ObjectiveTo decipher the mechanism by which Zuoguiwan （ZGW） treat hyperthyroidism in rats with kidney-Yin deficiency based on the dopamine receptor D4 （DRD4）/nicotinamide adenine dinucleotide phosphate （NADPH） oxidase 4 （NOX4） signaling pathway. MethodsThe rat model of kidney-Yin deficiency was induced by unilateral intramuscular injection of dexamethasone （0.35 mg·kg^-1）. After successful modeling， the rats were randomized into model， methimazole （positive control， 5 mg·kg^-1）， low-， medium-， and high-dose （1.85， 3.70， 7.40 g·kg^-1， respectively） ZGW， and normal control groups. After 21 days of continuous gavage， the behavioral indexes and body weight changes of rats were evaluated. The pathological changes of the renal tissue were observed by hematoxylin-eosin staining. The serum levels of thyroid hormones ［triiodothyronine （T₃）， thyroxine （T₄）， thyroid-stimulating hormone （TSH）］， renal function indexes ［serum creatine （Scr） and blood urea nitrogen （BUN）］， energy metabolism markers ［cyclic adenosine monophosphate （cAMP） and cyclic guanosine monophosphate （cGMP）］， and oxidative stress-related factors ［superoxide dismutase （SOD）， malondialdehyde （MDA）， and NADPH）］ were measured by enzyme-linked immunosorbent assay （ELISA）. Western blot was employed to analyze the expression of DRD4， NOX4， mitochondrial respiratory chain complex proteins ［NADH：ubiquinone oxidoreductase subunit S4 （NDUFS4） and cytochrome C oxidase subunit 4 （COX4）］， and inflammation-related protein ［tumor necrosis factor-alpha （TNF-α）， interleukin-6 （IL-6）， p38 mitogen-activated protein kinase （MAPK）］ pathway in the renal tissue. ResultsCompared with the normal group， the model group showed mental malaise， body weight decreases （P<0.01）， inflammatory cell infiltration in the renal tissue， a few residual parotid glands in the thyroid， elevations in serum levels of T₃， T₄， Scr， BUN， cAMP， cAMP/cGMP， MDA， and NADPH （P<0.01）， down-regulation in protein levels of TSH， SOD， and DRD4 （P<0.05， P<0.01）， and up-regulation in expression of NOX4， p-p38 MAPK/p38 MAPK， and inflammatory factors （P<0.01）. Compared with the model group， ZGW increased the body weight （P<0.05， P<0.01）， reduced the infiltration of renal interstitial inflammatory cells， restored the thyroid structure and follicle size， lowered the serum levels of T₃， T₄， Scr， BUN， cAMP， cAMP/cGMP， MDA and NADPH （P<0.05， P<0.01）， up-regulated the expression of TSH， SOD and DRD4 （P<0.05， P<0.01）， and down-regulated the expression of NOX4， p-p38 MAPK/p38 MAPK， and inflammatory factors （P<0.05， P<0.01）. Moreover， high-dose ZGW outperformed methimazole （P<0.05）. ConclusionBy activating DRD4， ZGW can inhibit the expression of NOX4 mediated by the p38 MAPK pathway， reduce oxidative stress and inflammatory response， thereby ameliorating the pathological state of hyperthyroidism due to kidney-Yin deficiency. This study provides new molecular mechanism support for the clinical application of ZGW.

Background/Aims: Liver transplantation (LT) following total hepatectomy is a life-saving treatment for hepatocellular carcinoma (HCC). The HCC recurrence after LT hinders the effectiveness of the procedure. The objective of this study is to develop a pre-operative risk stratification model based on a liquid biopsy. Methods: We conducted a comprehensive multi-omics study of 260 HCC patients from three centers, including clinical data, low-coverage whole-genome sequencing of cell-free DNA (cfDNA) from plasma, as well as whole-exome, single-nucleus RNA, and spatial transcriptomics from matched tumor and non-tumor tissues. Results: We identified cfDNA-derived copy number alteration (CNA) signatures associated with post-transplant recurrence. By integrating cfDNA-derived CNA profiles with single-cell transcriptomic data, we traced recurrence-associated cfDNA to a distinct subpopulation of malignant cells within the primary tumor. These cells were embedded in a pro-metastatic microenvironment of specialized endothelial subtypes and cancer-associated fibroblasts. Notably, most recurrence-associated lesions were detectable in cfDNA prior to liver transplantation (LT). Building on these insights, we developed the ZJU Criteria based on CNA fragments and tumor markers, a pre-LT risk prediction tool that integrates conventional clinical factors with cfDNA-derived CNA signatures, and validated it using internal and independent external cohorts. Conclusion Our findings suggest that post-transplant recurrence commonly originates from advanced subclones that emerge late during tumor evolution. The ZJU Criteria provides an accurate, non-invasive strategy that significantly improves pre-LT risk stratification and clinical decision-making for patients with HCC.

