An assay was established for detection of toxigenic Clostridioides difficile by combining microfluidic chip analysis with immunochromatography,and its performance was evaluated and compared with those of the Xpert C.difficile/Epi and VIDAS CD AB tests.Primer pairs were designed according to the tcdB and tpi genes in C.difficile.The specificity,limit of detection,reproducibility,and stability were evaluated.A total of 215 stool samples from patients with diarrhea were collected and tested in parallel with the Xpert C.difficile/Epi,VIDAS CDAB,and our assay.C.difficile was isolated from samples,and the tcdB gene was identified when discrepant results were obtained from the three above assays.Our assay showed no cross-reaction with other diarrhea-associated pathogens.Its reproducibility was 100％in testing of two standard plasmids containing tcdB and tpi genes at two concentrations(105 and 102 copies/μL).Two standard plasmids were detected after the PCR and immunochromatography reagents had been stored for 3,6,9,and 12 months,and all the results were posi-tive.The limit of detection was 10 copies/μL for toxigenic C.difficile.Testing of 33 samples positive for C.difficile with our assay(33/215,15.3％)yielded findings statistically coherent with those of the Xpert C.difficile/Epi test(kappa value=0.965).The sensitivity,specificity,positive predictive value,and negative predictive value of our assay,with respect to Xpert C.difficile/Epi as the standard,were 94.3％,100.0％,100.0％,and 98.9％;these values were significantly higher than those of VIDAS CDAB(60.0％,98.9％,91.3％,and 92.7％)(Kappa=0.714,OR=157.50,95％CI:62.03-847.28,P=0.013).In conclusion,our newly developed assay is specific,stable,and reproducible,and may be used for rapid and accu-rate detection of toxigenic C.difficile.The assay could be used for C.difficile infection screening in outpatient and emergen-cy,community medical service center,and epidemiological settings.

Objective:To investigate the changes and effects of the Wnt/β-catenin pathway during hypoxia-reoxygenation in human renal tubular epithelial(HK-2)cells.Methods:HK-2 cells were randomly divided into control group(Control),model group(Model),model+interference empty vector group(Model+NC),and model+interference group(Model+Si-β-catenin).Cells in the control group were cultured routinely.Cells in the Model group were intervened with hypoxia for 6 hours and reoxygenation for 2 hours to construct a hypoxia-reoxygenation model.HK-2 cells in the Model+NC group and the Model+Si-β-catenin group were transfected with β-catenin interference empty vector and β-catenin interference vector respectively,and then a hypoxia-reoxygenation model was constructed.After corresponding interventions,cells were collected.The apoptosis of cells in each group was detected by flow cytometry.The expression of matrix metalloproteinase-7(MMP-7)and α-smooth muscle actin(α-SMA)was detected by immunofluorescence.The expression level of Wnt pathway-related proteins was detected by Western blotting.Results:Compared with that in the control group,the apoptosis rate of cells in the Model group was significantly up-regulated,and there was no significant difference in cell apoptosis between the Model group,the Model+NC group and the Model+Si-β-catenin group.The results of WB detection showed that there was no significant difference in the expression of GSK-3β protein in the cells among the control group,the model group,the model+NC group and the model+Si-β-catenin group.Compared with that in the control group,the expression of p-GSK-3β protein in the Model group was significantly up-regulated,while there was no significant difference in the expression of p-GSK-3βprotein between the Model group,the Model+NC group and the Model+Si-β-catenin group.The results of immunofluorescence detection showed that MMP-7 and α-SMA in the Model group were significantly increased compared with that in the control group,while there was no significant difference between the Model group and the Model+NC group.Compared with the Model+NC group,MMP-7 and α-SMA in the Model+Si-β-catenin group were significantly down-regulated.Conclusion:The Wnt/β-catenin pathway is activated in HK-2 cells during hypoxia-reoxygenation and promotes the expression of fibrosis-related proteins.Inhibiting β-catenin can improve its fibrosis induction level.

