ObjectiveMagnetoencephalography (MEG), a non-invasive neuroimaging technique, meticulously captures the magnetic fields emanating from brain electrical activity. Compared with MEG based on superconducting quantum interference devices (SQUID), MEG based on optically pump magnetometer (OPM) has the advantages of higher sensitivity, better spatial resolution and lower cost. However, most of the current studies are clinical studies, and there is a lack of animal studies on MEG based on OPM technology. Pain, a multifaceted sensory and emotional phenomenon, induces intricate alterations in brain activity, exhibiting notable sex differences. Despite clinical revelations of pain-related neuronal activity through MEG, specific properties remain elusive, and comprehensive laboratory studies on pain-associated brain activity alterations are lacking. The aim of this study was to investigate the effects of inflammatory pain (induced by Complete Freund’s Adjuvant (CFA)) on brain activity in a rat model using the MEG technique, to analysis changes in brain activity during pain perception, and to explore sex differences in pain-related MEG signaling. MethodsThis study utilized adult male and female Sprague-Dawley rats. Inflammatory pain was induced via intraplantar injection of CFA (100 μl, 50% in saline) in the left hind paw, with control groups receiving saline. Pain behavior was assessed using von Frey filaments at baseline and 1 h post-injection. For MEG recording, anesthetized rats had an OPM positioned on their head within a magnetic shield, undergoing two 15-minute sessions: a 5-minute baseline followed by a 10-minute mechanical stimulation phase. Data analysis included artifact removal and time-frequency analysis of spontaneous brain activity using accumulated spectrograms, generating spectrograms focused on the 4-30 Hz frequency range. ResultsMEG recordings in anesthetized rats during resting states and hind paw mechanical stimulation were compared, before and after saline/CFA injections. Mechanical stimulation elevated alpha activity in both male and female rats pre- and post-saline/CFA injections. Saline/CFA injections augmented average power in both sexes compared to pre-injection states. Remarkably, female rats exhibited higher average spectral power 1 h after CFA injection than after saline injection during resting states. Furthermore, despite comparable pain thresholds measured by classical pain behavioral tests post-CFA treatment, female rats displayed higher average power than males in the resting state after CFA injection. ConclusionThese results imply an enhanced perception of inflammatory pain in female rats compared to their male counterparts. Our study exhibits sex differences in alpha activities following CFA injection, highlighting heightened brain alpha activity in female rats during acute inflammatory pain in the resting state. Our study provides a method for OPM-based MEG recordings to be used to study brain activity in anaesthetized animals. In addition, the findings of this study contribute to a deeper understanding of pain-related neural activity and pain sex differences.

ObjectiveTo observe the clinical effectiveness and safety of Xiaozhong Zhitong Mixture （消肿止痛合剂） combined with antibiotic bone cement in the treatment of diabetic foot ulcer （DFU） with damp-heat obstructing syndrome. MethodsA total of 72 DFU patients with damp-heat obstructing syndrome were randomly assigned to treatment group （36 cases） and the control group （36 cases）. Both groups received standard treatment and topical antibiotic bone cement for ulcer wounds， while the treatment group received oral Xiaozhong Zhitong Mixture （50 ml per time， three times daily） in additionally. Both groups underwent daily wound dressing changes for 21 consecutive days. Ulcer healing rate， serum levels of tumor necrosis factor-alpha （TNF-α）， interleukin-1 beta （IL-1β）， malondialdehyde （MDA）， superoxide dismutase （SOD）， C-reactive protein （CRP）， and white blood cell （WBC） count were observed before and after treatment， and visual analog scale （VAS） scores for wound pain， traditional Chinese medicine （TCM） syndrome scores， and the DFU Healing Scale （DMIST scale） were also compared. Liver and kidney function were evaluated before and after treatment， and adverse events such as allergic reactions， worsening ulcer pain were recorded. ResultsTotally 35 patients in the treatment group and 33 in the control group were included in the final analysis. The ulcer healing rate in the treatment group was （87.93±9.34）%， significantly higher than （81.82±12.02）% in the control group （P = 0.035）. Compared to pre-treatment levels， both groups showed significant reductions in serum CRP， WBC， MDA， IL-1β， and TNF-α levels， with an increase in SOD level （P＜0.05）. TCM syndrome scores， VAS， and DMIST scores also significantly decreased in both groups （P＜0.05）， with greater improvements in the treatment group （P＜0.05）. No significant adverse reactions were observed in either group during treatment. ConclusionXiaozhong Zhitong Mixture combined with antibiotic bone cement has significant advantages in promoting DFU healing， reducing inflammatory response， and alleviating oxidative stress in DFU patients with damp-heat obstructing syndrome， with good safety for DFU patients with damp-heat obstructing syndrome.

