Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.

Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

Background/Aims: Metabolic dysfunction-associated steatohepatitis (MASH) is a significant risk factor for gallstone formation, but mechanisms underlying MASH-related gallstone formation remain unclear. Golgi membrane protein 1 (GOLM1) participates in hepatic cholesterol metabolism and is upregulated in MASH. Here, we aimed to explore the role of GOLM1 in MASH-related gallstone formation. Methods: The UK Biobank cohort was used for etiological analysis. GOLM1 knockout (GOLM1-/-) and wild-type (WT) mice were fed with a high-fat diet (HFD). Livers were excised for histology and immunohistochemistry analysis. Gallbladders were collected to calculate incidence of cholesterol gallstones (CGSs). Biles were collected for biliary lipid analysis. HepG2 cells were used to explore underlying mechanisms. Human liver samples were used for clinical validation. Results: MASH patients had a greater risk of cholelithiasis. All HFD-fed mice developed MASH, and the incidence of gallstones was 16.7% and 75.0% in GOLM1-/- and WT mice, respectively. GOLM1-/- decreased biliary cholesterol concentration and output. In vivo and in vitro assays confirmed that GOLM1 facilitated cholesterol efflux through upregulating ATP binding cassette transporter subfamily G member 5 (ABCG5). Mechanistically, GOLM1 translocated into nucleus to promote osteopontin (OPN) transcription, thus stimulating ABCG5-mediated cholesterol efflux. Moreover, GOLM1 was upregulated by interleukin-1β (IL-1β) in a dose-dependent manner. Finally, we confirmed that IL-1β, GOLM1, OPN, and ABCG5 were enhanced in livers of MASH patients with CGSs. Conclusions In MASH livers, upregulation of GOLM1 by IL-1β increases ABCG5-mediated cholesterol efflux in an OPN-dependent manner, promoting CGS formation. GOLM1 has the potential to be a molecular hub interconnecting MASH and CGSs.

Humans ; PPAR gamma/metabolism* ; Multiple Sclerosis ; Transcription Factors/metabolism* ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha

OBJECTIVE To study the interventional effect and mechanism of 1，8-cineole on pancreatic β cell ferroptosis induced by type 2 diabetes. METHODS In vitro ferroptosis model was established in pancreatic β cells of mice by using high glucose. The effects of low-dose and high-dose 1，8-cineole （0.25， 0.5 μmol/L） on the level of Fe2+ in pancreatic β cells were investigated. The effects of 1，8-cineole （0.5 μmol/L） combined with ferroptosis inducer Erastin （20 μmol/L） and ferroptosis inhibitor Ferrostatin-1 （20 μmol/L） on the protein expressions of glutathione peroxidase-4 （GPX4） and cyclooxygenase-2 （COX2） were also detected. The type 2 diabetes model mice were established by feeding high-sugar and high-fat diet combined with intraperitoneal injection of streptozotocin. The effects of low-dose and high-dose 1，8-cineole （50， 200 mg/kg） on the pathological morphology of pancreatic tissue， the content of iron as well as the protein expressions of GPX4 and COX2 were investigated. RESULTS The results of the cell experiment showed that compared with the model group， pretreatment with 1，8-cineole significantly reduced intracellular Fe2+ levels and upregulated GPX4 protein expression， while downregulated COX2 protein expression in pancreatic β cells （P＜0.05）. After combining with Ferrostatin-1， the expression trends of the above two proteins were the same， while there was no statistically significant difference after combining with Erastin. The results of animal experiments showed that compared with the model group， after intervention with 1，8-cineole， the structure of the pancreatic islets in mice recovered intact and their morphology improved； the iron content of pancreatic tissue and protein expression of COX2 were decreased significantly （P＜0.05）， while protein expression of GPX4 was increased significantly （P＜0.05）. CONCLUSIONS 1，8-cineole could ameliorate pancreatic β cell injury induced by diabetes， the mechanism of which may be related to reducing intracellular iron deposition and regulating ferroptosis-related proteins.

