ObjectiveTo analyze the epidemiological and etiological characteristics of a cluster of human metapneumovirus (HMPV) infection in a kindergarten in Gongshu District of Hangzhou City in May 2024, and to provide reference for the prevention and control of similar outbreaks. MethodsAn on-site investigation was conducted using an epidemiological case investigation form. Throat swab specimens collected from cases were screened for 13 respiratory pathogens using real-time fluorescent polymerase chain reaction (PCR). For HMPV nucleic acid positive specimens, the F gene of HMPV was used as the target gene for amplification and sequencing. The sequencing results were then compared with sequences in GenBank database to determine the virus subtypes and perform phylogenetic analyses. ResultsThe outbreak occurred in a kindergarter junior class with a total of 28 preschoolers and 3 teachers and childcare workers. A total of 11 cases (10 preschoolers and 1 teacher) were identified, including 8 male cases and 3 female cases. Clinical manifestations included fever in all 11 cases (100.00%), cough in 8 cases (72.72%), catarrhal symptoms in 4 cases (36.36%), and headache in 3 cases (27.27%). All symptoms were mild, and no severe cases were observed. A total of 11 throat swab samples were collected. Real-time fluorescent PCR test results showed that 3 samples were positive for HMPV nucleic acid, 2 samples were positive for both HMPV and Streptococcus pneumoniae, and 1 sample was positive for both HMPV and rhinovirus. The sequences of the 6 HMPV nucleic acid positive specimens were amplified and analyzed using specific primers, and all were determined to be HMPV subtype A2b. The F gene fragment sequence showed the highest similarity to PV081665.1/Brazil/2024 (99.65%), and also exhibited high similarity to PP683455.1/Indonesia/2021 (99.48%), PV016275.1/Beijing/2024 (99.31%), and PV052230.1/USA/2024 (99.13%). ConclusionThis cluster of acute respiratory tract infection was caused by HMPV subtype A2b, with co-infection of rhinovirus and Streptococcus pneumoniae. The F gene fragment sequences of the HMPV in this outbreak were highly homologous to those of the A2b strains isolated from Brazil, Beijing, Indonesia, and the the United States.

Objective The prevalence of drug-resistant Mycobacterium tuberculosis (Mtb) strains is exacerbating the global burden of tuberculosis (TB), highlighting the urgent need for new treatment strategies for TB. Methods The recombinant adenovirus vaccine expressing cyclic di-adenosine monophosphate (c-di-AMP) phosphodiesterase B (CnpB) (rAd-CnpB), was administered to normal mice via mucosal immunization, either alone or in combination with drug therapy, to treat Mtb respiratory infections in mice.Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of antibodies in serum and bronchoalveolar lavage fluid (BALF). Real-time quantitative PCR was performed to assess the transcription levels of cytokines interferon γ(IFN-γ) and interleukin 10(IL-10) in mouse lungs. Flow cytometry was used to determine the proportions of CD4⁺ and CD8⁺ T cell subsets in the lungs and spleens. ELISA was employed to measure the levels of cytokines IFN-γ, IL-2, IL-10, inflammatory factors IL-6, and tumor necrosis factor α (TNF-α) secreted by spleen cells following antigen stimulation. The bacteria loads in the lungs and spleens of Mtb-infected mice were enumerated by plate counting methods. Resluts Intranasal immunization with rAd-CnpB induced high titers of IgG in mouse serum and the production of IgG and IgA in BALF, along with alterations in T lymphocyte subsets in the lungs and spleens. Administration of rAd-CnpB, either alone or in combination with drugs, to Mtb-infected mice significantly increased serum IgG levels as well as IgA and IgG levels in BALF. rAd-CnpB immunization promoted the secretion of CnpB-specific cytokines and inflammatory factors by splenocytes in Mtb-infected mice. However, rAd-CnpB immunotherapy, either alone or combined with drugs, did not significantly affect the bacterial loads in the lungs and spleens of mice with Mtb respiratory infections. Conclusion Mucosal immunization with rAd-CnpB induced significant mucosal, humoral and cellular immune responses in mice, and significantly enhanced CnpB-specific cellular immune responses in Mtb-infected mice.
