Abstract:Objective To investigate the role of Cav1 gene in regulating mitochondrial function and mRNA m6A modification and expressions of the key genes in mouse hepatocytes.Methods In mouse hepatocyte AML12 cells,the effects of lentivirus-mediated Cav1 knockdown and transfection with a negative control virus on mitochondrial membrane potential and mitochondrial respiratory function were analyzed using the TMRE fluorescent probe and the Seahorse extracellular flux analysis system.Methylated RNA immunoprecipitation(MeRIP)combined with m6A microarray was utilized to evaluate the changes in mRNA m6A modification and expression levels,followed by enrichment analysis to identify the functionally relevant genes.The m6A modification and expression levels of the mRNAs were validated by qPCR.Results Compared with the negative control group,the cells with Cav1 knockdown exhibited significantly reduced mitochondrial membrane potential and respiratory capacity.m6A microarray analysis revealed significant changes in m6A modification levels(fold change>1.5)of 7814 mRNAs,including 152 upregulated and 7662 downregulated mRNAs.Integrated expression analysis identified 2497 mRNAs showing coordinated changes in m6A modification and expression levels.These mRNAs were enriched in the oxidative phosphorylation pathway,with Usp15,Suclg2,and Ppa2 exhibiting the highest percent changes in m6A modification.Both microarray and qPCR results showed that the m6A modification and expression levels of Usp15,Suclg2 and Ppa2 mRNAs were significantly reduced in cells with Cav1 knockdown compared to the NC group.Conclusion Cav1 knockdown induces significant alterations in mRNA m6A modification as well as their expression levels.The regulatory effects of Cav1 on mitochondrial function may be mediated by modulation of m6A modification of Usp15,Suclg2,and Ppa2 mRNAs.

Abstract:Objective To develop and validate a risk prediction model for cognitive impairment in community-dwelling elderly individuals in China.Methods This cross-sectional study was based on data from the 2011 China Health and Retirement Longitudinal Study(CHARLS),and the data of 2228 individuals aged≥60 years were analyzed.The participants were randomly divided into a training set(n=1560)and an internal validation set(n=668)in a 7∶3 ratio.Thirty-eight candidate variables were collected,covering sociodemographic characteristics,lifestyle and behavioral habits,chronic disease history,physical function,and self-rated health status.Feature selection was performed using the least absolute shrinkage and selection operator(LASSO)regression,followed by multivariate logistic regression to identify independent risk factors for cognitive impairment.A nomogram was constructed based on these factors,its discrimination power and calibration were assessed using the receiver operating characteristic(ROC)curve and calibration plot,respectively,and its clinical utility was evaluated using decision curve analysis(DCA).Results Age,years of education,alcohol consumption,systolic blood pressure,grip strength,and depressive symptoms were identified as independent predictors of cognitive impairment in Chinese elderly individuals.The area under the ROC curve of the constructed nomogram was 0.839(95%CI:0.814-0.864)in the training set and 0.840(95%CI:0.801-0.879)in the validation set,indicating good predictive performance of the model.The calibration plots demonstrated good agreement between the predicted and observed outcomes,and the DCA showed good clinical utility of the model.Conclusion The nomogram developed in this study based on LASSO-selected predictors demonstrates high accuracy,discrimination power,and potential clinical applicability to facilitate early identification and intervention of cognitive impairment among rural elderly individuals in China.

Abstract:Objective To explore therapeutic mechanism of Hugan Tang(Hugan Decoction,HGT)for alleviating non-alcoholic fatty liver disease(NAFLD)in rats.Methods Network pharmacology analysis was used to predict the active components of HGT against NAFLD and their potential targets,and the core targets were identified using the protein-protein interaction network,followed by GO and KEGG pathway enrichment analyses.A rat model of high-fat diet(HFD)-induced NAFLD was used to test the effects of saline,silymarin,and low-,moderate-,and high-dose HGT on serum levels of ALT,AST,LDL,LDH,TG and TC,liver histopathology,and protein and mRNA expressions of ACC1,FASN,AMPK and m-TOR.In free fatty acid(FFA)-induced HepG2 cells,the effects of blank and HGT-medicated sera,compound C(an AMPK inhibitor),and MHY1485(a mTOR agonist)were tested on cell viability,intracellular lipid deposition,TC and TG levels,and expressions of ACC1,FASN,AMPK and m-TOR.Results We identified 130 active components in HGT,267 common targets with NAFLD,and 53 core gene nodes,nearly half of which were involved in lipid metabolism.HGT treatment of NAFLD was closely associated with lipid and atherosclerosis signaling,insulin resistance signaling,and AMPK signaling.In rat models of NAFLD,HGT significantly alleviated liver injury and lipid accumulation,and suppressed mRNA and protein expressions of ACC1 and FASN.In FFA-induced HepG2 cells,HGT-medicated serum obviously reduced TG and TC levels and inhibited ACC1 and FASN mRNA and protein expressions.The results of in vitro and in vivo experiments both demonstrated that HGT activated the AMPK/mTOR signaling pathway by promoting p-AMPK expression and suppressing p-mTOR expression,and its regulatory effects on p-AMPK,p-mTOR,ACC1,and FASN were differentially modulated by compound C and MHY1485.Conclusion HGT alleviates NAFLD in rats by activating the AMPK/m-TOR signaling pathway and reducing lipid synthesis.

