[Changes and significance of oxidized phospholipids and endothelial nitric oxide synthase in the acute stage of Kawasaki disease]. / å·å´Žç— æ€¥æ€§æœŸæ°§åŒ–ç£·è„‚å’Œå† çš®ä¸€æ°§åŒ–æ°®åˆé ¶çš„å˜åŒ–åŠæ„ä¹‰.

OBJECTIVES: To investigate the changes in the serum levels of oxidized phospholipids (OxPLs) and endothelial nitric oxide synthase (eNOS) and their association with coronary artery disease (CAL) in children in the acute stage of Kawasaki disease (KD), as well as the clinical significance of OxPLs and eNOS. METHODS: A prospective study was conducted on 95 children in the acute stage of KD (KD group). According to the presence of absence of CAL, the KD group was further divided into a CAL subgroup and a non-CAL (NCAL) subgroup. Thirty children with fever due to lower respiratory tract infection were enrolled as the fever group. Thirty healthy children who underwent physical examination were enrolled as the healthy control group. The above groups were compared in terms of general information and serum levels of OxPLs, eNOS and other laboratory indexes, and the correlation between OxPLs level and eNOS level was analyzed. RESULTS: The KD group had a significantly higher level of OxPLs and a significantly lower level of eNOS compared with the fever group and the healthy control group (P<0.05). After treatment, the children with KD had a significantly decreased OxPLs level and a significantly increased eNOS level (P<0.05). Compared with the NCAL subgroup, the CAL subgroup had a significantly higher level of OxPLs and a significantly lower level of eNOS (P<0.05). Among the children of KD, the level of OxPLs was negatively correlated with that of eNOS (rs=-0.353, P<0.05). CONCLUSIONS: Serum OxPLs and eNOS in the acute stage of KD may be involved in the development of CAL in children with KD, and therefore, they may be used as the biomarkers to predict CAL in these children.

Current career situations of Chinese pharmacovigilance professionals working for pharmaceutical companies: an exploratory survey.

BACKGROUND: Pharmacovigilance in China has experienced rapid development in the past 30 years. The implementation of Good Pharmacovigilance Practice in China since the end of 2021 heralds a new era of pharmacovigilance affairs, which puts forward higher requirements for the quantity and quality of pharmacovigilance personnel. This study aimed to preliminarily explore the current career situations of pharmacovigilance professionals working in China for pharmaceutical companies. METHODS: A questionnaire was adapted from research in the USA and Europe with the help of several pharmacovigilance experts. Snowball sampling was used to conduct an exploratory survey to obtain the frequency of basic demographic information, work status, and career expectations of pharmacovigilance professionals working for pharmaceutical companies. RESULTS: The personnel engaged in pharmacovigilance work for pharmaceutical companies were mainly medical or pharmaceutical undergraduates within 3 years of graduation. Their work intensity and pressure were relatively high. The training provided by their universities and enterprises could not well meet their needs to improve their job competence. Although they were optimistic about pharmacovigilance and will not change their career, most of them were planning to change their employers. CONCLUSION: There was a gap between the demand and supply of pharmacovigilance personnel. Relevant regulatory authorities and industry associations should guide higher education institutions to collaborate with pharmacovigilance specialists to strengthen pharmacovigilance education for medical or pharmaceutical students, on the basis of which pharmacovigilance certification courses and continuing education courses can be developed. Meanwhile, pharmaceutical enterprises should consider reasonably adjusting work intensity and income to avoid a high turnover rate.

A new biomimetic nanozyme of hemin/graphdiyne oxide with superior peroxidase-like activity for colorimetric bioassays.

