RESUMO
BACKGROUND: As promising biomarkers of diabetes, α-glucosidase (α-Glu) and ß-glucosidase (ß-Glu) play a crucial role in the diagnosis and management of diseases. However, there is a scarcity of techniques available for simultaneously and sensitively detecting both enzymes. What's more, most of the approaches for detecting α-Glu and ß-Glu rely on a single-mode readout, which can be affected by multiple factors leading to inaccurate results. Hence, the simultaneous detection of the activity levels of both enzymes in a single sample utilizing multiple-readout sensing approaches is highly attractive. RESULTS: In this work, we constructed a facile sensing platform for the simultaneous determination of α-Glu and ß-Glu by utilizing a luminescent covalent organic framework (COF) as a fluorescent indicator. The enzymatic hydrolysis product common to both enzymes, p-nitrophenol (PNP), was found to affect the fluorometric signal through an inner filter effect on COF, enhance the colorimetric response by intensifying the absorption peak at 400 nm, and induce changes in RGB values when analyzed using a smartphone-based color recognition application. By combining fluorometric/colorimetric measurements with smartphone-assisted RGB mode, we achieved sensitive and accurate quantification of α-Glu and ß-Glu. The limits of detection for α-Glu were determined to be 0.8, 1.22, and 1.85 U/L, respectively. Similarly, the limits of detection for ß-Glu were 0.16, 0.42, and 0.53 U/L, respectively. SIGNIFICANCE: Application of the proposed sensing platform to clinical serum samples revealed significant differences in the two enzymes between healthy people and diabetic patients. Additionally, the proposed sensing method was successfully applied for the screening of α-Glu inhibitors and ß-Glu inhibitors, demonstrating its viability and prospective applications in the clinical management of diabetes as well as the discovery of antidiabetic medications.
Assuntos
Inibidores de Glicosídeo Hidrolases , Estruturas Metalorgânicas , alfa-Glucosidases , beta-Glucosidase , Estruturas Metalorgânicas/química , Humanos , Inibidores de Glicosídeo Hidrolases/farmacologia , Inibidores de Glicosídeo Hidrolases/química , beta-Glucosidase/antagonistas & inibidores , beta-Glucosidase/metabolismo , alfa-Glucosidases/metabolismo , alfa-Glucosidases/sangue , Colorimetria/métodos , Limite de Detecção , Nitrofenóis/metabolismo , Nitrofenóis/química , Nitrofenóis/análise , Avaliação Pré-Clínica de Medicamentos , Corantes Fluorescentes/químicaRESUMO
As a key enzyme regulating postprandial blood glucose, α-Glucosidase is considered to be an effective target for the treatment of diabetes mellitus. In this study, a simple, rapid, and effective method for enzyme inhibitors screening assay was established based on α-glucosidase catalyzes reactions in a personal glucose meter (PGM). α-glucosidase catalyzes the hydrolysis of maltose to produce glucose, which triggers the reduction of ferricyanide (K3[Fe(CN)6]) to ferrocyanide (K4[Fe(CN)6]) and generates the PGM detectable signals. When the α-glucosidase inhibitor (such as acarbose) is added, the yield of glucose and the readout of PGM decreased accordingly. This method can achieve the direct determination of α-glucosidase activity by the PGM as simple as the blood glucose tests. Under the optimal experimental conditions, the developed method was applied to evaluate the inhibitory activity of thirty-four small-molecule compounds and eighteen medicinal plants extracts on α-glucosidase. The results exhibit that lithospermic acid (52.5 ± 3.0%) and protocatechualdehyde (36.8 ± 2.8%) have higher inhibitory activity than that of positive control acarbose (31.5 ± 2.5%) at the same final concentration of 5.0 mM. Besides, the lemon extract has a good inhibitory effect on α-glucosidase with a percentage of inhibition of 43.3 ± 3.5%. Finally, the binding sites and modes of four active small-molecule compounds to α-glucosidase were investigated by molecular docking analysis. These results indicate that the PGM method is feasible to screening inhibitors from natural products with simple and rapid operations.
