Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laboratory diagnosis of viral encephalitis is a worldwide challenge.Recently,high-throughput sequencing technology has provided new tools for diagnosing central nervous system infections.Thus,In this study,we established a multipathogen detection platform for viral encephalitis based on amplicon sequencing. Methods We designed nine pairs of specific polymerase chain reaction(PCR)primers for the 12 viruses by reviewing the relevant literature.The detection ability of the primers was verified by software simulation and the detection of known positive samples.Amplicon sequencing was used to validate the samples,and consistency was compared with Sanger sequencing. Results The results showed that the target sequences of various pathogens were obtained at a coverage depth level greater than 20×,and the sequence lengths were consistent with the sizes of the predicted amplicons.The sequences were verified using the National Center for Biotechnology Information BLAST,and all results were consistent with the results of Sanger sequencing. Conclusion Amplicon-based high-throughput sequencing technology is feasible as a supplementary method for the pathogenic detection of viral encephalitis.It is also a useful tool for the high-volume screening of clinical samples.

Activation of nuclear factor erythroid 2-related factor 2(Nrf2)by Kelch-like ECH-associated protein 1(Keap1)alkylation plays a central role in anti-inflammatory therapy.However,activators of Nrf2 through alkylation of Keap1-Kelch domain have not been identified.Deoxynyboquinone(DNQ)is a natural small molecule discovered from marine actinomycetes.The current study was designed to investigate the anti-inflammatory effects and molecular mechanisms of DNQ via alkylation of Keap1.DNQ exhibited signif-icant anti-inflammatory properties both in vitro and in vivo.The pharmacophore responsible for the anti-inflammatory properties of DNQ was determined to be the α,β-unsaturated amides moieties by a chemical reaction between DNQ and N-acetylcysteine.DNQ exerted anti-inflammatory effects through activation of Nrf2/ARE pathway.Keap1 was demonstrated to be the direct target of DNQ and bound with DNQ through conjugate addition reaction involving alkylation.The specific alkylation site of DNQ on Keap1 for Nrf2 activation was elucidated with a synthesized probe in conjunction with liquid chromatography-tandem mass spectrometry.DNQ triggered the ubiquitination and subsequent degra-dation of Keap1 by alkylation of the cysteine residue 489(Cys489)on Keap1-Kelch domain,ultimately enabling the activation of Nrf2.Our findings revealed that DNQ exhibited potent anti-inflammatory capacity through α,β-unsaturated amides moieties active group which specifically activated Nrf2 signal pathway via alkylation/ubiquitination of Keap1-Kelch domain,suggesting the potential values of targeting Cys489 on Keap1-Kelch domain by DNQ-like small molecules in inflammatory therapies.

Objective:To identify the differentially expressed proteins that caused hippocampal damage after liver ischemia-reperfusion (I/R) in rats.Methods:Eighteen clean-grade healthy juvenile male Sprague-Dawley rats, aged 2 weeks, weighing 20-30 g, were divided into 2 groups ( n=9 each) using a random number table method: sham operation group (S group) and liver I/R group (IR group). A rat model of liver I/R injury was prepared by restoring perfusion after 1 h of liver ischemia. The rats were sacrificed after being anesthetized at day 3 of reperfusion, and the hippocampal tissue was isolated and analyzed to obtain gene expression profiles. Differentially expressed genes were identified using the R software, and further protein interaction networks were constructed through Cytoscape and Kyoto Encyclopedia Genes and Genomes pathway analysis to determine the differentially expressed proteins. Quantitative real-time polymerase chain reaction and Western blot were used for validation. Results:A total of 45 differentially expressed proteins were identified by the proteomic analysis of hippocampal tissues, including 36 significantly up-regulated proteins and 9 significantly down-regulated proteins. The proteins with significant expression related to injury were identified from the PPI network complex using the CytoHubBA plug-in cystscape: Ras-related C3 botulinum toxin substrate (RAC2), HRAS, phosphatidylinositol-3-kinase inhibitor phosphatase and tensin homologue (PTEN), and N-methyl-D-aspartate ionotropic glutamate receptor 2b (GRIN2b). The results of quantitative real-time polymerase chain reaction and Western blot showed that the expression of RAC2, HRAS, PTEN, and GRIN2b in the hippocampal tissue was significantly up-regulated in IR group compared with S group ( P<0.05). The results of Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that differentially expressed proteins were significantly enriched in the expression of PD-L1 and its checkpoint pathway, long-term potentiation, and regulation of the Wnt signaling pathway in cancer. Conclusions:The mechanism by which liver I/R induces hippocampal injury may be related to the up-regulation of the expression of RAC2, HRAS, PTEN and GRIN2b in rats.

