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1.
Int J Mol Sci ; 20(21)2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31661772

RESUMO

Our previous research has shown that a fungal immunomodulatory protein from Nectria haematococca (FIP-nha) possesses a wide spectrum of anti-tumor activities, and FIP-nha induced A549 apoptosis by negatively regulating the PI3K/Akt signaling pathway based on comparative quantitative proteomics. This study further confirmed that the anti-lung cancer activity of FIP-nha was significantly stronger than that of the reported LZ-8 and FIP-fve. Subsequently, 1H NMR-based metabolomics was applied to comprehensively investigate the underlying mechanism, and a clear separation of FIP-nha-treated and untreated groups was achieved using pattern recognition analysis. Four potential pathways associated with the anti-tumor effect of FIP-nha on A549 cells were identified, and these were mainly involved in glycolysis, taurine and hypotaurine metabolism, fructose and mannose metabolism, and glycerolipid metabolism. Metabolic pathway analysis demonstrated that FIP-nha could induce A549 cell apoptosis partly by regulating the p53 inhibition pathway, which then disrupted the Warburg effect, as well as through other metabolic pathways. Using RT-PCR analysis, FIP-nha-induced apoptosis was confirmed to occur through upregulation of p53 expression. This work highlights the possible use of FIP-nha as a therapeutic adjuvant for lung cancer treatment.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Fúngicas/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Nectria , Proteína Supressora de Tumor p53/metabolismo , Células A549 , Antineoplásicos/uso terapêutico , Vias Biossintéticas , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/uso terapêutico , Células HEK293 , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Proteômica , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/genética
2.
Appl Microbiol Biotechnol ; 103(14): 5567-5581, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31147756

RESUMO

Bioactive peptide natural products are an important source of therapeutics. Prominent examples are the antibiotic penicillin and the immunosuppressant cyclosporine which are both produced by fungi and have revolutionized modern medicine. Peptide biosynthesis can occur either non-ribosomally via large enzymes referred to as non-ribosomal peptide synthetases (NRPS) or ribosomally. Ribosomal peptides are synthesized as part of a larger precursor peptide where they are posttranslationally modified and subsequently proteolytically released. Such peptide natural products are referred to as ribosomally synthesized and posttranslationally modified peptides (RiPPs). Their biosynthetic pathways have recently received a lot of attention, both from a basic and applied research point of view, due to the discoveries of several novel posttranslational modifications of the peptide backbone. Some of these modifications were so far only known from NRPSs and significantly increase the chemical space covered by this class of peptide natural products. Latter feature, in combination with the promiscuity of the modifying enzymes and the genetic encoding of the peptide sequence, makes RiPP biosynthetic pathways attractive for synthetic biology approaches to identify novel peptide therapeutics via screening of de novo generated peptide libraries and, thus, exploit bioactive peptide natural products beyond their direct use as therapeutics. This review focuses on the recent discovery and characterization of novel RiPP biosynthetic pathways in fungi and their possible application for the development of novel peptide therapeutics.


Assuntos
Produtos Biológicos/metabolismo , Vias Biossintéticas , Proteínas Fúngicas/isolamento & purificação , Fungos/metabolismo , Processamento de Proteína Pós-Traducional , Ribossomos/metabolismo , Descoberta de Drogas , Proteínas Fúngicas/uso terapêutico , Fungos/genética , Peptídeos Cíclicos , Biossíntese de Proteínas
3.
Clin Infect Dis ; 66(12): 1928-1936, 2018 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-29697768

RESUMO

Background: Recurrent vulvovaginal candidiasis (RVVC) is a problematic form of mucosal Candida infection, characterized by repeated episodes per year. Candida albicans is the most common cause of RVVC. Currently, there are no immunotherapeutic treatments for RVVC. Methods: This exploratory randomized, double-blind, placebo-controlled trial evaluated an immunotherapeutic vaccine (NDV-3A) containing a recombinant C. albicans adhesin/invasin protein for prevention of RVVC. Results: The study in 188 women with RVVC (n = 178 evaluable) showed that 1 intramuscular dose of NDV-3A was safe and generated rapid and robust B- and T-cell immune responses. Post hoc exploratory analyses revealed a statistically significant increase in the percentage of symptom-free patients at 12 months after vaccination (42% vaccinated vs 22% placebo; P = .03) and a doubling in median time to first symptomatic episode (210 days vaccinated vs 105 days placebo) for the subset of patients aged <40 years (n = 137). The analysis of evaluable patients, which combined patients aged <40 years (77%) and ≥40 years (23%), trended toward a positive impact of NDV-3A versus placebo (P = .099). Conclusions: In this unprecedented study of the effectiveness of a fungal vaccine in humans, NDV-3A administered to women with RVVC was safe and highly immunogenic and reduced the frequency of symptomatic episodes of vulvovaginal candidiasis for up to 12 months in women aged <40 years. These results support further development of NDV-3A vaccine and provide guidance for meaningful clinical endpoints for immunotherapeutic management of RVVC. Clinical Trials Registration: NCT01926028.