Assuntos
Benzaldeídos/farmacologia , Benzofuranos/farmacologia , Glicemia/análise , Catecóis/farmacologia , Depsídeos/farmacologia , Diabetes Mellitus Tipo 2/diagnóstico , Inibidores de Glicosídeo Hidrolases/farmacologia , Monitorização Ambulatorial/métodos , alfa-Glucosidases/sangue , Acarbose/química , Acarbose/farmacologia , Benzaldeídos/química , Benzaldeídos/isolamento & purificação , Benzofuranos/química , Benzofuranos/isolamento & purificação , Sítios de Ligação , Técnicas Biossensoriais/instrumentação , Catecóis/química , Catecóis/isolamento & purificação , Depsídeos/química , Depsídeos/isolamento & purificação , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/tratamento farmacológico , Inibidores de Glicosídeo Hidrolases/química , Humanos , Hidrólise , Cinética , Maltose/metabolismo , Simulação de Acoplamento Molecular , Monitorização Ambulatorial/instrumentação , Extratos Vegetais/química , Plantas Medicinais , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Termodinâmica , Dispositivos Eletrônicos Vestíveis , alfa-Glucosidases/químicaRESUMO
A target-dependent ratiometric fluorescence sensing strategy was designed and fabricated based on a redox reaction for highly sensitive detection of α-glucosidase (α-Glu) activity and its inhibitor. In this study, silicon quantum dots (SiQDs) with excellent optical properties and two-dimensional (2D) cobalt oxyhydroxide (CoOOH) nanosheets were successfully prepared and exploited for the detection of analytes. The CoOOH nanosheets are able to oxidize o-phenylenediamine (OPD), and the product 2,3-diaminophenazine (oxOPD) not only quenches the blue fluorescence of SiQDs (440 nm) by the inner filter effect (IFE) but also emits orange fluorescence (565 nm). α-Glu can catalytically hydrolyze l-ascorbic acid-2-O-α-d-glucopyranosyl (AA2G) to produce ascorbic acid (AA). The redox between AA and CoOOH could lead to the damage of CoOOH nanosheets, thereby inhibiting the oxidization of OPD and effectively preserving the fluorescence of SiQDs. Thus, ratiometric detection of α-Glu activity was achieved according to the AA-dependent dual-fluorescence signal responses. Under the optimal conditions, good linearity was obtained in the range of 0.01-6 U mL-1 with a detection limit of 0.004 U mL-1. The IC50 of α-Glu inhibitor acarbose was estimated to be 0.216 µM. The method provides high sensitivity and selectivity for the determination of α-Glu activity and its inhibitor, which has great application potential in clinical diagnosis and anti-diabetic drug screening. Furthermore, a logic gate analytical device was successfully established based on double fluorescence signals, which makes it possible to monitor α-Glu activity by intelligence equipment.
Assuntos
Acarbose/química , Inibidores de Glicosídeo Hidrolases/química , alfa-Glucosidases/química , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/química , Cobalto/química , Fluorescência , Humanos , Lógica , Nanoestruturas/química , Oxirredução , Óxidos/química , Fenilenodiaminas/química , Pontos Quânticos/química , Silício/química , alfa-Glucosidases/sangueRESUMO
Symptoms of autonomic dysfunction are common in Fabry disease. In this study we aimed to evaluate alterations in the pupillary response to white light stimulation in patients with Fabry disease and their association with the severity of autonomic symptoms. Fourteen consecutive patients with Fabry disease and 14 healthy control participants were enrolled in this cross-sectional study. The Mainz Severity Score Index (MSSI) was used to measure the severity of Fabry disease and the Composite Autonomic Symptom Scale 31 (COMPASS 31) questionnaire was used to evaluate the severity of autonomic symptoms. The pupil light responses were assessed with an infrared dynamic pupillometry unit. There were significant reductions in the amplitude (P = 0.048) and duration (P = 0.048) of pupil contraction, and the latency of pupil dilation (P = 0.048) in patients with Fabry disease compared to control subjects. The total weighted COMPASS 31 score correlated with MSSI (r = 0.592; P = 0.026) and the duration of pupil dilation (ρ = 0.561; P = 0.037). The pupillomotor weighted sub-score of the COMPASS 31 correlated inversely with the duration of pupil contraction (r = - 0.600; P = 0.023) and latency of pupil dilation (ρ = - 0.541; P = 0.046), and directly with the duration of pupil dilation (ρ = 0.877; P < 0.001) and MSSI (r = 0.533; P = 0.049). In conclusion, abnormal pupillary function is demonstrated in patients with Fabry disease, which is associated with the severity of autonomic symptoms.
