Objective:To analyze the mediating effect of cardiovascular health status (CVH) on the association between educational level and cardiovascular disease (CVD).Methods:The participants were from Shanghai Suburban Adult Cohort and Biobank, and questionnaire survey, physical examination, blood biochemistry were conducted from 2016 to 2020 for baseline information collection, and follow up was conducted until March 31, 2024 based on the medical data, CVD incidence data and death surveillance data at different levels. The associations of educational level, CVH and time to CVD onset of the study population were analyzed using the accelerated failure time model to analyze the mediating effects of CVH, health behaviors, and health factors in the association of educational level and time to CVD onset. The mediating effects of educational level, gender, and age moderated associations were also analyzed.Results:A total of 57 312 participants were included, with 2 780 new cases of CVD during a median follow-up of 6.71 (6.71-6.72) years, and a mean incidence density of 7.77/1 000 person-years (95% CI: 7.48/1 000 person-years -8.06/1 000 person-years). In total, the less educational level and the lower CVH, the higher CVD incidence density ( P<0.05). The results of accelerated failure time models showed that the time ratio for CVD-free survival was 1.15 (95% CI: 1.06-1.24) and 1.33 (95% CI: 1.10-1.60) for moderate and high educational level, respectively. The results of the mediation effect analysis showed that the association between moderate and high educational level and time to CVD onset was 29.60% (20.50%-50.00%) and 36.10% (23.80%-59.00%), 9.97% (5.07%-20.00%) and 13.84% (6.84%-29.00%), 15.24% (9.64%-27.00%) and 17.55% (11.58%-33.00%) of mediators mediated by CVH, health behaviors, health factors, respectively. Among them, there was an exposure-mediated interaction of educational level and a positive moderating effect of age. Conclusion:CVH, health behaviors and health factors had a proportionate mediating effect in the association between educational level and risk of CVD development.

Objective To map the genome-wide distribution profile of histone H3K27me3 modification in diffuse gastric cancer tissues,identify target genes regulated by H3K27me3,and primarily explore the potential mechanism of its modification reprogramming in the occurrence and development of the tumor.Methods Normal gastric mucosal tissues and diffuse gastric cancer tissues were harvested from the patients who underwent examinations or treatments in the departments of gastroenterology and gastrointestinal surgery of our medical center between 2021 and 2023.There were 14 patients in the normal group(6 males and 8 females,average age of 46 years)and 14 patients in the gastric cancer group(8 males and 6 females,average age of 63 years).Cleavage under target and tagmentation(CUT&Tag)technology was employed to capture genomic regions modified by H3K27me3,and analyze the reprogramming characteristics of these modifications.RNA sequencing data,data from high-throughput chromosome conformation capture(Hi-C)technology,and publicly available single-cell data were integrated to investigate the target genes regulated by the reprogramming of H3K27me3 modifications in diffuse gastric cancer cells.Results The quality of the CUT&Tag and RNA sequencing data met the standards required for subsequent analysis.Histone H3K27me3 modifications in normal gastric mucosa and diffuse gastric cancer tissues were primarily distributed in distal intergenic regions and intronic regions.In gastric cancer tissues,compared to normal tissues,there was significant reprogramming of H3K27me3 modifications,characterized by a marked reduction in overall H3K27me3 signal intensity.The loss of 2 912 H3K27me3 signal peaks might lead to the up-regulation of 822 tumor-associated genes.Among them,56 genes displayed the most significant up-regulation(fold change in signal intensity≥2,P<0.05),with notable enrichment in the mammalian target of rapamycin complex 1(mTORC1)signaling pathway.Specifically,the methionine transporter SLC7A5 and the cystine transporter SLC7A11 were found to have the highest expression levels in gastric cancer tissues.Single-cell data revealed that the abnormal overexpression of SLC7A11 in diffuse gastric cancer was primarily observed in tumor epithelial cells.Further validation using public data and immunohistochemical experiments confirmed the elevated expression of SLC7A11 in diffuse gastric cancer,which is associated with poor prognosis in gastric cancer patients.Conclusion The reprogramming of histone H3K27me3 modification is an important epigenetic characteristic in diffuse gastric cancer.Loss of H3K27me3 signal peaks may up-regulate the expression of SLC7A11 in diffuse gastric cancer cells,and thereby promote tumor progression.