Assuntos
Candidíase Vulvovaginal/terapia , Proteínas Fúngicas/uso terapêutico , Vacinas Fúngicas/uso terapêutico , Imunoterapia , Adolescente , Adulto , Linfócitos B/imunologia , Candida albicans/efeitos dos fármacos , Candidíase Vulvovaginal/imunologia , Método Duplo-Cego , Feminino , Vacinas Fúngicas/efeitos adversos , Humanos , Imunogenicidade da Vacina , Injeções Intramusculares , Pessoa de Meia-Idade , Recidiva , Linfócitos T/imunologia , Adulto Jovem
4.
Int J Antimicrob Agents ; 50(4): 529-535, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28669830

RESUMO

Invasive pulmonary aspergillosis (IPA) is an increasingly recognised problem in critically ill patients. Little is known about how intensivists react to an Aspergillus-positive respiratory sample or the efficacy of antifungal therapy (AFT). This study aimed to identify drivers of AFT prescription and diagnostic workup in patients with Aspergillus isolation in respiratory specimens as well as the impact of AFT in these patients. ICU patients with an Aspergillus-positive respiratory sample from the database of a previous observational, multicentre study were analysed. Cases were classified as proven/putative IPA or Aspergillus colonisation. Demographic, microbiological, diagnostic and therapeutic data were collected. Outcome was recorded 12 weeks after Aspergillus isolation. Patients with putative/proven IPA were more likely to receive AFT than colonised patients (78.7% vs. 25.5%; P <0.001). Patients with host factors for invasive fungal disease were more likely to receive AFT (72.5% vs. 37.4%) as were those with multiorgan failure (SOFA score >7) (68.4% vs. 36.9%) (both P <0.001). Once adjusted for disease severity, initiation of AFT did not alter the odds of survival (HR = 1.40, 95% CI 0.89-2.21). Likewise, treatment within 48 h following diagnosis did not change the clinical outcome (75.7% vs. 61.4%; P = 0.63). Treatment decisions appear to be based on diagnostic criteria and underlying disease severity at the time of Aspergillus isolation. IPA in this population has a dire prognosis and AFT is not associated with reduced mortality. This may be explained by delayed diagnosis and an often inevitable death due to advanced multiorgan failure.


Assuntos
Antifúngicos/uso terapêutico , Diagnóstico Tardio/mortalidade , Aspergilose Pulmonar Invasiva/diagnóstico , Aspergilose Pulmonar Invasiva/tratamento farmacológico , Idoso , Anfotericina B/uso terapêutico , Aspergillus/efeitos dos fármacos , Aspergillus/isolamento & purificação , Tomada de Decisão Clínica , Estado Terminal , Quimioterapia Combinada , Equinocandinas/uso terapêutico , Feminino , Proteínas Fúngicas/uso terapêutico , Humanos , Unidades de Terapia Intensiva , Aspergilose Pulmonar Invasiva/microbiologia , Aspergilose Pulmonar Invasiva/mortalidade , Masculino , Pessoa de Meia-Idade , Prognóstico , Sistema Respiratório/microbiologia , Resultado do Tratamento , Voriconazol/uso terapêutico
6.
Oncotarget ; 8(17): 28854-28864, 2017 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-28416764

RESUMO

Finding intracellular pathways and molecules that can prevent the proliferation of colon cancer cells can provide significant bases for developing treatments for this disease. Ostreolysin (Oly) is a protein found in the mushroom Pleurotus ostreatus, and we have produced a recombinant version of this protein (rOly).We measured the viability of several colon cancer cells treated with rOly. Xenografts and syngeneic colon cancer cells were injected into in vivo mouse models, which were then treated with this recombinant protein.rOly treatment induced a significant reduction in viability of human and mouse colon cancer cells. In contrast, there was no reduction in the viability of normal epithelial cells from the small intestine. In the search for cellular targets of rOly, we showed that it enhances the anti-proliferative activity of drugs targeting cellular tubulin. This was accompanied by a reduction in the weight and volume of tumours in mice injected with rOly as compared to their respective control mice in two in vivo models.Our results advance the functional understanding of rOly as a potential anti-cancer treatment associated with pro-apoptotic activities preferentially targeting colon cancer cells.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Proteínas Hemolisinas/uso terapêutico , Pleurotus/imunologia , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/patologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/uso terapêutico , Células HCT116 , Proteínas Hemolisinas/genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Terapia de Alvo Molecular , Proteínas Recombinantes/genética , Tubulina (Proteína)/metabolismo , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Enzyme Microb Technol ; 98: 9-17, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28110669

RESUMO

l-asparaginase is an enzyme of medical prominence and reputable as a chemotherapeutic agent. It also has immense potential to cure autoimmune and infectious diseases. The vast application of this enzyme in healthcare sector increases its market demand. However, presently the huge market demand is not achieved completely. This serves the basis to explore better producer microbial strains to bridge the gap between huge demand and supply of this therapeutic enzyme. The present study deals with the successful screening of potent microorganisms producing l-asparaginase. 47 microorganisms were screened including bacteria, fungi, and yeasts. Among all, Penicillium lilacinum showed the highest enzyme activity i.e., 39.67 IU/ml. Shigella flexneri has 23.21 IU/ml of enzyme activity (highest among all the bacterial strain tested). Further, the 3-D structure of l-asparaginase from higher producer strains was developed and validated in silico for its activity. l-asparagine (substrate for l-asparaginase) was docked inside the binding pocket of P. lilacinum and S. flexneri. Docking score for the most common substrate l-asparagine is -6.188 (P. lilacinum), -5.576 (S. flexneri) which is quite good. Moreover, the chemical property of the binding pocket revealed that amino acid residues Phe 243, Gln 260, Gly 365, Asp 386 in P. lilacinum and residues Asp 181, Thr 318, Asn 320 in S. flexneri have an important role in H-bonding. The in silico results supports and strengthen the wet lab results. The outcome obtained motivates to take the present study result from lab to industry for the economic/massive production of this enzyme for the diverse therapeutic application.


