Objective To investigate the role of quercetin(QUE)in ferroptosis during intestinal ischemia-reperfusion(IR)injury and elucidate its underlying mechanisms.Methods ① Potential target genes of QUE were predicted using the TCMSP,PharmMapper,and SwissTargetPredictive databases.Target genes associated with intestinal IR injury and ferroptosis were collected from GeneCards,PharmGKB,and OMIM databases.After overlapping genes were identified and analyzed,protein-protein interaction(PPI)networks were constructed using the STRING database and then visualized with Cytoscape 3.10.0.Molecular docking was performed to validate the binding conformations between QUE and key targets.② In vivo experiments were conducted to verify QUE's protective effects against intestinal IR injury.Thirty-six SPF-grade male C57BL/6J mice(6～8 weeks old,body weight:22±2 g)were randomly divided into Sham,Sham+QUE,IR,IR+QUE,IR+QUE+erastin(IR+QUE+Era),and IR+QUE+kevetrin hydrochloride(IR+QUE+KH)groups,with 6 mice in each group.Mouse model of intestinal IR injury was induced by 45 min ischemia of the superior mesenteric artery followed by 60 min reperfusion.HE staining was used to observe histopathological changes in the intestinal tissues.ELISA was employed to the serum or intestinal contents of diamine oxidase(DAO),pro-inflammatory cytokines(TNF-α,IL-6,IL-1β),and ferroptosis markers[glutathione(GSH)and Fe2+].Western blotting was utilized to detect the protein expression of glutathione peroxidase 4(GPX4),acyl-CoA synthetase long-chain family member 4(ACSL4),and tumor protein 53(p53).Results ① Network pharmacology identified 460 QUE targets,1 552 intestinal IR injury targets,and 1 967 ferroptosis-related targets,and 92 overlapping genes were identified as potential therapeutic targets.Molecular docking revealed a strong binding affinity between QUE and p53(binding energy:-6.8 kcal/mol).② In vivo experiments demonstrated that the IR+QUE group exhibited reduced intestinal damage and lower Chiu's score(P<0.05),decreased serum DAO content but elevated intestinal DAO content(P<0.05),decreased levels of TNF-α,IL-6,and IL-1β in the serum and intestinal tissues(P<0.05),reduced Fe2+accumulation,and increased GSH content(P<0.05),and up-regulated GPX4(P<0.05)and down-regulated ACSL4 and p53 expression(P<0.05)at protein level when compared with the IR group.While,the administration of ferroptosis agonist Era,or p53 agonist KH resulted in diminished therapeutic effects of QUE(P<0.05)when compared with the IR+QUE group.Conclusion QUE alleviates intestinal IR injury by inhibiting ferroptosis,which may be associated with its down-regulating p53 expression.

