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1.
Sci Rep ; 8(1): 16757, 2018 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-30425279

RESUMO

Directed evolution is an important research activity in synthetic biology and biotechnology. Numerous reports describe the application of tedious mutation/screening cycles for the improvement of proteins. Recently, knowledge-based approaches have facilitated the prediction of protein properties and the identification of improved mutants. However, epistatic phenomena constitute an obstacle which can impair the predictions in protein engineering. We present an innovative sequence-activity relationship (innov'SAR) methodology based on digital signal processing combining wet-lab experimentation and computational protein design. In our machine learning approach, a predictive model is developed to find the resulting property of the protein when the n single point mutations are permuted (2n combinations). The originality of our approach is that only sequence information and the fitness of mutants measured in the wet-lab are needed to build models. We illustrate the application of the approach in the case of improving the enantioselectivity of an epoxide hydrolase from Aspergillus niger. n = 9 single point mutants of the enzyme were experimentally assessed for their enantioselectivity and used as a learning dataset to build a model. Based on combinations of the 9 single point mutations (29), the enantioselectivity of these 512 variants were predicted, and candidates were experimentally checked: better mutants with higher enantioselectivity were indeed found.


Assuntos
Evolução Molecular Direcionada/métodos , Epóxido Hidrolases/genética , Epóxido Hidrolases/metabolismo , Aprendizado de Máquina , Aspergillus niger/enzimologia , Domínio Catalítico , Epóxido Hidrolases/química , Modelos Moleculares , Mutação , Estereoisomerismo , Especificidade por Substrato
2.
Angew Chem Int Ed Engl ; 57(51): 16678-16682, 2018 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-30383920

RESUMO

The properties and structures of viruses are directly related to the three-dimensional structure of their capsid proteins, which arises from a combination of hydrophobic and supramolecular interactions, such as hydrogen bonds. The design of synthetic materials demonstrating similar synergistic interactions still remains a challenge. Herein, we report the synthesis of a polymer/cyclic peptide conjugate that combines the capability to form supramolecular nanotubes via hydrogen bonds with the properties of an amphiphilic block copolymer. The analysis of aqueous solutions by scattering and imaging techniques revealed a barrel-shaped alignment of single peptide nanotubes into a large tubisome (length: 260 nm (from SANS)) with a hydrophobic core (diameter: 16 nm) and a hydrophilic shell. These systems, which have a structure that is similar to those of viruses, were tested in vitro to elucidate their activity on cells. Remarkably, the rigid tubisomes are able to perforate the lysosomal membrane in cells and release a small molecule into the cytosol.


Assuntos
Nanotubos/química , Peptídeos Cíclicos/química , Polímeros/química , Linhagem Celular Tumoral , Sobrevivência Celular , Células HEK293 , Humanos , Ligações de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Lisossomos/química , Substâncias Macromoleculares/síntese química , Substâncias Macromoleculares/química
3.
Soft Matter ; 14(30): 6320-6326, 2018 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-30019044

RESUMO

Self-assembling cyclic peptides (CP) consisting of amino acids with alternating d- and l-chirality form nanotubes by hydrogen bonding, hydrophobic interactions, and π-π stacking in solution. These highly dynamic materials are emerging as promising supramolecular systems for a wide range of biomedical applications. Herein, we discuss how varying the polymer conformation (linear vs. brush), as well as the number of polymer arms per peptide unimer affects the self-assembly of PEGylated cyclic peptides in different solvents, using small angle neutron scattering. Using the derived information, strong correlations were drawn between the size of the aggregates, solvent polarity, and its ability to compete for hydrogen bonding interactions between the peptide unimers. Using these data, it could be possible to engineer cyclic peptide nanotubes of a controlled length.