The objective of this study was to optimise the extraction process of peptide of Poecilobdella manillensis by the Box-Behnken design-response surface methodology, and to investigate its whitening and anti-aging effects. Based on single factor experiments, NaCl solution concentration, extracting time and extracting times were taken as influencing factors, and peptide yield was used as the response value. The response surface model was designed and used to obtain the optimal extraction conditions of peptides of Poecilobdella manillensis: NaCl solution concentration of 4.3%, ultrasonic time of 4 h, and ultrasonic times of 2 times (2 + 2 h), which was significant and well-fitted to the actual experiment. The tyrosinase inhibition activity of peptide of Poecilobdella manillensis was evaluated by measuring the oxidation rate of levodopa (L-DOPA) catalyzed by tyrosinase. Moreover, using Caenorhabditis elegans as a model organism, the effects of peptide of Poecilobdella manillensis on body length, locomotion, reproductive capacity, reactive oxygen species (ROS), and lipofuscin levels were determined. The results showed that peptide of Poecilobdella manillensis exhibited a significant inhibitory effect on tyrosinase, with an IC₅₀ of 0.58 mg·mL^-1, which was stronger than that of the positive control arbutin (2.24 mg·mL^-1). Compared to the control group, 0.1, 0.5 and 1.0 mg·mL^-1 peptide of Poecilobdella manillensis showed no significant differences in the body length, eggs, and body bending of Caenorhabditis elegans. However, 0.5 mg·mL^-1 peptide of Poecilobdella manillensis significantly reduced ROS levels in Caenorhabditis elegans; 0.5 and 1.0 mg·mL^-1 peptide of Poecilobdella manillensis significantly reduced lipofuscin levels in Caenorhabditis elegans. Peptide of Poecilobdella manillensis exhibits effective whitening and anti-aging activities, potentially mediated by its inhibition of tyrosinase activity and antioxidant effects.

Background/Aims: Despite the high efficacy of direct-acting antivirals (DAAs), approximately 1–3% of hepatitis C virus (HCV) patients fail to achieve a sustained virological response. We conducted a nationwide study to investigate risk factors associated with DAA treatment failure. Machine-learning algorithms have been applied to discriminate subjects who may fail to respond to DAA therapy. Methods: We analyzed the Taiwan HCV Registry Program database to explore predictors of DAA failure in HCV patients. Fifty-five host and virological features were assessed using multivariate logistic regression, decision tree, random forest, eXtreme Gradient Boosting (XGBoost), and artificial neural network. The primary outcome was undetectable HCV RNA at 12 weeks after the end of treatment. Results: The training (n=23,955) and validation (n=10,346) datasets had similar baseline demographics, with an overall DAA failure rate of 1.6% (n=538). Multivariate logistic regression analysis revealed that liver cirrhosis, hepatocellular carcinoma, poor DAA adherence, and higher hemoglobin A1c were significantly associated with virological failure. XGBoost outperformed the other algorithms and logistic regression models, with an area under the receiver operating characteristic curve of 1.000 in the training dataset and 0.803 in the validation dataset. The top five predictors of treatment failure were HCV RNA, body mass index, α-fetoprotein, platelets, and FIB-4 index. The accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of the XGBoost model (cutoff value=0.5) were 99.5%, 69.7%, 99.9%, 97.4%, and 99.5%, respectively, for the entire dataset. Conclusions Machine learning algorithms effectively provide risk stratification for DAA failure and additional information on the factors associated with DAA failure.

Background/Aims: Chronic hepatitis C (CHC) patients who failed antiviral therapy are at increased risk for hepatocellular carcinoma (HCC). This study assessed the potential role of metformin and statins, medications for diabetes mellitus (DM) and hyperlipidemia (HLP), in reducing HCC risk among these patients. Methods: We included CHC patients from the T-COACH study who failed antiviral therapy. We tracked the onset of HCC 1.5 years post-therapy by linking to Taiwan’s cancer registry data from 2003 to 2019. We accounted for death and liver transplantation as competing risks and employed Gray’s cumulative incidence and Cox subdistribution hazards models to analyze HCC development. Results: Out of 2,779 patients, 480 (17.3%) developed HCC post-therapy. DM patients not using metformin had a 51% increased risk of HCC compared to non-DM patients, while HLP patients on statins had a 50% reduced risk compared to those without HLP. The 5-year HCC incidence was significantly higher for metformin non-users (16.5%) versus non-DM patients (11.3%; adjusted sub-distribution hazard ratio [aSHR]=1.51; P=0.007) and metformin users (3.1%; aSHR=1.59; P=0.022). Statin use in HLP patients correlated with a lower HCC risk (3.8%) compared to non-HLP patients (12.5%; aSHR=0.50; P<0.001). Notably, the increased HCC risk associated with non-use of metformin was primarily seen in non-cirrhotic patients, whereas statins decreased HCC risk in both cirrhotic and non-cirrhotic patients. Conclusions Metformin and statins may have a chemopreventive effect against HCC in CHC patients who failed antiviral therapy. These results support the need for personalized preventive strategies in managing HCC risk.

