Journal Of Visualized Experiments[JOURNAL]
Sun
200 papers
Inducing Daytime Circadian Phase Shifts Using Chemogenetic and Spectral Approaches.
Bass S, Sun M, Gold E … +3 more , Wang H, Hattar S, Komal R
J Vis Exp · 2026 Jun · PMID 42371964 · Publisher ↗
In mammals, circadian phase shifting during the daytime is limited by reduced photic responsiveness of the suprachiasmatic nucleus (SCN), restricting the ability to experimentally manipulate the circadian clock during th...
In mammals, circadian phase shifting during the daytime is limited by reduced photic responsiveness of the suprachiasmatic nucleus (SCN), restricting the ability to experimentally manipulate the circadian clock during this phase. Reliable methods to induce daytime phase shifts are therefore essential for investigating mechanisms of circadian plasticity and photic entrainment. Complementary genetic and spectral strategies are described to enable robust, temporally precise manipulation of the circadian clock during the day. The first approach, currently limited to mice, employs a chemogenetic strategy involving intravitreal delivery of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) to selectively activate intrinsically photosensitive retinal ganglion cells (ipRGCs). This approach permits controlled activation of the retinohypothalamic pathway and induces reproducible daytime phase shifts independent of ambient lighting conditions. The second approach utilizes wavelength-specific optical stimulation as a non-invasive alternative. Exposure to violet light exploits the spectral sensitivity of ipRGCs to reduce depolarization block and promote sustained activation, enabling reliable phase resetting during the subjective day. This method is broadly applicable across mammalian systems and does not require genetic manipulation or pharmacological intervention. Detailed protocols are provided for experimental preparation, stimulation timing, validation of phase shifts using locomotor activity, and assessment of neuronal activation via c-Fos immunohistochemistry. Key considerations, including circadian timing, stimulus parameters, and experimental controls, are outlined to facilitate reproducibility. Together, these approaches provide versatile and experimentally tractable tools for inducing daytime circadian phase shifts and enable direct investigation of mechanisms underlying daytime circadian responsiveness.
Spatially Resolved, Integrated Single-Cell Multiomic Profiling of the Transcriptome and Epigenomic Targets in Frozen Tissue Sections.
Zhang T, Farassati FS, Stark-Quiroz A … +5 more , Liu Y, Huynh NM, Hayashi Y, Ordog T, Lee JH
J Vis Exp · 2026 Jun · PMID 42371963 · Publisher ↗
Spatial multiomic technologies combined with single-cell profiling offer unprecedented opportunities to elucidate the molecular and cellular architecture of complex tissues. Combining spatial barcoding of nuclei within i...
Spatial multiomic technologies combined with single-cell profiling offer unprecedented opportunities to elucidate the molecular and cellular architecture of complex tissues. Combining spatial barcoding of nuclei within individual cryosections prepared from tissue blocks embedded in optimal cutting temperature (OCT) freezing medium with single-cell multiomic assays into a simple and efficient workflow can facilitate the annotation and spatially resolved molecular analysis of cells in tissue types. This study reports a spatial cleavage under targets & tagmentation (CUT&Tag) multiome approach that simultaneously profiles histone H3K27ac modification and gene expression in a single frozen brain section. Following spatial barcoding, single nuclei are isolated and subjected to CUT&Tag, combined capture of tagmented DNAs, RNAs, and spatial barcodes, library preparation, and sequencing. The workflow generates spatially annotated epigenomic and transcriptomic profiles from the same nuclei, enabling assignment of the multiomic information to anatomical coordinates. Integrating epigenomic information with spatial transcriptomic data increases the fidelity of cell-type identification and reveals spatially organized regulatory programs and cell-cell interactions within intact tissues.
Confluence Protocol For XR-based Cricket Batting: A Framework For Integrated Performance Analysis.
Noorbhai H
J Vis Exp · 2026 Jun · PMID 42371961 · Publisher ↗
Extended reality (XR) technologies provide immersive training environments in sport, but their effectiveness for skill development and performance analysis requires careful and systematic evaluation. This protocol presen...