5.
Biomaterials ; 178: 570-582, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29680158

RESUMO

Size and shape have progressively appeared as some of the key factors influencing the properties of nanosized drug delivery systems. In particular, elongated materials are thought to interact differently with cells and therefore may allow alterations of in vivo fate without changes in chemical composition. A challenge, however, remains the creation of stable self-assembled materials with anisotropic shape for delivery applications that still feature the ability to disassemble, avoiding organ accumulation and facilitating clearance from the system. In this context, we report on cyclic peptide-polymer conjugates that self-assemble into supramolecular nanotubes, as confirmed by SANS and SLS. Their behaviour ex and in vivo was studied: the nanostructures are non-toxic up to a concentration of 0.5 g L-1 and cell uptake studies revealed that the pathway of entry was energy-dependent. Pharmacokinetic studies following intravenous injection of the peptide-polymer conjugates and a control polymer to rats showed that the larger size of the nanotubes formed by the conjugates reduced renal clearance and elongated systemic circulation. Importantly, the ability to slowly disassemble into small units allowed effective clearance of the conjugates and reduced organ accumulation, making these materials interesting candidates in the search for effective drug carriers.


Assuntos
Sistemas de Liberação de Medicamentos , Metacrilatos/química , Nanotubos/química , Peptídeos Cíclicos/química , Animais , Linhagem Celular Tumoral , Humanos , Masculino , Metacrilatos/farmacocinética , Difração de Nêutrons , Peptídeos Cíclicos/sangue , Peptídeos Cíclicos/farmacocinética , Polímeros/síntese química , Polímeros/química , Ratos Sprague-Dawley , Espalhamento de Radiação , Distribuição Tecidual
6.
J Allergy Clin Immunol Pract ; 6(3): 987-995, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29355645

RESUMO

BACKGROUND: Inefficient inhaler technique (IT) compromises the optimal delivery of medication. However, the IT knowledge of health care professionals (HCPs) has received scant attention. OBJECTIVE: The objective of this study was to perform a systematic review of published reports assessing the IT proficiency of HCPs in using pressurized metered dose (pMDI) and dry powder (DPI) inhalers. METHODS: Studies published between 1975 and 2014 that directly assessed the IT skills of HCPs were selected according to predefined selection criteria. RESULTS: Data were extracted from 55 studies involving 6,304 HCPs who performed 9,996 tests to demonstrate their IT proficiency. Overall, the IT was considered correct in 15.5% of cases (95% confidence interval [CI], 12-19.3), decreasing over time from 20.5% (95% CI, 14.9-26.8) from the early period (defined as 1975-1995) to 10.8% (95% CI, 7.3-14.8) during the late period (1996-2014). The most common errors in the use of pMDIs were as follows: not breathing out completely before inhalation (75%; 95% CI, 56-90), lack of coordination (64%; 95% CI, 29-92), and postinhalation breath-hold (63%; 95% CI, 52-72). The most common errors using DPI were deficient preparation (89%; 95% CI, 82-95), not breathing out completely before inhalation (79%; 95% CI, 68-87), and no breath-hold (76%; 95% CI, 67-84). CONCLUSIONS: HCPs demonstrated inadequate knowledge of the proper use of inhalers. The poor understanding of the correct use of these devices may prevent these professionals from being able to adequately assess and teach proper inhalation techniques to their patients.


Assuntos
Asma/epidemiologia , Uso de Medicamentos/estatística & dados numéricos , Inaladores de Pó Seco , Erros de Medicação/estatística & dados numéricos , Inaladores Dosimetrados , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Administração por Inalação , Asma/tratamento farmacológico , Bases de Dados Factuais , Pessoal de Saúde , Humanos
8.
Biomacromolecules ; 19(1): 239-247, 2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29156128

RESUMO

Functional drug carrier systems have potential for increasing solubility and potency of drugs while reducing side effects. Complex polymeric materials, particularly anisotropic structures, are especially attractive due to their long circulation times. Here, we have conjugated cyclic peptides to the biocompatible polymer poly(2-hydroxypropyl methacrylamide) (pHPMA). The resulting conjugates were functionalized with organoiridium anticancer complexes. Small angle neutron scattering and static light scattering confirmed their self-assembly and elongated cylindrical shape. Drug-loaded nanotubes exhibited more potent antiproliferative activity toward human cancer cells than either free drug or the drug-loaded polymers, while the nanotubes themselves were nontoxic. Cellular accumulation studies revealed that the increased potency of the conjugate appears to be related to a more efficient mode of action rather than a higher cellular accumulation of iridium.


