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1.
Chemosphere ; 364: 142988, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39103097

RESUMO

Increased awareness of the impact of human activities on the environment has emerged in recent decades. One significant global environmental and human health issue is the development of materials that could potentially have negative effects. These materials can accumulate in the environment, infiltrate organisms, and move up the food chain, causing toxic effects at various levels. Therefore, it is crucial to assess materials comprising nano-scale particles due to the rapid expansion of nanotechnology. The aquatic environment, particularly vulnerable to waste pollution, demands attention. This review provides an overview of the behavior and fate of metallic nanoparticles (NPs) in the aquatic environment. It focuses on recent studies investigating the toxicity of different metallic NPs on aquatic organisms, with a specific emphasis on thiol-biomarkers of oxidative stress such as glutathione, thiol- and related-enzymes, and metallothionein. Additionally, the selection of suitable measurement methods for monitoring thiol-biomarkers in NPs' ecotoxicity assessments is discussed. The review also describes the analytical techniques employed for determining levels of oxidative stress biomarkers.


Assuntos
Antioxidantes , Biomarcadores , Nanopartículas Metálicas , Estresse Oxidativo , Poluentes Químicos da Água , Nanopartículas Metálicas/toxicidade , Nanopartículas Metálicas/química , Biomarcadores/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Poluentes Químicos da Água/análise , Antioxidantes/metabolismo , Peptídeos/toxicidade , Animais , Metalotioneína/metabolismo , Monitoramento Ambiental/métodos , Organismos Aquáticos/efeitos dos fármacos , Glutationa/metabolismo , Humanos
2.
Heliyon ; 9(11): e21497, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-38027737

RESUMO

The Kékfrankos is the most frequently cultivated wine grape in Hungary, with a significant national and regional impact, resulting in considerable amounts of byproducts (e.g. pomace, seeds). To the best of our knowledge no research has been conducted on the antioxidant and antibacterial properties of its seed extracts (GSE). A novel apporach of applying direct microwave treatment on grape seeds was implemented for the first time to enhance antioxidant and antimicrobial properties of GSE. Antioxidant properties were assayed using the DPPH (2,2-diphenyl-1-picrylhydrazyl), FRAP (Ferric Reducing Antioxidant Power) and TPC (Folin-Ciocâlteu's Total Polyphenol Content) methods. Profile and content of polyphenols was studied using high-performance liquid chromatography/tandem mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry. Antibacterial properties were evaluated using Gram-positive Staphylococcus aureus (SA), methicillin-resistant Staphylococcus aureus (ST239) (MRSA) and Gram-negative Escherichia coli (EC) bacteria strains. Results proved that the mild direct microwave treatment of grape seeds significantly increased total polyphenol, (+)-catechin, (-)-epicatechin as well as antioxidant capacity levels by 20-30 % compared to untreated samples and resulted the best antibacterial properties based on bacterial growth curves (SA and MRSA: 0.015625 mg/mL, EC: 0.25 mg/mL). Results justify the importance of further pharmacological investigations on Kékfrankos grape seed extracts and that the direct microwave treatment of grape seeds is an innovative approach for the fast and cost efficient improvement of the antibacterial properties of grape seed extracts.

3.
Res Vet Sci ; 152: 1-9, 2022 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-35901636

RESUMO

Distinct monocyte subpopulations have been previously described in healthy pigs and pigs experimentally infected with Actinobacillus pleuropneumoniae (APP). The CD163+ subpopulation of bone marrow (BM), peripheral blood (PB) and lung monocytes was found to play an important role in the inflammatory process. The inflammation is accompanied by elevation of inflammatory cytokines. The aim of the study was to evaluate the contribution of CD163+ monocytes and macrophages to cytokine production during APP-induced lung inflammation. Cytokine production was assessed by flow cytometry (FC) and quantitative PCR (qPCR) in CD163+ monocytes and by qPCR, immunohistochemistry/fluorescence in lungs and tracheobronchial lymph nodes (TBLN). Despite the systemic inflammatory response after APP infection, BM and PB CD163+ monocytes did not express elevated levels of a wide range of cytokines compared to control pigs. In contrast, significant amounts of IL-1ß, IL-6, IL-8 and TNF-α were produced in lung lesions and IL-1ß in the TBLN. At the protein level, TNF-α was expressed by both CD163+ monocytes and macrophages in lung lesions, whereas IL-1ß, IL-6 and IL-8 expression was found only in CD163+ monocytes; no CD163+ macrophages were found to produce these cytokines. Furthermore, the quantification of CD163+ monocytes expressing the two cytokines IL-1ß and IL-8 that were most elevated was performed. In lung lesions, 36.5% IL-1ß positive CD163+ monocytes but only 18.3% IL-8 positive CD163+ monocytes were found. In conclusion, PB and BM CD163+ monocytes do not appear to contribute to the elevated cytokine levels in plasma. On the other hand, CD163+ monocytes contribute to inflammatory cytokine expression, especially IL-1ß at the site of inflammation during the inflammatory process.


