RESUMO
This study explored the potential of poly-(lactic-co-glycolic) acid (PLGA) nanoparticles to enhance the effectiveness of anticancer treatments through combination therapy with phytol and α-bisabolol. The encapsulation efficiency of the nanoparticles was investigated, highlighting the role of ionic interactions between the drugs and the polymer. Characterization of PLGA-Phy+Bis nanoparticles was carried out using DLS with zeta potential and HR-TEM for size determination. Spectrophotometric measurements evaluated the encapsulation efficiency, loading efficiency, and in vitro drug release. FTIR analysis assessed the chemical interactions between PLGA and the drug actives, ensuring nanoparticle stability. GC-MS was employed to analyze the chemical composition of drug-loaded PLGA nanocarriers. Cytotoxicity was evaluated via the MTT assay, while Annexin V-FITC/PI staining and western blot analysis confirmed apoptotic cell death. Additionally, toxicity tests were performed on L-132 cells and in vivo zebrafish embryos. The study demonstrates high encapsulation efficiency of PLGA-Phy+Bis nanoparticles, which exhibit monodispersity and sizes of 189.3±5nm (DLS) and 268±54 nm (HR-TEM). Spectrophotometric analysis confirmed efficient drug encapsulation and release control. FTIR analysis revealed nanoparticle structural stability without chemical interactions. MTT assay results demonstrated the promising anticancer potential of all the three nanoparticle types (PLGA-Phy, PLGA-Bis, and PLGA-Phy+Bis) against lung cancer cells. Apoptosis was confirmed through Annexin V-FITC/PI staining and western blot analysis, which also revealed changes in Bax and Bcl-2 protein expression. Furthermore, the nanoparticles exhibited non-toxicity in L-132 cells and zebrafish embryo toxicity tests. PLGA-Phy+Bis nanoparticles exhibited efficient encapsulation, controlled release, and low toxicity. Apoptosis induction in A549 cells and non-toxicity in healthy cells highlight their clinical potential.
Assuntos
Apoptose , Sinergismo Farmacológico , Neoplasias Pulmonares , Sesquiterpenos Monocíclicos , Nanopartículas , Fitol , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Peixe-Zebra , Apoptose/efeitos dos fármacos , Animais , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Humanos , Nanopartículas/química , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Sesquiterpenos Monocíclicos/farmacologia , Sesquiterpenos Monocíclicos/administração & dosagem , Fitol/administração & dosagem , Fitol/farmacologia , Fitol/química , Fitol/toxicidade , Linhagem Celular Tumoral , Portadores de Fármacos/química , Células A549 , Liberação Controlada de Fármacos , Sesquiterpenos/farmacologia , Sesquiterpenos/administração & dosagem , Sesquiterpenos/química , Sesquiterpenos/toxicidade , Sobrevivência Celular/efeitos dos fármacosRESUMO
Candidal infections, particularly vulvovaginal candidiasis (VVC), necessitate effective therapeutic interventions in clinical settings owing to their intricate clinical nature and elusive understanding of their etiological mechanisms. Given the challenges in developing effective antifungal therapies, the strategy of repurposing existing pharmaceuticals has emerged as a promising approach to combat drug-resistant fungi. In this regard, the current study investigates molecular insights on the anti-candidal efficacy of a well-proven anticancer small molecule -3-bromopyruvate (3BP) against three clinically significant VVC causing Candida species viz., C. albicans, C. tropicalis and C. glabrata. Furthermore, the study validates 3BP's therapeutic application by developing it as a vaginal cream for the treatment of VVC. 3BP exhibited phenomenal antifungal efficacy (killing >99%) with minimum inhibitory concentrations (MIC) and minimum fungicidal concentrations (MFC) of 256 µg/mL against all tested Candida spp. Time killing kinetics experiment revealed 20 min as the minimum time required for 3BP at 2XMIC to achieve complete-killing (99.9%) in all Candida strains. Moreover, the ergosterol or sorbitol experiment explicated that the antifungal activity of 3BP does not stem from targeting the cell wall or the membrane component ergosterol. Instead, 3BP was observed to instigate a sequence of pre-apoptotic cascade events, such as phosphatidylserine (PS) externalization, nuclear condensation and ROS accumulations, as evidenced by PI, DAPI and DCFH-DA staining methods. Furthermore, 3BP demonstrated a remarkable efficacy in eradicating mature biofilms of Candida spp., achieving a maximum eradication level of 90%. Toxicity/safety profiling in both in vitro erythrocyte lysis and in vivo Galleria mellonella survival assay authenticated the non-toxic nature of 3BP up to 512 µg/mL. Finally, a vaginal cream formulated with 3BP was found to be effective in VVC-induced female mice model, as it significantly decreasing fungal load and protecting vaginal mucosa. Concomitantly, the present study serves as a clear demonstration of antifungal mechanistic action of anticancer drug -3BP, against Candida species. This finding holds significant potential for mitigating candidal infections, particularly VVC, within healthcare environments.
