Journal Of Visualized Experiments[JOURNAL]
Sun
200 papers
A Video Protocol of a Randomized Controlled Clinical Trial - Electrochemotherapy of Cutaneous Metastases with Reduced Dose Bleomycin (BLESS Trial).
Tolstrup MA, Kjaer Lonkvist C, Emilie Rosted E … +8 more , Abdulmohsen M Alrasheed T, Armstrong Bastrup F, Mojsoska B, Plesnik H, Kosjek T, Rosenkrantz Hölmich L, Krezdorn N, Gehl J
J Vis Exp · 2026 Jun · PMID 42372073 · Publisher ↗
Cutaneous metastases in patients with incurable cancer represent a significant problem as they often cause pain, discomfort, and emotional distress that affect everyday life. Finding treatment options that are both effec...
Cutaneous metastases in patients with incurable cancer represent a significant problem as they often cause pain, discomfort, and emotional distress that affect everyday life. Finding treatment options that are both effective and gentle is essential. ECT offers one such possibility. Here, short, high-voltage electrical pulses are applied directly to the tumor, briefly opening tumor cells, allowing chemotherapy to enter more effectively and kill cancer cells. Traditionally, patients receive 15.000 IU/m2 of bleomycin intravenously, but emerging evidence suggests that a lower dose may be just as effective while causing fewer side effects. This protocol describes an ongoing double-blinded, randomized clinical trial that tests whether ECT with half the standard bleomycin dose is non-inferior to the conventional regimen for tumor control in patients with cutaneous metastases. The article outlines randomization and blinding procedures, pretreatment evaluation, bleomycin preparation and administration, electrode placement, pulse delivery, and response evaluation using the modified Response Evaluation Criteria in Solid Tumors (mRECIST) criteria. In addition to clinical response at three months, the protocol includes pharmacokinetic blood sampling and qualitative interviews with the patients to enable a comprehensive evaluation of treatment impact. Baseline tumor characteristics from the first 15 enrolled patients and an example of how mRECIST is applied are presented. Critical steps to ensure methodological rigor are discussed, including standardized tumor measurements, consistent electrode positioning, and predefined management of confluent or poorly demarcated tumors. By visually outlining the procedural workflow and key methodological considerations, this article provides a reproducible framework for dose optimization in ECT. It supports future implementation of reduced-dose regimens in clinical oncology practice.
A Standardized Ex Vivo Porcine Oromucosal Model for Evaluating Peptide Fluxes.
Ijaz M, Karki S, Malhotra S … +3 more , Bini EI, O'Cearbhaill ED, Brayden DJ
J Vis Exp · 2026 Jun · PMID 42372070 · Publisher ↗
Oromucosal administration of drugs in films, tablets, lozenges, and other dosage forms provides a convenient systemic administration route for pediatric and elderly patients, and for patients who cannot swallow capsules...
Oromucosal administration of drugs in films, tablets, lozenges, and other dosage forms provides a convenient systemic administration route for pediatric and elderly patients, and for patients who cannot swallow capsules or tablets easily. Developing buccal and sub-lingual peptide formulations requires preclinical models that align with the non-keratinized oromucosal epithelial structures of humans, of which pigs and dogs are the most suitable. We sourced discarded fresh porcine oromucosal tissues from an abattoir. The mucosae were dissected from the underlying smooth muscle using controlled heating and then mounted in either Franz Diffusion Cells or in novel low-volume 3D-printed resin-cured inserts. Peptide-permeation enhancer combinations were added to the donor side, and permeated molecules were sampled from the receiver side and quantified by ultra-high performance liquid chromatography (uHPLC). For the inserts, transepithelial electrical resistance (TEER) values of mounted mucosae were measured using EVOM chopstick electrodes. Tissue histology was examined following incubation in both Franz diffusion cells and 3D inserts. The optimal concentration of permeation enhancer was selected when aqueous mixtures of peptide and enhancer produced an increase in permeation. Peptide permeation reached 4.3% over 3 h in three-dimensional (3D) inserts and 1.7% in Hilltop chamber models, the latter simulating direct exposure from a buccal film. Layer-by-layer polymeric mucoadhesive films containing peptide-enhancer combinations were developed and applied to mucosal tissue for permeation studies using diffusion chambers. Ex vivo peptide permeation across porcine buccal mucosae reached 2% when mucosae were exposed to the optimal combination of glycodeoxycholate (GDC) incorporated into the buccal films. These ex vivo screening studies provide a pathway to subsequent pharmacokinetic evaluation of peptide-loaded film formulations administered via the oromucosal route in pigs and people.
Lightweight English Text Classification with Deep Learning Based on Complex System Theory.