Assuntos
Asparaginase/biossíntese , Sequência de Aminoácidos , Asparaginase/genética , Asparaginase/uso terapêutico , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/genética , Proteínas de Bactérias/uso terapêutico , Biotecnologia , Domínio Catalítico , Simulação por Computador , Proteínas Fúngicas/biossíntese , Proteínas Fúngicas/genética , Proteínas Fúngicas/uso terapêutico , Humanos , Técnicas In Vitro , Microbiologia Industrial , Cinética , Ligantes , Modelos Moleculares , Pectobacterium chrysanthemi/enzimologia , Pectobacterium chrysanthemi/genética , Penicillium/enzimologia , Penicillium/genética , Alinhamento de Sequência , Shigella flexneri/enzimologia , Shigella flexneri/genética
8.
Emerg Microbes Infect ; 5(11): e114, 2016 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-27826143

RESUMO

PAF, a small antifungal protein from Penicillium chrysogenum, inhibits the growth of several pathogenic filamentous fungi, including members of the Aspergillus genus. PAF has been proven to have no toxic effects in vivo in mice by intranasal application. To test its efficacy against invasive pulmonary aspergillosis (IPA), experiments were carried out in mice suffering from IPA. Adult mice were immunosuppressed and then infected with Aspergillus fumigatus. After stable infection, the animals were inoculated with PAF intranasally at a concentration of 2.7 mg/kg twice per day. At this concentration-which is highly toxic in vitro to A. fumigatus-the mortality of the animals was slightly delayed but finally all animals died. Histological examinations revealed massive fungal infections in the lungs of both PAF-treated and untreated animal groups. Because intranasally administered PAF was unable to overcome IPA, modified and combined therapies were introduced. The intraperitoneal application of PAF in animals with IPA prolonged the survival of the animals only 1 day. Similar results were obtained with amphotericin B (AMB), with PAF and AMB being equally effective. Combined therapy with AMB and PAF-which are synergistic in vitro-was found to be more effective than either AMB or PAF treatment alone. As no toxic effects of PAF in mammals have been described thus far, and, moreover, there are so far no A. fumigatus strains with reported inherent or acquired PAF resistance, it is worth carrying out further studies to introduce PAF as a potential antifungal drug in human therapy.


Assuntos
Antifúngicos/uso terapêutico , Aspergillus fumigatus/efeitos dos fármacos , Proteínas Fúngicas/uso terapêutico , Aspergilose Pulmonar Invasiva/tratamento farmacológico , Penicillium chrysogenum/química , Administração Intranasal , Anfotericina B/uso terapêutico , Animais , Antifúngicos/química , Antifúngicos/farmacologia , Antifúngicos/toxicidade , Aspergillus fumigatus/crescimento & desenvolvimento , Modelos Animais de Doenças , Quimioterapia Combinada , Humanos , Hospedeiro Imunocomprometido , Camundongos
9.
Protein Expr Purif ; 128: 8-13, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27476120

RESUMO

Huperzine A (HupA) is a drug used for the treatment of Alzheimer's disease. However, the biosynthesis of this medicinally important compound is not well understood. The HupA biosynthetic pathway is thought to be initiated by the decarboxylation of lysine to form cadaverine, which is then converted to 5-aminopentanal by copper amine oxidase (CAO). In this study, we cloned and expressed an SsCAO gene from a HupA-producing endophytic fungus, Shiraia sp. Slf14. Analysis of the deduced protein amino acid sequence showed that it contained the Asp catalytic base, conserved motif Asn-Tyr-Asp/Glu, and three copper-binding histidines. The cDNA of SsCAO was amplified and expressed in Escherichia coli BL21(DE3), from which a 76 kDa protein was obtained. The activity of this enzyme was tested, which provided more information about the SsCAO gene in the endophytic fungus. Gas Chromatograph-Mass Spectrometry (GC-MS) revealed that this SsCAO could accept cadaverine as a substrate to produce 5-aminopentanal, the precursor of HupA. Phylogenetic tree analysis indicated that the SsCAO from Shiraia sp. Slf14 was closely related to Stemphylium lycopersici CAO. This is the first report on the cloning and expression of a CAO gene from HupA-producing endophytic fungi. Functional characterization of this enzyme provides new insights into the biosynthesis of the HupA an anti-Alzheimer's drug.