Abstract:Objective To explore the role of Chi3l1 in human umbilical cord mesenchymal stem cell(hUC-MSCs)therapy of type 1 diabetes.Methods hUC-MSCs with stable Chi3l1 knockdown(sh-Chi3l1-MSCs)were constructed using a lentiviral vector and characterized by flow cytometry and adipogenic and osteogenic induction.In adult C57BL/6J mouse models of streptozotocin-induced T1DM,the therapeutic effects of sh-NC-MSCs and sh-Chi3l1-MSCs grafting were evaluated by observing changes in clinical manifestations,blood glucose,body weight and pancreatic tissue pathologies.Insulin content and macrophage infiltration in the islets were detected using immunohistochemistry and immunofluorescence staining.The effects of these two stem cells on induced polarization of co-cultured mouse bone marrow macrophages were assessed using flow cytometry by detecting the mRNA expressions of iNOS,Arg-1,TNF-α,IL-6,IL-10,IL-13,and IL-1β using qPCR.Results The constructed sh-Chi3l1-MSCs retained the characteristics of MSCs but showed reduced therapeutic efficacy in T1DM mice.Immunofluorescence staining showed that the number of macrophages in the pancreatic tissue of the mice treated with sh-Chi3l1-MSCs was higher than that in MSCs treatment group.In the co-culture experiments,sh-Chi3l1-MSCs exhibited a lowered capacity to suppress M1 polarization of the macrophages and a reduced efficacy to promote differentiation of M2-type macrophage subset.Analysis with qPCR showed that the expressions of M1 macrophage marker iNOS and the inflammatory factors TNF-α,IL-6,and IL-1β increased,while the expressions of M2 macrophage marker Arg-1 and the cytokines IL-13 and IL-10 were decreased significantly in sh-Chi3l1-MSCs group.Conclusion In T1DM mouse models,hUC-MSCs mitigate inflammatory responses by suppressing the production of pro-inflammatory M1-type macrophages via Chi3l1.

Abstract:Objective To investigate the mechanism of kahweol for promoting motor function recovery in mice with spinal cord injury(SCI).Methods Fifty-four 8-to 10-week-old C57BL/6J mice were randomized equally into sham operation(laminectomy only)group,SCI group(laminectomy with spinal cord contusion),and Kahweol treatment group(with daily intraperitoneal injection of 20 mg/kg Kahweol following SCI).Motor function of the mice was evaluated using BMS scores,footprint analysis,and swimming test,and SCI area,myelin integrity,and neuron survival were assessed using HE,LFB,and Nissl staining.In a co-culture system of lipopolysaccharide(LPS)-stimulated BV2 cells and HT22 neurons,the effects of different concentrations of Kahweol and PMA,a NF-κB pathway activator,on the number of activated microglia and apoptotic neurons were evaluated with immunofluorescence staining,and the changes in apoptosis-related proteins and IκBα/NF-κB pathway proteins were detected using Western blotting.The levels of inflammatory factors(TNF-α,IL-6,and IL-1β)were measured by qRT-PCR and ELISA.Results In the mice with SCI,kahweol treatment significantly promoted motor function recovery,reduced injury area in the spinal cord tissue,and increased the myelinated area and number of neurons.In both the mouse models and the cell co-culture system,kahweol treatment effectively alleviated neuronal apoptosis by inhibiting microglial activation and reducing the release of inflammatory factors.The results of Western blotting showed that kahweol significantly decreased the phosphorylation levels of NF-κB and IκBα.In the cell co-culture system,PMA obviously attenuated the inhibitory effect of kahweol on BV2 cell activation and neuronal apoptosis.Conclusion Kahweol promotes motor function recovery of mice with SCI by suppressing microglial activation via inhibiting the NF-κB pathway,which shed light on a new strategy for clinical treatment of SCI.

