ObjectiveTo compare the histological staining performance of four various tissue fixatives, including a self-developed fixative, for preparing paraffin sections of rat eyeball tissue. MethodsTwenty 5-week-old male Sprague-Dawley (SD) rats were randomly divided into 4 groups (n=5 per group). After anesthesia by intraperitoneal injection of Zoletil™ 50 at a dose of 45 mg/kg body weight, the rats were euthanized by exsanguination via the abdominal aorta. Bilateral eyeballs were carefully extracted intact and fixed for 72 h in 10% formaldehyde fixative, glutaraldehyde-formaldehyde mixed fixative, Davidson's fixative, and self-developed fixative, respectively. After fixation, the eyeballs were longitudinally sectioned along the optic nerve, with the portions containing the optic nerve preserved. The tissues were then dehydrated, embedded, and sectioned. Following hematoxylin and eosin (HE) staining, histological staining quality was compared among ocular structures including the cornea, lens, and retina. ResultsThe overall appearance of rat eyeballs showed marked shrinkage in the 10% formaldehyde group and the glutaraldehyde-formaldehyde group, whereas the eye morphology remained round and intact in the modified Davidson's fixative group and self-developed fixative group. The corneal stroma exhibited obvious rupture, and the cells were arranged in folded arrangement in the modified Davidson's fixative group, 10% formaldehyde group, and self-developed fixative group. In contrast, the corneal cells in the glutaraldehyde-formaldehyde group were neatly arranged, showing no rupture or folding, and exhibited clear staining, indicating the highest quality of corneal sectioning among all groups. In the 10% formaldehyde group, cracks were observed in the equatorial and cortical regions of the lens, but the lens fiber structure remained intact. The lenses in the modified Davidson's fixative group showed extensive rupture and detachment. The glutaraldehyde-formaldehyde group displayed only slight cracks at the equator. In the self-developed fixative group, mild red folding was limited to the peripheral lens region, with the remaining structures intact and unbroken. These findings indicated that the glutaraldehyde-formaldehyde and self-developed fixative groups achieved the best lens sectioning quality. The retina was severely detached from the choroid/sclera layer, with extensive rupture of each cellular layer in the 10% formaldehyde fixative group. In the glutaraldehyde-formaldehyde group, partial detachment between the retina and choroid/sclera was observed. The outer plexiform layer and nerve fiber layer showed separation with edema, while the cells in all layers were neatly arranged. In both the Davidson's fixative and the self-developed fixative groups, the retina remained intact without rupture, and no structural separation was observed in any layer. Both demonstrated advantages in preserving the integrity and orderly arrangement of all retinal layers; however, the self-developed fixative group exhibited greater contrast. ConclusionThe choice of fixative significantly affects the morphological preservation of various structures in the rat eye. The self-developed fixative demonstrates the best overall performance in maintaining overall eye morphology, the structural integrity of the lens, and retinal adhesion. For studies focusing solely on the corneal structure, the glutaraldehyde-formaldehyde mixed fixative is recommended. The 10% formaldehyde fixative exhibits unsatisfactory fixation effects for all the aforementioned ocular structures and is not recommended for detailed morphological studies of eyeball tissues.

Objective To evaluate the safety and efficacy of simultaneous resection for initially resectable rectal cancer with synchronous liver metastases. Methods A retrospective analysis was conducted on 305 patients with initially resectable rectal cancer with synchronous liver metastases. These patients were diagnosed at Zhongshan Hospital, Fudan University from January 2016 to June 2020. Among them, 191 underwent simultaneous rectum and liver resection and 114 underwent staged resection. Propensity score matching (PSM) was performed at a 1∶1 ratio. Clinical data were compared and Kaplan-Meier survival curves were plotted. Results After PSM, 85 patients were included in each group. General data showed no significant differences. Except for liver metastasis resection method, no statistical differences were found in primary tumor surgery approach, intraoperative blood loss, intraoperative complications, time to first flatus and defecation, 30-day mortality, and postoperative hospital stay between the simultaneous resection group and the staged resection group. The overall complication rate was higher in the simultaneous resection group (48.2% vs 29.4%, P＝0.04). Specifically, the grade Ⅱ complications were significantly higher (29.4% vs 14.1%, P＝0.016), but there’s no differences in severe complications (grade Ⅲ-Ⅴ). No statistically differences were observed in median progression-free survival (HR＝0.70, 95%CI 0.50-0.97, P＝0.103) and 5-year overall survival (HR＝0.95, 95%CI 0.63-1.44, P＝0.259). Conclusions Simultaneous resection demonstrates comparable safety and efficacy to staged resection for initially resectable rectal cancer with synchronous liver metastases.