Assuntos
Amina Oxidase (contendo Cobre) , Ascomicetos/genética , Proteínas Fúngicas , Huperzia/microbiologia , Doença de Alzheimer/tratamento farmacológico , Amina Oxidase (contendo Cobre)/biossíntese , Amina Oxidase (contendo Cobre)/genética , Amina Oxidase (contendo Cobre)/isolamento & purificação , Amina Oxidase (contendo Cobre)/uso terapêutico , Ascomicetos/metabolismo , Escherichia coli , Proteínas Fúngicas/biossíntese , Proteínas Fúngicas/genética , Proteínas Fúngicas/isolamento & purificação , Proteínas Fúngicas/uso terapêutico , Humanos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/uso terapêutico
10.
Br J Nutr ; 116(2): 360-74, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27198187

RESUMO

Dietary mycoprotein decreases energy intake in lean individuals. The effects in overweight individuals are unclear, and the mechanisms remain to be elucidated. This study aimed to investigate the effect of mycoprotein on energy intake, appetite regulation, and the metabolic phenotype in overweight and obese volunteers. In two randomised-controlled trials, fifty-five volunteers (age: 31 (95 % CI 27, 35) years), BMI: 28·0 (95 % CI 27·3, 28·7) kg/m2) consumed a test meal containing low (44 g), medium (88 g) or high (132 g) mycoprotein or isoenergetic chicken meals. Visual analogue scales and blood samples were collected to measure appetite, glucose, insulin, peptide tyrosine-tyrosine (PYY) and glucagon-like peptide-1 (GLP-1). Ad libitum energy intake was assessed after 3 h in part A (n 36). Gastric emptying by the paracetamol method, resting energy expenditure and substrate oxidation were recorded in part B (n 14). Metabonomics was used to compare plasma and urine samples in response to the test meals. Mycoprotein reduced energy intake by 10 % (280 kJ (67 kcal)) compared with chicken at the high content (P=0·009). All mycoprotein meals reduced insulin concentrations compared with chicken (incremental AUClow (IAUClow): -8 %, IAUCmedium: -12 %, IAUChigh: -21 %, P=0·004). There was no significant difference in glucose, PYY, GLP-1, gastric emptying rate and energy expenditure. Following chicken intake, paracetamol-glucuronide was positively associated with fullness. After mycoprotein, creatinine and the deamination product of isoleucine, α-keto-ß-methyl-N-valerate, were inversely related to fullness, whereas the ketone body, ß-hydroxybutyrate, was positively associated. In conclusion, mycoprotein reduces energy intake and insulin release in overweight volunteers. The mechanism does not involve changes in PYY and GLP-1. The metabonomics analysis may bring new understanding to the appetite regulatory properties of food.


Assuntos
Apetite/efeitos dos fármacos , Proteínas na Dieta/farmacologia , Ingestão de Energia/efeitos dos fármacos , Proteínas Fúngicas/farmacologia , Hormônios Gastrointestinais/sangue , Insulina/sangue , Obesidade , Adulto , Animais , Apetite/fisiologia , Regulação do Apetite/fisiologia , Proteínas na Dieta/uso terapêutico , Dipeptídeos/sangue , Ingestão de Alimentos/fisiologia , Feminino , Proteínas Fúngicas/uso terapêutico , Fusarium/química , Esvaziamento Gástrico/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Obesidade/sangue , Obesidade/dietoterapia , Obesidade/fisiopatologia , Peptídeo YY/sangue , Período Pós-Prandial , Aves Domésticas , Saciação/efeitos dos fármacos , Adulto Jovem
11.
Eur J Clin Microbiol Infect Dis ; 35(2): 187-93, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26634352

RESUMO

The incidence of Candida bloodstream infections (BSIs) has increased over time, especially in medical wards. The objective of this study was to evaluate the impact of different antifungal treatment strategies on 30-day mortality in patients with Candida BSI not admitted to intensive care units (ICUs) at disease onset. This prospective, monocentric, cohort study was conducted at an 1100-bed university hospital in Rome, Italy, where an infectious disease consultation team was implemented. All cases of Candida BSIs observed in adult patients from November 2012 to April 2014 were included. Patients were grouped according to the initial antifungal strategy: fluconazole, echinocandin, or liposomal amphotericin B. Cox regression analysis was used to identify risk factors significantly associated with 15-day and 30-day mortality. During the study period, 130 patients with candidemia were observed (58 % with C. albicans, 7 % with C. glabrata, and 23 % with C. parapsilosis). The first antifungal drug was fluconazole for 40 % of patients, echinocandin for 57.0 %, and liposomal amphotericin B for 4 %. During follow-up, 33 % of patients died. The cumulative mortality 30 days after the candidemia episode was 30.8 % and was similar among groups. In the Cox regression analysis, clinical presentation was the only independent factor associated with 15-day mortality, and Acute Physiology and Chronic Health Evaluation (APACHE) II score and clinical presentation were the independent factors associated with 30-day mortality. No differences in 15-day and 30-day mortality were observed between patients with and without C. albicans candidemia. In patients with candidemia admitted to medical or surgical wards, clinical severity but not the initial antifungal strategy were significantly correlated with mortality.