Ma C, Guo Z
J Vis Exp · 2026 Jun · PMID 42372063 · Publisher ↗
Globalization and the development of information technology have driven a surge in English text data, and there is an urgent need for efficient classification. To improve the generalization ability of English text classi...
Globalization and the development of information technology have driven a surge in English text data, and there is an urgent need for efficient classification. To improve the generalization ability of English text classification in small-sample and cross-domain scenarios and to achieve lightweight models, this study combines complex systems theory with deep learning to construct a graph neural network model that fully accounts for the distributional characteristics of text samples. A two-level meta-distillation method, combined with meta-learning strategies, dynamically adjusts the teacher model's parameters and optimizes the entire knowledge-transfer process. The experimental results show that the classification performance of the proposed model in few-shot and cross-domain text classification tasks is significantly superior to that of traditional graph neural networks and other mainstream comparative models. In terms of lightweighting, the SM generated by the two-stage meta-distillation mechanism maintains an extremely high level of performance retention on multi-topic text classification datasets, while significantly reducing computational time. In addition, this method can maintain high efficiency and stable classification performance in long text processing scenarios and offers clear advantages in computational efficiency compared with traditional distillation methods and other methods reported in the literature. The proposed method effectively enhances the classification performance of English text in low-sample and cross-domain application scenarios while significantly reducing the computational cost, thus providing a feasible and efficient technical solution for English text classification in resource-constrained environments.
Integrating Artificial Intelligence-Assisted Translation Support into English Courses: Effects on Translation Accuracy, Perceived Stress, and Anxiety.
Luo R
J Vis Exp · 2026 Jun · PMID 42372062 · Publisher ↗
English translation competence is an important academic skill in higher education, yet many students experience substantial linguistic difficulty, cognitive burden, and psychological pressure during translation tasks. Th...
English translation competence is an important academic skill in higher education, yet many students experience substantial linguistic difficulty, cognitive burden, and psychological pressure during translation tasks. This mixed-methods study examined whether structured artificial intelligence-assisted translation support integrated into English courses was associated with changes in translation accuracy, perceived stress, and anxiety among university students. A total of 525 undergraduate and postgraduate students participated in an eight-week instructional intervention. Baseline and post-intervention assessments included a translation accuracy test, the Perceived Stress Scale, and the Generalized Anxiety Disorder-7 scale. Quantitative analysis was conducted using paired comparisons, correlation analysis, regression analysis, and group-based comparisons, and was complemented by qualitative interviews to explore students' learning experiences. After the intervention, participants showed improved translation accuracy and lower levels of perceived stress and anxiety. Higher frequency of tool use was associated with better translation performance, although the study design does not support strong causal inference. Qualitative findings further indicated that structured tool-assisted practice helped reduce cognitive overload, supported revision processes, and improved engagement with translation tasks. These findings suggest that integrating artificial intelligence-assisted translation support into English courses may offer a practical instructional approach for strengthening translation performance while supporting student well-being. The study contributes empirical evidence to current discussions of technology-enhanced translation pedagogy and provides an adaptable teaching framework for higher education contexts.
A Toxin-Based Counter-Selection System for Markerless Gene Deletion and High-Density Tn5 Transposon Mutagenesis in Pectobacterium brasiliense.
Xie B, Xie M, Handique U … +5 more , Zhang Y, Teper D, Li L, Wu J, Zhou X
J Vis Exp · 2026 Jun · PMID 42372061 · Publisher ↗
Pectobacterium brasiliense is a highly pathogenic bacterium responsible for potato tuber soft rot, blackleg, and aerial stem rot. Elucidation of its pathogenic mechanisms is essential for disease control; however, geneti...
Pectobacterium brasiliense is a highly pathogenic bacterium responsible for potato tuber soft rot, blackleg, and aerial stem rot. Elucidation of its pathogenic mechanisms is essential for disease control; however, genetic tools for this organism remain limited. A toxin-based counter-selection system was developed to enable markerless gene deletion and high-density Tn5 transposon mutagenesis. The vmi480 toxin derived from the pLP12 plasmid was used as a counter-selection marker to construct the suicide vector pKV2. Using this system, the recA gene in the P. brasiliense SM strain was successfully deleted. The resulting recA mutant served as the parental strain for the construction of a Tn5 insertion library comprising over 12,000 mutants through conjugation with the RHO3 helper strain carrying pKV2-LLtnp. In this derivative plasmid, the vmi480 toxin was replaced with a hyperactive Tn5 transposase under the control of an outward-facing lac promoter to reduce polar effects. High-throughput tuber inoculation assays enabled the identification of pathogenicity-deficient mutants, including insertions in the type II secretion system, a key virulence determinant that mediates the secretion of plant cell wall-degrading enzymes. This platform supports functional genomics and facilitates the investigation of virulence mechanisms in this economically important pathogen.
