Inteins are unique protein insertion sequences capable of self-excision, enabling the covalent ligation of flanking extein peptides via amide bond formation. This process proceeds spontaneously without requiring external enzymes, cofactors, or chemical reagents, granting inteins exceptional biocompatibility and traceless performance in protein engineering applications. Split inteins represent a specialized and versatile subclass whose splicing domains are encoded by two separate gene fragments rather than a single continuous open reading frame. These fragments, known as the N-terminal (IntN) and C-terminal (IntC) split inteins, associate through non-covalent interactions including hydrophobic forces, hydrogen bonds, and van der Waals forces to assemble into an active three-dimensional structure, which then drives efficient extein ligation and enables protein trans-splicing. Protein trans-splicing mediated by split inteins has become a cornerstone for traceless protein ligation owing to its high specificity and irreversibility, fundamentally reshaping strategies for protein modification, assembly, and functional regulation. Compared with traditional chemical ligation methods, split intein systems require no complex chemical derivatization of peptide fragments and can operate efficiently at micromolar concentrations under physiological conditions, thus avoiding structural and functional damage caused by organic reagents. In contrast to enzymatic ligation tools such as sortase, split inteins eliminate the need for additional enzymes or cofactors, simplifying reaction systems, reducing costs, and minimizing non-specific side products. These distinctive advantages render split inteins highly promising for applications in chemical biology, synthetic biology, and biopharmaceutical development. In recent years, deepened mechanistic understanding has established structure-guided rational design as the primary approach to overcoming key limitations of split inteins, including intrinsic aggregation propensity, strict extein sequence dependence, and limited splicing efficiency. Bioinformatic tools have been used to identify aggregation-prone regions in the IntN fragment, and site-directed mutagenesis of hydrophobic residues, relocation of split sites, or removal of misfolding-prone sequences has substantially reduced in vitro aggregation and improved soluble expression and assembly activity. Rational engineering of catalytic residues and adjacent flexible loops has relaxed strict amino acid preferences at extein junctions, enhancing sequence tolerance and reducing the risk of functional impairment in target proteins. Consensus design based on multiple sequence alignments has yielded ultra-fast splicing variants such as Cfa DnaE and Cat-TerL, which exhibit significantly accelerated kinetics and improved tolerance to denaturing conditions. Meanwhile, advances in structural biology have further clarified the conformational dynamics and catalytic mechanisms of splicing, supporting the precise design of high-performance intein modules. On this basis, electrostatic interaction tuning and metagenomic screening have yielded multiple mutually orthogonal split intein pairs, enabling selective multi-fragment protein ligation and providing new routes for the efficient synthesis of large multi-domain functional proteins. With these engineered split inteins offering continuously improved performance and expanded applicability, protein trans-splicing has been widely applied in numerous cutting-edge areas of protein research and biomedicine. In gene delivery, split intein-based systems overcome the packaging limit of adeno-associated viral vectors, enabling the accurate reconstitution of large therapeutic proteins and base editors in target cells, thereby enhancing the efficacy and scope of gene therapy for genetic diseases. In internal protein sequence editing, split inteins mediate precise sequence replacement and modification in flexible regions or loops of target proteins, without the need for complex multi-step ligation and protein refolding involved in traditional protein semisynthesis. In protein-protein interaction studies, intein-mediated splicing covalently captures transient and weak intracellular complexes, enabling sensitive, high-throughput interaction detection and drug screening. In synthetic biology, conditionally controllable splicing systems support the construction of diverse intracellular and cell-surface biological logic gates for the precise regulation of cellular behavior. In mechanistic biochemical research, split inteins enable photocatalytic proximity labeling and site-specific tagging, allowing the preparation of homogeneous protein samples carrying precise post-translational modifications such as ubiquitination and polyglutamylation for chromatin interactome analysis and epigenetic studies. Moreover, covalent trapping strategies using split inteins stabilize transient enzymatic intermediates, providing unprecedented insights into molecular mechanisms such as nucleosome ubiquitination that are difficult to elucidate using conventional methods. This review systematically summarizes key technological advances in split inteins over the past decade, highlighting engineering strategies, mechanistic insights, and the development of orthogonal components. It comprehensively surveys emerging applications at the frontiers of protein research, analyzes current core challenges, and proposes future directions, particularly emphasizing artificial intelligence-driven de novo design and novel splicing pathways to break existing technical bottlenecks. By enabling traceless, efficient, and versatile protein manipulation, split inteins continue to serve as indispensable tools that drive innovation in protein engineering and fundamental life science research.