Objective To explore the role of miR-429 in synovial mesenchymal stem cell-derived exosomes(SMSC-Exos)in repairing cartilage damage in temporomandibular joint osteoarthritis(TMJ OA)by extracting SMSC-Exos from human synovial tissue and screening differentially expressed microRNA(miRNA)through transcriptome sequencing.Methods Human synovial tissues were obtained from 6 patients who underwent surgery at the First Medical Center of the Chinese PLA General Hospital from June to December 2023,including 3 patients with osteoarthritis(OA group)and 3 control patients(control group),all of whom were male.SMSC-Exos were extracted from the synovial tissues for miRNA sequencing and differential expression analysis.Further,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were performed on the target genes of differentially expressed miRNA to identify key functional miRNA and construct miRNA-target gene regulatory networks and protein-protein interaction(PPI)networks of target genes.An in vitro model of rabbit condylar cartilage cell inflammatory microenvironment induced by interleukin-1β(IL-1β)was established,with the control group cultured in DMEM/F12 basic medium and the inflammation-induced group cultured in DMEM/F12 basic medium containing 10 ng/ml IL-1β.RT-qPCR was used to detect the effects of overexpressed target miRNA on the mRNA expression levels of cartilage phenotype factors such as type Ⅱ collagen α1 chain(Col2a1),aggrecan(Acan),as well as inflammatory factors including a disintegrin and metalloproteinase with thrombospondin motifs 5(Adamts5)and cyclooxygenase-2(Cox-2).Results(1)SMSC-Exos were successfully isolated,cultured,and identified.(2)miRNA sequencing of SMSC-Exos from OA and control groups revealed 16 differentially expressed miRNAs(|log2FC|>2,P<0.05).Compared with control group,7 miRNAs were up-regulated and 9 were down-regulated in OA group.GO and KEGG analysis indicated that the target genes of miR-429 were mainly involved in development process,anatomical structure development,system development,cell development and differentiation,and were enriched in inflammation-related pathways such as mitogen-activated protein kinase(MAPK)and phosphatidylinositol 3-kinase-protein kinase B(PI3K-Akt).(3)Functional validation of miR-429 in the rabbit condylar cartilage cell inflammatory model showed that overexpression of miR-429 increased the mRNA expression levels of Col2a1 and Acan(P<0.05)and decreased the mRNA expression levels of Adamts5 and Cox-2(P<0.05)in the inflammation-induced group.Conclusions miRNA sequencing of SMSC-Exos isolated and identified from human synovial tissues reveals a specific miRNA expression profile in OA patients,with miR-429 significantly down-regulated.Functional validation demonstrates that overexpression of miR-429 has reparative and anti-inflammatory effects on condylar cartilage cells in an inflammatory microenvironment.

Objective To investigate the clinical prognosis value of the positivity rate of oligoclonal bands(OCB)and immunoglobulin G(IgG)level of cerebrospinal fluid(CSF)in severe encephalitis.Methods A total of 699 cases of encephalitis patients admitted to the Department of Neurology of the First Affiliated Hospital of Air Force Military Medical University,and Xijing 986 Hospital from January 2016 to October 2020 were enrolled.According to the severity of their diseases,these patients were divided into a mild(n=360)group and a severe(n=339)group.CSF and serum samples were collected from the patient at the time of admission,and the differences in cerebrocyte count,glucose contem,glucose content,chlorine content,IgG of CSF and OCB of CSF and serum were compared.According to the GOS score of patients with severe encephalitis at discharge,the patients were divided into good prognosis group(n=259)and poor prognosis group(n=80),and multivariate Logistic regression analysis was used to analyze factors that affected the prognosis of severe encephalitis patients,and the correlation between the OCB and IgG of CSF and prognosis of patients with severe encephalitis.The predictive value of CSF IgG for the prognosis of patients with severe encephalitis was tested,and receiver operating characteristic(ROC)curve was plotted.Results Compared to patients with mild encephalitis,patients with severe encephalitis had a higher proportion of fever,pulmonary infection,status epilepticus,and mechanical ventilation,and were more likely to be complicated by stroke and hydrocephalus,and the differences were statistically significant(χ2=5.319～245.179,all P<0.05).There were significant differences in the positive rate of cerebrocyte count,chlorine content,IgG content and OCB in cerebrospinal fluid between the two groups(Z=-3.623,-4.875,-3.518,χ2=6.581,all P<0.05).CSF OCB and CSF IgG were independent risk factors for poor prognosis in patients with severe encephalitis(Wald χ2=7.295,0.001,all P<0.05).A restrictive cubic spline plot showed a linear relationship between CSF IgG and poor prognosis.The AUC(95%CI)of CSF IgG was 0.754(0.632～0.876).Conclusion The CSF IgG content and positive rate of CSF OCB in patients with severe encephalitis with poor prognosis are higher than those in patients with good prognosis,and detecting these two indicators has certain reference value for the prognosis prediction of patients with severe encephalitis.

