Objective: To explore the association between the HLA antibody positivity rate in female blood donors and pregnancy history, number of pregnancies, interval from the last pregnancy to blood donation, and age, to identify associated variables using a univariate generalized additive model (GAM), and to further analyze the predictive role of characteristic variables for HLA antibody positivity using a random forest model. Methods: HLA antibody detection was performed on 391 female blood donors using the Luminex immunomagnetic bead method. The correlation between pregnancy-related factors and HLA antibodies was analyzed using the Chi-square test. Based on R software, a univariate GAM was first constructed to analyze the association types between characteristic variables and the HLA antibody positivity rate, followed by the construction of a random forest model to evaluate the predictive value of the variables. Results: Among the 391 female blood donors without a transfusion history, the overall HLA antibody positivity rate was 26.34%. The positivity rate in donors with a pregnancy history was significantly higher than that in those without (30.09% vs 9.72%, P<0.05), and HLA antibody positivity rate increased linearly with the number of pregnancies (P<0.05). In the univariate GAM, age and number of deliveries exhibited a non-linear association with the HLA antibody positivity rate (the positivity rate increased sharply between 25-35 years of age and stabilized after 3 deliveries). Besides, the interval from the last pregnancy to blood donation showed a linear association with the HLA antibody positivity rate, and the positivity rate decreased as the interval prolonged (P<0.05). In the random forest model, age (mean decrease gini=29.26) and interval from the last pregnancy to blood donation (mean decrease gini=22.02) were core predictive variables: age was more conducive to identifying positive samples, while the interval from the last pregnancy to blood donation was more helpful for excluding negative samples. The number of deliveries (mean decrease accuracy=16.98) made a significant contribution to predicting positive samples, whereas the number of abortions had no impact. The model had an AUC of 0.583 (95% CI: 0.593 8-0.770 2), indicating a certain predictive value. Conclusion: The associated variables identified by the univariate GAM model, including age, interval from the last pregnancy to blood donation, and number of deliveries, provide a basis for key variables in the random forest model. All three variables have predictive value for HLA antibody positivity, which can provide evidence-based support for personalized transfusion management and stratified screening of female blood donors in this region.

ObjectiveTo systematically assess the public health governance capacity in Zhejiang Province, to conduct an in-depth analysis of its strengths and weaknesses, so as to provide scientific basis and strategic recommendations for further enhancement. MethodsA systematic collection of policy documents, public information reports, and research literature related to public health governance capacity in Zhejiang Province from 2002 to 2023 was conducted (encompassing a total of 1 263 policy documents, 138 pieces of information reports and 631 research articles). Based on the evaluation criteria suitable for public health systems previously developed by the research team, the basic status and magnitude of change in public health governance capacity in Zhejiang Province was evaluated. Additionally, normative gap analyses were employed to identify the strengths and weaknesses. ResultsZhejiang Province ranked 4th nationwide in terms of public health governance capacity with a score of 733.4 points (1 000.0-point maximum). The province has effectively implemented the principle of health first (scoring 698.5 points in the assessment of health-first strategy implementation) and attached sufficient importance to health-related goals (scoring 658.2 points in the scientific rationality of goal setting). However, the implementation of inter-departmental coordination and incentive mechanisms only scored 178.7 points, the feasibility of management and monitoring mechanisms scored even lower at only 144.0 points, and the coverage of incentive mechanisms scored 286.0 points. ConclusionZhejiang Province has effectively implemented its health first strategy and attached great importance to health targets, but still needs to strengthen cross-departmental coordination mechanisms and health-oriented incentives.