Abstract:Objective To investigate the inhibitory effect of polyphyllin VII(PP7)on osteosarcoma xenograft growth in mice and explore the underlying molecular mechanism.Methods Ultra-performance liquid chromatography-tandem mass spectrometry was used to analyze the main active components of Paris polyphylla.Six nude mice bearing patient-derived xenograft(PDX)were randomized into two groups for treatment with 2 mg/kg PP7 gavage or saline every other day for 28 days,and the changes in tumor volume and mass were measured.In cultured 143B and HOS cells,the effect of PP7 treatment(0,1.25,2.5,5,and 10 μmol/L)on cell proliferation was assessed with CCK-8 assay,and Transwell assays were employed to examine the changes in cell migration and invasion.The target of PP7 was predicted by integrated analyses with single-cell RNA sequencing(scRNA-seq),bulk RNA sequencing(bulk RNA-seq)and molecular docking and verified using Western blotting.In osteosarcoma cells transfected with SOHLH1 siRNAs or a negative control sequence,the effects of PP7 treatment(5 μmol/L)on cell migration,invasion,ferroptosis,reactive oxygen species(ROS)production and lipid peroxidation(LPO)were analyzed.Results PP7 was identified as one of the major active constituents of Paris polyphylla.In the tumor-bearing mice,PP7 treatment significantly lower the tumor volume and mass.In 143B and HOS cells,PP7 concentration-dependently inhibited cell proliferation,and at 5 μmol/L,PP7 significantly inhibited cell proliferation,migration and invasion.Multi-omics analysis identified SOHLH1 as a potential target of PP7,and Western blotting confirmed that PP7 upregulated SOHLH1 expressions at both the mRNA and protein levels.SOHLH1 silencing obviously attenuated the inhibitory effects of PP7 on cell migration and invasion and reduced PP7-induced ferroptosis.Conclusion PP7 suppresses osteosarcoma xenograft growth in mice by inducing ferroptosis via upregulating SOHLH1 expression.

Abstract:Objective To investigate the mechanism by which Morinda officinalis polysaccharide(MOP)delays osteoarthritis chondrocyte degeneration.Methods In primary cultures of chondrocytes from 4-week-old C57BL/6 mice,the effects of IL-1β and MOP treatment at different concentrations on cell viability were assessed with CCK-8 assay.The treated cells were examined for protein expressions of PKM2,caspase-1,and GSDMD using Western blotting and for XIST expression using fluorescence in situ hybridization(FISH).In IL-1β-induced mouse chondrocytes,the effects of MOP,transfection for XIST overexpression or knockdown,and MOP treatment after the transfection were tested by detecting mRNA levels of GluT1,HK2,PKM2,LDHA,PFKFB3,NLRP3,caspase-1,and GSDMD;flow cytometry,Western blotting,and toluidine blue staining were used to analyze chondrocyte apoptosis,expressions of glycolysis and pyroptosis regulators,and glycosaminoglycan expression.Results The second-passage chondrocytes showed good viability and positive collagen II staining.IL-1β induction caused degenerative morphological changes of the cells,decreased collagen II expression,and upregulated cellular expressions of PKM2,caspase-1,and GSDMD proteins.MOP treatment(especially at 4 mg/mL)significantly enhanced cell viability and reduced HK2,PKM2,caspase-1 and GSDMD expressions in IL-1β-induced mouse chondrocytes.XIST was localized predominantly in the nuclei of the chondrocytes,and its expression increased significantly in IL-1β-treated cells,and was attenuated by MOP treatment.XIST overexpression synergized with IL-1β to upregulate mRNA and protein expressions of glycolysis-and pyroptosis-related factors in the chondrocytes,and such effects were obviously attenuated by MOP.Conversely,XIST knockdown significantly inhibited chondrocyte apoptosis and glycosaminoglycan expression,and down-regulated glycolysis-and pyroptosis-related proteins.MOP treatment exhibited similar protective effects to XIST knockdown,and their combination significantly augmented these protective effects.Conclusions MOP mitigates IL-1β-induced mouse chondrocyte degeneration by modulating glycolysis and pyroptosis via targeting XIST.

