Objective To explore the construction of heart preservation model of empty beating donor based on extracorporeal membrane oxygenation (ECMO). Methods From January 2022 to August 2023, 20 Guangxi Bama miniature pigs weighing 25-30 kg were selected, half male and half female. Under general anesthesia and heparinization, a midline thoracotomy was performed. The pericardium was cut after freeing the anterior and posterior vena cavae, and a perfusion needle was inserted near the brachiocephalic artery in the ascending aorta, connected to a blood collection bag to collect 500-600 mL of blood. The anterior and posterior vena cavae were ligated, the aorta was blocked and perfused with HTK solution to stop the heart beating. The superior and inferior vena cavae were cut off, the right pulmonary vein was decompressed, the aorta and left and right pulmonary arteries and veins were cut off, and the whole heart was removed. An ECMO device was used to continuously perfuse a cardioprotective solution mainly composed of oxygenated warm blood, maintaining the isolated pig heart beating for 8 hours, monitoring (once/hour) ECMO perfusion parameters, blood gas indicators, perfusate electrolytes, inflammatory factors, myocardial enzymes, myoglobin, and troponin levels. Myocardial tissue was taken for hematoxylin-eosin (HE) staining to observe myocardial cell damage and evaluate the quality of heart preservation. Results Among the 20 isolated beating pig hearts, 17 successfully resumed beating, 3 experienced ventricular fibrillation, resuscitated after intracardiac electrical defibrillation, and all 20 pig hearts successfully beat for 8 hours. There was no statistical difference in ECMO perfusion parameters, blood gas indicators, perfusate electrolytes, and inflammatory factors at each time point (P>0.05). There were statistical increases in myocardial enzymes, myoglobin, and troponin levels (P<0.05). HE staining results suggested that there was no severe myocardial damage. Conclusion ECMO technology can be used for pig heart preservation with good results, and this study provides experimental evidence for improving heart preservation research in clinical heart transplantation.

OBJECTIVE To investigate the clinical distribution and dynamic change of drug resistance of K.pneu-moniae and carbapenem-resistant Klebsiella pneumoniae(CRKP)isolated from a three-A hospital of Suzhou so as to provide scientific bases for prevention and control of hospital-associated infections and reasonable application of antibiotics.METHODS The K.pneumoniae and CRKP strains that were isolated from the submitted specimens were collected from the patients who treated in the Affiliated Suzhou Hospital of Nanjing Medical University from 2019 to 2023.The clinical characteristics of the patients with infection and the trend of drug resistance were statis-tically analyzed.RESULTS Totally 5631 strains of K.pneumoniae were isolated,1205(21.40％)of which were CRKP,and the isolation rate of CRKP showed an upward trend in the five years(x2=236.352,P＜0.001).Among the K.pneumoniae isolates,51.59％were isolated from sputum,13.51％from urine;19.43％were isolated from intensive care unit(ICU),7.64％from emergency department,and 7.19％from respiratory department.There were significant differences in gender,age and season between the patients detected with CRKP and the patients detected with non-CRKP(P＜0.05).The drug resistance rates of the K.pneumoniae strains to cephalosporins,quinolones and carbapenems con-tinuously increased from 2019 to 2023(P＜0.001),the drug resistance rate to imipenem increased from 11.69％to 34.24％,meropenem from 10.92％to 34.24％.CONCLUSIONS The K.pneumoniae isolates show severe drug re-sistance from 2019 to 2023,and the isolation rate of CRKP strains rises increasingly.It is necessary for the hospi-tal to focus on the continuous monitoring of key populations and departments and optimize the management of an-tibiotics and infection control strategies so as to provide guidance for reasonable clinical use of antibiotics,effective control of transmission of drug-resistant strains and cope with the increasingly severe drug resistance.

