ObjectiveTo detect the proportion of splenic follicular helper T （Tfh） cells and their functionally related cytokine expression levels in the incomplete embryo implantation disorder （EID） model mice， and to explore the immunological mechanism of Tfh in infertility caused by embryo implantation disorder. MethodsSixteen female Kunming mice were randomly divided into two groups， with eight mice in each group. On day 4 of pregnancy， an incomplete EID mouse model was established by oral gavage of mifepristone suspension， while an equal volume of saline was administered to the control group. On day 8 of pregnancy， the mice were euthanized. Flow cytometry was used to detect the levels of Tfh cells in the spleen lymphocytes of both incomplete EID mice and normal control mice. qRT-PCR was performed to measure the mRNA levels of B-cell lymphoma 6 （Bcl-6） and C-X-C chemokine receptor type 5 protein （CXCR5） in the spleen lymphocytes of both groups. Western blot was employed to assess the protein expression levels of Bcl-6 and CXCR5 in the spleen lymphocytes of both groups. Serum levels of interleukin-4 （IL-4）， IL-6， and IL-21 were measured by ELISA. Immunohistochemistry （IHC） was used to observe the expression levels of progesterone receptor （PR）， Bcl-6， and CXCR5 proteins in the uterine endometrial tissue of mice in both groups. ResultsIncomplete-type EID mice had a reduced number of embryo implantation points and reduced endometrial PR expression. Flow assay results showed that the proportion of CD4⁺CXCR5⁺Tfh cells in splenic lymphocytes of incomplete-type EID mice was significantly higher than that of normal controls （P<0.05）. Compared with the normal control group， Bcl-6 and CXCR5 mRNA levels and protein levels were elevated in splenic lymphocytes of incomplete EID mice， with statistically significant differences （P<0.05）； serum IL-4， IL-6， and IL-21 levels were elevated in incomplete EID mice， and Bcl-6 and CXCR5 proteins in the endometrium were significantly elevated （P<0.05）. ConclusionThe increase of Tfh cells and their associated cytokines Bcl-6 and CXCR5 is associated with the development of incomplete EID， and may be involved in the development of female immune infertility.

Skull segmentation in magnetic resonance image(MRI)provides realistic skull models for MEG and EEG positive problems.An MRI skull segmentation method based on bi-stream pyramid decoder and deep supervision(BiNETR)is proposed to solve the problem of difficult segmentation due to the blurred and complex structure of MRI skull imaging.The method uses a bi-stream pyramid decoder as the main decoder in the network structure of encoding-decoding,including serial dual decoders for edge information oriented and precise feature merging.Specifically,edge information oriented pyramid decoder effectively enhances the edge information based on feature sharpening to improve the edge segmentation accuracy,and the precise feature merging pyramid decoder further refines and reuses the edge-enhanced features to promote the fusion of deep and shallow features.In addition,deep supervised computation of intermediate feature loss is introduced to implant the gradient into the deep network for enhancing network training.The segmentation algorithm is validated on the skull dataset,achieving a Dice similarity coefficient of 0.880±0.039 and an average symmetric surface distance of(0.931±0.286)mm,outperforming other state-of-the-art methods.The experimental results demonstrate the effectiveness and accuracy of the proposed algorithm in MRI skull segmentation.

Objective:To explore the application value of problem-based learning (PBL) combined with mini-clinical evaluation exercise (Mini-CEX) in the development of post competency for interns in the Department of Neurology.Methods:A total of 56 interns rotating at the Department of Neurology of The First Affiliated Hospital of Chongqing Medical University from June 2023 to January 2024 were enrolled as the study subjects. They were randomly divided into a control group and an experiment group using the random number table method, with 28 interns in each group. The control group received traditional methods including small lectures and teaching rounds, while the experimental group received the PBL teaching method combined with Mini-CEX. The teaching effectiveness was evaluated through theoretical assessments, practical skill evaluations, teacher and student satisfaction surveys, and Mini-CEX scale assessments conducted at the beginning, middle, and end of the rotation for the experimental group. The data were analyzed using SPSS 23.0 software. For continuous data, the independent-samples t test or Mann-Whitney U test was used for comparison between groups. The chi-square test was used for categorical data and the Kruskal-Wallis H test for repeated-measurement data. Results:The theoretical scores [(45.36±2.67) vs. (42.00±4.29), P<0.01] and practical skill scores [(45.11±2.53) vs. (42.39±4.53), P<0.01] were significantly higher in the experimental group compared to the control group. The Mini-CEX score of the experimental group at the end of the rotation was notably higher than that at the beginning of rotation ( P<0.05), and their abilities improved continuously. The satisfaction rates of teachers and students in the experimental group were 71.43% (20/28) and 67.86% (19/28), respectively, which were significantly higher than those in the control group [39.29% (11/28) and 35.71% (10/28), P<0.05]. Conclusions:The teaching model integrating PBL and Mini-CEX can effectively enhance the post competency of interns in the Department of Neurology, thus offering a new perspective for clinical undergraduate teaching.