Assuntos
Candidíase Vulvovaginal , Candidíase , Feminino , Camundongos , Humanos , Animais , Candidíase Vulvovaginal/tratamento farmacológico , Candidíase Vulvovaginal/prevenção & controle , Candidíase Vulvovaginal/microbiologia , Antifúngicos/farmacologia , Antifúngicos/uso terapêutico , Espécies Reativas de Oxigênio/farmacologia , Cremes, Espumas e Géis Vaginais/farmacologia , Candida , Candidíase/tratamento farmacológico , Candidíase/prevenção & controle , Candida glabrata , Candida tropicalis , Ergosterol/farmacologia , Candida albicans , Testes de Sensibilidade MicrobianaRESUMO
The anti-biofilm and anti-virulence potential of the essential oil (E.O.) extracted from Hedychium larsenii M. Dan & Sathish was determined against Streptococcus pyogenes. A crystal violet assay was employed to quantify the biofilm. Linalool, a monoterpene alcohol from the E.O., showed concentration-dependent biofilm inhibition, with a maximum of 91% at a concentration of 0.004% (v/v). The AlamarBlueTM assay also confirmed Linalool's non-bactericidal anti-biofilm efficacy (0.004%). Linalool treatment impeded micro-colony formation, mature biofilm architecture, surface coverage, and biofilm thickness and impaired cell surface hydrophobicity and EPS production. Cysteine protease synthesis was quantified using the Azocasein assay, and Linalool treatment augmented its production. This suggests that Linalool destabilizes the biofilm matrix. It altered the expression of core regulons covRS, mga, srv, and ropB, and genes associated with virulence and biofilm formation, such as speB, dltA, slo, hasA, and ciaH, as revealed by qPCR analysis. Cytotoxicity analysis using human kidney cells (HEK) and the histopathological analysis in Danio rerio proved Linalool to be a druggable molecule against the biofilms formed by S. pyogenes. This is the first report on Linalool's anti-biofilm and anti-virulence potential against S. pyogenes.
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Previous reports have shown that consumption of wine has several health benefits; however, there are different types of wine. In the present study, red wines were investigated for their compositions of active ingredients. The interaction of each component in terms of its binding mode with different serum proteins was unraveled, and the components were implicated as drug candidates in clinical settings. Overall, the study indicates that red wines have a composition of flavonoids, non-flavonoids, and phenolic acids that can interact with the key regions of proteins to enhance their biological activity. Among them, rutin, resveratrol, and tannic acid have shown good binding affinity and possess beneficial properties that can enhance their role in clinical applications.