Assuntos
Infecções por Actinobacillus , Actinobacillus pleuropneumoniae , Suínos , Animais , Actinobacillus pleuropneumoniae/fisiologia , Monócitos/metabolismo , Citocinas , Fator de Necrose Tumoral alfa/metabolismo , Interleucina-8/metabolismo , Interleucina-6/metabolismo , Infecções por Actinobacillus/veterinária , Inflamação/metabolismo , Inflamação/veterinária
4.
J Am Chem Soc ; 143(40): 16486-16501, 2021 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-34477370

RESUMO

Mammalian metallothioneins (MTs) are a group of cysteine-rich proteins that bind metal ions in two α- and ß-domains and represent a major cellular Zn(II)/Cu(I) buffering system in the cell. At cellular free Zn(II) concentrations (10-11-10-9 M), MTs do not exist in fully loaded forms with seven Zn(II)-bound ions (Zn7MTs). Instead, MTs exist as partially metal-depleted species (Zn4-6MT) because their Zn(II) binding affinities are on the nano- to picomolar range comparable to the concentrations of cellular Zn(II). The mode of action of MTs remains poorly understood, and thus, the aim of this study is to characterize the mechanism of Zn(II) (un)binding to MTs, the thermodynamic properties of the Zn1-6MT2 species, and their mechanostability properties. To this end, native mass spectrometry (MS) and label-free quantitative bottom-up and top-down MS in combination with steered molecular dynamics simulations, well-tempered metadynamics (WT-MetaD), and parallel-bias WT-MetaD (amounting to 3.5 µs) were integrated to unravel the chemical coordination of Zn(II) in all Zn1-6MT2 species and to explain the differences in binding affinities of Zn(II) ions to MTs. Differences are found to be the result of the degree of water participation in MT (un)folding and the hyper-reactive character of Cys21 and Cys29 residues. The thermodynamics properties of Zn(II) (un)binding to MT2 are found to differ from those of Cd(II), justifying their distinctive roles. The potential of this integrated strategy in the investigation of numerous unexplored metalloproteins is attested by the results highlighted in the present study.


Assuntos
Metalotioneína
5.
Sci Rep ; 11(1): 5496, 2021 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-33750814

RESUMO

Metallothionein-3 has poorly characterized functions in neuroblastoma. Cisplatin-based chemotherapy is a major regimen to treat neuroblastoma, but its clinical efficacy is limited by chemoresistance. We investigated the impact of human metallothionein-3 (hMT3) up-regulation in neuroblastoma cells and the mechanisms underlying the cisplatin-resistance. We confirmed the cisplatin-metallothionein complex formation using mass spectrometry. Overexpression of hMT3 decreased the sensitivity of neuroblastoma UKF-NB-4 cells to cisplatin. We report, for the first time, cisplatin-sensitive human UKF-NB-4 cells remodelled into cisplatin-resistant cells via high and constitutive hMT3 expression in an in vivo model using chick chorioallantoic membrane assay. Comparative proteomic analysis demonstrated that several biological pathways related to apoptosis, transport, proteasome, and cellular stress were involved in cisplatin-resistance in hMT3 overexpressing UKF-NB-4 cells. Overall, our data confirmed that up-regulation of hMT3 positively correlated with increased cisplatin-chemoresistance in neuroblastoma, and a high level of hMT3 could be one of the causes of frequent tumour relapses.