Objective To investigate the effect and mechanism of actin gamma1(ACTG1)downregulation on apoptosis in gastric cancer cells line HGC-27.Methods Stable ACTG1 knockdown HGC-27 cell line was constructed by CRISPR/Cas9,ACTG1 knockdown was verified by DNA sequencing and Western blot,apoptosis rate was detected by flow cytometry,Western blot detected the expression level of apoptosis-related proteins Bcl2-associated X protein(Bax),B cell lymphoma 2 family protein(Bcl2)and PI3K/AKT signal pathway-related proteins AKT and p-AKT.Results HGC-27 cell lines with stable knockdown of ACTG1 were constructed,and the ACTG1 protein levels were decreased in the sgACTG1-1 and sgACTG1-2 groups compared with the Control group.Compared with the apoptosis rate in the Control group(6.54%±0.67%),cell apoptosis rate in the sgACTG1-1 and sgACTG1-2 groups(10.11%±0.46%,14.67%±0.17%)were significantly increased,the differences were statistically significant(t=-7.58,-20.28,all P<0.01).Compared the Control group,the Bax protein levels in the sgACTG1-1 and sgACTG1-2 groups(0.89±0.02,1.00±0.08 vs 0.71±0.03)were significantly increased(t=-8.14,-5.87),the level of Bcl2 protein(0.49±0.06,0.39±0.06 vs 0.65±0.07)were significantly decreased(t=3.09,5.35),the differences were statistically significant(all P<0.05).Compared with the Control group,the AKT protein levels in the sgACTG1-1 and sgACTG1-2 groups(0.95±0.10,0.43±0.09 vs 1.17±0.06)and the P-AKT protein level(0.38±0.08,0.28±0.12 vs 0.70±0.14)were significantly decreased,the differences were statistically significant(t=3.20,12.13;3.44,3.85,all P<0.05).Conclusion Downregulation of ACTG1 inhibits the expression of PI3K/Akt signaling pathway related proteins AKT,p-AKT and promotes gastric cancer cells line HGC-27 apoptosis.

AIM To observe the effects of Hugan Tablets on high-fat diet induced non-alcoholic fatty liver disease(NAFLD)in a mouse model,and the autophagy,pyroptosis and the PI3K/Akt/mTOR signaling pathway as well.METHODS The C57BL/6 mice were randomly divided into the normal group,the model group,the Hugan Tablets group(0.7 g/kg)and the Yishanfu group(0.23 g/kg),with 10 mice in each group.The NAFLD mouse model was established by 16 weeks feeding of high-fat diet.From the 13th week,the mice started their corresponding dosing of the drug by gavage followed by killing of the mice at the end of 16th week and collection of their serum and liver tissue samples.The mice had their serum ALT,AST,TG,TC,LDL levels,liver TG,TC,NEFA,MDA levels and activities of SOD and GSH-Px detected;their serum levels of IL-1β,IL-6 and TNF-αdetected by ELISA;their hepatic pathological changes observed using HE staining and oil red O staining;and their hepatic protein expressions of ACC,CPT1A,FAS,p-PI3K,p-Akt,p-mTOR,P62,LC3,NLRP3,GSDMD and Caspase1 detected by Western blot.RESULTS Compared with the model group,the Hugan Tablets group displayed decreased body weight and hepatosmatic index level(P＜0.01);decreased levels of serum ALT,AST,TG,TC,LDL,IL-6,IL-1β and TNF-α(P＜0.05,P＜0.01);increased hepatic levels of TG,TC,NEFA and MDA(P＜0.05);decreased activities of SOD and GSH-Px(P＜0.05);improved pathological changes of hepatic lipid deposition and hepatocytic ballooning and decreased NAS score and oil red O staining area(P＜0.01);decreased hepatic protein expressions of ACC1,FAS,NLRP3,Caspase1,GSDMD,P62,p-PI3K,p-Akt and p-mTOR(P＜0.05,P＜0.01);and increased protein expressions of CPT1A and LC3(P＜0.01).CONCLUSION Hugan Tablets can effectively prevent and control the development of high-fat diet induced NAFLD in mice,and the mechanism may be associated with the promotion of autophagy in hepatocytes and the inhibition of pyroptosis via the inhibition of PI3K/Akt/mTOR signaling pathway.

OBJECTIVE To investigate the clinical distribution and dynamic change of drug resistance of K.pneu-moniae and carbapenem-resistant Klebsiella pneumoniae(CRKP)isolated from a three-A hospital of Suzhou so as to provide scientific bases for prevention and control of hospital-associated infections and reasonable application of antibiotics.METHODS The K.pneumoniae and CRKP strains that were isolated from the submitted specimens were collected from the patients who treated in the Affiliated Suzhou Hospital of Nanjing Medical University from 2019 to 2023.The clinical characteristics of the patients with infection and the trend of drug resistance were statis-tically analyzed.RESULTS Totally 5631 strains of K.pneumoniae were isolated,1205(21.40％)of which were CRKP,and the isolation rate of CRKP showed an upward trend in the five years(x2=236.352,P＜0.001).Among the K.pneumoniae isolates,51.59％were isolated from sputum,13.51％from urine;19.43％were isolated from intensive care unit(ICU),7.64％from emergency department,and 7.19％from respiratory department.There were significant differences in gender,age and season between the patients detected with CRKP and the patients detected with non-CRKP(P＜0.05).The drug resistance rates of the K.pneumoniae strains to cephalosporins,quinolones and carbapenems con-tinuously increased from 2019 to 2023(P＜0.001),the drug resistance rate to imipenem increased from 11.69％to 34.24％,meropenem from 10.92％to 34.24％.CONCLUSIONS The K.pneumoniae isolates show severe drug re-sistance from 2019 to 2023,and the isolation rate of CRKP strains rises increasingly.It is necessary for the hospi-tal to focus on the continuous monitoring of key populations and departments and optimize the management of an-tibiotics and infection control strategies so as to provide guidance for reasonable clinical use of antibiotics,effective control of transmission of drug-resistant strains and cope with the increasingly severe drug resistance.

