Magnetic covalent organic frameworks for extraction and determination of endocrine-disrupting chemicals in beverage and water samples.

BACKGROUND: Phenolic endocrine-disrupting chemicals (EDCs) are widespread and easily ingested through the food chain. They pose a serious threat to human health. Magnetic solid-phase extraction (MSPE) is an effective sample pre-treatment technology to determine traces of phenolic EDCs. RESULTS: Magnetic covalent organic framework (COF) (Fe3 O4 @COF) nanospheres were prepared and characterized. The efficient and selective extraction of phenolic EDCs relies on a large specific surface and the inherent porosity of COFs and hydrogen bonding, π-π, and hydrophobic interactions between COF shells and phenolic EDCs. Under optimal conditions, the proposed magnetic solid-phase extraction-high-performance liquid chromatography-ultra violet (MSPE-HPLC-UV) based on the metallic covalent organic framework method for phenolic EDCs shows good linearities (0.002-6 µg mL-1 ), with R2 of 0.995 or higher, and low limits of detection (6-1.200 ng mL-1 ). CONCLUSION: Magnetic covalent organic frameworks (Fe3 O4 @COFs) with good MSPE performance for phenolic EDCs were synthesized by the solvothermal method. The magnetic covalent organic framework-based MSPE-HPLC-UV method was applied successfully to determine phenolic EDCs in beverage and water samples with satisfactory recoveries (90.200%-123%) and relative standard deviations (2.100%-12.100%). © 2023 Society of Chemical Industry.

A Fluorescent Glucose Transport Assay for Screening SGLT2 Inhibitors in Endogenous SGLT2-Expressing HK-2 Cells.

The sodium-dependent glucose transporters 2 (SGLT2) plays important role in renal reabsorption of urinal glucose back to plasma for maintaining glucose homeostasis. The approval of SGLT2 inhibitors for treatment of type 2 diabetes highlights the SGLT2 as a feasible and promising drug target in recent years. Current methods for screening SGLT2 inhibitors are complex, expensive and labor intensive. Particularly, these methods cannot directly measure nonradioactive glucose uptake in endogenous SGLT2-expressing kidney cells. In present work, human kidney cells, HK-2, was incubated with a fluorescent D-glucose derivant 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG) and the fluorescent intensity of 2-NBDG was employed to measure the amount of glucose uptake into the cells. By optimizing the passages of HK-2 cells, 2-NBDG concentration and incubation time, and by measuring glucose uptake treated by Dapagliflozin, a clinical drug of SGLT2 inhibitors, we successfully developed a new assay for measuring glucose uptake through SGLT2. The nonradioactive microplate and microscope-based high-throughput screening assay for measuring glucose can be a new method for screening of SGLT2 inhibitors and implied for other cell assays for glucose measurement extensively.

Study on the effect of reactive oxygen species-mediated oxidative stress on the activation of mitochondrial apoptosis and the tenderness of yak meat.

This study investigated the effect of reactive oxygen species-mediated oxidative stress on activation of mitochondrial apoptosis and tenderness of yak meat during postmortem ageing. Oxidative stress degree, Ca2+ levels, membrane permeability transition pore opening, mitochondrial membrane potential, apoptotic factors and the shear force were examined. Results showed that the ROS generated by H2O2 significantly increased mitochondrial oxidative stress by decreasing the activities of superoxide dismutase, catalase and glutathione peroxidase, and increasing lipid peroxidation. Furthermore, oxidative stress enhanced Ca2+ production and cytochrome c release, changed the levels of Bcl-2 family proteins and activated caspase-9 and -3 activities. Ultimately, oxidative stress increased the apoptosis rate and tenderness of yak meat. These observations confirmed that ROS-mediated oxidative stress participates in the activation of the apoptotic cascade reaction involving Ca2+ and Bcl-2 family proteins. The results further suggested that ROS-mediated oxidative stress plays a significant role in meat tenderization through the mitochondrial apoptotic pathway.

Effect of mitochondrial apoptotic activation through the mitochondrial membrane permeability transition pore on yak meat tenderness during postmortem aging.