Seamless Multimodal Human-Robot Communication: Integration Techniques in Human-Computer Interaction.
Zheng W, Gao Y
J Vis Exp · 2026 Jun · PMID 42372057 · Publisher ↗
Seamless multimodal human-robot communication has become essential as social, assistive, and educational robots move into everyday settings like homes, healthcare facilities, and classrooms, where they need to integrate...
Seamless multimodal human-robot communication has become essential as social, assistive, and educational robots move into everyday settings like homes, healthcare facilities, and classrooms, where they need to integrate speech, gesture, gaze, facial expressions, and tactile cues with high precision, low latency, and real-world robustness. This review systematically examines how current techniques achieve real-time, reciprocal multimodal human-robot interaction (HRI), focusing on fusion strategies, system architectures, and applications in specific domains. We searched the Web of Science Core Collection for English-language empirical studies from 2015 to 2025, selecting 148 papers that address communicative integration. The most important insights include a clear shift toward hybrid and attention-based fusion since 2022 (about 43% of approaches), which better handles temporal asynchrony and embodied interactions than earlier methods; widespread inconsistency in evaluation metrics that hinders cross-study comparisons; and a persistent gap between strong laboratory performance and weaker real-world robustness under noise, occlusion, or user variability. Key challenges include real-time processing, semantic alignment, data efficiency, deployment robustness, and the under-explored integration of tactile signals with affect. Looking ahead, the review suggests prioritizing adaptive fusion policies, standardized benchmarks for synchronization and fluency, and continual learning to enable user-personalized adaptation. Ultimately, it aims to guide the development of more human-centered robotic agents that can engage collaboratively, meaningfully, and are socially acceptable in daily life.
A Randomized Controlled Trial of Auricular Acupressure Combined with Sodium Hyaluronate for Abnormal Blink Reflex in Children.
Wang J, Yang G, Yu N … +4 more , Liao W, Zhang X, Zhou D, Liu P
J Vis Exp · 2026 Jun · PMID 42372056 · Publisher ↗
Abnormal blink reflex in children is a common ocular symptom with complex causes, including ocular surface diseases like dry eye and inflammation, as well as neuromuscular dysfunction. Current treatments show limited eff...
Abnormal blink reflex in children is a common ocular symptom with complex causes, including ocular surface diseases like dry eye and inflammation, as well as neuromuscular dysfunction. Current treatments show limited efficacy, highlighting the need for improved therapeutic options. This study evaluates the clinical efficacy and safety of auricular acupressure combined with sodium hyaluronate eye drops. A randomized controlled design included 97 children aged 3-10 years with abnormal blink reflex. Participants were randomly assigned to three groups: the control group (Group A, n = 32), treated with sodium hyaluronate eye drops; the experimental group (Group B, n = 33), receiving eye drops combined with auricular acupressure using Vaccaria seed patches; and the sham stimulation group (Group C, n = 32), receiving ear patches without Vaccaria seed. The treatment lasted for three cycles. The primary outcome was the blink rate, and secondary outcomes included tear film break-up time (FBUT), corneal fluorescein staining (FL) score, Visual Analog Scale (VAS) scores for ocular dryness and photophobia, clinical efficacy, and adverse events. Compared with Groups A and C, Group B showed a significant reduction in blink rate (P<0.05), a significant increase in FBUT (P<0.05), a notable decrease in FL score (P<0.05), and a significant reduction in VAS scores for ocular dryness and photophobia (P<0.05). No statistical differences in clinical symptoms and signs were observed between Groups A and C (P>0.05). The total effective rate was 78.13% for Group A, 93.94% for Group B, and 84.38% for Group C, with Group B showing significantly higher efficacy. No serious adverse events were observed in any group. Auricular acupressure with Vaccaria seed improves the clinical symptoms of abnormal blink reflex in children, enhances tear film stability, and provides corneal protection. It is a safe and effective new intervention for the treatment of abnormal blink reflex in children.
Standardized Protocol to Evaluate the Effect of Surface Treatments, Luting Cements, And Thermocycling On PEEK-Composite Bond Strength.
Cigdem KZ, Idil D, Erkul S
J Vis Exp · 2026 Jun · PMID 42372055 · Publisher ↗
Polyetheretherketone (PEEK) has gained increasing attention as a biomaterial in prosthodontics; however, achieving durable adhesion to veneering composite materials remains a major challenge due to its chemically inert s...
