BACKGROUND:Agomelatine is a clinically proven treatment for neuropsychiatric symptoms,such as anxiety and depression.Furthermore,our previous study has demonstrated that agomelatine ameliorates cognitive behaviors,hippocampal synaptic plasticity,and brain pathology in a mouse model of Alzheimer's disease.However,it remains unclear whether agomelatine can improve anxiety and depression-like behaviors in Alzheimer's disease model mice. OBJECTIVE:To investigate the improving effects of agomelatine on anxiety-and depression-like behaviors in APP/PS1 transgenic mice and its underlying molecular mechanisms. METHODS:(1)Eighteen APP/PS1 transgenic mice were randomly divided into model control group(n=9)and model intervention group(n=9).Another wild-type mice were randomized into control group(n=9)and intervention group(n=9).Model intervention group and intervention group were intraperitoneally injected with 10 mg/kg agomelatine per day for 31 continuous days.Behavioral experiments,including the elevated cross maze and forced swimming tests,and mRNA sequencing of the hippocampus were then performed.(2)Mouse hippocampal neuronal cell lines(HT22)and brain microvascular endothelial cell lines(bEnd.3)were cultured and divided into four groups:blank group without any drug,drug group with 20 μmol/L agomelatine,model group with 10 μmol/L β-amyloid 1-42,and experimental group with 10 μmol/L β-amyloid 1-42+20 μmol/L agomelatine.After 24 hours of incubation,protein expression of S416p-tau and S9p-GSK3β in HT22 cells was detected by immunoblotting,and protein expression of low-density lipoprotein receptor-related protein 1 and glycosylation end-product receptor in bEnd.3 cells was detected by immunoblotting. RESULTS AND CONCLUSION:In the elevated plus maze test,the time spent in the open arms(P＜0.01)and the entries into open arms(P＜0.05)in the mice of model control group were evidently lower than those in the control group,whereas those were obviously increased in the model intervention group compared with the model control group(P＜0.05).Forced swimming test results showed that the immobile time exhibited a marked increase in the model control group compared with the control group(P＜0.05),but it was significantly decreased in the model intervention group compared with the model control group(P＜0.05).Hippocampal tissue mRNA sequencing showed that agomelatine enhanced the expression of low-density lipoprotein receptor-related protein 1 in the hippocampus of APP/PS1 mice.Western blot analysis revealed that the level of S416p-tau in HT22 cells was higher in the model group than the blank group(P＜0.05),while it was markedly decreased in the experimental group compared with the model group(P＜0.05);the level of S9p-GSK3β in HT22 cells was higher in the drug group than the blank group(P＜0.05)as well as higher in the experimental group than the model group(P＜0.05).Moreover,the expression of low-density lipoprotein receptor-related protein 1 in bEnd.3 cells was higher in the experimental group than the model group(P＜0.05).To conclude,agomelatine can alleviate anxiety-and depression-like behaviors in Alzheimer's disease mice by promoting the clearance of β-amyloid and phosphorylated tau.