Assuntos
Anfotericina B/uso terapêutico , Antifúngicos/uso terapêutico , Candidemia/tratamento farmacológico , Candidemia/mortalidade , Equinocandinas/uso terapêutico , Fluconazol/uso terapêutico , Proteínas Fúngicas/uso terapêutico , Adulto , Idoso , Candida albicans/isolamento & purificação , Candida glabrata/isolamento & purificação , Candidemia/microbiologia , Estudos de Coortes , Feminino , Hospitalização , Humanos , Unidades de Terapia Intensiva , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Estudos Prospectivos , Índice de Gravidade de Doença , Síndrome de Resposta Inflamatória Sistêmica/tratamento farmacológico , Síndrome de Resposta Inflamatória Sistêmica/microbiologia , Síndrome de Resposta Inflamatória Sistêmica/mortalidade
12.
PLoS One ; 10(11): e0140069, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26540166

RESUMO

Malformin C, a fungal cyclic pentapeptide, has been claimed to have anti-cancer potential, but no in vivo study was available to substantiate this property. Therefore, we conducted in vitro and in vivo experiments to investigate its anti-cancer effects and toxicity. Our studies showed Malformin C inhibited Colon 38 and HCT 116 cell growth dose-dependently with an IC50 of 0.27±0.07µM and 0.18±0.023µM respectively. This inhibition was explicated by Malformin C's effect on G2/M arrest. Moreover, we observed up-regulated expression of phospho-histone H2A.X, p53, cleaved CASPASE 3 and LC3 after Malformin C treatment, while the apoptosis assay indicated an increased population of necrotic and late apoptotic cells. In vivo, the pathological study exhibited the acute toxicity of Malformin C at lethal dosage in BDF1 mice might be caused by an acute yet subtle inflammatory response, consistent with elevated IL-6 in the plasma cytokine assay. Further anti-tumor and toxicity experiments proved that 0.3mg/kg injected weekly was the best therapeutic dosage of Malformin C in Colon 38 xenografted BDF1 mice, whereas 0.1mg/kg every other day showed no effect with higher resistance, and 0.9mg/kg per week either led to fatal toxicity in seven-week old mice or displayed no advantage over 0.3mg/kg group in nine-week old mice. Overall, we conclude that Malformin C arrests Colon 38 cells in G2/M phase and induces multiple forms of cell death through necrosis, apoptosis and autophagy. Malformin C has potent cell growth inhibition activity, but the therapeutic index is too low to be an anti-cancer drug.


Assuntos
Antineoplásicos/uso terapêutico , Proteínas Fúngicas/uso terapêutico , Células HCT116/efeitos dos fármacos , Peptídeos Cíclicos/uso terapêutico , Animais , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Relação Dose-Resposta a Droga , Feminino , Proteínas Fúngicas/farmacologia , Fase G2/efeitos dos fármacos , Humanos , Camundongos , Transplante de Neoplasias , Peptídeos Cíclicos/farmacologia
13.
Int J Biol Macromol ; 79: 761-78, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26047901

RESUMO

Dental plaque is a highly diverse biofilm, which has an important function in maintenance of oral and systemic health but in some conditions becomes a cause of oral diseases. In addition to mechanical plaque removal, current methods of dental plaque control involve the use of chemical agents against biofilm pathogens, which however, given the complexity of the oral microbiome, is not sufficiently effective. Hence, there is a need for development of new anti-biofilm approaches. Polysaccharides, especially (1→3),(1→6)-α-D-glucans, which are key structural and functional constituents of the biofilm matrix, seem to be a good target for future therapeutic strategies. In this review, we have focused on (1→3)-α-glucanases, which can limit the cariogenic properties of the dental plaque extracellular polysaccharides. These enzymes are not widely known and have not been exhaustively described in literature.


Assuntos
Proteínas de Bactérias/uso terapêutico , Placa Dentária/prevenção & controle , Proteínas Fúngicas/uso terapêutico , Glicosídeo Hidrolases/uso terapêutico , Streptococcus mutans/enzimologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Sequência de Carboidratos , Placa Dentária/microbiologia , Placa Dentária/patologia , Ativação Enzimática , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Expressão Gênica , Glucanos/química , Glucanos/metabolismo , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Isoenzimas/uso terapêutico , Dados de Sequência Molecular , Filogenia , Streptococcus mutans/genética
14.
PLoS One ; 10(5): e0125774, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25946033

RESUMO

Chemoresistance in cancer therapy is an unfavorable prognostic factor in non-small cell lung cancer (NSCLC). Elevation of intracellular calcium level in multidrug resistant (MDR) sublines leads to sensitization of MDR sublines to cell death. We demonstrated that a fungal protein from Ganoderma microsporum, GMI, elevates the intracellular calcium level and reduces the growth of MDR subline via autophagy and apoptosis, regardless of p-glycoprotein (P-gp) overexpression, in mice xenograft tumors. In addition, we examined the roles of autophagy in the death of MDR A549 lung cancer sublines by GMI, thapsigargin (TG) and tunicamycin (TM) in vitro. Cytotoxicity of TG was inhibited by overexpressed P-gp. However, TM-induced death of MDR sublines was independent of P-gp level. Combinations of TG and TM with either docetaxel or vincristine showed no additional cytotoxic effects on MDR sublines. TG- and TM-mediated apoptosis of MDR sublines was demonstrated on Annexin-V assay and Western blot and repressed by pan-caspase inhibitor (Z-VAD-FMK). Treatment of MDR sublines with TG and TM also augmented autophagy with accumulation of LC3-II proteins, breakdown of p62 and formation of acidic vesicular organelles (AVOs). Inhibition of ATG5 by shRNA silencing significantly reduced autophagy and cell death but not apoptosis following TG or TM treatment. GMI treatment inhibited the phosphorylation of Akt/S473 and p70S6K/T389. Interestingly, the phosphorylation of ERK was not associated with GMI-induced autophagy. We conclude that autophagy plays a pro-death role in acquired MDR and upregulation of autophagy by GMI via Akt/mTOR inhibition provides a potential strategy for overcoming MDR in the treatment of lung cancers.