Polyetheretherketone (PEEK) has gained increasing attention as a biomaterial in prosthodontics; however, achieving durable adhesion to veneering composite materials remains a major challenge due to its chemically inert structure. This study presents and validates a standardized, reproducible protocol for evaluating shear bond strength (SBS) between PEEK and an indirect composite resin under different surface treatments, luting cements, and thermocycling conditions. A total of 240 PEEK discs were divided into four groups according to surface treatment: no surface treatment, sandblasting, sulfuric acid etching, and laser irradiation. Surface morphology was analyzed using atomic force microscopy. Each group was then subdivided according to the luting cement used (zinc oxide non-eugenol cement or self-adhesive resin cement; n = 30), and further subdivided into thermocycled and non-thermocycled groups (n = 15). SBS was measured using a universal testing machine, and failure modes were analyzed under a stereomicroscope. Among all experimental groups, specimens luted with self-adhesive resin cement and not subjected to thermocycling demonstrated the highest SBS values (25.134 ± 1.665 MPa), whereas specimens luted with zinc oxide non-eugenol cement and subjected to thermocycling exhibited the lowest SBS values (1.958 ± 0.345 MPa). For all surface treatment protocols, SBS values were significantly higher in specimens luted with self-adhesive resin cement compared with zinc oxide non-eugenol cement (p < 0.001). Thermocycling significantly reduced SBS values across all groups (p < 0.001). This protocol highlights critical procedural steps, including surface treatment selection and cement type, that significantly influence bonding outcomes. The standardized workflow and visual demonstration of key steps provide a reproducible framework for evaluating PEEK-composite bonding.
Real-Time Label-Free Imaging and Quantitative Analysis of Macrophage Morphodynamics Using Optical Diffraction Tomography.
Sun C, Zhu JJ, Yang YQ … +3 more , Liu SY, Shi KB, Peng C
J Vis Exp · 2026 Jun · PMID 42372054 · Publisher ↗
This study establishes a label-free optical diffraction tomography (ODT)-based workflow for live-cell imaging and analysis to observe time-dependent morphological changes in macrophages. The method enables continuous rec...
This study establishes a label-free optical diffraction tomography (ODT)-based workflow for live-cell imaging and analysis to observe time-dependent morphological changes in macrophages. The method enables continuous recording of single-cell morphology and movement over extended periods under stable environmental conditions and allows extraction of quantitative parameters, including projected area, perimeter, and average migration speed. This workflow provides a practical approach for capturing dynamic cellular behaviors at the single-cell level without exogenous labeling. Using RAW264.7 macrophages as a model, time-lapse imaging was performed under lipopolysaccharide stimulation with baicalin pretreatment to capture dynamic cellular changes under different conditions. Representative cells were selected for tracking and quantitative analysis. The results show that this workflow supports stable long-term single-cell tracking and reflects temporal changes in cell morphology and motility. This approach provides a label-free method for observing dynamic cellular behaviors in response to different stimuli and can serve as a useful complement to conventional endpoint-based assays. It may also be applicable to other adherent cell types for studies of cell morphodynamics.
Development of a Rat Caudal Interbody Fusion Model: A Pilot Feasibility Study.
Ikegami T, Ushiku C, Saito M
J Vis Exp · 2026 Jun · PMID 42372053 · Publisher ↗
Rat caudal vertebrae are widely used in bone regeneration and spinal fusion research as convenient sites for testing bone graft materials and implants. Prior rat tail fusion models have achieved intervertebral fusion thr...
Rat caudal vertebrae are widely used in bone regeneration and spinal fusion research as convenient sites for testing bone graft materials and implants. Prior rat tail fusion models have achieved intervertebral fusion through disc removal and bone grafting, or with external fixation devices. Nonetheless, no such small-animal model has been developed that uses an interbody fusion cage analogous to that used clinically. It was hypothesized that an evaluable bone fusion environment could be established without neurological complications by performing a posterior cage insertion mimicking clinical procedures, supplemented with plate fixation, in the rat caudal vertebrae. This preliminary pilot study (n = 1) was designed to evaluate technical feasibility. A surgical approach was developed in which a miniaturized titanium cage was precisely inserted into the resected disc space between the adjacent caudal vertebrae, closely mirroring the human surgical procedure. Titanium was selected for the cage owing to its widespread clinical use and biocompatibility. The initial results demonstrated the technical feasibility of this approach. The implanted cage maintained disc height and spinal alignment. Progressive bone ingrowth through the cage was observed, culminating in confirmed continuous bone bridging across the intervertebral space by 12 weeks postoperatively. The presence of continuous trabecular structure and the absence of intervening fibrous tissue were confirmed radiologically and histologically. Overall, this cage-based rat tail fusion model provides a promising platform for evaluating novel biomaterials and investigating mechanisms of intervertebral bone fusion. However, the lack of a control group and statistical analysis, due to the single-subject design, limits the generalizability of these findings and necessitates further validation.