Animals ; Adenoviridae/immunology* ; Mycobacterium tuberculosis/genetics* ; Mice ; Female ; Phosphoric Diester Hydrolases/genetics* ; Tuberculosis Vaccines/administration & dosage* ; Tuberculosis/prevention & control* ; Mice, Inbred BALB C ; Cytokines ; Lung/microbiology* ; Immunization ; Bronchoalveolar Lavage Fluid/immunology* ; Immunity, Mucosal

Debates persist regarding the efficacy and safety of azvudine, particularly its real-world outcomes. This study involved patients aged ≥60 years who were admitted to 25 hospitals in mainland China with confirmed SARS-CoV-2 infection between December 1, 2022, and February 28, 2023. Efficacy outcomes were all-cause mortality during hospitalization, the proportion of patients discharged with recovery, time to nucleic acid-negative conversion (T _NANC), time to symptom improvement (T _SI), and time of hospital stay (T _HS). Safety was also assessed. Among the 5884 participants identified, 1999 received azvudine, and 1999 matched controls were included after exclusion and propensity score matching. Azvudine recipients exhibited lower all-cause mortality compared with controls in the overall population (13.3% vs. 17.1%, RR, 0.78; 95% CI, 0.67-0.90; P = 0.001) and in the severe subgroup (25.7% vs. 33.7%; RR, 0.76; 95% CI, 0.66-0.88; P < 0.001). A higher proportion of patients discharged with recovery, and a shorter T _NANC were associated with azvudine recipients, especially in the severe subgroup. The incidence of adverse events in azvudine recipients was comparable to that in the control group (2.3% vs. 1.7%, P = 0.170). In conclusion, azvudine showed efficacy and safety in older patients hospitalized with COVID-19 during the SARS-CoV-2 omicron wave in China.

BACKGROUND:Wogonin is a flavonoid extracted from the root of Scutellaria baicalensis.Previous studies have shown that baicalein has protective effects against cerebral ischemia-reperfusion injury,and can also reduce blood sugar and complications in diabetic mice,but its role and mechanism in diabetic cerebral infarction remain unclear. OBJECTIVE:To explore the effect of wogonin on nerve injury in rats with diabetic cerebral infarction and its mechanism. METHODS:Sprague-Dawley rats were randomly divided into six groups:control group,model group,low-dose wogonin group,medium-dose wogonin group,high-dose wogonin group,and high-dose wogonin+Ras homolog gene family member A(RhoA)activator group.Except for the control group,the other rats were established with diabetes and cerebral ischemia models using intraperitoneal injection of streptozotocin and middle cerebral artery occlusion.Low,medium-and high-dose wogonin groups were intragastrically given 10,20,40 mg/kg wogonin,respectively;high-dose wogonin+RhoA activator group was intragastrically given 40 mg/kg wogonin and intraperitoneally injected 10 mg/kg lysophosphatidic acid;control group and model group were given the same amount of normal saline once a day for 7 consecutive days.Rats in each group were evaluated for neurological deficits and their blood glucose levels were measured after the last dose.TTC staining was applied to detect the volume of cerebral infarction.Hematoxylin-eosin staining was applied to observe pathological changes in brain tissue.ELISA kit was applied to detect tumor necrosis factor-α,interleukin-6,malondialdehyde,and superoxide dismutase levels in brain tissue.Western blot was applied to detect the protein expression of RhoA and Rho-associated protein kinase(ROCK)2 in brain tissue. RESULTS AND CONCLUSION:Compared with the control group,the neuronal structure of rats in the model group was severely damaged,with cell necrosis and degeneration,the neurological deficit score,blood glucose level,and infarct volume were significantly elevated(P<0.05),the levels of tumor necrosis factor-α,interleukin-6,and malondialdehyde,and the protein expression of RhoA and ROCK2 in brain tissue were significantly increased(P<0.05),and the superoxide dismutase level was decreased(P<0.05).Compared with the model group,the low-,medium-,and high-dose wogonin groups showed improved neuronal damage,reduced cell degeneration and necrosis,a significant reduction in neurological deficit score,blood glucose level,infarct volume,and the levels of tumor necrosis factor-α,interleukin-6,and malondialdehyde,and the protein expression of RhoA and ROCK2 in brain tissue,and an increase in the superoxide dismutase level(P<0.05).Compared with the high-dose wogonin group,the high-dose wogonin+RhoA activator group significantly weakened the improvement in the above indexes of rats with diabetic cerebral infarction(P<0.05).To conclude,wogonin can improve the blood glucose level in rats with diabetic cerebral infarction,reduce cerebral infarction and nerve injury,and its mechanism may be related to the inhibition of RhoA/ROCK signaling pathway.