Assuntos
Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Metalotioneína 3/biossíntese , Proteínas de Neoplasias/biossíntese , Animais , Linhagem Celular Tumoral , Embrião de Galinha , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Metalotioneína 3/genética , Proteínas de Neoplasias/genética
6.
J Proteome Res ; 20(1): 776-785, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-32924499

RESUMO

Identification of metal-binding sites in proteins and understanding metal-coupled protein folding mechanisms are aspects of high importance for the structure-to-function relationship. Mass spectrometry (MS) has brought a powerful adjunct perspective to structural biology, obtaining from metal-to-protein stoichiometry to quaternary structure information. Currently, the different experimental and/or instrumental setups usually require the use of multiple data analysis software, and in some cases, they lack some of the main data analysis steps (MS processing, scoring, identification). Here, we present a comprehensive data analysis pipeline that addresses charge-state deconvolution, statistical scoring, and mass assignment for native MS, bottom-up, and native top-down with emphasis on metal-protein complexes. We have evaluated all of the approaches using assemblies of increasing complexity, including free and chemically labeled proteins, from low- to high-resolution MS. In all cases, the results have been compared with common software and proved how MetaOdysseus outperformed them.


Assuntos
Cisteína , Proteínas , Sítios de Ligação , Espectrometria de Massas , Software
7.
Int J Mol Sci ; 21(21)2020 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-33114430

RESUMO

Dietary supplementation with polyunsaturated fatty acids (PUFA) n-3 can affect cutaneous wound healing; however, recent findings demonstrate the variable extent of their influence on the quality of healing. Here, we compare the effect of several dietary oils, containing different levels of PUFA n-3 and PUFA n-6, on wound healing in the rat model. Rats were fed the feed mixture with 8% palm oil (P), safflower oil (S), fish oil (F) or Schizochytrium microalga extract (Sch) and compared to the animals fed by control feed mixture (C). Dorsal full-thickness cutaneous excisions were performed after 52 days of feeding and skin was left to heal for an additional 12 days. Histopathological analysis of skin wounds was performed, including immune cells immunolabeling and the determination of hydroxyproline amount as well as gene expression analyses of molecules contributing to different steps of the healing. Matrix-assisted-laser-desorption-ionization mass-spectrometry-imaging (MALDI-MSI) was used to determine the amount of collagen α-1(III) chain fragment in healing samples. Treatment by Schizochytrium extract resulted in decrease in the total wound area, in contrast to the safflower oil group where the size of the wound was larger when comparing to control animals. Diet with Schizochytrium extract and safflower oils displayed a tendency to increase the number of new vessels. The number of MPO-positive cells was diminished following any of oil treatment in comparison to the control, but their highest amount was found in animals with a fish oil diet. On the other hand, the number of CD68-positive macrophages was increased, with the most significant enhancement in the fish oil and safflower oil group. Hydroxyproline concentration was the highest in the safflower oil group but it was also enhanced in all other analyzed treatments in comparison to the control. MALDI-MSI signal intensity of a collagen III fragment decreased in the sequence C > S > Sch > P > F treatment. In conclusion, we observed differences in tissue response during healing between dietary oils, with the activation of inflammation observed following the treatment with oil containing high eicosapentaenoic acid (EPA) level (fish oil) and enhanced healing features were induced by the diet with high content of docosahexaenoic acid (DHA, Schizochytrium extract).


Assuntos
Gorduras Insaturadas na Dieta/administração & dosagem , Ácidos Graxos Ômega-3/análise , Ácidos Graxos Ômega-6/análise , Pele/lesões , Cicatrização/efeitos dos fármacos , Animais , Antígenos CD8/metabolismo , Colágeno Tipo III/metabolismo , Gorduras Insaturadas na Dieta/farmacologia , Modelos Animais de Doenças , Óleos de Peixe/administração & dosagem , Óleos de Peixe/química , Óleos de Peixe/farmacologia , Indóis/química , Macrófagos/imunologia , Masculino , Óleo de Palmeira/administração & dosagem , Óleo de Palmeira/química , Óleo de Palmeira/farmacologia , Ratos , Óleo de Cártamo/administração & dosagem , Óleo de Cártamo/química , Óleo de Cártamo/farmacologia , Pele/efeitos dos fármacos , Pele/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
8.
Anal Chem ; 92(19): 12950-12958, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32786475