Objective To investigate whether remimazolam attenuates intestinal ischemia-reperfusion(I/R)injury in mice by regulating ferroptosis through connexin-43(CX43).Methods Molecular docking was applied to predict the binding affinity of remimazolam to CX43.A total of 72 SPF-grade adult male C57BL/6J mice(6～8 weeks old,weighing 20～25 g)were subjected.Thirty of them were randomly divided into sham operation group(Sham group),I/R group 1,and I/R+10,20 and 40 mg/kg remimazolam groups(RM10,RM20 and RM40 groups),with 6 mice in each group.Another 30 mice were randomly assigned into 5 groups(n=6),I/R group 2,erastin group(E group),I/R+40 mg/kg remimazolam group 2(RM40 group 2),I/R+Fer-1 group(Fer-1 group),and erastin+40 mg/kg remimazolam group(ERM group).The left 12 mice were randomly and equally grouped into I/R+RM+oe-NC group(oe-NC group)and I/R+RM+oe-CX43 group(oe-CX43 group).The Fer-1 group was given an intraperitoneal injection of 5 mg/kg Fer-1 in 1 h prior to reperfusion,the E group was given 10 mg/kg erastin intraperitoneally 1 d before modeling,and all the remimazolam groups,the oe-NC group and the oe-CX43 group were injected intravenously with corresponding doses of remimazolam 30 min pre-modeling,while the oe-NC and oe-CX43 groups were injected with empty vector virus and overexpression of CX43 vector virus,respectively,48 h before the administration of remimazolam.A mouse intestinal I/R injury model was constructed by clamping the superior mesenteric artery for 45 min and reperfusion for 30 min.The small intestine tissues were harvested and observed for pathological changes,and the intestinal mucosal damage was assessed with Chiu's score.The contents of Fe2+,total iron,malondialdehyde(MDA),glutathione(GSH),and superoxide dismutase(SOD)were detected by colorimetric assay;the production of reactive oxygen species(ROS)was determined by DHE probe;the expression of ferroptosis-related genes was determined by RT-qPCR;and the expression levels of CX43,GPX4,and SLC7A11 were detected by Western blotting.Results Molecular docking indicated that remimazolam had a binding energy of-6.699 kcal/mol with CX43 protein,suggesting good binding affinity between them.Compared with the Sham group,the I/R group 1 showed increases in Chiu's scores and CX43 expression(P<0.05),along with pathological damage to intestinal tissues,and elevated contents of Fe2+,total iron,ROS and MDA(P<0.05),and down-regulated GPX4 and SLC7A11(P<0.05).Compared with the I/R group 1,Chiu's score was reduced in the RM40 group,and CX43 was significantly down-regulated(P<0.05),contents of Fe2+,total iron,ROS,and MDA were decreased(P<0.05),and expression levels of GPX4 and SLC7A11 were enhanced(P<0.05),and severity of intestinal histological damage was attenuated in both the RM40 and Fer-1 groups.Compared with the E group,the ERM group had the decreases in CX43 expression level(P<0.05),Fe2+,total iron,ROS,and MDA contents(P<0.05),and increases in GPX4 and SLC7A11 expression levels(P<0.05),with the improvement in intestinal tissue.Compared with the oe-NC group,overexpression of CX43 resulted in the increased CX43 expression,elevated contents of Fe2+,total iron,ROS and MDA(P<0.05)and decreased expression of GPX4 and SLC7A11(P<0.05),leading to the exacerbated injury in intestinal tissue.Conclusion Remimazolam attenuates intestinal I/R injury by inhibiting ferroptosis through down-regulating CX43 expression.

AIM:To investigate the mechanism by which resveratrol(RSV)alleviates intestinal ischemia-re-perfusion(I/R)injury through modulation of the Wnt/β-catenin signaling pathway.METHODS:Twenty-four male C57BL/6J mice were randomly allocated into four experimental groups:the sham group,I/R group,I/R+RSV(treatment)group,and I/R+RSV+XAV939(inhibitor)group,with 6 mice per group.In the I/R,treatment,and inhibitor groups,the superior mesenteric artery was clamped for 45 minutes and then reperfused for 2 hours to establish the intestinal I/R injury model.The sham group underwent vascular dissection without occlusion.The treatment and inhibitor groups received dai-ly intraperitoneal injections of 15 mg/kg resveratrol for 5 d preoperatively,and the other two groups were intraperitoneally injected with an equal volume of 0.1%dimethyl sulfoxide.The inhibitor group additionally received 10 mg/kg XAV939 30 minutes pre-ischemia.HE staining was used to observe the pathological changes in the intestinal tissues,and the Chiu score standard was used to evaluate the pathological damage.Apoptosis was evaluated through TUNEL staining and the apoptotic index(AI).Diamine oxidase(DAO)level was measured by ELISA.The expression levels of B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein,cleaved caspase-3,Wnt3a and β-catenin proteins were detected by Western blot.RESULTS:Compared with sham group,the mice in I/R group showed significantly increased Chiu score,elevated DAO level and AI(P<0.05),up-regulated cleaved caspase-3 and Bax protein levels,and down-regulated Bcl-2,Wnt3a and β-catenin protein levels(P<0.05).Treatment with RSV reversed these alterations(P<0.05).XAV939 co-administration abolished the protective effects of RSV(P<0.05).CONCLUSION:Resveratrol ameliorates intestinal I/R injury by acti-vating the Wnt/β-catenin signaling pathway and suppressing apoptosis.

