimization approach focuses on critical experimental parameters and data quality, aiming to achieve a level of measurement imprecision of less than 20%. In conclusion, to address the critical issues of reproducibility and reliability, protocol standardization, such as the one described in this manuscript, can greatly enhance the credibility of cytotoxicity studies, ultimately advancing drug safety assessments.


Assuntos
Sobrevivência Celular , Oxazinas , Xantenos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Bioensaio/métodos , Reprodutibilidade dos Testes
7.
Spectrochim Acta A Mol Biomol Spectrosc ; 314: 124180, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38522378

RESUMO

N2H4 is a common raw material used in the production of pesticides and has good water solubility, so it may contaminate water sources and eventually enter living organisms, causing serious health problems. Viscosity is an important indicator of the cellular microenvironment and an early warning signal for many diseases. The high reactivity of hydrazine depletes glutathione (GSH) in hepatocytes, causing oxidative stress ultimately leading to significant changes in intracellular viscosity and even death. Therefore, it is particularly important to develop an effective method to detect N2H4 and viscosity in environmental and biological systems. On this basis, we developed two fluorescent probes, BDD and BHD, based on xanthene and 2-benzothiazole acetonitrile. The experimental results show that BHD and BDD have good imaging capabilities for N2H4 in cells, zebrafish and Arabidopsis. BHD and BDD also showed sensitive detection and fluorescence enhancement in the near-infrared region when the intracellular viscosity was changed. Notably, the probe BDD has also successfully imaged N2H4 in a variety of real water samples.


Assuntos
Corantes Fluorescentes , Peixe-Zebra , Animais , Humanos , Viscosidade , Xantenos , Água , Hidrazinas , Células HeLa , Espectrometria de Fluorescência
8.
Diagn Microbiol Infect Dis ; 109(2): 116236, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38537506

RESUMO

We proposed a new methodology, the microelution ATM/CZA (mATM/CZA), based on the antibiotic disc elution and the use of resazurin, for rapid (<4h) determination of in vitro susceptibility to aztreonam combined with ceftazidime-avibactam among Enterobacterales. The mATM/CZA presented excellent accuracy with 1.9 %, 98.1 % and 100 % of major error, specificity and sensitivity, respectively. Furthermore, we assessed synergism between aztreonam and ceftazidime-avibactam in Enterobacterales and Pseudomonas aeruginosa, which was observed in 37/55 Enterobacterales and 31/56 P. aeruginosa. As reference methodologies (checkerboard, time-kill curve) are not compatible with the routine of the clinical microbiology laboratories, mATM/CZA is an important alternative to evaluate susceptibility of the combination in a scenario where its clinical use is increasingly important.


Assuntos
Antibacterianos , Compostos Azabicíclicos , Aztreonam , Ceftazidima , Combinação de Medicamentos , Sinergismo Farmacológico , Testes de Sensibilidade Microbiana , Aztreonam/farmacologia , Compostos Azabicíclicos/farmacologia , Testes de Sensibilidade Microbiana/métodos , Testes de Sensibilidade Microbiana/normas , Antibacterianos/farmacologia , Ceftazidima/farmacologia , Humanos , Pseudomonas aeruginosa/efeitos dos fármacos , Enterobacteriaceae/efeitos dos fármacos , Sensibilidade e Especificidade , Xantenos , Oxazinas
9.
Braz J Microbiol ; 55(2): 1349-1357, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38438831