Assuntos
Antineoplásicos/administração & dosagem , Sistemas de Liberação de Medicamentos , Nanotubos/química , Compostos Organometálicos/administração & dosagem , Peptídeos Cíclicos/química , Polímeros/química , Antineoplásicos/farmacocinética , Linhagem Celular Tumoral , Humanos , Espectroscopia de Ressonância Magnética , Nêutrons , Compostos Organometálicos/farmacocinética , Espalhamento a Baixo Ângulo , Espectroscopia de Infravermelho com Transformada de Fourier
9.
Macromol Biosci ; 18(1)2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29211345

RESUMO

Polymer masked-unmasked protein therapy (PUMPT) employs polymer conjugation to protect therapeutic proteins during transit through the bloodstream and allow controlled release at a disease site via triggered degradation of the polymeric component. Most reported PUMPT systems are based on the specific enzymatic degradation of the polymeric component to release the protein and reinstate its activity. In these cases, therapeutic output is dependent on the presence of the required enzyme at the disease site at a sufficiently high concentration. The present study aims to overcome this design limitation by using pH as the protein release trigger. An acidic-pH triggered PUMPT system is described herein employing biodegradable polyacetals (PAs) and trypsin as a model protein. While this system protects trypsin activity at the neutral pH of the bloodstream, acidic pH (characteristic of disease sites, tissue damage, or lysosomal compartments) contributes to PA degradation and the "unmasking" of protein activity.


Assuntos
Acetais/síntese química , Sistemas de Liberação de Medicamentos , Polímeros/síntese química , Proteínas/síntese química , Acetais/química , Acetais/farmacologia , Linhagem Celular Tumoral , Liberação Controlada de Fármacos , Humanos , Concentração de Íons de Hidrogênio , Neoplasias/tratamento farmacológico , Polímeros/química , Polímeros/farmacologia , Proteínas/química , Proteínas/farmacologia , Tripsina/síntese química , Tripsina/química , Tripsina/farmacologia
10.
Chem Commun (Camb) ; 53(87): 11901-11904, 2017 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-29043301

RESUMO

The combination of the reversible addition fragmentation chain transfer (RAFT) polymerization R-group grafting from approach and RAFT one-pot acrylamide multiblock methodology is used to synthesise complex bottle-brush architectures. This is demonstrated for block copolymer grafted side chains and the insertion of ungrafted blocks into the backbone to yield multi-segmented bottle-brushes.

11.
J Drug Target ; 25(9-10): 829-844, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28737432

RESUMO

The generation of rationally designed polymer therapeutics via the conjugation of low molecular weight anti-cancer drugs to water-soluble polymeric nanocarriers aims to improve the therapeutic index. Here, we focus on applying polymer therapeutics to target two cell compartments simultaneously - tumour cells and angiogenic endothelial cells. Comparing different polymeric backbones carrying the same therapeutic agent and targeting moiety may shed light on any correlation between the choice of polymer and the anti-cancer activity of the conjugate. Here, we compared three paclitaxel (PTX)-bound conjugates with poly-l-glutamic acid (PGA, 4.9 mol%), 2-hydroxypropylmethacrylamide (HPMA, 1.2 mol%) copolymer, or polyethyleneglycol (PEG, 1:1 conjugate). PGA and HPMA copolymers are multivalent polymers that allow the conjugation of multiple compounds within the same polymer backbone, while PEG is a bivalent commercially available Food and Drug Administration (FDA)-approved polymer. We further conjugated PGA-PTX and PEG-PTX with the integrin αvß3-targeting moiety RGD (5.5 mol% and 1:1 conjugate, respectively). We based our selection on the overexpression of integrin αvß3 on angiogenic endothelial cells and several types of cancer cells. Our findings suggest that the polymer structure has major effect on the conjugate's activity on different tumour compartments. A multivalent PGA-PTX-E-[c(RGDfK)2] conjugate displayed a stronger inhibitory effect on the endothelial compartment, showing a 50% inhibition of the migration of human umbilical vein endothelial cell cells, while a PTX-PEG-E-[c(RGDfK)2] conjugate possessed enhanced anti-cancer activity on MDA-MB-231 tumour cells (IC50 = 20 nM versus IC50 300 nM for the PGA conjugate).


