BACKGROUND:Previous studies have found that neuronal damage caused by continuous excessive fluoride exposure is related to Ca2+overload,but the mechanism of Ca2+flow conversion between intracellular calcium stores and cell apoptosis damage is still unclear.OBJECTIVE:To investigate the effect of fluoride exposure on Ca2+transport channel proteins and apoptosis levels in the mitochondria-associated endoplasmic reticulum membrane of primary cultured neural cells.METHODS:Primary nerve cells of neonatal SD rats were cultured in vitro and identified by immunofluorescence staining with neuronal nucleus-specific antibody up to day 7.The nerve cells were divided into control group(containing 0 mmol/L sodium fluoride),low fluoride group(containing 0.5 mmol/L sodium fluoride),and high fluoride group(containing 1 mmol/L sodium fluoride).The cell morphological changes were observed by light microscope 24 hours after fluorine exposure.The expression levels of apoptosis-related protein BAX/BCL-2 and calcium transfer-related pathways VDAC1,GRP 75,and IP3R were detected using western blot assay.The expression levels of VDAC1,GRP 75,and IP3R mRNA were detected by RT-PCR.Ca2+levels were detected by Rhood-2AM Ca2+probe.Mitochondrial membrane potential detection kit was used to detect the change in mitochondrial membrane potential.The level of apoptosis was determined by flow cytometry and TUNEL staining.RESULTS AND CONCLUSION:(1)The purity of neurons cultured on day 7 had been determined to be over 90％,with few impurities,good growth status,and tight cell network connections,meeting the requirements of subsequent experiments.(2)Compared with the control group,growth of neural cell clusters in the low-fluoride group and the high-fluoride group increased;the processes were broken;the cell body was rounded,and the connection network between cells was destroyed.Compared with the low-fluoride group,the cell damage changes in the high-fluoride group were more obvious.(3)Compared with the control group,the protein expressions of VDAC1,GRP75,and IP3R were increased in the low-fluoride group and the high-fluoride group(P＜0.05),and the ratio of apoptosis-related protein BAX/BCL-2 was increased(P＜0.05).Compared with the control group,the expression of VDAC1 and GRP75 mRNA in the low-fluoride group was significantly increased(P＜0.05);the expression levels of VDAC1,GRP75,and IP3R mRNA in the high-fluoride group were significantly increased(P＜0.01).(4)The level of cell apoptosis increased significantly after fluoride exposure,and the high-fluoride group was significantly higher than the control and low-fluoride groups(P＜0.01).(5)After fluoride exposure,the concentration of mitochondrial Ca2+in nerve cells increased significantly(P＜0.05),the mitochondrial membrane potential decreased(P＜0.01),and the degree of damage in the high-fluoride group was more obvious(P＜0.05).The results show that fluoride exposure impairs the morphological structure of primary neural cells,resulting in upregulation of Ca2+transfer pathway protein expression between the endoplasmic reticulum and mitochondria,mitochondrial Ca2+overload,mitochondrial damage,and increased levels of apoptosis.

Objective To construct machine learning models based on preoperative inflammatory and radiological features for the prediction of glioma grading and isocitrate dehydrogenase (IDH) mutation status, and to analyze application values of these models and identify the optimal predictive models. Methods A retrospective analysis was conducted on the data of pathologically confirmed glioma patients admitted to Tongji Hospital Affiliated to Tongji University from March 2019 to March 2023. LASSO regression was used to screen feature variables, and predictive models were constructed based on logistic regression (LR), random forest (RF), support vector machine (SVM), gradient boosting decision tree (XGBoost) and K-nearest neighbor (KNN) algorithms. The model performance was comprehensively evaluated using metrics including discrimination ability, area under the precision-recall curve (AUC), accuracy, F1 score and Brier score. The DeLong test was adopted to compare the AUC values among different models; Friedman rank-sum test was used to determine the overall performance differences of the models, with the Nemenyi test applied for multiple comparison correction. Results In the task of glioma grading prediction, the LR model achieved the highest comprehensive score (0.726), and no significant difference was observed between the LR model and the other four models; age was positively correlated with glioma grading (P=0.003). In the task of IDH mutation status prediction, the XGBoost model obtained the highest comprehensive score (0.832), which was superior to the LR (0.762, P=0.035) and KNN models (0.754, P=0.025), while no statistical differences were found between the XGBoost model and the RF or SVM models. Conclusions The LR model for glioma grading prediction and XGBoost model for IDH mutation prediction constructed based on a task-oriented strategy achieve a favorable interpretability while ensuring optimized performance, thereby providing reliable decision support for the individualized diagnosis and treatment of glioma.