RESUMO

Chromoblastomycosis is a fungal chronic disease, which affects humans, especially in cutaneous and subcutaneous tissues. There is no standard treatment for Chromoblastomycosis, and it is a therapeutic challenge, due natural resistance of their causative agents, inadequate response of patients and common cases of relapse. Protocols for determination of antifungal drugs susceptibility are not standardized for chromoblastomycosis agents and endpoint definition is usually based on visual inspection, which depends on the analyst, making it sometimes inaccurate. We presented a colorimetric and quantitative methodology based on resazurin reduction to resofurin to determine the metabolic status of viable cells of Fonsecaea sp. Performing antifungal susceptibility assay by a modified EUCAST protocol allied to resazurin, we validated the method to identify the minimum inhibitory concentrations of itraconazole, fluconazole, amphotericin B, and terbinafine for eight Fonsecaea clinical isolates. According to our data, resazurin is a good indicator of metabolic status of viable cells, including those exposed to antifungal drugs. This work aimed to test resazurin as an indicator of the metabolic activity of Fonsecaea species in susceptibility assays to antifungal drugs. Species of this genus are the main causative agents of Chromoblastomycosis, which affects humans.


Assuntos
Antifúngicos , Cromoblastomicose , Fonsecaea , Testes de Sensibilidade Microbiana , Oxazinas , Xantenos , Xantenos/metabolismo , Oxazinas/metabolismo , Antifúngicos/farmacologia , Humanos , Fonsecaea/efeitos dos fármacos , Fonsecaea/genética , Fonsecaea/metabolismo , Cromoblastomicose/microbiologia , Cromoblastomicose/tratamento farmacológico , Colorimetria/métodos
10.
Anal Chem ; 96(13): 5134-5142, 2024 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-38507805

RESUMO

Mitochondria are important organelles that provide energy for cellular physiological activities. Changes in their structures may indicate the occurrence of diseases, and the super-resolution imaging of mitochondria is of great significance. However, developing fluorescent probes for mitochondrial super-resolution visualization still remains challenging due to insufficient fluorescence brightness and poor stability. Herein, we rationally synthesized an ultrabright xanthene fluorescence probe Me-hNR for mitochondria-specific super-resolution imaging using structured illumination microscopy (SIM). The rigid structure of Me-hNR provided its ultrahigh fluorescence quantum yield of up to 0.92 and ultrahigh brightness of up to 16,000. Occupying the para-position of the O atom in the xanthene skeleton by utilizing the smallest methyl group ensured its excellent stability. The study of the photophysical process indicated that Me-hNR mainly emitted fluorescence via radiative decay, and nonradiative decay and inter-system crossing were rare due to the slow nonradiative decay rate and large energy gap (ΔEst = 0.55 eV). Owing to these excellent merits, Me-hNR can specifically light up mitochondria at ultralow concentrations down to 5 nM. The unprecedented spatial resolution for mitochondria with an fwhm of 174 nm was also achieved. Therefore, this ultrabright xanthene fluorescence probe has great potential in visualizing the structural changes of mitochondria and revealing the pathogenesis of related diseases using SIM.


Assuntos
Corantes Fluorescentes , Xantenos , Corantes Fluorescentes/química , Mitocôndrias , Organelas , Microscopia de Fluorescência/métodos
11.
Bioorg Chem ; 145: 107182, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38359707

RESUMO

Gambogenic acid (GNA), a caged xanthone derived from Garcinia hanburyi, exhibits a wide range of anti-cancer properties. The caged skeleton of GNA serves as the fundamental pharmacophore responsible for its antitumor effects. However, limited exploration has focused on the structural modifications of GNA. This study endeavors to diversify the structure of GNA and enhance its anti-cancer efficacy. Sulfoximines, recognized as pivotal motifs in medicinal chemistry due to their outstanding properties, have featured in several anti-cancer drugs undergoing clinical trials. Accordingly, a series of 33 GNA derivatives combined with sulfoximines were synthesized and evaluated for their anti-cancer effects against MIAPaCa2, MDA-MB-231, and A549 cells in vitro. The activity screening led to the identification of compound 12k, which exhibited the most potent anti-cancer effect. Mechanistic studies revealed that 12k primarily induced pyroptosis in MIAPaCa2 and MDA-MB-231 cells by activating the caspase-3/gasdermin E (GSDME) pathway. These findings suggested that 12k is a promising drug candidate in cancer therapy and highlighted the potential of sulfoximines as a valuable functional group in drug discovery.