Graphdiyne oxide (GDYO) is a novel type of two-dimensional carbon allotrope nanomaterial consisting of a large conjugated system and excellent chemical stability. To date, application of GDYO as a nanozyme in biosensing has been rarely reported. In this study, a novel ultrasensitive colorimetric bioassay was constructed using a hemin/GDYO nanocomposite (H/GDYO) as a new nanozyme with superior peroxidase-like activity for the detection of H2O2 and glucose. It was discovered that H/GDYO exhibited 6-fold higher peroxidase-like activity than pure hemin. Catalytic kinetic analysis showed that H/GDYO had a much higher affinity for H2O2 and glucose than that of hemin. The designed colorimetric bioassay displayed excellent sensitivity for H2O2 and glucose detection with a wide linear range of 0.015-0.5 mM and 0.1-10 mM, respectively, while the limit of detection (LOD) was as low as 4.39 µM and 38 µM, respectively. Moreover, it was successfully applied for the analysis of H2O2 in milk and glucose in real human serum samples with acceptable recoveries. Importantly, the developed colorimetric bioassay shows good agreement with the results obtained from a commercial blood glucose meter. We believe that the proposed method could provide a promising prospect for medical diagnosis and biotechnology.

Electrochemical aptasensor for ultrasensitive detection of lipopolysaccharide using silver nanoparticles decorated titanium dioxide nanotube/functionalized reduced graphene oxide as a new redox nanoprobe.

A novel and relatively simple signal-off electrochemical aptasensor was constructed for highly sensitive detection of lipopolysaccharide (LPS). For the first time, silver nanoparticles (AgNPs) decorated titanium dioxide nanotube (TNT) was conjugated with polydiallyldimethylammonium chloride (PDDA) functionalized reduced graphene oxide (rGO) to form a new nanohybrid of Ag-TNT/P-rGO. This nanohybrid with a large specific surface area exhibited excellent electrochemical activity, which not only served as the sensing platform to immobilize LPS binding aptamer (LBA) but was also employed as the redox probe to monitor the change of the electrochemical signal. The electrochemical signal responses were measured by cyclic voltammetry (CV) in the potential range -0.3 to 0.5 V at a scan rate of 0.1 V/s. The proposed aptasensor exhibited acceptable stability, reproducibility, and specificity for LPS detection with a wide linear range from 17 fg/mL to 100 ng/mL. The limit of detection (LOD) was 5 fg/mL. Furthermore, the prepared aptasensor showed acceptable recovery ranging from 96% to 103%, and the RSD varied between 1.4% and 8.5% for determining LPS in real samples.Graphical abstract.

Tumour necrosis factor-α and myoglobin associated with the recovery time of coronary artery lesions in Kawasaki disease patients.

AIM: To assess the relationship between clinical parameters and medium term recovery time of coronary artery lesions (CALs). METHODS: In total, 344 Kawasaki disease patients were screened and 311 Kawasaki disease patients were included and followed-up for the next 2 years. Clinical records, clinical parameters and inflammatory biomarkers were collected for all subjects. RESULTS: Tumour necrosis factor (TNF)-α and myoglobin (MYO) levels in patients without recovery from CALs were significantly higher than those without CALs and with recovery from CALs. Kaplan-Meier survival analysis showed that in the high-TNF-α group, the estimated median time to recovery (5.0 months, 95% confidence interval (CI) 1.436-8.564) is significantly longer than the low-TNF-α group (2.00 months, 95% CI: 0.633-3.367, P = 0.044). Also, the estimated median time (5.0 months, 95% CI: 1.836-8.164) in the high-MYO group is significantly longer than the low-MYO group (2.00 months, 95% CI: 0.405-3.595, P = 0.002). Cox regression analysis showed independent factors for recovery of CALs included age, left coronary artery to aortic annulus ratio, TNF-α and MYO levels. CONCLUSIONS: These findings suggest that clinical parameters such as age, left coronary artery to aortic annulus ratio, TNF-α and MYO levels associate with medium term recovery time of CALs and could help in the design of a clinical strategy for the surveillance and prevention of late cardiovascular events.

An aptamer based voltammetric biosensor for endotoxins using a functionalized graphene and molybdenum disulfide composite as a new nanocarrier.