Abstract:Objective To identify the differentially expressed genes in obese asthma versus non-obese asthma and evaluate the effect of acupuncture on chronic airway inflammation in obese mice with asthma.Methods The key genes of obesity-related asthma were screened using GSE110551 dataset from the GEO database,and the characteristic genes were selected from the genes with the highest correlation with T cells using Lasso regression and SVM feature selection algorithms.Fifty C57BL/6J mice(5-6 weeks old)were randomized equally into 5 groups,including a normal feeding(control)group,a high-fat feeding group,and 3 high-fat feeding and ovalbumin sensitization groups with intraperitoneal injections with saline or dexamethasone(DEX),or treated with acupuncture.Western blotting,qPCR,and flow cytometry were used to analyze the changes in the expressions of the key genes and inflammation in the airway of the mice.Results FAM126B and VNN1 were identified as the characteristic genes in obesity-related asthma for subsequent analysis.The mice with high-fat feeding and ovalbumin sensitization showed the highest expression levels of Vnn1 and FAM126B among the 5 groups,with also significantly decreased Treg cell percentage and obvious inflammatory cell infiltration in the lungs.Treatment with DEX and acupuncture both significantly decreased the number of infiltrating inflammatory cells and increased the percentage of Treg cells in airway of the mouse models of obesity-related asthma.HIF-1α was identified as a key regulatory factor for asthmatic inflammation,and its expression level was significantly increased in the asthmatic mouse models but obviously lowered after acupuncture treatment or dexamethasone therapy.Conclusion Vnn1 and FAM126B may serve as the key therapeutic targets for treatment of obese asthma patients.Acupuncture treatment may downregulate airway HIF-1α by reducing the expressions of Vnn1 and FAM126B and increasing the number of Treg cells.

Abstract:Objective To evaluate the effect of resveratrol(RES)on barrier function of mouse brain microvascular endothelial cell monolayers exposed to oxygen/glucose deprivation/reoxygenation(OGD/R)and PM2.5 and explore the role of mitochondrial fission and fusion in protecting endothelial barrier function.Methods Cultured mouse brain microvascular endothelial cells were exposed to OGD/R,treated with PM2.5(100 μg/mL)before OGD/R,or pretreated with RES(40 mg/mL)prior to OGD/R+PM2.5 exposures.The changes in cell viability were examined with CCK-8 assay,and cell permeability was assessed by measuring transendothelial electrical resistance(TEER)and FITC-dextran permeation.Malondialdehyde(MDA)content and superoxide dismutase(SOD)activity were measured,and intracellular and mitochondrial ROS levels were detected using fluorescent probes.Mitochondrial morphology in the treated cells was observed using Mito-Tracker Red CMXRos.Western blotting was performed to detect the changes in cellular expressions of the tight junction proteins(ZO-1,occludin,and claudin-5)and mitochondrial dynamics-associated proteins(Drp1,Fis1,Mfn2,and OPA1).Results Compared with the normal control cells,the cells exposed to OGD/R or both OGD/R and PM2.5 showed significantly decreased TEER levels,increased permeability,elevated oxidative stress,and increased ROS fluorescence intensities.Obvious mitochondrial fragmentation and morphological changes in the mitochondria were observed in the exposed cells,which also showed decreased expressions of tight junction proteins and mitochondrial fusion proteins with increased expressions of mitochondrial fission proteins.RES pretreatment of the endothelial cells before the exposures significantly reduced membrane permeability,lowered ROS levels,improved mitochondrial morphology,increased expressions of tight junction and fusion proteins,and decreased fission protein expressions.Conclusion RES can protect barrier function of mouse brain microvascular endothelial cell monolayers exposed to OGD/R and PM2.5 by modulating mitochondrial dynamics,potentially through promoting mitochondrial fusion and inhibiting mitochondrial fission.

Abstract:Objective To investigate the molecular mechanism of helicid for improving depressive-like behaviors in rats exposed to chronic unpredictable mild stress(CUMS).Methods SD rats were randomly divided into normal control group(n=20)and CUMS group(n=70)to receive no stimulation and mild unpredictable stress for 6 weeks,respectively.After successful modeling,CUMS rats were further divided into 7 subgroups for intracerebroventricular injection with saline,adeno-associated virus(AAV)vector,or AAV carrying si-NCALD(NCALD silencing experiment,n=10);or intracerebroventricular injection with saline,saline with daily helicid gavage,AAV vector with helicid gavage,or NCALD-overexpressing AAV with helicid gavage(NCALD overexpresison experiment,n=10).The depressive state of the rats was evaluated by assessing changes in body weight,sucrose preference,and open field test.The expressions of NCALD,sGCα1,sGCβ1,PKG1/2,and cleaved-caspase 3 in the hippocampus of the rats were detected by Western blotting,and hippocampal cGMP level was determined with ELISA.Results Compared with the normal control rats,CUMS rats showed significantly increased hippocampal expressions of NCALD and cleaved caspase-3 and abnormal activation of the sGC/cGMP/PKG pathway.Silencing NCALD by intracerebroventricular injection of AAV-si-NCALD significantly reduced cleaved caspase-3 and inhibited sGC/cGMP/PKG pathway activation in the hippocampus,and improved depressive-like behaviors of the rats.Helicid treatment produced similar effects,but its effect was abolished by intracerebroventricular injection of NCALD-overex-pressing AAV.Conclusion Helicid relieves depressive-like behaviors in CUMS rats by downregulating NCALD,inhibiting abnormal sGC/cGMP/PKG activation,and reducing hippocampal apoptosis.