Construction and Validation of A Nomogram to Identify Mucus Obstruction In Patients With Chronic Obstructive Pulmonary Disease.
Su W, Zhang Y, Yang Z … +5 more , Zhou Y, Gao B, Shi R, Wu L, Huang H
J Vis Exp · 2026 Jun · PMID 42372050 · Publisher ↗
Small airway mucus impaction in chest computed tomography (CT) is a clinically significant finding in chronic obstructive pulmonary disease (COPD), associated with accelerated pulmonary function decline, increased freque...
Small airway mucus impaction in chest computed tomography (CT) is a clinically significant finding in chronic obstructive pulmonary disease (COPD), associated with accelerated pulmonary function decline, increased frequency of acute exacerbations, and higher susceptibility to respiratory infections. However, a validated predictive tool for identifying patients at risk of CT-detected mucus plugs is currently lacking. This study aimed to develop and validate a nomogram to predict small airway mucus obstruction in patients with COPD. We retrospectively enrolled 212 COPD patients from Shenzhen Second People's Hospital (January 2021 to June 2022), of whom 47 had CT-confirmed mucus plugs (mucus plug group, MP) and 165 did not (non-mucus plug group, NMP). Univariate and receiver operating characteristic (ROC) analyses were used to identify candidate predictors. Multivariate logistic regression was conducted to construct the final predictive model, which was then transformed into a nomogram. Internal validation was performed using bootstrap sampling (1000 iterations). Bronchiectasis, chronic rhinosinusitis (CRS), body mass index (BMI), forced expiratory flow at 25-75% of predicted (FEF25-75%pred), residual volume-to-total lung capacity ratio (RV/TLC), and serum 25-hydroxyvitamin D [25(OH)D] were identified as independent risk factors for CT mucus plugs. The nomogram demonstrated excellent predictive value with an AUC of 0.9611. Calibration curves and decision curve analyses demonstrated good clinical utility. Bootstrap internal validation further supported the model's predictive stability. This nomogram provides a practical, individualized tool to facilitate early identification and personalized management of COPD patients at risk of small-airway mucus obstruction.
Decoding The Epitranscriptome: In Silico Insights Into m6A Regulatory Network In Breast Cancer.
Abdulkarim S, Akhtar S, Ur Rehman M … +3 more , Abu-Zaid A, Zhang D, Mehmood R
J Vis Exp · 2026 Jun · PMID 42372047 · Publisher ↗
N6-methyladenosine (m6A) is the most abundant internal RNA modification in eukaryotic transcripts and plays a critical role in RNA metabolism, gene expression, and cellular homeostasis. Dysregulation of m6A regulators, i...
N6-methyladenosine (m6A) is the most abundant internal RNA modification in eukaryotic transcripts and plays a critical role in RNA metabolism, gene expression, and cellular homeostasis. Dysregulation of m6A regulators, including "writers," "erasers," and "readers", has been increasingly implicated in cancer biology; however, their comprehensive roles in breast cancer remain to be understood. The primary objective of this methods article is to provide bioinformatics beginners with a step-by-step framework for utilizing publicly available cancer datasets to perform mutational analyses, assess gene expression alterations, and examine their associations with patient survival. As a case study, m6A regulators in breast cancer were analyzed using datasets from the Cancer Genome Atlas (TCGA), the Genotype-Tissue Expression (GTEx) project, and microarray platforms. Transcriptomic profiles were systematically analyzed to demonstrate workflows for evaluating the prognostic relevance of m6A regulatory components in breast cancer. Using this analytical framework, distinct patterns of genetic alterations and differential expression among key m6A regulators were identified. Several regulators, including METTL14, CBLL1, YTHDC1, HNRNPC, HNRNPA2B1, and RBMX, were associated with better patient survival, while YWHAG was associated with poor overall survival. This study provides a comprehensive systems genomics overview of m6A regulatory genes in breast cancer while demonstrating a practical and reproducible web-based bioinformatics workflow. These findings advance the understanding of epitranscriptomic regulation in breast cancer and offer a foundation for the development of novel m6A-based diagnostic and therapeutic strategies.
Measuring Inspiratory Effort In Mechanically Ventilated Children.
Vandebergh V, Feder J, Reise K … +6 more , Evans R, Shurdha E, Nallasamy K, Kushal UD, Guerineau LR, Schepens T
J Vis Exp · 2026 Jun · PMID 42372045 · Publisher ↗
An accurate assessment of inspiratory effort in mechanically ventilated children is essential for optimizing ventilator support and avoiding lung- and diaphragmatic injury. Over-assistance leads to diaphragmatic disuse a...
