Objective This study aims to achieve high-yield functional expression of the human voltage-gated potassium channel K_V3.1 using an Escherichia coli cell-free protein synthesis system, thereby providing a novel synthetic approach for drug screening, structural analysis and functional characterization of K_V3.1. Methods K_V3.1 was expressed in an Escherichia coli cell-free protein synthesis system for 10 hours in the presence of peptide surfactant A₆K. The secondary structure of K_V3.1 was analyzed by circular dichroism spectroscopy. The potassium channel activity of the recombinant protein liposome K_V3.1-A₆K was investigated using fluorescent dyes Oxonol VI as indicators, which are capable of reflecting alterations in membrane potential. Results Soluble K_V3.1 protein was successfully synthesized, achieving a purified yield of up to 1.2 mg/mL via an Escherichia coli cell-free protein synthesis system. Circular dichroism spectroscopy revealed that K_V3.1 exhibited characteristic α-helical secondary structures. Membrane potential fluorescence assays demonstrated that the K_V3.1-A₆K proteoliposomes, which were reconstructed with surfactant peptide A₆K, exhibited remarkable potassium ion permeability. Conclusion This study successfully achieved high-yield expression of human K_V3.1 with activity using an Escherichia coli-based cell-free protein synthesis system. This innovative method not only significantly enhances the expression yield of K_V3.1, but also maintains its functional activity, thereby establishing a novel and efficient synthetic platform for drug screening and advancing our understanding of structure-function relationships in K_V3.1 research.
Humans ; Escherichia coli/metabolism* ; Shaw Potassium Channels/biosynthesis* ; Cell-Free System ; Circular Dichroism ; Protein Biosynthesis ; Recombinant Proteins/metabolism* ; Membrane Potentials ; Shab Potassium Channels

[Objectives]To investigate the effect of METTL3 on the malignant biological behaviors of uveal melanoma cells and to verify whether this effect is related to m6A methylation.[Methods]Uveal melanoma cell models with METTL3 knockdown,overexpression,and point mutations at m6A-related catalytic sites were constructed via lentivirus transfection.Transwell assays were used to assess cell migration and invasion;CCK8 assays were used to measure cell proliferation and flow cytometry was used to analyze apoptosis and cycle changes.[Results]The proliferation,migration and invasion abilities of C918 and MUM-2B cells with METTL3 knockdown were significantly decreased(P<0.001),while the apoptosis rate was increased.The proportion of cells in G1 phase significantly increased,whereas the proportion in the S phase significantly decreased.Cells overexpressing METTL3 showed significantly enhanced proliferation,migration and invasion abilities(P<0.001),along with a decreased apoptosis rate.In C918 cells,the proportion of cells in G1 phase decreased significantly,while the proportion in S phase increased significantly.The cell cycle distribution of MUM-2B cells did not change remarkably.Following point mutation of m6A-related catalytic sites,cell proliferation,migration and invasion decreased,and the apoptosis rate increased.In MUM-2B cells,the percentage of cells in G1 phase significantly increased;the percentage in S phase significantly decreased and the percentage in G2 phase slightly decreased.In C918 cells,the percentage of G1 phase cells significantly increased,with no significant changes in the proportions of S and G2 phases.[Conclusions]The proliferation,invasion and metastasis of uveal melanoma cells were positively correlated with the expression of METTL3,while the apoptosis rate was negatively correlated.Changes in METTL3 levels differentially affect the cell cycles in different cell lines.The effects of METTL3 on the proliferation,migration and invasion of uveal melanoma cells are related to m6A methylation modification.

Objective:To investigate the use of non-vitamin K antagonist oral anticoagulants(NOACs)and their associated comorbidities in patients aged 80 years and older with non-valvular atrial fibrillation(NVAF), as well as to understand the challenges faced by elderly patients receiving NOAC therapy.Methods:We retrospectively enrolled elderly patients(≥80 years old)with NVAF who were treated with NOACs at a hospital in Beijing from January 2018 to August 2023.Patients were categorized into two age groups: 80-89 years and ≥90 years.We collected baseline data, including demographic characteristics, details of atrial fibrillation, comorbidities, laboratory test results, and medication combinations, for descriptive statistical analysis and intergroup comparisons.Results:A total of 695 elderly patients with NVAF receiving NOACs were included in the study, with a median age of 84 years.Among these patients, there were 328 males(47.19%, 328/695)and 422 cases of paroxysmal atrial fibrillation(60.72%, 422/695).The age group of 80-89 years comprised 640 cases(92.09%, 640/695), while the group aged 90 years and above included 55 cases(7.91%, 55/695).The use of NOACs in patients aged 90 and older exhibited an increasing trend over the years.Inter-group comparisons indicated that the ≥90 years group had lower body mass index, longer hospital stays, increased bedridden time, poorer renal function, lower levels of albumin and hemoglobin, and higher D-dimer levels.Inappropriate dosing of DOACs occurred in 49.64%(345/695)of cases, with 90.72%(313/345)receiving doses lower than recommended.Lower-than-recommended doses were more prevalent in the ≥90 years group, while higher-than-recommended doses were more common in the 80-89 years group.Polypharmacy was noted in 61.29%(426/695)of patients.The concurrent use of antiplatelet drugs, rhythm control medications, and ventricular rate control drugs was observed in 12.52%(87/695), 19.57%(136/695), and 54.53%(379/695)of patients, respectively, with no significant differences between groups.Conclusions:Inappropriate dosing and polypharmacy are prevalent issues among elderly NVAF patients.Therefore, it is essential to enhance multidisciplinary collaboration to optimize anticoagulation treatment strategies.

