Sustainable Endospore-Based Microreactor System for Antioxidant Capacity Assay

A novel endospore-based microbial method for “post-additional” antioxidant capacity assay was developed. The technique was based on oxidation and catalysis of the 2,2′-azinobis­(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS) by Bacillus subtilis 168 endospores in the presence of dissolved oxygen. Coat protein A (CotA), which belongs to the endospore coat, was expressed, purified, and assessed for its ability to oxidize ABTS into the ABTS^•+ radical cation. The wild-type endospore necessary for oxidizing ABTS into ABTS^•+ radical cation was confirmed by knocking out the <i>cotA</i> gene from B. subtilis 168 by homologous double exchange. Findings revealed that the catalytic activity of the endospores may be attributed to the presence of the CotA protein. The use of endospores instead of purified enzymes to prepare ABTS^•+ greatly reduced the assay cost and eliminated the need to purify and store of enzymes. The self-life of the radical cation was kept stable for at least 12 days without addition of a stabilizer and laccase inhibitor. This behavior enables the large-scale preparation of ABTS^•+. The antioxidant capacities of the individual antioxidants and fruit samples were easily quantified and compared using the proposed method. The developed technique can be further developed as a high-throughput screening technique for antioxidants

Grand average ERP waveforms elicited by masked fearful faces and masked neutral faces in the face task and letter task across low or high perceptual load at fronto-central and central electrodes.

<p>The topographic maps at the bottom display differences between the ERPs for fearful and neutral stimuli in the time window of 250-330ms and 450–700 ms.</p

Grand average ERP waveforms elicited by masked fearful faces and masked neutral faces in the face task and letter task across low or high perceptual load at lateral occipito-temporal electrodes.

<p>Grand average ERP waveforms elicited by masked fearful faces and masked neutral faces in the face task and letter task across low or high perceptual load at lateral occipito-temporal electrodes.</p

Reaction times and percentages of correct responses in the face task and letter task across low or high perceptual load and fearful or neutral peripheral facial expressions.

<p>Error bars show one standard error (S.E).</p

Example of stimulus sequence.

<p>On each trial, subjects fixated and viewed a string of 6 letters presented in the centre of the screen, and facial expressions were presented peripherally for 16.7 ms and masked by scrambled faces. The letter string comprised six identical (low load) or six different letters (high load). Participants were instructed to discriminate the letters (X or N) at fixation or the facial expression (fearful or neutral) in the periphery. Please note that schematic faces displayed in the figure were not employed in the experiment but were only used for illustration purpose due to issues of copyrights. Real facial expressions were employed as stimuli in the experiment.</p

The possible causal relationship between COVID-19 and imaging markers of cerebral small vessel disease: a Mendelian randomization study

Observational studies have suggested that SARS-CoV-2 infection may increase the burden of cerebral small vessel disease (CSVD). This study aims to explore the causal correlation between COVID-19 and the imaging markers of CSVD using Mendelian randomization (MR) methods. Summary-level genome-wide association study (GWAS) statistics for COVID-19 susceptibility, hospitalization, and severity were utilized as proxies for exposure. Large-scale meta-analysis GWAS data on three neuroimaging markers of white matter hyperintensity, lacunar stroke, and brain microbleeds, were employed as outcomes. Our primary MR analysis employed the inverse variance weighted (IVW) approach, supplemented by MR-Egger, weighted median, and MR-PRESSO methods. We also conducted multivariable MR analysis to address confounding bias and validate the robustness of the established causal estimates. Comprehensive sensitivity analyses included Cochran’s Q test, Egger-intercept analysis, MR-PRESSO, and leave-one-out analysis. The MR analysis revealed a significant causal correlation between the severity of COVID-19 and an increased risk of lacunar stroke, as demonstrated by the IVW method (ORivw = 1.08, 95% CI: 1.03–1.16, pivw = 0.005, FDR = 0.047). Nevertheless, no causal correlations were observed between COVID-19 susceptibility or hospitalization and any CSVD imaging markers. The robustness and stability of these findings were further confirmed by multivariable MR analysis and comprehensive sensitivity analyses. This study provides compelling evidence of a potential causal effect of severe COVID-19 on the incidence of lacunar stroke, which may bring fresh insights into the understanding of the comorbidity between COVID-19 and CSVD.</p