Objective To systematically identify the key genes regulating the exit from na?ve pluripotency in embryonic stem cells(ESCs)in order to provide novel targets and theoretical insights into the mechanisms for pluripotency transition and early cell fate determination.Methods Nanog-green fluorescent protein(Nanog-GFP)reporter-labeled ESCs were infected with a genome-wide Brie knockout library,and further cultured under leukemia inhibitory factor/serum(LIF/S)conditions for 14 d.Flow cytometry was used to sort Nanog-GFP?(na?ve-state)and Nanog-GFP-(primed state)cell populations,followed by genomic DNA extraction and high-throughput sequencing.Model-based Analysis of Genome-wide CRISPR/Cas9 Knockout(MAGeCK)was applied to identify differential genes between GFP?/Input,GFP?/Input,and GFP?/GFP? groups.Metascape and Gene Set Enrichment Analysis(GSEA)were conducted for functional enrichment analysis.Then the obtained candidate genes were employed to construct knockout models,and their roles were assessed through cell morphology observation,Nanog-positive rate detection,colony formation assays,and pluripotency gene expression analysis.Results The GFP?/Input screening revealed 2 921 negatively regulated genes(mainly enriched in basic life processes,such as RNA metabolism and cell cycle)and 1 393 positively regulated genes(enriched in the processes of nervous system development,carbohydrate metabolism,and vascular system development).In the GFP?/Input screening,2 765 negatively regulated genes(enriched in RNA metabolism,cell cycle,and other fundamental processes)and 1 303 positively regulated genes(enriched in neural development,cell survival,and endothelial migration)were identified.The GFP?/GFP? comparison identified 1 001 negatively regulated genes[involved in stress response and inhibition of mitogen-activated protein kinase(MAPK)signaling]and 983 positively regulated genes[related to fibroblast growth factor/extracellular signal-regulated kinase(FGF/ERK)signaling pathway and glucose metabolism).These genes,were not only known pluripotency regulators(e.g.,Nanog,Nr5a2,Klf2,Klf4)and exit-associated genes(e.g.,Gata6,Grb2,Zeb1,Fgfr1),but also some novel candidates(e.g.,Dmrt1,Rxra,Zbtb14 and Tmem41b).Functional validation showed that transient knockout of Dmrt1,Tmem41b,and Hic2 significantly increased the proportion of Nanog? cells(P<0.01),suggesting their role in suppressing ground-state exit.ESCs with stable Dmrt1 knockout exhibited a more na?ve-state phenotype,presenting compact,dome-shaped colonies,with increased ratio of undifferentiated colonies(P<0.01),up-regulation of ground-state markers(Nanog,Nr5a2,Dppa3,P<0.01),and down-regulation of primed-state markers(Fgf5,Lefty1,Dnmt3b,P<0.01).Rescue experiments for Dmrt1 expression reversed these above phenotypes.Conclusion A candidate gene set regulating exit from na?ve pluripotency in ESC is screened out and identified with genome-wide CRISPR.Our findings implicate Dmrt1 plays a critical role in promoting the exit.

Objective To understand the changing composition and antibiotic resistance of bacterial species in the clinical isolates from outpatient and emergency department(hereinafter referred to as outpatients)and inpatient children over time in various hospitals,and to provide laboratory evidence for rational antibiotic use.Methods The data on clinically isolated pathogenic bacteria and antimicrobial susceptibility of isolates from outpatients and inpatient children in the CHINET program from 2015 to 2021 were collected and analyzed.Results A total of 278 471 isolates were isolated from pediatric patients in the CHINET program from 2015 to 2021.About 17.1％of the strains were isolated from outpatients,primarily group A β-hemolytic Streptococcus,Escherichia coli,and Staphylococcus aureus.Most of the strains(82.9％)were isolated from inpatients,mainly SS.aureus,E.coli,and H.influenzae.The prevalence of methicillin-resistant S.aureus(MRSA)in outpatients(24.5％)was lower than that in inpatient children(31.5％).The MRSA isolates from outpatients showed lower resistance rates to the antibiotics tested than the strains isolated from inpatient children.The prevalence of vancomycin-resistant Enterococcus faecalis or E.faecium and penicillin-resistant S.pneumoniae was low in either outpatients or inpatient children.S.pneumoniae,β-hemolytic Streptococcus and S.viridans showed high resistance rates to erythromycin.The prevalence of erythromycin-resistant group A β-hemolytic Streptococcus was higher in outpatients than that in inpatient children.The prevalence of β-lactamase-producing H.influenzae showed an overall upward trend in children,but lower in outpatients(45.1％)than in inpatient children(59.4％).The prevalence of carbapenem-resistant Klebsiella pneumoniae(CRKpn),carbapenem-resistant Pseudomonas aeruginosa(CRPae)and carbapenem-resistant Acinetobacter baumannii(CRAba)was 14％,11.7％,47.8％in outpatients,but 24.2％,20.6％,and 52.8％in inpatient children,respectively.The prevalence of multidrug-resistant E.coli,K.pneumoniae,Proteus mirabilis,P.aeruginosa and A.baumannii strains was lower in outpatients than in inpatient children.The prevalence of fluoroquinolone-resistant E.coli,ESBLs-producing K.pneumoniae,ESBLs-producing P.mirabilis,carbapenem-resistant E.coli(CREco),CRKpn,and CRPae was lower in children in outpatients than in inpatient children,but the prevalence of CRAba in 2021 was higher than in inpatient children.Conclusions The distribution of clinical isolates from children is different between outpatients and inpatients.The prevalence of MRSA,ESBL,and CRO was higher in inpatient children than in outpatients.Antibiotics should be used rationally in clinical practice based on etiological diagnosis and antimicrobial susceptibility test results.Ongoing antimicrobial resistance surveillance and prevention and control of hospital infections are crucial to curbing bacterial resistance.

