Objective:To establish a microfluidic chip-based digital PCR (cdPCR) method for detecting hepatitis B virus (HBV) RNA and evaluate its application in patients with chronic HBV infection.Methods:A total of 135 patients with chronic HBV infection were recruited from the First Affiliated Hospital of Fujian Medical University and stratified into two groups based on HBV DNA levels: HBV DNA>100 IU/ml ( n=85) and HBV DNA≤100 IU/ml ( n=50). Additionally, healthy individuals and subjects infected with other viruses ( n=15) served as controls. Primers and probes targeting the HBV pre-C/C region were designed to optimize the microfluidic cdPCR method, and its linear range, limit of detection (LOD), specificity, and precision were assessed. Serum HBV RNA levels were measured using the self-developed method and two commercial kits. Pearson correlation was applied to evaluate the relationships between HBV RNA and other HBV markers. Results:The optimized microfluidic cdPCR method featured a denaturation time of 10 seconds, an annealing/extension temperature of 62 ℃, and primer and probe concentrations of 0.3 μmol/L and 0.2 μmol/L, respectively. The method demonstrated a linear range of 103-10? copies/ml and an LOD of 102 copies/ml. The intra-assay coefficient of variation ( CV) for plasmid standards at 10? and 10? copies/ml were 1.06% and 0.82%, respectively, while the inter-assay CVs were 0.75% and 0.44%. Specificity analysis confirmed the absence of positive signals in sera from healthy controls and subjects infected with other pathogens. In the HBV DNA>100 IU/ml group, the detection rate of the self-developed cdPCR method was 81.18% (69/85), significantly higher than the 64.71% (55/85) achieved by commercial kit B ( P<0.016 7). However, in the HBV DNA≤100 IU/ml group, no significant differences were observed among the three methods ( P>0.05). HBV RNA levels were positively correlated with HBV DNA ( r=0.67), hepatitis B surface antigen ( r=0.53), and hepatitis B e antigen ( r=0.44) (all P<0.001). Conclusion:A microfluidic cdPCR assay for the quantitative detection of HBV RNA has been successfully developed. This assay demonstrates high sensitivity, specificity, and robust detection capability, even for low HBV DNA-concentration samples.

Objective:To investigate the role of TcpC, a virulence factor of uropathogenic Escherichia coli (UPEC), in inhibiting the formation of neutrophil extracellular traps (NETs) in mouse bone marrow neutrophils, and to analyze its pathogenic mechanism. Methods:C57BL/6 mice were injected with either wild-type (CFT073 wt) or tcpc gene-knockout UPEC CFT073(CFT073 Δ tcpc) to establish a mouse model of cystitis. Mice were sacrificed 3 d post-infection, and their bladders were collected to observe gross pathological changes. Hematoxylin-eosin (HE) staining was used to assess histopathological changes in bladder tissues, and immunohistochemistry was performed to localize TcpC in bladder tissues. Bacterial loads in urine samples were quantified using the ten-fold dilution method, and the presence of tcpc gene in genomic DNA from bladder or urine samples was confirmed by PCR. The expression of TcpC at mRNA and protein levels in mouse bone marrow nuetrophils infected with CFT073 wt was detected by qRT-PCR and Western blot. The effects of UPEC infection on expression of NETs-related proteins and the production of pro-inflammatory factors in mouse bone marrow neutrophils were analyzed by Western blot and ELISA, respectively. Reactive oxygen species(ROS) level and bacterial viability in mouse bone marrow nuetrophils were measured using ROS and bacterial viability detection kits. Results:Compared to the CFT073 Δ tcpc group, the bladder of CFT073 wt group mice exhibited significant enlargement, extensive inflammatory cell infiltration, and the presence of TcpC in bladder tissue. The bacterial load in the urine of CFT073 wt -infected mice was significantly higher than that in the CFT073 Δ tcpc group ( P<0.01). PCR confirmed the presence of the tcpc gene in bladder and urine samples from CFT073 wt-infected mice. Increased expression of TcpC at both mRNA and protein levels was observed in CFT073 wt-infected mouse bone marrow neutrophils. CFT073 wt infection inhibited the mRNA and protein expression of NETs-related proteins and reduced the production of pro-inflammatory factors in mouse bone marrow neutrophils. TcpC suppressed ROS level and promoted the survival of CFT073 wt in mouse bone marrow neutrophils. Conclusions:TcpC enhances the pathogenicity of UPEC CFT073 by inhibiting the formation and activation of NETs in mouse bone marrow neutrophils. This study provides new insights into the pathogenic mechanisms of UPEC and the immune evasion strategies of other pathogenic bacteria, as well as potential targets for clinical prevention and treatment of UPEC-induced urinary tract infections.