Assuntos
Biomarcadores/sangue , Proteínas Sanguíneas/antagonistas & inibidores , Flavonoides/farmacologia , Vinho/análise , Bebidas Alcoólicas , Antioxidantes/análise , Sítios de Ligação , Proteínas Sanguíneas/química , Proteínas Sanguíneas/metabolismo , Flavonoides/química , Flavonoides/farmacocinética , Fluorometria/métodos , Humanos , Ligantes , Modelos Moleculares , Conformação Molecular , Fenóis , Ligação Proteica , Relação Estrutura-Atividade , Vinho/efeitos adversosRESUMO
SARS-CoV-2, an etiological agent of COVID-19, has been the reason for the unexpected global pandemic, causing severe mortality and imposing devastative effects on public health. Despite extensive research work put forward by scientist around globe, so far, no suitable drug or vaccine (safe, affordable, and efficacious) has been identified to treat SARS-CoV-2. As an alternative way of improvising the COVID-19 treatment strategy, that is, strengthening of host immune system, a great deal of attention has been given to phytocompounds from medicinal herbs worldwide. In a similar fashion, the present study deliberately focuses on the phytochemicals of three Indian herbal medicinal plants viz., Mentha arvensis, Coriandrum sativum, and Ocimum sanctum for their efficacy to target well-recognized viral receptor protein through molecular docking and dynamic analyses. Nucleocapsid phosphoprotein (N) of SARS-CoV-2, being a pivotal player in replication, transcription, and viral genome assembly, has been recognized as one of the most attractive viral receptor protein targets for controlling the viral multiplication in the host. Out of 127 phytochemicals screened, nine (linarin, eudesmol, cadinene, geranyl acetate, alpha-thujene, germacrene A, kaempferol-3-O-glucuronide, kaempferide, and baicalin) were found to be phenomenal in terms of exhibiting high binding affinity toward the catalytic pocket of target N-protein. Further, the ADMET prediction analysis unveiled the non-tumorigenic, noncarcinogenic, nontoxic, non-mutagenic, and nonreproductive nature of the identified bioactive molecules. Furthermore, the data of molecular dynamic simulation validated the conformational and dynamic stability of the docked complexes. Concomitantly, the data of the present study validated the anti-COVID efficacy of the bioactives from selected medicinal plants of Indian origin.
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Biomaterial associated microbial infections are complicated and mostly lead to revision surgery or removal which are painful to the patients and quite expensive. These infections are difficult to treat with antibiotics as it is often related to biofilm formation. Methicillin resistant Staphylococcus aureus (MRSA) is the leading pathogen in biomaterial associated infections and well known to form biofilm on foreign materials. To reduce the risk of biomaterial associated infections, recent treatment strategies focus on modification of the implant surface to prevent the adhesion of bacteria. Antibiofilm coating is the effective approach than coating with antimicrobials as antibiofilm agents will not create selective pressure thereby excludes possibility of drug resistance. The current study identified and validated the synergistic antibiofilm activity of citral (CIT) and thymol (THY) by crystal violet quantification and microscopic analysis without alteration in growth and metabolic viability of MRSA. Polymeric antibiofilm coating with CIT + THY as active ingredients was formulated and coated on titanium surface by the process of spin coating. Fourier-transform infrared spectroscopy (FTIR) analysis confirmed the effective blending of polymeric formulation and the presence of CIT and THY. Atomic force microscopy (AFM) images revealed the homogenous coating and reduced surface roughness and thickness of the coating was measured by surface profilometer. Antibiofilm coating released CIT and THY in a sustained manner for 60 days. Antibiofilm coating effectively inhibited MRSA adherence in vitro and antibiofilm activity of coating was not affected by plasma conditioning. In addition, antibiofilm coating was non-hemolytic and non-toxic to PBMC. Thus, the current study demonstrated the effectual strategy to prevent biomaterial associated infections and proposes the prospective role of antibiofilm coating in clinical applications.
Assuntos
Staphylococcus aureus Resistente à Meticilina , Monoterpenos Acíclicos , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Biofilmes , Humanos , Leucócitos Mononucleares , Testes de Sensibilidade Microbiana , Estudos Prospectivos , Timol , Titânio/farmacologiaRESUMO
Antiplatelet drugs reduce the risks associated with atherothrombotic events and show various applications in diverse cardiovascular diseases including myocardial infarctions. Efficacy of the current antiplatelet medicines including aspirin, clopidogrel, prasugrel and ticagrelor, and the glycoprotein IIb/IIIa antagonists, are limited due to their increased risks of bleeding, and antiplatelet drug resistance. Hence, it is important to develop new effective antiplatelet drugs, with fewer side-effects. The vast repertoire of natural peptides can be explored towards this goal. Proteins and peptides derived from snake venoms and plants represent exciting candidates for the development of novel and potent antiplatelet agents. Consequently, this review discusses multiple peptides that have displayed antiplatelet aggregation activity in preclinical drug development stages. This review also describes the antiplatelet mechanisms of the peptides, emphasizing the signaling pathways intervened by them. Also, the hurdles encountered during the development of peptides into antiplatelet drugs have been listed. Finally, hitherto unexplored peptides with the potential to prevent platelet aggregation are explored.