Objective:To investigate the status of medical care seeking delay and diagnosis delay of pulmonary tuberculosis (PTB) patients in Tongzhou District and Changping District of Beijing, analyze the related factors and put forward suggestions for early detection and scientific management of PTB patients.Methods:A retrospective epidemiological survey was conducted to collect the incidence data of PTB registered in Tongzhou and Changping from January 1 to December 31, 2021 by using the Chinese Tuberculosis Information Management System, and telephone interview were used for information supplement. Multivariate logistic regression model and decision tree model were used to analyze the influencing factors of medical care seeking delay and diagnosis delay of PTB patients.Results:In 2021, the medical care seeking delay time M( Q1, Q3) in the PTB patients in Tongzhou and Changping was 11 (5, 26) days, with a delay rate of 41.71%. Results from multivariate logistic regression model analysis revealed that factors influencing the medical care seeking delay included regular health check-up ( OR=0.033, 95% CI: 0.008-0.147), coughing for less than 2 weeks or showing any symptom of PTB before medical care seeking ( OR=0.378, 95% CI: 0.215-0.665), showing other symptoms before medical care seeking( OR=2.791, 95% CI: 1.710-4.555), no work or school in medical care seeking ( OR=2.990, 95% CI: 1.419-6.298). The diagnosis delay time M( Q1, Q3) in the PTB patients was 8 (0, 18) days, with a delay rate of 35.20%. Multivariate logistic regression model analysis revealed that the factors influencing the diagnosis delay of PTB included being diagnosed at a specialized tuberculosis hospital ( OR=0.426, 95% CI: 0.236-0.767) or a tuberculosis prevention and control institution ( OR=1.843, 95% CI: 1.061-3.202) and being traced as a source of infection ( OR=2.632, 95% CI: 1.062-6.521). The overall performance of the multivariate logistic regression model was comparable to that of the decision tree model, with the decision tree model exhibiting higher sensitivity but lower specificity. Conclusions:The medical care seeking delay rate and diagnosis delay rate of tuberculosis in Tongzhou and Changping were at low levels in 2021. However, it is still necessary to strengthen the health education and active screening, improve the public awareness of PTB prevention and control, and further improve the level of medical services and medical access to reduce the medical care seeking delay and diagnosis delay of PTB patients.

Objective: This study aimed to evaluate the effects of 2 endoscopic spine surgeries on the biomechanical properties of normal and osteoporotic spines. Methods: Based on computed tomography images of a healthy adult volunteer, 6 finite element models were created. After validating the normal intact model, a concentrated force of 400 N and a moment of 7.5 Nm were exerted on the upper surface of L3 to simulate 6 physiological activities of the spine. Five types of indices were used to assess the biomechanical properties of the 6 models, range of motion (ROM), maximum displacement value, intervertebral disc stress, maximum stress value, and articular protrusion stress, and by combining them with finite element stress cloud. Results: In normal and osteoporotic spines, there was no meaningful change in ROM or disc stress in the 2 surgical models for the 6 motion states. Model N1 (osteoporotic percutaneous transforaminal endoscopic discectomy model) showed a decrease in maximum displacement value of 20.28% in right lateral bending. Model M2 (unilateral biportal endoscopic model) increased maximum displacement values of 16.88% and 17.82% during left and right lateral bending, respectively. The maximum stress value of L4–5 increased by 11.72% for model M2 during left rotation. In addition, using the same surgical approach, ROM, maximum displacement values, disc stress, and maximum stress values were more significant in the osteoporotic model than in the normal model. Conclusion In both normal and osteoporotic spines, both surgical approaches were less disruptive to the physiologic structure of the spine. Furthermore, using the same endoscopic spine surgery, normal spine biomechanical properties are superior to osteoporotic spines.