Objective:To investigate molecular mechanism of Helicobacter pylori(H.pylori)-mediated neutrophil apoptosis disorder.Methods:Neutrophils extracted from peripheral blood of healthy adults were used as research subjects,and H.pylori stan-dard strain NCTC11637 was added to culture system to construct a model of in vitro infection,which was divided into control group(with PBS or medium treatment),H.pylori group(H.pylori co-cultivated with neutrophils at ratio of 10∶1).After a certain time of in-fection,apoptosis-related molecules of neutrophils were detected by flow cytometry,ELISA and Western blot,respectively.Results:Cytokines IL-1β,IL-6 and TNF-α were secreted significantly higher in H.pylori group than control group,apoptosis rate was signifi-cantly lower than control group,apoptosis-related proteins Caspase-3 and Caspase-8 were higher than control group,but their activa-tion level were lower than control group.Conclusion:H.pylori infection may prolong neutrophil survival by inhibiting apoptotic pro-cess of neutrophils through inhibiting activation of Caspase-3/8.

BACKGROUND:Osteoporotic fracture is the most serious complication of osteoporosis.Previous studies have demonstrated that gut microbiota has a regulatory effect on skeletal tissue and that gut microbiota has an important relationship with osteoporotic fracture,but the causal relationship between the two is unclear. OBJECTIVE:To explore the causal relationship between gut microbiota and osteoporotic fractures using Mendelian randomization method. METHODS:The genome-wide association study(GWAS)datasets of gut microbiota and osteoporotic fracture were obtained from the IEU Open GWAS database and the Finnish database R9,respectively.Using gut microbiota as the exposure factor and osteoporotic fracture as the outcome variable,Mendelian randomization analyses with random-effects inverse variance weighted,MR-Egger regression,weighted median,simple model,and weighted model methods were performed to assess whether there is a causal relationship between gut microbiota and osteoporotic fracture.Sensitivity analyses were performed to test the reliability and robustness of the results.Reverse Mendelian randomization analyses were performed to further validate the causal relationship identified in the forward Mendelian randomization analyses. RESULTS AND CONCLUSION:The results of this Mendelian randomization analysis indicated a causal relationship between gut microbiota and osteoporotic fracture.Elevated abundance of Actinomycetales[odds ratio(OR)=1.562,95%confidence interval(CI):1.027-2.375,P=0.037),Actinomycetaceae(OR=1.561,95%CI:1.027-2.374,P=0.037),Actinomyces(OR=1.544,95%CI:1.130-2.110,P=0.006),Butyricicoccus(OR=1.781,95%CI:1.194-2.657,P=0.005),Coprococcus 2(OR=1.550,95%CI:1.068-2.251,P=0.021),Family ⅩⅢ UCG-001(OR=1.473,95%CI:1.001-2.168,P=0.049),Methanobrevibacter(OR=1.274,95%CI:1.001-1.621,P=0.049),and Roseburia(OR=1.429,95%CI:1.015-2.013,P=0.041)would increase the risk of osteoporotic fractures in patients.Elevated abundance of Bacteroidia(OR=0.660,95%CI:0.455-0.959,P=0.029),Bacteroidales(OR=0.660,95%CI:0.455-0.959,P=0.029),Christensenellacea(OR=0.725,95%CI:0.529-0.995,P=0.047),Ruminococcaceae(OR=0.643,95%CI:0.443-0.933,P=0.020),Enterorhabdus(OR=0.558,95%CI:0.395-0.788,P=0.001),Eubacterium rectale group(OR=0.631,95%CI:0.435-0.916,P=0.016),Lachnospiraceae UCG008(OR=0.738,95%CI:0.546-0.998,P=0.048),and Ruminiclostridium 9(OR=0.492,95%CI:0.324-0.746,P=0.001)would reduce the risk of osteoporotic fractures in patients.We identified 16 gut microbiota associated with osteoporotic fracture by the Mendelian randomization method.That is,using gut microbiota as the exposure factor and osteoporotic fracture as the outcome variable,eight gut microbiota showed positive causal associations with osteoporotic fracture and another eight gut microbiota showed negative causal associations with osteoporotic fracture.The results of this study not only identify new biomarkers for the early prediction of osteoporotic fracture and potential therapeutic targets in clinical practice,but also provide an experimental basis and theoretical basis for the study of improving the occurrence and prognosis of osteoporotic fracture through gut microbiota in bone tissue engineering.