With the rapid development of competitive sports, the incidence of anterior cruciate ligament (ACL) injury is on the rise. Such injuries may shorten athletes′ career and lead to other long-term adverse consequences. Although athletes generally recover well after ACL reconstruction, many still struggle to return to their pre-injury performance levels. Advances in the understanding of ACL anatomy and injury mechanisms, along with the evolution of surgical techniques and rehabilitation methods, have provided more individualized and tailored options for athletes following ACL injuries. However, there is currently no consensus in China regarding surgical and rehabilitation strategies for competitive athletes aiming to return to sports after ACL injuries. To this end, the Sports Medicine Committee of the Chinese Research Hospital Association and the Editorial Board of the Chinese Journal of Trauma jointly formulated the Expert consensus on surgical treatment and rehabilitation for competitive sports athletes returning to sports after anterior cruciate ligament injury ( version 2025), and presented 14 recommendations covering surgical indications, preoperative rehabilitation, surgical timing, surgical strategies and postoperative rehabilitation strategies, aiming to improve the surgical treatment and rehabilitation system for ACL injuries in competitive athletes and facilitate their return to high-level sports performance after injury.

Objective To investigate the comparative neuroprotective effects of human umbilical cord mesenchymal stem cells(hUC-MSCs-Exos)administered via different routes on hypoxic ischemic brain damage(HIBD)in neonatal mice.Methods Healthy one-week-old SPF-grade BALB/c mice were randomly divided into 4 groups:sham operation group(n=6),model group(n=6),exosome group 1(n=8),exosome group 2(n=8).HIBD was induced using the Rice-Vannucci method.Exosome group 1 and Exosome group 2 were intraperitoneal injection/intranasal drip of phosphate buffer(PBS)100 μl containing 10 μl exosomes within 24 h after successful modeling,respectively.Sham operation and model groups were intraperitoneal injection of PBS 100 μl.On the 7th day after the intervention,neuromotor function was assessed using the horizontal grid test and pole climbing test.On the 2nd day after the evaluation,all mice were killed and their brains were removed by decapitation.HE staining was used to observe the pathological injury of brain tissue,toluidine blue staining was used to observe the survival of neurons in cerebral cortex,and TUNEL staining was used to observe the apoptosis of cerebral cortex cells.Results Compared with sham operation group,model group,exosome group 1 and exosome group 2 exhibited increased hind limb drops in horizontal grid test and climbing scores(P<0.05).No significant difference was found in model group,exosome group 1 and exosome group 2 in these measures(P<0.05).Significant pathology was observed in model group,exosome group 1 and exosome group 2 compared to sham operation group(P<0.05),with significantly reduced damage in exosome group 1 and exosome group 2 compared to model group(P<0.05).Compared with sham operation group,Nissl body count was lower in model group and exosome group 1 and exosome group 2,with a higher count in exosome group 2 compared to exosome group 1(P<0.05).Compared with sham operation group,apoptotic cells were higher in model group and exosome group 1 and exosome group 2,with a significant reduction in exosome group 1 and exosome group 2 compared to model group,and the lowest in exosome group 2(P<0.05).Conclusions hUC-MSCs-Exos can improve the neuronal motor function,promote neuron repair and inhibit apoptosis in HIBD mice.Intranasal administration of hUC-MSCs-Exos is more effective than intraperitoneal administration for reducing neuronal apoptosis in HIBP neonatal mice,offering a convenient and rapid method suitable for clinical application.