The epidemiological and spatiotemporal clustering characteristics of dengue fever in Fujian Province were ana-lyzed,to provide a scientific basis for dengue fever prevention and control.Descriptive epidemiology,spatial autocorrelation a-nalysis,and spatiotemporal scanning were used to analyze dengue fever cases in Fujian Province from 2011 to 2023.In this peri-od,a total of 3 586 cases of dengue fever were reported in Fujian Province,including 2 360 local cases,1 134 imported cases from abroad,and 92 imported cases from China.Cases were reported in ten prefectures and cities of the province,and 81 out of 88 counties reported cases.Imported cases were reported throughout the year in Fujian Province,but the occurrence of local ca-ses showed clear seasonality.Local cases and domestic imports were concentrated in August to October,whereas overseas im-ports occurred primarily from June to October.The imported cases were mainly from Southeast Asian countries,but a trend of spreading from Southeast Asian countries to South Asia,Africa,the Americas,and other regions,was observed.Spatio-tem-poral clustering of dengue fever was found in Fujian Province(Moran's I value 0.14-0.66,P＜0.05),and the high-high ag-gregation areas were distributed primarily in Fuzhou,Quanzhou,and Putian.Spatio-temporal scanning detected three aggrega-tion areas:one main and two secondary.The aggregation time was from the end of July to October,and the distribution was primarily in Fuzhou,Quanzhou,Putian,Zhangzhou,and Xiamen.The distribution of dengue fever in Fujian Province showed clear spatial and temporal clustering from the end of July to October,and the distribution was primarily in Fuzhou,Quanzhou,Putian,Zhangzhou,and Xiamen.For high concentration areas,national health campaigns,mosquito prevention and control,epidemic surveillance,medical personnel training,and other relevant measures could be carried out in advance before local cases appear every year.Reduce local transmission of dengue fever due to importation.

Objective To study the expression characteristics of osteoprotegerin(OPG)/receptor activator of nuclear factor-κB ligand(RANKL)/receptor activator of nuclear factor-κB(RANK)system and the relationship with fibrosis in myocardial tissues of rats with chronic heart failure.Methods SD rats were randomly divided into sham-operation group(12 rats)and model group.In sham-operation group,surgical thread was passed through the abdominal aorta without constricting it after laparotomy;in model group,establish the heart failure model by abdominal aorta coarctation.The successful model rats were randomly divided into model 1 week(12 rats),model 2 weeks(11 rats),model 4 weeks(11 rats),model 8 weeks(11 rats)and model 12 weeks groups(11 rats).The end point of the study is at week 12.The contents of hydroxyproline(HYP),total myocardial collagen and collagen volume fraction(CVF)were compaired in all proups.The expression levels of OPG,RANKL and RANK proteins in cardiomyocytes were determined by Western blot.Results The contents of HYP in sham-operation,model 1 week,model 2 weeks,model 4 weeks,model 8 weeks and model 12 weeks group were(0.25±0.04),(0.37±0.05),(0.45±0.04),(0.60±0.05),(0.82±0.10)and(1.03±0.07)μg·mg-1;the total myocardial collagen contents were(1.87±0.31),(2.73±0.38),(3.36±0.31),(4.47±0.37),(6.08±0.74)and(7.67±0.49)μg·mg-1;the CVF were(1.95±0.23)％,(2.40±0.25)％,(3.65±0.25)％,(5.43±0.29)％,(6.97±0.36)％and(9.38±0.49)％;the relative expression levels of OPG protein were 0.64±0.07,0.80±0.07,1.02±0.07,1.32±0.11,2.13±0.12 and 2.84±0.16;the relative expression levels of RANKL protein were 0.71±0.08,1.06±0.07,1.53±0.07,2.62±0.12,4.46±0.14 and 6.11±0.16;the relative expression levels of RANK protein were 0.30±0.05,0.45±0.05,0.63±0.06,0.98±0.07,1.43±0.10 and 1.63±0.10.With the extention of time,the above indexs of all model groups were significantly higher than those in the sham-operation group(all P＜0.05).There were positive linear correlation between the relative expression levels of OPG,RANKL,RANK protein and the levels of CVF and total contents in cardiomyocytes of rats with chronic heart failure(allP＜0.01).Conclusions In the process of chronic heart failure,the expression of OPG/RANKL/RANK axis is obviously enhanced,in which the up-regulation of RANKL level is most obvious.The expression level of OPG/RANKL/RANK is positively correlated with CVF and total myocardial collagen content.