Assuntos
Antineoplásicos/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Integrina alfaVbeta3/antagonistas & inibidores , Nanopartículas/administração & dosagem , Paclitaxel/administração & dosagem , Polímeros/administração & dosagem , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Integrina alfaVbeta3/metabolismo , Nanopartículas/metabolismo , Paclitaxel/química , Polímeros/química
12.
Chest ; 150(2): 394-406, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27060726

RESUMO

BACKGROUND: Problems with the use of inhalers by patients were noted shortly after the launch of the metered-dose inhaler (MDI) and persist today. We aimed to assess the most common errors in inhaler use over the past 40 years in patients treated with MDIs or dry powder inhalers (DPIs). METHODS: A systematic search for articles reporting direct observation of inhaler technique by trained personnel covered the period from 1975 to 2014. Outcomes were the nature and frequencies of the three most common errors; the percentage of patients demonstrating correct, acceptable, or poor technique; and variations in these outcomes over these 40 years and when partitioned into years 1 to 20 and years 21 to 40. Analyses were conducted in accordance with recommendations from Preferred Reporting Items for Systematic Reviews and Meta-Analyses and Strengthening the Reporting of Observational Studies in Epidemiology. RESULTS: Data were extracted from 144 articles reporting on a total number of 54,354 subjects performing 59,584 observed tests of technique. The most frequent MDI errors were in coordination (45%; 95% CI, 41%-49%), speed and/or depth of inspiration (44%; 40%-47%), and no postinhalation breath-hold (46%; 42%-49%). Frequent DPI errors were incorrect preparation in 29% (26%-33%), no full expiration before inhalation in 46% (42%-50%), and no postinhalation breath-hold in 37% (33%-40%). The overall prevalence of correct technique was 31% (28%-35%); of acceptable, 41% (36%-47%); and of poor, 31% (27%-36%). There were no significant differences between the first and second 20-year periods of scrutiny. CONCLUSIONS: Incorrect inhaler technique is unacceptably frequent and has not improved over the past 40 years, pointing to an urgent need for new approaches to education and drug delivery.


Assuntos
Asma/tratamento farmacológico , Inaladores de Pó Seco , Inaladores Dosimetrados , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Autoadministração/normas , Administração por Inalação , Humanos , Autoadministração/tendências
13.
Biomacromolecules ; 16(7): 1924-37, 2015 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-25988940

RESUMO

New pH-responsive polymersomes for active anticancer oligonucleotide delivery were prepared from triblock copolymers. The delivery systems were formed by two terminal hydrophilic blocks, PEG and polyglycerolmethacrylate (poly-GMA), and a central weakly basic block, polyimidazole-hexyl methacrylate (poly-ImHeMA), which can complex with oligonucleotides and control vesicle formation/disassembly via pH variations. Targeted polymersomes were prepared by mixing folate-derivatized and underivatized copolymers. At pH 5, ds-DNA was found to complex with the pH-responsive copolymers at a N/P molar ratio above ∼2:1, which assisted the encapsulation of ds-DNA in the polymersomes, while low association was observed at pH 7.4. Cytotoxicity studies performed on folate receptor overexpressing KB and B16-F10 cells and low folate receptor expressing MCF-7 cells showed high tolerance of the polymersomes at up to 3 mg/mL concentration. Studies performed with red blood cells showed that at pH 5.0 the polymersomes have endosomolytic properties. Cytofluorimetric studies showed a 5.5-fold higher uptake of ds-DNA loaded folate-functional polymersomes in KB cells compared to nontargeted polymersomes. In addition, ds-DNA was found to be localized both in the nucleus and in the cytosol. The incubation of luciferase transfected B16-F10 cells with targeted polymersomes loaded with luciferase and Hsp90 expression silencing siRNAs yielded 31 and 23% knockdown in target protein expression, respectively.