BACKGROUND:Previous studies have found that neuronal damage caused by continuous excessive fluoride exposure is related to Ca2+overload,but the mechanism of Ca2+flow conversion between intracellular calcium stores and cell apoptosis damage is still unclear.OBJECTIVE:To investigate the effect of fluoride exposure on Ca2+transport channel proteins and apoptosis levels in the mitochondria-associated endoplasmic reticulum membrane of primary cultured neural cells.METHODS:Primary nerve cells of neonatal SD rats were cultured in vitro and identified by immunofluorescence staining with neuronal nucleus-specific antibody up to day 7.The nerve cells were divided into control group(containing 0 mmol/L sodium fluoride),low fluoride group(containing 0.5 mmol/L sodium fluoride),and high fluoride group(containing 1 mmol/L sodium fluoride).The cell morphological changes were observed by light microscope 24 hours after fluorine exposure.The expression levels of apoptosis-related protein BAX/BCL-2 and calcium transfer-related pathways VDAC1,GRP 75,and IP3R were detected using western blot assay.The expression levels of VDAC1,GRP 75,and IP3R mRNA were detected by RT-PCR.Ca2+levels were detected by Rhood-2AM Ca2+probe.Mitochondrial membrane potential detection kit was used to detect the change in mitochondrial membrane potential.The level of apoptosis was determined by flow cytometry and TUNEL staining.RESULTS AND CONCLUSION:(1)The purity of neurons cultured on day 7 had been determined to be over 90％,with few impurities,good growth status,and tight cell network connections,meeting the requirements of subsequent experiments.(2)Compared with the control group,growth of neural cell clusters in the low-fluoride group and the high-fluoride group increased;the processes were broken;the cell body was rounded,and the connection network between cells was destroyed.Compared with the low-fluoride group,the cell damage changes in the high-fluoride group were more obvious.(3)Compared with the control group,the protein expressions of VDAC1,GRP75,and IP3R were increased in the low-fluoride group and the high-fluoride group(P＜0.05),and the ratio of apoptosis-related protein BAX/BCL-2 was increased(P＜0.05).Compared with the control group,the expression of VDAC1 and GRP75 mRNA in the low-fluoride group was significantly increased(P＜0.05);the expression levels of VDAC1,GRP75,and IP3R mRNA in the high-fluoride group were significantly increased(P＜0.01).(4)The level of cell apoptosis increased significantly after fluoride exposure,and the high-fluoride group was significantly higher than the control and low-fluoride groups(P＜0.01).(5)After fluoride exposure,the concentration of mitochondrial Ca2+in nerve cells increased significantly(P＜0.05),the mitochondrial membrane potential decreased(P＜0.01),and the degree of damage in the high-fluoride group was more obvious(P＜0.05).The results show that fluoride exposure impairs the morphological structure of primary neural cells,resulting in upregulation of Ca2+transfer pathway protein expression between the endoplasmic reticulum and mitochondria,mitochondrial Ca2+overload,mitochondrial damage,and increased levels of apoptosis.