Objective: To develop a prediction model for mild cognitive impairment (MCI) risk among the elderly, so as to provide a tool for MCI early screening. Methods : From July 2022 to September 2024, a multi-stage stratified random cluster sampling method was used to recruit permanent residents aged ≥65 years from the Xinjiang Uygur Autonomous Region as study participants. Data on sociodemographic characteristics, nutritional status, body composition indices, bone mineral density, and handgrip strength were collected through questionnaires and physical examinations. Sarcopenia was defined based on appendicular skeletal muscle index and handgrip strength. MCI was assessed using the Mini-Mental State Examination, with adjustments for educational level. Participants were randomly divided into a training set and a validation set in a 7∶3 ratio. LASSO regression and multivariable logistic regression models were employed to screen for predictors and construct an MCI risk prediction model. The predictive performance of the model was evaluated using receiver operating characteristic (ROC) curve and decision curve analysis (DCA). Results: A total of 1 641 participants were surveyed, including 755 males (46.01%) and 886 females (53.99%). The majority of participants were aged 65-<75 years, comprising 1 154 individuals (70.32%). MCI was detected in 517 participants, corresponding to a detection rate of 31.51%. Resultsfrom LASSO regression and multivariate logistic regression analysis showed that residence (rural, OR = 2.323, 95% CI: 1.682-3.210), age (75-<85 years, OR = 1.405, 95% CI: 1.019-1.937; ≥85 years, OR = 3.655, 95% CI: 1.696-7.875), educational level (primary school, OR = 0.341, 95% CI: 0.247-0.472; junior high school, OR = 0.255, 95% CI: 0.160-0.408; high school, OR = 0.286, 95% CI: 0.154-0.531; bachelor's degree or above, OR = 0.120, 95% CI: 0.041-0.351), history of alcohol consumption (yes, OR = 3.216, 95% CI: 2.164-4.779), risk of malnutrition (yes, OR = 1.464, 95% CI: 1.064-2.014), sarcopenia (yes, OR = 3.197, 95% CI: 2.332-4.385), and waist-to-hip ratio (abnormal, OR = 1.540, 95% CI: 1.159-2.048) were identified as predictive factors for MCI among the elderly. In the training set, the area under the ROC curve, sensitivity, and specificity were 0.788, 0.719, and 0.712, respectively. In the validation set, the corresponding values were 0.784, 0.913, and 0.542, respectively. DCA demonstrated that the model provided a higher clinical net benefit for predicting MCI risk when the risk threshold probability ranged from 0.124 to 0.764. Conclusion The prediction model developed in this study demonstrates good discriminative ability and clinical utility, indicating its substantial value for predicting the MCI risk among the elderly.

Driven by the growing practical need to accelerate drug development and the continuous innovation of trial design in recent years， the number of protocol amendments during clinical trials have gradually increased， and the changed contents have become more flexible and complex， which significantly heightens the difficulty of ethical review on amendments. Against this backdrop， it is of great importance to fully leverage the role and responsibilities of ethics committees， effectively control clinical trial risks， and ensure subject safety. This paper analyzed development trends of protocol amendments in recent years， sorted out requirements for protocol amendments in Chinese regulations and guiding principles， and examined difficulties of amendment ethical review in practical work. Based on these， targeted strategies and recommendations were proposed， namely， strengthening the integration with scientific review， enhancing the formal review， adjusting the scope of review according to approval notifications， and adopting appropriate review methods， with a view to providing insights and references for the management of the amendment ethical review in drug clinical trials.

BACKGROUND:Fibromyalgia syndrome,as a common rheumatic disease,is related to central sensitization and immune abnormalities.However,the specific mechanism has not been elucidated,and there is a lack of specific diagnostic markers.Exploring the possible pathogenesis of this disease has important clinical significance. OBJECTIVE:To screen the potential diagnostic marker genes of fibromyalgia syndrome and analyze the possible immune infiltration characteristics based on bioinformatics methods,such as weighted gene co-expression network analysis(WGCNA),and machine learning. METHODS:Gene expression profiles in peripheral serum of fibromyalgia syndrome patients and healthy controls were obtained from the gene expression omnibus(GEO)database.The differentially co-expressed genes were screened in the expression profile by differential analysis and WGCNA analysis.Least absolute shrinkage and selection operator(LASSO)and support vector machine-recursive feature elimination(SVM-RFE)machine learning algorithm were further used to identify hub biomarkers,and draw receiver operating characteristic curve(ROC)to evaluate the accuracy of diagnosing fibromyalgia syndrome.Finally,single sample gene set enrichment analysis(ssGSEA)and gene set enrichment analysis(GSEA)were used to evaluate the immune cell infiltration and pathway enrichment in patients with fibromyalgia syndrome. RESULTS AND CONCLUSION:Eight down-regulated differentially expressed genes(DEGs)were obtained after differential analysis of the GSE67311 dataset according to the conditions of log2|(FC)|>0 and P<0.05.After WGCNA analysis,497 genes were included in the module(MEdarkviolet)with the highest positive correlation(r=0.22,P=0.04),and 19 genes were included in the module(MEsalmon2)with the highest negative correlation(r=-0.41,P=6×10-5).After intersecting DEGs and the module genes of WGCNA,seven genes were obtained.Four genes were screened out by LASSO regression algorithm and five genes were screened out by SVM-RFE machine learning algorithm.After the intersection of the two,three core genes were identified,which were germinal center associated signaling and motility like,integrin beta-8,and carboxypeptidase A3.The areas under the ROC curve of the three core genes were 0.744,0.739,and 0.734,respectively,indicating that they have good diagnostic value and can be used as biomarkers for fibromyalgia syndrome.The results of immune infiltration analysis showed that memory B cells,CD56 bright NK cells,and mast cells were significantly down-regulated in patients with fibromyalgia syndrome compared with the control group(P<0.05),and were significantly positively correlated with the above three biomarkers(P<0.05).The enrichment analysis suggested that there were nine fibromyalgia syndrome enrichment pathways,mainly related to olfactory transduction pathway,neuroactive ligand-receptor interaction,and infection pathway.The above results showed that the occurrence and development of fibromyalgia syndrome are related to the involvement of multiple genes,abnormal immune regulation,and multiple pathways imbalance.However,the interactions between these genes and immune cells,as well as their relationships with various pathways need to be further investigated.