Objective To investigate the clinical efficacy of oblique lumbar interbody fusion(OLIF)combined with lateral plate fixation in the treatment of single-level adjacent segment disease(ASDis)following lumbar fusion surgery so as to evaluate the safety and effectiveness of this surgical approach.Methods A retrospective analysis was conducted on 46 patients with single-level ASDis after lumbar fusion surgery from August 2022 to October 2024.Twenty-three patients underwent OLIF combined with lateral plate fixation(OLIF group),while 23 patients received posterior lumbar interbody fusion(PLIF)(PLIF group).The following parameters were compared between the two groups:operative time,intraoperative blood loss,visual analogue scale(VAS)for pain,Oswestry disability index(ODI),disc height(DH),intervertebral foramen height(IFH),and interbody fusion status.Results All the 46 patients successfully completed surgery for single-level ASDis and were followed up for(13.7±1.1)months.The OLIF group had significantly shorter operative time[(70.7±4.6)min vs.(128.6±12.0)min]and less intraoperative blood loss[(58.6±5.7)mL vs.(313.3±47.5)mL]compared to the PLIF group(all P＜0.05).Both groups showed significant improvements in postoperative lumbar VAS and ODI scores at all follow-up time points compared to preoperative values(P＜0.05).The OLIF group exhibited significantly lower lumbar VAS scores at 3 days and 3 months postoperatively than those of the PLIF group(P＜0.05),and there was no statistical difference in VAS scores at the other follow-up time points(P＞0.05).There was no significant difference in postoperative ODI between OLIF group and PLIF group at each time point(P＞0.05).Postoperative DH and IFH were significantly improved in both groups compared to preoperative measurements(P＜0.05).In OLIF group,1 case of transient left thigh numbness resolved with conservative treatment within 2 weeks;1 case of cage subsidence was observed at 1 month postoperatively,achieving fusion without further displacement by 13 months.All the OLIF cases achieved complete fusion(fusion rate:100％).In PLIF group,2 cases of cerebrospinal fluid leakage healed with bed rest,1 case of wound exudation resolved with intensive dressing changes,and 1 case failed to achieve fusion(fusion rate:96％).Conclusion OLIF combined with lateral plate fixation demonstrates satisfactory early clinical outcomes for single-level ASDis after lumbar fusion,with significant advantages in operative efficiency(shorter time plus reduced blood loss)and short-term pain relief.Therefore,it is a safe and effective surgical approach.

