Objective To construct a risk prediction model for delayed medical treatment in patients with acute myocardial infarction(AMI),and to evaluate the predictive performance of the model.Methods Convenience sampling method was used to select 219 patients with AMI who were hospitalized in Yancheng Third People's Hospital from March 2023 to May 2024 as investigation objects.The patients with AMI were divided into delayed group(n=106)and undelayed group(n=1 13)with the 6-hour interval.Logistic regression analysis was used to establish a risk prediction model for AMI patients with delayed medical treatment.Hosmer-Lemeshow test and receiver operating characteristic curve were used to evaluate the goodness of fit and prediction ability of the model.Results Binary Logistic regression analysis showed that age,unknown heart disease at the time of onset,first chest pain and low score of brief health literacy screen were all risk factors for delayed medical treatment in AMI patients(P＜0.05).The model predicted that the area under the curve of AMI patients with delayed hospitalization was 0.771,the Youden index was 0.562,the optimal cutoff value was 0.514,and the sensitivity and specificity were 77.3％and 86.5％,respectively.Conclusion The constructed risk prediction model can effectively predict and screen the high-risk groups of AMI patients with delayed medical treatment,reduce the risk of AMI patients with delayed medical treatment,and provide a scientific basis for taking reasonable intervention measures to shorten the time of medical treatment for AMI patients.

ObjectiveTo investigate the effect and mechanism of Polygonatum neutral polysaccharides from sibiricum （PSP-NP） on colon injury in mice with chronic obstructive pulmonary disease （COPD）. MethodsMale C57BL/6J mice were randomly divided into a control group， a COPD model group， and a PSP-NP group. The COPD model was established using smoke exposure combined with intranasal LPS administration. The PSP-NP group was simultaneously treated daily with 200 mg/kg of PSP-NP via intragastric gavage， while the other groups received an equal volume of saline. HE staining was used to observe the pathological changes in the colon. ELISA was employed to detect the levels of LPS in serum and the expressions of ZO-1， Occludin， IL-6， and TNF-α in colon tissue. UPLC-MS was used to detect the types and contents of bile acids in colonic content， and to screen for differential bile acids. Differential microbial flora were identified using 16S rRNA gene sequencing， and correlation analysis was conducted with differential bile acids. PSP-NP was combined with the differential bile acids cholic acid （CA）， and deoxycholic acid （DCA） in vitro to analyze the binding capacity of PSP-NP for CA and DCA. PSP-NP was applied to NCM460 normal colonic epithelial cells cultured in CA and DCA. Cell migration ability was assessed using the scratch assay， and the mRNA expression levels of inflammatory cytokines TNF-α， IL-6， and NF-κB were measured by RT-qPCR. ResultsPSP-NP effectively improved colonic damage in COPD model mice， enhanced mechanical barrier function， alleviated inflammatory response， and regulated abnormal changes in colonic flora and bile acid metabolism. Correlation analysis further revealed that PSP-NP regulated colonic bile acid metabolism and reduced the redundancy of secondary bile acids by increasing the relative abundance of Bacteroidota， Verrucomicrobiota， Bacteroides， and Akkermansia， while decreasing the relative abundance of Lactobacillus and Bifidobacterium. Notably， in vitro binding assays demonstrated that PSP-NP bound to differential bile acids DCA and CA， with the strongest binding capacity for DCA at 58.2%. In cellular functional studies， DCA inhibited the migration ability of colonic epithelial cells NCM460 and significantly increased the relative mRNA expression levels of inflammatory factors TNF-α， IL-6， and NF-κB. Importantly， co-treatment with PSP-NP significantly ameliorated the impact of DCA on NCM460 cells. ConclusionsPSP-NP may significantly improve colonic damage in COPD model mice. The mechanism may involve the regulation of colonic bile acid metabolism and bile acid profiles through both microbial modulation and direct binding， thereby reducing the damage caused by secondary bile acids such as DCA to colonic epithelial cells.

