RESUMO
In this work, a novel, rapid and enzyme-free colorimetric biosensor for the detection of nucleic acids has been developed based on entropy-driven (EDC) circuit and DNAzyme-mediated autocatalytic reactions. On sensing the target DNA, the EDC reaction could be initiated and the intact Mg2+-dependent DNAzyme was formed in the reaction product; then, a "mimic target" DNA was generated during the cleavage process of DNAzyme, which in turn catalyzed the EDC reaction corresponding to an autocatalytic process. Meanwhile, numerous G-quadruplex structures were released and further interacted with hemin to form peroxidase-mimicking DNAzyme, inducing a remarkably amplified colorimetric signal. This autocatalytic EDC (AEDC) sensing system exhibited a linear relationship in the range from 20 pM to 10 nM with a detection limit of 10.2 pM. More importantly, benefitting from the Mg2+-dependent DNAzyme-mediated autocatalytic reaction, the detection time (20 min) was significantly reduced compared to that for the reported EDC strategies. In addition, this sensing system has been applied for the detection of target DNA in human serum samples, indicating that it is promising for the on-site and real-time detection of nucleic acids in biomedical research and disease diagnosis.
Assuntos
DNA Catalítico , Quadruplex G , Colorimetria , DNA Catalítico/metabolismo , Entropia , Hemina , HumanosRESUMO
Glutaraldehyde (GLUT) crosslinked bioprosthetic heart valves (BHVs) might fail within ten years due to progressive degradation and calcification. GLUT cannot stabilize glycosaminoglycans (GAGs), one important component of BHVs. In this current study we developed a new BHVs preparation strategy named as 'HPA/NT/HRP' treatment that utilized 3,4-hydroxyphenylpropionic acid (HPA)/tyramine (TRA)/neomycin trisulfate (NT) conjugated pericardiums and horseradish peroxidase (HRP)/H2O2 enzyme-oxidative-polymerization method. HPA/TRA-pericardium and HPA-NT conjugation would provide extra phenol groups for enzymatic crosslinking. HPA/TRA conjugated pericardium could be crosslinked by HRP/H2O2 enzyme-oxidative-polymerization. The feeding ratio of HPA-NT was optimized. The GAGs content, collagenase and elastase degradation in vitro, the in vivo GAGs stability and anti-calcification potential of HPA/NT/HRP treated pericardiums were characterized. We demonstrated that HPA/NT/HRP treated pericardiums had sufficiently increased GAGs stabilization and decreased calcification. This new HPA/NT/HRP enzyme-oxidative-polymerization strategy would be a promising method to make BHVs with better GAGs stability and anti-calcification properties.
Assuntos
Bioprótese , Calcinose , Glicosaminoglicanos/química , Próteses Valvulares Cardíacas , Oxigênio/química , Animais , Valva Aórtica , Ácidos Cafeicos/química , Calcificação Fisiológica , Cálcio/química , Varredura Diferencial de Calorimetria , Colagenases/química , Enzimas/química , Hexosaminas/química , Peroxidase do Rábano Silvestre/química , Humanos , Peróxido de Hidrogênio/química , Hidroxiprolina/química , Neomicina/química , Pericárdio/metabolismo , Pericárdio/patologia , Polímeros/química , Ratos , Ratos Sprague-Dawley , Estresse Mecânico , Suínos , Temperatura , Resistência à Tração , Tiramina/químicaRESUMO
Valvular heart diseases lead to over 300,000 heart valve replacements worldwide each year. Commercially available bioprosthetic heart valves (BHVs) are mostly made from porcine or bovine pericardiums which were crosslinked by glutaraldehyde (GLUT). However, valve failures can occur within 10 years due to progressive degradation and calcification. GLUT could crosslink collagen but it fails to stabilize elastin. In this current study, we developed a new BHVs preparation strategy named as "GLUT/TE/LOXL/EGCG" that utilizes exogenous tropoelastin (TE)/lysyl oxidase (LOXL) and epigallocatechin gallate (EGCG) to increase the elastin content as well as the stabilization of elastin. The feeding ratios of tropoelastin and lysyl oxidase were optimized. The contents of desmosine and insoluble elastin, biomechanics, cytotoxicity, hemocompatibility, in vivo componential stability and anti-calcification potential were characterized. Pericardiums with increased elastin content had improved the mechanical properties. GLUT/TE/LOXL/EGCG-treated pericardiums had similar cytotoxicity and coagulation properties compared to GLUT and GLUT/EGCG control. We demonstrated that GLUT/TE/LOXL/EGCG-treated pericardiums had high amount of insoluble elastin in 90 days' rat subdermal implantation model, and better resistance for calcification. This new tropoelastin and lysyl oxidase treatments strategy would be a promising method to make BHVs which have better structural stability and anti-calcification properties.