Objective To investigate the prognostic value of remnant lipoprotein cholesterol(RLP-C)and platelet-lymphocyte ratio(PLR)for elderly patients with acute myocardial infarction(AMI)stratified by body mass index(BMI).Methods A total of 130 elderly AMI patients admitted to our hospital from January 2022 to January 2024 were recruited,and then based on their BMI,they were categorized into low(n=18),normal(n=60),and high BMI groups(n=52).Multivariate logistic regression analysis was used to identify the risk factors for poor prognosis in elderly AMI patients.Spearman correlation analysis was conducted to examine the relationship of RLP-C and PLR with prognostic outcomes.ROC curve analysis was performed to evaluate the predictive per-formance of the two indicators for prognosis in elderly AMI patients of different BMI.Results Significant differences in RLP-C and PLR levels were observed among the above three groups(P＜0.05,P＜0.01).In the overall population,as well as in the low,normal and high BMI groups,both RLP-C and PLR were positively correlated with adverse outcomes in elderly AMI patients(P＜0.01).PLR and RLP-C were independent risk factors for poor prognosis in elderly AMI pa-tients(P＜0.01),and they were independently associated with prognosis in each BMI group(P＜0.01).The combined the three indicators together significantly improved the sensitivity and speci-ficity in predicting adverse outcomes in elderly AMI patients when compared to single indicator alone(sensitivity:54.29％,75.94％,64.18％vs 82.61％;specificity:68.83％,70.22％,72.15％vs 80.49％;P＜0.01).Conclusion RLP-C and PLR are influencing factors for poor prognosis in eld-erly AMI patients,and can serve as predictive factors for prognosis,which are helpful for provi-ding valuable guidance for early treatment and prevention of adverse outcomes.

Objective To develop a tool for assessing the needs for care services for adults with disabilities in welfare institutions,and formulate criteria for classifying care needs.Methods Based on the theory of the International Classification of Functioning,Disability and Health,the core contents of domestic and international care service assessment standards were analysed,and the first draft of the assess-ment tool was formed integrating with the current practice of welfare institutions.A total of eleven experts in the field of care services validated the first draft,and the assessment tool was revised according to the experts'opin-ions.From May to June,2024,340 adults with disabilities from Second Social Welfare Institute of Shanghai were graded to analyze the reliability and validity of the assessment tool.And then,1 046 subjects from Second Social Welfare Institute of Shanghai were graded using the assessment tool,and the scoring intervals for the five levels of care were determined using cluster analysis.Results The assessment tool included four first-level dimensions,eleven second-level dimensions,39 third-level dimen-sions and 43 indicators.The coefficients of variation of each dimension and indicator were less than 0.25 for ex-pert validation,while the average score of the importance of each indicator was above 4.63,with a full score rate more than 0.86.The overall Cronbach's α coefficient was 0.956,the test-retest reliability was 0.997,and the con-tent validity index of each indicator was 0.83 to 1.00.The exploratory factor analysis extracted four common fac-tors,with a cumulative variance contribution of 86.679%;the loading coefficients of indicators were 0.675 to 0.878,and the correlation coefficients among the dimensions were 0.325 to 0.476.Cluster analysis determined the level of institutional care needs as level 1(severe),level 2(medium-heavy),level 3(medium),level 4(light-moderate),and level 5(light).Conclusion The needs assessment tool for care services for adults with disabilities in welfare institutions developed in this study is good in reliability and validity,and can be used in guiding the care services provided for people with disabilities more efficient and targeted.