Objective To comparatively analyze the clinical characteristics of primary bilateral macronodular adrenal hyperplasia(PBMAH)and adrenal cortisol-producing Adenoma(CPA),and enhance the understanding of two diseases.Methods The clinical data of 85 PBMAH patients(PBMAH group)and 195 CPA patients(CPA group)diagnosed at Department of Endocrinology,the First Medical Center of Chinese PLA General Hospital,from September 2014 to August 2024 were retrospectively analyzed.The demographic characteristics,comorbidities,biochemical indicators,adrenocorticotropic hormone-cortisol(ACTH-F)levels,and adrenal imaging features and treatment conditions were compared between the two groups.Results(1)General characteristics:Compared with CPA group,PBMAH group had older age at diagnosis and a higher proportion of male patients.(2)Clinical characteristics:Compared with CPA group,PBMAH group had a longer disease duration,a higher proportion of subclinical Cushing's syndrome(CS),and a higher proportion of hypertension,impaired glucose tolerance/diabetes,bone mass reduction or osteoporosis,with higher serum potassium levels,and the differences were statistically significant(P<0.01).(3)Hormone levels:Both PBMAH and CPA groups showed ACTH-F rhythm disorder,significantly increased cortisol levels and suppressed ACTH.Compared with PBMAH group,CPA group had stronger autonomous cortisol secretion ability,manifested by increased midnight serum cortisol(F0:00),16:00 serum cortisol(F16:00),24-hour urinary free cortisol(24 h UFC)levels and lower 8:00 serum ACTH(ACTH8:00)and 16:00 serum ACTH(ACTH16:00)(P<0.01).After low-dose dexamethasone suppression test(LDDST),CPA group showed lower suppression rates of ACTH and cortisol,and higher proportions of paradoxical elevation in serum cortisol and 24 h UFC compared with PBMAH(P<0.01).Conclusions PBMAH has a longer disease course and higher proportions of comorbid metabolic disorders than CPA,mostly manifested as subclinical Cushing's syndrome.CPA has stronger autonomous cortisol secretion ability,with cortisol less likely to be suppressed after LDDST and more obvious paradoxical elevation of cortisol and 24 h UFC.

Objective To specifically extract and analyze nascent proteins synthesized by bone marrow mesenchymal stem cells(BMSCs)after transplantation into ischemic hearts using a technique employing mutant methionyl-tRNA synthetase(MetRSL247G)for nascent protein labeling,in order to explore the potential mechanisms of action in BMSCs post-transplantation.Methods Point mutation at position 274 of the MetRS gene in BMSCs was induced via lentiviral infection to enable azidonorleucine(ANL)-mediated labeling of nascent proteins in BMSCs.The labeling efficiency was verified by means of fluorescent non-canonical amino-acid tagging(FUNCAT).Thirty healthy female C57BL/6J mice(8-10 weeks old)were divided into control and experimental groups,with 15 mice in each group.The acute myocardial infarction model was constructed by ligating the left anterior descending coronary artery in experimental group,while control mice underwent only thoracotomy without coronary ligation.After modeling,both groups received intramyocardial injections of MetRSL247G-modified BMSCs(MetRSL247G-BMSCs)at 3 different sites in the peri-infarct ischemic region.Mice were intraperitoneally injected with ANL every 6 hours for 4 times on postoperative days 0,2,and 6(n=5 for each time point)respectively,euthanized 24 h after the last injection,and cardiac tissues were isolated.The newly synthesized and labeled proteins produced by BMSCs after transplantation into the myocardium of experimental and control groups were collected,using an enrichment technique for ANL-tagged proteins and liquid chromatography-tandem mass spectrometry(LC-MS)analysis.Gene ontology(GO)analysis,Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis,protein-protein interaction(PPI)analysis,and heatmap visualization analysis were performed to identify differentially expressed proteins at the 3 time points and screen key pathways and genes.Results Under fluorescence microscopy,the MetRSL247G lentivirus-infected BMSCs were observed to be labelled with mCherry signals,confirming the successful construction of the MetRSL247G-BMSCs cell line.Green fluorescent signals were detected only in nascent proteins in culture medium containing both MetRSL247G-BMSCs and ANL,validating the sensitivity and specificity of the labeling method.GO analysis revealed that differentially expressed proteins were primarily involved in basic cellular biological processes such as extracellular exosome formation,extracellular matrix organization,and focal adhesion.KEGG and PPI analyses indicated that the differential proteins were mainly involved in complement and coagulation cascade pathway,actin cytoskeleton regulation pathway,and apoptosis pathway.Heatmap analysis showed significantly upregulated expression of anti-apoptosis and cell adhesion-related factors in experimental group on day 1(P＜0.05),upregulated anti-apoptotic factors,pro-apoptotic factors,and cell adhesion-related factors on day 3(P＜0.05),and upregulated anti-apoptotic factors,cell differentiation-related factors,and cell adhesion-related factors on day 7(P＜0.05)compared with control group.Expression of apoptosis-inducing factor 1 was significantly downregulated on days 1 and 7(P＜0.05).On day 3,most differentially expressed proteins,including anti-apoptosis factors(Protein S100-A11,Clusterin,Gelsolin),pro-apoptosis factor(Cathepsin B),cell differentiation-related factor(Transgelin-2),and cell adhesion-related factors(Cofilin-1,Periostin,Fibronectin)were significantly upregulated(P＜0.05).Conclusions The MetRSL247G mutation enables BMSCs to incorporate ANL and synthesize labeled proteins,confirming the feasibility of this nascent protein labeling technique.Nascent proteins of BMSCs in ischemic myocardium primarily contribute to extracellular exosome secretion and extracellular matrix organization.BMSCs may adapt to and respond to ischemic and hypoxic environments by influencing complement and coagulation cascades,activating inflammatory factors,regulating actin cytoskeleton structure,and modulating apoptosis,thereby maintaining the survival of BMSCs.