Objective To assess the efficacy of clear lens extraction combined with extended depth of focus(EDOF)intraocular lens(IOL)implantation in enhancing visual quality and concurrently opening peripheral angle structures in patients diagnosed with primary angle closure suspect(PACS)and presbyopia.Methods A total of 80 patients with PACS,presbyopia,and transparent lenses were enrolled and divided into two groups based on the type of implanted lens:a control group(n=46)and an experimental group(n=34).The control group underwent clear lens extraction followed by monofocal intraocular lens implantation,while the experimental group underwent clear lens extraction followed by extended depth of focus(EDOF)intraocular lens implantation.Preoperative and postoperative evaluations at 3 months included visual acuity,intraocular pressure,anterior segment structure,defocus curve,and Catquest 9SF-CN questionnaire scores.Adverse events were recorded to assess safety.Results Regarding the angular structure,after 3 months of treatment,results indicated significant increases in central ante-rior chamber depth,angle opening distance at 750 μ m,angle recess area at 750 μ m,trabecular-iris space area at 750 μm,and trabecular-iris angle at 750 μm in both groups compared to preoperative values(all P<0.01).However,no significant differences were observed between the two groups(all P>0.05).For the defocusing curve,visual acuity in the experimental group improved from+1.0 D to-4.0 D after 3 months of treatment(all P<0.05),while the control group showed improvements at+0.5 D,-0.5 D,-1.0 D,-2.0 D,-3.0 D,-3.5 D,and-4.0 D(all P<0.05).Postoperatively at 3 months,the experimental group exhibited superior visual acuity to the control group at-1.0 D to-4.0 D(all P<0.05).According to the Catquest 9SF-CN scale,scores in both groups significantly im-proved after 3 months of treatment(P<0.01),with the experimental group scoring higher than the control group(P<0.01).During the study,transient intraocular pressure elevation occurred in 7 patients(3 in the control group and 4 in the experimental group),but no serious adverse reactions were reported in either group.Conclusions In patients with pseudoexfoliation glaucoma combined with presbyopia,significant improvements in angle structures were observed following clear lens extraction.The implantation of an EDOF IOL notably enhanced postoperative visual quality compared to monofocal IOLs.

Objective To develop and validate a transgenic mouse model enabling specific and inducible optogenetic activation of oligodendrocyte precursor cells(OPCs).Methods A conditional allele for the photosensitive opsin chicken opsin 5(cOpn5)(Rosa26-LSL-cOpn5)was generated using CRISPR/Cas9 technology.These mice were subsequently crossed with NG2-CreERT transgenic mice to produce NG2-CreERT;cOpn5 animals.In this model,tamoxifen administration induces Cre-mediated recombination,leading to specific expression of cOpn5 in NG2-positive OPCs.The specificity and efficiency of cOpn5 expression in OPCs were confirmed by immunofluorescent staining.Functional validation of light-induced OPC activation was performed by using calcium imaging in acute brain slices after stimulation with 470 nm blue light.Results Immunofluorescence analysis confirmed robust and specific expression of cOpn5 within NG2-positive OPCs in the brains of tamoxifen-treated NG2-CreERT;cOpn5 mice.Crucially,calcium imaging of acute brain slices from these mice demonstrated a significant increase in intracellular calcium levels in cOpn5-expressing OPCs upon stimulation with 470 nm blue light,indicating successful optogenetic activation.Conclusion We have successfully generated and validated a novel transgenic mouse model(NG2-CreERT;cOpn5)that permits specific and inducible optogenetic activation of OPCs.This model provides a novel tool for subsequent in vivo studies of the role and regulating mechanisms of OPCs in the central nervous system.

Randomized controlled trials have long been regarded as the gold standard for evaluating vaccine efficacy. However, conducting randomized controlled trials to assess vaccines in real-world settings is often impractical due to limitations in research resources and ethical considerations. In recent years, the emulated target trial has emerged as a novel methodological framework that applies the design principles of randomized trials to observational studies. By using observational data to emulate the randomization and treatment assignment of a clinical trial, this approach enables the evaluation of vaccine effectiveness in the real world. The application of emulated target trials enhances the quality and applicability of real-world evidence. This article explains the fundamental principles, design elements, advantages, and limitations of using emulated target trials to evaluate vaccine effectiveness, providing methodological guidance for conducting such studies in China.