Neurocritical care involves complex pathophysiological mechanisms, and its incidence is higher, injuries are more severe, and treatment is more challenging in high-altitude environments. This consensus, based on the latest domestic and international evidence-based medical data, establishes a standardized, goal-oriented framework for neurocritical care management applicable in high-altitude regions and nationwide. The consensus was developed following international standards for evidence quality assessment and underwent two rounds of Delphi expert consultation, resulting in 32 recommendation statements covering three parts: management systems, monitoring and assessment, and core strategies. Key updates include: advocating for the establishment of independent neurocritical care units and implementing precise tiered diagnosis and treatment based on the "Five Differences in Critical Care" concept; constructing a "trinity" multimodal brain monitoring system centered on cerebral blood flow, cerebral oxygenation, and brain function, emphasizing routine bedside transcranial Doppler ultrasound, cerebral oximetry, and continuous electroencephalography monitoring; shifting management strategies from mild hypothermia therapy to targeted temperature management, and defining the "446" target management pathway for the supercritical stage; emphasizing the assessment of static and dynamic cerebrovascular autoregulation functions through multimodal methods to achieve individualized optimal mean arterial pressure management; elevating cerebrospinal fluid management goals to the level of "glymphatic system" function maintenance; implementing a multidisciplinary collaborative, whole-process management model focusing on patients' long-term neurological functional outcomes; de-escalation criteria include multidimensional indicators such as recovery of brain structure, restoration of cerebrovascular autoregulation, improvement in cerebrospinal fluid dynamics, and reduction in biomarker levels; and integrating cutting-edge technologies like artificial intelligence into post-critical care management and rehabilitation planning. This consensus systematically integrates the entire process of neurocritical care management, reflecting the modern connotation of goal-oriented, dynamic, and multimodal integration in neurocritical care medicine. It aims to adapt to new trends such as deepening understanding of pathophysiological mechanisms, the integration of medicine and engineering, and the empowerment of artificial intelligence, thereby further advancing the discipline of critical care medicine.

Prostate-specific membrane antigen (PSMA) is expressed in the neovasculature of various malignancies, such as colorectal cancer (CRC) and hepatocellular carcinoma (HCC). However, PSMA expression in hepatic CRC metastasis has not been studied in detail. Methods: The PSMA expression in primary CRC and corresponding hepatic metastasis was evaluated by immunohistochemistry in a metastatic CRC cohort (n = 56), which was divided into subgroups according to treatment history and timing of metastasis. Demographic and histological characteristics of primary CRC were collected and their relationships with PSMA expression were examined. Additionally, the PSMA expression in resected HCC (n = 76) was compared with that of hepatic CRC metastasis. Results: In primary CRC, PSMA level showed a positive association with tumor size. Lower PSMA expression in hepatic metastasis was associated with higher primary CRC grade, advanced pTNM stage at the time of CRC resection, presence of tumor deposit, and unresectability of metastatic lesion. PSMA expression in primary CRC correlated with that in hepatic metastasis only in concurrent and untreated metastasis subgroup. PSMA expression in primary CRC and hepatic metastasis, regardless of treatment history and timing of metastasis, was not significantly different from that of HCC. Conclusions: Several adverse pathological features of primary CRC were associated with a lower PSMA expression in hepatic metastasis. PSMA expression in hepatic metastasis correlated with that of primary CRC only in concurrent and untreated subgroup. Primary HCC and hepatic CRC metastasis show comparable levels of PSMA expression.