Assuntos
Sistema Nervoso Autônomo/fisiopatologia , Doença de Fabry/fisiopatologia , Distúrbios Pupilares/fisiopatologia , Adulto , Estudos de Casos e Controles , Estudos Transversais , Doença de Fabry/sangue , Feminino , Humanos , Masculino , Reflexo Pupilar , Índice de Gravidade de Doença , Inquéritos e Questionários , Centros de Atenção Terciária , alfa-Glucosidases/sangueRESUMO
In studies on the degradation of glycogen by rhGAA, a glycosylated protein core material was found which consists of about 5-6% of the total starting glycogen. There was an additional 25% of the glycogen unaccounted for based on glucose released. After incubation of glycogen with rhGAA until no more glucose was released, no other carbohydrate was detected on HPAEC-PAD. Several oligosaccharides are then detectable if the medium is first boiled in 0.1 N HCl or incubated with trypsin. It is present in serum either in an HCl extract or in a trypsin digest. The characteristics of the in vivo serum material are identical to the material in the in vitro incubation medium. One oligosaccharide cannot be further degraded by rhGAA, from the incubation medium as well as from serum co-elute on HPAEC-PAD. Several masked oligosaccharides in serum contain m-inositol, e-inositol, and sorbitol as the major carbohydrates. The presence of this glycosylated protein in serum is a fraction of glycogen that is degraded outside the lysosome and the cell. The glycosylated protein in the serum is not present in the serum of Pompe mice not on ERT, but it is present in the serum of Pompe disease patients who are on ERT, so it is a biomarker of GAA degradation of lysosomal glycogen.
Assuntos
Doença de Depósito de Glicogênio Tipo II/metabolismo , Glicogênio/metabolismo , alfa-Glucosidases/metabolismo , Feminino , Glicogênio/sangue , Doença de Depósito de Glicogênio Tipo II/sangue , Glicosilação , Humanos , Lactente , Lisossomos/metabolismo , Proteínas Recombinantes/sangue , Proteínas Recombinantes/metabolismo , Solubilidade , alfa-Glucosidases/sangueRESUMO
A turn-on method for determining α-glucosidase activity is described using a chemical redox strategy in which the fluorescence of red fluorescent carbon dots (CDs) is modulated. The red fluorescent CDs were prepared using a solvothermal method with p-phenylenediamine and sodium citrate. The excitation and emission maxima of the CDs were 490 and 618 nm, respectively. Ce4+ ions catalyze the oxidation of the colorless substrate 3,3',5,5'-tetramethylbenzidine (TMB) to give a blue oxidized TMB product (oxTMB). Absorption by oxTMB overlaps with the red light emitted by the CDs because of the fluorescence inner filter effect; therefore the presence of oxTMB decreases the intensity of fluorescence emission by the CDs. However, hydrolysis of L-ascorbic acid-2-O-α-D-glucopyranosyl by the enzyme α-glucosidase causes formation of ascorbic acid . Ascorbic acid reduces oxTMB to TMB, so that the inner filter effect disappeared and the fluorescence recovered. The strategy allows α-glucosidase activity to be successfully determined down to 0.02 U mL-1 and gives a dynamic linear range of 0-5.5 U mL-1. The strategy is very selective for α-glucosidase activity in the presence of potentially interfering substances. The method has been successfully applied to the determination of α-glucosidase activity in spiked human serum samples and gave satisfactory results. Graphical Abstract Schematic of the method used to prepare the carbon dots and the mechanisms involved in determining α-glucosidase activity.
Assuntos
Benzidinas/química , Ensaios Enzimáticos/métodos , Corantes Fluorescentes/química , Pontos Quânticos/química , alfa-Glucosidases/sangue , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/química , Carbono/química , Cério/química , Compostos Cromogênicos/química , Cor , Fluorescência , Humanos , Limite de Detecção , Oxirredução , Espectrometria de Fluorescência , alfa-Glucosidases/químicaAssuntos
Erros de Diagnóstico , Terapia de Reposição de Enzimas/métodos , Doença de Depósito de Glicogênio Tipo II/diagnóstico , alfa-Glucosidases/uso terapêutico , Feminino , Doença de Depósito de Glicogênio Tipo II/tratamento farmacológico , Humanos , Masculino , Pessoa de Meia-Idade , alfa-Glucosidases/sangueRESUMO
BACKGROUND: Interpretation of genetic variants detected by sequencing of genomic DNA, which may cause splicing defects, regularly requires mRNA analysis. Usually, only bioinformatic testing is provided, because simple and non-invasive assay protocols are lacking. Furthermore, the detection of mis-splicing is often hampered by nonsense mediated mRNA decay (NMD). METHODS: Starting from a case of Pompe disease with two potential splicing variants an assay for the analysis of splice defects in general was developed. We analyzed the transcripts from the gene of interest by standard methods after short-term culture of the patient's lymphocytes in the presence and absence of a NMD inhibitor. Variant and wild type transcript expression were quantified by allele specific PCR in the patient and both parents and the expression ratio with/without NMD inhibition was calculated for each transcript. RESULTS: NMD detection in lymphocytes was optimized and evaluated by analyzing a naturally occurring NMD transcript. Several compounds inhibited NMD successfully, including potential therapeutic agents. Sample storage for up to 4â¯days at room temperature prior to lymphocyte isolation did not affect results. In a proof of concept we identified two candidate variants as severe splicing variants in a patient with Pompe disease, but the strategy can also be used to screen for any mis-spliced transcripts prone to NMD. CONCLUSIONS: We developed a simple, non-invasive assay for the detection and characterization of potential splicing variants. This is essential, because early and near-term diagnosis and disease classification is required to facilitate therapy in many genetic diseases.