Lipopolysaccharides (LPS), known as endotoxins, can cause a strong inflammatory response and lead to multiple organ failure in severe cases. This work reports a simple label-free voltammetric aptasensor for highly sensitive determination of LPS using a polyethyleneimine (PEI) functionalized reduced graphene oxide (rGO) and molybdenum disulfide (MoS2) composite (PEI-rGO-MoS2) as a new nanocarrier for electroactive toluidine blue (TB). The PEI-rGO-MoS2 nanocomposite with high electrical conductivity and large specific surface area can greatly increase the loading of TB and facilitate electron transfer from TB to an electrode. Then gold nanoparticles (AuNPs) were utilized to immobilize a thiolated LPS binding aptamer (LBA), which not only exhibited excellent biocompatibility, but also significantly amplified the electrochemical signal of TB. The proposed aptasensor exhibited high sensitivity for LPS and showed a wide linear range from 5.0 × 10-5 ng mL-1 to 2.0 × 102 ng mL-1 with a low limit of detection (LOD) of 3.01 × 10-5 ng mL-1, which overcame the shortcomings of traditional detection methods and achieved fast and accurate detection of LPS. Moreover, it exhibited excellent recovery and specificity upon spiking LPS in serum samples, indicating that this method has promising application in the field of trace analysis of LPS in clinical detection.

Adherence to Adjuvant Imatinib Therapy in Patients with Gastrointestinal Stromal Tumor in Clinical Practice: A Cross-Sectional Study.

BACKGROUND: Adherence to imatinib therapy has been significantly associated with disease progression and direct medical costs in gastrointestinal stromal tumor (GIST) patients. However, adherence to oral anticancer drugs is frequently hindered by the influence of various factors. The aim of this study was to evaluate the prevalence of imatinib adherence and its influencing factors among GIST patients in the adjuvant setting. METHODS: Adherence of GIST patients (receiving imatinib for ≥1 month) was assessed using the 8-item Morisky Medication Adherence Scale (MMAS), with a score <8 indicating nonadherence. Quality of life and social support were evaluated by the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire C30 (EORTC QLQ C30) and Social Support Rating Scale (SSRS). Factors associated with nonadherence were identified by multivariate logistic regression analysis. Imatinib plasma concentrations were determined and compared between adherent and nonadherent groups. RESULTS: A total of 158 GIST patients were enrolled, 92 (58.2%) patients were considered nonadherent. Intentional nonadherence, especially feeling hassled by treatment plan (34.2% of patients), was common. In the multivariate logistic regression analysis, gender (OR 2.68, 95% CI 1.33-5.41; p = 0.0058), place of residence (OR 3.20, 95% CI 1.39-7.35; p = 0.0061), and global health status (OR 1.02, 95% CI 1.00-1.04; p = 0.0378) were significantly associated with nonadherence. Moreover, imatinib plasma concentrations in nonadherent patients were significantly lower than that in the good adherence group (p = 0.0338). CONCLUSIONS: Poor adherence to imatinib is a notable problem in Chinese GIST patients in the adjuvant therapy setting. The predominant indicators of nonadherence in this study were gender (female), living in a rural area, and harboring a low global health status score. These indicators may aid clinicians in determining where increased efforts in promoting adherence may be beneficial.

Risk Factors for New-Onset Diabetes Mellitus after Heart Transplantation in Chinese Patients: A Single Center Experience.

BACKGROUND: New-onset diabetes after transplantation -(NODAT) is a frequent complication after heart transplantation (HT) and is associated with graft loss and patient survival. OBJECTIVES: The aim of this study was to identify the incidence and associated factors contributing to NODAT in Chinese heart transplant recipients. METHODS: Adult patients without diabetes mellitus before HT were enrolled in this study. NODAT was diagnosed according to the criteria recommended by the American Diabetes Association. The cumulative incidence was determined at 3, 6, and 12 months, respectively. The risk factors of NODAT were estimated by logistic regression analysis. RESULTS: A total of 154 adults who first received HT were included. Among them, 50 (32.5%) recipients were diagnosed with NODAT after a median follow-up time of 611 days. The cumulative incidence of NODAT was 27.3% at 3 months, 29.9% at 6 months, and 30.5% at 12 months, respectively. Independent risk factors for NODAT included age ≥45 years (OR 3.82, 95% CI 1.57-9.31; p = 0.003), hypertension (OR 3.28, 95% CI 1.17-9.20; p= 0.024), and transient hyperglycemia (OR 12.13, 95% CI 3.35-43.92; p < 0.001). Moreover, recipients treated with both acarbose and insulin for transient hyperglycemia had a significantly higher prevalence of NODAT than those without any anti-diabetic agents (OR 5.35, 95% CI 1.21-23.64; p = 0.027). CONCLUSIONS: Age ≥45 years, hypertension, transient hyperglycemia, and associated treatment strategies are imperative to identify recipients at high risk of developing -NODAT.