RESUMO

Here, using human metallothionein (MT2) as an example, we describe an improved strategy based on differential alkylation coupled to MS, assisted by zinc probe monitoring, for identification of cysteine-rich binding sites with nanomolar and picomolar metal affinity utilizing iodoacetamide (IAM) and N-ethylmaleimide reagents. We concluded that an SN2 reaction provided by IAM is more suitable to label free Cys residues, avoiding nonspecific metal dissociation. Afterward, metal-bound Cys can be easily labeled in a nucleophilic addition reaction after separation by reverse-phase C18 at acidic pH. Finally, we evaluated the efficiency of the method by mapping metal-binding sites of Zn7-xMT species using a bottom-up MS approach with respect to metal-to-protein affinity and element(al) resolution. The methodology presented might be applied not only for MT2 but to identify metal-binding sites in other Cys-containing proteins.


Assuntos
Metalotioneína/química , Zinco/análise , Sítios de Ligação , Humanos , Concentração de Íons de Hidrogênio , Metalotioneína/genética
9.
Colloids Surf B Biointerfaces ; 182: 110391, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31377608

RESUMO

Nanoparticular form of titanium dioxide (TiO2 NPs) belongs to important industrial material. Despite being widely used, serious contradictions regarding biosafety of TiO2 NPs remain. We anticipate that such discrepancies could be due to a lack of understanding of a linkage between TiO2 NPs phase composition and cytotoxicity. Therefore, we synthesized two types of biphasic TiO2 NPs differing in an anatase-brookite phase composition. The study presents an array of in vitro data suggesting that TiO2 NPs with a prevailing anatase phase composition possess higher cytotoxicity compared to TiO2 NPs with an equal anatase-brookite crystallinity. This phenomenon was evidenced by significantly higher inhibition of metabolic activity and growth of epithelial and neuroblast-like cells. Moreover, anatase-prevailing TiO2 NPs tend to produce higher amount of reactive oxygen species resulting in DNA fragmentation. Further insights into the molecular aspects of cytotoxicity of anatase-prevailing TiO2 NPs were obtained by comparative proteomics delineating that TiO2 NPs deregulate expression of a variety of proteins and associated pathways. This inevitably results in a decreased cellular ability to detoxify reactive oxygen species and respond to various stress conditions. The study provides novel data that add another piece to the jigsaw of the relation between structural features of NPs and biosafety.


Assuntos
Nanopartículas Metálicas/química , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Titânio/química , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Humanos , Nanopartículas Metálicas/toxicidade , Nanopartículas Metálicas/ultraestrutura , Microscopia Eletrônica de Transmissão , Titânio/toxicidade
10.
Mol Oncol ; 13(5): 1002-1017, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30628163

RESUMO

DNA hypermethylation is one of the most common epigenetic modifications in prostate cancer (PCa). Several studies have delineated sarcosine as a PCa oncometabolite that increases the migration of malignant prostate cells while decreasing their doubling time. Here, we show that incubation of prostate cells with sarcosine elicited the upregulation of sarcosine N-demethylation enzymes, sarcosine dehydrogenase and pipecolic acid oxidase. This process was accompanied by a considerable increase in the production of the major methyl-donor S-adenosylmethionine (SAMe), together with an elevation of cellular methylation potential. Global DNA methylation analyses revealed increases in methylated CpG islands in distinct prostate cell lines incubated with sarcosine, but not in cells of nonprostate origin. This phenomenon was further associated with marked upregulation of DNA methyltransferases (Dnmts). Epigenetic changes were recapitulated through blunting of Dnmts using the hypomethylating agent 5-azacytidine, which was able to inhibit sarcosine-induced migration of prostate cells. Moreover, spatial mapping revealed concomitant increases in sarcosine, SAMe and Dnmt1 in histologically confirmed malignant prostate tissue, but not in adjacent or nonmalignant tissue, which is in line with the obtained in vitro data. In summary, we show here for the first time that sarcosine acts as an epigenetic modifier of prostate cells and that this may contribute to its oncometabolic role.