Abstract:Objective To evaluate the protective effect of Lonicerae Japonicae Flos(LJF)extract against doxorubicin(DOX)-induced cardiotoxicity(DIC)and explore the possible mechanisms.Methods Network pharmacology,bioinformatics analysis and molecular docking were used to predict the targets of the core components of LJF.In a mouse model of DOX-induced myocardial injury,the protective effects of different doses of LJF extract were evaluated and the underlying mechanisms were explored by detecting the changes in mouse myocardial functions,myocardial enzymes,myocardial pathologies,and the expressions of inflammatory factors and pyroptosis-related proteins.Results The 10 core ingredients of LJF showed strong binding to AKT,EGFR,and GSK3β.In the animal experiment,the DOX-treated mice,compared with the sham-treated mice,had significantly decreased cardiac output,stroke volume,left ventricular ejection fraction,left ventricular fraction shorting,elevated serum levels of CK-MB and LDH,increased myocardial expressions of IL-18 and IL-1β,obvious myocardial damage,increased expression levels of NLRP3,caspase-1,GSDMD and GSDMD-N,and reduced expressions of EGFR,p-AKT and p-GSK3β proteins in the myocardial tissues.LJF treatment obviously improved myocardial function,decreased myocardial expressions of IL-18,IL-1β,NLRP3,caspase-1,GSDMD and GSDMD-N proteins,and increased the expressions EGFR,p-AKT and p-GSK3β proteins in DOX-treated mice.Conclusion LJF extract alleviates DOX-induced myocardial injury in mice possibly by reducing myocardial inflammation and pyroptosis via targeting EGFR,AKT and GSK3β to regulate the ErbB signaling pathway.

Abstract:Objective To explore the mechanism of Shenqi Xiezhuo Decoction(SQXZD)for improving renal fibrosis(RF)in rats.Methods The chemical components of SQXZD were identified using UPLC-Q Exactive/MS,and component-disease target network and enrichment analyses were conducted to screen the key pathways and targets.In the animal experiment,49 male SD rats were randomized equally into blank control group,sham operation group,unilateral ureteral obstruction-induced RF model group,losartan treatment(daily dose 4.6 mg/kg)group,and low-,medium-,and high-dose SQXZD(9.7,19.4,and 38.8 g/kg,respectively)treatment groups.After 14 days'treatment,renal pathologies and collagen deposition of the rats were examined with HE and Masson staining,and serum levels of BUN,Cr,SOD,MDA,GSH-px,IL-6,and TNF-α were detected.Western blotting and qRT-PCR were used to detect renal protein and mRNA expressions of α-SMA,Col-I,NAKED2,Rap1,B-raf,Raf-1,MEK3/6,p38MAPK,MEK,ERK1/2,p-ERK1/2,FoxO3a,p-FoxO3a,and MnSOD.Results A total of 263 chemical components were identified in SQXZD.Network pharmacology revealed 170 intersecting targets between the components and RF enriched in the MAPK,Rap1,and FoxO pathways.The rat models of RF showed abnormal renal structural changes,increased fibrosis area,elevated serum BUN,Cr,MDA,IL-6,and TNF-α levels,reduced SOD and GSH-px levels,upregulated renal expressions of α-SMA,Col-I,NAKED2,Rap1,B-raf,MEK,ERK1/2,p-ERK1/2,MEK3/6,and p38MAPK,and downregulated Raf-1,FoxO3a,p-FoxO3a,and MnSOD expressions.Treatment with losartan and SQXZD(especially at the medium dose)obviously lessened renal pathologies,improved renal functions,alleviated oxidative stress and inflammation,and ameliorated abnormal changes in the Rap1/MAPK/FoxO3a signaling pathway in the rat models.Conclusion SQXZD alleviates RF and improves renal function in rats possibly by ameliorating renal oxidative stress and inflammation via regulating the Rap1/MAPK/FoxO3a signaling pathway.