Objective To understand the changing composition and antibiotic resistance of bacterial species in the clinical isolates from outpatient and emergency department(hereinafter referred to as outpatients)and inpatient children over time in various hospitals,and to provide laboratory evidence for rational antibiotic use.Methods The data on clinically isolated pathogenic bacteria and antimicrobial susceptibility of isolates from outpatients and inpatient children in the CHINET program from 2015 to 2021 were collected and analyzed.Results A total of 278 471 isolates were isolated from pediatric patients in the CHINET program from 2015 to 2021.About 17.1％of the strains were isolated from outpatients,primarily group A β-hemolytic Streptococcus,Escherichia coli,and Staphylococcus aureus.Most of the strains(82.9％)were isolated from inpatients,mainly SS.aureus,E.coli,and H.influenzae.The prevalence of methicillin-resistant S.aureus(MRSA)in outpatients(24.5％)was lower than that in inpatient children(31.5％).The MRSA isolates from outpatients showed lower resistance rates to the antibiotics tested than the strains isolated from inpatient children.The prevalence of vancomycin-resistant Enterococcus faecalis or E.faecium and penicillin-resistant S.pneumoniae was low in either outpatients or inpatient children.S.pneumoniae,β-hemolytic Streptococcus and S.viridans showed high resistance rates to erythromycin.The prevalence of erythromycin-resistant group A β-hemolytic Streptococcus was higher in outpatients than that in inpatient children.The prevalence of β-lactamase-producing H.influenzae showed an overall upward trend in children,but lower in outpatients(45.1％)than in inpatient children(59.4％).The prevalence of carbapenem-resistant Klebsiella pneumoniae(CRKpn),carbapenem-resistant Pseudomonas aeruginosa(CRPae)and carbapenem-resistant Acinetobacter baumannii(CRAba)was 14％,11.7％,47.8％in outpatients,but 24.2％,20.6％,and 52.8％in inpatient children,respectively.The prevalence of multidrug-resistant E.coli,K.pneumoniae,Proteus mirabilis,P.aeruginosa and A.baumannii strains was lower in outpatients than in inpatient children.The prevalence of fluoroquinolone-resistant E.coli,ESBLs-producing K.pneumoniae,ESBLs-producing P.mirabilis,carbapenem-resistant E.coli(CREco),CRKpn,and CRPae was lower in children in outpatients than in inpatient children,but the prevalence of CRAba in 2021 was higher than in inpatient children.Conclusions The distribution of clinical isolates from children is different between outpatients and inpatients.The prevalence of MRSA,ESBL,and CRO was higher in inpatient children than in outpatients.Antibiotics should be used rationally in clinical practice based on etiological diagnosis and antimicrobial susceptibility test results.Ongoing antimicrobial resistance surveillance and prevention and control of hospital infections are crucial to curbing bacterial resistance.

Objective To investigate the changing antibiotic resistance profiles of E.coli isolated from patients in the 52 hospitals participating in the CHINET program from 2015 to 2021.Methods Antimicrobial susceptibility was tested for clinical isolates of E.coli according to the unified protocol of CHINET program.WHONET 5.6 and SPSS 20.0 software were used for data analysis.Results Atotal of 289 760 nonduplicate clinical strains ofE.coli were isolated from 2015 to 2021,mainly from urine samples(44.7±3.2)％.The proportion of E.coli strains isolated from urine samples was higher in females than in males(59.0％vs 29.5％).The proportion of E.coli strains isolated from respiratory tract and cerebrospinal fluid samples was significantly higher in children than in adults(16.7％vs 7.8％,0.8％vs 0.1％,both P＜0.05).The isolates from internal medicine department accounted for the largest proportion(28.9±2.8)％with an increasing trend over years.Overall,the prevalence of ESBLs-producing E.coli and carbapenem resistant E.coli(CREco)was 55.9％and 1.8％,respectively during the 7-year period.The prevalence of ESBLs-producing E.coli was the highest in tertiary hospitals each year from 2015 to 2021 compared to secondary hospitals.The prevalence of CREco was higher in children's hospitals compared to secondary and tertiary hospitals each year from 2015 to 2021.The prevalence of ESBLs-producing E.coli in tertiary hospitals and children's hospitals and the prevalence of CREco in children's hospitals showed a decreasing trend over the 7-year period.The prevalence of CREco in secondary and tertiary hospitals increased slowly.Antibiotic resistance rates changed slowly from 2015 to 2021.Carbapenem drugs(imipenem,meropenem)were the most active drugs amongβ-lactams against E.coli(resistance rate≤2.1％).The resistance rates of E.coli to β-lactam/β-lactam inhibitor combinations(piperacillin-tazobactam,cefoperazone-sulbactam),aminoglycosides(amikacin),nitrofurantoin and fosfomycin(for urinary isolates only)were all less than 10％.The resistance rate of E.coli strains to antibiotics varied with the level of hospitals and the departments where the strains were isolated,especially for cefazolin and ciprofloxacin,to which the resistance rate of E.coli strains from children in non-ICU departments was significantly lower than that of the strains isolated from other departments(P＜0.05).The E.coli isolates from ICU showed higher resistance rate to most antimicrobial agents tested(excluding tigecycline)than the strains isolated from other departments.The E.coli strains isolated from tertiary hospitals showed higher resistance rates to the antimicrobial agents tested(excluding tigecycline,polymyxin B,cefepime and carbapenems)than the strains from secondary hospitals and children's hospitals.Conclusions E.coli is an important pathogen causing clinical infection.More than half of the clinical isolates produced ESBL.The prevalence of CREco is increasing in secondary and tertiary hospitals over the 7-year period even though the overall prevalence is still low.This is an issue of concern.