Objective To map the super-enhancers remodeling of intestinal gastric cancer and reveal the tumor biological functions of the super-enhancers and the downstream target genes that may be activated.Methods A total of 31 normal gastric mucosal tissues,23 intestinal gastric cancer tissues and 9 intestinal gastric cancer organoids were collected from the Department of Gastroenterology of Army Medical Center of PLA from January to December 2022.Chromatin targeting histone H3K27ac modified chromatin targeting cleavage under targets and tagmentation(CUT&Tag)sequencing was conducted on above tissues.The remodeling profiles of super-enhancers in intestinal gastric cancer were analyzed and the key target genes were identified based on bioinformation tools.CRISPRi technology was used to intervene with the super-enhancers,the expression of target genes was detected with Western blotting,and the proliferation,migration and invasion abilities were detected by CCK-8 assay and Transwell chambers in the control group and the intervention group.Results There was a significant difference in the signal of super-enhancers between intestinal gastric cancer tissues and normal gastric mucosal tissues(P<0.05),and the active super-enhancers in cancer tissues may be involved in biological processes such as negative regulation of the immune system and cell adhesion.The expression of up-regulated cell migration-inducing protein(CEMIP)in tumor cells was regulated by the super-enhancers,and intervening the super-enhancers down-regulated the expression of CEMIP(P<0.05),and inhibited the cell proliferation,invasion and migration abilities of tumor cells(P<0.05).Conclusion Super-enhancer remodeling is observed in intestinal gastric cancer,and they can up-regulate the expression of CEMIP gene and promote the growth,migration and invasion of cancer cells.

Objective To draw the genome-wide distribution and remodeling characteristics of H3K27me3 silencers in signet-ring cell carcinoma of the stomach(SRCC)through epigenetic sequencing technology,and to investigate their roles in transcriptional regulation in order to elucidate the regulatory mechanism of SRCC malignant progression.Methods The study was conducted on 35 gastric samples obtained by gastroendoscopic biopsy(15 normal and 20 SRCC tissues)from Department of Gastroenterology of Army Medical Center of PLA between January 2021 and December 2023.Multi-omics analyses,including assay for transposase-accessible chromatin with high-throughput sequencing(ATAC-seq),cleavage under targets and tagmentation(CUT&Tag)and transcriptome sequencing(RNA-seq),were performed to identify chromatin accessibility,H3K27me3 silencer regions,and transcriptional changes,with aid of Illumina NovaSeq 6000.H3K27me3 related differentially expressed genes(|Log2FC|>1,FDR<0.05)were screened using DESeq2.Gene Ontology(GO)analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis were employed to analyze the enrichment function,and Homer was employed to identify transcription factor motifs.A regulatory network was constructed using Cytoscape,and then validated using immunohistochemistry to explore its regulatory mechanism.Results H3K27me3 silencers were primarily located in distal intergenic regions(37.06%)in SRCC.Compared with the normal tissues,SRCC showed a significant reduction in H3K27me3 silencer signals(95%CI:1.34～2.30,P=0.007)with 6 257 lost sites(FDR<0.01).Integrating CUT&Tag and RNA-seq revealed 380 up-regulated immune-related genes,particularly in T cell receptor signaling(OR=4.2,95%CI:2.8～6.3,P=0.002).Immunohistochemistry confirmed elevated expression of transcription factor EHF(P<0.05).Conclusion There is the remodeling of H3K27me3 silencers in SRCC,and EHF may potentially play a crucial role in the SRCC malignant progression.

Objective To identify the enhancer profile marked by histone H3K27ac modification in high-grade intraepithelial neoplasia(HGIN)in order to reveal the novel regulatory mechanism of HGIN pathogensis.Methods Gastric tissue samples were collected from Department of Gastroenterology of Army Medical Center of PLA between June 2022 and June 2023,including 14 normal gastric tissues(Nor group),31 HGIN tissues(HGIN group)and 17 gastric cancer tissues(GC group).Cleavage under targets and tagmentation(CUT&Tag)technique was employed to capture enhancer regions modified by histone H3K27ac.Multi-omics analysis was performed to identify HGIN-specific active enhancers and their potentially regulated genes.Immunohistochemical profiling was performed to assess differential expression of the gene of interest across clinically stratified specimens,combined with CRISPR-dCas9-mediated ablation of active enhancers to monitor the gene of interest transcriptional dynamics and validate enhancer-mediated regulatory mechanisms.Results Epigenomic sequencing obtained the data with excellent quality,and indicated that obvious remodeling was observed in H3K27ac enhancers in HGIN and GC groups(P<0.05),though no significant difference in the genome-wide distribution of H3K27ac modification among the 3 groups.Combining transcriptome data revealed that enhancer remodeling may up-regulate the expression of the proliferation-related target gene,CD24,in the HGIN tissue;while,inhibiting enhancer activity can notably reduce CD24 expression level(P<0.05).Immunohistochemical assay displayed a positive correlation between the expression levels of CD24 and Ki-67(P<0.001).Conclusion The remodeling of H3K27ac enhancer represents a significant epigenetic feature of the transformation from normal condition to HGIN.Remodeling of H3K27ac enhancer up-regulates CD24,which may facilitate the abnormal proliferation of gastric epithelial cells.