Objective:To evaluate the effectiveness of entrustable professional activities (EPAs) in the general practice internship of undergraduate clinical medicine students, identify issues that need improvement in the internship, and enhance medical students' competence.Methods:A total of 75 students in the five-year (English class) clinical medicine program enrolled in 2018 and 2019 who participated in general practice internship in Renji Hospital affiliated to Shanghai Jiao Tong University School of Medicine from October 2021 to October 2023 were selected as study subjects. The design of core EPAs was adopted to assess the correlation among different EPA dimensions and to analyze the qualified rates.Results:The evaluation of EPAs showed that EPA2 (practicing respect, understanding and teamwork) had the highest mean score of 9.33, and EPA10 (chronic disease management and management of key populations) had the lowest mean score of 8.08. A supervision level of 3a and above was used as the criterion for qualification. The supervision levels of the students' EPAs were mostly concentrated at levels 3a and 3b. The highest qualified rate was for EPA2 (practicing respect, understanding and teamwork) at 85.33%, followed by EPA1 (complying with the rules of the profession and demonstrating professionalism) at 80.00% and EPA8 (reviewing information and solving clinical problems) at 72.00%. The lowest qualified rate was for EPA10 (chronic disease management and management of key populations) at 33.33%, followed by EPA4 (analyzing and interpreting test results) at 57.33%.Conclusions:EPAs concretize competency evaluation, which can effectively reflect the "competency-oriented" training objectives encompassing multiple elements such as knowledge, skills, values, and attitudes, while maintaining professional specificity. Undergraduates demonstrated strengths in professionalism and academics, but showed deficiencies in community chronic disease management and management of key populations. These findings suggest the need to strengthen the training in health and social care to better align with the competencies required during standardized residency training.

Objective To investigate the mechanism of the miR-518a-5p/histone deacetylase 6(HDAC6)axis in DNA oxidative damage in ovarian cancer(OC)SKOV3 cells.Methods Expression levels of miR-518a-5p and HDAC6 mRNA in OC tissues and in various cancer cells(A2780,SKOV3,CAOV3)were detected by qRT-PCR.SKOV3 cells were separated into Control,miR-NC,miR-518a-5p mimics,miR-518a-5p mimics+pcDNA-NC,and miR-518a-5p mimics+pc-HDAC6 groups.Cell proliferation and apoptosis were analyzed by colony-forming assay and Hoechst 33258 staining.Expression of phosphorylated histone H2AX(γ-H2AX)was detected by immunofluorescence assay and reactive oxygen species(ROS)were detected by flow cytometry.HDAC6,Bcl-2-associated X protein(Bax),and B-cell lymphoma-2(Bcl-2)protein expression were analyzed by Western blot.The regulatory relationship between miR-518a-5p and HDAC6 was analyzed by dual luciferase assay.The effect and mechanism of miR-518a-5p on oxidative DNA damage in OC cells were studied in a xenotransplantation tumor model.Results miR-518a-5p expression was decreased and HDAC6 expression was increased in OC tissues and A2780,SKOV3,and CAOV3 cells(P＜0.001).Expression levels of miR-518a-5p were lowest and expression levels of HDAC6 were highest in SKOV3 cells,and SKOV3 cells were therefore selected for subsequent experiments.miR-518a-5p expression,apoptosis rate,number of γ-H2AX-positive cells,relative ROS fluorescence intensity,and expression of Bax were all higher in the miR-518a-5p mimics group compared with the miR-NC group,while HDAC6 mRNA and protein expression,Bcl-2 expression,and colony-formation number were all lower(P＜0.001).HDAC6 mRNA and protein expression,colony-formation number,and expression of Bcl-2 were higher in the miR-518a-5p mimics+pc-HDAC6 group compared with the miR-518a-5p mimics+pcDNA-NC group,and the apoptosis rate,number of γ-H2AX-positive cells,relative ROS fluorescence intensity,and expression of Bax were all lower(P＜0.001).HDAC6 had a targeted regulatory relationship with miR-518a-5p.Overexpression of miR-518a-5p decreased tumor volume,weight,and HDAC6 protein expression in tumor tissues,and increased γ-H2AX expression in vivo(P＜0.001).Upregulation of HDAC6 expression by overexpression of miR-518a-5p increased graft tumor volume,weight,and HDAC6 protein expression and decreased γ-H2AX-positive expression(P＜0.05).Conclusions miR-518a-5p expression is reduced and HDAC6 expression is increased in OC tissues and cells.Overexpression of miR-518a-5p can induce oxidative DNA damage in SKOV3 cells by inhibiting HDAC6 expression,thereby inhibiting cell proliferation and promoting cell apoptosis.