Assuntos
Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Proteínas Fúngicas/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Proteína 5 Relacionada à Autofagia , Linhagem Celular Tumoral , Docetaxel , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Ganoderma , Humanos , Masculino , Medicina Tradicional Chinesa , Camundongos , Camundongos Endogâmicos NOD , Proteínas Associadas aos Microtúbulos/genética , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Interferência de RNA , RNA Interferente Pequeno , Proteínas Quinases S6 Ribossômicas 70-kDa/antagonistas & inibidores , Serina-Treonina Quinases TOR/antagonistas & inibidores , Taxoides/uso terapêutico , Tapsigargina/uso terapêutico , Tunicamicina/uso terapêutico , Vincristina/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
15.
J Oral Maxillofac Surg ; 72(9): 1703.e1-1703.e10, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24836420

RESUMO

PURPOSE: This study evaluated the osteogenetic capability of Ling Zhi-8 (LZ-8; a protein purified from traditional Chinese medicine [lingzhi]) compared with recombinant human bone morphogenic protein-2 (rhBMP-2) in a standardized bony defect using a rabbit sinus model. MATERIALS AND METHODS: Twelve male New Zealand white rabbits (18 to 24 weeks old, 3.3 to 3.8 kg) were included in the study. Implants of normal saline 0.1 mg, rhBMP-2 0.1 mg, and LZ-8 0.1 mg were each mixed with a uniform biodegradable polyurethane-based material (Nasopore). The implants were inserted in a standardized bony defect of the nasal bone created by a 2.5-mm trephine bur. The rabbits were sacrificed at 1, 2, 4, and 8 weeks postoperatively. Volume computerized tomographic and histomorphometric examinations were used to evaluate the quantity and quality of regenerated bone. RESULTS: At postoperative week 4, radiography showed that the new bone volume was significantly larger in the rhBMP-2 group compared with the LZ-8 group (P = .041) and the control group (P = .015). Histomorphometrically, better wound healing of the rhBMP-2 group was found during the healing phase compared with the other 2 groups. CONCLUSION: The biomaterial implants using rhBMP-2 and LZ-8 had good biocompatibility and osteogenetic capabilities in the rabbit sinus model. Bone healing in rhBMP-2-treated defects was excellent and showed a significant difference compared with LZ-8. However, LZ-8-treated defects also exhibited bone regeneration, and this traditional Chinese medicine may possess osteogenic potential. Further investigations of the mechanism and application of this protein in osteogenesis are needed.


Assuntos
Proteína Morfogenética Óssea 2/uso terapêutico , Medicamentos de Ervas Chinesas/uso terapêutico , Proteínas Fúngicas/uso terapêutico , Osso Nasal/cirurgia , Doenças Nasais/cirurgia , Osteogênese/efeitos dos fármacos , Fator de Crescimento Transformador beta/uso terapêutico , Implantes Absorvíveis , Animais , Materiais Biocompatíveis/química , Densidade Óssea/efeitos dos fármacos , Regeneração Óssea/efeitos dos fármacos , Modelos Animais de Doenças , Portadores de Fármacos , Humanos , Imageamento Tridimensional/métodos , Masculino , Osso Nasal/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Poliuretanos/química , Coelhos , Proteínas Recombinantes/uso terapêutico , Fatores de Tempo , Tomografia Computadorizada por Raios X/métodos , Cicatrização/efeitos dos fármacos
16.
World J Gastroenterol ; 19(35): 5837-47, 2013 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-24124328