Assuntos
Peptídeos/uso terapêutico , Inibidores da Agregação Plaquetária/uso terapêutico , Proteínas/uso terapêutico , Animais , Proteínas Alimentares/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Humanos , Peptídeos/farmacocinética , Plantas/químicaRESUMO
Flavonoids are natural polyphenolic compounds which are included in a panoply of drugs and used to treat and/or manage human ailments such as metabolic, cardiovascular, neurological disorders and cancer. Thus, the purpose of this review is to emphasize the importance of flavonoids for the treatment of autoimmune diseases and put into the limelight of the scientific community several health-promoting effects of flavonoids which could be beneficial for the development of novel drugs from natural products. Despite available reviews on flavonoids targeting various disease conditions, a comprehensive review of flavonoids for autoimmune diseases is still lacking. To the best of our knowledge, this is the first attempt to review the potential of flavonoids for autoimmune diseases. The structure-activity relationship of flavonoids in this review revealed that the rearrangement and introduction of other functional groups into the basic skeleton of flavonoids might lead to the development of new drugs which will be helpful in relieving the painful symptoms of various autoimmune diseases.
Assuntos
Anti-Inflamatórios/uso terapêutico , Doenças Autoimunes/tratamento farmacológico , Flavonoides/uso terapêutico , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacocinética , Anti-Inflamatórios/toxicidade , Doenças Autoimunes/genética , Flavonoides/química , Flavonoides/farmacocinética , Flavonoides/toxicidade , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Relação Estrutura-AtividadeRESUMO
Tanshinone IIA is one of the most predominant bioactive constituents of Danshen, a traditional Chinese medicinal plant with multiple cardiovascular protective actions. Although Tanshinone IIA has been well documented for its endothelial protective efficacy, studies unveiling the mechanism and/or molecular targets for its pharmacological activity are still inadequate. In recent studies, it has been envisaged that the expression of pentraxin 3 (PTX3) was associated with atherosclerotic cardiovascular diseases (ACVD). Therefore, the current study was designed to evaluate the possible role of Tanshinone IIA in influencing the expression of PTX3 in endothelial cells and thereby prevents endothelial dysfunction. Molecular analyses through real-time PCR, western blot, and ELISA revealed that Tanshinone IIA down-regulates PTX3 gene expression as well as protein secretion in human endothelial cells in the presence or absence of TNF-α. Besides, Tanshinone IIA inhibits the adhesion of THP1 cells (a monocytic cell line) to activated-endothelial cells stimulated with TNF-α. Furthermore, mechanistic studies uncovered the role of p38 MAPK/NF-κB pathway in Tanshinone II-A mediated pharmacological effects. Thus, the present study exemplifies the manifestation of Tanshinone IIA as a plausible alternative natural remedy for ACVD by targeting PTX3.
Assuntos
Abietanos/farmacologia , Proteína C-Reativa/metabolismo , Células Endoteliais/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Componente Amiloide P Sérico/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Proteína C-Reativa/genética , Adesão Celular , Linhagem Celular , Sobrevivência Celular , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Estrutura Molecular , Monócitos/fisiologia , NF-kappa B/metabolismo , Interferência de RNA , Componente Amiloide P Sérico/genéticaRESUMO
Pseudomonas aeruginosa is a nosocomial pathogen colonizing patients with chronic infectious diseases and has gained resistance to all the known broad spectrum antibiotics available today. The present study showcases the antibiofilm potential of an essential oil (EO) from an underexplored Cinnamomum species namely, C. tamala, against P. aeruginosa biofilms. Furthermore, the synergistic effects of the EO along with a commercially available DNase (DNaseI) and a DNase (MBD) isolated from a marine bacterium were explored for its antibiofilm activity. The results showed that the synergized action has maximum efficacy in inhibiting young and preformed biofilms. The synergized effect of EO and DNaseI showed 70% inhibition against matured biofilms of P. aeruginosa. The EO from C. tamala also showed quorum sensing inhibitory potential as it could inhibit the swarming motility behavior of P. aeruginosa. The synergistic action of EO and DNases offers a novel alternate therapeutic strategy for combating P. aeruginosa biofilm associated infections.