Objective:The transcriptome data was utilized to screen cuproptosis-related genes(CRGs)in oral squamous cell car-cinoma(OSCC),and the characteristic genes were identified for constructing a prognostic model for predicting patients'survival time.Methods:OSCC transcriptome gene expression and clinical data were obtained from TCGA and GEO.Through Lasso regres-sion analysis and Cox regression analysis,relevant prognostic genes were screened and prognostic models were constructed.Ac-cording to the median value of risk scores,patients were divided into high and low risk groups,and their survival rates were com-pared.Finally,the predictive performance of the model was verified.Results:In this study,9 characteristic genes with prognostic value(ENO2,P4HA1,SLC2A3,AQP1,PLS1,NXPH4,CTSG,TRAC,THBS1)were screened out and a 9-gene prognostic model was constructed.The survival rate of high-risk group based on prognostic model was significantly lower than that of low-risk group.The area under curve(AUC)of receiver operating characteristic curve(ROC)was 0.701,0.729 and 0.702 at 1 year,3 years and 5 years,respectively,which verifies that the risk model has good predictive performance.The nomogram predicted that the 1-year,3-year,and 5-year survival probabilities of OSCC patients are 89.6％,72.4％,and 63.9％respectively,and the cal-ibration curve confirmed the accuracy of the nomogram prediction.Conclusion:The 9-gene prognostic model based on CRGs screening could predict the prognosis of OSCC patients,which is helpful for clinical personalized treatment of OSCC patients and prediction of their survival rate.

Objective:To compare the efficacy of different doses of tranexamic acid (TXA) for traumatic hemorrhagic shock (THS) in the early phase of trauma following acute exposure to high altitude in rabbits.Methods:Twenty-five healthy male New Zealand rabbits were randomly divided into plain control group ( n=5) and acute high-altitude THS group ( n=20) according to the random number table method. The plain control group did not undergo THS modeling throughout the experiment while the acute high-altitude THS group was raised in a hypoxia simulation chamber with a volume fraction of 10% for 3 days to establish the THS model. Based on the different doses of TXA administered intravenously at 30 minutes after THS modeling, the acute high-altitude THS group was further divided into four subgroups: acute high-altitude THS+0 mg/kg TXA subgroup, acute high-altitude THS+45 mg/kg TXA subgroup, acute high-altitude THS+90 mg/kg TXA subgroup and acute high-altitude THS+135 mg/kg TXA subgroup, with 5 rabbits in each. The vital signs [mean arterial pressure (MAP), heart rate, rectal temperature] and blood cell counts [red blood cell count (RBC), platelet count (PLT)], 4 coagulation parameters [fibrinogen (FIB), D-dimer, activated partial thromboplastin time (APTT), prothrombin time (PT)], thromboelastography [clotting reaction time (R value), clot formation time (K value), maximum amplitude (MA value)], syndecan-1, inflammatory factors [interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α)], and plasminogen activator inhibitor-1 (PAI-1) were recorded before blood loss, at 30 minutes and 120 minutes after blood loss. At 6 hours after THS, the lungs, terminal ileum, and kidneys of the rabbits were collected to observe tissue damage, and the wet/dry weight ratio (W/D) and total water content (TLW) of the lung tissue were measured. Results:(1) Vital signs: Before blood loss, there were no significant differences in MAP, heart rate, or rectal temperature between the acute high-altitude THS subgroups and the plain control group ( P>0.05). At 30 minutes and 120 minutes after blood loss, the acute high-altitude THS subgroups exhibited significantly lower MAP, heart rate, and rectal temperature compared to those in the plain control group ( P<0.05). No significant differences were observed in MAP, heart rate or rectal temperature among the acute high-altitude THS subgroups at any time point ( P>0.05). In the acute high-altitude THS subgroups, MAP, heart rate and rectal temperature were significantly decreased at 30 minutes and 120 minutes after blood loss compared to those before blood loss ( P<0.05); At 120 minutes after blood loss, these parameters were further significantly decreased compared to those at 30 minutes after blood loss ( P<0.05). (2) Blood cell counts: Before blood loss, the RBC count was significantly higher in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), while the PLT was significantly lower ( P<0.05). At 30 minutes after blood loss, there was no significant difference in RBC count between the acute high-altitude THS subgroups and the plain control group ( P>0.05), but the PLT remained significantly lower in the acute high-altitude THS subgroups ( P<0.05). At 120 minutes after blood loss, the RBC count was significantly lower in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), with no significant differences among the acute high-altitude THS subgroups ( P>0.05). The PLT count was significantly lower in the acute high-altitude THS+0 mg/kg TXA subgroup compared to the other subgroups ( P<0.05). The PLT count in the acute high-altitude THS+45 mg/kg TXA subgroup was significantly lower than those in the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P<0.05), with no significant differences between the latter two