The effect of membrane permeability transition pore dependent mitochondrial apoptotic activation on yak meat tenderness was investigated. Results indicate that MPTP opening increased significantly and the mitochondrial membrane potential decreased markedly in the early aging process (P<0.05). Cytochrome c was released from the mitochondria to the cytoplasm via the MPTP in the early period. Meanwhile, the activation of procaspase-9 occurred earlier than that of procaspase-3. Cyclosporin A suppressed the MPTP opening, depolarization of the mitochondrial membrane potential, activities of caspase-9 and caspase-3, apoptosis rate, myofibril fragmentation index, reactive oxygen species generation, and Ca2+ levels. These results demonstrated that MPTP mediated the release of cytochrome c in the mitochondrial apoptotic pathway. Furthermore, yak meat tenderness was improved by mitochondrial apoptotic pathway during aging. MPTP opening may be influenced by the ROS generation and Ca2+ overloading in yak meat during postmortem aging.

Comparative analysis of selected innate immune-related genes following infection of immortal DF-1 cells with highly pathogenic (H5N1) and low pathogenic (H9N2) avian influenza viruses.

H5N1 and H9N2 viruses are important causes of avian influenza in China. H5N1 is typically associated with severe to fatal disease in poultry, while H9N2 is usually associated with mild disease. Differences in viral virulence prompted us to investigate whether innate immune responses would be differentially regulated following infection by H5N1 and H9N2 viruses. To address this hypothesis, expression of a panel of innate immune-related genes including IFN-α, IFN-ß, Mx1, OASL, ISG12, IFIT5, IRF7, USP18, SST, and KHSRP in immortal DF-1 cells following H5N1 and H9N2 infection was analyzed and compared by real-time quantitative RT-PCR. Cells infected by either virus overall exhibited a similar expression profile for four ISGs (Mx1, OASL, ISG12, and IFIT5), IFN-α, IFN-ß, and SST gene. However, two immune-regulatory genes (IRF7 and KHSRP) were not responsive to highly pathogenic H5N1 infection but were strongly up-regulated in DF-1 cells infected with low pathogenic H9N2 infection. The subtype-dependent host response observed in this study offers new insights into the potential roles of IRF7 and KHSRP in control and modulation of the replication and virulence of different subtypes or strains of avian influenza A virus.

[Serum levels of 25-(OH)D(3) and total IgE in children with asthma].

OBJECTIVE: To study the changes and clinical significance of serum levels of 25-(OH)D(3) and total IgE in children with asthma. METHODS: Thirty children with asthma, 40 children with asthmatic bronchitis, and 40 healthy children were enrolled. Double-antibody radioimmunoassay was used to detect the levels of serum 25-(OH)D(3) and total IgE. RESULTS: Serum 25-(OH)D(3) levels (18±3 ng/Ml)decreased significantly in the asthmatic group compared with those in the asthmatic bronchitis group (43±3 ng/mL) and the control group (43±3 ng/mL) (P<0.01). In contrast, serum total IgE levels (192±16 IU/mL) increased significantly in the asthmatic group compared with those in the asthmatic bronchitis group (123±14 IU/mL) and the control group (118±15 IU/mL) (P<0.01). Serum 25-(OH)D(3) levels were negatively correlated with serum total IgE levels in asthmatic children (r=-0.783, P<0.01). There were no correlation between serum 25-(OH)D(3) levels and serum total IgE level in the asthmatic bronchitis and the control groups. CONCLUSIONS: 25-(OH)D(3) may play an important role in the pathogenesis of asthma. The increased serum 25-(OH)D(3) level may inhibit total IgE expression, suggesting that increasing serum 25-(OH)D(3) level might be a new option for the prevention and treatment of asthma.

The role of MKK1/2 kinase activity in human cytomegalovirus infection.

Human cytomegalovirus infection of quiescent fibroblasts was found to induce a bi-phasic activation of mitogen-activated protein kinase (MAPK) kinase 1 and 2 (MKK1/2) and two of their downstream targets, extracellular signal regulated kinase 1 and 2 (ERK1/2), as determined by Western blot analysis using phospho-specific antibodies. Treatment of infected fibroblasts with U0126, a potent and specific inhibitor of MKK1/2 kinase activity, completely blocked ERK1/2 activation following HCMV infection without affecting cell viability. Anti-viral studies demonstrate that in the presence of U0126, viral titres are reduced and viral DNA replication is inhibited. In addition, protein levels of two viral early genes that are required for viral DNA replication, UL44 and UL84, are significantly decreased in the presence of U0126. These results suggest that HCMV-mediated activation of MKK1/2 kinase activity enhances virus infectivity by ensuring timely initiation of viral DNA replication, possibly by regulating early gene expression.