Objective To investigate the effect of mutation human proprotein convertase subtilism/kexin type 9(hPCSK9D374Y)in PCSK9 gene on methionine choline deficiency diet(MCD)-induced nonalcoholic steato-hepatitis(NASH)in mice.Methods Sixteen C57BL/6J wild-type mice were selected and randomly divided into the hPCSK9D374Y group and the control GFP group.MCD was fed for 6 weeks,and then the serum level of hepatic triglyceride,alanine aminotransferase(ALT)and aspartate aminotransferase(AST)was examined.Oil Red O and Sirius Red staining microscopy were used to identify hepatic lipid infiltration and fibrosis severity.F4/80-positive cell infiltration was analyzed using immunohistochemistry.Lipid synthesis and inflammatory response-related proteins were detected by Western blot and related mRNA expression was analyzed by RT-qPCR.Results Hepatic hPCSK9 protein and mRNA were significantly up-regulated,LDLR protein expression was down-regulated,and ser-um level of ALT and AST was significantly elevated in the hPCSK9D374Y group of mice(P＜0.05).The degree of he-patic steatosis and fibrosis increased and F4/80-positive cells were significantly increased(P＜0.01).FASN and SCD1 proteins were significantly up-regulated and PPARα was down-regulated in the hPCSK9D374Y group;The ex-pression of TLR4 and p-P65 was elevated,whereas the expression of Iκ Bα was decreased(P＜0.001).RT-qPCR re-sults showed a significant increase of mRNA coding inflammatory factors TNF-α,IL-1β,IL-6,and MCP-1,and a significant up-regulation of fibrosis-associated mRNAs(collagen Ⅰα and collagen Ⅲα)was found(P＜0.001).Conclusions Functionally acquired mutation in the PCSK9 gene(hPCSK9D374Y)exacerbates MCD-induced hepatic steatosis,inflammatory response and fibrosis in mice.

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 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 construct a risk prediction model for delayed medical treatment in patients with acute myocardial infarction(AMI),and to evaluate the predictive performance of the model.Methods Convenience sampling method was used to select 219 patients with AMI who were hospitalized in Yancheng Third People's Hospital from March 2023 to May 2024 as investigation objects.The patients with AMI were divided into delayed group(n=106)and undelayed group(n=1 13)with the 6-hour interval.Logistic regression analysis was used to establish a risk prediction model for AMI patients with delayed medical treatment.Hosmer-Lemeshow test and receiver operating characteristic curve were used to evaluate the goodness of fit and prediction ability of the model.Results Binary Logistic regression analysis showed that age,unknown heart disease at the time of onset,first chest pain and low score of brief health literacy screen were all risk factors for delayed medical treatment in AMI patients(P＜0.05).The model predicted that the area under the curve of AMI patients with delayed hospitalization was 0.771,the Youden index was 0.562,the optimal cutoff value was 0.514,and the sensitivity and specificity were 77.3％and 86.5％,respectively.Conclusion The constructed risk prediction model can effectively predict and screen the high-risk groups of AMI patients with delayed medical treatment,reduce the risk of AMI patients with delayed medical treatment,and provide a scientific basis for taking reasonable intervention measures to shorten the time of medical treatment for AMI patients.

Objective To investigate the effect of mutation human proprotein convertase subtilism/kexin type 9(hPCSK9D374Y)in PCSK9 gene on methionine choline deficiency diet(MCD)-induced nonalcoholic steato-hepatitis(NASH)in mice.Methods Sixteen C57BL/6J wild-type mice were selected and randomly divided into the hPCSK9D374Y group and the control GFP group.MCD was fed for 6 weeks,and then the serum level of hepatic triglyceride,alanine aminotransferase(ALT)and aspartate aminotransferase(AST)was examined.Oil Red O and Sirius Red staining microscopy were used to identify hepatic lipid infiltration and fibrosis severity.F4/80-positive cell infiltration was analyzed using immunohistochemistry.Lipid synthesis and inflammatory response-related proteins were detected by Western blot and related mRNA expression was analyzed by RT-qPCR.Results Hepatic hPCSK9 protein and mRNA were significantly up-regulated,LDLR protein expression was down-regulated,and ser-um level of ALT and AST was significantly elevated in the hPCSK9D374Y group of mice(P＜0.05).The degree of he-patic steatosis and fibrosis increased and F4/80-positive cells were significantly increased(P＜0.01).FASN and SCD1 proteins were significantly up-regulated and PPARα was down-regulated in the hPCSK9D374Y group;The ex-pression of TLR4 and p-P65 was elevated,whereas the expression of Iκ Bα was decreased(P＜0.001).RT-qPCR re-sults showed a significant increase of mRNA coding inflammatory factors TNF-α,IL-1β,IL-6,and MCP-1,and a significant up-regulation of fibrosis-associated mRNAs(collagen Ⅰα and collagen Ⅲα)was found(P＜0.001).Conclusions Functionally acquired mutation in the PCSK9 gene(hPCSK9D374Y)exacerbates MCD-induced hepatic steatosis,inflammatory response and fibrosis in mice.

