Human infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) and there is no cure currently. The 3CL protease (3CLpro) is a highly ...conserved protease which is indispensable for CoVs replication, and is a promising target for development of broad-spectrum antiviral drugs. In this study we investigated the anti-SARS-CoV-2 potential of Shuanghuanglian preparation, a Chinese traditional patent medicine with a long history for treating respiratory tract infection in China. We showed that either the oral liquid of Shuanghuanglian, the lyophilized powder of Shuanghuanglian for injection or their bioactive components dose-dependently inhibited SARS-CoV-2 3CLpro as well as the replication of SARS-CoV-2 in Vero E6 cells. Baicalin and baicalein, two ingredients of Shuanghuanglian, were characterized as the first noncovalent, nonpeptidomimetic inhibitors of SARS-CoV-2 3CLpro and exhibited potent antiviral activities in a cell-based system. Remarkably, the binding mode of baicalein with SARS-CoV-2 3CLpro determined by X-ray protein crystallography was distinctly different from those of known 3CLpro inhibitors. Baicalein was productively ensconced in the core of the substrate-binding pocket by interacting with two catalytic residues, the crucial S1/S2 subsites and the oxyanion loop, acting as a "shield" in front of the catalytic dyad to effectively prevent substrate access to the catalytic dyad within the active site. Overall, this study provides an example for exploring the in vitro potency of Chinese traditional patent medicines and effectively identifying bioactive ingredients toward a specific target, and gains evidence supporting the in vivo studies of Shuanghuanglian oral liquid as well as two natural products for COVID-19 treatment.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Alzheimer's disease (AD) is an intractable neurodegenerative disorder in the elderly population, currently lacking a cure. Trichostatin A (TSA), a histone deacetylase inhibitor, showed some ...neuroprotective roles, but its pathology-improvement effects in AD are still uncertain, and the underlying mechanisms remain to be elucidated. The present study aims to examine the anti-AD effects of TSA, particularly investigating its underlying cellular and molecular mechanisms.
Novel object recognition and Morris water maze tests were used to evaluate the memory-ameliorating effects of TSA in APP/PS1 transgenic mice. Immunofluorescence, Western blotting, Simoa assay, and transmission electron microscopy were utilized to examine the pathology-improvement effects of TSA. Microglial activity was assessed by Western blotting and transwell migration assay. Protein-protein interactions were analyzed by co-immunoprecipitation and LC-MS/MS.
TSA treatment not only reduced amyloid β (Aβ) plaques and soluble Aβ oligomers in the brain, but also effectively improved learning and memory behaviors of APP/PS1 mice. In vitro study suggested that the improvement of Aβ pathology by TSA was attributed to the enhancement of Aβ clearance, mainly by the phagocytosis of microglia, and the endocytosis and transport of microvascular endothelial cells. Notably, a meaningful discovery in the study was that TSA dramatically upregulated the expression level of albumin in cell culture, by which TSA inhibited Aβ aggregation and promoted the phagocytosis of Aβ oligomers.
These findings provide a new insight into the pathogenesis of AD and suggest TSA as a novel promising candidate for the AD treatment.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
•Chronic sleep deprivation (SD) aggravates the cognitive impairments in APP/PS1 mice.•Chronic SD exacerbates in vivo hippocampal LTP suppression in APP/PS1 mice.•Chronic SD down-regulates the ...expression level of PSD-95 in the hippocampus of APP/PS1 mice.•Chronic SD increases Aβ deposition and microglia activation in the hippocampus of APP/PS1 mice.
Alzheimer’s disease (AD) is a progressive neurodegenerative disorder characterized by cognitive deficits. Sleep deprivation (SD) could lead to memory deficits, and it was a candidate risk factor for AD. However, the effects of chronic SD on the cognitive functions of AD model mice and its possible mechanism are still unclear. In the present study, 8-month-old male APP/PS1 transgenic mice and wild type (WT) littermates were subjected to chronic SD by using the modified multiple platform method (MMPM), with 20 h of SD each day for 21 days. Then, the effects of chronic SD on cognitive functions in APP/PS1 mice were tested by using behavioral tests, the potential mechanisms were investigated by in vivo electrophysiological recording, western blot and immunochemistry. The results showed that chronic SD obviously aggravated the cognitive impairments, exacerbated in vivo hippocampal long-term potentiation (LTP) suppression, reduced the expression level of PSD95, increased amyloid-β (Aβ) protein deposition and overactivated microglia in the hippocampus of APP/PS1 mice. These results indicate that chronic SD exacerbates the cognitive deficits in APP/PS1 mice by accelerating the development of AD pathologies, reducing the expression of PSD95 and aggravating the LTP suppression in hippocampus. At the same time, chronic SD also impaired cognitive functions and synaptic plasticity in WT mice through down-regulating the level of PSD95 and activating microglia. These findings further clarify the electrophysiological and molecular mechanisms of exacerbated cognitive deficits in AD caused by chronic SD.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
To establish cholyglycine (CG) detection via enzyme‐multiplied immunoassay technique (EMIT), glucose‐6‐phosphate dehydrogenase (G6PD) was used as a reporter enzyme to prepare hapten–enzyme conjugate. ...Gel electrophoresis and UV scanning demonstrated that G6PD was successfully labeled with cholyglycine, and CG‐G6PD conjugate was obtained. Furthermore, the effects of various parameters on the preparation of CG‐G6PD conjugates were investigated. Consequently, CG amount, nicotinamide adenine dinucleotide, d‐glucose‐6‐phosphate (G6P), phosphate buffer and the pH, and ionic strength of solution had important effects on the residual activity of CG‐G6PD. Moreover, CG amount, the pH, and G6P played important roles in changing CG labeling location on G6PD. Using the CG‐G6PD conjugate as test kit, the cholyglycine–EMIT calibration curve was established, which could be employed in clinical detection of cholyglycine. This study provides some valuable information for preparing hapten–G6PD conjugates.