Organ damage from cancer treatment remarkably effects patients’ prognosis and quality of life. In recent years, preventive organ protection strategies, such as interdisciplinary collaboration, early prevention, precision interventions, psychological support, and the integrated application of traditional Chinese medicine, have demonstrated substantial clinical value and achieved notable progress. However, these approaches still encounter multiple challenges. Establishing multidisciplinary teams, optimizing therapeutic balance, and strengthening evidence-based research are essential for addressing the challenges related to treatment balance optimization, multidisciplinary coordination, and clinical translation of novel technologies. This review systematically summarizes recent advancements in preventive organ protection, analyzes existing challenges and potential solutions, and offers forward-looking recommendations. It aims to provide valuable insights for optimizing comprehensive cancer treatment strategies and improving long-term patient outcomes.

The intestinal microecology is closely related to the occurrence and development of hepatocellular carcinoma (HCC). The intestinal microbiota and its metabolites can regulate the tumor immune microenvironment through the "gut-liver axis", promoting cancer progression. Therefore, the intestinal microbiome is gradually demonstrating the potential as a biomarker for early diagnosis of HCC and prediction of the efficacy of immunotherapy. Targeted intervention on the intestinal microecology (such as probiotics, fecal microbiota transplantation, dietary regulation, etc.) may enhance the efficacy of immune checkpoint inhibitors (ICIs) and is becoming a promising combination therapy strategy. In the future, HCC treatment will rely on multi-omics integration, artificial intelligence-assisted diagnosis, and synthetic biology tools to promote the translation of precise gut flora intervention strategies from basic research to the clinic. This article summarized the latest research progress of intestinal microecology in HCC, explored its potential value and development direction for precision diagnosis and treatment of HCC, and provided a theoretical basis for the clinical application of related intervention strategies.

Objective:To analyze the reoperation rate and causes during the early adoption phase of unilateral biportal endoscopy (UBE).Methods:The clinical data of 180 patients who underwent UBE performed by a single surgeon at Beijing Friendship Hospital, Capital Medical University from October 2021 to June 2023 were retrospectively analyzed. Clinical and imaging data of patients who underwent reoperation were collected to analyze the causes of reoperation, and the clinical efficacy of the reoperations was also followed up. Measurement data were expressed as mean ± standard deviation ( ± s), and t-test was used before and after treatment. Results:A total of 180 patients who underwent UBE were included in this study, of which 6 patients underwent reoperation, and the reoperation rate was 3.33%. Among them, 3 cases occurred in the first 90 surgeries and the other 3 occurred in the subsequent 90 surgeries. The causes of reoperation were as follows: recurrent lumbar disc herniation at the same segment postoperatively in 2 cases, insufficient decompression in 2 cases, disc herniation following isolated decompression in 1 case, and immediate postoperative perianal numbness in 1 case. The time between the initial surgery and reoperation ranged from 0 to 187 days, with an average of 63.3 days. The average follow-up time after reoperation was 18.3 months. The visual analogue scale (VAS) and Oswestry disability index (ODI) scores of the patients at the last follow-up were significantly improved compared with those before operation (VAS score of low back pain: 5.2 ± 1.7 before operation, 1.2 ± 0.8 at the last follow-up, P<0.001; VAS score of leg pain: 7.2 ± 1.5 before operation, 1.2 ± 1.2 at the last follow-up, P<0.001; ODI score: 67.3 ± 5.7 before operation, 20.2 ± 8.2 at the last follow-up, P<0.001). The postoperative modified MacNab scores were generally satisfactory (4 cases were rated as excellent, accounting for 66.7%; 2 cases were rated as good, accounting for 33.3%). Except for one patient who experienced dural injury during open revision surgery, there were no serious complications such as nerve damage. Conclusions:In the early stages of UBE surgery, recurrent lumbar disc herniation and inadequate decompression are the primary reasons for reoperation, typically occurring within the first three months postoperatively. Reoperation does not significantly increase the risk of nerve injury. Enhanced early postoperative follow-up is recommended. For symptomatic patients, a second surgery with thorough decompression can yield satisfactory treatment outcomes.