Assuntos
Antineoplásicos/farmacologia , Núcleo Celular/genética , Polietilenoglicóis/química , RNA Interferente Pequeno/farmacologia , Citosol/metabolismo , Portadores de Fármacos/química , Receptores de Folato com Âncoras de GPI/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Células KB , Células MCF-7 , Metacrilatos/síntese química , Metacrilatos/química , Nanopartículas/química
14.
J Am Chem Soc ; 136(49): 17262-72, 2014 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-25394568

RESUMO

Structure-based directed evolution utilizing iterative saturation mutagenesis (ISM) has been applied to phenyl acetone monooxygenase (PAMO), a thermally robust Baeyer-Villiger monooxygenase, in the quest to access a mutant which displays reversed enantioselectivity in the asymmetric sulfoxidation of prochiral thioethers. Whereas WT PAMO leads to 90% ee in the sulfoxidation of p-methylbenzyl methyl thioether with preference for the (S)-sulfoxide, the evolved mutant I67Q/P440F/A442N/L443I is 95% (R)-selective in the reaction of this and other thioethers. Partial deconvolution of the (R)-selective mutant with generation of the respective four single mutants shows that all of them are (S)-selective, which points to pronounced synergism (cooperative nonadditivity) when they interact in concert. Complete deconvolution with formation of all combinatorial forms of the respective double and triple mutants allows the designed construction of a fitness landscape featuring all 24 upward pathways leading from WT to the (R)-selective quadruple mutant. In all 24 trajectories strong cooperative mutational effects were found as well, which indicates that such mutational changes in enzymes constitute nonlinear systems. A theoretical analysis based on induced fit docking explains many of the observed effects on a molecular level.

15.
Respir Care ; 59(12): 1832-7, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25233388

RESUMO

BACKGROUND: Spirometry is an apparently simple test, yet the recommended criteria for acceptability and reproducibility can be difficult to fulfill. This study aimed (1) to prospectively assess the number of tests that meet the American Thoracic Society/European Respiratory Society (ATS/ERS) 2005 acceptability and repeatability criteria in the routine practice of an experienced technician at a referral hospital's lung function laboratory, (2) to identify the most common errors, and (3) to explore patient characteristics possibly associated with failure to meet standards. METHODS: We prospectively evaluated 257 consecutive spirometries supervised by the same technician, who gave priority to achieving a minimum of 3 correct maneuvers within a maximum of 8 attempts. We recorded FVC, FEV1, expiratory time (TE), back-extrapolated volume (VE), end-of-test volume (VEOT), number of maneuvers with and without errors, and errors (VE > 0.15 L or 5% of FVC, TE < 6 s, and VEOT ≥ 0.025 L for ≥ 1 s). RESULTS: Two-hundred and fifteen spirometries (83.7%, 95% CI 78.6-87.7%) met the ATS/ERS 2005 criteria. Acceptability criteria were met in 73.9% (95% CI 71.2-76.3%) of the maneuvers and repeatability criteria in 90.7% (95% CI 86.5-93.6%). A mean ± SD of 3.3 ± 1.4 per subject was acceptable, and a mean ± SD of 4.5 ± 1.9 was obtained. TE and VEOT errors were the most common. CONCLUSIONS: Nearly 15% of the subjects failed to fulfill all the ATS/ERS 2005 criteria for spirometry performed even though they were coached by a qualified and regularly trained technician in a hospital lung function laboratory. The fact that the ATS/ERS 2005 criteria cannot be met by all patients in optimal technical conditions should be further considered and explored.