Objective To establish a rapid immunological detection method for MPT64 protein based on red microspheres and select highly-sensitive and highly-specific antibody pairs.Methods A His-tagged prokaryotic expression vector was constructed for expression of MPT64 protein that was used to immunize New Zealand white rabbits after purification and validation.Peripheral blood mononuclear cells(PBMCs)were isolated from the rabbits,and antigen-specific B cells expressing antibodies were sorted using single B-cell flow cytometry.mRNA in B cells was reverse-transcribed into cDNA,and paired antibody heavy-and light-chain sequences were amplified via nested PCR.Expression vectors were constructed,and recombinant antibodies were produced in Expi293F cells.Fluorescent immunochromatography was employed to screen for matched antibody pairs.The selected antibodies were used to establish a rapid detection method based on red microsphere immunochromatography.Results Ten high-affinity monoclonal antibodies against MPT64 were generated.Two antibody pairs were selected for MPT64 immunodetection that reached a sensitivity of 0.0125 ng/mL.Conclusion High-affinity rabbit monoclonal antibodies against MPT64 are obtained via single B-cell technology,and a rapid red microspheres-based immunodetection method is established,enabling highly sensitive detection of Mycobacterium tuberculosis MPT64 protein.

Objective To summarize the changing prevalence of carbapenem resistance in Enterobacterales based on the data of CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021 for improving antimicrobial treatment in clinical practice.Methods Antimicrobial susceptibility testing was performed using a commercial automated susceptibility testing system according to the unified CHINET protocol.The results were interpreted according to the breakpoints of the Clinical & Laboratory Standards Institute(CLSI)M100 31st ed in 2021.Results Over the seven-year period(2015-2021),the overall prevalence of carbapenem-resistant Enterobacterales(CRE)was 9.43％(62 342/661 235).The prevalence of CRE strains in Klebsiella pneumoniae,Citrobacter freundii,and Enterobacter cloacae was 22.38％,9.73％,and 8.47％,respectively.The prevalence of CRE strains in Escherichia coli was 1.99％.A few CRE strains were also identified in Salmonella and Shigella.The CRE strains were mainly isolated from respiratory specimens(44.23±2.80)％,followed by blood(20.88±3.40)％and urine(18.40±3.45)％.Intensive care units(ICUs)were the major source of the CRE strains(27.43±5.20)％.CRE strains were resistant to all the β-lactam antibiotics tested and most non-β-lactam antimicrobial agents.The CRE strains were relatively susceptible to tigecycline and polymyxins with low resistance rates.Conclusions The prevalence of CRE strains was increasing from 2015 to 2021.CRE strains were highly resistant to most of the antibacterial drugs used in clinical practice.Clinicians should prescribe antimicrobial agents rationally.Hospitals should strengthen antibiotic stewardship in key clinical settings such as ICUs,and take effective infection control measures to curb CRE outbreak and epidemic in hospitals.