Objective To investigate the changing antibiotic resistance profiles of E.coli isolated from patients in the 52 hospitals participating in the CHINET program from 2015 to 2021.Methods Antimicrobial susceptibility was tested for clinical isolates of E.coli according to the unified protocol of CHINET program.WHONET 5.6 and SPSS 20.0 software were used for data analysis.Results Atotal of 289 760 nonduplicate clinical strains ofE.coli were isolated from 2015 to 2021,mainly from urine samples(44.7±3.2)％.The proportion of E.coli strains isolated from urine samples was higher in females than in males(59.0％vs 29.5％).The proportion of E.coli strains isolated from respiratory tract and cerebrospinal fluid samples was significantly higher in children than in adults(16.7％vs 7.8％,0.8％vs 0.1％,both P＜0.05).The isolates from internal medicine department accounted for the largest proportion(28.9±2.8)％with an increasing trend over years.Overall,the prevalence of ESBLs-producing E.coli and carbapenem resistant E.coli(CREco)was 55.9％and 1.8％,respectively during the 7-year period.The prevalence of ESBLs-producing E.coli was the highest in tertiary hospitals each year from 2015 to 2021 compared to secondary hospitals.The prevalence of CREco was higher in children's hospitals compared to secondary and tertiary hospitals each year from 2015 to 2021.The prevalence of ESBLs-producing E.coli in tertiary hospitals and children's hospitals and the prevalence of CREco in children's hospitals showed a decreasing trend over the 7-year period.The prevalence of CREco in secondary and tertiary hospitals increased slowly.Antibiotic resistance rates changed slowly from 2015 to 2021.Carbapenem drugs(imipenem,meropenem)were the most active drugs amongβ-lactams against E.coli(resistance rate≤2.1％).The resistance rates of E.coli to β-lactam/β-lactam inhibitor combinations(piperacillin-tazobactam,cefoperazone-sulbactam),aminoglycosides(amikacin),nitrofurantoin and fosfomycin(for urinary isolates only)were all less than 10％.The resistance rate of E.coli strains to antibiotics varied with the level of hospitals and the departments where the strains were isolated,especially for cefazolin and ciprofloxacin,to which the resistance rate of E.coli strains from children in non-ICU departments was significantly lower than that of the strains isolated from other departments(P＜0.05).The E.coli isolates from ICU showed higher resistance rate to most antimicrobial agents tested(excluding tigecycline)than the strains isolated from other departments.The E.coli strains isolated from tertiary hospitals showed higher resistance rates to the antimicrobial agents tested(excluding tigecycline,polymyxin B,cefepime and carbapenems)than the strains from secondary hospitals and children's hospitals.Conclusions E.coli is an important pathogen causing clinical infection.More than half of the clinical isolates produced ESBL.The prevalence of CREco is increasing in secondary and tertiary hospitals over the 7-year period even though the overall prevalence is still low.This is an issue of concern.