[Role of apoptosis signal-regulating kinase 1 in left ventricular remodeling in mice].

OBJECTIVE: To study the changes and significance of apoptosis signal-regulating kinase 1 (ASK1) in left ventricular remodeling in FVB/N mice. METHODS: A total of 54 FVB/N mice were randomly divided into 4 groups: 0 d group with 8 mice, 7 d group with 10 mice, 14 d group with 16 mice, and 21 d group with 20 mice. A model of cardiac remodeling was established by intraperitoneal injection of isoproterenol (ISO) at a daily dose of 30 mg/kg, and the 7 d, 14 d, and 21 d groups were injected for 7, 14, and 21 consecutive days respectively. The 0 d group was given intraperitoneal injection of an equal volume of normal saline. Echocardiography was used to measure left ventricular posterior wall thickness at end diastole (dLVPW) and the ratio of heart weight to tibia length (HW/TL) was measured. Hematoxylin-eosin staining was used to measure left ventricular myocardial fiber diameter. Picric-Sirius red staining was used to measure myocardial collagen deposition area in the left ventricle. Quantitative real-time PCR was used to measure the mRNA expression of ASK1, type I collagen (collagen I), and B-type natriuretic peptide (BNP). The mortality rate was observed for each group. RESULTS: There were gradual increases in HW/TL, myocardial fiber diameter, and dLVPW after 0, 7, and 14 days of ISO injection (P<0.05). There were no significant changes in HW/TL ratio and dLVPW from days 14 to 21 of ISO injection (P>0.05), while there was a significant reduction in myocardial fiber diameter (P<0.05), which was similar to the value on day 7 (P>0.05). There were significant increases in myocardial collagen deposition area and the mRNA expression of collagen I, ASK1, and BNP after 0, 7, 14, and 21 days of ISO injection, which reached the peaks on day 21 (P<0.01). The mRNA expression of ASK1 was positively correlated with myocardial collagen deposition area and the mRNA expression of collagen I and BNP and had a weak correlation with HW/TL, myocardial fiber diameter, and dLVPW. There was a significant increase in the mortality rate of the mice over the time of ISO injection. CONCLUSIONS: The expression of ASK1 in the myocardium is closely associated with left ventricular remodeling. The increase of ASK1 expression may lead to the aggravation of left ventricular remodeling, and the mechanism of which needs further study.

[Expression of Nod-like receptor protein 3 inflammasome in peripheral blood mononuclear cells of children with Kawasaki disease in the acute stage].

OBJECTIVE: To study the association of Nod-like receptor protein 3 (NLRP3) inflammasome with inflammatory response in the acute stage and coronary artery lesion (CAL) in children with Kawasaki disease (KD). METHODS: A total of 42 children with KD who were hospitalized from January to October 2017 were enrolled as the KD group, among whom 9 had CAL (CAL group) and 33 had no CAL (NCAL group). Fifteen age- and gender-matched children with pneumonia and pyrexia were enrolled as the pneumonia-pyrexia group. Fifteen healthy children were enrolled as the healthy control group. Real-time PCR was used to measure the mRNA expression of NLRP3 inflammasome (NLRP3, ASC and caspase-1) in peripheral blood mononuclear cells. The Spearman rank correlation test was used to investigate the correlation of NLRP3 mRNA expression with serum levels of C-reactive protein, erythrocyte sedimentation rate, interleukin-6, interleukin-1ß, procalcitonin, albumin and prealbumin. RESULTS: The KD group had significantly higher mRNA expression of NLRP3, ASC and caspase-1 in the acute stage than the pneumonia-pyrexia and healthy control groups (P<0.05). The CAL group had significantly higher mRNA expression of NLRP3 than the NCAL group (P<0.05). NLRP3 mRNA expression was correlated with C-reactive protein, interleukin-6, interleukin-1ß, and prealbumin levels in children with KD in the acute stage (rs=0.449, 0.376, 0.427, and -0.416 respectively; P<0.05). CONCLUSIONS: NLRP3 inflammasome may participate in inflammatory response in the acute stage and the development of CAL in children with KD.