BACKGROUND:Rev-erbα is involved in the regulation of inflammation,but pharmacological activation of Rev-erbα increases the risk for cardiovascular diseases.To reduce the relevant risk,an exploration on SR9009,a Rev-erbα agonist,combined with other drugs to relieve inflammation in skeletal myoblasts was conducted,laying the theoretical foundation for the treatment of inflammation-associated skeletal muscle atrophy. OBJECTIVE:To investigate the relationship of SR9009,indolepropionic acid and nuclear factor-κB signaling pathways in lipopolysaccharide-induced C2C12 myoblasts. METHODS:(1)C2C12 myoblasts were induced to differentiate in the presence of lipopolysaccharide(1 μg/mL).RNA-seq and KEGG pathway analysis were used to study signaling pathways.(2)C2C12 myoblast viability was assessed using the cell counting kit-8 assay to determine optimal concentrations of indolepropionic acid.Subsequently,cells were categorized into control group,lipopolysaccharide(1 μg/mL)group,SR9009(10 μmol/L)+lipopolysaccharide group,indolepropionic acid(80μmol/L)+lipopolysaccharide group,and SR9009+indolepropionic acid+lipopolysaccharide group.ELISA was employed to measure protein expression levels of interleukin-6 in the cultured supernatant.Real-time quantitative PCR were employed to measure mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.Western blot assay were employed to measure protein expression levels of NF-κB p65 and p-NF-κB p65.(3)After Rev-erbα was knocked down by siRNA,knockdown efficiency was assessed by RT-qPCR.And mRNA levels of interleukin-6 and tumor necrosis factor α were also measured. RESULTS AND CONCLUSION:Compared with the blank control group,lipopolysaccharide time-dependently inhibited myofibroblast fusion to form myotubes,the mRNA expression levels of interleukin-6 and tumor necrosis factor α were elevated,and the level of interleukin-6 in the cell supernatant was significantly increased.The results of KEGG pathway showed that the nuclear factor-κB signaling pathway was activated by lipopolysaccharide.Indolepropionic acid exhibited significant suppression of C2C12 myoblasts viability when its concentration exceeded 80 μmol/L.Indolepropionic acid and SR9009 inhibited the activation of NF-κB signaling pathway,thereby played an anti-inflammatory role,and suppressed the mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.Compared with the lipopolysaccharide group,the ratio of p-NF-κB p65/NF-κB p65 protein expression were downregulated.SR9009 combined with indolepropionic acid notably reduced lipopolysaccharide-induced inflammation,further downregulated the mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.The ratio of p-NF-κB p65/NF-κB p65 protein expression was significantly lower than that in the SR9009+lipopolysaccharide group or indolepropionic acid+lipopolysaccharide group.Rev-erbα increases time-dependently with lipopolysaccharide induction.The knockdown efficiency of Rev-erbα by siRNA reached over 58%,and lipopolysaccharide was added after Rev-erbα was successfully knocked down.Compared with the lipopolysaccharide group,the mRNA expression levels of interleukin-6 and tumor necrosis factor α were significantly up-regulated.These results conclude that Rev-erbα may act as a promising pharmacological target to reduce inflammation.SR9009 targeted activation of Rev-erbα combined with indolepropionic acid significantly inhibits the nuclear factor-κB signaling pathway and attenuates the inflammatory response of C2C12 myofibroblasts.Moreover,the combined anti-inflammatory effect is superior to that of the intervention alone.