Assuntos
Processamento Alternativo/genética , Doença de Depósito de Glicogênio Tipo II/diagnóstico , Doença de Depósito de Glicogênio Tipo II/genética , Linfócitos/metabolismo , Degradação do RNAm Mediada por Códon sem Sentido/efeitos dos fármacos , RNA Mensageiro/genética , Alelos , Processamento Alternativo/efeitos dos fármacos , Anisomicina/farmacologia , Células Cultivadas , Pré-Escolar , Cromatografia Líquida , Códon sem Sentido , Éxons , Feminino , Doença de Depósito de Glicogênio Tipo II/sangue , Doença de Depósito de Glicogênio Tipo II/fisiopatologia , Heterozigoto , Humanos , Lactente , Linfócitos/efeitos dos fármacos , Masculino , Mutação , Degradação do RNAm Mediada por Códon sem Sentido/genética , Inibidores da Síntese de Proteínas/farmacologia , RNA Mensageiro/efeitos dos fármacos , Espectrometria de Massas em Tandem , alfa-Glucosidases/sangue , alfa-Glucosidases/genéticaAssuntos
Doença de Depósito de Glicogênio Tipo II/complicações , Doença de Depósito de Glicogênio Tipo I/complicações , Doença de Depósito de Glicogênio/diagnóstico , Feminino , Glucose-6-Fosfatase/sangue , Doença de Depósito de Glicogênio/classificação , Doença de Depósito de Glicogênio Tipo I/diagnóstico , Doença de Depósito de Glicogênio Tipo I/genética , Doença de Depósito de Glicogênio Tipo I/terapia , Doença de Depósito de Glicogênio Tipo II/diagnóstico , Doença de Depósito de Glicogênio Tipo II/terapia , Humanos , Hipoglicemia/etiologia , Hipoglicemia/prevenção & controle , Masculino , alfa-Glucosidases/sangueRESUMO
The objective of the present study was to evaluate the effects of low-molecular-weight chitosan (LMWC) on the growth performance, immune responses and intestinal health of weaned pigs challenged by enterotoxigenic Escherichia coli (ETEC). A total of 32 weaned pigs were randomly allocated to four treatments: non-challenged (fed with basal diet), ETEC-challenged (fed with basal diet) and ETEC-challenged plus 50 or 100 mg/kg LMWC supplementation, respectively. After 11â¯days feeding, the non-challenged pigs were infused with sterilised Luria-Bertani culture, while the remaining pigs were infused with 2.6â¯×â¯1011 colony-forming units of ETEC. At 3â¯days post-challenge, all pigs were administered d-xylose at 0.1â¯g/kg body weight. One hour later, blood samples were obtained, and the pigs then euthanised to collect intestinal samples. Data showed that only 100â¯mg/kg LMWC supplementation attenuated (Pâ¯<â¯0.05) the average daily gain reduction caused by ETEC. Furthermore, besides the decreased (P < 0.05) serum tumour necrosis factor-α and immunoglobulin (Ig) G concentrations detected in ETEC-challenged pigs supplemented with LMWC at 50 or 100 mg/kg, the higher dose (100 mg/kg) also decreased (P < 0.05) the serum IgM concentration and increased (P < 0.05) the villus height and villus height-to-crypt depth ratio in both the jejunum and ileum, and the sucrase activity in the ileal mucosa. Moreover, LMWC supplementation (50 or 100 mg/kg) in ETEC-challenged pigs elevated (Pâ¯<â¯0.05) the mRNA levels of jejunal mucosal peptide transporter 1 and ileal mucosal peptide transporter 1, divalent metal transporter 1 and zinc transporter 1, and decreased (Pâ¯<â¯0.05) the ileal and caecal E. coli abundances, while 100 mg/kg LMWC additionally elevated (Pâ¯<â¯0.05) the ileal Bacillus abundance, and caecal and colonic Bifidobacterium abundances. These results suggest that LMWC helps alleviate ETEC-induced growth retardation in weaned pigs, which could be associated with the inhibition of the immune responses and improved intestinal health.