Objective To explore an experimental protocol for differentiating human-induced pluripotent stem cells(iPSCs)into highly pure midbrain dopaminergic(DA)neurons.Methods By optimizing a blend of small molecules and recombinant human growth factors,iPSCs were induced to differentiate into ventral midbrain floor plate DA progenitor cells and subsequently into mature substantia nigra pars compacta DA neurons.Throughout the differentiation process,Real-time PCR and immunofluorescent staining were utilized as a method for quality assessment.Results iPSCs firstly differentiate into dopaminergic precursor cells,and then gradually differentiate into DA neurons expressing tyrosine hydroxylase(TH).Conclusion The protocol successfully yields approximately high purity tyrosine hydroxylase-positive(TH+)DA neurons.This differentiation technique offers an effective cellular model for studying the physiological mechanisms and pathogenesis of Parkinson's disease,providing valuable insights for future research and potential therapeutic strategies.

Objective To explore the regulatory role of farnesol in Candida albicans(C.albicans)biofilm cAMP-PKA signaling pathway and its correlation with drug resistance.Methods Standard,fluconazole-resistant,wild and high RAS1 gene expression strains of C.albicans were cultured to different phases of the biofilm(6,12,24,36 h),and the sessile minimal inhibitory concentration 50％(SMIC50)of fluconazole were determined by XTT reduction after farnesol treatment.The regulatory effects of farnesol on the ex-pression of genes related to the cAMP-PKA signaling pathway in standard and fluconazole-resistant strains of C.albicans,such as RAS1,CYR1,PDE2 were examined using qPCR;the effects of farnesol on the protein expression of the pathway were analyzed by Western blot.RAS1 gene expression of the wild and high RAS1 gene expression strains was measured by qPCR.Results ① Compared with the standard strain,resistant strains of C.albicans had higher levels of biofilm SMIC50 at 6,12 and 24 h;there was no significant difference in RAS1 expression(P＞0.05),while CYR1 expression increased significantly at 6 and 24 h in the biofilm(P＜0.01),and PDE2 expression decreased at 6 h in the biofilm(P＜0.01).②After treatment with farnesol,the resistance of the biofilm of the standard strain and drug-resistant strain decreased.Compared with no treatment with farnesol,the expression of RAS1 in the biofilm of the standard strain and drug-resistant strain decreased at all time points(P＜0.01);CYR1 expression decreased in the biofilm at 6,24 and 36 h,and in-creased in the biofilm at 12 h(P＜0.01);PDE2 expression increased in the 12 h biofilm(P＜0.01).③Compared with the wild strain,the high expression strain of RAS1 gene showed higher SMIC50 in the biofilm at 12 and 24 h,and significantly higher expression of RAS1 gene in the biofilm at 12,24 and 36 h(P＜0.01).④After treatment with farnesol,the resistance of wild-type strains and high expres-sion strains of RAS1 gene decreased.Compared with the untreated group,the expression of RAS1 gene in the biofilm of wild-type and RAS1 gene high expression strain decreased at 12 and 24 h(P＜0.01).Conclusion Farnesol can affect the sensitivity of C.albicans biofilm to fluconazole by regulating the expression of resistance molecules RAS1,CYR1 and PDE2 in the cAMP-PKA pathway.The regulatory effect varies at different stages of biofilm formation.