Aptamer based voltammetric biosensor for Mycobacterium tuberculosis antigen ESAT-6 using a nanohybrid material composed of reduced graphene oxide and a metal-organic framework.

The 6-kDa early secretory antigenic target referred to as ESAT-6 is a virulence factor secreted by Mycobacterium tuberculosis (MTB). This work describes a voltammetric aptasensor for ultrasensitive detection of ESAT-6. Reduced graphene oxide doped with metal-organic framework (MOF-rGO) was deposited on a glassy carbon electrode (GCE). This increases the immobilization of electroactive Toluidine Blue (TB) and facilitates the electron transfer from TB to the modified GCE. Platinum/gold core/shell (Pt@Au) nanoparticles were used to assemble thiolated ESAT-6 binding aptamer (EBA) on a modified electrode and to further amplify the response to TB. The modified GCE, typically operated at -0.36 V (vs. SCE), has a linear response in 1.0 × 10-4 to 2.0 × 102 ng⋅mL-1 ESAT-6 concentration range, and the limit of detection (LOD) for ESAT-6 is as low as 3.3 × 10-5 ng⋅mL-1. It exhibits satisfactory specificity and reproducibility when analyzing spiked human serum. Graphical abstract Schematic presentation of a voltammetric aptasensor for Mycobacterium tuberculosis antigen ESAT-6 using a glassy carbon electrode modified with reduced graphene oxide (rGO) and a metal-organic framework (MOF). The limit of detection for ESAT-6 is as low as 3.3 × 10-5 ng/mL.

[Establishment of cardiac remodeling model in FVB/N mice by intraperitoneal injection of isoproterenol].

OBJECTIVE: To explore the feasibility of intraperitoneal injection of isoproterenol (ISO) to induce cardiac remodeling in FVB/N mice. METHODS: Forty-eight FVB/N mice were divided into back subcutaneous saline group (subcutaneous saline group), intraperitoneal saline group, back subcutaneous ISO group (subcutaneous ISO group), and intraperitoneal ISO group according to the route of administration of saline or ISO. ISO (30 µg/g body weight/day) was given to the subcutaneous ISO group and the intraperitoneal ISO group, twice daily with an interval of 12 hours, for 14 consecutive days. The subcutaneous saline group and the intraperitoneal saline group were injected with an equal volume of saline. The left ventricular end-diastolic posterior wall thickness was measured by echocardiography, and the ratio of heart weight to tibia length was determined. Hematoxylin-eosin staining was used to determine the myocardial fiber diameter. Picric-sirius red staining was used to determine the myocardial collagen deposition area. Quantitative real-time PCR was used to measure the mRNA expression of collagen I. RESULTS: Compared with the subcutaneous ISO, subcutaneous saline, and intraperitoneal saline groups, the intraperitoneal ISO group had increased sizes of the cardiac cavity and the heart. Compared with the subcutaneous saline and intraperitoneal saline groups, the subcutaneous ISO group showed no significant changes in the gross morphology of the cardiac cavity and the heart. The intraperitoneal ISO group showed significant increases in the ratio of heart weight to tibia length, myocardial fiber diameter, left ventricular end-diastolic posterior wall thickness, myocardial collagen area percentage, and the mRNA expression of collagen I compared with the subcutaneous ISO, subcutaneous saline, and intraperitoneal saline groups (P<0.01). There were no significant differences in the above five indices between the subcutaneous ISO group and the subcutaneous saline and intraperitoneal saline groups (P>0.05). No significant difference in the mortality rate was found between the subcutaneous ISO and intraperitoneal ISO groups (P>0.05). CONCLUSIONS: Intraperitoneal injection of ISO can induce cardiac hypertrophy and fibrosis in FVB/N mice.

[Association between use of antibacterial agents in the first year of life and childhood asthma: a Meta analysis].