Objective:To observe any effect of supplementing treatment according to the Early Start Denver model (ESDM) with repeated transcranial magnetic stimulation (rTMS) in the treatment of children with autism spectrum disorder (ASD).Methods:Sixty-seven children on the autism spectrum aged 2 or 3 years were randomly divided into a control group of 33 and an observation group of 34. Both groups were treated as specified by the ESDM for 24 weeks, but the observation group additionally received rTMS. At 12 and 24 weeks, both groups were evaluated using the Autism Behavior Checklist, the Childhood Autism Rating Scale (CARS), the revised version of the Repetitive Behavior Scale (RBS-R), Gesell Development Schedules, and the Autism Treatment Evaluation Checklist (ATEC).Results:The CARS, Gesell, RBS-R and ATEC results of both groups had improved significantly after 12 weeks, with further improvements observed another 12 weeks later, when the average Autism Behavior Checklist scores had also improved significantly. At that point the results of the observation group were significantly better than those of the control group, on average.Conclusions:Combining ESDM and rTMS can significantly relieve the main symptoms of autism and improve the comprehensive development of children on the autism spectrum 2 or 3 years old. Therefore, such combination is worthy of application in clinical practice.

Objective:To explore the method for selecting characteristic prices of Chinese patent medicines based on informa-tion entropy theory.It involves analyzing the connotative differences among various price indicators and utilizing information entropy metrics to validate the scientific rigor of characteristic price selection so as to optimize the pricing model for Chinese patent medi-cines and improve the accuracy of price evaluation.Methods:A correlation analysis and information entropy calculation are con-ducted on the median price of the smallest preparation unit,average daily cost,and average course cost of TCM.It compares the information diversity and uncertainty of different pricing indicators.Results:The average daily cost exhibits the highest information diversity and uncertainty among all the pricing indicators examined.Conclusion:It is recommended that the average daily cost be used as the dependent variable for characteristic prices in TCM pricing research.This choice plays an important role in optimizing TCM pricing models and enhancing the accuracy of price evaluation.

Objective:To establish a scientific,systematic,and objective indicator system for the characteristic price variables of Traditional Chinese Patent Medicines(TCPM),providing a reference framework for the pricing mechanism of TCPM.Methods:The brainstorming method was initially used to screen related variable indicators.The Nominal Group Technique(NGT)and Delphi methods were applied to gather expert opinions,and SPSS 28.0 was employed for data statistical analysis.It led to the development of a TCPM characteristic price variable indicator system consisting of 6 dimensions,14 characteristic variables and 26 measurement indicators.Results:The authority coefficient of the experts exceeded 0.7,indicating the representativeness of the results.Expert opinions were generally concentrated.Based on the collected opinions and statistical analysis,the scope of selected TCPM characteristic price variables was preliminarily established.Conclusion:The TCPM characteristic price variable indicator system was initially developed.However,due to the complexity of the pricing mechanism and divergent expert opinions,further qualitative and quantitative research methods,along with a dynamic adjustment mechanism,are needed to verify and refine the system.

Objective To summarize the changing prevalence of carbapenem resistance in Enterobacterales based on the data of CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021 for improving antimicrobial treatment in clinical practice.Methods Antimicrobial susceptibility testing was performed using a commercial automated susceptibility testing system according to the unified CHINET protocol.The results were interpreted according to the breakpoints of the Clinical & Laboratory Standards Institute(CLSI)M100 31st ed in 2021.Results Over the seven-year period(2015-2021),the overall prevalence of carbapenem-resistant Enterobacterales(CRE)was 9.43％(62 342/661 235).The prevalence of CRE strains in Klebsiella pneumoniae,Citrobacter freundii,and Enterobacter cloacae was 22.38％,9.73％,and 8.47％,respectively.The prevalence of CRE strains in Escherichia coli was 1.99％.A few CRE strains were also identified in Salmonella and Shigella.The CRE strains were mainly isolated from respiratory specimens(44.23±2.80)％,followed by blood(20.88±3.40)％and urine(18.40±3.45)％.Intensive care units(ICUs)were the major source of the CRE strains(27.43±5.20)％.CRE strains were resistant to all the β-lactam antibiotics tested and most non-β-lactam antimicrobial agents.The CRE strains were relatively susceptible to tigecycline and polymyxins with low resistance rates.Conclusions The prevalence of CRE strains was increasing from 2015 to 2021.CRE strains were highly resistant to most of the antibacterial drugs used in clinical practice.Clinicians should prescribe antimicrobial agents rationally.Hospitals should strengthen antibiotic stewardship in key clinical settings such as ICUs,and take effective infection control measures to curb CRE outbreak and epidemic in hospitals.