Ankylosing spondylitis (AS) is linked to an increased prevalence of myocardial infarction (MI). However, research dedicated to elucidating the pathogenesis of AS-MI is lacking. In this study, we explored the biomarkers for enhancing the diagnostic and therapeutic efficiency of AS-MI. Datasets were obtained from the Gene Expression Omnibus database. We employed weighted gene co-expression network analysis and machine learning models to screen hub genes. A receiver operating characteristic curve and a nomogram were designed to assess diagnostic accuracy. Gene set enrichment analysis was conducted to reveal the potential function of hub genes. Immune infiltration analysis indicated the correlation between hub genes and the immune landscape. Subsequently, we performed single-cell analysis to identify the expression and subcellular localization of hub genes. We further constructed a transcription factor (TF)-microRNA (miRNA) regulatory network. Finally, drug prediction and molecular docking were performed. S100A12 and MCEMP1 were identified as hub genes, which were correlated with immune-related biological processes. They exhibited high diagnostic value and were predominantly expressed in myeloid cells. Furthermore, 24 TFs and 9 miRNA were associated with these hub genes. Enzastaurin, meglitinide, and nifedipine were predicted as potential therapeutic agents. Our study indicates that S100A12 and MCEMP1 exhibit significant potential as biomarkers and therapeutic targets for AS-MI, offering novel insights into the underlying etiology of this condition.
Humans ; Spondylitis, Ankylosing/complications* ; Systems Biology/methods* ; Myocardial Infarction/diagnosis* ; Biomarkers/metabolism* ; MicroRNAs/genetics* ; Gene Regulatory Networks ; Gene Expression Profiling ; Machine Learning

BACKGROUND: Macrophage polarization anomalies and dysfunction play a crucial role in the pathogenesis of immune thrombocytopenia (ITP). Itaconate is a Krebs cycle-derived immunometabolite synthesized by myeloid cells to modulate cellular metabolism and inflammatory responses. This study aimed to evaluate the immunoregulatory effects of an itaconate derivative on macrophages in patients with ITP. METHODS: Peripheral blood-derived macrophages from patients with ITP and healthy controls were treated with 4-octyl itaconate (4-OI), a derivative of itaconate that can penetrate the cell membrane. Macrophage polarization, antigen-presenting functions, and phagocytic capability were measured via flow cytometry and enzyme-linked immunosorbent assay (ELISA). Macrophage glycolysis in patients with ITP and the metabolic regulatory effect of 4-OI were detected using a Seahorse XFe96 Analyzer. An active murine model of ITP was used to evaluate the therapeutic effects of 4-OI in vivo . RESULTS: 4-OI reduced the levels of CD80 and CD86 in M1 macrophages and suppressed the release of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 pro-inflammatory cytokines, suggesting that 4-OI could hinder the polarization of macrophages toward an M1 phenotype. We found that 4-OI pretreated M1 macrophages reduced the proliferation of CD4 + T cells and promoted the differentiation of regulatory T cells. In addition, after 4-OI treatment, the phagocytic capacity of M1 macrophages toward antibody-coated platelets decreased significantly in patients with ITP. In addition, the glycolytic function of M1 macrophages was elevated in individuals with ITP compared to those in healthy controls. 4-OI treatment downregulated glycolysis in M1 macrophages. The glycolysis inhibitor 2-deoxy-d-glucose (2-DG) also inhibited the polarization of M1 macrophages and restored their functions. In vivo , 4-OI treatment significantly increased platelet counts in the active ITP murine model. CONCLUSIONS Itaconate derivative 4-OI inhibited M1 macrophage polarization and restored impaired functions through metabolic reprogramming. This study provides a novel therapeutic option for ITP.
Macrophages/metabolism* ; Humans ; Animals ; Succinates/pharmacology* ; Mice ; Male ; Female ; Adult ; Middle Aged ; Flow Cytometry ; Tumor Necrosis Factor-alpha/metabolism* ; Enzyme-Linked Immunosorbent Assay ; Purpura, Thrombocytopenic, Idiopathic/metabolism* ; Glycolysis/drug effects* ; Metabolic Reprogramming