RESUMO

AIM: To assesses the safety and efficacy of Aspergillus niger prolyl endoprotease (AN-PEP) to mitigate the immunogenic effects of gluten in celiac patients. METHODS: Patients with initial diagnosis of celiac disease as confirmed by positive serology with subtotal or total villous atrophy on duodenal biopsies who adhere to a strict gluten-free diet (GFD) resulting in normalised antibodies and mucosal healing classified as Marsh 0 or I were included. In a randomised double-blind placebo-controlled pilot study, patients consumed toast (approximately 7 g/d gluten) with AN-PEP for 2 wk (safety phase). After a 2-wk washout period with adherence of the usual GFD, 14 patients were randomised to gluten intake with either AN-PEP or placebo for 2 wk (efficacy phase). Measurements at baseline included complaints, quality-of-life, serum antibodies, immunophenotyping of T-cells and duodenal mucosa immunohistology. Furthermore, serum and quality of life questionnaires were collected during and after the safety, washout and efficacy phase. Duodenal biopsies were collected after the safety phase and after the efficacy phase. A change in histological evaluation according to the modified Marsh classification was the primary endpoint. RESULTS: In total, 16 adults were enrolled in the study. No serious adverse events occurred during the trial and no patients withdrew during the trial. The mean score for the gastrointestinal subcategory of the celiac disease quality (CDQ) was relatively high throughout the study, indicating that AN-PEP was well tolerated. In the efficacy phase, the CDQ scores of patients consuming gluten with placebo or gluten with AN-PEP did not significantly deteriorate and moreover no differences between the groups were observed. During the efficacy phase, neither the placebo nor the AN-PEP group developed significant antibody titers. The IgA-EM concentrations remained negative in both groups. Two patients were excluded from entering the efficacy phase as their mucosa showed an increase of two Marsh steps after the safety phase, yet with undetectable serum antibodies, while 14 patients were considered histologically stable on gluten with AN-PEP. Also after the efficacy phase, no significant deterioration was observed regarding immunohistological and flow cytometric evaluation in the group consuming placebo compared to the group receiving AN-PEP. Furthermore, IgA-tTG deposit staining increased after 2 wk of gluten compared to baseline in four out of seven patients on placebo. In the seven patients receiving AN-PEP, one patient showed increased and one showed decreased IgA-tTG deposits. CONCLUSION: AN-PEP appears to be well tolerated. However, the primary endpoint was not met due to lack of clinical deterioration upon placebo, impeding an effect of AN-PEP.


Assuntos
Aspergillus niger/enzimologia , Doença Celíaca/terapia , Terapia Enzimática , Proteínas Fúngicas/uso terapêutico , Glutens/metabolismo , Serina Endopeptidases/uso terapêutico , Adulto , Idoso , Anticorpos/sangue , Atrofia , Biópsia , Doença Celíaca/diagnóstico , Doença Celíaca/enzimologia , Doença Celíaca/imunologia , Método Duplo-Cego , Duodeno/efeitos dos fármacos , Duodeno/patologia , Feminino , Proteínas Fúngicas/efeitos adversos , Proteínas Fúngicas/isolamento & purificação , Glutens/imunologia , Humanos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Masculino , Pessoa de Meia-Idade , Países Baixos , Projetos Piloto , Qualidade de Vida , Serina Endopeptidases/efeitos adversos , Serina Endopeptidases/isolamento & purificação , Fatores de Tempo , Resultado do Tratamento , Adulto Jovem
17.
Food Chem Toxicol ; 59: 470-5, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23831311

RESUMO

In order to degrade aflatoxin B1 (AFB1), AFB1-degrading microbes (probiotics) such as Lactobacillus casei, Bacillus subtilis and Pichia anomala, and the AFB1-degrading enzyme from Aspergillus oryzae were selected and combined to make feed additive. Seventy-five 43-day-old male Arbor Acres broilers were randomly divided into 5 groups, 15 broilers for each group. The broilers were given with 5 kinds of diets such as the basal diet, 400 µg/kg AFB1 supplement without feed additive, and 200, 400, 800 µg/kg AFB1 supplement with 0.15% feed additive. The feeding experimental period was 30 d, which was used to determine production performance of broilers. In addition, serum, liver and chest muscle were selected for measuring AFB1 residues, gene expressions, microscopic and antioxidant analyses. The results showed that adding 0.15% feed additive in broiler diets could significantly relieve the negative effect of AFB1 on chicken's production performance and nutrient metabolic rates (P<0.05). It could also improve AFB1 metabolism, hepatic cell structure, antioxidant activity, and many hepatic enzyme gene expressions involved in oxidoreductase, apoptosis, cell growth, immune system and metabolic process (P<0.05). It could be concluded that the feed additive was able to degrade AFB1 and improve animal production.


Assuntos
Aflatoxina B1/antagonistas & inibidores , Aldeído Redutase/uso terapêutico , Antitoxinas/uso terapêutico , Proteínas Fúngicas/uso terapêutico , Regulação Enzimológica da Expressão Gênica , Fígado/metabolismo , Probióticos/uso terapêutico , Aflatoxina B1/metabolismo , Aflatoxina B1/toxicidade , Aldeído Redutase/administração & dosagem , Aldeído Redutase/metabolismo , Animais , Animais Endogâmicos , Antitoxinas/administração & dosagem , Antitoxinas/metabolismo , Aspergillus flavus/enzimologia , Aspergillus flavus/crescimento & desenvolvimento , Proteínas Aviárias/biossíntese , Proteínas Aviárias/genética , Proteínas Aviárias/metabolismo , Bacillus subtilis/crescimento & desenvolvimento , Carcinógenos/antagonistas & inibidores , Carcinógenos/metabolismo , Carcinógenos/toxicidade , Galinhas , China , Ingestão de Energia , Contaminação de Alimentos , Doenças Transmitidas por Alimentos/etiologia , Doenças Transmitidas por Alimentos/metabolismo , Doenças Transmitidas por Alimentos/patologia , Doenças Transmitidas por Alimentos/prevenção & controle , Proteínas Fúngicas/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Lactobacillus casei/crescimento & desenvolvimento , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Pichia/crescimento & desenvolvimento , Probióticos/administração & dosagem , Probióticos/metabolismo , Ganho de Peso
18.
Immunopharmacol Immunotoxicol ; 35(3): 426-33, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23656432