Objective To develop and validate a transgenic mouse model enabling specific and inducible optogenetic activation of oligodendrocyte precursor cells(OPCs).Methods A conditional allele for the photosensitive opsin chicken opsin 5(cOpn5)(Rosa26-LSL-cOpn5)was generated using CRISPR/Cas9 technology.These mice were subsequently crossed with NG2-CreERT transgenic mice to produce NG2-CreERT;cOpn5 animals.In this model,tamoxifen administration induces Cre-mediated recombination,leading to specific expression of cOpn5 in NG2-positive OPCs.The specificity and efficiency of cOpn5 expression in OPCs were confirmed by immunofluorescent staining.Functional validation of light-induced OPC activation was performed by using calcium imaging in acute brain slices after stimulation with 470 nm blue light.Results Immunofluorescence analysis confirmed robust and specific expression of cOpn5 within NG2-positive OPCs in the brains of tamoxifen-treated NG2-CreERT;cOpn5 mice.Crucially,calcium imaging of acute brain slices from these mice demonstrated a significant increase in intracellular calcium levels in cOpn5-expressing OPCs upon stimulation with 470 nm blue light,indicating successful optogenetic activation.Conclusion We have successfully generated and validated a novel transgenic mouse model(NG2-CreERT;cOpn5)that permits specific and inducible optogenetic activation of OPCs.This model provides a novel tool for subsequent in vivo studies of the role and regulating mechanisms of OPCs in the central nervous system.

ObjectiveTo explore the regulatory effects and mechanisms of Astragali Radix polysaccharide （APS） on inhibitor of differentiation1 （ID1） and protein kinase B （Akt） in gastric cancer. MethodsImmunohistochemical staining was used to detect the expression of ID1 and Akt in 61 gastric cancer tissue samples and 20 adjacent normal gastric tissue samples. Immunofluorescence was used to detect the localization of ID1 and Akt. The effects of APS at the concentrations of 0.625， 1.25， 2.5， 5， 10， 20 mg·L^-1 on the proliferation of gastric cancer MGC-803 cells were examined by the cell counting kit-8（CCK-8） method and the colony formation assay. The target information of APS was retrieved from the Traditional Chinese Medicine Systems Pharmacology and Analysis Platform and Swiss Target Prediction. Keywords such as gastric cancer， gastric tumor， and stomach cancer were searched against GeneCards， UniProt， DisGeNET， and Online Mendelian Inheritance in Man （OMIM） for the screening of gastric cancer-related targets. The online tool jvenn was used to create the Venn diagram to identify the common targets， and STRING and Cytoscape were used to construct the protein-protein interaction network. Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analyses were conducted via R 4.2.2 to predict the potential roles of APS in the development of gastric cancer. The cell scratch assay was employed to assess the effect of APS on the migration of MGC-803 cells. The protein and mRNA levels of ID1 and Akt in the cells treated with APS were determined by Western blot and Real-time PCR， respectively. ResultsCompared with the adjacent normal gastric tissue， the gastric adenocarcinoma tissue showed increased positive expression of ID1 （χ² =81.00， P<0.01）. Immunofluorescence detection showed that ID1 and Akt were mainly located in the cytoplasm of gastric adenocarcinoma cells. Bioinformatics analysis identified 14 common genes shared between APS and gastric cancer. The average degree of protein-protein interaction network nodes was 14.29. GO and KEGG pathway enrichment results showed that ID1 and Akt were significantly enriched in the Rap1 and phosphatidylinositol-3-kinase （PI3K） /Akt signaling pathways. Cell experiments demonstrated that 5-fluorouracil （0.1 mg·L^-1） and APS （10， 20 mg·L^-1） groups showed decreased cell proliferation， migration， and colony formation. Compared with the control group， 10， 20 mg·L^-1 APS inhibited the proliferation of MGC-803 cells （P<0.01）， with 10 mg·L^-1 APS demonstrating stronger inhibitory effect. In addition， APS at 10， 20 mg·L^-1 inhibited the migration （P<0.01） and colony formation （P<0.05， P<0.01） of MGC-803 cells. Compared with the control group， APS at 10， 20 mg·L^-1 down-regulated the protein levels of ID1 （P<0.01） and Akt （P<0.05） and the mRNA levels of ID1 （P<0.05， P<0.01） and Akt （P<0.05， P<0.01） in MGC-803 cells. ConclusionID1 and Akt are highly expressed in the gastric adenocarcinoma tissue， which may be related to the development of gastric cancer. APS can down-regulate the protein and mRNA levels of ID1 and Akt to exert anti-tumor effects， which is expected to provide new therapeutic targets for gastric cancer treatment.