ObjectiveBy comparing the composition and content of volatile components in raw products， wine-washed products and wine-fried products of Ligusticum sinense cv. Chaxiong rhizome（LSCR）， to investigate the influence of wine processing on the volatile components of LSCR， in order to provide a basis for the development of quality standards for LSCR and its processed products. MethodsElectronic nose was used to identify the odors of LSCR， wine-washed and wine-fried LSCR， and their volatile components were detected by headspace gas chromatography-mass spectrometry（HS-GC-MS）， and the relative mass fractions of these components were determined by peak area normalization method. Principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA） were performed on the obtained sample data by SIMCA 14.1 software， and the differential components of LSCR， wine-washed and wine-fried LSCR were screened according to the variable importance in the projection（VIP） value>1. Pearson correlation analysis was used to explore the relationship between volatile differential flavor components and electronic nose sensors. ResultsElectronic nose detection results showed that there were significant differences in the odors of LSCR， wine-washed and wine-fried LSCR， mainly reflected in the sensors S2， S4， S5， S6， S11， S12， S13. And a total of 62 compounds were identified from LSCR and its wine-processed products， among which 46， 50 and 51 compounds were identified from LSCR， wine-fried and wine-washed LSCR， respectively. There were 21 differential components between the raw products and wine-fried products， of which 10 components were increased and 11 were decreased after processing. There were 20 differential components between the raw products and wine-washed products， of which 11 constituents increased and 9 decreased after processing. There were 17 differential components between the wine-wash products and wine-fried products. Compared with the wine-washed products， the contents of 13 components in the wine-fried products increased， and the contents of 4 components decreased. The increasing trend of the content of phthalides in the wine-washed products was more obvious than that in the wine-fried products， but the content of total volatile components was higher in the wine-fried products than the wine-washed products. Correlation analysis showed that there were different degrees of correlation between the 7 differential sensors of electronic nose and 24 differential volatile components， mainly phthalides and olefins. ConclusionThe odor and the content of volatile components in LSCR changed obviously after wine processing， and n-butylphthalide， Z-butylidenephthalide and E-ligustilide can be used as the candidate differential markers of volatile components in LSCR before and after wine processing.