Assuntos
Quitosana/uso terapêutico , Suplementos Nutricionais , Escherichia coli Enterotoxigênica , Infecções por Escherichia coli/dietoterapia , Transtornos do Crescimento/dietoterapia , Animais , Quitosana/química , Citocinas/sangue , Infecções por Escherichia coli/sangue , Infecções por Escherichia coli/complicações , Infecções por Escherichia coli/patologia , Transtornos do Crescimento/sangue , Transtornos do Crescimento/etiologia , Transtornos do Crescimento/patologia , Imunoglobulinas/sangue , Intestinos/efeitos dos fármacos , Intestinos/enzimologia , Intestinos/patologia , Lactase/sangue , Peso Molecular , Sacarase/sangue , Suínos , Desmame , alfa-Glucosidases/sangueRESUMO
Pomegranate juice (PJ) has gained popularity attributed to its phenolic compounds and their medicinal properties. Its potential hypoglycemic effect has been related to enzymatic inhibition, insulin release, and the protection of pancreatic tissue. These effects depend on several aspects, such as the content and composition of phenols in pomegranate and characteristics of the organism that consumes the juice. The objective of this study was to systematically review scientific evidence investigating the hypoglycemic effect of PJ; the factors that affect bioactive compounds; and the mechanisms of action attributed to this effect. Human and rodent in vivo and in vitro studies were retrieved from PubMed, Scopus, and ScienceDirect databases. After reviewing the articles, it was identified that the methodologies and results among the scientific evidence were quite heterogeneous. Despite these limitations, many of the in vivo and in vitro studies found important hypoglycemic effects from PJ, as well as an increase in the function of ß-cell, insulin secretion, a significantly lower activity of α-amylase enzyme, an inhibition of the enzyme α-glucosidase and dipeptidyl peptidase-4 (DPP-4), and the protection against DNA damage. Determining the potential health benefits of polyphenols contained in the pomegranate is limited for multiple factors that could affect the efficacy of PJ. Overall, the results of this review suggest the need for further experimentation, using controlled variable factors and testing the effect of PJ under similar experimental conditions.
Assuntos
Sucos de Frutas e Vegetais/análise , Hipoglicemiantes/farmacologia , Polifenóis/farmacologia , Punica granatum/química , Animais , Dipeptidil Peptidase 4/sangue , Humanos , Insulina/sangue , Compostos Fitoquímicos/farmacologia , Ensaios Clínicos Controlados Aleatórios como Assunto , Ratos , alfa-Glucosidases/sangueRESUMO
A turn-on ratiometric fluorescent assay is described for the determination of the activity of enzymes participating in ascorbic acid-forming reactions. Blue-emitting carbon dots (bCDs; with excitation/emission wavelength at 380/450 nm) serve as fluorescent indicator. Their fluorescence is reduced by Fe3+ ions via an inner filter effect. Yellow-emitting CDs (yCDs; with excitation/emission wavelength at 380/550 nm) serve as internal reference because their fluorescence is insensitive to Fe3+. Upon exposure to ascorbic acid (AA), Fe3+ is reduced to Fe2+. Hence, the fluorescence of the bCDs is restored. Thus, enzymes participating in AA-related reactions such as α-glucosidase (α-Glu) and alkaline phosphatase (ALP) can be determined. α-Glu activity was quantified in the range from 0.13 to 6.70 U mL-1, and ALP activity was determined between 2.0 and 130 U L-1. Endowed with excellent sensitivity, selectivity and low background signals, the method may also be used to screen the inhibitors of α-Glu and ALP. Graphical abstractSchematic representation of a redox modulated ratiometric fluorometric method based on the use of dual-color carbon dots for determination of the activity of enzymes participating in ascorbic acid-related reactions. Blue-emitting carbon dots (bCDs) serve as fluorescent indicator while yellow-emitting CDs (yCDs) serve as internal reference.
