Preliminary Validation of Tumor Cell Attachment Inhibition Assay for Developmental Toxicants With Mouse S180 Cells

This study was designed to explore the possibility of using ascitic mouse sarcoma cell line (S180) to validate the mouse tumor cell attachment assay for developmental toxicants, and to test the inhibitory effects of various developmental toxicants. The results showed that 2 of 3 developmental toxicants under consideration, sodium pentobarbital and ethanol, significantly inhibited S180cells attachment to Concanavalin A-coaed surfaces. Inhibition was dependent on concentration, and the IC50 (the concentration tha reduced attachment by 50% ), of these 2 chemicals was 1.2×10-3mol/L and 1 .0 mol/L, respectively. Anoher developmental toxiant, hydmiortisone, did not show inhibitory activity. Two non-developmental toxicants, sodium chloride and glycine were also tested and these did not decrease attachment rates. The main results reported here were generally sindlar to those obtained with ascitic mouse ovdrian tumor cells as a model. Therefore, this study added further evidence to the conclusion that cell specificity does not lindt attachment inhibition to Con A-coated surfaces, so S180 cell may serve as an altemative cell model, especially when other cell lines are unavailable. Furthermore, after optimal validation, it can be suggested that an S180 cell attachment assay may be a candidate for a series of assays to detect developmental toxicants.

Synthesized flavanoid-derived ligand reduced dengue virus type-2 replication in vitro

Objective: To investigate the antiviral property of a lead ligand, YK51 that was synthesized based on the flavanoid of a natural product toward dengue virus type-2(DENV2)replication.Methods: c RNA was isolated from HepG2 cells inoculated with 1 000 median tissue culture infective dose of DENV2 and treated with different doses of the ligand followed by RT-PCR to quantify the virus gene copies. Confocal microscopy of actin and tubulin redistribution was also performed.Results: The quantitative RT-PCR result showed reduction of the DENV2 gene copies as the ligand concentration was increased. The confocal microscopy result showed increase in the tubulin intensity(79.6%) of infected BHK21 cells treated with the ligand,compared with the non-treated cells(54.8%). The 1.5-fold increase in the intensity of tubulin suggested that the ligand inhibitory effect stabilized the cellular microtubule structure.Conclusions: The synthesized ligand YK51 reduced DENV2 viral load by inhibiting virus replication thus is highly potential to be developed as antiviral agent.

Degradation and detoxification of microcystin-LR in drinking water by sequential use of UV and ozone

Microcystins （MCs） produced by cyanobacteria are strong hepatotoxins and classified as possible carcinogens. MCs pose a considerable threat to human health through tainted drinking and surface waters. Herein filtrated water from a waterworks in Harbin, China, was spiked with microcysfin-LR （MC-LR） extracted from a toxic scum of microcystis aeruginosa, and the spiked sample waters were treated using UV irradiation with consequent ozonation process （UV/O3）, compared with ozonation at a dose range commonly applied in water treatment plants, UV irradiation at 254 nm and UV irradiation combined with ozonation （UV＋O3）, respectively. The remaining of toxins were analyzed using high-performance liquid chromatography and also determined using a protein phosphatase type 2A inhibition assay, which was utilized to evaluate the reduction in toxicity. Results indicated that in comparison to other three processes （O3, UV, and UV＋O3）, UV/O3 process could effectively decrease both the concentration and toxicity of MC-LR at 100 μg/L level after 5 min UV irradiation with consequent 5 min ozonation at 0.2 mg/L （below 1 μg/L ）, while 0.5 mg/L ozone dose was required for the level below 0.1 μg/L. The addition of an UV treatment step to the existing treatment train may induce significant transformation of micropollutants and breaks down the natural organic matters into moieties unfavorable for ozone decomposition, stabilizing the ozone residual. These findings suggested that sequential use of UV and ozone may be a suitable method for the removal of these potentially hazardous microcystins from drinking water.

Organophosphate esters cause thyroid dysfunction via multiple signaling pathways in zebrafish brain

Organophosphate esters(OPEs)are widespread in various environmental media,and can disrupt thyroid endocrine signaling pathways.Mechanisms by which OPEs disrupt thyroid hormone(TH)signal transduction are not fully understood.Here,we present in vivo-in vitro-in silico evidence establishing OPEs as environmental THs competitively entering the brain to inhibit growth of zebrafish via multiple signaling pathways.OPEs can bind to transthyretin(TTR)and thyroxine-binding globulin,thereby affecting the transport of TH in the blood,and to the brain by TTR through the blood-brain barrier.When GH3 cells were exposed to OPEs,cell proliferation was significantly inhibited given that OPEs are competitive inhibitors of TH.Cresyl diphenyl phosphate was shown to be an effective antagonist of TH.Chronic exposure to OPEs significantly inhibited the growth of zebrafish by interfering with thyroperoxidase and thyroglobulin to inhibit TH synthesis.Based on comparisons of modulations of gene expression with the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases,signaling pathways related to thyroid endocrine functions,such as receptor-ligand binding and regulation of hormone levels,were identified as being affected by exposure to OPEs.Effects were also associated with the biosynthesis and metabolism of lipids,and neuroactive ligand-receptor interactions.These findings provide a comprehensive understanding of the mechanisms by which OPEs disrupt thyroid pathways in zebrafish.

Accelerated Evolution of H7N9 Subtype Influenza Virus under Vaccination Pressure

No avian H7 N9 outbreaks have occurred since the introduction of H7 N9 inactivated vaccine in the fall of 2017.However,H7 N9 is still prevalent in poultry.To surveil the prevalence,genetic characteristics,and antigenic changes of H7 N9,over7000 oropharyngeal and cloaca swab specimens were collected from live poultry markets and farms in 15 provinces of China from 2017 to 2019.A total of 85 influenza virus subtype H7 N9 strains were isolated and 20 representative strains were selected for genetic analysis and antigenicity evaluation.Results indicated the decreased prevalence of low-pathogenic H7 N9 strains while highly-pathogenic H7 N9 strains became dominated since the introduction of vaccine.Phylogenetic analysis showed that strains from 2019 formed an independent small branch and were genetically distant to strains isolated in 2013–2018.Analysis of key amino acid sites showed that the virus strains may adapt to the host environment evolutionally through mutation.Our analysis predicted additional potential glycosylation sites for HA and NA genes in the 2019 strains.Sequence analysis of HA gene in strains isolated from 2018 to 2019 showed that there were an increased nucleotide substitution rate and an increased mutation rate in the first and second nucleotides of coding codons within the open reading frame.The hemagglutination inhibition(HI)assay showed that H7-Re1 and H7-Re2 exhibited a lower HI titer for isolates from 2019,while H7-Re3 and r LN79 showed a high HI titer.The protective effect of the vaccine decreased after15 months of use.Overall,under vaccination pressure,the evolution of influenza virus subtype H7 N9 has accelerated.