ObjectiveTo investigate whether the effect of Danggui Shaoyaosan （DSS） on cognitive impairment is related to its promotion of ketone body metabolism in the brain and regulation of glutamate/α-amino-3-hydroxy-5-methyl-4-isox-azolepropionic acid receptor （AMPAR） pathway. MethodsMale APP/PS1 mice were randomized into model， low-dose （3.2 g·kg^-1） DSS， medium-dose （6.4 g·kg^-1） DSS， high-dose （12.8 g·kg^-1） DSS， ketogenic diet， and inhibitor （talampanel， 5 mg·kg^-1） groups. C57BL/6J wild-type mice were selected as the normal group. The talampanel group was intragastrically administered for 2 consecutive weeks from the 6th week， and the other groups were continuously treated for 8 weeks before behavioral testing. The ultrastructure of synapses in hippocampal CA1 region was observed by transmission electron microscopy， and calcium/calmodulin-dependent kinase Ⅱ （CaMKⅡ） expression was detected by immunohistochemistry （IHC）. The levels of β-hydroxybutyrate （BHB） and lactic acid （LAC） were measured by biochemical kits， and those of glutamate （Glu）， glutamine （Gln）， γ-aminobutyric acid （GABA）， and synaptophysin （SYP） by enzyme-linked immunosorbent assay （ELISA）. The mRNA levels of glutamine synthetase （GS）， glutaminase （GLS）， glutamate decarboxylase （GAD）， CaMKⅡ， and AMPAR subunits GluA1 and GluA2 were quantified by Real-time quantitative polymerase chain reaction （Real-time PCR）， and the protein levels of GS， GLS， GAD， GluA1， and GluA2 by Western blot. ResultsCompared with the normal group， the model group exhibited decreased recognition index， spontaneous alternation rate， and residence time in the target quadrant （P<0.01）， increased escape latency and ratio of the time in the opposite side of the target quadrant to the time in the target quadrant （P<0.05， P<0.01）， damaged synaptic ultrastructure of hippocampal CA1 region， increased positive expression of CaMKⅡ， no significant difference in BHB level， elevated LAC and Glu levels （P<0.01）， declined SYP， Gln， and GABA levels （P<0.01）， downregulated mRNA and protein levels of GS， GAD， and GluA2 （P<0.05， P<0.01）， and upregulated mRNA and protein levels of GLS and GluA1 （P<0.05， P<0.01） and mRNA level of CaMKⅡ （P<0.01）. Compared with the model group， DSS intervention improved the behavioral indexes of mice （P<0.05， P<0.01）， enhanced the synaptic ultrastructure plasticity in hippocampal CA1 region， decreased the positive expression of CaMKⅡ， raised the BHB level （P<0.01）， reduced the LAC and Glu levels （P<0.01）， elevated the SYP， Gln， and GABA levels （P<0.01）， upregulated the mRNA and protein levels of GS， GAD， and GluA2， and downregulated the mRNA and protein levels of GLS and GluA1 （P<0.05， P<0.01） and the mRNA level of CaMKⅡ （P<0.01）. ConclusionDSS can improve the learning and memory and enhance the synaptic plasticity of APP/PS1 mice. Its neuroprotective effect is coupled with the promotion of ketone body formation and remodeling of brain energy metabolism， and is related to the regulation of glutamate/AMPAR pathway.

ObjectiveTo elucidate the critical role of rehabilitation medical records (including electronic records) in rehabilitation medicine's clinical practice and management, comprehensively analyzed the structure, core content and data standards of rehabilitation medical records, to develop a standardized medical record data architecture and core dataset suitable for rehabilitation medicine and to explore the application of rehabilitation data in performance evaluation and payment. MethodsBased on the regulatory documents Basic Specifications for Medical Record Writing and Basic Specifications for Electronic Medical Records (Trial) issued by National Health Commission of China, and referencing the World Health Organization (WHO) Family of International Classifications (WHO-FICs) classifications, International Classification of Diseases (ICD-10/ICD-11), International Classification of Functioning, Disability and Health (ICF), and International Classification of Health Interventions (ICHI Beta-3), this study constructed the data architecture, core content and data standards for rehabilitation medical records. Furthermore, it explored the application of rehabilitation record summary sheets (home page) data in rehabilitation medical statistics and payment methods, including Diagnosis-related Groups (DRG), Diagnosis-Intervention Packet (DIP) and Case Mix Index. ResultsThis study proposed a systematic standard framework for rehabilitation medical records, covering key components such as patient demographics, rehabilitation diagnosis, functional assessment, rehabilitation treatment prescriptions, progress evaluations and discharge summaries. The research analyzed the systematic application methods and data standards of ICD-10/ICD-11, ICF and ICHI Beta-3 in the fields of medical record terminology, coding and assessment. Constructing a standardized data structure and data standards for rehabilitation medical records can significantly improve the quality of data reporting based on the medical record summary sheet, thereby enhancing the quality control of rehabilitation services, effectively supporting the optimization of rehabilitation medical insurance payment mechanisms, and contributing to the establishment of rehabilitation medical performance evaluation and payment based on DRG and DIP. ConclusionStructured rehabilitation records and data standardization are crucial tools for quality control in rehabilitation. Systematically applying the three reference classifications of the WHO-FICs, and aligning with national medical record and electronic health record specifications, facilitate the development of a standardized rehabilitation record architecture and core dataset. Standardizing rehabilitation care pathways based on the ICF methodology, and developing ICF- and ICD-11-based rehabilitation assessment tools, auxiliary diagnostic and therapeutic systems, and supporting terminology and coding systems, can effectively enhance the quality of rehabilitation records and enable interoperability and sharing of rehabilitation data with other medical data, ultimately improving the quality and safety of rehabilitation services.