Abstract:Objective We propose an efficient deep learning model to improve the classification accuracy in automatic classification tasks of 12-lead electrocardiogram(ECG)signals.Methods We designed a new ResLSTM-TemporalSE network architecture by incorporating a multi-layer Residual Long Short-Term Memory(ResLSTM)structure and introducing skip connections between LSTM layers to establish residual learning pathways for the temporal features.A temporal attention mechanism was integrated into the traditional Squeeze-and-Excitation(SE)module to enhance channel-wise feature representation while capturing long-term temporal dependencies within ECG signals,thereby an efficient hierarchical feature extraction framework was constructed.The model was validated using the public CPSC2018 dataset and a private clinical dataset from the Seventh Affiliated Hospital of Southern Medical University.Results The experimental results demonstrated that the model achieved a classification accuracy of 99.70%on the CPSC2018 test set,with precision,recall,and F1-score values of 0.9966,0.9370,and 0.9653,respectively.On the private clinical dataset,it attained an accuracy of 82.77%,with precision,recall,and F1-score values of 0.6811,0.8961,and 0.7723.Ablation studies confirmed the significant contributions of both the residual connections and the temporal attention module to model performance.Conclusion The ResLSTM-TemporalSE model effectively integrates spatiotemporal features of the ECG signals and demonstrates superior classification performance on the CPSC2018 benchmark while maintaining strong generalization capabilities in real-world clinical settings.This framework provides a robust solution for automated ECG analysis and holds significant promise for clinical applications.

Abstract:Objective To investigate the causal relationship between gut microbiota,T-cell function,and the risk of colorectal cancer.Methods Gut microbiota data from the MiBioGen database and T-cell and colorectal cancer data from publicly available GWAS datasets were obtained for analyzing the causality between gut microbiota,T-cell subsets,and the risk of colorectal cancer with two-sample Mendelian randomization(MR)analyses,using inverse variance weighting as the primary analytical method supplemented with MR-Egger,weighted median,simple mode,and weighted mode methods.Horizontal pleiotropy was assessed using MR-PRESSO and MR-Egger regression.Cochran's Q test was used to evaluate heterogeneity,and sensitivity analysis was performed using the leave-one-out method.Results In the Forward MR analysis of gut microbiota and T cells,11 gut microbiota species showed causal relationships.Six of these species exhibited positive correlations with T cells,including Prevotella7(P=0.003),Ruminococcaceae UCG011(P=0.033),Ruminococcaceae UCG004(0.010),Ebacterium brachy group(P=0.005),Lachnospiraceae FCS020 group(P=0.028),and Coprobacter(P=0.033),and the remaining 5 species showed negative correlations with T cells.Forward MR analysis of T cells and colorectal cancer suggested that CD25++CD45RA-CD4+non-regulatory T cells were negatively correlated with colorectal cancer risk(IVW:OR=0.935,95%CI:0.878-0.995;P=0.035).The analysis of gut microbiota and colorectal cancer suggested that 11 gut microbiota species were causally associated with colorectal cancer,and 6 of them(Eubacterium xylanophilum group,P=0.039;Selenomonadales,P=0.014;Negativicutes,P=0.014;Bifidobacteriaceae,P=0.048;Bifidobacteriales,P=0.048;and Coprococcus1,P=0.033)showed positive correlations and the remaining 5 showed negative correlations.Conclusion Coprobacter spp.and Eubacterium xylanophilum group spp.are causally associated with both T cell activity and colorectal cancer risk,and the former bacteria induce inactivation of CD25++CD45RA-CD4+non-regulatory T cells to promote colorectal cancer progression,whereas the latter bacteria promote CD25++CD45RA-CD4+non-regulatory T cell activity to inhibit colorectal cancer development.

Abstract:Objective To quantitatively analyze setup errors of 4 immobilization devices in precision radiotherapy for prostate cancer,their accuracy differences,and the factors affecting their setup precisions.Methods We conducted a retrospective analysis of 240 prostate cancer patients undergoing image-guided radiotherapy at Sun Yat-sen University Cancer Center from May,2016 to May,2024.According to the immobilization devices used,the patients were divided into 1.2 m vacuum bag group(n=60),1.8 m vacuum bag group(n=60),Orfit frame group(n=60),and customized prone board group(n=60).All the patients received pre-treatment cone-beam CT(CBCT)scans,and setup errors in the right-left(RL),superior-inferior(SI),and anterior-posterior(AP)directions were obtained through XVI system grayscale registration.Further subgroup analyses were performed based on patient stratifications by lymph node irradiation status(n=120 each),age(<65 years,n=80;≥65 years,n=160),and BMI(BMI<24 kg/m²,n=120;BMI≥24 kg/m²,n=120).Results The setup errors differed significantly among the 4 groups in three-dimensional directions(P<0.05).The customized prone board group showed minimal errors in the RL(0.02±0.25 cm)and SI(0.01±0.32 cm)directions,but demonstrated the largest error in the AP direction(-0.28±0.36 cm).The patients with lymph node irradiation had significantly greater AP directional errors(-0.22±0.36 cm)than those without(-0.01±0.43 cm;P<0.001).BMI showed a negative correlation with SI directional errors(R=-0.45,P<0.001),while age was not significantly correlated with the setup errors(P>0.05).Conclusion The customized prone board demonstrates clinically significant advantages for its high setup accuracies in RL and SI directions in spite of its systematic AP directional errors.The setup accuracy in the SI direction is especially important for patients with lymph node irradiation or low BMI.Our findings provide quantitative evidence for immobilization device selection and individualized optimization of precision radiotherapy for prostate cancer.