RESUMO

The aim of the study is to investigate the effect of recombinant Ganoderma lucidum immunoregulatory protein (rLZ-8) on mouse models of cyclophosphamide-induced leukopenia, which we have established with both single-phase and multi-phase administration methods. Treatment with rLZ-8 had a strong effect on both models of cyclophosphamide-induced leukopenia. In particular, it increased the number of neutrophils, lymphocytes and monocytes. rLZ-8 treatment also increased the percentage of CD4(+) T cells and the levels of secreted IL-3 and IL-4, which contributed to the cyclophosphamide-induced immune dysfunction and immune system imbalance. In conclusion, rLZ-8 treatment benefitted mice with cyclophosphamide-induced leukopenia by improving overall immune function and by specifically increasing the number of white blood cells.


Assuntos
Antineoplásicos Alquilantes/efeitos adversos , Ciclofosfamida/efeitos adversos , Proteínas Fúngicas/uso terapêutico , Leucopenia/tratamento farmacológico , Reishi/química , Animais , Citocinas/imunologia , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Proteínas Fúngicas/administração & dosagem , Proteínas Fúngicas/genética , Contagem de Leucócitos , Leucócitos/citologia , Leucopenia/sangue , Leucopenia/induzido quimicamente , Leucopenia/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C
19.
Acta Trop ; 127(3): 174-80, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23639468

RESUMO

This study investigated the immunotherapeutic potential of the protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride immuno-modulator (P-MAPA) on canine visceral leishmaniasis. Twenty mongrel dogs presenting clinical symptoms compatible with leishmaniasis and diagnosis confirmed by the detection of anti-leishmania antibodies were studied. Ten dogs received 15 doses of the immunomodulator (2.0 mg/kg) intramuscularly, and 10 received saline as a placebo. Skin and peripheral blood samples were collected following administration of the immunomodulator. The groups were followed to observe for clinical signals of remission; parasite load in the skin biopsies using real-time PCR, the cytokines IL-2, IL-10 and IFN-γ in the supernatant of peripheral blood mononuclear cells stimulated in vitro with either total promastigote antigen or phytohemagglutinin measured by capture ELISA, and changes in CD4⁺ and CD8⁺ T cell subpopulations evaluated by flow cytometry. Comparison between the groups showed that treatment with the immunomodulator promoted improvement in clinical signs and a significant reduction in parasite load in the skin. In peripheral blood mononuclear cell cultures, supernatants showed a decrease in IL-10 levels and an increase in IL-2 and IFN-γ. An increase in CD8⁺ T cells was observed in peripheral blood. In addition, the in vitro leishmanicidal action of P-MAPA was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and no leishmanicidal activity was detected. These findings suggest that P-MAPA has potential as an immunotherapeutic drug in canine visceral leishmaniasis, since it assists in reestablishing partial immunocompetence of infected dogs.


Assuntos
Antiprotozoários/uso terapêutico , Doenças do Cão/patologia , Proteínas Fúngicas/uso terapêutico , Fatores Imunológicos/uso terapêutico , Leishmaniose Visceral/veterinária , Animais , Antiprotozoários/efeitos adversos , Doenças do Cão/imunologia , Cães , Feminino , Proteínas Fúngicas/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Fatores Imunológicos/efeitos adversos , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-10/genética , Interleucina-10/metabolismo , Interleucina-2/genética , Interleucina-2/metabolismo , Leishmania infantum , Leishmaniose Visceral/imunologia , Leishmaniose Visceral/patologia , Fígado/efeitos dos fármacos , Masculino
20.
Toxicon ; 67: 12-6, 2013 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-23462379

RESUMO

Ribosome-inactivating proteins (RIPs) either single-chain (type 1) or two-chain (type 2) are frequent in plants, often in multiple forms. They are RNA N-glycosidases, have antiviral, antifungal and insecticidal activity. Their expression in plants is increased under stressful conditions. They are investigated for practical applications in medicine and in agriculture. In medicine, RIPs have been linked to, or fused with, appropriate antibodies or other carriers to form "immunotoxins" or other conjugates specifically toxic to the cells target of the carrier, with the aim of eliminating malignant or other undesired cells. In agriculture, it has been observed that an enhanced expression of RIPs confers to plants an increased resistance to viruses, fungi, insects, and also to drought and salinity.


Assuntos
Proteínas Inativadoras de Ribossomos/toxicidade , Proteínas Inativadoras de Ribossomos/uso terapêutico , Proteínas de Algas/uso terapêutico , Proteínas de Algas/toxicidade , Animais , Antifúngicos , Antivirais , Proteínas de Bactérias/uso terapêutico , Proteínas de Bactérias/toxicidade , Proteínas Fúngicas/uso terapêutico , Proteínas Fúngicas/toxicidade , Humanos , Imunotoxinas/uso terapêutico , Imunotoxinas/toxicidade , Inseticidas , Controle Biológico de Vetores , Proteínas de Plantas/uso terapêutico , Proteínas de Plantas/toxicidade
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