Objective To analyze clinical characteristics of mesenchymal-subtype gastric cancer(Mes-GC)by integrating multi-omics data and explore the characteristics of tumor cells and microenvironment associated with the risk for recurrence.Methods Gastric tumor tissue samples were collected from the patients who visited Department of Gastroenterology of Army Medical Center of PLA from January 2022 to December 2023.Transcriptome and genome sequencing were applied for these tissue samples,including 19 cases of diffuse-type gastric cancer,22 cases of intestinal-type gastric cancer,and 23 cases of mixed-type gastric cancer patients.Bioinformatics analysis was employed to investigate the differences in clinical characteristics and tumor microenvironment between Mes-GC and non-mesenchymal-subtype gastric cancer(non-Mes-GC)by integrating data resources including The Cancer Genome Atlas(TCGA),Gene Expression Omnibus(GEO),and National Genomics Data Center(NGDC).Results Compared to non-Mes-GC patients,Mes-GC ones were characterized by later clinical stages,deeper tumor infiltration,and higher rates of lymph node metastasis.Kaplan-Meier survival analysis confirmed that Mes-GC patients were associated with shorter survival time,poor prognosis as well as increased risk of cancer recurrence(P<0.05).Single-cell RNA sequencing data revealed that tumor cells in Mes-GC showed higher expression levels of the genes related to stemness,metastasis(P<0.05),and epithelial-mesenchymal transition(EMT).And in the tumor microenvironment,there were significant more myeloid cells,smooth muscle cells,endothelial cells and fibroblasts,with the most pronounced elevation in the proportion of fibroblasts(P<0.05).Moreover,the patients with larger proportion of fibroblasts were associated with poorer prognosis.Conclusion Mes-GC tumor cells exhibit higher stemness and EMT characteristics,and stromal cells such as myeloid cells,endothelial cells,and fibroblasts are enriched in the tumor microenvironment.These features may be key factors contributing to poor prognosis and high recurrence rate of Mes-GC.

Objective To identify the enhancer landscape marked by histone H3K27ac modifications in diffuse-type gastric cancer(DGC)tissues,and to elucidate the epigenetic remodeling mechanisms by which active enhancers regulate cachexia-related genes.Methods Gastric mucosal tissue samples were collected from Department of Gastroenterology of Army Medical Center of PLA during January 2022 to March 2023,including 10 normal gastric mucosa tissues(Normal group),10 DGC tissues diagnosed with cachexia(DGC group),and 10 organoids derived from DGC tissues(Organoid group).Using H3K27ac chromatin targeting cleavage and tagmentation(CUT&Tag)technology,genomic modification regions were captured to screen specific active enhancers and their potential target genes in DGC tissues.CRISPR-dCas9 gene editing technology was used to intervene with the enhancers,and the expression of target genes was detected with Western blotting and qRT-PCR.Sixteen female SPF-grade BALB/c Nude mice(6～8 weeks old,weighing 18～21 g)were utilized to establish an orthotopic xenograft tumor model using the human diffuse-type gastric cancer cell line MKN45.Cachexia-related phenotypes were evaluated in 3 groups:normal group(n=4),silencing group(n=6),and control group(n=6).Results Significant differential enhancer regions were identified between DGC and normal gastric mucosa tissues.DGC tissues exhibited a marked increase in enhancer abundance(P<0.05)and signal intensity when compared with the normal counterparts.Integrated analysis of transcriptome data revealed that some of these active enhancers up-regulated the expression of GDF15,a cachexia-associated target gene in DGC.Targeted silencing of the active enhancer of GDF15 using CRISPR/dCas9-KRAB plasmid technology resulted in a significant reduction in GDF15 expression at both mRNA levels(P<0.05)and protein.Results from orthotopic transplantation experiments of DGC demonstrated that silencing of active enhancers alleviated the cachexia phenotype in nude mice(P<0.05).Conclusion DGC exhibits enhancer remodeling,which regulates the expression of the cachexia-associated gene GDF15,and thereby contributes to the pathogenesis and progression of cancer cachexia.