The incidence of thrombosis-induced cardiovascular diseases is increasing worldwide and poses a serious threat to human health. Three factors, slow speed of blood flow, hypercoagulable blood and vascular damage, have been considered to be causes of thrombosis. Antithrombotic drugs have been classified into three categories based on the mechanism of thrombosis, including anticoagulants, platelet inhibitors and fibrinolytics. The coagulation and anticoagulation systems have drawn increasing attention because of the important role they play in the process of thrombosis. Novel compounds with anticoagulant activity are now emerging, alleviating to some extent some of the problems associated with the clinical use of early approved thrombotic drugs, such as high bleeding risk, slow onset of action and narrow therapeutic windows. In this review, we initially describe the mechanisms of coagulation as well as thrombosis. Meanwhile, a wide range of bioactive compounds and potential antithrombotic candidates reported in recent years have been summarized. In addition, the structure-activity relationship of certain compounds has been discussed, expecting to facilitate the development of molecules with anticoagulant biological activity for the treatment of thrombotic diseases.

Uric acid (UA) is the final product of purine metabolism in human body,and its metabolic disorder will induce hyperuricemia (HUA).The occurrence and development of HUA are associated with a variety of pathological mechanisms such as oxidative stress injury,activation of inflammatory cytokines,and activation of renin-angiotensin-aldosterone system.These mechanisms directly or indirectly affect the bioavailability of endogenous nitric oxide (NO).The decrease in NO bioavailability is common in the diseases with high concentration of UA as an independent risk factor.In this review,we summarize the mechanisms by which high concentrations of UA affect the endogenous NO bioavailability,with a focus on the mechanisms of high-concentration UA in decreasing the synthesis and/or increasing the consumption of NO.This review aims to provide references for alleviating the multisystem symptoms and improving the prognosis of HUA,and lay a theoretical foundation for in-depth study of the correlations between HUA and other metabolic diseases.
Humans ; Nitric Oxide ; Uric Acid ; Hyperuricemia ; Biological Availability ; Cytokines

OBJECTIVE This study aims to elucidate the mechanism of action of Isoliquiritigenin(ILG)in the treatment of Dia-betic Encephalopathy(DE)based on network pharmacological analysis and in-vitro experiments.METHODS The potential targets of ILG were predicted using the HERB database and SwissTargetPrediction database.DE-associated disease targets were obtained from GeneCards,OMIM,and PharmGkb,and the intersecting targets between ILG and DE were identified using the Venny software.A PPI network was constructed using the STRING database,and core targets were screened out using Cytoscape software.GO function and KEGG pathway enrichment analyses were undertaken using R 4.0.3,followed by validation via molecular docking techniques and in vitro experiments.RESULTS 65 intersecting targets between ILG and DE were identified in this study.Topological analysis yielded eight core targets namely,EGFR,ESR1,PTGS2,PPARG,GSK3β,CDK2,PIK3R1,and F3.GO function and KEGG pathway en-richment analyses revealed that ILG antagonizes DE through several biological processes which impact numerous cellular components and molecular functions such as response to lipopolysaccharides,protein phosphorylation,protein kinase activity,and serine/threo-nine/tyrosine kinase activity.Pathways implicated included the PI3K-Akt signaling pathway,protein polysaccharide signaling pathway in cancer,and endocrine resistance pathway.The molecular docking results showed that all eight core targets had a good binding with ILG,especially with GSK3β,with a binding energy of-7.22 kcal·mol-1.In vitro experiments indicated that ILG could improve high glucose-induced cell damage and activate the PI3K/AKT/GSK3β signaling pathway.CONCLUSION ILG is likely to exert its effects on GSK3β to regulate the PI3K/AKT/GSK3β signaling pathway,thereby alleviating DE.

Diabetic retinopathy(DR)is a prevalent microvascular complication of diabetes and the leading cause of blindness and severe visual impairment in adults.The high levels of glucose trigger multiple intracellular oxidative stress pathways,such as POLDIP2,resulting in excessive reactive oxygen species(ROS)pro-duction and increased expression of vascular cell adhesion molecule-1(VCAM-1),hypoxia-inducible factor 1α(HIF-1α),and vascular endothelial growth factor(VEGF),causing microvascular dysfunction.Dihydromyricetin(DMY)is a natural flavonoid small molecule antioxidant.However,it exhibits poor solubility in physiological environments,has a short half-life in vivo,and has low oral bioavailability.In this study,we present,for the first time,the synthesis of ultra-small Fe-DMY nano-coordinated polymer particles(Fe-DMY NCPs),formed by combining DMY with low-toxicity iron ions.In vitro and in vivo experiments confirm that Fe-DMY NCPs alleviate oxidative stress-induced damage to vascular endo-thelial cells by high glucose,scavenge excess ROS,and improve pathological features of DR,such as retinal vascular leakage and neovascularization.Mechanistic validation indicates that Fe-DMY NCPs can inhibit the activation of the Poldip2-Nox4-H2O2 signaling pathway and downregulate vital vascular function indicators such as VCAM-1,HIF-1α,and VEGF.These findings suggest that Fe-DMY NCPs could serve as a safe and effective antioxidant and microangio-protective agent,with the potential as a novel multimeric drug for DR therapy.