AIM:This study aims to explore the mechanisms of influenza A virus(IAV)-induced macrophage inflammatory injury based on the interleukin-6(IL-6)/signal transducer and activator of transcription 3(STAT3)signaling loop and investigate the intervention effects of Ma-Xing-Shi-Gan decoction(MXSGD)-medicated serum.METHODS:RAW264.7 and BV2 cells were divided into control,Janus kinase(JAK)/STAT signaling pathway activator,inhibitor,model,oseltamivir,antiviral particle,and MXSGD groups.After IAV modeling and serum interventions,the cells were cultured for 24 and 48 h,and the indicators were detected and analyzed.ELISA,RT-qPCR,Western blot,and immuno-fluorescence assay were used to detect the secretion levels of IL-6 in the cell culture supernatant,IL-6 and STAT3 mRNA expressions,protein expression of STAT3,and expression levels of phosphorylated STAT3(p-STAT3),respectively.Pearson correlation analysis was used to evaluate the correlation between p-STAT3 and IL-6 in the two cell types.A co-cul-ture model of the two cells was constructed,and the secretion levels of IL-6 in the cell culture supernatant was measured.Molecular docking analyses were performed for STAT3 and MXSGD.RESULTS:After IAV simulation,the secretion lev-els of IL-6 in the cell culture supernatant,mRNA expression levels of IL-6 and STAT3,and protein expression levels of STAT3 and p-STAT3 in both cell lines were elevated(P<0.05 or P<0.01).Pearson correlation analysis revealed that p-STAT3 expression was positively correlated with IL-6 expression.The secretion levels of IL-6 in the co-culture model in-creased(P<0.01).MXSGD down-regulated the secretion levels of IL-6 in the cell culture supernatant mRNA,expression levels of IL-6 and STAT3,and protein expression levels of STAT3 and p-STAT3 in two kinds of cells(P<0.05 or P<0.01),and inhibited the secretion levels of IL-6 in co-culture models.STAT3 demonstrated good binding energies for liquiritin,amygdalin,and ephedrine.CONCLUSION:IAV can induce inflammatory injury in macrophages,and its mechanism may be related to activation of the IL-6/STAT3 signaling loop.MXSGD may alleviate the pathogenic effects of IAV by modulating the signaling loop.

Objective To investigate the effects of paeoniflorin(PF)on lipopolysaccharide(LPS)-induced aerobic glycolysis in renal tubular epithelial cell line HK-2 and its underlying mechanism of action.Methods This study consists of a preliminary experiment and a formal experiment.Preliminary experiment:CCK-8 assay and RT-qPCR were used respectively to measure cell viability and mRNA expression levels of inflammatory factors in HK-2 cells after LPS stimulation to determine the optimal LPS concentration for modeling as well as to evaluate the toxicity of PF and screen for its appropriate concentration.Formal experiment:HK-2 cells were divided into control group(CON group),LPS group,LPS+PF group and LPS+PF+740Y-P group.LPS was used to establish a cell model of sepsis associated-acute kidney injury(SA-AKI)in HK-2 cells,and then the cell model was treated with PF and PI3K activator 740Y-P,correspondingly for 24 h.CCK-8 assay was employed to detect cell viability,and Extracellular Acidification Rate(ECAR)Kit was utilized to measure the rate.The contents of IL-1β,IL-18,lactic acid(Lac)and lactate dehydrogenase A(LDHA)were determined with ELISA.Western blotting was applied to detect the expression of p-PI3K,p-AKT,HIF-1α,pyruvate kinase 2(PKM2,a key enzyme in aerobic glycolysis)and NOD-like receptor thermal protein domain associated protein 3(NLRP3),and immunofluorescence assay was performed to observe the expression and distribution of PKM2.Results ① Our preliminary experiment identified that the optimal concentration of LPS for modeling was 20.0 μg/mL,a safe dosage range of PF was 0～100.0 μmol/L,and its optimal therapeutic concentration was 25.0 μmol/L.② Compared with the CON group,LPS stimulation resulted in significantly decreased cell viability(P<0.05),increased ECAR(P<0.05),elevated contents of IL-1β,IL-18,Lac and LDHA(P<0.05),up-regulated protein levels of p-PI3K,p-AKT,HIF-1α,p-PKM2 and NLRP3(P<0.05),and enhanced fluorescence intensity of PKM2 in the nucleus of cells(P<0.05).Compared with the model group,PF treatment reversed all above effects induced by LPS stimulation(all P<0.05).Compared with the LPS+PF group,in the LPS+PF+740Y-P group,ECAR was elevated(P<0.05),the contents of IL-1β,IL-18,Lac and LDHA were increased(P<0.05),and the relative expression levels of p-PI3K,p-AKT,HIF-1α,p-PKM2 and NLRP3 were increased(P<0.05),and the fluorescence intensity of PKM2 was strengthened(P<0.05)and enhanced in the nucleus(P<0.05).Conclusion PF reduces aerobic glycolysis in HK-2 cells and alleviates the inflammatory response by inhibiting the PI3K/AKT/HIF-1α signaling pathway.