Assuntos
Fidelidade a Diretrizes/estatística & dados numéricos , Espirometria/normas , Adulto , Idoso , Feminino , Volume Expiratório Forçado , Humanos , Masculino , Pessoa de Meia-Idade , Guias de Prática Clínica como Assunto , Estudos Prospectivos , Reprodutibilidade dos Testes , Capacidade Vital
16.
Respirology ; 19(6): 900-6, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24890124

RESUMO

BACKGROUND AND OBJECTIVE: Transbronchial lung biopsy (TBLB) is required for evaluation in selected patients with interstitial lung disease (ILD). The diagnostic yield of histopathologic assessment is variable and is influenced by factors such as the size of samples and the presence of crush artefacts left by conventional biopsy forceps. We compared the diagnostic yield and safety of TBLB with cryoprobe sampling versus conventional forceps sampling. METHODS: This randomized clinical trial analysed data for 77 patients undergoing TBLB for evaluation of ILD; patients were assigned to either a conventional-forceps group or a cryoprobe group. Two pathologists assessed the tissue samples and agreed on histopathologic diagnoses. We also compared the duration of procedures, complications and sample-quality variables. RESULTS: The most frequent diagnosis observed in the cryoprobe group was non-specific interstitial pneumonia. Histopathologic diagnoses were identified in more cases in the cryoprobe group (74.4%) than in the conventional-forceps group (34.1%) (P < 0.001), and the diagnostic yield was higher in the cryoprobe group (51.3% vs 29.1% in the conventional forceps group; P = 0.038). A larger mean area of tissue was harvested by cryoprobe (14.7 ± 11 mm(2) ) than by conventional forceps (3.3 ± 4.1 mm(2)) (P < 0.001). More grade 2 bleeding (not statistically significant) occurred in the cryoprobe group (56.4%) than in the conventional-forceps group (34.2%). No differences in other complications were observed. CONCLUSIONS: TBLB by cryoprobe is safe and potentially useful in the diagnosis of ILD. Larger multisite randomized trials are required to confirm the potential benefits of this procedure. Clinical trial registration at ClinicalTrials.gov: NCT01064609.


Assuntos
Biópsia/métodos , Broncoscopia/instrumentação , Criopreservação/instrumentação , Técnicas Histológicas/instrumentação , Doenças Pulmonares Intersticiais/diagnóstico , Doenças Pulmonares Intersticiais/patologia , Idoso , Biópsia/efeitos adversos , Biópsia/instrumentação , Broncoscopia/efeitos adversos , Broncoscopia/métodos , Criopreservação/métodos , Feminino , Hemorragia/epidemiologia , Hemorragia/etiologia , Técnicas Histológicas/métodos , Humanos , Incidência , Pulmão/patologia , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos Testes , Instrumentos Cirúrgicos
17.
Respir Med ; 107(4): 495-502, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23290591

RESUMO

This brief overview of the factors determining lung deposition of aerosols provides background information required by health care providers when instructing patients to use their prescribed inhalers. We discuss differences in the optimal inhalation manoeuvres for each type of aerosol generator and the difficulties patients face. Provision of short, clear instructions with demonstration of critical steps and checking technique during later clinical visits are necessary if these aerosolised medications are to be fully beneficial.


Assuntos
Antiasmáticos/administração & dosagem , Nebulizadores e Vaporizadores , Administração por Inalação , Aerossóis/administração & dosagem , Asma/tratamento farmacológico , Inaladores de Pó Seco , Desenho de Equipamento , Humanos , Erros de Medicação/estatística & dados numéricos , Inaladores Dosimetrados , Educação de Pacientes como Assunto/métodos
18.
Arch. bronconeumol. (Ed. impr.) ; 48(6): 189-196, jun. 2012. graf, tab
Artigo em Espanhol | IBECS | ID: ibc-99442