Background:The core circadian clock gene Bmal1 has been shown to be involved in the formation of visceral sensitization in irritable bowel syndrome(IBS)by affecting the tryptophan hydroxylase 1(TPH1)-5-hydroxytryptamine(5-HT)pathway,but the exact mechanism of its regulation is unknown.Aims:To investigate the molecular mechanism by which Bmal1 regulates the TPH1-5-HT pathway through the clock-controlled gene Piezo1.Methods:Dexamethasone was used to synchronize the expression of the circadian clock genes in RIN-14B cells.Bmal1 expression was up-regulated or down-regulated in RIN-14B cells by plasmid and siRNA transfection of the enterochromaffin cell(EC)model.The expression levels of target genes and proteins were detected by immunofluorescence staining,RT-qPCR,and Western blotting.5-HT content was detected by ELISA method.Results:(1)This study was the first to report the oscillation characteristics of RIN-14B circadian clock genes in EC model,among which the oscillation of Bmal1 was the most significant.Immunofluorescence showed that RIN-14B cells expressed CGA,Bmal1 and Piezo1.(2)After transfected with the Bmal1 overexpression plasmid,the mRNA and protein expression of Bmal1 were significantly up-regulated in RIN-14B cells compared with the negative control group(all P<0.001);while transfected with Bmal1 siRNA significantly decreased the mRNA and protein expression of Bmal1 in RIN-14B cells compared with the negative control group(all P<0.05).(3)After transfected with Bmal1 overexpression plasmid,the mRNA and protein expression of Piezo1,the protein expression of TPH1,and the intracellular content of 5-HT were significantly increased(all P<0.051).(4)After transfected with Bmal siRNA,mRNA expression of Piezo1 and TPH1 in RIN-14B cells was significantly down-regulated(all P<0.05),and the protein expression of Piezo1,TPH1,the intracellular 5-HT content tended to be decreased by 21%,31%,and 10%,respectively.Conclusions:RIN-14B cells have the characteristics of rhythmic oscillation of circadian clock genes,and Bmal1 overexpression and underexpression EC models can be successfully established by using RIN-14B cells.Overexpression and underexpression of Bmal1 can lead to significant changes in the clock-controlled Piezo1-TPH1-5-HT signaling pathway,suggesting that Bmal1 can be expressed by clock-control signals through the Piezo1-TPH1 pathway.This suggests that Bmal1 may be involved in the development of visceral hypersensitivity in IBS through regulation of 5-HT synthesis by the clock-controlled gene Piezo1.

In order to investigate the effects of porin genes ompW,ompS and ompD on the biological properties and virulence of Salmonella typhimurium,the corresponding mutant strains were con-structed using the λ Red homologous recombination system,and the growth curves,motility,bio-chemical properties,in vitro genetic stability,biofilm-forming ability,drug resistance,and lethal dose at half capacity(LD50)between the standard strain and each mutant strain were detected by comparative assays for Salmonella typhimurium.The results showed that,compared with the standard strain,the ompD and ompW mutation had less effect on the growth rate and motility of the bacteria,while the ompS mutation significantly reduced the growth rate and motility;none of the three genetic mutation affected the biochemical characteristics of Salmonella typhimurium,nor the genetic stability,but affected its susceptibility to a variety of commonly used antibiotics to varying degrees and caused a highly significant decrease(P＜0.01)in the ability to form a biofilm,and the results showed that the three mutant strains had a significant reduction in the ability to form a biofilm.The result of LD50 virulence assay showed that all three genetic mutation led to a decrease in the virulence of Salmonella typhimurium,among which the ompS mutant strain showed the most obvious decrease in virulence,LD50 was 25 times that of the standard strain.In conclusion,mutations of the pore protein ompW,ompS,and ompD genes can affect some biological properties of Salmonella typhimurium.The results of this study laid an experimental foundation for further research on the biological functions of the pore protein ompW,ompS and ompD genes and Salmonella pathogenicity.

Objective To establish and evaluate a gas chromatography-tandem mass spectrometry(GC-MS/MS)method for the detection of six common drugs(methamphetamine,meperidine,caffeine,codeine,cocaine and ketamine)in blood,and to improve the determination basis of results.Methods The above six drugs were added into the blank blood,and GC-MS/MS was used for detection after ether extraction.The collection,quantification and confirmation were carried out under the mode of multi-reaction monitoring(MRM).The qualitative results of the above six drugs were evaluated based on the maximum allowable deviation of the retention time and relative ion abundance ratio in the qualitative results of GC-MS/MS.Results There was a good linear relationship between the six common drugs,among which ketamine and caffeine had the lowest detection limit(0.01 μg/mL),methamphetamine had the highest detection limit(0.5 μg/mL).The retention time(RT)and relative retention time(RRT)of the target substance were stable under the six supplemental levels,and the absolute deviation(ΔRTabsolute)of RT was within±0.025 min.The absolute deviation of RRT(ΔRRTabsolute)was within±0.004.The relative ion abundance ratio absolute deviation(ΔIabsolute)is±20%,and the relative ion abundance ratio relative deviation(ΔIrelative)is±50%.Conclusion This study clarified the reference range for qualitative determination of six common drugs in blood matrix detected by GC-MS/MS,and effectively supplemented the qualitative determination indicators of existing instrumental analysis methods.