Objective:To present a set of clinically representative quality assurance (QA) test cases for stereotactic radiosurgery (SRT) plans of multiple intracranial metastases, in order to assess the plan quality and machine execution capabilities.Methods:Based on the clinical characteristics of multiple brain metastases, four groups of test cases with three target volumes (TVs), six TVs, nine TVs, and TVs near organs at risk (OARs) were designed. For these cases, SRT plans were developed, and plan quality was assessed using metrics including the Radiation Therapy Oncology Group conformality index (RTOG CI), gradient index (GI), homogeneity index (HI), and the volume of normal brain tissue receiving a dose of 24 Gy ( V24 Gy), which was defined as the volume enclosed by the 24 Gy isodose line around the Brain-PTV ( V24 Gy of Brain-PTV). Verification plans were generated for each test case, including the verification of point doses, planar doses (PD), and SRS MapCHECK (SMC) semiconductor matrix planar doses. Compared with the calculated result of the treatment planning system (TPS), the criteria for the γ analysis of planar doses were set at 1 mm/2% and 2 mm/2%. Results:For the four groups of test cases, the mean CI, GI, HI, and V24 Gy of Brain-PTV were 1.04±0.03, 3.79±0.40, 0.73±0.01 and (7.46±3.80) cm 3, respectively. The mean deviations of the point doses were 0.88%±0.98%, 1.47%±0.79%, 1.52%± 0.76%, and 1.17% ± 0.38%, respectively. The mean γ passing rates of the single fields for PDs were greater than 98% at 2 mm/2% and exceeding 96% at 1 mm/2%, and the mean γ pass rates of the SMC semiconductor matrix for PDs were 97.75% ± 2.31% and 99.33% ± 0.62%, at 1 mm/2% and 2 mm/2% respectively. Conclusions:The proposed QA test cases for SRT of multiple intracranial metastases allow for the effective assessments of the plan quality and machine execution capabilities and, thus, can assist various centers in clinical applications.

Objective:To investigate the clinicopathological features and differential diagnosis of male genital system lympho-ma(MGSL).Methods:We retrospectively analyzed the clinicopathological and immunophenotypic features and prognosis of 80 ca-ses of MGSL.Results:The onset age of the MGSL patients ranged from 4 to 85(median 62)years old.All the cases showed non-specificity of the imaging features and clinical manifestations.MGSL was located mainly in the testis(n=66),followed by the pros-tate(n=7),epididymis(n=3),scrotum(n=3)and penile glans(n=1).Diffused large B cell lymphoma(DLBCL)was the most common pathological type(n=62),next came extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue(MALT)(n=7)and other rare types(n=12).During the 1-112-month follow-up of 10 of the 19 patients,1 died at 1 month af-ter diagnosed with prostatic B-lymphoblastic lymphoma(B-LBL)and another 1 died at 50 months after diagnosed with testicular DLBCL.Conclusion:MGSL is rare clinically,mainly of the DLBCL type pathologically,lacking specificity in clinical symptoms and imaging manifestation.The definite diagnosis of the malignancy depends on histopathology combined with related molecular exami-nation and immunohistochemical labeling,and R-CHOP chemotherapy is the first choice for its treatment.