AIM:To construct a network calcula-tor based on machine learning(ML)models to pre-dict the risk of delayed awakening from anaesthesia in breast cancer(BC)patients.METHODS:A total of 435 BC patients surgically treated at our hospital from January 2023 to June 2024 were selected.The Boruta algorithm was used to screen for important characteristic variables for the risk of delayed awak-ening from anaesthesia.All patients were randomly assigned to a training set(n=261)and a test set(n=174)based on a 3:2 ratio and nine ML models were constructed and trained.Nine ML models were evaluated on the basis of receiver operating charac-teristic(ROC)curves for a random sample of 10 sub-jects and the clinical utility of the models was as-sessed using decision curve analysis.Combined with SHapley Additive exPlanations(SHAP)bar graphs,summary graphs and force diagrams additional in-terpretation and visualization of the ML model.Con-struction of a network calculator for predicting the risk of delayed awakening from anesthesia in BC pa-tients using the R package.RESULTS:Of the 435 BC patients,25.1%experienced delayed awakening from anesthesia.Boruta algorithm screened seven feature variables.The ROC curve shows that the XG-Boost model has the highest area under the curve(AUC)for 10 random samples among the 9 ML mod-els,and the decision curve shows that the XGBoost model has a significant clinical net benefit.The SHAP bar graph shows the importance of ASA classi-fication,surgery time,anesthesia time,intraopera-tive blood loss,propofol,preoperative anemia,and intraoperative hypothermia,and the SHAP summa-ry graph reflects the distribution of the ranges of in-fluence of the seven important characteristic vari-ables,which are"separated at the ends."The SHAP force diagram visualization XGBoost model predict-ed the risk of delayed awakening from anesthesia for individual patients with a predictive value of 0.998 for patients with delayed awakening from an-esthesia and 0.008 91 for patients without delayed awakening from anesthesia.A web-based calculator(https://xz-nomogram.shinyapps.io/DE_web/)based on an interpretable XGBoost model effective-ly predicts the risk of delayed awakening from anes-thesia in BC patients.CONCLUSION:ASA classifica-tion,surgery time,propofol,intraoperative blood loss,anaesthesia time,preoperative anaemia and intraoperative hypothermia are important charac-teristic variables for the risk of delayed awakening from anaesthesia in BC patients.The network calcu-lator based on the interpretable XGBoost model can accurately and quickly quantify the risk of de-layed awakening from anaesthesia,which can help clinicians to effectively adjust the treatment strate-gy and better improve the prognosis of patients.