Abstract:Objective To investigate the potential mechanism underlying the protective effect of secoisolariciresinol diglucoside(SDG)against chronic kidney disease(CKD)in offspring mice caused by maternal exposure to trans fatty acids(TFA)during pregnancy and lactation.Methods Thirty female C57BL/6 mice were randomized into control group,TFA model group,and 3 TFA model groups treated with SDG at low,medium and high doses(10,20 and 30 mg/kg,respectively).The changes in blood urea nitrogen(BUN)and serum creatinine(CRE)levels of the mice were measured.Network pharmacology analysis was conducted to explore protective mechanism of SDG against TFA-induced renal injury,and molecular docking was used to assess the binding affinity of SDG to Bcl-2,Bax,and caspase-3.The protein expressions of cleaved caspase-3,Bax,and Bcl-2 in the renal tissues of the offspring mice were detected with Western blotting.Result The mice in TFA group showed significantly higher BUN and CRE levels than those in the control group.Treatment with SDG at the medium and high doses significantly reduced BUN and CRE levels in the mouse models.Network pharmacology and molecular docking suggested that SDG ameliorated renal injury by targeting the apoptosis-related Bcl-2/Bax/caspase-3 axis.The results of Western blotting showed the mouse models in TFA exposure group had increased renal cell apoptosis with elevated expression levels of cleaved caspase-3 protein and a decreased Bcl-2/Bax ratio(P<0.05),and intervention with SDG at all the 3 doses significantly reduced renal cell apoptosis and renal expression of cleaved caspase-3 and increased the Bcl-2/Bax ratio in the mouse models.Conclusion Maternal TFA exposure during gestation and lactation induces renal injury in offspring mice.Dietary SDG intervention can mitigate TFA-induced renal injury in offspring mice possibly by suppressing renal cell apoptosis via regulating the Bcl-2/Bax/caspase-3 signaling axis.

Abstract:Objective To investigate the clinical significance of abnormal expression of angio-associated migratory cell protein(AAMP)in hepatocellular carcinoma(HCC).Methods Bioinformatics analyses were used to analyze AAMP expression level in HCC and its prognosis value.In 60 pairs of HCC and adjacent tissues,AAMP expression was detected immunohistochemically and its correlation with clinicopathological characteristics of the patients was analyzed.In cultured Mahlavu and Huh-7 cells with lentivirus-mediated AAMP knockdown,the changes in cell proliferation,apoptosis,migration and invasion were observed,and their lung metastasis following tail vein injection in nude mice were assessed.In HCC cells with AAMP knockdown,Western blotting,immunofluorescence staining or RT-qPCR were used to examine the changes in expression levels of E-cadherin,N-cadherin,Vimentin and Snail and the effects of MG-132 and CHX on RhoA expression.The correlation between the expressions of AAMP and RhoA in HCC tissues was analyzed by immunohistochemistry.Results Bioinformatics analysis showed that AAMP expression was elevated in HCC tissues(P<0.05)in correlation with advanced clinical stage and poor prognosis(P<0.05).Immunohistochemistry results confirmed significant correlations of high AAMP expression with Edmondson-Steiner grade(III+IV),venous infiltration and TNM stage(III+IV)of HCC(P<0.05).In cultured HCC cells,AAMP knockdown did not significantly affect cell proliferation or apoptosis,but obviously suppressed cell migration and invasion in vitro and lung metastasis in nude mice.AAMP knockdown significantly increased E-cadherin expression,decreased N-cadherin,Vimentin and Snail expressions,and reduced RhoA protein levels without obviously affecting RhoA mRNA levels.MG-132 treatment blocked the inhibitory effect of AAMP knockdown on RhoA protein expression.The expressions of AAMP and RhoA showed a significant positive correlation in HCC tissues(P<0.05).Conclusion AAMP overexpression is associated with malignant clinical features of HCC and promotes epithelial-mesenchymal transition and metastasis of HCC cells partly by stabilizing RhoA expression.