RESUMO

El conocimiento de los profesionales y la destreza de los pacientes en el uso de los dispositivos inhalados continúa siendo deficiente. Objetivo: Validación externa por un grupo de expertos de unas recomendaciones clínicas prácticas elaboradas para la mejora del conocimiento y del uso de la terapia inhalada. Métodos: Tras una revisión bibliográfica sobre terapia inhalada se propusieron 40 recomendaciones clínicas. Se utilizó el método Delphi modificado en dos rondas para contrastar las opiniones de un panel de 59 expertos sobre dichas recomendaciones. Estas estaban agrupadas en 8 áreas: aspectos generales (4), fármacos inhalados (9), cartucho presurizado y cámaras espaciadoras (6), inhaladores de polvo (4), nebulizadores (3), dispositivos para ventilación mecánica (3), inhaladores para el niño (5) y aspectos relacionados con el cumplimiento y la educación (6). Resultados: Tras la primera ronda del panel se apreció un consenso en la aceptación de 35 de las 40 recomendaciones analizadas. Al final de la segunda ronda se alcanzó el acuerdo en 39 (97,5%). En 8 (20%) el consenso fue por unanimidad (100%). El ítem 14 fue suprimido de las recomendaciones al no alcanzarse el consenso. Conclusiones: La valoración externa por expertos en terapia inhalada constató un elevado nivel de acuerdo con las recomendaciones clínicas propuestas. Este consenso proporciona un instrumento que podría contribuir a la mejora futura en el uso de los inhaladores en nuestro ámbito(AU)


The understanding of physicians and the skill of patients in the use of inhalers continues to be inadequate. Objective: The external validation, by an expert panel, of practical clinical recommendations that had been developed in order to improve the knowledge and understanding of correct inhaled therapy use. Methods: After a bibliographic review about inhaled therapies, 40 clinical recommendations were proposed. A two-round modified Delphi consensus was used to compare the opinions of a panel of 59 experts about the recommendations, which were grouped into 8 areas: general aspects (4), inhaled drugs (9), pressurized metered-dose and spacer inhalers (6), powder inhalers (4), nebulizers (3), devices for mechanical ventilation (3), inhalers for children (5) and issues related with compliance and education (6).Results: After the first round of the consensus panel, 35 of the 40 recommendations analyzed were accepted. At the end of round 2, agreement was reached in 39 (97.5%). In 8 (20%), the consensus was unanimous (100%). Item 14 was deleted from the recommendations as consensus was not reached. Conclusions: The external validation by experts in inhaled therapy found a high level of agreement with the clinical recommendations proposed. This consensus provides a tool that could contribute to the improved use of inhalers in our country in the future(AU)


Assuntos
Humanos , Administração por Inalação , Inaladores Dosimetrados/normas , Inaladores de Pó Seco/métodos , Obstrução das Vias Respiratórias/tratamento farmacológico , Pneumopatias Obstrutivas/tratamento farmacológico , Terapia Respiratória/normas , Consenso
19.
Chembiochem ; 13(7): 1060-6, 2012 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-22522601

RESUMO

Directed evolution is a method to tune the properties of enzymes for use in organic chemistry and biotechnology, to study enzyme mechanisms, and to shed light on darwinian evolution in nature. In order to enhance its efficacy, iterative saturation mutagenesis (ISM) was implemented. This involves: 1) randomized mutation of appropriate sites of one or more residues; 2) screening of the initial mutant libraries for properties such as enzymatic rate, stereoselectivity, or thermal robustness; 3) use of the best hit in a given library as a template for saturation mutagenesis at the other sites; and 4) continuation of the process until the desired degree of enzyme improvement has been reached. Despite the success of a number of ISM-based studies, the question of the optimal choice of the many different possible pathways remains unanswered. Here we considered a complete 4-site ISM scheme. All 24 pathways were systematically explored, with the epoxide hydrolase from Aspergillus niger as the catalyst in the stereoselective hydrolytic kinetic resolution of a chiral epoxide. All 24 pathways were found to provide improved mutants with notably enhanced stereoselectivity. When a library failed to contain any hits, non-improved or even inferior mutants were used as templates in the continuation of the evolutionary pathway, thereby escaping from the local minimum. These observations have ramifications for directed evolution in general and for evolutionary biological studies in which protein engineering techniques are applied.