One of the earliest hallmarks of intervertebral disc degeneration is the decrease in nucleus pulposus (NP) water content, which is fundamentally driven by the loss and structural alteration of extracellular matrix (ECM) aggrecan. At its core, the true determinant of NP degeneration lies in the imbalance between cells and the ECM. During the transition from notochordal cells to mature NP cells, reductions in cell density and matrix synthesis capacity lead to an inherently diminished ability to produce and renew aggrecan. Subsequently, inflammation, acidification, abnormal mechanical loading, and hypotonic microenvironments further disturb aggrecan’s structure, expression, and aggregation state through cellular mechanosensing and transcriptional regulation. These processes decrease the fixed charge density and osmotic pressure of the NP, thereby weakening its hydration, load-bearing, and resilience capacities. This review focuses on the NP microenvironment, using aggrecan as the central molecular anchor, to summarize recent advances in NP research. By integrating insights from microstructural alterations, imaging changes, and therapeutic strategies, this work aims to provide a comprehensive reference for understanding and managing the progression of NP degeneration.

Objective:To analyze the learning curve of TiRobot-assisted lumbar pedicle screw fixation (LPSF) by cumulative sum (CUSUM) test method.Methods:The clinical data of 50 patients who underwent TiRobot-assisted LPSF from January 2020 to December 2022 in Beijing Friendship Hospital, Capital Medical University were retrospectively analyzed. CUSUM analysis and learning curve fitting were performed with robot usage time as the main indicator with the time for each step refined (robot registration time, path planning time and guide wire placement time), to select the best learning curve fitting model with the R2 value closest to 1. Using the turning point of the learning curve as the boundary, the learning curve was divided into two stages as learning stage and maturity stage, and then the observation indexes were compared between the two stages. Results:All 50 patients successfully completed the surgery without perioperative complications, with a total of 244 pedicle screws implanted. The total robot usage time and robot registration time showed a gradually decreasing trend with the increase of case number, and the learning curves were successfully fitted and reached their peaks at the seventeenth and thirteenth cases respectively. The entire learning process was divided into learning stage (17 cases) and maturity stage (33 cases) based on the turning point of the learning curve of total robot usage time. The path planning time and guide wire placement time did not show significant changes with the increase in the case number. The total robot usage time, robot registration time and the intraoperative blood loss in the learning stage were significantly higher than those in the maturity stage: (35.35 ± 1.58) min vs. (30.61 ± 0.43) min, (20.83 ± 1.56) min vs. (14.94 ± 0.29) min and 400 (150, 500) ml vs. 200 (110, 300) ml, the guide wire placement time of per screw was significantly lower than that in the maturity stage: 2.00 (1.83, 2.34) min/screw vs. 2.33 (2.13, 2.69) min/screw, and there were statistical differences ( P<0.05 or <0.01). There were no statistical difference in the path planning time, path planning time of per screw, guide wire placement time and the accuracy of screw placement between two stages ( P>0.05). Conclusions:TiRobot-assisted LPSF is a new technology with safety and effectiveness, and it has a relatively short learning curve. To achieve technological maturity, at least 17 surgeries are required with accumulated experience, and the robot registration is the main step of the learning process. After reaching maturity stage, the robot usage time is significantly shortened and intraoperative trauma is significantly reduced while the relatively high screw placement accuracy is ensured.

Objective To quantitatively assess the correlation between the skeletal muscle index(SMI)of patients and the occur-rence of anastomotic leakage(AL)in rectal cancer patients after surgery,and to analyze the risk factors for AL in rectal cancer patients and the influencing factors of sarcopenia.Methods The clinical,pathological,and related imaging data of 362 patients who under-went radical surgery for rectal cancer were retrospectively analyzed.All patients underwent pelvic MRI and abdominal CT scans(plain/enhanced)within one month before surgery,and the third lumbar vertebra skeletal muscle area(L3-SMA)was measured from the images.All patients were divided into AL group(56 cases)and control group(306 cases)based on the presence or absence of postoperative complications.The differences in clinical characteristics and imaging parameters between the two groups were analyzed.A logistic risk prediction model was established.Results Significant differences were observed between the two groups in sarcopenia,type of surgery,surgical approach,serum albumin level,operation duration,stoma type,and extramural vascular invasion(EMVI)(P＜0.05).These factors were incorporated in a multivariate logistic regression analysis model,the area under the curve(AUC)of receiver operating characteristic(ROC)curve of the model was 0.810[95％confidence interval(CI)0.743-0.876,P＜0.001],with a sensitivity of 0.865 and specificity of 0.669.Conclusion Sar-copenia is a significant risk factor for AL after rectal cancer surgery.It enhances the predictive efficacy for postoperative AL and serves as a basis for identifying high-risk populations for AL in clinical practice.