BACKGROUND:In recent years,with the deepening of cell death research,necroptosis has gradually become a hot and difficult topic in academic research.Its various signaling pathways and proteins play an important role in the occurrence and development of periodontitis. People try to delay the process of periodontal tissue inflammation by preventing the occurrence of necroptosis. OBJECTIVE:To provide a review on the molecular mechanism of necroptosis and its correlation with periodontitis in the hope of providing new ideas and directions for the prevention,diagnosis,treatment and evaluation of periodontitis. METHODS:The first author searched PubMed,CNKI,and other databases in October 2023 for relevant literature published up to April 2024 with the search terms of "necroptosis,programmed cell death,periodontitis,periodontal,immunity,inflammation,receptor interacting protein kinase 1,receptor interacting protein kinase 3,mixed lineage kinase domain-like" in Chinese and "necroptosis,programmed cell death,periodontitis,periodontal,immunity,inflammation,RIP1,RIP3,MLKL" in English,respectively. The titles and abstracts of each document were read for preliminary screening,and 56 documents were finally selected for generalization and analysis. RESULTS AND CONCLUSION:(1) The main pathogenesis of periodontitis is that periodontal pathogenic bacteria stimulate the organism,leading to changes in the periodontal microenvironment,breaking the original immune balance and releasing a variety of inflammatory factors,triggering an inflammatory response resulting in destruction of periodontal tissues. Necroptosis can modulate the immuno-inflammation in periodontal tissues,thus affecting the development of periodontitis. (2) The occurrence of necroptosis is related to a variety of proteins,signaling pathways,and signaling molecules. Animal experiments have confirmed that the expression of phosphorylated mixed lineage kinase domain-like protein (pMLKL) is related to the degree of inflammation,and RIP3 and pMLKL are highly expressed in the periodontal tissues of mice suffering from periodontitis. Blocking the RIP3/MLKL pathway is conducive to the reduction of inflammation in periodontal tissues and the control of periodontitis progression. (3) To explore whether pMLKL can be used as a diagnostic indicator of periodontitis is of great value in the prevention,diagnosis and treatment of periodontitis.

Objective:To evaluate strategies for screening and diagnosing Lynch syndrome associated endom-etrial carcinoma(LS-EC)in clinical practice and analyze clinicopathological characteristics of LS-EC.Methods:A total of 258 patients with endometrial carcinoma who underwent surgery in The Fourth Hospital of Hebei Medical University from January 2019 to January 2022 were included.All enrolled patients underwent postoperative immu-nohistochemical testing for mismatch repair(MMR)proteins.Based on the expression status of the four MMR proteins,the patients were divided into deficient mismatch repair(d-MMR)group(57 cases)and proficient mis-match repair(p-MMR)group(201 cases).Among them,23 patients in the d-MMR group underwent germline gene testing for Lynch syndrome(LS).According to germline gene testing results,these patients were further classified into LS-EC(n=8)and non-LS-EC(n=15)groups.Clinicopathological features of LS-EC patients were analyzed.Results:Among the 258 endometrial carcinoma patients,57cases(22.1%)exhibited d-MMR,with MLH1 and PMS2 co-deletion being the most common(61.4%,35/57).Among the23 d-MMR patients who under-went genetic testing,8 cases(34.8%)were identified as having LS-EC,including 5 cases(62.5%)of MLH1 gene mutation,1 case(12.5%)of MSH2 gene mutation,1 case(12.5%)of PMS2 gene mutation and 1 case(12.5%)of MSH6 gene mutation.Compared with the non-LS-EC group,the LS-EC patients showed significant familial aggregation and higher pathological grade(P<0.05).Conclusions:Immunohistochemical analysis of MMR proteins combined with family history represents an effective screening strategy for LS-EC,however defini-tive diagnosis requires germline genetic testing.Among LS-EC cases,MLH1 is the most frequently mutated gene.LS-EC patients are characterized by familial clustering and high pathological grade.Discrepancies between immu-nohistochistochemistry and genetic testing results present challenges in the definitive diagnosis of LS-EC.