An accurate assessment of inspiratory effort in mechanically ventilated children is essential for optimizing ventilator support and avoiding lung- and diaphragmatic injury. Over-assistance leads to diaphragmatic disuse and atrophy, whereas under-assistance may result in fatigue, diaphragm weakness and patient self-inflicted lung injury (P-SILI). This manuscript demonstrates validated bedside techniques to quantify inspiratory effort. We describe the reference standard for respiratory muscle effort quantification, esophageal manometry, and present alternative approaches for routine clinical practice, given its practical limitations. We distinguish between respiratory drive, tidal inspiratory effort, and maximal inspiratory effort. Indirect parameters of effort include diaphragm electrical activity (EAdi), airway occlusion maneuvers (P0.1, Pocc, PMI) and diaphragm ultrasound parameters. The maximal inspiratory pressure (MIP) is discussed as a measure of maximal effort. Representative pediatric data are provided where available. Although reference values are included for selected parameters, evidence defining safe thresholds in children remains limited. Combined monitoring using these techniques allows individualized titration of mechanical ventilation and supports lung- and diaphragm-protective strategies in the pediatric ICU.
Identification of Hub Genes, Single-Nucleotide Polymorphisms, and Potential Drug Targets In Breast Cancer Using Transcriptomic Analysis.
Jia X, Su H, Zhang J … +1 more , Liu Z
J Vis Exp · 2026 Jun · PMID 42372043 · Publisher ↗
HER2-positive (HER2) breast cancer often develops resistance to therapies like Lapatinib, potentially involving mitochondrial metabolic and redox reprogramming. This study aimed to identify mitochondrial oxidative stress...
HER2-positive (HER2) breast cancer often develops resistance to therapies like Lapatinib, potentially involving mitochondrial metabolic and redox reprogramming. This study aimed to identify mitochondrial oxidative stress-related genes (MOS-DEGs) as biomarkers of resistance and to characterize their functional nsSNPs and structural impacts. RNA-seq datasets (GSE231524, GSE231525) were analyzed using DESeq2 to identify DEGs, which were intersected with mitochondrial oxidative stress genes to obtain MOS-DEGs. Functional enrichment, PPI analysis, ROC analysis, expression profiling, and survival analysis were performed. nsSNPs were evaluated using multiple predictive tools, and structural impacts were assessed through secondary and 3D modeling. Integrated transcriptomic and clinical analysis identified MTHFD2 and PRDX3 as central MOS-DEGs displaying opposing regulatory roles in mitochondrial redox metabolism. MTHFD2 was significantly upregulated and associated with poor prognosis (HR = 1.53, p = 1.1 × 10⁻), while PRDX3 exhibited protective expression patterns (HR = 0.73, p = 7.7×10⁻). ROC analysis suggested their potential as predictors of therapeutic response. nsSNP analysis revealed five deleterious variants in MTHFD2 and four deleterious variants in PRDX3, with rs1471336772 (MTHFD2) and rs747786383 (PRDX3.) identified as the most pathogenic. These variants were predicted to be damaging based on multiple computational scoring systems, including SIFT ≤ 0.05, PolyPhen-2 ≥ 0.85, deleterious CADD scores, and high REVEL scores, and were located within catalytic or redox-active domains. Structural modeling suggested that these substitutions may destabilize conformation, disrupt metal- and cofactor-binding sites, and affect NADPH regeneration or thioredoxin-dependent peroxidase activity. Molecular dynamics predictions indicated potential loss of structural stability and altered flexibility, suggesting possible functional impairment of mitochondrial redox control. This study identifies MTHFD2 and PRDX3 as regulators of mitochondrial oxidative stress in HER2 breast cancer. Deleterious nsSNPs in these genes may contribute to altered redox balance, potentially influencing metabolic adaptation (MTHFD2) and antioxidant defense (PRDX3).
Changes In Lower Respiratory Tract Bacterial Isolates In ICU Patients Before and After the COVID-19 Pandemic.
Ulger M, Kurtulus S, Can R … +4 more , Yakut S, Celik M, Kalaycioglu H, Zeyrek FY
J Vis Exp · 2026 Jun · PMID 42372042 · Publisher ↗
To compare the distribution of bacterial pathogens isolated from lower respiratory tract specimens in COVID-19-negative intensive care unit patients before and after the COVID-19 pandemic, culture results from patients a...