AIM:To investigate the mechanism by which resveratrol(RSV)alleviates intestinal ischemia-re-perfusion(I/R)injury through modulation of the Wnt/β-catenin signaling pathway.METHODS:Twenty-four male C57BL/6J mice were randomly allocated into four experimental groups:the sham group,I/R group,I/R+RSV(treatment)group,and I/R+RSV+XAV939(inhibitor)group,with 6 mice per group.In the I/R,treatment,and inhibitor groups,the superior mesenteric artery was clamped for 45 minutes and then reperfused for 2 hours to establish the intestinal I/R injury model.The sham group underwent vascular dissection without occlusion.The treatment and inhibitor groups received dai-ly intraperitoneal injections of 15 mg/kg resveratrol for 5 d preoperatively,and the other two groups were intraperitoneally injected with an equal volume of 0.1%dimethyl sulfoxide.The inhibitor group additionally received 10 mg/kg XAV939 30 minutes pre-ischemia.HE staining was used to observe the pathological changes in the intestinal tissues,and the Chiu score standard was used to evaluate the pathological damage.Apoptosis was evaluated through TUNEL staining and the apoptotic index(AI).Diamine oxidase(DAO)level was measured by ELISA.The expression levels of B-cell lymphoma 2(Bcl-2),Bcl-2-associated X protein,cleaved caspase-3,Wnt3a and β-catenin proteins were detected by Western blot.RESULTS:Compared with sham group,the mice in I/R group showed significantly increased Chiu score,elevated DAO level and AI(P<0.05),up-regulated cleaved caspase-3 and Bax protein levels,and down-regulated Bcl-2,Wnt3a and β-catenin protein levels(P<0.05).Treatment with RSV reversed these alterations(P<0.05).XAV939 co-administration abolished the protective effects of RSV(P<0.05).CONCLUSION:Resveratrol ameliorates intestinal I/R injury by acti-vating the Wnt/β-catenin signaling pathway and suppressing apoptosis.

AIM:To construct a network calcula-tor based on machine learning(ML)models to pre-dict the risk of delayed awakening from anaesthesia in breast cancer(BC)patients.METHODS:A total of 435 BC patients surgically treated at our hospital from January 2023 to June 2024 were selected.The Boruta algorithm was used to screen for important characteristic variables for the risk of delayed awak-ening from anaesthesia.All patients were randomly assigned to a training set(n=261)and a test set(n=174)based on a 3:2 ratio and nine ML models were constructed and trained.Nine ML models were evaluated on the basis of receiver operating charac-teristic(ROC)curves for a random sample of 10 sub-jects and the clinical utility of the models was as-sessed using decision curve analysis.Combined with SHapley Additive exPlanations(SHAP)bar graphs,summary graphs and force diagrams additional in-terpretation and visualization of the ML model.Con-struction of a network calculator for predicting the risk of delayed awakening from anesthesia in BC pa-tients using the R package.RESULTS:Of the 435 BC patients,25.1%experienced delayed awakening from anesthesia.Boruta algorithm screened seven feature variables.The ROC curve shows that the XG-Boost model has the highest area under the curve(AUC)for 10 random samples among the 9 ML mod-els,and the decision curve shows that the XGBoost model has a significant clinical net benefit.The SHAP bar graph shows the importance of ASA classi-fication,surgery time,anesthesia time,intraopera-tive blood loss,propofol,preoperative anemia,and intraoperative hypothermia,and the SHAP summa-ry graph reflects the distribution of the ranges of in-fluence of the seven important characteristic vari-ables,which are"separated at the ends."The SHAP force diagram visualization XGBoost model predict-ed the risk of delayed awakening from anesthesia for individual patients with a predictive value of 0.998 for patients with delayed awakening from an-esthesia and 0.008 91 for patients without delayed awakening from anesthesia.A web-based calculator(https://xz-nomogram.shinyapps.io/DE_web/)based on an interpretable XGBoost model effective-ly predicts the risk of delayed awakening from anes-thesia in BC patients.CONCLUSION:ASA classifica-tion,surgery time,propofol,intraoperative blood loss,anaesthesia time,preoperative anaemia and intraoperative hypothermia are important charac-teristic variables for the risk of delayed awakening from anaesthesia in BC patients.The network calcu-lator based on the interpretable XGBoost model can accurately and quickly quantify the risk of de-layed awakening from anaesthesia,which can help clinicians to effectively adjust the treatment strate-gy and better improve the prognosis of patients.