Objective : To investigate the expression and interaction of activating transcription factor 3(ATF3) and smad family member 4(Smad4) in pterygium. Methods: In this study, two sets of data GSE51995 and GSE2513 in NCBI database were analyzed by bioinformatics methods to screen out the key difference-expressed gene ATF3 in pterygium when normal conjunctiva was used as the control. The differential genes identified by bioinformatics analysis were further detected by reverse transcription quantitative polymerase chain reaction(RT-qPCR). Finally, the literatures were searched and The Gene Transcription Regulation Database(GTRD) was used to predict whether there was a target gene Smad4 that might bind to the target gene. The expression of Smad4 in normal conjunctival and pterygium tissues was verified by RT-qPCR, and the interaction between the target protein ATF3 and Smad4 in the pterygium was investigated through Co-Immunoprecipitation(Co-IP). Results : The results of bioinformatics a- nalysis showed that compared with normal conjunctival tissue , there was significantly different expression of ATF3 gene in pterygium tissue (P < 0. 05) . RT-qPCR confirmed that ATF3 was less expressed in pterygium tissue than normal conjunctival tissue (P < 0. 001) . Genomic data from the GTRD database revealed that Smad4 contains two ATF3 binding motifs , suggesting functional interaction potential . Compared with normal conjunctiva , RT-qPCR re- vealed Smad4 downregulation in pterygium tissues (P < 0. 01) ; Co-IP demonstrated enhanced interaction between Smad4 and ATF3 in pterygium tissues following immunoprecipitation with anti-ATF3 antibodies (P < 0. 05) . Conclusion ATF3 interacts with Smad4 in pterygium , and the low expression of both in pterygium tissues and their in- teraction may be associated with the pathogenesis of pterygium .

BackgroundChildhood represents a critical stage for cognitive development. Accurate assessment of children's cognitive abilities and understanding their developmental characteristics are essential for promoting healthy growth. However， traditional cognitive assessment methods typically rely on manual administration， presenting limitations such as low efficiency and insufficient engagement. These methods struggle to meet the assessment needs of children and are difficult to scale up for large-scale applications. ObjectiveTo develop a digital cognitive assessment tool for children， so as to provide a more convenient approach for evaluating children's cognitive functions. MethodsBased on classic psychological paradigms （Stroop Task， N-back， digit span， spatial orientation， and face-name matching）， a digital cognitive assessment tool was developed. This tool includes five tasks including color matching， shape matching， greening the home， great collector， and face-name matching， designed to assess core cognitive functions such as inhibitory control， working memory， short-term memory， spatial orientation， and semantic processing， respectively. From August 2024 to March 2025， a total of 750 students aged 9–12 yeas old from a primary school in Chengdu were enrolled and assessed using the digital cognitive assessment tool. Three months later， 40 children were randomly selected for retesting using both the digital tool and its corresponding standardized psychological paradigms. Pearson correlation analysis was conducted to examine the correlation between the pre-test and retest scores of the digital cognitive assessment tool， as well as the correlation between the digital cognitive task scores and the corresponding psychological paradigm assessment results， in order to evaluate the reliability and validity of the digital cognitive assessment tool. Additionally， differences in scores across the cognitive tasks were compared among children of different age groups and genders. ResultsA total of 699 valid samples were included. The younger age group consisted of children aged 9–10 years old （n=460）， while the older age group comprised those aged 11–12 years old （n=239）. There were 356 boys （50.93%） and 343 girls （49.07%）. In the reliability analysis， the Pearson correlation coefficients between the pre-test and retest scores of each assessment task ranged from 0.732 to 0.970 （P<0.01）， indicating statistically significant results. In the validity analysis， the Pearson correlation coefficients between each task and its corresponding standard cognitive test ranged from 0.679 to 0.988 （P<0.01）. In the color-matching task， both the main effects of age and gender were statistically significant （F=31.071， 21.198， P<0.01）. In the shape-matching task， the main effects of age， gender， and their interaction were all statistically significant （F=20.933， 5.926， 4.318， P<0.05 or 0.01）. In the greening the home task， the main effect of age was significant （F=5.243， P=0.023）. In the great collector task， the main effect of age was significant （F=33.697， P<0.01）. In the face-name matching task， only the main effect of gender was significant （F=27.016， P<0.01）. Further analysis showed that within the female group， older group scored significantly higher than younger group in five tasks（P<0.05 or 0.01）. Within the male group， younger group scored lower than older group in both the color-matching and great collector tasks （P<0.05 or 0.01）. Within the younger group， boys scored significantly higher than girls in color-matching and shape-matching tasks （P<0.01）. In the older group， girls scored significantly higher than boys in face-name matching task （P<0.01）. ConclusionThe digital cognitive assessment tool developed in this study demonstrates good reliability and validity. The development of cognitive functions in children aged 9–12 years old showed significant differences in age and gender， with specific developmental trajectories across different cognitive dimensions. At younger ages， boys outperformed girls in inhibitory control and working memory tasks， though this advantage diminished with age. At older ages， girls exhibited superior performance in semantic processing compared with boys.