Objective:To investigate the association between high-risk human papillomavirus (hrHPV) persistent infection and vaginal microecology.Methods:A total of 53 women were enrolled in the gynecological clinic of Beijing Obstetrics and Gynecology Hospital from January 2020 to January 2021, including 7 women without HPV and 46 women with hrHPV infection. Among the hrHPV infected women, 24 woemn who did not use any drugs were classified as the observation group and the other 22 women who were given standardized interferon vaginal administration for 3 months were regarded as the treatment group. Vaginal secretions of all women were taken for Gram-stained microecological test at the time of enrollment and at the 4, 8, and 12 month follow-up. HPV turning negative was taken as the end point of follow-up.Results:(1) Women of hrHPV persistent infection in the observation and treatmnet groups had more times of abortions ( P=0.180). (2) The hrHPV negative conversion rate was 17% (4/24) in the observation group and 36% (8/22) in the treatment group, but the difference was not significant ( P=0.183). The median hrHPV negative conversion time were 11.0 months and 7.5 months in the observation and treatment groups, respectively, and the difference was statistically significant ( P=0.001). (3) Vaginal microecology was generally normal at the time of enrollment and at the end of follow-up in women with HPV natural negative conversion in the observation group. While vaginal microecological disorders were more common in women with hrHPV persistent infection in the observation and treatmnet groups, including high vaginal pH value, poor vaginal cleanliness, poor grade of Lactobacillus and increased vaginal clutter bacteria, and the vaginal microecological situation did not improve after the 12-month follow-up. (4) In the treatment group, women who turned HPV negative within six months all had normal vaginal microecology when enrollment (5/5). While those who turned negative six months later had a higher proportion of vaginal clutter bacteria (2/3), a poor grade of Lactobacillus (2/3) and a higher proportion of vaginal dysbiosis (2/3). Conclusions:(1) Interferon therapy could shorten the negative turning time of hrHPV. (2) Women with normal vaginal microecology have the ability to naturally clear hrHPV. (3) The vaginal microecological Gram-stain test has limited value in predicting hrHPV clearance, perhaps due to its inability to detect Lactobacillus subtypes.

Objective:To establish a rat model of vulvovaginal candidiasis (VVC) and to directly observe the histopathological and ultrastructural characteristics of vaginal mucosal barrier after Candida albicans infection and treatment with Lactobacillus crispatus.Methods:Female unmated SD rats were used to establish the VVC model and divided into three groups (normal group, VVC group, and Lactobacillus group; n=6 per group). Lactobacillus group received intravaginal administration of Lactobacillus crispatus suspension, while rats in VVC group and normal group were infused with phosphate buffered solution instead. Vaginal tissues were collected on day 4 post-treatment for HE staining and transmission electron microscopy (to observe ultrastructural pathological changes). Results:The results of HE staining revealed the disruption and desquamation of vaginal epithelium, necrotic epithelial tissues, neutrophil infiltration in Candida albicans-infected rats. Lactobacillus crispatus intervention restored the damaged vaginal mucosal structure (mucosal layers and thickness) to normal levels, mucosal layers of Lactobacillus group and normal group were 9.50±1.38 vs 10.67±1.03 ( P=0.226), mucosal thickness of Lactobacillus group and normal group were (116.50±12.14) vs (130.33±13.91) μm ( P=0.211). The results of transmission electron microscopy revealed intercellular desmosome rupture, loss of microvilli and glycocalyx on superficial cells, and mitochondrial swelling in Candida albicans-infected rats. Lactobacillus crispatus intervention restored the damaged vaginal mucosal ultrastructures (mitochondria and intercellular connections, etc.) to normal levels. Conclusions:Fungal infection severely disrupte the vaginal mucosal barrier in rats. Lactobacillus crispatus could restore the vaginal mucosal barrier and epithelial ultrastructures.