Assuntos
Apoptose , Piroptose , Humanos , Xantenos/farmacologia , Xantenos/química , Células A549 , Linhagem Celular Tumoral
12.
Anal Chim Acta ; 1294: 342292, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38336413

RESUMO

BACKGROUND: Hypochlorous acid (HClO) is an important biomarker for inflammation, cardiovascular disease, and even cancer. It is of great significance to accurately monitor and quantitatively analyze the fluctuations of HClO to better understand their physiological functions. Traditional HClO detection methods such as high-performance liquid chromatography (HPLC), and mass spectrometry are preferred, but are costly and unsuitable in vivo. Near-infrared (NIR) fluorescence imaging has the advantages of high sensitivity, high temporal and spatial resolutions, minimal autofluorescence, and deep tissue penetration, which facilitates its application in biological systems. Therefore, the development of sensitivity and simple NIR fluorescence monitoring HClO methods in vivo and in vitro is essential and desirable. RESULTS: Herein, we present a NIR probe NOF3 by integrating the rhodamine scaffold and HClO-triggered moiety for the real-time detection of HClO in vitro and in vivo. NOF3 reacts with the HClO and releases the NOF-OH fluorophore of emitted signals at 730 nm, which is in the NIR region. The designed probe detected concentrations of HClO ranging from 0 to 17 µM with a low detection limit of 0.146 µM, presenting excellent sensitivity and selectivity toward HClO over other species. NOF3 manifests significantly turn-on NIR fluorescent signals in response to HClO concentration, which makes it favorable for monitoring dynamic HClO distribution in vivo. We exemplify NOF3 for the tracking of endogenously overexpressed HClO distribution in RAW 264.7 cells, and further realize real-time in vivo bioimaging of HClO activity in inflammation mice. SIGNIFICANCE: The facile NIR NOF3 probe was successfully applied to visualize endogenous and exogenous HClO in living cells and mice. This study provides not only an effective tool for spatial and temporal resolution HClO bioimaging in vivo but also possesses great potential for use in future research on HClO-related biology and pathology.


Assuntos
Ácido Hipocloroso , Xantenos , Camundongos , Animais , Ácido Hipocloroso/análise , Rodaminas/química , Corantes Fluorescentes/química , Inflamação/diagnóstico por imagem
13.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 55(1): 198-203, 2024 Jan 20.
Artigo em Chinês | MEDLINE | ID: mdl-38322510

RESUMO

Objective: To establish and evaluate a microbial sensitivity test method for Neisseria gonorrhoeae based on resazurin coloration. Methods: Based on the broth microdilution method, resazurin was added as a live bacteria indicator. WHO G, a WHO gonococcal reference strain, was used to optimize the incubation time for resazurin-stained bacteria and the color change was visually observed to obtain the results. Agar dilution method (the gold standard) and resazurin-based microdilution assay were used to determine the minimum inhibitory concentration (MIC) of azithromycin, ceftriaxone, and spectinomycin for 3 reference strains and 32 isolates of Neisseria gonorrhoeae. The results were analyzed based on essential agreement (EA), which reflected the consistency of the MIC values, category agreement (CA), which reflected the consistency in the determination of drug resistance, intermediary, and sensitivity, very major error (VME), which reflected false sensitivity, and major error (ME), which reflected pseudo drug resistance, to evaluate the accuracy of resazurin-based microdilution assay as a microbial sensitivity test of of Neisseria gonorrhoeae. CA and EA rates≥90% and VME and ME rates≤3% were found to be the acceptable performance rates. Results: The results obtained 6 hours after resazurin was added were consistent with those of the agar dilution method and the resazurin-based microdilution assay was established accordingly based on this parameter. The EA of resazurin-based microdilution assay for measuring the MIC results of azithromycin, ceftriaxone, and spectinomycin was 97.1%, 91.5%, and 94.3%, respectively, and the CA was 88.6%, 94.3%, and 94.3%, respectively. The VME was 0% for all three antibiotics, while the ME was 11.4%, 5.7%, and 5.7%, respectively. Conclusion: The resazurin-based microdilution assay established in this study showed good agreement with agar dilution method for measuring the MIC of antibiotics against Neisseria gonorrhoeae. Moreover, the sensitivity results of this method were highly reliable and could be easily obtained through naked eye observation. Nonetheless, the results of drug resistance should be treated with caution and the optimization of parameters should be continued.