To compare the distribution of bacterial pathogens isolated from lower respiratory tract specimens in COVID-19-negative intensive care unit patients before and after the COVID-19 pandemic, culture results from patients admitted to a university hospital chest diseases intensive care unit were retrospectively analyzed. Patients without clinical or radiological suspicion of COVID-19 and with negative PCR results were included and classified as pre-COVID-19 or post-COVID-19 based on March 2020. Specimens were cultured on 5% sheep blood agar, eosin methylene blue agar, and chocolate agar, and evaluated after 24-48 h incubation at 37 °C. Only dominant microorganisms demonstrating growth in at least the second streaking sector were considered clinically significant. A total of 1,665 patients were included, with microbial growth detected in 48.9% of samples, the majority originating from the pre-COVID-19 period. Acinetobacter baumannii was the most frequently isolated organism in both periods. Logistic regression analysis showed reduced detection rates of Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus (odds ratio ≈0.3), and Escherichia coli (odds ratio ≈0.4) in the post-COVID-19 period. Mortality rates were also lower in the post-COVID-19 period, although no significant difference in discharge type was observed. Overall pathogen diversity remained similar across periods; however, decreases in selected bacterial detections were observed following the onset of the pandemic. These findings may reflect changes in infection control practices during the pandemic period, although causality cannot be established due to the retrospective study design and absence of direct measurements of contributing factors.
Standardized Flow Cytometry Assay Using Dried Recombinant Antibody Panels for CAR T Cell Characterization.
Ridzal A, Brauchle A, Missing D … +5 more , Hudecek M, Mues M, Luu M, Quintarelli C, Wilmes S
J Vis Exp · 2026 Jun · PMID 42372040 · Publisher ↗
Flow cytometry is commonly used to assess critical quality attributes of Chimeric Antigen Receptor T (CAR T) cells, including viability, phenotype, and transduction efficiency. Conventional liquid reagent workflows requi...
Flow cytometry is commonly used to assess critical quality attributes of Chimeric Antigen Receptor T (CAR T) cells, including viability, phenotype, and transduction efficiency. Conventional liquid reagent workflows require multiple preparation steps, which can introduce variability and differences in operator-dependent handling. Preformulated dried recombinant antibody panels provide an alternative format that combines recombinant antibody technology with a ready-to-use configuration. Recombinant antibodies offer defined specificity and reduced lot-to-lot variability, while the dried format reduces preparation steps and simplifies reagent handling. Here, we present a step-by-step workflow for CAR T cell analysis using dried recombinant antibody panels. The protocol includes sample staining, data acquisition, and analysis, and is accompanied by a schematic overview of the workflow. Representative staining results are shown to illustrate panel application, and differences in hands-on time compared to a conventional liquid reagent workflow are highlighted. The examples provided demonstrate how these panels can be used to streamline the multiparametric characterization of CAR T cells. Overall, this work provides a practical framework for implementing dried recombinant antibody panels in flow cytometry-based CAR T cell analysis.
Current Incidence and Risk Factors for Polymyxin B-Associated Acute Kidney Injury In Patients With Septic Shock.
Teng Y, Ma L, Li Q
J Vis Exp · 2026 Jun · PMID 42372039 · Publisher ↗
This study evaluates the incidence and risk factors of polymyxin B-associated acute kidney injury (AKI) in patients with septic shock, a population at high risk of nephrotoxicity due to critical illness and exposure to l...
This study evaluates the incidence and risk factors of polymyxin B-associated acute kidney injury (AKI) in patients with septic shock, a population at high risk of nephrotoxicity due to critical illness and exposure to last-line antibiotics. A prospective cohort of 200 adult patients with microbiologically confirmed multidrug-resistant gram-negative infections was analyzed. All patients received intravenous polymyxin B with standardized dosing and renal monitoring based on Kidney Disease Improving Global Outcomes (KDIGO) criteria. AKI occurred in 25% of patients, with a median onset of 4 days following initiation of therapy. Significant risk factors included higher baseline body mass index, elevated serum urea and creatinine, reduced glomerular filtration rate, presence of comorbidities, and concomitant use of nephrotoxic agents. Structured nursing interventions and standardized monitoring enabled early detection of renal dysfunction and timely dose adjustment. These findings highlight the importance of systematic renal surveillance and individualized therapeutic strategies, including therapeutic drug monitoring, to minimize nephrotoxicity in septic shock patients receiving polymyxin B.
Physical Activity and Perceived Barriers In Individuals With Spinal Cord Injury In Saudi Arabia: A Cross-Sectional Study.
Almutairi SM, Alnaim LH, Alobaysi JI … +1 more , Basha MA
J Vis Exp · 2026 Jun · PMID 42372035 · Publisher ↗
Spinal cord injury (SCI) impairs physical function and quality of life (QoL), and physical activity is important for health maintenance; however, participation may be influenced by multiple barriers. This cross-sectional...