Objectives:To explore the clinical efficacy and the postoperative recurrence rate of the vertebral"Collapse"lesion resection for treating thoracolumbar spinal metastases compared to separation surgery.Methods:A retrospective study was conducted on 78 patients with thoracolumbar spinal metastases treated at Tianjin University General Hospital from December 2018 to December 2022.There were 42 males(53.8％)and 36 females(46.2％),with a mean age of 61.9±10.6 years(range 34-76 years).The patients were divided into separation surgery group(21 cases)and"Collapse"resection group(57 cases).Two to three weeks after surgery,radiotherapy,chemotherapy or targeted drugs were given according to the type of primary tumor after wound healing.The data on operative time,intraoperative blood loss,drainage volume,Frankel classification for spinal cord injury,and visual analogue scale(VAS)scores for pain preoperatively,7d after operation and at final follow-up,as well as local recurrence rates and perioperative complications were collected.Results:The operative time and intraoperative blood loss in the"Collapse"resection group(328.7±89.2min,782.5±89.2mL)were significantly higher than those in the separation surgery group(243.2±89.2min,585.7±89.2mL)(P＜0.05).There was no statistically significant difference in postoperative drainage volume between the two groups("Collapse"resection group 698.2±230.2mL vs separation surgery group 674.7±180.7mL)(P＞0.05).During follow-up,all the patients in the separation surgery group received stereotactic radiotherapy postoperatively;While 10 patients(21.2％)in the"Collapse"resection group underwent radiotherapy,and the remaining 47(78.8％)patients received targeted,immunological,or chemotherapy as part of comprehensive treatment.The recurrence rate in the separation surgery group was approximately 9.5％(2/21).Among the patients who received"Collapse"resection plus radiotherapy,there was 1 local recurrence,resulting in a recurrence rate of 10％(1/10),while the patients who didn't receive radiotherapy experienced 3 local recurrences,with a recurrence rate of 6.4％(3/47).The overall recurrence rate for patients in the"Collapse"resection group was 7.0％(4/57).There was no statistically significant difference in recurrence rates between the two surgical procedures(P=0.708).In the separation surgery group,postoperative complications included one case of wound infection,one case of wound dehiscence two months after postoperative radiotherapy,and one case of epidural hematoma causing compression.In the group of"Collapse"resection,complications included one case of cerebrospinal fluid leakage with secondary infection and one case of superficial wound infection.VAS scores on 7d postoperatively significantly decreased compared to preoperative scores in both groups(P＜0.05),and there were significant differences in VAS scores at final follow-up compared to preoperative scores(P＜0.05).No statistical difference in VAS scores was observed between the two groups on 7d postoperatively and at final follow-up(P＞0.05).Except for 5 Frankel A-D grade patients with no significant improvement in neurological function,the remaining 73 patients showed no deterioration in neurological function,with varying degrees of recovery of at least 1 grade,yielding an improvement rate of 93.6％(73/78).Conclusions:The vertebral"Collapse"resection for thoracolumbar spinal tumors does not require postoperative radiotherapy,as the tumor recurrence rate is comparable to that of patients treated with separation surgery followed by radiotherapy.

Aim To study the effect of p-benzoyl sali-droside(pOBz)on angiogenesis after ischemic stroke and to explore the underlying mechanism.Methods The MCAO model was prepared by suture method.Rats were divided into four groups:sham,MCAO,pOBz administration,and edaravone positive control,treated for seven days.The mNSS was used to assess the neurological impairment.Western blotting was em-ployed to detect CD31,NICD,and Hes-1 protein ex-pression,while immunofluorescence staining was ap-plies to quantify CD31-positive cells in ischemic brain tissue.In vitro an OGD/R model was established in HUVECs.Following treatment with varying pOBz con-centrations(0.01,0.1,1 μmol·L-1),the CCK-8 as-say was uses to measure cell viability,and in vitro tube formation assay was utilized to evaluate angiogenesis.Western blotting was employed again to assess CD31,NICD and Hes-1 protein levels.To further elucidate the mechanism,HUVEC were treated with the Notch inhibitor DAPT prior to grouping and pOBz administra-tion,and the same parameters were evaluated.Results pOBz significantly reduced the mNSS score of MCAO rats,increased CD31-positive cell counts,and upregu-lated CD31,NICD,and Hes-1 protein expression(P＜0.01).In vitro results further showed that pOBz could dose-dependently increase the survival rate and angio-genesis ability of HUVEC induced by OGD/R,and promote CD31,NICD and Hes-1 proteins(P＜0.01),and Notch inhibitor DAPT could reverse the above effects of pOBz.Conclusion pOBz promotes angio-genesis in HUVEC,and its mechanism involves activa-tion of the Notch signaling pathway.