Objective:To explore the effects of neutrophil extracellular traps (NET) on the proliferation and migration of breast cancer cells and the related mechanisms.Methods:The peripheral blood neutrophils were separated using density gradient centrifugation, 100 nmol/L phorbol 12-myristate 13-acetate (PMA) was used to stimulate the formation of NET. Breast cancer cell line MDA-MB-231 was selected and treated with NET (NET group) or deoxyribonucleaseⅠ (DNase Ⅰ) digested NET (NET+DNase Ⅰ group),normal cultured MDA-MB-231 cells were used as the control group. In MDA-MB-231 cells of each group; cell proliferation ability was detected using CCK-8 assay; cell migration was detected by cell scratch assay and Transwell assay; real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expressions of frizzled homolog 10 (FZD10), cyclin D1, E-cadherin and Vimentin immunofluorescence assay was performed to detect the formation of NET and the expressions of E-cadherin and Vimentin; Western blotting was used to detected the expression of FZD10-Wnt-β-catenin signaling pathway-related proteins.Results:Neutrophils were stimulated with PMA for 4 hours, and immunofluorescence assay results showed that NET exhibited fibrous reticular structure with high expression of citrullinated histone (citH3) and myeloperoxidase (MPO). The CCK-8 assay results showed that the absorbance values of cells in the control group, NET group and NET+DNase Ⅰ group were 1.25±0.06, 2.14±0.15 and 1.47±0.11, respectively, and the difference was statistically significant ( F = 80.60, P < 0.001). The results of scratch assay displayed that the percentage of wound closure in the control group, NET group and NET+DNase Ⅰ group were (36.2±1.3)%, (67.2±2.0)% and (46.3±3.2)%, respectively, and the difference was statistically significant ( F = 140.50, P < 0.001). Transwell assay indicated that the number of migrating cells in the control group, NET group and NET+DNase Ⅰ group were 317±18, 512±23 and 356±23, respectively, and the difference was statistically significant ( F = 75.39, P < 0.001). qRT-PCR assay revealed that the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET group were higher than those in the control group, while the relative expression of E-cadherin mRNA was lower than that in the control group, and the differences were statistically significant (all P < 0.001); the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin mRNA was higher than that in the NET group, and the differences were statistically significant (all P < 0.001). Immunofluorescence assay indicated that the number of cells expressing Vimentin in the control group, NET group and NET+DNase Ⅰ group were 35±6, 86±13 and 51±6, respectively, and the difference was statistically significant ( F = 24.65, P = 0.001); the number of cells expressing E-cadherin were 31±4, 11±3 and 24±2, respectively, and the difference was statistically significant ( F = 38.36, P < 0.001). Western blotting results demonstrated that the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET group were higher than those in the control group, while the relative expression of E-cadherin protein was lower than that in the control group; the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin protein was higher than that in the NET group. Conclusions:NET may promote the proliferation and migration of breast cancer cells by upregulating the expression of FZD10 and subsequently activating the Wnt-β-catenin signal pathway.