Assuntos
Ilhas de CpG , Epigênese Genética/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Próstata/metabolismo , Neoplasias da Próstata/metabolismo , Sarcosina/farmacologia , Regulação para Cima/efeitos dos fármacos , Linhagem Celular , Humanos , Masculino , Próstata/patologia , Neoplasias da Próstata/patologia
11.
Int J Biol Macromol ; 125: 270-277, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30517841

RESUMO

Looking insight pathological processes, metallothioneins (MTs) are considered to be potential biomarkers for monitoring of a development of various types of diseases, such as cancer. The early identification of the MTs in biological tissues could be important tool for the estimation of appropriate clinical therapy. Therefore, here we investigated the application of matrix assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) together with immunohistochemical analyses (IHC) using MT-1/2 antibody for MT detection in formalin-fixed paraffin-embedded (FFPE) biopsy specimens of human skin. Principal component analyses revealed differences in the peptide/protein profiles separating healthy skin from the carcinoma specimens. Statistically significant ion peaks at m/z 6038, 6300, 6676, and 7026 were more frequently detected in squamous cell carcinoma (SCC), basal cell carcinoma (BCC) and melanoma. Using IHC, we found that MT-1/2 was significantly higher in SCC and melanoma compared to healthy skin. Surprisingly, significantly low levels of MT-1/2 were found in BCC. On one side, the results indicate important role of MTs in melanoma occurrence and progression, as on the second side, there are hidden processes associated with MTs based on differences of the occurrence of the MS peaks, which could be associated with cycling of MTs isoforms.


Assuntos
Imuno-Histoquímica , Proteínas/química , Proteínas/metabolismo , Pele/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Humanos
12.
Bioconjug Chem ; 29(9): 2954-2969, 2018 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-30086240

RESUMO

Novicidin (NVC), is a membrane-penetrating peptide, which forms a stable complex with Zn-Schiff base with interesting antitumor selectivity. We studied NVC derivatives to determine functional roles of key amino acids in toxicity, helicity, and binding of the Zn-Schiff base complex. Trimmed derivatives highlighted the role of peptide length and helicity in toxicity and membrane penetration. The removal of Lys from position 1 and 2 strongly increases the ability to disrupt the membranes. The trimming of the N-terminal residues significantly increases the stability of peptide helicity enhancing penetrating properties. Gly residue derivatives undermined a role of peptide bending in membrane penetration and toxicity. After the substitution of the central Gly derivatives with Ile or Lys, the peptides retained toxicity. These results illustrate the minor role of central helix bending in NVC toxicity. Binding-site-peptide derivatives identified His residue as the sole Zn-Schiff base binding site and eliminated the role of other aromatic residues.


Assuntos
Peptídeos Catiônicos Antimicrobianos/administração & dosagem , Sistemas de Liberação de Medicamentos , Bases de Schiff/química , Zinco/administração & dosagem , Sequência de Aminoácidos , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/toxicidade , Sítios de Ligação , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Glicina/química , Humanos , Ligantes , Conformação Proteica , Espectroscopia de Prótons por Ressonância Magnética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectroscopia de Infravermelho com Transformada de Fourier , Zinco/química
13.
PLoS One ; 12(12): e0189305, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29220390

RESUMO

BACKGROUND: Treatment of advanced cutaneous melanoma remains challenging, and new data on melanoma biology are required. The most widely accepted criteria for the prognostic evaluation of melanoma are histopathological and clinical parameters, and the identification of additional tumor markers is thus of paramount importance. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI), an important tool in cancer research, is useful for unraveling the molecular profile of melanoma. METHODOLOGY/PRINCIPAL FINDINGS: In this report, we used the melanoma-bearing Libechov minipig (MeLiM), a unique animal model that allows observation of the complete spontaneous regression of invasive cutaneous melanoma, to investigate i) the differences between melanoma and healthy skin protein profiles and ii) the proteins potentially involved in spontaneous regression. The MeLiM tissues were cryosected, histologically characterized, analyzed by MALDI MSI, and immunohistologically stained. Multivariate statistical analyses of the MALDI MSI data revealed ten relevant m/z ions, of which the expression levels varied significantly among the studied MeLiM tissues. These ion peaks were used to create mass ion images/maps and visualize the differences between tumor and healthy skin specimens, as well as among histologically characterized tissue regions. CONCLUSIONS/SIGNIFICANCE: Protein profiles comprising ten statistically significant mass ion peaks useful for differentiating cutaneous melanoma and healthy skin tissues were determined. Peaks at m/z 3044, 6011, 6140 and 10180 were overexpressed in melanoma compared with healthy skin tissue. More specifically, m/z 6140 was expressed at significantly (p < 0.05) higher levels in normally growing melanoma regions than in regions with early and late spontaneous regression. This study demonstrates the clinical utility of MALDI MSI for the analysis of tissue cryosections at a molecular level.