BACKGROUND:The blood-brain barrier is an important structure that protects the central nervous system,and the study of the effects of high glucose on the blood-brain barrier is important for the prevention of high glucose-induced damage to the central nervous system.OBJECTIVE:To investigate the potential effect of high glucose on the blood-brain barrier function of hCMEC/D3 human brain microvascular endothelial cells.METHODS:hCMEC/D3 cells were cultured in regular sugar medium(glucose concentration of 25 mmol/L)and high-sugar medium(glucose concentration of 55 mmol/L).The morphology of cells in each group was observed by light microscopy.CCK-8 assay was used to detect changes in cell viability.A monolayer blood-brain barrier model was established using hCMEC/D3 cell line with Transwell chamber device.Changes in cell transmembrane resistance were monitored daily.The permeability of cell monolayers was detected by phenol red permeability.Flow cytometry was used to detect the apoptosis rate of the cells.Western blot assay was used to detect the expression of Bcl-2,Bax,Caspase-3,ZO-1,Occludin,Claudin-1,and histone deacetylase 4.The levels of histone deacetylase in cell supernatant were detected by ELISA.The expression of histone deacetylase 4 in cells was detected by immunofluorescence.RESULTS AND CONCLUSION:(1)The cell viability of high sugar group was significantly lower than that of control group(P＜0.000 1).(2)The cells of the control group were in a good growth state,interwoven into a dense mesh,with interconnections between synapses.The cell growth of high glucose group was suppressed,and the connection of inter-cellular synapses was reduced.(3)Compared with the control group,the transmembrane resistance value of the high glucose group was reduced(P＜0.05);phenol-red permeability of the monolayer cell membrane was increased(P＜0.05);cell apoptosis rate was increased(P＜0.01);the expression of Bax protein was increased(P＜0.000 1);the expression of Caspase-3 protein had no significant change(P＞0.05);the expression of Bcl-2,ZO-1,Occludin,Claudin-1,and histone deacetylase 4 proteins was decreased(P＜0.01,P＜0.001,P＜0.01,P＜0.000 1,P＜0.01);the fluorescence expression of histone deacetylase 4 was decreased(P＜0.001)in the high glucose group.(4)The level of histone deacetylase 4 in the cell supernatant of the high glucose group was lower than that of the control group(P＜0.05).The results show that high glucose induces the increased apoptosis and enhances permeability of hCEMCE/D3 cells,and its mechanism may be related to the decreased expression level of histone deacetylase 4.