Assuntos
Azitromicina , Neisseria gonorrhoeae , Oxazinas , Xantenos , Azitromicina/farmacologia , Ceftriaxona/farmacologia , Espectinomicina , Ágar , Antibacterianos/uso terapêutico , Testes de Sensibilidade Microbiana , Farmacorresistência Bacteriana
14.
Anal Methods ; 16(9): 1409-1414, 2024 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-38369924

RESUMO

Peroxynitrite (ONOO-) is a critical ROS in living systems, and could induce lipid peroxidation which is the driver of ferroptotic cell death. Therefore, precise and rapid detection of cellular ONOO- is critical for the deep study of the biological functions of ONOO- during ferroptosis. Herein, we developed fluorescent probes (Rh-1, Rh-2 and Rh-3) for the rapid detection of intracellular ONOO- during ferroptosis. These probes used bishydrazide groups as the reactive sites for ONOO-. The response of these probes to ONOO- resulted in the production of the emissive xanthene fluorophore, providing a marked enhancement in the fluorescence intensity at 561 nm. The probe Rh-3 exhibited prominent selectivity and sensitivity towards ONOO-. Bioimaging experiments suggested that Rh-3 could be applied to image exogenous and endogenous ONOO- in living cells. By fluorescence imaging, it was demonstrated that erastin-induced ferroptosis caused increased levels of the endogenous ONOO-, and ferrostatin-1 (Fer-1) and vitamin E (VE) could markedly inhibit the excessive production of ONOO- during ferroptosis in living cells.


Assuntos
Ferroptose , Corantes Fluorescentes , Corantes Fluorescentes/química , Ácido Peroxinitroso/química , Ácido Peroxinitroso/metabolismo , Imagem Óptica , Xantenos
15.
Planta Med ; 90(5): 353-367, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38295847

RESUMO

Gambogenic acid is a derivative of gambogic acid, a polyprenylated xanthone isolated from Garcinia hanburyi. Compared with the more widely studied gambogic acid, gambogenic acid has demonstrated advantages such as a more potent antitumor effect and less systemic toxicity than gambogic acid according to early investigations. Therefore, the present review summarizes the effectiveness and mechanisms of gambogenic acid in different cancers and highlights the mechanisms of action. In addition, drug delivery systems to improve the bioavailability of gambogenic acid and its pharmacokinetic profile are included. Gambogenic acid has been applied to treat a wide range of cancers, such as lung, liver, colorectal, breast, gastric, bladder, and prostate cancers. Gambogenic acid exerts its antitumor effects as a novel class of enhancer of zeste homolog 2 inhibitors. It prevents cancer cell proliferation by inducing apoptosis, ferroptosis, and necroptosis and controlling the cell cycle as well as autophagy. Gambogenic acid also hinders tumor cell invasion and metastasis by downregulating metastasis-related proteins. Moreover, gambogenic acid increases the sensitivity of cancer cells to chemotherapy and has shown effects on multidrug resistance in malignancy. This review adds insights for the prevention and treatment of cancers using gambogenic acid.