Assuntos
Melanoma/metabolismo , Proteínas de Neoplasias/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Animais , Melanoma/patologia , Suínos
14.
Colloids Surf B Biointerfaces ; 156: 123-132, 2017 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-28527356

RESUMO

Multiwall carbon nanotubes (MWCNTs) are among the frequently studied carbon materials, particularly because of their physical and chemical properties and high potential for application in materials chemistry, industry, and medicine. MWCNTs are very promising as transporters of bioactive molecules because of their π electrons and large surface area, which can be easily modified, mostly by the application of inorganic acids for the introduction of carboxylic moieties on the surface. In the present study, we designed an oxidised MWCNTs (oMWCNTs) transporter for the targeted delivery of doxorubicin (Dox). The modification of oMWCNTs with prostate-homing peptide (SMSIARL) promotes increased cytotoxicity for prostate cancer cells. Using advanced analytical techniques, we studied the loading efficiency, stability, and release kinetics of Dox from a oMWCNTs-Dox-Pep nanoconstruct. We show that pH strictly drives Dox release, and imitating the pH of intracellular acidic compartments, 60% of Dox is released from oMWCNTs-Dox-Pep, while in plasma conditions, only a 14% release of Dox was found during 24h. The nanoconstruct displayed no cytotoxicity in non-malignant prostate cells (PNT1A), while in metastatic prostate cancer cells (LNCaP), the cytotoxic effects were close to the cytotoxicity of free Dox. This indicates that peptide modification promotes interactions with malignant cells, resulting in efficient internalisation into the intracellular region. Overall, we show that oMWCNTs are exceptional platforms for simple and stable non-covalent modification with bioactive molecules.


Assuntos
Antibióticos Antineoplásicos/química , Doxorrubicina/química , Nanotubos de Carbono , Próstata/metabolismo , Linhagem Celular Tumoral , Humanos , Cinética , Masculino , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectroscopia de Infravermelho com Transformada de Fourier
15.
J Biotechnol ; 241: 193-200, 2017 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-27984119

RESUMO

Nanobiosynthesis belongs to the most recent methods for synthesis of nanoparticles. This type of synthesis provides many advantages including the uniformity in particle shape and size. The biosynthesis has also a significant advantage regarding chemical properties of the obtained particles. In this study, we characterized the basic properties and composition of quantum dots (QDs), obtained by the extracellular biosynthesis by Escherichia coli. Furthermore, the toxicity of the biosynthesized QDs was compared to QDs prepared by microwave synthesis. The obtained results revealed the presence of cyan CdTe QDs after removal of substantial amounts of organic compounds, which stabilized the nanoparticle surface. QDs toxicity was evaluated using three cell lines Human Foreskin Fibroblast (HFF), Human Prostate Cancer cells (PC-3) and Breast Cancer cells (MCF-7) and the MTT assay. The test revealed differences in the toxicity between variants of QDs, varying about 10% in the HFF and 30% in the MCF-7 cell lines. The toxicity of the biosynthesized QDs to the PC-3 cell lines was about 35% lower in comparison with the QDs prepared by microwave synthesis.