RESUMO
UNLABELLED: The aim of the present study was to evaluate the effect of known quorum sensing inhibitors (QSIs) against reference strain Pseudomonas aeruginosa PAO1 and 32 clinical isolates. Among the evaluated QSIs, curcumin effectively inhibited the production of quorum sensing (QS) regulated virulence factors and biofilm production in the reference strain as well as all the clinical isolates. Hence, we sought to unearth the underlying molecular mechanism responsible for QS inhibition by curcumin. Proteomic, mass spectrometric and gene ontology analysis revealed that the differentially regulated proteins are indeed involved in iron acquisition, iron storage, detoxification of reactive oxygen species and synthesis of metabolic intermediates for virulence factor production. In vitro assays also confirmed the alterations in catalase, superoxide dismutase, pyocyanin and pyoverdine production and sensitivity of PAO1 to H2O2 upon curcumin exposure. All these results suggest that curcumin attenuates the QS and biofilm formation by affecting iron homeostasis and oxidative stress response of PAO1. Furthermore, successive exposure of PAO1 to curcumin for 20 successive passages does not affect the efficacy of curcumin to inhibit virulence factor and biofilm production. Hence, it is hypothesized that curcumin inhibits the virulence factor production and biofilm formation without obviously inciting any selection pressure from PAO1. BIOLOGICAL SIGNIFICANCE: Most of the well-known QSIs, which are effective against standard strains of P. aeruginosa, are found ineffective against QS regulated virulence determinants of clinical isolates of P. aeruginosa, suggesting the existence of resistance towards QS inhibitors. The non-toxic nature, wide range of pharmacological benefits and the ability to inhibit the QS regulated virulence factors of all the tested clinical isolates of P. aeruginosa in the current study makes curcumin as a potent antagonistic agent against P. aeruginosa. The study on proteomics of P. aeruginosa against curcumin is expected to hold a greater significance in the development of antipseudomonal regimen. The results revealed the ability of curcumin to attenuate the virulence by targeting antioxidant enzymes, iron transport and biosynthesis of metabolic intermediates involved in virulence factors production.
Assuntos
Curcumina/farmacologia , Ferro/metabolismo , Proteômica/métodos , Pseudomonas aeruginosa/fisiologia , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Homeostase , Estresse Oxidativo/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , Fatores de Virulência/antagonistas & inibidores , Fatores de Virulência/biossínteseRESUMO
Infectious diseases caused by bacteria and fungi are the major cause of morbidity and mortality across the globe. Multi-drug resistance in these pathogens augments the complexity and severity of the diseases. Various studies have shown the role of biofilms in multi-drug resistance, where the pathogen resides inside a protective coat made of extracellular polymeric substances. Since biofilms directly influence the virulence and pathogenicity of a pathogen, it is optimal to employ a strategy that effectively inhibits the formation of biofilm. Pomegranate is a common food and is also used traditionally to treat various ailments. This study assessed the anti-biofilm activity of a methanolic extract of pomegranate against bacterial and fungal pathogens. Methanolic extract of pomegranate was shown to inhibit the formation of biofilms by Staphylococcus aureus, methicillin resistant S. aureus, Escherichia coli, and Candida albicans. Apart from inhibiting the formation of biofilm, pomegranate extract disrupted pre-formed biofilms and inhibited germ tube formation, a virulence trait, in C. albicans. Characterization of the methanolic extract of pomegranate revealed the presence of ellagic acid (2,3,7,8-tetrahydroxy-chromeno[5,4,3-cde]chromene-5,10-dione) as the major component. Ellagic acid is a bioactive tannin known for its antioxidant, anticancer, and anti-inflammatory properties. Further studies revealed the ability of ellagic acid to inhibit the growth of all species in suspension at higher concentrations (>75 µg ml(-1)) and biofilm formation at lower concentrations (<40 µg ml(-1)) which warrants further investigation of the potential of ellagic acid or peel powders of pomegranate for the treatment of human ailments.
Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Candida albicans/fisiologia , Escherichia coli/fisiologia , Lythraceae , Extratos Vegetais/farmacologia , Staphylococcus aureus/fisiologia , Candida albicans/efeitos dos fármacos , Candida albicans/crescimento & desenvolvimento , Cromatografia em Camada Fina , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Frutas/química , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Microscopia Confocal , Extratos Vegetais/química , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/crescimento & desenvolvimentoRESUMO
Caenorhabditis elegans has been increasingly used to study the innate immunity and for the screening of microbe/host-specific pathogenic factors. Staphylococcus aureus-mediated infections with live C. elegans were performed on solid (full-lawn) and liquid assays. S. aureus required 90 ± 10 h for the complete killing of C. elegans, but the infection was started only after 32 h of exposure with 20% inoculum of S. aureus. The short time exposure studies revealed that, in 20% of inoculum, continuous exposure to the pathogen was required for the killing of nematode. In 100% of inoculum, only 8 h of exposure was sufficient to kill the C. elegans. To evaluate kinetically at the innate immune level, the regulation of representative candidate antimicrobial genes was investigated. Both semi-quantitative reverse transcriptase polymerase chain reaction (PCR) and real-time PCR analyses indicated the regulation of candidate immune regulatory genes of lysozyme (lys-7), cysteine protease (cpr-2), and C-type lectin (clec-60 and clec-87) family members during the course of S. aureus infections, indicating the possible contribution of the above players during the host immune response against S. aureus exposures.
Assuntos
Proteínas de Caenorhabditis elegans/imunologia , Caenorhabditis elegans/imunologia , Cisteína Proteases/imunologia , Imunidade Inata , Lectinas Tipo C/imunologia , Muramidase/imunologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Contagem de Colônia Microbiana , Cisteína Proteases/genética , Cisteína Proteases/metabolismo , Farmacorresistência Bacteriana , Expressão Gênica , Interações Hospedeiro-Patógeno , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Muramidase/genética , Muramidase/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/microbiologia , VirulênciaRESUMO
The protective effect of hydroxytyrosol (HT), a strong antioxidant compound from extra virgin olive oil, against TCDD induced toxicity was investigated in human peripheral blood mononuclear cells (PBMC). PBMC (1 × 10(6)cellsmL(-1)) were divided into four groups and were incubated in a CO(2) incubator (5% CO(2)) for 12h with vehicle, TCDD (10 nM), TCDD+HT (10 nM+100 µM) and HT alone (100 µM) respectively. To clarify the role of HT against TCDD induced cytotoxicity, oxidative stress and the levels of antioxidant enzymes were assessed. Incubation of PBMC with TCDD significantly decreased cell viability, catalase (CAT) and glutathione peroxidase (GPx) and increased the levels of superoxide dismutase (SOD), glutathione reductase (GR) and oxidative stress markers such as lipid peroxidation products (LPO), protein carbonyl content (PCC) and reactive oxygen species (ROS). Whereas, HT had an effective antioxidant property as observed by the increased cell viability, normalization of antioxidant enzymes and decreased levels of LPO, PCC and ROS in PBMC co-treated with HT and TCDD. Apoptosis detection and comet assay results shows that HT, by acting as an antioxidant, prevents the damage to DNA induced by TCDD. In addition light microscopic and histopathological observations revealed that the cells are apoptotic and degenerated during TCDD treatment, whereas cells showed intact morphology during co-treatment with HT. On the whole, the results reveal that HT exerts a promising antioxidant potential in protecting the PBMC against TCDD induced oxidative stress, which might be due to the presence of catechol moiety in its structure.