Assuntos
Fosfatase Alcalina/metabolismo , Ácido Ascórbico/metabolismo , Carbono/química , Cor , Fluorometria , Pontos Quânticos/química , alfa-Glucosidases/metabolismo , Fosfatase Alcalina/antagonistas & inibidores , Fosfatase Alcalina/sangue , Ácido Ascórbico/química , Humanos , Oxirredução , Tamanho da Partícula , Propriedades de Superfície , alfa-Glucosidases/sangue , alfa-Glucosidases/químicaRESUMO
α-Glucosidase and its inhibitors play a key role in diagnosis and treatment of diabetes. In the present work, we established a facile, sensitive and selective fluorescence method based on silicon quantum dots (SiQDs) and MnO2 nanosheets for the determination of α-glucosidase and one of its inhibitors acarbose. The fluorescence of SiQDs was greatly quenched by MnO2 nanosheets due to the inner filter effect. α-Glucosidase could easily catalyze the hydrolysis of l-ascorbic acid-2-O-α-d-glucopyranosyl (AAG) to produce ascorbic acid (AA), which could reduce MnO2 nanosheets to Mn2+, resulting in dramatic recovery of the fluorescence of SiQDs. The proposed sensing platform could provide a good linear relationship between the fluorescence intensity of SiQDs and the concentration of α-glucosidase in the range of 0.02-2.5 U mL-1 with a detection limit of 0.007 U mL-1. In addition, the sensing platform could be used for α-glucosidase inhibition. Acarbose was one of the most common and typical inhibitors, and this sensing platform can be utilized to detect acarbose in the range of 1-1000 µM. The developed fluorescence method was successfully validated for the determination of α-glucosidase in human serum samples.
Assuntos
Corantes Fluorescentes/química , Inibidores de Glicosídeo Hidrolases/análise , Nanocompostos/química , Pontos Quânticos/química , alfa-Glucosidases/sangue , Humanos , Compostos de Manganês/química , Óxidos/química , Silício/química , Espectrometria de FluorescênciaRESUMO
Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide. Interestingly, the great majority of individuals affected by the tumor have underlying liver disease, therefore narrowing the population to be screened. Still, however, there is a clear lack of blood biomarkers, and surveillance in those at risk is performed by frequent imaging of the liver. A variety of multinational collaborations are currently invested in finding biomarkers for HCC based on liver-produced proteins. A new approach with assessment of peripheral proteins might be necessary for the successful early detection of this malignancy.
Assuntos
Biomarcadores Tumorais/sangue , Carcinoma Hepatocelular/diagnóstico , Neoplasias Hepáticas/diagnóstico , Biomarcadores/sangue , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/sangue , Detecção Precoce de Câncer , Glipicanas/sangue , Humanos , Cirrose Hepática , Neoplasias Hepáticas/sangue , Precursores de Proteínas/sangue , Protrombina , Sensibilidade e Especificidade , Ultrassonografia , alfa-Fetoproteínas/metabolismo , alfa-Glucosidases/sangueRESUMO
OBJECTIVE: Pu-erh tea was presumed to have anti-hyperglycemic effects via inhibition on alpha-amylase and alpha-glucosidase. However, no integerated literatures were published to substantiate such presumption. METHODS: Current study adopted systemic review method to validate inhibitory effects on alpha amylase and alpha-glucosidase. Five English databases (PubMed, EBSCO, SCOPUS, Cochrane Library, Web of Science) and three Chinese ones (Airti Library, CNKI Library, and Google Scholar) were searched up to 22 March 2018 for eligible literatures, using keywords of Pu-erh, Pu'er, alpha-amylase or alpha-glucosidase. RESULTS: Six studies exploring inhibitory effects on alpha-glucosidase and seven on alpha-amylase were included for systemic review. Though results showed pu-erh tea has significant inhibitory effects on alpha-amylase and alpha-glucosidase, high heterogeneity was detected among studies included. CONCLUSIONS: High heterogeneity may be due to complex alterations of chemicals under different degrees of fermentation. More future studies are required to further identify principal bioactive component(s) at work.