OBJECTIVE: To evaluate the association between the use of antibacterial agents in the first years of life and childhood asthma. METHODS: The Chinese and English databases CNKI, Wanfang Data, VIP, PubMed, and EBSCO were searched for prospective cohort studies on the association between the use of antibacterial agents in the first years of life and childhood asthma. Stata12.0 software was used to analyze the association through a Meta analysis. RESULTS: The articles with a high quality score and adjusted effective values for factors for lower respiratory tract infection were pooled, and a total of 8 studies were included. The results of the Meta analysis showed that the use of antibacterial agents in the first years of life increased the risk of childhood asthma (OR=1.14, 95%CI: 1.10-1.17, P<0.05). Compared with the children who used antibacterial agents 0-1 times in the first years of life, those who used more than 4 times had an increased risk of asthma (OR=1.28, 95%CI: 1.19-1.38, P<0.05). High-risk children (at least one immediate family member had asthma) who used antibacterial agents had an increased risk of asthma (OR=1.47, 95%CI: 1.20-1.81, P<0.05). CONCLUSIONS: The use of antibacterial agents in the first years of life increases the risk of childhood asthma. High-risk children who use antibacterial agents have an increased risk of asthma. The increased frequency of use of antibacterial agents in the first years of life is associated with an increased risk of childhood asthma, but the detailed dose relationship needs further investigation.

Microplate-based method to screen inhibitors of isozymes of cyclic nucleotide phosphodiesterase fused to SUMO.

The feasibility for microplate-based screening of inhibitors of isozymes of cyclic nucleotide phosphodiesterase (PDE) was tested via the coupled action of a phosphatase on adenosine-5'-monophosphate and an improved malachite green assay of phosphate. Human full-length PDE4B2 and truncated mutant (152-528aa) were expressed in Escherichia coli via fusion to SUMO, which after purification through Ni-NTA column exhibited specific activities >0.017 U mg(-1). In the presence of proteins <30 mg L(-1), absorbance for 10 µΜ phosphate was measurable; a PDE isozyme of specific activity over 0.008 U mg(-1) after reaction for 20 min thus suited for microplate-based screening of inhibitors. By using Biotek ELX 800 microplate reader, affinities of two forms of PEDE4B2 for cAMP, rolipram and papaverine varied over three magnitudes and were consistent with those by routine assay, respectively. Hence, the proposed method was promising for high-throughput-screening of inhibitors of phosphate-releasing enzymes bearing specific activities over 0.008 U mg(-1).

Identification of diagnostic model in heart failure with myocardial fibrosis and conduction block by integrated gene co-expression network analysis.

BACKGROUND: Despite the advancements in heart failure(HF) research, the early diagnosis of HF continues to be a challenging issue in clinical practice. This study aims to investigate the genes related to myocardial fibrosis and conduction block, with the goal of developing a diagnostic model for early treatment of HF in patients. METHOD: The gene expression profiles of GSE57345, GSE16499, and GSE9128 were obtained from the Gene Expression Omnibus (GEO) database. After merging the expression profile data and adjusting for batch effects, differentially expressed genes (DEGs) associated with conduction block and myocardial fibrosis were identified. Gene Ontology (GO) resources, Kyoto Encyclopedia of Genes and Genomes (KEGG) resources, and gene set enrichment analysis (GSEA) were utilized for functional enrichment analysis. A protein-protein interaction network (PPI) was constructed using a string database. Potential key genes were selected based on the bioinformatics information mentioned above. SVM and LASSO were employed to identify hub genes and construct the module associated with HF. The mRNA levels of TAC mice and external datasets (GSE141910 and GSE59867) are utilized for validating the diagnostic model. Additionally, the study explores the relationship between the diagnostic model and immune cell infiltration. RESULTS: A total of 395 genes exhibiting differential expression were identified. Functional enrichment analysis revealed that these specific genes primarily participate in biological processes and pathways associated with the constituents of the extracellular matrix (ECM), immune system processes, and inflammatory responses. We identified a diagnostic model consisting of 16 hub genes, and its predictive performance was validated using external data sets and a transverse aortic coarctation (TAC) mouse model. In addition, we observed significant differences in mRNA expression of 7 genes in the TAC mouse model. Interestingly, our study also unveiled a correlation between these model genes and immune cell infiltration. CONCLUSIONS: We identified sixteen key genes associated with myocardial fibrosis and conduction block, as well as diagnostic models for heart failure. Our findings have significant implications for the intensive management of individuals with potential genetic variants associated with heart failure, especially in the context of advancing cell-targeted therapy for myocardial fibrosis.