Assuntos
Compostos de Cádmio/toxicidade , Escherichia coli/metabolismo , Pontos Quânticos/toxicidade , Telúrio/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos
16.
Sci Rep ; 6: 33379, 2016 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-27646588

RESUMO

Herein, we describe the preparation of liposomes with folate-targeting properties for the encapsulation of anti-sarcosine antibodies (antisarAbs@LIP) and sarcosine (sar@LIP). The competitive inhibitory effects of exogenously added folic acid supported the role of folate targeting in liposome internalization. We examined the effects of repeated administration on mice PC-3 xenografts. Sar@LIP treatment significantly increased tumor volume and weight compared to controls treated with empty liposomes. Moreover, antisarAbs@LIP administration exhibited a mild antitumor effect. We also identified differences in gene expression patterns post-treatment. Furthermore, Sar@LIP treatment resulted in decreased amounts of tumor zinc ions and total metallothioneins. Examination of the spatial distribution across the tumor sections revealed a sarcosine-related decline of the MT1X isoform within the marginal regions but an elevation after antisarAbs@LIP administration. Our exploratory results demonstrate the importance of sarcosine as an oncometabolite in PCa. Moreover, we have shown that sarcosine can be a potential target for anticancer strategies in management of PCa.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Ácido Fólico/metabolismo , Lipossomos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Sarcosina/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Humanos , Lipossomos/química , Lipossomos/ultraestrutura , Masculino , Metalotioneína/metabolismo , Camundongos , Modelos Biológicos , Fosfatidiletanolaminas , Neoplasias da Próstata/tratamento farmacológico , Sarcosina/química , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Zinco/metabolismo
17.
PLoS One ; 10(12): e0145426, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26695684

RESUMO

BACKGROUND: The environmental impacts of various substances on all levels of organisms are under investigation. Among these substances, endocrine-disrupting compounds (EDCs) present a threat, although the environmental significance of these compounds remains largely unknown. To shed some light on this field, we assessed the effects of 17ß-oestradiol on the growth, reproduction and formation of free radicals in Eisenia fetida. METHODOLOGY/PRINCIPAL FINDINGS: Although the observed effects on growth and survival were relatively weak, a strong impact on reproduction was observed (50.70% inhibition in 100 µg/kg of E2). We further demonstrated that the exposure of the earthworm Eisenia fetida to a contaminant of emerging concern, 17ß-oestradiol (E2), significantly affected the molecules involved in antioxidant defence. Exposure to E2 results in the production of reactive oxygen species (ROS) and the stimulation of antioxidant systems (metallothionein and reduced oxidized glutathione ratio) but not phytochelatins at both the mRNA and translated protein levels. Matrix-assisted laser desorption/ionization (MALDI)-imaging revealed the subcuticular bioaccumulation of oestradiol-3,4-quinone, altering the levels of local antioxidants in a time-dependent manner. CONCLUSIONS/SIGNIFICANCE: The present study illustrates that although most invertebrates do not possess oestrogen receptors, these organisms can be affected by oestrogen hormones, likely reflecting free diffusion into the cellular microenvironment with subsequent degradation to molecules that undergo redox cycling, producing ROS, thereby increasing environmental contamination that also perilously affects keystone animals, forming lower trophic levels.


Assuntos
Estradiol/farmacologia , Oligoquetos/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Oligoquetos/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Reprodução/efeitos dos fármacos , Poluentes do Solo/farmacologia
18.
Braz. j. infect. dis ; Braz. j. infect. dis;19(6): 604-613, Nov.-Dec. 2015. tab, graf
Artigo em Inglês | LILACS | ID: lil-769627

RESUMO

ABSTRACT BACKGROUND: Infections, mostly those associated with colonization of wound by different pathogenic microorganisms, are one of the most serious health complications during a medical treatment. Therefore, this study is focused on the isolation, characterization, and identification of microorganisms prevalent in superficial wounds of patients (n = 50) presenting with bacterial infection. METHODS: After successful cultivation, bacteria were processed and analyzed. Initially the identification of the strains was performed through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry based on comparison of protein profiles (2-30 kDa) with database. Subsequently, bacterial strains from infected wounds were identified by both matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and sequencing of 16S rRNA gene 108. RESULTS: The most prevalent species was Staphylococcus aureus (70%), and out of those 11% turned out to be methicillin-resistant (mecA positive). Identified strains were compared with patients' diagnoses using the method of artificial neuronal network to assess the association between severity of infection and wound microbiome species composition. Artificial neuronal network was subsequently used to predict patients' prognosis (n = 9) with 85% success. CONCLUSIONS: In all of 50 patients tested bacterial infections were identified. Based on the proposed artificial neuronal network we were able to predict the severity of the infection and length of the treatment.