Traumatic supraorbital fissure syndrome (TSOFS) is a symptom complex caused by nerve entrapment in the supraorbital fissure after skull base trauma. If the compressed cranial nerve in the supraorbital fissure is not decompressed surgically, ptosis, diplopia and eye movement disorder may exist for a long time and seriously affect the patients′ quality of life. Since its overall incidence is not high, it is not familiarized with the majority of neurosurgeons and some TSOFS may be complicated with skull base vascular injury. If the supraorbital fissure surgery is performed without treatment of vascular injury, it may cause massive hemorrhage, and disability and even life-threatening in severe cases. At present, there is no consensus or guideline on the diagnosis and treatment of TSOFS that can be referred to both domestically and internationally. To improve the understanding of TSOFS among clinical physicians and establish standardized diagnosis and treatment plans, the Skull Base Trauma Group of the Neurorepair Professional Committee of the Chinese Medical Doctor Association, Neurotrauma Group of the Neurosurgery Branch of the Chinese Medical Association, Neurotrauma Group of the Traumatology Branch of the Chinese Medical Association, and Editorial Committee of Chinese Journal of Trauma organized relevant experts to formulate Chinese expert consensus on the diagnosis and treatment of traumatic supraorbital fissure syndrome ( version 2024) based on evidence of evidence-based medicine and clinical experience of diagnosis and treatment. This consensus puts forward 12 recommendations on the diagnosis, classification, treatment, efficacy evaluation and follow-up of TSOFS, aiming to provide references for neurosurgeons from hospitals of all levels to standardize the diagnosis and treatment of TSOFS.

Objective:To explore the role of c-Jun N-terminal kinase (JNK) /c-Jun signaling pathway mediated by endoplasmic reticulum stress in triptolide-induced apoptosis of melanoma A375 cells.Methods:Nude mice were subcutaneously inoculated with melanoma A375 cells on the back to establish the animal model of melanoma. Tumor formation could be observed at approximately 3 weeks after inoculation, and then the mice were divided into 4 groups (4 mice in each group) : control group (injected with sodium chloride physiological solution via the tail vein), 0.1-, 0.2-, and 0.4-mg/kg triptolide groups (injected with 0.1, 0.2, and 0.4 mg/kg triptolide via the tail vein, respectively). Injections were performed twice a week. After 3 weeks of injections, tumors were resected, and their size and weight were measured. The apoptosis levels of tumor xenografts were detected by the terminal deoxyribonucleotide transferase-mediated dUTP nick end labeling (TUNEL) assay. qPCR was conducted to determine the mRNA expression of inositol-requiring enzyme 1 (IRE1), JNK, and c-Jun, and Western blot analysis to determine the protein expression of IRE1, JNK, c-Jun, phosphorylated-JNK (p-JNK), and phosphorylated-c-Jun (p-c-Jun). Comparisons among multiple groups were performed using one-way analysis of variance, and multiple comparisons were performed using Dunnett's test.Results:Significant differences were observed in the tumor mass, volume and tumor suppression rate among the control group, 0.1-, 0.2-, and 0.4-mg/kg triptolide groups (all P < 0.05) ; all the triptolide groups showed significantly decreased tumor masses and volumes (all P < 0.05), but significantly increased tumor suppression rates compared with the control group (all P < 0.05). The tumor apoptosis index significantly differed among the control group, 0.1-, 0.2-, and 0.4-mg/kg triptolide groups (7.67% ± 1.15%, 9.67% ± 3.21%, 62.00% ± 6.08%, and 85.67% ± 5.51%, respectively; F = 305.91, P < 0.001), and the 0.2- and 0.4-mg/kg triptolide groups showed significantly increased tumor apoptosis indices compared with the control group ( t = 17.56, 27.72, respectively, both P < 0.05). qPCR and Western blot analysis revealed significant differences in the mRNA expression of IRE1, JNK, and c-Jun among the control group, 0.1-, 0.2-, and 0.4-mg/kg triptolide groups ( F = 112.23, 27.51, 112.37, respectively, all P < 0.05), as well as in the relative protein expression levels of IRE1, JNK, c-Jun, p-JNK, and p-c-Jun among the above 4 groups (all P < 0.05). Additionally, the 0.4-mg/kg triptolide group showed significantly increased mRNA and protein expression of IRE1, JNK and c-Jun (including p-JNK, p-c-Jun) compared with the control group (all P < 0.05). The mRNA and protein expression levels of IRE1, JNK, and c-Jun in the tumor tissues tended to increase with the rise in drug concentrations, and the protein expression levels of p-JNK and p-c-Jun showed the same trend. Conclusion:Triptolide could activate the JNK/c-Jun signaling pathway mediated by the endoplasmic reticulum stress, and then induce apoptosis of melanoma A375 cells in mice.