Assuntos
Chá , alfa-Amilases/sangue , alfa-Glucosidases/sangue , Humanos , Extratos VegetaisRESUMO
A sensitive nanocomplex probe prepared from fluorescent polydopamine nanoparticles (F-PDA) and cobalt oxyhydroxide (CoOOH) nanosheets was established for the determination of α-glucosidase activity. In this detection system, the fluorescence of F-PDA was firstly quenched by CoOOH nanosheets based on fluorescence resonance energy transfer (FRET). Subsequently, ascorbic acid was produced from 2-O-α-d-glucopyranosyl-l-ascorbic acid which was selectively hydrolyzed by α-glucosidase. CoOOH was reduced to Co2+ by the released ascorbic acid, which resulted in the recovery of F-PDA nanoparticles fluorescence. In consequence, α-glucosidase activity was determined by the fluorescence recovery degree of the F-PDA nanoparticle. This fluorescent method showed a good linear relationship with the activity of α-glucosidase from 2 to 80 Uâ¯L-1 and low detection limit of 1.65 Uâ¯L-1â¯(S/Nâ¯=â¯3). This fluorescence probe with high selectivity and sensitivity demonstrated a remarkable applicability in human serum samples and provided an alternative for α-glucosidase inhibitors screening in the discovery of anti-diabetes drugs.
Assuntos
Cobalto/química , Corantes Fluorescentes/química , Indóis/química , Nanopartículas/química , Óxidos/química , Polímeros/química , alfa-Glucosidases/sangue , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/química , Ensaios Enzimáticos/métodos , Fluorescência , Transferência Ressonante de Energia de Fluorescência/métodos , Humanos , Hidrólise , Limite de Detecção , OxirreduçãoRESUMO
Pompe disease is a rare inherited disorder of lysosomal glycogen metabolism due to acid α-glucosidase (GAA) deficiency. Enzyme replacement therapy (ERT) using alglucosidase alfa, a recombinant human GAA (rhGAA), is the only approved treatment for Pompe disease. Although alglucosidase alfa has provided clinical benefits, its poor targeting to key disease-relevant skeletal muscles results in suboptimal efficacy. We are developing an rhGAA, ATB200 (Amicus proprietary rhGAA), with high levels of mannose-6-phosphate that are required for efficient cellular uptake and lysosomal trafficking. When administered in combination with the pharmacological chaperone AT2221 (miglustat), which stabilizes the enzyme and improves its pharmacokinetic properties, ATB200/AT2221 was substantially more potent than alglucosidase alfa in a mouse model of Pompe disease. The new investigational therapy is more effective at reversing the primary abnormality - intralysosomal glycogen accumulation - in multiple muscles. Furthermore, unlike the current standard of care, ATB200/AT2221 dramatically reduces autophagic buildup, a major secondary defect in the diseased muscles. The reversal of lysosomal and autophagic pathologies leads to improved muscle function. These data demonstrate the superiority of ATB200/AT2221 over the currently approved ERT in the murine model.
Assuntos
Terapia de Reposição de Enzimas/métodos , Doença de Depósito de Glicogênio Tipo II/tratamento farmacológico , alfa-Glucosidases/farmacologia , alfa-Glucosidases/uso terapêutico , 1-Desoxinojirimicina/análogos & derivados , Animais , Modelos Animais de Doenças , Feminino , Glicogênio/metabolismo , Doença de Depósito de Glicogênio Tipo II/genética , Doença de Depósito de Glicogênio Tipo II/patologia , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Manosefosfatos/metabolismo , Camundongos , Camundongos Knockout , Músculo Esquelético/metabolismo , Ratos , Ratos Sprague-Dawley , alfa-Glucosidases/sangue , alfa-Glucosidases/genéticaRESUMO
Pompe disease is an inherited disease caused by acid alpha-glucosidase (GAA) deficiency. A single center observational study aimed at assessing the prevalence of late-onset Pompe disease in a high-risk Brazilian population, using the dried blood spot test to detect GAA deficiency as a main screening tool. Dried blood spots were collected for GAA activity assay from 24 patients with "unexplained" limb-girdle muscular weakness without vacuolar myopathy in their muscle biopsy. Samples with reduced enzyme activity were also investigated for GAA gene mutations. Of the 24 patients with dried blood spots, one patient (4.2%) showed low GAA enzyme activity (NaG/AaGIA: 40.42; %INH: 87.22%). In this patient, genetic analysis confirmed two heterozygous mutations in the GAA gene (c.-32-13T>G/p.Arg854Ter). Our data confirm that clinicians should look for late-onset Pompe disease in patients whose clinical manifestation is an "unexplained" limb-girdle weakness even without vacuolar myopathy in muscle biopsy.