Objective To characterize the changing species distribution and antibiotic resistance profiles of respiratory isolates in hospitals participating in the CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021.Methods Commercial automated antimicrobial susceptibility testing systems and disk diffusion method were used to test the susceptibility of respiratory bacterial isolates to antimicrobial agents following the standardized technical protocol established by the CHINET program.Results A total of 589 746 respiratory isolates were collected from 2015 to 2021.Overall,82.6％of the isolates were Gram-negative bacteria and 17.4％were Gram-positive bacteria.The bacterial isolates from outpatients and inpatients accounted for(6.0±0.9)％and(94.0±0.1)％,respectively.The top microorganisms were Klebsiella spp.,Acinetobacter spp.,Pseudomonas aeruginosa,Staphylococcus aureus,Haemophilus spp.,Stenotrophomonas maltophilia,Escherichia coli,and Streptococcus pneumoniae.Each microorganism was isolated from significantly more males than from females(P＜0.05).The overall prevalence of methicillin-resistant S.aureus(MRSA)was 39.9％.The prevalence of penicillin-resistant S.pneumoniae was 1.4％.The prevalence of extended-spectrum β-lactamase(ESBL)-producing E.coli and K.pneumoniae was 67.8％and 41.3％,respectively.The overall prevalence of carbapenem-resistant E.coli,K.pneumoniae,Enterobacter cloacae,Pseudomonas aeruginosa,and Acinetobacter baumannii was 3.7％,20.8％,9.4％,29.8％,and 73.3％,respectively.The prevalence of β-lactamase was 96.1％in Moraxella catarrhalis and 60.0％in Haemophilus influenzae.The H.influenzae isolates from children(＜18 years)showed significantly higher resistance rates to β-lactam antibiotics than the isolates from adults(P＜0.05).Conclusions Gram-negative bacteria are still predominant in respiratory isolates associated with serious antibiotic resistance.Antimicrobial resistance surveillance should be strengthened in clinical practice to support accurate etiological diagnosis and appropriate antimicrobial therapy based on antimicrobial susceptibility testing results.

Objective To investigate the role and mechanism of AMP-activated protein kinase(AMPK)in regulating mitochondrial function and cardiomyocyte injury in sepsis.Methods Forty Balb/c mice were randomly divided into four groups:Sham group(n=10),Sham+5-aminoimidazole-4-carboxamide ribonucleotide(AICAR)group(n=10),CLP group(n=10),and CLP+AICAR group(n=10).A sepsis mouse model was established through cecal ligation and puncture(CLP).Echocardiography and histological analysis were used to assess sepsis-induced cardiac injury.Neonatal rat cardiomyocytes(NRCMs)were incubated with lipopolysaccharide(LPS)(10 μg/mL)for 24 h to induce an in vitro sepsis model,and treated with AICAR.Mitochondrial function and dynamics were assessed by using Western blotting,enzyme-linked immunosorbent assay(ELISA),and immunofluorescence assays.Results Compared with the Sham group,AMPK expression in the myocardial tissue of CLP mice was significantly reduced(P<0.05).Compared with the CLP group,AMPK expression in the CLP+AICAR group was significantly increased(P<0.05).Survival analysis showed that CLP led to a high mortality rate(～60％ ),while AICAR treatment significantly improved the survival rate of CLP mice(P<0.05).Compared with the Sham group,cardiac output(CO),stroke volume(SV),and left ventricular end-diastolic volume(LVEDV)were significantly decreased in the CLP group(P<0.05),while left ventricular posterior wall systolic(LVPWs)and left ventricular posterior wall thickness(LVPWd)were significantly increased(P<0.05).AICAR treatment alleviated the cardiac dysfunction induced by CLP.Compared with the CLP group,mitochondrial size and the number of mitochondrial cristae in the myocardial tissue of CLP+AICAR group mice were significantly increased(P<0.05),while DHE fluorescence intensity and the number of TUNEL-positive cells were significantly reduced(P<0.05).Compared with the LPS group,ATP production,mitochondrial respiration rate,and complex Ⅰ,Ⅱ,and Ⅲ activities in NRCMs of the LPS+AICAR group were significantly increased(P<0.05),while mitochondrial and cytoplasmic ROS levels were significantly reduced(P<0.05).Compared with the control group,mitochondrial size in NRCMs of the LPS group was significantly reduced(P<0.05),while Bax and Caspase-3 expression,as well as mitochondrial fission index,were significantly increased(P<0.05),and these changes were mitigated by AICAR(P<0.05).Conclusion AMPK plays a crucial role in maintaining cardiac function and mitigating septic myocardial injury by regulating mitochondrial structure and function,energy metabolism,oxidative stress,and apoptosis.