Objective:To explore the relationship between coronary artery measurements combined with C-reactive protein(CRP), erythrocyte sedimentation rate(ESR) and coronary artery lesionsin(CAL) with incomplete Kawasaki disease (IKD) in children.Methods:Sixty-eight cases of pediatric IKD admitted to Huaibei Maternal and Child Health Care Hospital from January 2020 to April 2024 were selected as the IKD group, and 34 healthy children undergoing physical examination during the same period were selected as the control group according to the principle of 2∶1 pairing. Echocardiographic Z values including left coronary artery (LCA)-Z, right coronary artery (RCA)-Z, left anterior descending branch (LAD)-Z, left circumflex branch (LCX)-Z and CRP, ESR were compared between the two groups. General information of children with or without CAL were compared. The multivariate Logistic regression was used to screen the risk factors of IKD combined with CAL, and receiver operating characteristic (ROC) curve was used to analyze the predictive value of each index to IKD combined with CAL.Results:The LCA-Z, RCA-Z, LAD-Z, LCX-Z and the levels of CRP, ESR in the IKD group were higher than those in the control group: 3.26 ± 0.97 vs. 1.35 ± 0.28, 3.46 ± 0.92 vs. 1.01 ± 0.29, 3.18 ± 0.69 vs. 1.18 ± 0.34, 2.97 ± 0.68 vs.1.27 ± 0.34, (39.97 ± 9.03) mg/L vs. (4.21 ± 1.05) mg/L, (59.67 ± 16.34) mm/1 h vs. (12.85 ± 2.43) mm/1 h, there were statistical differences ( P<0.05). The duration of fever, intravenous immunoglobulin (IVIG) treatment response, IVIG start time, creatine kinase (CK), creatine kinase-MB(CK-MB), and N-terminal pro-B-type natriuretic peptide (NT-proBNP) between the children with IKD and CAL, IKD without CAL: (10.02 ± 2.45) d vs. (7.68 ± 1.43) d, 65.22%(15/23) vs. 88.89%(40/45), 60.87%(14/23) vs. 86.67%(39/45), (236.78 ± 59.74) U/L vs. (192.67 ± 35.41) U/L, (45.19 ± 9.85) U/L vs. (33.18 ± 9.87) U/L, (1.78 ± 0.32) μg/L vs. (0.92 ± 0.20) μg/L, there were statistical differences ( P<0.05). Logistic regression analysis showed that the duration of fever, IVIG treatment response, IVIG initiation time, CK, CK-MB, NT-proBNP, echocardiographic Z value, CRP and ESR levels were independent risk factors for IKD combined with CAL ( P<0.05). ROC curve analysis results showed that the area under the curve of IKD combined with CAL predicted by LCA-Z, RCA-Z, LADE-Z, LCX-Z, CRP and ESR was the largest, which was 0.917 (95% CI 0.824 - 0.970), which was higher than that diagnosed by each index alone ( P<0.05). Conclusions:There is a certain relationship between the echocardiographic Z values, CRP and ESR in children with IKD and CAL, which can be used to assist in the evaluation of IKD complicated with CAL.

Objective To explore methods for point estimation in adaptive enrichment design,providing a reference for effect estimation inn this context.Methods We simulated efficacy data under adaptive enrichment design and subsequentially estimated the effects using five methods:maximum likelihood method,stage 2 method,Lindley's method,parametric Bootstrap method,and the uniformly minimum variance conditionally unbiased estimated(UMVCUE)method.The performance of these methods was further evaluated.Results The simulation study showed that the maximum likelihood method has a large bias and is very sensitive to some parameters,while Lindley's method tends to underestimate and the parametric bootstrap method sometimes overestimates.The stage 2 method and UMVCUE are almost unbiased.In terms of mean squared error(MSE),all methods perform well except stage 2 estimator.Conclusion In adaptive enrichment design,the UMVCUE performs well in terms of bias and MSE.Therefore,we recommend it as the point estimator for the treatment effect in adaptive enrichment design.