Surface-Activated Ti3C2Tx Adsorption of Acetylene Black Coupled with Polyaniline as a Signal Tag for the Detection of the ESAT-6 Antigen of Mycobacterium tuberculosis.

Early secretory antigenic target-6 (ESAT-6) is regarded as the most immunogenic protein produced by Mycobacterium tuberculosis, whose detection is of great clinical significance for tuberculosis diagnosis. However, the detection of the ESAT-6 antigen has been hampered by the expensive cost and complex experimental procedures, resulting in low sensitivity. Herein, we developed a titanium carbide (Ti3C2Tx)-based aptasensor for ESAT-6 detection utilizing a triple-signal amplification strategy. First, acetylene black (AB) was immobilized on Ti3C2Tx through a cross-linking reaction to form the Ti3C2Tx-AB-PAn nanocomposite. Meanwhile, AB served as a conductive bridge, and Ti3C2Tx can synergistically promote the electron transfer of PAn. Ti3C2Tx-AB-PAn exhibited outstanding conductivity, high electrochemical signals, and abundant sites for the loading of ESAT-6 binding aptamer II (EBA II) to form a novel signal tag. Second, N-CNTs were adsorbed on NiMn layered double hydride (NiMn LDH) nanoflowers to obtain NiMn LDH/N-CNTs, exhibiting excellent conductivity and preeminent stability to be used as electrode modification materials. Third, the biotinylated EBA (EBA I) was immobilized onto a streptavidin-coated sensing interface, forming an amplification platform for further signal enhancement. More importantly, as a result of the synergistic effect of the triple-signal amplification platform, the aptasensor exhibited a wide detection linear range from 10 fg mL-1 to 100 ng mL-1 and a detection limit of 4.07 fg mL-1 for ESAT-6. We envision that our aptasensor provides a way for the detection of ESAT-6 to assist in the diagnosis of tuberculosis.

Spectrophotometric-dual-enzyme-simultaneous assay in one reaction solution: chemometrics and experimental models.

Spectrophotometric-dual-enzyme-simultaneous assay in one reaction solution (SDESA) is proposed. SDESA requires the following: (a) Enzyme A acts on Substrate A to release Product A bearing the longest difference absorbance peak (λ(A)) much larger than that of Product B (λ(B)) formed by Enzyme B action on Substrate B; λ(B) is close to the longest isoabsorbance wavelength of Product A and Substrate A (λ(0)); (b) absorbance at λ(A) and λ(0) is quantified via swift alternation of detection wavelengths and corrected on the basis of absorbance additivity; (c) inhibition/activation on either enzyme by any substance is eliminated; (d) Enzyme A is quantified via an integration strategy if levels of Substrate A are lower than the Michaelis constant. Chemometrics of SDESA was tested with γ-glutamyltransferase and lactate-dehydrogenase of complicated kinetics. γ-Glutamyltransferase releases p-nitroaniline from γ-glutamyl-p-nitroaniline with λ(0) at 344 nm and λ(A) close to 405 nm, lactate-dehydrogenase consumes reduced nicotinamide dinucleotide bearing λ(B) at 340 nm. Kinetic analysis of reaction curve yielded lactate-dehydrogenase activity free from inhibition by p-nitroaniline; the linear range of initial rates of γ-glutamyltransferase via the integration strategy, and that of lactate-dehydrogenase after interference elimination, was comparable to those by separate assays, respectively; the quantification limit of either enzyme by SDESA at 25-fold higher activity of the other enzyme remained comparable to that by a separate assay. To test potential application, SDESA of alkaline phosphatase (ALP) and ß-D-galactosidase as enzyme-linked-immunoabsorbent assay (ELISA) labels were examined. ALP releases 4-nitro-1-naphthol from 4-nitronaphthyl-1-phosphate with λ(0) at 405 nm and λ(A) at 458 nm, ß-D-galactosidase releases 4-nitrophenol from ß-D-(4-nitrophenyl)-galactoside with λ(B) at 405 nm. No interference from substrates/products made SDESA of ß-galactosidase and ALP simple for ELISA of penicillin G and clenbuterol in one well, and the quantification limit of either hapten was comparable to that via a separate assay. Hence, SDESA is promising.