subgroups ( P>0.05). (3) Four Coagulation parameters: Before blood loss, D-dimer level was significantly higher in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), while no significant difference was observed in FIB ( P>0.05). APTT and PT were significantly shortened in the acute high-altitude THS subgroups ( P<0.05). At 30 minutes after blood loss, D-dimer level remained significantly higher in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), while FIB was significantly lower ( P<0.05), with significant increase of APTT and PT compared to those before blood loss ( P<0.05). At 120 minutes after blood loss, the acute high-altitude THS+0 mg/kg TXA subgroup exhibited significantly higher D-dimer level compared to the other subgroups ( P<0.05), with significantly lower FIB and higher APTT and PT ( P<0.05). The acute high-altitude THS+45 mg/kg TXA subgroup also showed significantly higher D-dimer level compared to those in the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P<0.05), with significantly lower FIB and increased APTT and PT ( P<0.05). No significant differences were observed in D-dimer, FIB, APTT or PT between the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P>0.05). (4) Thromboelastography parameters: Before blood loss, the R value was significantly shorter in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), while no significant differences were observed in K value or MA value ( P>0.05). At 30 minutes after blood loss, both R value and K value were significantly shorter in the acute high-altitude THS subgroups compared to those in the plain control group ( P<0.05), with no significant differences in MA value ( P>0.05). At 120 minutes after blood loss, the acute high-altitude THS+0 mg/kg TXA subgroup exhibited significantly increased R value and K value compared to those in the other subgroups ( P<0.05), while MA value was significantly decreased ( P<0.05). The remaining acute high-altitude THS subgroups showed significant decrease of R value and K value compared to those in the plain control group ( P<0.05), while MA value was significantly lower ( P<0.05). The acute high-altitude THS+45 mg/kg TXA subgroup exhibited significantly lower R value and K value compared to those in the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P<0.05), with no significant differences in R value, K value and MA value between the later two groups ( P<0.05). (5) Changes in Syndecan-1, inflammatory factors and PAI-1: Before blood loss, syndecan-1 was significantly higher in the acute high-altitude THS subgroups compared to that in the plain control group ( P<0.05), while no significant differences were observed in IL-6, TNF-α, or PAI-1 ( P>0.05). At 30 minutes after blood loss, syndecan-1, IL-6, TNF-α, and PAI-1 were significantly higher in the acute high-altitude THS subgroups compared to those in the plain control group ( P<0.05). At 120 minutes after blood loss, syndecan-1, IL-6, TNF-α, and PAI-1 were significantly higher in the acute high-altitude THS subgroups compared to those in the plain control group ( P<0.05). Among them, the acute high-altitude THS+0 mg/kg TXA group exhibited significantly higher levels of syndecan-1, IL-6, TNF-α, and PAI-1 compared to the other acute high-altitude THS subgroups ( P<0.05). The acute high-altitude THS+45 mg/kg TXA subgroup had significantly higher syndecan-1, IL-6, and TNF-α compared to those in the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P<0.05), with no significant difference in PAI-1 ( P>0.05). No significant differences were observed in syndecan-1, IL-6, TNF-α or PAI-1 between the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA subgroups ( P>0.05). (6) Tissue injury: At 6 hours after THS, acute high-altitude THS+0 mg/kg TXA group exhibited significant interstitial thickening of the lung with extensive inflammatory cell infiltration, localized loss of intestinal brush border accompanied by cellular disruption, and marked structural disruption of renal corpuscles with focal cellular injury and necrosis. At 6 hours after THS, the acute high-altitude THS+0 mg/kg TXA subgroup exhibited significantly higher lung injury scores, Chiu′s intestinal injury scores, and kidney injury scores compared to those of the other subgroups ( P<0.05). No significant differences were observed in the tissue injury scores of the lungs, intestines and kidneys among the other subgroups ( P>0.05). The acute high-altitude THS+0 mg/kg TXA subgroup also had significantly higher lung W/D and TLW compared to those in the other subgroups ( P<0.05). At 6 hours after THS, the acute high-altitude THS+45 mg/kg TXA group exhibited significantly higher W/D and TLW of the lung tissues compared to those in the acute high-altitude THS+90 mg/kg TXA and acute high-altitude THS+135 mg/kg TXA groups ( P<0.05), with no significant differences between the latter two subgroups ( P>0.05). Conclusions:At 3 days after acute exposure to high altitude, rabbits show a hypercoagulable state of the blood, accompanied by endothelial barrier dysfunction. At 30 minutes after the induction of acute high-altitude THS, a single slow intravenous bolus injection of TXA at doses of 90 mg/kg and 135 mg/kg is more effective in improving coagulation and fibrinolysis function, inflammatory response, endothelial injury, and reduced the risk of pulmonary edema than that at a dose of 45 mg/kg.