Spinal cord injury (SCI) impairs physical function and quality of life (QoL), and physical activity is important for health maintenance; however, participation may be influenced by multiple barriers. This cross-sectional study (May-September 2024) recruited 50 individuals with SCI in Saudi Arabia using convenience sampling via outpatient clinics and online distribution. Data were collected through structured interviews or self-administered surveys using identical questionnaires, including demographics, lesion characteristics, physical activity levels using the Arabic Physical Activity Scale for Individuals with Physical Disabilities (PASIPD-AR), awareness of physical activity recommendations, and perceived barriers and facilitators. Descriptive analyses were performed. The PASIPD score was 12.94 (4.48-26.86) MET-hr/day (median [IQR]) and 17.25 ± 15.35 MET-hr/day (mean ± SD). Awareness of recommendations was reported by 32% of participants, while 10% and 4% correctly identified aerobic and strength recommendations, respectively. The most frequently reported barrier was lack of suitable facilities (42%), and the most commonly reported facilitator was maintaining a healthy body (74%). Individuals with SCI in Saudi Arabia reported lower PASIPD scores compared with previous studies and limited awareness of recommendations, alongside commonly reported environmental and informational barriers.
In Vitro Functional Analysis of Regulatory T cells: Focus On Proliferation And Differentiation.
Zhang W, Cheng H, Ma X … +2 more , Zhang D, Nie C
J Vis Exp · 2026 Jun · PMID 42372030 · Publisher ↗
Regulatory T (Treg) cells play key roles in maintaining immune homeostasis. Abnormalities in Treg cell function are associated with various diseases, including autoimmune diseases, organ transplant rejection, infection,...
Regulatory T (Treg) cells play key roles in maintaining immune homeostasis. Abnormalities in Treg cell function are associated with various diseases, including autoimmune diseases, organ transplant rejection, infection, and cancer. Manipulating Treg cells is considered a potential therapeutic strategy for treating inflammatory diseases, including autoimmune disorders, solid organ transplant rejection, graft-versus-host disease, and allergic diseases. Classic in vitro Treg cell suppression assays mainly assess the inhibitory effect of Treg cells on effector T (Teff) cell proliferation. However, in addition to proliferation, T cell differentiation is important for T cells to perform their immune functions. Therefore, this protocol provides a modified in vitro Treg cell suppression assay for investigating the effects of Treg cells on the proliferation and differentiation of Teff cells. In this protocol, the preparation of mouse Treg cells and naive T cells is first introduced, followed by the co-culture strategy for studying the regulatory effects of Treg cells on the proliferation and differentiation of Teff cells, including Th1 and Th17 cells, and finally, the staining strategy for flow cytometry after co-culture is presented. This in vitro functional analysis of Treg cells can be used to investigate the effects of Treg cells on T cell proliferation and polarization.
Development and Validation of a Service Quality Assessment System for Chronic Disease Management in Primary Healthcare.
Tian Y, Yuan S, Yu D … +1 more , Liu Y
J Vis Exp · 2026 Jun · PMID 42372029 · Publisher ↗
Chronic non-communicable diseases require high-quality primary healthcare management, yet existing evaluation systems often fail to capture the multidimensional nature of patient-perceived service quality in chronic care...
Chronic non-communicable diseases require high-quality primary healthcare management, yet existing evaluation systems often fail to capture the multidimensional nature of patient-perceived service quality in chronic care settings. This study developed and validated a service quality assessment system (SERVQUAL) tailored for primary chronic disease management based on the SERVQUAL framework. A multi-stage methodology integrating the Delphi method and analytic hierarchy process (AHP) was employed to construct and weight the scale. The instrument was empirically validated using cross-sectional data from 433 patients across five primary healthcare institutions. Construct validity and reliability were evaluated through exploratory factor analysis (EFA) and confirmatory factor analysis (CFA). Furthermore, the predictive performance of the AHP-SERVQUAL model was compared with that of alternative models (KANO, TOPSIS-RSR) for patient compliance, problem resolution rates, and complaint frequencies. Psychometric evaluation demonstrated that the adapted five-dimensional scale (tangibility, reliability, responsiveness, assurance, and empathy) possessed high internal consistency and structural validity. Application across the centers revealed significant variations in service quality, particularly in the assurance and empathy dimensions. Comparative modeling indicated that the AHP-SERVQUAL framework yielded higher predictive power for patient adherence and overall service quality than the alternative models. Additionally, the quality indices generated by this model were significantly correlated with higher problem-resolution rates and lower complaint frequencies. The AHP-SERVQUAL-based system offers a reliable and valid metric for evaluating chronic disease management in primary care. By accurately capturing patient-perceived quality, this instrument provides administrators with an evidence-based tool to target quality improvement initiatives.
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