The objective of this experiment was to investigate the inhibitory effect and mechanism of mammary-derived extracellular vesicles(MmEVs)from mastitis dairy cows on the biofilm for-mation of Staphylococcus aureus SA1.The biofilm-forming ability of Staphylococcus aureus SA1 was confirmed using Congo red staining,and the biofilm growth curve of S.aureus SA1 was plot-ted using the crystal violet staining method.The minimum inhibitory concentration(MIC)and minimum biofilm inhibitory concentration(MBIC)of MmEVs against S.aureus SA1 were deter-mined.After treating S.aureus SA1 with different concentrations of MmEVs,the cell morphology of S.aureus SA1 was observed using transmission electron microscopy.The effects of MmEVs on S.aureus SA1 under low pH(pH value=5)or heat stress(58℃)were investigated.The hydro-phobicity index was explored using the microbial adhesion to hydrocarbons(MATH)assay.Bacte-rial conductivity was measured.The expression levels of biofilm-related genes(SarA,icaB,FnbA,ClfB,CidA,and gyrB)were detected using quantitative real-time PCR(qPCR).The results showed that MIC of MmEVs against the biofilm of S.aureus SA1 was 1 000 mg/L,and the MBIC was 500 mg/L.Under the influence of MmEVs,the internal substances of S.aureus SA1 leaked,the biofilm boundary became blurred,and the cell wall separated.At the MBIC concentration,MmEVs significantly reduced the tolerance of S.aureus SA1 to low pH(P＜0.001)and high tem-perature(P＜0.001),decreased hydrophobicity(P＜0.001),and increased bacterial conductivity(P＜0.001).At the MBIC concentration,MmEVs significantly downregulated the gene expression of Sa rA(P＜0.001),icaB(P＜0.001),FnbA(P＜0.001),ClfB(P＜0.001),and CidA(P＜0.001)in S.aureus SA1,while no significant effect was observed on the expression of the gyrB gene.In summary,MmEVs inhibit the formation of Staphylococcus aureus SA1 biofilms by sup-pressing the gene expression of SarA,icaB,FnbA,ClfB,and CidA within the biofilm.This dis-ruption damages the biofilm's morphological structure,reduces its tolerance to low pH and high temperature,decreases hydrophobicity,and increases bacterial conductivity,thereby ultimately in-hibiting the formation of S.aureus SA1 biofilms.

Objective To investigate the correlation of peripheral blood follicle suppressor-like protein L(FSTL-1)and chitosanidase 1(CHIT-1)levels with cardiovascular diseases(CVD)in hemodialysis patients and their clinical value.Methods A total of 150 hemodialysis patients treated in Chongqing Liangjiang New Area People's Hospital from January 2021 to June 2023(case group)and 60 healthy people who underwent medical checkups(control group)were selected.The case group consisted of 62 patients who had CVD(CVD group)and 88 patients who did not have CVD(non-CVD group).Enzyme-linked immunosorbent assay was used to detect the levels of FSTL-1 and CHIT-1 in peripheral blood.The levels of FSTL-1 and CHIT-1 among the groups were compared,and the correlation between the levels of FSTL-1 and CHIT-1 with the Gensini scores,high-sensitivity C-reactive protein(hs-CRP),troponin T(cTnT),and left ventricular ejection fraction(LVEF)were analyzed by Pearson's correlation test.Multivariate Logistic regression was used to analyze the influencing factors affecting the occurrence of CVD.Receiver operating characteristic(ROC)curve was used to assess the predictive efficacy of FSTL-1 and CHIT-1 for the occurrence of CVD.Results The levels of FSTL-1 and CHIT-1 were higher in the CVD group and non-CVD group than those in the control group(F=229.200,273.200,P＜0.05),and the levels of FSTL-1 and CHIT-1 in the CVD group were higher than those in the non-CVD group(P＜0.05).The peripheral blood FSTL-1 and CHIT-1 levels in hemodialysis patients were positively correlated with Gensini score,cTnT,hs-CRP(r=0.662,0.724,0.712,r=0.679,0.502,0.341,P＜0.05)and negatively correlated with LVEF(r=-0.463,-0.581,P＜0.05).Compared with the non-CVD group,CVD group had longer chronic kidney disease duration,higher hs-CRP,cTnT,Gensini score,FSTL-1,CHIT-1(P＜0.05),and lower LVEF(P＜0.05).High levels of cTnT,FSTL-1,and CHIT-1 were independent risk factors for CVD in hemodialysis patients(P＜0.05).The area under the curve of the com-bined detection of peripheral blood FSTL-1 and CHIT-1 levels for predicting the occurrence of CVD in hemo-dialysis patients was 0.852(95％CI:0.789-0.915),which was significantly larger than those of the single index of FSTL-1 and CHIT-1(Z=-2.084,-3.112,P=0.038,0.002).Conclusion The increased levels of peripheral blood FSTL-1 and CHIT-1 levels in CVD patients are correlated with cardiovascular injury,cardiac function,and inflammatory indexes in hemodialysis patients,which are independent risk factors affecting the occurrence of CVD in hemodialysis patients,and the combined detection of the two could improve the predic-tive efficacy for the occurrence of CVD in hemodialysis patients.