Aim To study the renal protective effect of balanophora polysaccharide(BPS)on diabetic nephrop-athy(DN)mice and explore the related mechanisms.Methods A DN mouse model was induced using a high-fat diet combined with intraperitoneal injection of streptozotocin(STZ),which was indicated by fasting blood glucose higher than 11.1 mmol·L-1,accompa-nied by diabetic symptoms such as polydipsia,polydia-gia,polyuria and weight loss,then BPS intervention was performed.Body weight and fasting blood glucose of each group mice were detected;automatic biochemical analyzer was used to detect blood creatinine(SCr),blood urea nitrogen(BUN),24 h urinary protein(24 h UP),triglycerides(TG),total cholesterol(TC),alanine aminotransferase(ALT)content;ELISA was applied to determine serum inflammatory factor interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)level;HE and Masson staining were employed to observe renal his-topathological morphology;Western blot was used to de-tect Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),nuclear factor κB(NF-κB)for pro-tein expression.Results Compared with the model group,after BPS,body weight and fasting blood glucose decreased(P＜0.01 or P＜0.05);SCr,BUN,24 h UP,TC,TG and ALT significantly decreased(P＜0.01 or P＜0.05);the levels of the proinflammatory factors TNF-α and IL-6 were significantly reduced(P＜0.01 or P＜0.05);renal tissue injury and fibrosis decreased;TLR4,MyD88,NF-κB protein expression significantly decreased(P＜0.01 or P＜0.05).Conclusion BPS has a protective effect on the kidneys of DN mice,re-ducing the blood glucose level,improving liver and kid-ney function,alleviating renal tissue damage and renal fibrosis,and reducing inflammation response.Its mecha-nism may be related to the regulation of TLR4/MyD88/NF-κB signaling pathway.

Hepatocellular carcinoma(HCC)is difficult to detect in its early stages and current treatment methods are associated with significant side effects and a high risk of developing drug resistance.This study aims to investigate the effect of phycocyanin(PC)on the apoptosis of human HCC HepG2 cells and its potential mechanism.HepG2 cells were treated with PC at concentrations of 0.1,0.25,0.5,1,2.5,5,and 10 μg/mL for 12 h,and with 10 μg/mL PC and 2.5 μmol/L Wip1 inhibitor(Wip1i)alone or in combination for 12 and 24 h,respectively.Cell proliferation levels were assessed using the CCK-8 cell proliferation-toxicity assay kit.Apoptosis levels were measured by Annexin V-FITC/Propidium Iodide double staining combined with flow cytometry.TMT(Tandem Mass Tag)proteomics quantitative technol-ogy was applied to analyze differential protein expression.Western blotting was used to detect the expres-sion levels of Wip1,p53,and phosphorylated-p53(Ser15)proteins.The CCK-8 assay revealed that PC effectively inhibited HepG2 cell proliferation in a concentration-dependent manner,with a half-maximal inhibitory concentration(IC50)of 19.37 μg/mL.Flow cytometry results showed that PC significantly in-duced apoptosis,with an apoptosis rate of 30.40％.Quantitative proteomics analysis indicated that PC induced activation of the p53 pathway.The CCK-8 assay showed that Wip1i enhanced the cytotoxic effect of PC on HepG2 cells.Western blotting confirmed that PC inhibited Wip1 expression,induced p53 pro-tein phosphorylation,and promoted the expression of total p53 protein.Additionally,Wip1i further en-hanced PC-mediated activation of the p53 pathway,increasing the expression of p53 and pP53(S15).In conclusion,PC may induce apoptosis by inhibiting the activity of the p53 negative regulator Wip1,thereby promoting apoptosis through the Wip1/p53 pathway.