Objective:To explore the possibility of using a inferior gluteal artery perforator flap (IGAPF) for breast reconstruction in the patient who did not have suitable donor site in back and abdomen.Methods:In November 2024, a 25-year-old unmarried and childless woman with right breast cancer received immediate right breast reconstruction by a right free IGAPF after modified right mastectomy in the Department of Breast and Thyroid Surgery, Second Affiliated Hospital of Hainan Medical University. The locations of perforators were confirmed by both Multi-detector computed tomography angiography (MDCTA) and portable Doppler blood flow detector before surgery. The IGAPF was designed to take the inferior gluteal wrinkle as the lower edge, the axis of the flap was parallel to the inferior gluteal wrinkle, and the width of the flap was estimated where the incision could be directly closed. The size of right IGAPF was 6.0 cm×19.0 cm. Sharp dissection was performed between the sarcolemma and muscle fibres of gluteus, then the perforators were dissected along the direction of muscle fibres of gluteus. The vascular pedicle was kept at about 8.0 cm in length. The diameter of artery was about 2.0 mm and that for the veins was about 1.5 mm. End-to-end anastomoses with the right thoracodorsal artery and vein were successfully carried out. The donor site was directly closed, and it was hidden in the inferior gluteal wrinkle. Postoperative outpatient clinical review was made.Results:Pathological examination reported: an invasive carcinoma of right breast, axillary lymph node metastasis (2/10). The patient recovered well and the flap survived without any complication, i.e. ischemic necrosis, infection and haematoma. The patient was off-bed at 3 days and discharged at 13 days after surgery. At the 40 days of postoperative follow-up, the patient achieved a good recovery and the lower limb activity was not affected by the surgery. The patient was satisfied with the reconstructed breast and donor site recovery. The patient followed with scheduled chemotherapy and subsequent radiotherapy. The volume of reconstructed breast was smaller than the other breast, of which the patient was fully informed before the surgery.Conclusion:A free IGAPF provides an alternative donor sites for achieving a breast reconstruction due to the reliable pedicle vessels and invisible donor scars.

OBJECTIVE: To investigate the clinical efficacy of percutaneous prying combined with modified rotary reduction with needle in the treatment of supracondylar fracture of humerus in Gartland type Ⅳ children, and to evaluate the postoperative elbow joint function, the incidence of elbow varus deformity and the application prospect of this technique. METHODS: A total of 98 children diagnosed with Gartland type Ⅳ supracondylar humeral fractures between June 2020 and January 2023 were included in this study, comprising of 57 males and 41 females. The age ranged from 2 to 14 years old with an average of (6.24±4.76) years old. There were 48 cases on the left side and 50 on the right side. The time interval from injury to surgery ranged from 2 hours to 2 days, and emergency operation was performed in 26 cases. During the operation, the Kirschner wire was inserted bluntly into the broken end of the fracture using the lever principle, and the reduction was performed by percutaneous prying. Then the modified rotary reduction method with kirschner wire was used to close and reset the Kirschner wire internal fixation, and zero incision was achieved in all cases. Flynn score was used to evaluatal the function and appearance of the elbow joint at 6 months after operation. RESULTS: The operation was successfully completed by all 98 children, and they were followed up for a duration from 6 to14 months with an average of (9.82±3.51) months. The fractures in all cases healed, the time ranged from 6 to 8 weeks with an average of (6.72±1.17) weeks. The Flynn function score of the elbow joint was rated as excellent in 95 cases and good in 3 cases at the 6-month postoperative evaluation. No related complications occurred, such as cubitus varus deformity, osteofascial compartment syndrome, Kirschner wire rupture or rejection, iatrogenic vascular and nerve injury, Volkmann's contracture or ossification myositis. CONCLUSION Percutaneous pry extraction combined with modified rotary reduction with needle in the treatment of humeral supracondylar fracture in extremely instability Gartland type Ⅳ children has the advantages of zero incision, little trauma, safety and good efficacy, and minimization of surgical trauma and scar formation. The postoperative elbow joint function recovery is good.
Humans ; Female ; Child ; Male ; Humeral Fractures/physiopathology* ; Child, Preschool ; Bone Wires ; Adolescent ; Fracture Fixation, Internal/instrumentation*

Liquiritigenin(LG)is a flavone of pharmacological importance,however,its application potential is severely limited due to its poor water solubility.LG could be disassociated slightly in water to form phenolate anion,therefore,better solubilization effect is expected by inclusion with cationic cyclodextrins(CCDs).In this work,four kinds of CCDs modified with amino groups at the primary face were synthesized,and their solid inclusion complexes with LG were successfully prepared by preparing their saturated solutions.The formation of the solid inclusion complexes was confirmed by scanning electron microscopy(SEM)and powder X-ray diffraction(PXRD),and their supramolecular binding behavior in solution was studied using multiple techniques.A 1∶1 inclusion stoichiometry of inclusion complexation was defined using Job plot by ultraviolet-visible(UV-vis)spectroscopy,and their binding stability constants(Ks)were determined as 2862.77,3494.70,6521.85 and 9599.48 L/mol using UV-vis spectroscopic titration,far more superior to that of nativeβ-CD(Ks=236.79 L/mol).This indicated that the amino side chains on CCDs could actively participate in the inclusion complexation through anion-cation interactions,significantly strengthening the host-guest binding between CCDs and LG.The inclusion modes were further elucidated based on proton and two-dimensional rotating-frame overhauser enhancement spectroscopy(2D-ROESY)nuclear magnetic resonance(NMR)experiments and molecular docking.Water solubility of LG was dramatically promoted up to 4.9 mg/mL,which was 70-fold higher than that of native LG.This study could draw inspiration for the binding and solubilization of phenols such as flavones by design of cationic macrocyclic molecules.