Chromogenic substrate from 4-nitro-1-naphthol for hydrolytic enzyme of neutral or slightly acidic optimum pH: 4-nitro-1-naphthyl-ß-D-galactopyranoside as an example.

At pH from 5.5 to 7.6, absorptivity of 4-nitro-1-naphthol at 450 nm is over 2.1-fold of that of para-nitrophenol at 405 nm and over 9.6-fold of that of ortho-nitrophenol at 415 nm. On 4-nitro-1-naphthyl-ß-D-galactopyranoside at pH 7.4, catalytic efficiency of Escherichia coli ß-D-galactosidase is 3-fold of that on para-nitrophenyl-ß-D-galactopyranoside and about 40% of that on ortho-nitrophenyl-ß-D-galactopyranoside, and produces a lower quantification limit of penicillin G by enzyme-linked-immunoabsorbent-assay. Hence, 4-nitro-1-naphthol is favorable to prepare chromogenic substrates of hydrolytic enzymes of neutral or slightly acidic optimum pH.

Identification of aneuploidy-related gene signature to predict survival in head and neck squamous cell carcinomas.

BACKGROUND: To parse the characteristics of aneuploidy related riskscore (ARS) model in head and neck squamous cell carcinomas (HNSC) and their predictive ability on patient prognosis. METHODS: Molecular subtyping of HNSC specimens was clustered by Copy Number Variation (CNV) data from The Cancer Genome Atlas (TCGA) dataset applying consistent clustering, followed by immune condition evaluation, differentially expressed genes (DEGs) analysis and DEGs function annotation. Weighted gene co-expression network analysis (WGCNA), protein-protein interaction, Univariate Cox regression analysis, least absolute shrinkage and selection operator (LASSO) and stepwise multivariate Cox regression analysis were implemented to construct an ARS model. A nomogram for clinic practice was designed by rms package. Immunotherapy evaluation and drug sensitivity prediction were also carried out. RESULTS: We stratified HNSC patients into three different molecular subgroups, with the best prognosis in C1 cluster among 3 clusters. C1 cluster displayed greatest immune infiltration status. The most DEGs between C1 and C2 groups, mainly enriched in cell cycle and immune function. We constructed a nine-gene ARS model (ICOS, IL21R, CCR7, SELL, CYTIP, ZAP70, CCR4, S1PR4 and CD79A) that effectively differentiates between high- and low-risk patients. Patients in low ARS group showed a higher sensitivity to immunotherapy. A nomogram built by integrating ARS and clinic-pathological characteristics helped predict clinic survival benefit. Drug sensitivity evaluation found that 4/9 inhibitor drugs (MK-8776, AZD5438, PD-0332991, PHA-665752) acted on the cell cycle. CONCLUSIONS: We classified 3 molecular subtypes for HNSC patients and established an ARS prognostic model, which offered a prospective direction for prognosis in HNSC.

An enzymatic activity regulation-based clusterzyme sensor array for high-throughput identification of heavy metal ions.

The accurate identification and sensitive quantification of heavy metal ions are of great significance, considering that pose a serious threat to environment and human health. Most array-based sensing platforms, to date, utilize nanozymes as sensing elements, but few studies have explored the application of the peroxidase-like activity of clusterzymes in identification of multiple analytes. Herein, for the first time, we developed a clusterzyme sensor array utilizing gold nanoclusters (AuNCs) as sensing elements for five heavy metal ions identification including Hg2+, Pb2+, Cu2+, Cd2+ and Co2+. The heavy metal ions can differentially regulate the peroxidase-like activity of AuNCs, and that can be converted into colorimetric signals with 3,3',5,5'-tetramethylbenzidine (TMB) as the chromogenic substrate. Subsequently, the generated composite responses can be interpreted by combining pattern recognition algorithms. The developed clusterzyme sensor array can identify five heavy metal ions at concentrations as low as 0.5 µM and their multi-component mixtures. Especially, we demonstrated the successful identification of multiple heavy metal ions in tap water and traditional Chinese medicine, with an accuracy of 100% in blind test. This study provided a simple and effective method for identification and quantification of heavy metal ions, rendering a promising technique for environmental monitoring and drug safety assurance.