Assuntos
Antineoplásicos , Xantenos , Animais , Apoptose , Linhagem Celular Tumoral , Xantenos/farmacologia , Xantenos/uso terapêutico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico
16.
Reprod Domest Anim ; 59(1): e14534, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38268217

RESUMO

The present work was designed for a thorough investigation into the sperm morphology and morphometry of Kurdish stallions. The semen samples were collected from 10 Kurdish stallions. Three preparations from each ejaculate were stained with eosin-nigrosin (EN), Diff-Quik (DQ) and Rose Bengal (RB). The area, perimeter, length and width of the sperm head as well as tail length and total sperm length were measured. The parameters ellipticity, elongation, roughness and regularity were calculated. The morphology of sperm was also investigated under scanning and transmission electron microscopes. DQ and RB provided more clarified images for examining sperm structures compared to the EN method. The head length, head width, area and perimeter in EN were significantly higher than those in DQ and RB (p ≤ .05). Furthermore, the difference in head width, head area and head perimeter between DQ and RB was not significant (p ≥ .05). The tail length and total sperm length in all methods were close together (p ≥ .05). The highest percentage of normal sperm was seen in DQ and RB methods (82.55 ± 2.88 and 88.31 ± 5.19) respectively. The highest values for ellipticity, elongation and regularity were found in RB, whereas the highest value for roughness was measured in EN. Tail defects including coiled tails, and folded midpieces were the most frequent. Scanning electron microscope revealed two types of head shapes: heads with round anterior border, and heads with flat anterior border. The results indicated that despite the routine use of EN for morphological assessment of stallion sperm, RB and DQ can be considered for more clarified details of sperm structure including acrosome and midpiece. Furthermore, the Kurdish stallion sperm has morphometric traits in the normal range established for stallions; yet, some traits were larger than those reported for other breeds. It seems that the sperm of the Kurdish stallion has a longer head and tail in comparison with other horse breeds.


Assuntos
Compostos de Anilina , Corantes Azur , Azul de Metileno , Sêmen , Espermatozoides , Xantenos , Masculino , Cavalos , Animais , Irã (Geográfico) , Acrossomo , Cabeça do Espermatozoide , Amarelo de Eosina-(YS) , Rosa Bengala
17.
Chemistry ; 30(1): e202303038, 2024 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-37852935

RESUMO

Photoacoustic imaging (PAI) is an emerging imaging technique that uses pulsed laser excitation with near-infrared (NIR) light to elicit local temperature increases through non-radiative relaxation events, ultimately leading to the production of ultrasound waves. The classical xanthene dye scaffold has found numerous applications in fluorescence imaging, however, xanthenes are rarely utilized for PAI since they do not typically display NIR absorbance. Herein, we report the ability of Nebraska Red (NR) xanthene dyes to produce photoacoustic (PA) signal and provide a rational design approach to reduce the hydrolysis rate of ester containing dyes, affording cell permeable probes. To demonstrate the utility of this approach, we construct the first cell permeable rhodamine-based, turn-on PAI imaging probe for hypochlorous acid (HOCl) with maximal absorbance within the range of commercial PA instrumentation. This probe, termed SNR700 -HOCl, is capable of detecting exogenous HOCl in mice. This work provides a new set of rhodamine-based PAI agents as well as a rational design approach to stabilize esterified versions of NR dyes with desirable properties for PAI. In the long term, the reagents described herein could be utilized to enable non-invasive imaging of HOCl in disease-relevant model systems.


Assuntos
Corantes Fluorescentes , Técnicas Fotoacústicas , Animais , Camundongos , Rodaminas , Ésteres , Técnicas Fotoacústicas/métodos , Xantenos , Imagem Óptica/métodos
18.
Chemistry ; 30(12): e202303208, 2024 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-38038726

RESUMO

Fluorophores are considered powerful tools for not only enabling the visualization of cell structures, substructures, and biological processes, but also making for the quantitative and qualitative measurement of various analytes in living systems. However, most fluorophores do not meet the diverse requirements for biological applications in terms of their photophysical and biological properties. Hybridization is an important strategy in molecular engineering that provides fluorophores with complementarity and multifunctionality. This review summarizes the basic strategies of hybridization with four classes of fluorophores, including xanthene, cyanine, coumarin, and BODIPY with a focus on their structure-property relationship (SPR) and biological applications. This review aims to provide rational hybrid ideas for expanding the reservoir of knowledge regarding fluorophores and promoting the development of newly produced fluorophores for applications in the field of life sciences.