Objective To investigate the clinical prognosis value of the positivity rate of oligoclonal bands(OCB)and immunoglobulin G(IgG)level of cerebrospinal fluid(CSF)in severe encephalitis.Methods A total of 699 cases of encephalitis patients admitted to the Department of Neurology of the First Affiliated Hospital of Air Force Military Medical University,and Xijing 986 Hospital from January 2016 to October 2020 were enrolled.According to the severity of their diseases,these patients were divided into a mild(n=360)group and a severe(n=339)group.CSF and serum samples were collected from the patient at the time of admission,and the differences in cerebrocyte count,glucose contem,glucose content,chlorine content,IgG of CSF and OCB of CSF and serum were compared.According to the GOS score of patients with severe encephalitis at discharge,the patients were divided into good prognosis group(n=259)and poor prognosis group(n=80),and multivariate Logistic regression analysis was used to analyze factors that affected the prognosis of severe encephalitis patients,and the correlation between the OCB and IgG of CSF and prognosis of patients with severe encephalitis.The predictive value of CSF IgG for the prognosis of patients with severe encephalitis was tested,and receiver operating characteristic(ROC)curve was plotted.Results Compared to patients with mild encephalitis,patients with severe encephalitis had a higher proportion of fever,pulmonary infection,status epilepticus,and mechanical ventilation,and were more likely to be complicated by stroke and hydrocephalus,and the differences were statistically significant(χ2=5.319～245.179,all P<0.05).There were significant differences in the positive rate of cerebrocyte count,chlorine content,IgG content and OCB in cerebrospinal fluid between the two groups(Z=-3.623,-4.875,-3.518,χ2=6.581,all P<0.05).CSF OCB and CSF IgG were independent risk factors for poor prognosis in patients with severe encephalitis(Wald χ2=7.295,0.001,all P<0.05).A restrictive cubic spline plot showed a linear relationship between CSF IgG and poor prognosis.The AUC(95%CI)of CSF IgG was 0.754(0.632～0.876).Conclusion The CSF IgG content and positive rate of CSF OCB in patients with severe encephalitis with poor prognosis are higher than those in patients with good prognosis,and detecting these two indicators has certain reference value for the prognosis prediction of patients with severe encephalitis.

An assay was established for detection of toxigenic Clostridioides difficile by combining microfluidic chip analysis with immunochromatography,and its performance was evaluated and compared with those of the Xpert C.difficile/Epi and VIDAS CD AB tests.Primer pairs were designed according to the tcdB and tpi genes in C.difficile.The specificity,limit of detection,reproducibility,and stability were evaluated.A total of 215 stool samples from patients with diarrhea were collected and tested in parallel with the Xpert C.difficile/Epi,VIDAS CDAB,and our assay.C.difficile was isolated from samples,and the tcdB gene was identified when discrepant results were obtained from the three above assays.Our assay showed no cross-reaction with other diarrhea-associated pathogens.Its reproducibility was 100％in testing of two standard plasmids containing tcdB and tpi genes at two concentrations(105 and 102 copies/μL).Two standard plasmids were detected after the PCR and immunochromatography reagents had been stored for 3,6,9,and 12 months,and all the results were posi-tive.The limit of detection was 10 copies/μL for toxigenic C.difficile.Testing of 33 samples positive for C.difficile with our assay(33/215,15.3％)yielded findings statistically coherent with those of the Xpert C.difficile/Epi test(kappa value=0.965).The sensitivity,specificity,positive predictive value,and negative predictive value of our assay,with respect to Xpert C.difficile/Epi as the standard,were 94.3％,100.0％,100.0％,and 98.9％;these values were significantly higher than those of VIDAS CDAB(60.0％,98.9％,91.3％,and 92.7％)(Kappa=0.714,OR=157.50,95％CI:62.03-847.28,P=0.013).In conclusion,our newly developed assay is specific,stable,and reproducible,and may be used for rapid and accu-rate detection of toxigenic C.difficile.The assay could be used for C.difficile infection screening in outpatient and emergen-cy,community medical service center,and epidemiological settings.