Aim To study the renal protective effect of balanophora polysaccharide(BPS)on diabetic nephrop-athy(DN)mice and explore the related mechanisms.Methods A DN mouse model was induced using a high-fat diet combined with intraperitoneal injection of streptozotocin(STZ),which was indicated by fasting blood glucose higher than 11.1 mmol·L-1,accompa-nied by diabetic symptoms such as polydipsia,polydia-gia,polyuria and weight loss,then BPS intervention was performed.Body weight and fasting blood glucose of each group mice were detected;automatic biochemical analyzer was used to detect blood creatinine(SCr),blood urea nitrogen(BUN),24 h urinary protein(24 h UP),triglycerides(TG),total cholesterol(TC),alanine aminotransferase(ALT)content;ELISA was applied to determine serum inflammatory factor interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)level;HE and Masson staining were employed to observe renal his-topathological morphology;Western blot was used to de-tect Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),nuclear factor κB(NF-κB)for pro-tein expression.Results Compared with the model group,after BPS,body weight and fasting blood glucose decreased(P＜0.01 or P＜0.05);SCr,BUN,24 h UP,TC,TG and ALT significantly decreased(P＜0.01 or P＜0.05);the levels of the proinflammatory factors TNF-α and IL-6 were significantly reduced(P＜0.01 or P＜0.05);renal tissue injury and fibrosis decreased;TLR4,MyD88,NF-κB protein expression significantly decreased(P＜0.01 or P＜0.05).Conclusion BPS has a protective effect on the kidneys of DN mice,re-ducing the blood glucose level,improving liver and kid-ney function,alleviating renal tissue damage and renal fibrosis,and reducing inflammation response.Its mecha-nism may be related to the regulation of TLR4/MyD88/NF-κB signaling pathway.

Objective:Varicocele(VC)induces male infertility by mediating changes in the testicular microenvironment,in which testicular hypoxia and high-pressure are important pathological conditions.This study aims to compare the mouse spermatogenesis(GC-2spd)cells and Sertoli(TM4)cells of mouse testis after hypoxic modeling and hypoxic and high-pressure combined modeling,and to explore the feasibility of establishing a hypoxic and high-pressure combined cell model.Methods:On the basis of cell hypoxia induced by CoCl2,the complex model of testicular cell hypoxia and high pressure was constructed by changing the osmotic pressure of GC-2 and TM4 cell medium with a high concentration of NaCl solution.After selecting the intervention concentration of CoCl2 by MTT test and detecting the expression level of HIF-1α for the determination of the optimal osmotic pressure conditions of the cell model,the cells were divided into normal group,hypoxia model group and composite model group.And the levels of OS,programmed cell death,inflammatory factors,and the expression levels of pyroptosis-related proteins were compared between the normal group and the groups with different modeling methods.Results:The optimal intervention concentration of CoCl2 in GC-2 and TM4 cells was 150 and 250μmol/L,respectively,and the expression of HIF-1α was the highest in both cells under osmotic pressure of 500 mOsmol/kg(P＜0.05).Compared with the normal group,the SOD levels of GC-2 and TM4 cells decreased(all P＜0.05),CAT level decreased(all P＜0.05),and MDA level increased(all P＜0.01),and the OS level of GC-2 and TM4 cells was more obvious than that of the hy-poxia model group(all P＜0.05).Compared with the normal group,apoptosis occurred in GC-2 and TM4 cells after composite model-ing(all P＜0.05).Compared with the normal group,the mRNA expressions of IL-1β,IL-18,TNF-α and COX-2 in GC-2 and TM4 cells significantly increased(P＜0.01)and higher than those in hypoxia model group(P＜0.05)and induced pyroptosis(P＜0.01).The expression level of GSDMD increased(P＜0.05).Conclusion:The cell model with hypoxia and high pressure com-bined modeling can not only induce oxidative stress and apoptosis of cells better than that with hypoxia alone,but also further cause in-flammatory response damage and pyroptosis,which simulates the changes of testis microenvironment mediated by hypoxia and high pressure combined conditions in VC.This cell model can be used for studying the pathogenesis of VC-associated male infertility,evalu-ating drug efficacy,and exploring pharmacological mechanisms.