Objective To investigate the impact of risk prevention and control on the quality efficiency of medical equipment procurement.Methods The medical equipment procurement projects with enhanced risk prevention measures during 2023(119 items)were assigned to an observation group,while those without strengthened risk control during 2022(118 items)were enrolled into a control group.The two groups were compared in terms of bid failure/rejection rates,procurement completion cycles(including procurement execution cycle and total procurement cycle)and procurement satisfaction.Statistical analysis was performed using SPSS 20.0 software.Results The observation group demonstrated significantly lower bid failure/rejection rates when compared with the control group(≥2 occurrences:5.88％vs.15.25％,P＜0.05).The procurement execution cycle was notably shorter in the observation group((69.16±78.65)d vs.(97.67±49.84)d,P＜0.05),though no significant difference was observed in total procurement cycle(P＞0.05).The observation group behaved better significantly than the control group in procurement efficiency,equipment performance,procedural compliance and post-purchase service satisfaction(all P＜0.05).Conclusion Risk prevention and control measures in medical equipment procurement effectively reduce bid failure/rejection risks while enhancing procurement quality efficiency.[Chinese Medical Equipment Journal,2025,46(3):81-85]

Objective To elucidate the role of let-7 in the pathogenesis of oral squamous cell carcinoma(OSCC)and its underlying molecular mechanism.Methods The expression of let-7 in human oral keratinocyte(HOK),OSCC tissues,and OSCC cells(CAL-27,SCC-25,and SCC-9)was detected by quantitative real-time PCR.Colony formation,scratch,Transwell,and flow cytometry assays were used to examine the effects of let-7 on the proliferation,migration,invasion,and apoptosis of SCC-25 cells.Western blotting was used to detect the expression of Wnt and β-catenin proteins.The correlation between let-7 expression and the clinicopathological cha-racteristics and prognosis of OSCC patients was analyzed.Results let-7 was downregulated in OSCC tissues and cells.let-7 over-expression effectively inhibited the proliferation,migration,and invasion of SCC-25 cells while promoting apoptosis by suppressing acti-vation of the Wnt/β-catenin pathway.let-7 expression was closely associated with tumor stage,lymph node metastasis,and patient survival status.Patients with low let-7 expression exhibited lower overall survival rate.Conclusion let-7 targets and regulates the Wnt/β-catenin pathway,thereby inhibiting proliferation,migration,and invasion of OSCC cells while promoting apoptosis.

Objective:To establish a mouse model of liver cancer resistant to PD-1 monoclonal antibody and analyze the changes in its metabolomics to explore the potential mechanism of drug resistance.Methods:BALB/c mice were randomly divided into control and treatment groups after being loaded with tumor,and a normal group was additionally set up.The normal and control groups were injected with saline,and the treatment group was injected with PD-1 monoclonal antibody,after which the mice in the treatment group were screened for drug resistant and response groups.Observed the drug-resistant situation,body mass,tumor growth and survival rate of mice in each group,calculate the spleen index.The pathological features of tumor tissues were observed by HE staining method.Serum metabolites were detected by non-targeted metabolomics.Finally,a bivariate Pearson correlation analysis was conducted between the differential serum metabolites and tumor size.Results:The tumor-bearing mouse model with PD-1 monoclonal antibody resistance was successfully established,and the drug resistance rate of the mice was 50％.Compared with the normal and response groups,mice in the resistant group showed an increase in body weight,a significant increase in tumor volume,a decrease in survival rate,and a significant increase in splenic index.There was less lymphocyte infiltration in the tumor tissue.Metabolomics analysis showed that the serum levels of glutamic acid and aspartic acid increased and malic acid decreased in the resistant mice compared with the response group,and these changes were closely related to the arginine biosynthesis pathway.Conclusions:The tumor-bearing mouse model with PD-1 monoclonal antibody resistance was successfully established.The changes in its peripheral serum metabolomics mainly involve arginine metabolism and the related changes of aspartate,malate and glutamate.