Assuntos
Evolução Molecular Direcionada/métodos , Enzimas/química , Enzimas/metabolismo , Sequência de Aminoácidos , Enzimas/genética , Biblioteca Gênica , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese , Mutação , Engenharia de Proteínas/métodos , Estereoisomerismo
20.
Arch. bronconeumol. (Ed. impr.) ; 48(4): 114-119, abr. 2012. tab
Artigo em Espanhol | IBECS | ID: ibc-101368

RESUMO

Si bien el asma es una de las enfermedades respiratorias crónicas más prevalentes, existe la sospecha de que la participación de los centros de neumología españoles en su manejo es potencialmente mejorable. Objetivo: Determinar el grado de implicación de los servicios de neumología españoles en la asistencia, docencia e investigación relacionadas con la enfermedad asmática y específicamente con la de control difícil. Método: Se remitió una encuesta formada por 75 ítems que cumplimentaron los responsables de los centros entrevistados. La encuesta interrogaba sobre la asistencia neumológica general y la del asma en particular, así como la actividad docente e investigadora en la enfermedad asmática en los cinco años previos. Resultados: De las 107 encuestas enviadas, 69 (64,5%) centros las retornaron cumplimentadas. De estos, 47 (68,1%) reunieron criterios de actividad asistencial destacada en asma. Sin embargo, solo 29 (42%) disponían de una consulta monográfica de asma de control difícil y 37 (53,6%) utilizaban un programa de educación. En el ámbito docente, únicamente 31 (44,9%) proporcionaban formación específica sobre asma a sus médicos residentes. Y en el de investigación, 12 (17,4%) referían tener proyectos subvencionados por SEPAR y 25 (36,2%) habían publicado trabajos en revistas con factor de impacto. Conclusiones: Si bien la mayoría de los centros neumológicos entrevistados refieren una notable actividad en la asistencia del asma, su implicación en el abordaje especializado del asma de control difícil es insuficiente. Así mismo, su participación en actividades docentes y de investigación relacionadas con la enfermedad es desigual y limitada a escasos centros(AU)


Although asthma is one of the most prevalent chronic respiratory diseases, the participation of Spanish pulmonology units in the management of asthma may have room for improvement. Objective: To determine the degree of involvement of the Spanish pulmonology services in the patient care, education and research related with asthma disease and especially in difficult-to-control asthma. Methods: A survey made up of 75 questions was sent to the heads of several pulmonology departments. The survey asked questions pertaining to respiratory disease care in general, and in asthma in particular, as well as the educational activities and research in asthma during the previous five years. Results: Out of the 107 surveys sent, 69 (645%) centers filled them out and returned them. Forty-seven (681%) met the criteria for an important level of health-care activity in asthma. However, only 29 (42%) had a monographic consultation for difficult-to-control asthma and 37 (536%) used an education program. As for postgraduate education, only 31 (449%) provided their resident physicians with specific asthma training. And in the research field, 12 (174%) reported having projects funded by SEPAR and 25 (362%) had published studies in journals with an impact factor. Conclusions: Although the majority of the pulmonology centers interviewed report a notable activity in asthma patient care, their involvement in the specialized approach for difficult-to-control asthma is insufficient. Likewise, participation in educational activities and research related with the disease is inconsistent and limited to few centers(AU)


Assuntos
Humanos , Asma/prevenção & controle , Serviço Hospitalar de Educação , Pesquisa sobre Serviços de Saúde/estatística & dados numéricos , Pesquisa sobre Serviços de Saúde , Coleta de Dados/métodos , Coleta de Dados , Pesquisa sobre Serviços de Saúde , Epidemiologia Descritiva
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