Assuntos
Antioxidantes/farmacologia , Poluentes Ambientais/toxicidade , Leucócitos Mononucleares/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Álcool Feniletílico/análogos & derivados , Dibenzodioxinas Policloradas/toxicidade , Catalase/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Glutationa Peroxidase/metabolismo , Humanos , Leucócitos Mononucleares/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Azeite de Oliva , Álcool Feniletílico/farmacologia , Óleos de Plantas/química , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismoRESUMO
Leather industries release a large amount of pollution-causing chemicals which creates one of the major industrial pollutions. The development of enzyme based processes as a potent alternative to pollution-causing chemicals is useful to overcome this issue. Proteases are enzymes which have extensive applications in leather processing and in several bioremediation processes due to their high alkaline protease activity and dehairing efficacy. In the present study, we report cloning, characterization of a Mn2+ dependent alkaline serine protease gene (MASPT) of Bacillus pumilus TMS55. The gene encoding the protease from B. pumilus TMS55 was cloned and its nucleotide sequence was determined. This gene has an open reading frame (ORF) of 1,149 bp that encodes a polypeptide of 383 amino acid residues. Our analysis showed that this polypeptide is composed of 29 residues N-terminal signal peptide, a propeptide of 79 residues and a mature protein of 275 amino acids. We performed bioinformatics analysis to compare MASPT enzyme with other proteases. Homology modeling was employed to model three dimensional structure for MASPT. Structural analysis showed that MASPT structure is composed of nine α-helices and nine ß-strands. It has 3 catalytic residues and 14 metal binding residues. Docking analysis showed that residues S223, A260, N263, T328 and S329 interact with Mn2+. This study allows initial inferences about the structure of the protease and will allow the rational design of its derivatives for structure-function studies and also for further improvement of the enzyme.
Assuntos
Bacillus/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Endopeptidases/química , Endopeptidases/metabolismo , Serina Endopeptidases/química , Serina Endopeptidases/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Clonagem Molecular , Endopeptidases/genética , Dados de Sequência Molecular , Serina Endopeptidases/genéticaRESUMO
According to the Centers for Disease Control and Prevention, biofilms cause 65% of infections in developed countries. Pseudomonas aeruginosa biofilm cause life threatening infections in cystic fibrosis infection and they are 1,000 times more tolerant to antibiotic than the planktonic cells. As quorum sensing, hydrophobicity index and extracellular polysaccharide play a crucial role in biofilm formation, extracts from 46 marine bacterial isolates were screened against these factors in P. aeruginosa. Eleven extracts showed antibiofilm activity. Extracts of S6-01 (Bacillus indicus = MTCC 5559) and S6-15 (Bacillus pumilus = MTCC 5560) inhibited the formation of PAO1 biofilm up to 95% in their Biofilm Inhibitory Concentration(BIC) of 50 and 60 microg/ml and 85% and 64% in the subinhibitory concentrations (1/4 and 1/8 of the BIC, respectively). Furthermore, the mature biofilm was disrupted to 70-74% in their BIC. The antibiofilm compound from S6-15 was partially purified using solvent extraction followed by TLC and silica column and further characterized by IR analysis. Current study for the first time reveals the antibiofilm and antiquorum-sensing activity of B. pumilus, B. indicus, Bacillus arsenicus, Halobacillus trueperi, Ferrimonas balearica, and Marinobacter hydrocarbonoclasticus from marine habitat.
Assuntos
Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Bactérias/química , Biofilmes/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Antibacterianos/química , Bactérias/isolamento & purificação , Biofilmes/crescimento & desenvolvimento , Cromatografia Líquida , Cromatografia em Camada Fina , Pseudomonas aeruginosa/crescimento & desenvolvimento , Água do Mar/microbiologia , Espectrofotometria InfravermelhoRESUMO
Plasmid pSET152 is a broad host range mobilizable vector which integrates into streptomyces chromosome utilizing att site and int function of slashed circleC31. Transformation of this plasmid into Mycobacterium smegmatis mc2 155 SMR5 gave stable transformants carrying the pSET152 as an integrated copy. Integration occurred at the cross over sequence 5'TTG disrupting the gatA gene (Glu-tRNA(Gln) amidotransferase subunitA), which is non-essential under conditions used. Recombinant pSET152 plasmids carrying mce1 locus of Mycobacterium leprae were used to construct M. smegmatis transformants carrying the mce1 locus in their chromosome. RT-PCR analysis revealed specific transcripts of M. leprae mce in M. smegmatis. The transcribed mRNA carried intergenic regions between genes of mce1 locus indicating that mce1 locus is an operon. Examination of M. leprae specific mRNA from lepromatous leprosy patient's biopsy showed that mce locus is transcribed as an operon in the pathogen also.