Abstract:Objective To investigate the association of preoperative serum magnesium(sMg)level with postoperative delirium(POD)in elderly surgical patients and the mediating role of systemic inflammation.Methods This retrospective cohort study was conducted among 12 876 patients aged≥65 years undergoing non-cardiac,non-neurological surgeries at Chinese PLA General Hospital between January,2014 and December,2021.Preoperative sMg and C-reactive protein(CRP)levels were measured within 30 days before surgery.POD was identified within 7 days postoperatively using structured chart review based on the Confusion Assessment Method.Multivariate logistic regression and restricted cubic spline(RCS)models were used to evaluate the association between sMg and POD.Mediation analysis with structural equation modeling was used to quantify the indirect effect of CRP after adjusting for the confounding factors.Results POD was identified in 685(5.3%)of the patients.A significant nonlinear association was observed between preoperative sMg levels and POD risk,and POD incidence was the lowest in patients with sMg levels of 0.90-0.94 mmol/L.Compared with those in the 4th quintile,the patients in the lowest quintile exhibited a markedly increased risk of POD(OR=1.81,95%CI:1.41-2.35)even after adjustment for multiple confounding factors.Mediation analysis suggested that CRP explained 17.1%of the total effect of sMg on POD risk,and a stronger mediating effect was observed in cancer as compared with the non-cancer patients(24.1%vs 11.9%).Subgroup analyses revealed a significant nonlinear relationship between sMg and POD particularly in cancer patients and patients beyond 75 years of age.Conclusion Preoperative sMg level is independently associated with an increased POD risk in elderly patients,mediated partly by systemic inflammation.sMg may serve as a modifiable biomarker for early risk stratification and prevention for POD in perioperative care.

Abstract:Objective To investigate the impact of high expression of transmembrane and coiled helix structural domain 1(TMCO1)on prognosis of gastric cancer and the possible mechanisms.Methods TMCO1 expression in gastric cancer and its effect on gastric cancer progression and prognosis were analyzed using publicly available databases and clinical data of patients undergoing radical surgery in our hospital,and its possible biological functions were explored using KEGG and GO analyses.In gastric cancer HGC-27 cells,the effects of lentivirus-mediated TMCO1 overexpression and TMCO1 silencing on cell apoptosis,proliferation,invasion and migration were examined.Results TMCO1 expression was significantly elevated in gastric cancer tissues(P<0.05),and its high expression was positively correlated with cancer progression(P<0.001)and a lowered postoperative 5-year survival rate of the patients(P<0.05).Bioinformatic analyses suggested that TMCO1 may affect gastric cancer cell apoptosis via Wnt signaling.In HGC-27 cells,TMCO1 overexpression significantly promoted tumor cell proliferation,inhibited cell apoptosis,and enhanced cell migration and invasion,whereas TMCO1 silencing produced the opposite effects.Western blotting showed that β-catenin levels were significantly upregulated in TMCO1-overexpressing cells and downregulated in cells with TMCO1 silencing.Conclusion TMCO1 is overexpressed in gastric cancer tissues,and its high expression promotes gastric cancer progression and affects long-term prognosis of the patients possibly by activating the Wnt/β-catenin signaling pathway to inhibit apoptosis of gastric cancer cells.

Abstract:Objective To develop a heterogeneous graph prediction method based on the fusion of multi-layer semantics and topological information for addressing the challenges in drug-target interaction prediction,including insufficient modeling of high-order semantic dependencies,lack of adaptive fusion of semantic paths,and over-smoothing of node features.Methods A heterogeneous graph network with multiple types of entities such as drugs,proteins,side effects,and diseases was constructed,and graph embedding techniques were used to obtain low-dimensional feature representations.An adaptive metapath search module was introduced to automatically discover semantic path combinations for guiding the propagation of high-order semantic information.A semantic aggregation mechanism integrating multi-head attention was designed to automatically learn the importance of each semantic path based on contextual information and achieve differentiated aggregation and dynamic fusion among paths.A structure-aware gated graph convolutional module was then incorporated to regulate the feature propagation intensity for suppressing redundant information and redcuing over-smoothing.Finally,the potential interactions between drugs and targets were predicted through an inner product operation.Results Compared with existing drug-target interaction prediction methods,the proposed method achieved an average improvement of 3.4%and 2.4%,3.0%and 3.8%in terms of the area under the receiver operating characteristic curve(AUC)and the area under the precision-recall curve(AUPRC)on public datasets,respectively.Conclusion The drug-target interaction prediction method developed in this study can effectively extract complex high-order semantic and topological information from heterogeneous biological networks,thereby improving the accuracy and stability of drug-target interaction prediction.This method provides technical support and theoretical foundation for precise drug target discovery and targeted treatment of complex diseases.