Objective To describe the clinical features of patients with primary sclerosing cholangitis(PSC)in China based on a nationwide multicenter patient cohort,and to investigate the risk factors for prognosis.Methods A retrospective cohort study was conducted among the patients with a confirmed diagnosis of PSC based on the electronic medical record system of seven grade A tertiary hospitals across the country,and related data were extracted.The Mann-Whitney U test was used for comparison of continuous data between groups,and the chi-square test was used for comparison of categorical data between groups.The Kaplan-Meier method was used to estimate liver transplant-free survival,and the log-rank test was used for comparison of survival rate between PSC patients with different features.The Cox regression model was used to identify independent risk factors for the prognosis of PSC patients and the interactions between key factors.Results A total of 107 patients were enrolled,among whom 55.6%(55/99)had large-duct PSC and 29.0%(31/107)had comorbidity with inflammatory bowel disease(IBD).The positivity rate of anti-neutrophil cytoplasmic antibody(ANCA)was 32.9%(24/73),and 50.0%(40/80)of the patients had an increase in IgG/IgM.The median symptom-to-diagnosis interval was 1 year(<1-4.0),and 38.3%(41/107)of the patients had progressed to decompensated cirrhosis at the time of diagnosis.The median liver transplant-free survival time was 114 months(95%confidence interval[CI]:62-166),with a 5-year survival rate of 65.7%.The multivariate analysis showed that an increase in total bile acid(TBA)(hazard ratio[HR]=1.006,95%CI:1.002-1.010,P=0.001)and a prolonged symptom-to-diagnosis interval(HR=1.252,95%CI:1.059-1.480,P=0.009)were independent risk factors for prognosis.The interaction analysis showed that compared with the female patients with TBA<50 μmol/L,both male and female patients with TBA≥50 μmol/L had a significant increase in the risk of liver transplantation or death(male:HR=16.563,95%CI:2.103-130.449,P<0.001;female:HR=17.009,95%CI:2.113-136.934,P<0.001),and compared with the patients with an age of<45 years and a TBA level of<50 μmol/L,the patients with an age of≥45 years and a TBA level of≥50 μmol/L had a significant increase in the risk of liver transplantation or death(HR=10.729,95%CI:1.325-86.859,P=0.026).Compared with the female patients with an symptom-to-diagnosis interval of≤2 years,the male patients with a symptom-to-diagnosis interval of>2 years had an increased risk of liver transplantation or death(HR=4.825,95%CI:1.725-13.644,P=0.003),and compared with the patients with an age of<45 years and a symptom-to-diagnosis interval of≤2 years,the patients with an age of<45 years and a symptom-to-diagnosis interval of>2 years had an increased risk of liver transplantation or death(HR=4.983,95%CI:1.366-18.173,P=0.015).Conclusion Compared with the reports from Western countries,large-duct PSC is also the main type of PSC in China,but with a relatively low proportion,and there is also a relatively low proportion of patients with IBD or positive ANCA.An increase in TBA and a prolonged symptom-to-diagnosis interval are independent risk factors for prognosis,with significant interactions with age and sex.This suggests that early screening and intervention should be enhanced to improve prognosis.