Drug addiction,also referred to as drug dependence or substance use disorder,is a chronic and recurrent brain disease.Its main characteristics are compulsive drug-seeking behavior,continued use of drugs,and a loss of control over intake.Prolonged use of addictive substances can result in both physiological and psychological dependence.When usage is ceased,individuals may experience intense discomfort,including anxiety,insomnia,nausea,vomiting,and a strong craving for the substances.Drug dependence is classified into two types:physical dependence and psychological dependence.Physical dependence describes a pathological state of adaptation that results from the repeated use of addictive substances,leading to severe withdrawal syndrome upon cessation.Psychological dependence involves a mental craving for addictive substances,which is needed to experience the specific euphoria that follows consumption.Regular or continuous use is required to sustain these euphoric effects.The mechanisms of addiction are complex and influenced by genetic,environmental,and various other factors.They involve higher-level neurological activities,such as memory,reward,and decision-making.Currently,effective treatment methods for drug addiction are insufficient.Due to the complexity of drug addiction,laboratory animal research is essential.Using animal behavioral models to simulate human drug addiction can enhance our understanding of the mechanisms of addiction.This research offers a comprehensive overview of various animal experimental models that explore both physical and psychological dependence.It includes detailed descriptions of the methods and procedures used to assess physical dependence,behavioral sensitization,conditioned place preference,drug discrimination,and self-administration experiments.Additionally,the characteristics of each experimental model are compared,and the relevance of these models is discussed,aiming to provide support for the research on addiction mechanisms and the development of therapeutic methods.

Objective:To analyze the mediating effect of cardiovascular health status (CVH) on the association between educational level and cardiovascular disease (CVD).Methods:The participants were from Shanghai Suburban Adult Cohort and Biobank, and questionnaire survey, physical examination, blood biochemistry were conducted from 2016 to 2020 for baseline information collection, and follow up was conducted until March 31, 2024 based on the medical data, CVD incidence data and death surveillance data at different levels. The associations of educational level, CVH and time to CVD onset of the study population were analyzed using the accelerated failure time model to analyze the mediating effects of CVH, health behaviors, and health factors in the association of educational level and time to CVD onset. The mediating effects of educational level, gender, and age moderated associations were also analyzed.Results:A total of 57 312 participants were included, with 2 780 new cases of CVD during a median follow-up of 6.71 (6.71-6.72) years, and a mean incidence density of 7.77/1 000 person-years (95% CI: 7.48/1 000 person-years -8.06/1 000 person-years). In total, the less educational level and the lower CVH, the higher CVD incidence density ( P<0.05). The results of accelerated failure time models showed that the time ratio for CVD-free survival was 1.15 (95% CI: 1.06-1.24) and 1.33 (95% CI: 1.10-1.60) for moderate and high educational level, respectively. The results of the mediation effect analysis showed that the association between moderate and high educational level and time to CVD onset was 29.60% (20.50%-50.00%) and 36.10% (23.80%-59.00%), 9.97% (5.07%-20.00%) and 13.84% (6.84%-29.00%), 15.24% (9.64%-27.00%) and 17.55% (11.58%-33.00%) of mediators mediated by CVH, health behaviors, health factors, respectively. Among them, there was an exposure-mediated interaction of educational level and a positive moderating effect of age. Conclusion:CVH, health behaviors and health factors had a proportionate mediating effect in the association between educational level and risk of CVD development.

Drug addiction,also referred to as drug dependence or substance use disorder,is a chronic and recurrent brain disease.Its main characteristics are compulsive drug-seeking behavior,continued use of drugs,and a loss of control over intake.Prolonged use of addictive substances can result in both physiological and psychological dependence.When usage is ceased,individuals may experience intense discomfort,including anxiety,insomnia,nausea,vomiting,and a strong craving for the substances.Drug dependence is classified into two types:physical dependence and psychological dependence.Physical dependence describes a pathological state of adaptation that results from the repeated use of addictive substances,leading to severe withdrawal syndrome upon cessation.Psychological dependence involves a mental craving for addictive substances,which is needed to experience the specific euphoria that follows consumption.Regular or continuous use is required to sustain these euphoric effects.The mechanisms of addiction are complex and influenced by genetic,environmental,and various other factors.They involve higher-level neurological activities,such as memory,reward,and decision-making.Currently,effective treatment methods for drug addiction are insufficient.Due to the complexity of drug addiction,laboratory animal research is essential.Using animal behavioral models to simulate human drug addiction can enhance our understanding of the mechanisms of addiction.This research offers a comprehensive overview of various animal experimental models that explore both physical and psychological dependence.It includes detailed descriptions of the methods and procedures used to assess physical dependence,behavioral sensitization,conditioned place preference,drug discrimination,and self-administration experiments.Additionally,the characteristics of each experimental model are compared,and the relevance of these models is discussed,aiming to provide support for the research on addiction mechanisms and the development of therapeutic methods.