CG‐G6PD conjugates for CG detection.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The biological heterogeneity of hepatocellular carcinoma (HCC) makes prognosis difficult. Although many molecular tools have been developed to assist in stratification and prediction of patients by ...using microarray analysis, the classification and prediction are still improvable because the high-through microarray contains a large amount of information. Meanwhile, gene expression patterns and their prognostic value for HCC have not been systematically investigated. In order to explore new molecular diagnostic and prognostic biomarkers, the gene expression profiles between HCCs and adjacent nontumor tissues were systematically analyzed in the present study.
In this study, gene expression profiles were obtained by repurposing five Gene Expression Omnibus databases. Differentially expressed genes were identified by using robust rank aggregation method. Three datasets (GSE14520, GSE36376, and GSE54236) were used to validate the associations between cytochrome P450 (CYP) family genes and HCC. GSE14520 was used as the training set. GSE36376 and GSE54236 were considered as the testing sets.
From the training set, a four-CYP gene signature was constructed to discriminate between HCC and nontumor tissues with an area under curve (AUC) of 0.991. Accuracy of this four-gene signature was validated in two testing sets (AUCs for them were 0.973 and 0.852, respectively). Moreover, this gene signature had a good performance to make a distinction between fast-growing HCC and slow-growing HCC (AUC = 0.898), especially for its high sensitivity of 95%. At last, CYP2C8 was identified as an independent risk factor of recurrence-free survival (hazard ratio HR =0.865, 95% confidence interval CI, 0.754-0.992, P = 0.038) and overall survival (HR = 0.849; 95% CI, 0.716-0.995, P = 0.033).
In summary, our results confirmed for the first time that a four-CYP gene (CYP1A2, CYP2E1, CYP2A7, and PTGIS) signature is associated with fast-growing HCC, and CYP2C8 is associated with patient survival. Our findings could help to identify HCC patients at high risk of rapid growth and recurrence.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Tuberculosis (TB), caused by Mycobacterium tuberculosis, is one of the most fatal diseases in the world. Methylenetetrahydrofolate reductase (MTHFR) catalyzes the production of ...5-methyltetrahydrofolate (5-CH
-THF), which is required for the
biosynthesis of methionine in bacteria. Here, we identified Rv2172c as an MTHFR in M. tuberculosis through
and
analyses and determined that the protein is essential for the
growth of the bacterium. Subsequently, we constructed
R159N and L214A mutants in M. tuberculosis and found that these mutants were more sensitive to the antifolates
-aminosalicylic acid (PAS) and sulfamethoxazole (SMX). Combining biochemical and genetic methods, we found that
R159N or L214A mutation impaired methionine production, leading to increased susceptibility of M. tuberculosis to PAS, which was largely restored by adding exogenous methionine. Moreover, overexpression of
in M. tuberculosis could increase methionine production and lead to PAS resistance. This research is the first to identify an MTHFR in M. tuberculosis and reveals that the activity of this enzyme is associated with susceptibility to antifolates. These findings have particular value for antitubercular drug design for the treatment of drug-resistant TB.