Objective To establish a laser diffraction method for determining globule size distribution in medium and long chain fat emulsion injection(C8～24Ve).Methods The Mastersizer 3000 laser particle size analyzer and Hydro EV wet sampling device were used,with water as the dispersion medium.Critical parameters,including refractive index,absorption coefficient,obscuration,measurement duration,stirring speed,and sonication parameters,were systematically optimized.Comparative evaluations were conducted across different computational models(Mie vs.Fraunhofer theories)and instrument platforms.Results The optimized optical parameters for particle size distribution analysis of medium and long chain fat emulsion injection(C8～24Ve)were determined as follows:absorption rate 0.001,refractive index 1.52,measurement duration 10 s,stirring speed 1 200 r·min-1,with no sonication applied.Under these conditions,the method demonstrated satisfactory precision,repeatability,and robustness.Conclusions This method demonstrates operational simplicity,high accuracy,and excellent reproducibility,making it suitable for particle size distribution analysis of medium and long chain fat emulsion injection(C8～24Ve).The established protocol provides a valuable reference for the development of particle size determination methods for emulsion formulations.

Objective To establish a laser diffraction method for determining globule size distribution in medium and long chain fat emulsion injection(C8～24Ve).Methods The Mastersizer 3000 laser particle size analyzer and Hydro EV wet sampling device were used,with water as the dispersion medium.Critical parameters,including refractive index,absorption coefficient,obscuration,measurement duration,stirring speed,and sonication parameters,were systematically optimized.Comparative evaluations were conducted across different computational models(Mie vs.Fraunhofer theories)and instrument platforms.Results The optimized optical parameters for particle size distribution analysis of medium and long chain fat emulsion injection(C8～24Ve)were determined as follows:absorption rate 0.001,refractive index 1.52,measurement duration 10 s,stirring speed 1 200 r·min-1,with no sonication applied.Under these conditions,the method demonstrated satisfactory precision,repeatability,and robustness.Conclusions This method demonstrates operational simplicity,high accuracy,and excellent reproducibility,making it suitable for particle size distribution analysis of medium and long chain fat emulsion injection(C8～24Ve).The established protocol provides a valuable reference for the development of particle size determination methods for emulsion formulations.

Purpose To analyze the impact of early renal complication on brain function in diabetes using the amplitude of low frequency fluctuations(ALFF)and functional connectivity(FC).Materials and Methods A total of 39 early diabetic kidney disease patients and 49 diabetic patients without kidney disease were included at the First Affiliated Hospital of Army Medical University from September 2023 to June 2024.Cognitive assessments were conducted using the Montreal cognitive assessment(MoCA)and mini-mental state examination(MMSE).All subjects underwent resting-state functional magnetic resonance imaging.Differences in brain function between the two groups were analyzed by ALFF and FC.In the early diabetic kidney disease group,the correlation and stepwise multiple linear regression analysis were performed between imaging difference indicators and clinical variables.Results In the early diabetic kidney disease patients,there was a decrease in ALFF values in the left calcarine(P<0.05,corrected for FWE),but an increase in FC with the left putamen(P<0.05,corrected for FWE).The ALFF values showed a negative correlation with urinary albumin-to-creatinine ratio and total cholesterol(r=-0.595,-0.351,both P<0.05)and a positive correlation with MoCA and MMSE scores(r=0.596,0.591,both P<0.001).The FC values were positively correlated with urinary albumin-to-creatinine ratio(r=0.552,P<0.001),while negatively correlated with MoCA and MMSE scores(r=-0.497,P=0.011;r=-0.529,P<0.001).Conclusion Early renal complications can affect changes in brain function and cognitive abilities in diabetic patients;alterations in the ALFF of the left calcarine and its FC with left putamen may serve as imaging biomarkers for monitoring cognitive impairment and brain injury in early diabetic kidney disease.