Low SP1 Expression Differentially Affects Intestinal-Type Compared with Diffuse-Type Gastric Adenocarcinoma

<div><p>Specificity protein 1 (SP1) is an essential transcription factor that regulates multiple cancer-related genes. Because aberrant expression of SP1 is related to cancer development and progression, we focused on SP1 expression in gastric carcinoma and its correlation with disease outcomes. Although patient survival decreased as SP1 expression increased (P<0.05) in diffuse-type gastric cancer, the lack of SP1 expression in intestinal-type gastric cancer was significantly correlated with poor survival (P<0.05). The knockdown of SP1 in a high SP1-expressing intestinal-type gastric cell line, MKN28, increased migration and invasion but decreased proliferation. Microarray data in <i>SP1</i> siRNA-transfected MKN28 revealed that the genes inhibiting migration were downregulated, whereas the genes negatively facilitating proliferation were increased. However, both migration and invasion were decreased by forced SP1 expression in a low SP1-expressing intestinal-type gastric cell line, AGS. Unlike the intestinal-type, in a high SP1-expressing diffuse-type gastric cell line, SNU484, migration and invasion were decreased by <i>SP1</i> siRNA. In contrast to previous studies that did not identify differences between the 2 histological types, our results reveal that low expression of SP1 is involved in cancer progression and metastasis and differentially affects intestinal-type compared with diffuse-type gastric adenocarcinoma.</p> </div

The transfection of the intestinal-type cell lines, MKN28 and AGS, with <i>SP1</i> siRNA or <i>SP1</i> plasmid controls cell proliferation, cell migration, and invasion.

<p>A) MKN28 cells were transfected with <i>SP1</i> siRNA or control siRNA. Total mRNA was extracted to determine the expression of endogenous VEGF and CDH1 using qRT-PCR. <i>HPRT</i> was used as a control. *, <i>P</i><0.05; **, <i>P</i><0.01. B) Western blot data reveal that <i>SP1</i> siRNA causes the knockdown of SP1 and reduces the expression of both VEGF and CDH1 in MKN28. GAPDH was used as a control. C) Cell proliferation, migration, and invasion were assessed after transfection with <i>SP1</i> or control siRNA in MKN28. **, <i>P</i><0.01. D) Western blot data demonstrated that SP1 protein levels were increased by transfection with the <i>SP1</i> plasmid. ACTB served as the control in AGS. E) Cell proliferation, migration, and invasion assays were performed after transfection with the <i>SP1</i> plasmid or a control vector in AGS. **, <i>P</i><0.01.</p

Gene list divided into categories based on gene expression array data of <i>SP1</i> siRNA-treated MKN28 cells.

<p>Gene list divided into categories based on gene expression array data of <i>SP1</i> siRNA-treated MKN28 cells.</p

Immunohistochemical and survival analysis of SP1 expression in gastric cancer patients and SP1 expression in gastric cancer cell lines.

<p>A) The expression of SP1 increases with tumor progression from precancerous to malignant lesions. Positive staining is determined by a reddish-brown precipitate in the nuclei. B) The Kaplan-Meier survival probability with respect to SP1 expression levels in gastric carcinomas. Log-rank tests were performed to compare the survival of SP1 subgroups both within and between subtypes. C) The Kaplan-Meier survival probability with respect to SP1 expression levels in intestinal-type gastric carcinomas. Log-rank tests were performed to compare the survival of SP1 subgroups both within and between subtypes. <i>P</i><0.05 was considered to be statistically significant. D) The Kaplan-Meier survival probability with respect to SP1 expression levels in diffuse-type gastric carcinomas. Log-rank tests were performed to compare the survival of SP1 subgroups both within and between subtypes. <i>P</i><0.05 was considered to be statistically significant. E) The SP1 expression in 15 gastric cancer cell lines was measured by western blot analysis. The numbers indicate semi-quantified protein expression based on scanning densitometry normalized to GAPDH (a control). VEGF was also measured to compare its expression with SP1 in each cell type.</p