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.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.

To reduce the subjectivity and uncertainty present in the current international methods of drug value pricing when converting value into monetary prices,based on the hedonic pricing theory,it considers the post-negotiation price between manufacturers and payers as a reasonable price reference in the value pricing of Chinese patent medicine.By constructing an indicator system for the characteristics of Chinese patent medicine,it selects and measures the value characteristic variables that affect the price of Chinese patent medicine.It serves as the theoretical foundation and research basis for establishing a Hedonic price model between characteristic price variables and negotiation prices,thereby promoting the enhancement of rationality and objectivity in value-guided pricing of Chinese patent medicine.