Assuntos
Doença de Depósito de Glicogênio Tipo II/diagnóstico , Distrofia Muscular do Cíngulo dos Membros/sangue , Distrofia Muscular do Cíngulo dos Membros/diagnóstico , alfa-Glucosidases/sangue , Adulto , Biópsia , Feminino , Doença de Depósito de Glicogênio Tipo II/sangue , Doença de Depósito de Glicogênio Tipo II/patologia , Humanos , Masculino , Distrofia Muscular do Cíngulo dos Membros/patologia , PrevalênciaRESUMO
BACKGROUND: Diagnosis of Pompe disease is sometimes challenging because it exhibits clinical similarities to muscular dystrophy. CASE: We describe a case of Becker muscular dystrophy (BMD) with a remarkable reduction in activity of the acid α-glucosidase (GAA) enzyme, caused by a combination of pathogenic mutation and polymorphism variants resulting in pseudodeficiency in GAA. The three-year-old boy demonstrated asymptomatic creatine kinase elevation. Neither exon deletion nor duplication was detected on multiplex ligation-dependent probe amplification (MLPA) of DMD. GAA enzyme activity in both dried blood spots and lymphocytes was low, at 11.7% and 7.7% of normal, respectively. However, genetic analysis of GAA detected only heterozygosity for a nonsense mutation (c.118Câ¯>â¯T, p.Arg40∗). Muscle pathology showed no glycogen deposits and no high acid phosphatase activity. Hematoxylin-eosin staining detected scattered regenerating fibers; the fibers were faint and patchy on immunochemistry staining of dystrophin. The amount of dystrophin protein was reduced to 11.8% of normal, on Western blotting analysis. Direct sequencing analysis of DMD revealed hemizygosity for a nonsense mutation (c.72Gâ¯>â¯A, p.Trp24∗). The boy was diagnosed with BMD, despite remarkable reduction in GAA activity; further, he demonstrated heterozygosity for [p.Gly576Ser; p.Glu689Lys] polymorphism variants that indicated pseudodeficiency on another allele in GAA. CONCLUSIONS: Pseudodeficiency alleles are detected in approximately 4% of the Asian population; these demonstrate low activity of acid α-glucosidase (GAA), similar to levels found in Pompe disease. Clinicians should be careful in their interpretations of pseudodeficiency alleles that complicate diagnosis in cases of elevated creatine kinase.
Assuntos
Creatina Quinase/sangue , Distrofia Muscular de Duchenne/enzimologia , alfa-Glucosidases/sangue , Pré-Escolar , Diagnóstico Diferencial , Distrofina/genética , Distrofina/metabolismo , Humanos , Masculino , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patologia , alfa-Glucosidases/genéticaRESUMO
ABSTRACT Pompe disease is an inherited disease caused by acid alpha-glucosidase (GAA) deficiency. A single center observational study aimed at assessing the prevalence of late-onset Pompe disease in a high-risk Brazilian population, using the dried blood spot test to detect GAA deficiency as a main screening tool. Dried blood spots were collected for GAA activity assay from 24 patients with "unexplained" limb-girdle muscular weakness without vacuolar myopathy in their muscle biopsy. Samples with reduced enzyme activity were also investigated for GAA gene mutations. Of the 24 patients with dried blood spots, one patient (4.2%) showed low GAA enzyme activity (NaG/AaGIA: 40.42; %INH: 87.22%). In this patient, genetic analysis confirmed two heterozygous mutations in the GAA gene (c.-32-13T>G/p.Arg854Ter). Our data confirm that clinicians should look for late-onset Pompe disease in patients whose clinical manifestation is an "unexplained" limb-girdle weakness even without vacuolar myopathy in muscle biopsy.
RESUMO A doença de Pompe é uma doença hereditária causada pela deficiência da enzima alfa-glicosidase ácida (GAA). Estudo observacional foi realizado, em um único centro, para determinar a prevalência da doença de Pompe de início tardio (LOPD) em uma população brasileira de alto risco, usando teste em gota seca (DBS) como ferramenta principal de triagem para detectar a deficiência da GAA. DBS foi coletado para avaliar a atividade da GAA em 24 pacientes com fraqueza muscular de cinturas "não explicada" sem miopatia vacuolar na biópsia muscular. As amostras com atividade enzimática reduzida foram também submetidas a análise de mutações no gene GAA. Dos 24 pacientes com DBS, baixa atividade da enzima GAA (NaG/AaGIA: 40.42; %INH: 87.22%) foi encontrada em um paciente (4.2%). Nessa paciente, a análise genética confirmou duas mutações em heterozigose composta no gene GAA (c.-32-13T > G/p.Arg854Ter). Nossos resultados confirmam que LOPD deve ser investigada quando a manifestação clínica é uma fraqueza muscular de cinturas "não explicada", mesmo na ausência de miopatia vacuolar na biópsia muscular.