BACKGROUND:In previous studies,the equivalent structure of three-dimensional cell reconstruction of tissue engineering oral mucosa is similar to normal oral mucosa,including epithelial-like structure,lamina propria-like structure,and vascular lumen-like structure,and has initially achieved the establishment of vascular equivalent,but its vascularization characteristics are not very clear.OBJECTIVE:Vascular-like structures of vascularized oral mucosa equivalent were obtained by targeting vascular endothelial cells specific marker expression profiles correlated with laser capture microdissection system,and their vascularization ability was evaluated to reveal their vascularization characteristics.METHODS:Human gingival epithelial cells were cultured from human gingival epithelium and human gingival fibroblasts,human gingival mesenchymal stem cells were cultured from human gingival lamina propria.Human gingival mesenchymal stem cells were induced to differentiate into vascular endothelial-like cells after monoclonal expansion culture.Human gingival epithelial cells,human gingival fibroblasts,and vascular endothelial-like cells were loaded with acellular vascular matrix-0.25%human-like collagen type Ⅰ scaffold to construct the vascularized oral mucosa equivalent.The layered structure of oral mucosa equivalent(experimental group)and the acellular vascular matrix-0.25%human-like collagen type Ⅰ scaffold(control group)were implanted subcutaneously into the back of nude mice,respectively.14 days later,the incision surface of the two groups was coated with biogel.The biogel surface of the experimental group was inoculated with human gingival epithelial cells,while the control group was not inoculated with cells.The samples were collected after 14 days of feeding.The layered structure of oral mucosa equivalent was observed by morphology.The neovascular-like structures in oral mucosa equivalents were labeled by immunohistochemistry and immunofluorescence with a more comprehensive expression profile of vascular endothelial cells,and the vascularization characteristics were analyzed.A laser capture microdissection system was used to capture the neovascularization structures in the oral mucosa equivalents specifically labeled by immunohistochemistry and analyze their vascularization characteristics.RESULTS AND CONCLUSION:(1)The morphology showed that the cell level of oral mucosa equivalent was clear,and the structure was similar to that of normal oral mucosa,that is,there were epithelioid structures,lamina-like structures,and vascular cavelike structures,and there were scattered erythrocytes in the vascular cavelike structures.(2)The results of EdU Apollo tracer seed cells in the oral mucosa equivalent group showed that human gingival epithelial cells labeled with EdU Apollo 488 showed green fluorescence expression.DAPI labeled human gingival fibroblasts showed blue fluorescence expression and formed lamina-like structures in vivo.EdU Apollo 567 labeled vascular endothelial-like cells showed red fluorescence expression and formed a vascular-like structure in vivo.(3)Vascular endothelial cell specific marker expression profile immunofluorescence labeling of vascular structure showed that compared with normal oral mucosa,the expressions of CD31,CD51,CD54,CD105,Tie-2,VWF,vascular endothelial growth factor receptor 1,and vascular endothelial growth factor receptor 2 in oral mucosa equivalents were increased(P<0.000 1).There were no significant changes in CD34 expression(P>0.05).(4)Compared with the specifically labeled oral mucosal vascular structures,the expression levels of CD51,CD54,CD105,Tie-2,VWF,vascular endothelial growth factor receptor 1,and vascular endothelial growth factor receptor 2 of the oral mucosa equivalents targeted by the laser capture microdissection system were increased(P<0.000 1).There were no significant changes in expression of CD31 and CD34(P>0.05).(5)The results showed that the oral mucosa equivalent reconstructed by three-dimensional cell stratification could achieve good vascularization,and its vascularization characteristics were consistent with the immunological function and characteristics of neovascularization.Vascularization helps three-dimensional cell layer reconstruction of oral mucosa equivalent regeneration.

Objective:To explore the spatial correlation between gray matter volume (GMV) changes and neurotransmitter receptors/transporters in patients with first-episode schizophrenia (FES) .Methods:Fifty-four FES patients(FES group) and fifty-nine healthy controls (HC group) were selected from June 2014 to May 2020 in the Psychiatry Department of Renmin Hospital of Wuhan University. Structural magnetic resonance imaging (sMRI) was conducted on all subjects. Differences of GMV were compared across 400 cortical regions and 32 subcortical regions. Based on the positron emission tomography(PET) data from Neuromaps, which provides the density of 19 different neurotransmitter receptors and transporters, Spearman correlation analysis was performed to evaluate the spatial correlation between GMV changes and neurotransmitter systems.Results:Compared to the HC group, FES group exhibited significant GMV reductions in widespread cortical (90/400) and subcortical (6/32) regions (all FDR-corrected P<0.05). The effect size of GMV reduction (Cohen’s d) showed significant positive correlations with the density of 5-hydroxytryptamine 1a(5HT1a) ( r=0.400, Pspin=0.002), γ-aminobutyric acid type A receptor(GABA A)( r=0.307, Pspin=0.002), and metabotropic glutamate receptor 5(mGluR5) ( r=0.275, Pspin=0.020) receptors (all FDR-corrected P<0.05). Conclusion:GMV reductions in a wide range of brain regions existed in patients with FES. There are significant correlations between 5HT1a, GABA A and mGluR5 receptors and gray matter reduction in patients with FES. The disorder of these neurotransmitter receptors may be the potential neurobiological mechanism of gray matter structural abnormalities in the early stage of schizophrenia.