Objective To describe the clinical features of patients with primary sclerosing cholangitis(PSC)in China based on a nationwide multicenter patient cohort,and to investigate the risk factors for prognosis.Methods A retrospective cohort study was conducted among the patients with a confirmed diagnosis of PSC based on the electronic medical record system of seven grade A tertiary hospitals across the country,and related data were extracted.The Mann-Whitney U test was used for comparison of continuous data between groups,and the chi-square test was used for comparison of categorical data between groups.The Kaplan-Meier method was used to estimate liver transplant-free survival,and the log-rank test was used for comparison of survival rate between PSC patients with different features.The Cox regression model was used to identify independent risk factors for the prognosis of PSC patients and the interactions between key factors.Results A total of 107 patients were enrolled,among whom 55.6%(55/99)had large-duct PSC and 29.0%(31/107)had comorbidity with inflammatory bowel disease(IBD).The positivity rate of anti-neutrophil cytoplasmic antibody(ANCA)was 32.9%(24/73),and 50.0%(40/80)of the patients had an increase in IgG/IgM.The median symptom-to-diagnosis interval was 1 year(<1-4.0),and 38.3%(41/107)of the patients had progressed to decompensated cirrhosis at the time of diagnosis.The median liver transplant-free survival time was 114 months(95%confidence interval[CI]:62-166),with a 5-year survival rate of 65.7%.The multivariate analysis showed that an increase in total bile acid(TBA)(hazard ratio[HR]=1.006,95%CI:1.002-1.010,P=0.001)and a prolonged symptom-to-diagnosis interval(HR=1.252,95%CI:1.059-1.480,P=0.009)were independent risk factors for prognosis.The interaction analysis showed that compared with the female patients with TBA<50 μmol/L,both male and female patients with TBA≥50 μmol/L had a significant increase in the risk of liver transplantation or death(male:HR=16.563,95%CI:2.103-130.449,P<0.001;female:HR=17.009,95%CI:2.113-136.934,P<0.001),and compared with the patients with an age of<45 years and a TBA level of<50 μmol/L,the patients with an age of≥45 years and a TBA level of≥50 μmol/L had a significant increase in the risk of liver transplantation or death(HR=10.729,95%CI:1.325-86.859,P=0.026).Compared with the female patients with an symptom-to-diagnosis interval of≤2 years,the male patients with a symptom-to-diagnosis interval of>2 years had an increased risk of liver transplantation or death(HR=4.825,95%CI:1.725-13.644,P=0.003),and compared with the patients with an age of<45 years and a symptom-to-diagnosis interval of≤2 years,the patients with an age of<45 years and a symptom-to-diagnosis interval of>2 years had an increased risk of liver transplantation or death(HR=4.983,95%CI:1.366-18.173,P=0.015).Conclusion Compared with the reports from Western countries,large-duct PSC is also the main type of PSC in China,but with a relatively low proportion,and there is also a relatively low proportion of patients with IBD or positive ANCA.An increase in TBA and a prolonged symptom-to-diagnosis interval are independent risk factors for prognosis,with significant interactions with age and sex.This suggests that early screening and intervention should be enhanced to improve prognosis.

Objective : To explore the causal relationship between 46 phenotypes ( including 15 seropositive case- control phenotypes and 31 quantitative antibody-measurement phenotypes) and ulcerative colitis( UC) using two- sample bidirectional Mendelian randomization( TSMR) . Methods: Single nucleotide polymorphisms ( SNPs) sig- nificantly associated with the relative abundance of the 46 antibody sera were extracted as instrumental variables ac- cording to preset thresholds . Summary statistics for UC were obtained from the OPEN GWAS database ( n = 47 745) . MR-Egger regression , weighted median method ( WME) , inverse variance weighting ( IVW) , the simple mode method (SM) , and weighted multitude method (WM) were used to estimate the causal relationship between antibody levels and UC , primarily using the IVW method . The results were assessed according to the effect indica- tor dominance ratios (OR) and 95% confidence intervals (CI) . Sensitivity analysis , heterogeneity test , gene plei- otropy test , and outlier test (MR-PRESSO) were combined to verify the stability and reliability of the results , and the causal association study was performed again using reverse Mendelian randomization(MR) . Results : IVW re- sults showed that Epstein-Barr( EB) virus EA-D antibody levels ( OR = 0. 806 , 95% CI = 0. 693 - 0. 939 , P < 0. 01) , Epstein-Barr virus EBNA-1 antibody levels ( OR = 1 . 870% , 95% CI = 1 . 480 - 2. 360 , P < 0. 000 1) , Anti-polyomavirus 2 IgG seropositivity (OR = 0. 570 , 95% CI = 0. 435 - 0. 746 , P < 0. 000 1) were associated with UC . The inverse MR analysis revealed a causal effect on anti-polyomavirus 2 IgG seropositivity , and none of the a- bove revealed genetic pleiotropy or significant heterogeneity of IVs . Conclusion EB virus EBNA-1 antibody levels are positively associated with the risk of UC , while EB virus EA-D antibody levels and anti-polyomavirus 2 IgG se- ropositivity are negatively associated with the risk of UC , indicating that they are protective factors for UC .