In our previous studies, we have shown that (D-Ser2) oxyntomodulin (Oxm), a glucagon-like peptide 1 (GLP-1) receptor (GLP1R)/glucagon receptor (GCGR) dual agonist peptide, protects hippocampal ...neurons against Aβ1-42-induced cytotoxicity, and stabilizes the calcium homeostasis and mitochondrial membrane potential of hippocampal neurons. Additionally, we have demonstrated that (D-Ser2) Oxm improves cognitive decline and reduces the deposition of amyloid-beta in Alzheimer's disease model mice. However, the protective mechanism remains unclear. In this study, we showed that 2 weeks of intraperitoneal administration of (D-Ser2) Oxm ameliorated the working memory and fear memory impairments of 9-month-old 3×Tg Alzheimer's disease model mice. In addition, electrophysiological data recorded by a wireless multichannel neural recording system implanted in the hippocampal CA1 region showed that (D-Ser2) Oxm increased the power of the theta rhythm. In addition, (D-Ser2) Oxm treatment greatly increased the expression level of synaptic-associated proteins SYP and PSD-95 and increased the number of dendritic spines in 3×Tg Alzheimer's disease model mice. These findings suggest that (D-Ser2) Oxm improves the cognitive function of Alzheimer's disease transgenic mice by recovering hippocampal synaptic function and theta rhythm.
Serine/threonine phosphatase (Stp1) is a member of the bacterial Mg
- or Mn
- dependent protein phosphatase/protein phosphatase 2C family, which is involved in the regulation of Staphylococcus aureus ...virulence. Aurintricarboxylic acid (ATA) is a known Stp1 inhibitor with an IC50 of 1.03 μM, but its inhibitory mechanism has not been elucidated in detail because the Stp1-ATA cocrystal structure has not been determined thus far. In this study, we performed 400 ns molecular dynamics (MD) simulations of the apo-Stp1 and Stp1-ATA complex models. During MD simulations, the flap subdomain of the Stp1-ATA complex experienced a clear conformational transition from an open state to a closed state, whereas the flap domain of apo-Stp1 changed from an open state to a semi-open state. In the Stp1-ATA complex model, the hydrogen bond (H-bond) between D137 and N142 disappeared, whereas critical H-bond interactions were formed between Q160 and H13, Q160/R161 and ATA, as well as N162 and D198. Finally, four residues (D137, N142, Q160, and R161) in Stp1 were mutated to alanine and the mutant enzymes were assessed using phosphate enzyme activity assays, which confirmed their important roles in maintaining Stp1 activity. This study indicated the inhibitory mechanism of ATA targeting Stp1 using MD simulations and sheds light on the future design of allosteric Stp1 inhibitors.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Triglycerides account for 99% of milk fat and play a central role in determining dairy product quality. Many factors influence triglyceride synthesis and milk fat secretion. MicroRNAs have been ...verified to be involved in numerous biological processes, but little is known about their roles in milk fat biosynthesis. In this study, we aim to explore whether miR-454 could regulate triglyceride synthesis in bovine mammary epithelial cells (BMECs) by targeting PPAR-γ. A luciferase reporter assay showed that the predicted target site was correct and that miR-454 and PPAR-γ had a direct interaction. In addition, miR-454 mimics and inhibitors were transfected into BMECs. The results showed that both the mRNA and protein levels of PPAR-γ were negatively correlated with miR-454 expression. Fat droplet accumulation and triglyceride production were also inversely correlated with miR-454 expression. Our results indicate that miR-454 regulates triglyceride synthesis by directly targeting the PPAR-γ 3′ UTR in BMECs, suggesting that miR-454 could potentially be a new factor to elevate dairy product quality.
•PPAR-γ is a target gene of miR-454 in bovine mammary epithelial cells.•miR-454 reduced the mRNA and protein expression levels of PPAR-γ.•miR-454 suppresses lipogenesis in bovine mammary epithelial cells.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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•Magnetic microspheres were modified with Cibacron Blue via three spacer arms.•Spacer arms have important effect on CB modification and protein adsorption.•Maximum adsorption capacity ...was found to be 114.0mg/g microspheres.
In order to obtain a deep insight into the effects of spacer arms on performing CB modification and dye-affinity adsorption, poly(methyl methacrylate) (PMMA) magnetic microspheres were prepared and modified respectively with poly(ethylene glycol) (PEG), poly(vinyl alcohol) (PVA) and ethylenediamine (EDA) as spacer arms. Then, an affinity dye-ligand, Cibacron Blue F3GA (CB), was coupled with these spacer arms to synthesize three kinds of CB-attached magnetic microsphere including CB-PEG-PMMA, CB-PVA-PMMA and CB-EDA-PMMA. CB density of the resulting microspheres was determined to be 138.1μmol/g for CB-PEG-PMMA, 209.7μmol/g for CB-PVA-PMMA, and 266.0μmol/g for CB-EDA-PMMA, respectively. The affinity adsorption performances of the resulting microspheres were evaluated using bovine serum albumin (BSA). The results showed that the adsorption capacity of BSA increased with the increase in CB density, and CB-EDA-PMMA microspheres exhibited the highest adsorption capacity. Furthermore, the effects of pH, BSA concentration and ionic strength on BSA adsorption were investigated and the maximum adsorption capacity was found to be 114.0mg/g CB-EDA-PMMA microspheres. Moreover, the reusability of the microspheres was also studied.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
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