Objective:To investigate the effect of dexmedetomidine(DEX)on the polarization of alveolar macrophages in-duced by lipopolysaccharide(LPS)and to explore the related mechanisms.Methods:Rat alveolar macrophages NR8383 were cul-tured in vitro.Experiment one was divided into control group,model group(1 μg/ml LPS),DEX low,medium and high dose groups(1,5,10 mg/kg DEX+10 mg/kg LPS).Experiment two was divided into DEX high dose group(10 mg/kg)and DEX high dose+Colive-lin(JAK2/STAT3 signaling pathway activator)group(10 mg/kg DEX+0.5 μmol/L Colivelin).The morphological changes of rat alveo-lar macrophages NR8383 were observed by inverted microscope;RT-PCR method was used to detect the expression levels of iNOS and Arg1 mRNA in NR8383 cells,and flow cytometry was used to detect the expression levels of CD86 and CD163 proteins in NR8383 cells;Western blot was used to detect the expression levels of surface marker proteins TNF-α,iNOS,SOCS,Arg1,TGF-β and JAK2/STAT3 signaling pathway related proteins in NR8383 cells.Results:Compared with control group,there were a lot of cell debris in the intercellular space of NR8383 in the model group,the proportions of iNOS mRNA,CD86 positive cells,and the expression levels of TNF-α,p-JAK2/JAK2,p-STAT3/STAT3 were significantly increased,the proportions of Arg1 mRNA,CD163 positive cells,and the expression levels of SOCS and TGF-β were significantly reduced(P<0.05);compared with the model group,the NR8383 intercellular cell debris in the DEX low,medium,and high dose groups were decreased,the proportions of iNOS mRNA,CD86 positive cells,and the expression levels of TNF-α,p-JAK2/JAK2,p-STAT3/STAT3 were significantly reduced,the proportions of Arg1 mRNA,CD163 positive cells,and the expression levels of SOCS and TGF-β were significantly increased(P<0.05).The reactivation of the JAK2/STAT3 signal pathway by Colivelin could weaken the role of DEX in LPS induced NR8383 cell polarization.Conclusion:DEX can inhibit the M1 polarization of NR8383 cells induced by LPS,which may be achieved by inhibiting the JAK2/STAT3 signaling pathway.

Objective To investigate potential causative associations between immunophenotype traits and cervical cancer by using two-sample Mendelian randomization(MR)analysis.Methods The genetic instrumental variables(IVs)of 731 immunophenotypes of peripheral blood were obtained from the GWAS Catalog database.The GWAS summary data of cervical cancer were obtained from FinnGen database.The inverse-variance weighted(IVW),weighted mode,weighted median,and MR Egger methods were used for evaluations.The sensitivity analysis and reverse Mendelian randomization analysis were conducted to eliminate bias and reverse causality.The MR Steiger directionality test was further used to ascertain the reverse causal relationship between immune cells and cervical cancer.Results A total of 71 immune cell subtypes associated with cervical cancer were identified,of which 31 had a strong association.The majority of the B cell panel was protective factors for cervical cancer.B-cell activating factor receptor(BAFF-R)was the most frequently expressed molecule in this analysis.It is expressed on several B cell subtypes.The CD20 on IgD+CD38+B cell(OR=1.887,95%CI:1.078-3.306,P=0.026)is the risk factor for cervical cancer.In cDC panels,the CD123 expression on plasmacytoid dendritic cell(OR=2.48,95%CI:1.229-5.003,P=0.011),CD123 expression on CD62L+plasmacytoid dendritic cell(OR=2.5,95%CI:1.231-5.077,P=0.011),CD80 expression on plasmacytoid dendritic cell(OR=2.62,95%CI:1.244-5.515,P=0.011),and CD80 expression on CD62L+plasmacytoid dendritic cell(OR=2.641,95%CI:1.246-5.596,P=0.011)were positively associated with the incidence of cervical cancer.All gynecological cancers in this study have no statistically significant effect on immune cells,according to reverse MR analysis.Conclusion This study emphasized the genetically predicted causality between immune cells and cervical cancer.In clinical practice,it is important to pay attention to the screening of peripheral blood immune cells for patients with cervical cancer.