Objective To characterize the changing species distribution and antibiotic resistance profiles of respiratory isolates in hospitals participating in the CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021.Methods Commercial automated antimicrobial susceptibility testing systems and disk diffusion method were used to test the susceptibility of respiratory bacterial isolates to antimicrobial agents following the standardized technical protocol established by the CHINET program.Results A total of 589 746 respiratory isolates were collected from 2015 to 2021.Overall,82.6％of the isolates were Gram-negative bacteria and 17.4％were Gram-positive bacteria.The bacterial isolates from outpatients and inpatients accounted for(6.0±0.9)％and(94.0±0.1)％,respectively.The top microorganisms were Klebsiella spp.,Acinetobacter spp.,Pseudomonas aeruginosa,Staphylococcus aureus,Haemophilus spp.,Stenotrophomonas maltophilia,Escherichia coli,and Streptococcus pneumoniae.Each microorganism was isolated from significantly more males than from females(P＜0.05).The overall prevalence of methicillin-resistant S.aureus(MRSA)was 39.9％.The prevalence of penicillin-resistant S.pneumoniae was 1.4％.The prevalence of extended-spectrum β-lactamase(ESBL)-producing E.coli and K.pneumoniae was 67.8％and 41.3％,respectively.The overall prevalence of carbapenem-resistant E.coli,K.pneumoniae,Enterobacter cloacae,Pseudomonas aeruginosa,and Acinetobacter baumannii was 3.7％,20.8％,9.4％,29.8％,and 73.3％,respectively.The prevalence of β-lactamase was 96.1％in Moraxella catarrhalis and 60.0％in Haemophilus influenzae.The H.influenzae isolates from children(＜18 years)showed significantly higher resistance rates to β-lactam antibiotics than the isolates from adults(P＜0.05).Conclusions Gram-negative bacteria are still predominant in respiratory isolates associated with serious antibiotic resistance.Antimicrobial resistance surveillance should be strengthened in clinical practice to support accurate etiological diagnosis and appropriate antimicrobial therapy based on antimicrobial susceptibility testing results.

Objective To investigate the distribution and antimicrobial resistance profiles of common pathogens isolated from cerebrospinal fluid(CSF)in CHINET program from 2015 to 2021.Methods The bacterial strains isolated from CSF were identified in accordance with clinical microbiology practice standards.Antimicrobial susceptibility test was conducted using Kirby-Bauer method and automated systems per the unified CHINET protocol.Results A total of 14 014 bacterial strains were isolated from CSF samples from 2015 to 2021,including the strains isolated from inpatients(95.3％)and from outpatient and emergency care patients(4.7％).Overall,19.6％of the isolates were from children and 80.4％were from adults.Gram-positive and Gram-negative bacteria accounted for 68.0％and 32.0％,respectively.Coagulase negative Staphylococcus accounted for 73.0％of the total Gram-positive bacterial isolates.The prevalence of MRSA was 38.2％in children and 45.6％in adults.The prevalence of MRCNS was 67.6％in adults and 69.5％in children.A small number of vancomycin-resistant Enterococcus faecium(2.2％)and linezolid-resistant Enterococcus faecalis(3.1％)were isolated from adult patients.The resistance rates of Escherichia coli and Klebsiella pneumoniae to ceftriaxone were 52.2％and 76.4％in children,70.5％and 63.5％in adults.The prevalence of carbapenem-resistant E.coli and K.pneumoniae(CRKP)was 1.3％and 47.7％in children,6.4％and 47.9％in adults.The prevalence of carbapenem-resistant Acinetobacter baumannii(CRAB)and Pseudomonas aeruginosa(CRPA)was 74.0％and 37.1％in children,81.7％and 39.9％in adults.Conclusions The data derived from antimicrobial resistance surveillance are crucial for clinicians to make evidence-based decisions regarding antibiotic therapy.Attention should be paid to the Gram-negative bacteria,especially CRKP and CRAB in central nervous system(CNS)infections.Ongoing antimicrobial resistance surveillance is helpful for optimizing antibiotic use in CNS infections.