Assuntos
Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Técnicas de Tipagem Bacteriana/métodos , Microbiota , /genética , Infecção dos Ferimentos/microbiologia , Redes Neurais de Computação , Filogenia , Índice de Gravidade de Doença , Fatores de Tempo
19.
Oncol Rep ; 34(6): 3247-55, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26503549

RESUMO

Presented exploratory pilot study was aimed at evaluation of proteins present in urinary specimens collected from prostate cancer suffering subjects after radical prostatectomy, divided into two experimental cohorts: positive (n=15) and negative (n=15) surgical margins (PSM/NSM). The presence of PSM suggests inadequate cancer clearance and the possible need for additional treatment. Proper identification of these risk-patients is therefore of a paramount importance. Total protein profiles were firstly identified by using SDS-PAGE and compared by using partial least square discrimination analysis (PLS-DA), which revealed differences in molecular weights of 80-99 and 150-235 kDa between the experimental groups. For further identification of proteins, comparative proteomic technologies were employed. Two-dimensional gel electrophoresis with subsequent identification of protein spots by using MALDI-TOF mass fingerprinting revealed differential expression of proteins between NSM/PSM cohorts. Moreover, in PSM group, three uniquely identified proteins (cyclin-dependent kinase 6, galectin-3-binding protein and L-lactate dehydrogenase C chain) were found, which show tight connection with prostate cancer and presence of all of them was previously linked to certain aspects of prostate cancer. These proteins may be associated with the molecular mechanisms of prostate cancer development; hence, their identification may be helpful for the assessment of disease progression risk after radical prostatectomy, but also for possible early diagnosis.


Assuntos
Antígenos de Neoplasias/biossíntese , Biomarcadores Tumorais/biossíntese , Proteínas de Transporte/biossíntese , Quinase 6 Dependente de Ciclina/biossíntese , Glicoproteínas/biossíntese , L-Lactato Desidrogenase/biossíntese , Neoplasias da Próstata/genética , Neoplasias da Próstata/cirurgia , Idoso , Antígenos de Neoplasias/urina , Biomarcadores Tumorais/urina , Proteínas de Transporte/urina , Quinase 6 Dependente de Ciclina/urina , Intervalo Livre de Doença , Eletroforese em Gel de Poliacrilamida , Glicoproteínas/urina , Humanos , Isoenzimas/biossíntese , Isoenzimas/urina , L-Lactato Desidrogenase/urina , Masculino , Pessoa de Meia-Idade , Prognóstico , Prostatectomia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/urina , Proteômica
20.
Electrophoresis ; 36(16): 1894-904, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26033737

RESUMO

A novel microfluidic label-free bead-based metallothionein immunosensors was designed. To the surface of superparamagnetic agarose beads coated with protein A, polyclonal chicken IgY specifically recognizing metallothionein (MT) were immobilized via rabbit IgG. The Brdicka reaction was used for metallothionein detection in a microfluidic printed 3D chip. The assembled chip consisted of a single copper wire coated with a thin layer of amalgam as working electrode. Optimization of MT detection using designed microfluidic chip was performed in stationary system as well as in the flow arrangement at various flow rates (0-1800 µL/min). In stationary arrangement it is possible to detect MT concentrations up to 30 ng/mL level, flow arrangement allows reliable detection of even lower concentration (12.5 ng/mL). The assembled miniature flow chip was subsequently tested for the detection of MT elevated levels (at approx. level 100 µg/mL) in samples of patients with cancer. The stability of constructed device for metallothionein detection in flow arrangement was found to be several days without any maintenance needed.


Assuntos
Técnicas Eletroquímicas/instrumentação , Separação Imunomagnética/instrumentação , Metalotioneína/sangue , Animais , Anticorpos Imobilizados/química , Anticorpos Imobilizados/metabolismo , Galinhas , Técnicas Eletroquímicas/métodos , Eletrodos , Desenho de Equipamento , Neoplasias de Cabeça e Pescoço/sangue , Humanos , Imunoglobulina G/química , Imunoglobulina G/metabolismo , Imunoglobulinas/química , Imunoglobulinas/metabolismo , Separação Imunomagnética/métodos , Masculino , Pessoa de Meia-Idade , Coelhos
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