RESUMO
Objective: To develop a detection method for single nucleotide polymorphisms (SNPs) of bronchial asthma (BA) susceptibility genes (IL-13, IL-33, and GSDMA) based on fluorescence PCR melting curves.Methods: Peripheral blood samples from 33 patients with BA were collected. DNA was extracted, and positive plasmids were constructed. Probes and primers for fluorescence polymerase chain reaction (PCR) were designed according to IL-13, IL-33, and GSDMA sequences, and the SNPs were separately detected by gene sequencing and fluorescence PCR melting curve.Results: The system was successfully divided into 3 SNPs, including IL-13, IL-33, and GSDMA, and a comparison of sequencing methods showed that the results were completely consistent. The lowest detection limit was 1 ng/reaction, the sensitivity and specificity were 100%, and this method had high repeatability (CV = 2.8%).Conclusion: The fluorescence PCR melting curve method is suitable for the rapid and accurate classification of SNPs. The method is economical, simple, and efficient, and is suitable for the screening of the susceptible gene SNPs in a large-scale population of patients with BA.
Assuntos
Asma/diagnóstico , Testes Genéticos/métodos , Técnicas de Genotipagem/métodos , Programas de Rastreamento/métodos , Reação em Cadeia da Polimerase/métodos , Adolescente , Adulto , Asma/sangue , Asma/genética , Estudos de Viabilidade , Feminino , Fluorescência , Predisposição Genética para Doença , Técnicas de Genotipagem/economia , Humanos , Interleucina-13/genética , Interleucina-33/genética , Masculino , Programas de Rastreamento/economia , Proteínas de Neoplasias/genética , Reação em Cadeia da Polimerase/economia , Polimorfismo de Nucleotídeo Único , Sensibilidade e Especificidade , Adulto JovemRESUMO
Biopharmaceutics classification system of Chinese materia medica (CMMBCS) emphasizes characteristic of the multi-component environment based on the drug solubility and permeability. In this study, the in situ closed-loop method combined with LC-MS technique was utilized to study the intestinal absorption and metabolism of Puerariae Lobatae Radix decoction (PLRD), providing selection basis for intestinal permeability components in CMMBCS. A total of 36 components were identified from PLRD. Among them, 17 components could be detected in the plasma sample, indicating that 17 components could be absorbed into blood, so these 17 components could be used as intestinal permeability evaluation components in CMMBCS. The other 19 components were not detected in the plasma sample, suggesting that they may not be absorbed or metabolized by the gut wall enzymes.
RESUMO
<p><b>AIM</b>To study the regular pattern and mechanism of positive inotropic effect after washout of ACh (rebound of myocardial contractile force) in isolated rabbit hearts.</p><p><b>METHODS</b>The changes of myocardial contractile force after perfusion and washout of ACh were observed in isolated Langendorff perfused rabbit hearts.</p><p><b>RESULTS</b>Maximum rebound rate induced by ACh of 10(-8)-10(-3) mol/L were 2.20% +/- 1.70%, 6.71% +/- 3.40%, 9.18% +/- 3.54%, 14.16% +/- 3.27%, 4.37% +/- 5.86% and 1.03% +/- 6.86%, respectively. Compared with the ACh of 10(-5) mol/L in control group, adrenaline enhanced rebound of myocardial contractile force, maximum rebound rate in adrenaline group was 29.25% +/- 5.83% (P < 0.05), propranolol reduced rebound, and maximum rebound rate in propranolol group was 5.15% +/- 4.45% (P < 0.05), we had not detected rebound of myocardial contractile force in 800 s after addition ACh in verapamil group.</p><p><b>CONCLUSION</b>In isolated rabbit heart, positive inotropic effect after washout of ACh has relevance to the activities of calcium current channel and beta-adrenergic receptor. Perhaps there are some different aspects in the mechanism of positive inotropic effect between perfusion of high concentration and after washout of ACh.</p>