Assuntos
Corantes Fluorescentes , Xantenos , Corantes Fluorescentes/química , Xantenos/química , Ionóforos
19.
Spectrochim Acta A Mol Biomol Spectrosc ; 305: 123522, 2024 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-37852120

RESUMO

In this paper, an Hg2+ detection probe, HOS, was prepared with a xanthene as the parent fluorophore. Hg2+-initiated thioacetal deprotection reaction is the detection mechanism of this probe. After testing, the probe HOS was able to accurately determine Hg2+ with a detection limit of 36 nM. It was successfully applied to the detection of Hg2+ in different water samples and shrimp samples, meanwhile, the filter paper strips prepared by HOS were obviously changed from light yellow to dark yellow under daylight, and from green to yellow under 365 nm UV light. Furthermore, probe HOS enabled Hg2+ bioimaging experiments on HepG2 cells, zebrafish and tobacco seedlings under laser confocal microscopy.


Assuntos
Mercúrio , Água , Animais , Corantes Fluorescentes , Peixe-Zebra , Nicotiana , Plântula , Xantenos , Imagem Óptica/métodos , Espectrometria de Fluorescência/métodos
20.
Parasit Vectors ; 16(1): 243, 2023 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-37468906

RESUMO

BACKGROUND: Helminth infections are an important public health problem in humans and have an even greater impact on domestic animal and livestock welfare. Current readouts for anthelmintic drug screening assays are stage development, migration, or motility that can be subjective, laborious, and low in throughput. The aim of this study was to apply and optimize a fluorometric technique using resazurin for evaluating changes in the metabolic activity of Ascaris suum third-stage larvae (L3), a parasite of high economic relevance in swine. METHODS: Ascaris suum L3 were mechanically hatched from 6- to 8-week embryonated and sucrose-gradient-enriched eggs. Resazurin dye and A. suum L3 were titrated in 96-well microtiter plates, and resazurin reduction activity was assessed by fluorometry after 24 h of incubation. Fluorescence microscopy was used to localize the resazurin reduction site within the larvae. Finally, we exposed A. suum L3 to various stress conditions including heat, methanol, and anthelmintics, and investigated their impact on larval metabolism through resazurin reduction activity. RESULTS: We show that the non-fluorescent dye resazurin is reduced inside vital A. suum L3 to fluorescent resorufin and released into the culture media. Optimal assay parameters are 100-1000 L3 per well, a resazurin concentration of 7.5 µg/ml, and incubation at 37 °C/5% CO2 for 24 h. An intact L2 sheath around the L3 of A. suum completely prevents the uptake of resazurin, while in unsheathed L3, the most intense fluorescence signal is observed along the larval midgut. L3 exposed to methanol or heat show a gradually decreased resazurin reduction activity. In addition, 24 h exposure to ivermectin at 0.625 µM, mebendazole at 5 µM, and thiabendazole from 10 to 100 µM significantly decreased larval metabolic activity by 55%, 73%, and 70% to 89%, respectively. CONCLUSIONS: Together, our results show that both metabolic stressors and anthelmintic drugs significantly and reproducibly reduce the resazurin reduction activity of A. suum L3, making the proposed assay a sensitive and easy-to-use method to evaluate metabolic activity of A. suum L3 in vitro.


Assuntos
Anti-Helmínticos , Ascaríase , Ascaris suum , Humanos , Animais , Suínos , Metanol/farmacologia , Metanol/uso terapêutico , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Xantenos/farmacologia , Xantenos/uso terapêutico , Ascaríase/parasitologia , Larva
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