Objective:To investigate the status of sedentary behavior and its influencing factors among inpatients with stable schizophrenia, providing empirical evidence for developing interventions to reduce sedentary behavior.Methods:A cross-sectional survey design was used to prospectively collect clinical data from 166 inpatients with stable schizophrenia (97 males, 69 females, mean age 56.4±8.4 years) hospitalized at the Shanghai Mental Health Center affiliated with Shanghai Jiao Tong University School of Medicine from February 2024 to May 2024. Sedentary behavior time was assessed using the Sedentary Behavior Questionnaire (SBQ), daily step count was measured via pedometers, and negative schizophrenic symptoms were evaluated using the Scale for the Assessment of Negative Symptoms (SANS). Patients were divided into a non-sedentary behavior group (≥5 000 steps/day, 66 cases) and a sedentary behavior group (<5 000 steps/day, 100 cases). Clinical variables were compared between the two groups, and binary logistic regression was used to identify influencing factors of sedentary behavior.Results:Stable inpatients with schizophrenia exhibited high levels of sedentary behavior, with an average daily sedentary time of (8.03±2.33) hours and a median daily step count of 3 352 (1 258-5 506) steps. Significant differences were observed between sedentary and non-sedentary behavior groups in Age ( t=-2.38),hospitalization duration ( Z=-1.53),blunted affect ( t=-8.37),poverty of thought ( t=-2.45),avolition ( t=-2.45),impoverished interests/social engagement ( t=-2.41),abdominal obesity ( χ2=9.18),and open vs. restricted hospital/wards environment ( χ2=8.88)(all P<0.05). Binary logistic regression analysis identified that hospital/wards environment ( OR=0.314, 95 %CI: 0.125-0.787),hospitalization duration ( OR=1.001, 95 %CI: 1.000-1.001),and the negative symptom of blunted affect ( OR=3.256, 95 %CI: 1.960-5.407)(all P<0.05) were significantly influencing factors for sedentary behavior in patients with stable schizophrenia. Conclusion:Hospitalized patients with stable schizophrenia exhibit high levels of sedentary behavior. Hospital/wards environment and blunted affect are significant factors influencing sedentary behavior.

BACKGROUND:The blood-brain barrier is an important structure that protects the central nervous system,and the study of the effects of high glucose on the blood-brain barrier is important for the prevention of high glucose-induced damage to the central nervous system.OBJECTIVE:To investigate the potential effect of high glucose on the blood-brain barrier function of hCMEC/D3 human brain microvascular endothelial cells.METHODS:hCMEC/D3 cells were cultured in regular sugar medium(glucose concentration of 25 mmol/L)and high-sugar medium(glucose concentration of 55 mmol/L).The morphology of cells in each group was observed by light microscopy.CCK-8 assay was used to detect changes in cell viability.A monolayer blood-brain barrier model was established using hCMEC/D3 cell line with Transwell chamber device.Changes in cell transmembrane resistance were monitored daily.The permeability of cell monolayers was detected by phenol red permeability.Flow cytometry was used to detect the apoptosis rate of the cells.Western blot assay was used to detect the expression of Bcl-2,Bax,Caspase-3,ZO-1,Occludin,Claudin-1,and histone deacetylase 4.The levels of histone deacetylase in cell supernatant were detected by ELISA.The expression of histone deacetylase 4 in cells was detected by immunofluorescence.RESULTS AND CONCLUSION:(1)The cell viability of high sugar group was significantly lower than that of control group(P＜0.000 1).(2)The cells of the control group were in a good growth state,interwoven into a dense mesh,with interconnections between synapses.The cell growth of high glucose group was suppressed,and the connection of inter-cellular synapses was reduced.(3)Compared with the control group,the transmembrane resistance value of the high glucose group was reduced(P＜0.05);phenol-red permeability of the monolayer cell membrane was increased(P＜0.05);cell apoptosis rate was increased(P＜0.01);the expression of Bax protein was increased(P＜0.000 1);the expression of Caspase-3 protein had no significant change(P＞0.05);the expression of Bcl-2,ZO-1,Occludin,Claudin-1,and histone deacetylase 4 proteins was decreased(P＜0.01,P＜0.001,P＜0.01,P＜0.000 1,P＜0.01);the fluorescence expression of histone deacetylase 4 was decreased(P＜0.001)in the high glucose group.(4)The level of histone deacetylase 4 in the cell supernatant of the high glucose group was lower than that of the control group(P＜0.05).The results show that high glucose induces the increased apoptosis and enhances permeability of hCEMCE/D3 cells,and its mechanism may be related to the decreased expression level of histone deacetylase 4.