Extended reality (XR) technologies provide immersive training environments in sport, but their effectiveness for skill development and performance analysis requires careful and systematic evaluation. This protocol presents an integrated XR training methodology for cricket batting that combines immersive simulation with physiological and biomechanical measurements. It outlines procedures to collect and analyze muscle activity, metabolic responses, and motion-capture data during XR-based batting to characterize physiological and biomechanical responses. The goal is to evaluate batting performance (including power hitting) within a controlled virtual scenario while measuring muscle activation via electromyography (EMG), metabolic demands via respiratory gas analysis (VO₂), heart rate (HR), and kinematic outputs via motion capture. A single batter (n = 1) wears a VR headset and inertial sensors to face a virtual bowler, with EMG electrodes on key upper-body muscles and a portable metabolic system for breath-by-breath analysis. The protocol outlines the setup of the XR system, sensor calibration, and execution of standardized batting trials, with representative results presented as time-synchronized outputs. EMG traces indicated peak muscle activation during stroke execution, while VO₂ and HR fluctuations suggested a moderate aerobic response. Joint kinematics, including elbow extension and trunk rotation, were captured through motion data. These results illustrate the feasibility of using XR-based training to capture physical and physiological responses, offering a novel framework for enhancing performance and analyzing responses. The critical implementation steps, advantages over traditional drills, potential troubleshooting issues, and considerations for integrating XR into cricket training programs are discussed. This confluence of immersive technology with sports science measurements provides a framework for objectively assessing and exploring batting performance metrics in an XR setting and could be replicated in other sports.
A Low-Cost Markerless DeepLabCut-Based Workflow for Spontaneous Gait and Locomotion Analysis in Freely Moving Mice.
Andrade-Guerrero J, Meda-Hernández A, Sasia-Saldivar V … +2 more , Díaz-Miranda SY, Soto-Rojas LO
J Vis Exp · 2026 Jun · PMID 42371955 · Publisher ↗
Gait is a widely used functional biomarker for detecting motor alterations across various diseases and conditions, as it reflects changes in coordination, strength, balance, and sensorimotor integration. However, traditi...
Gait is a widely used functional biomarker for detecting motor alterations across various diseases and conditions, as it reflects changes in coordination, strength, balance, and sensorimotor integration. However, traditional methods to analyze gait in animal models often require expensive equipment, complex setups, or invasive procedures that can alter natural behavior. Here, we present a low-cost, markerless workflow based on DeepLabCut, an open-source pose estimation software, for the quantitative analysis of gait and spontaneous locomotion in freely moving mice. The method relies on single-camera video acquisition, markerless tracking of anatomical landmarks, and extraction of spatiotemporal locomotor parameters, without the need for physical markers or specialized hardware. To demonstrate the protocol's applicability, it was implemented in the triple transgenic mouse model of Alzheimer's disease (3xTg-AD) as a representative example of application. This approach preserves free movement and minimizes handling-related stress, enabling non-invasive assessment of motor behavior. The protocol is compatible with standard behavioral testing environments. Overall, this method provides an accessible and non-invasive framework for quantitative analysis of gait and locomotion in preclinical research.
Long Short-Term Memory Modeling of the Chinese Traditional Mongolian Osteopathy Rolling Method Using Adam and the Hodrick-Prescott Filter.
Ji Y, Pang J
J Vis Exp · 2026 Jun · PMID 42371954 · Publisher ↗
The acquisition and prediction of mechanical information were important for the digitization of Chinese Traditional Mongolian Osteopathy. The extraction of periodic and trend-oriented mechanical information from the forc...
The acquisition and prediction of mechanical information were important for the digitization of Chinese Traditional Mongolian Osteopathy. The extraction of periodic and trend-oriented mechanical information from the force exertion process presented a substantial challenge in achieving precise mechanical information prediction. To tackle this challenge, this article introduced a double-layer Long Short-Term Memory (LSTM) network integrated with Hodrick-Prescott (HP) filtering, which combined HP filtering techniques with LSTM to enhance the accuracy of mechanical information prediction. This research was grounded in experimentally collected data for mechanical information analysis, wherein dimensionality reduction was performed on the acquired data based on the primary joints involved in force application. Specifically, three sensor data points were selected from a total of 24 as pivotal input features. LSTM was employed to capture long-term dependencies inherent in sequential data. In this investigation, the Adam optimization algorithm was utilized to fine-tune the model's hyperparameters, ensuring optimal performance. The experimental outcomes underscored the efficacy of the proposed methodology, evidenced by a coefficient of determination (R) of 0.897, a mean absolute error of 0.025, and a root mean square error of 0.031. A comparative analysis with alternative estimation methods further attested to the stability, accuracy, and generalization capabilities of the two-layer LSTM network augmented with HP filtering.
Standardized Ileal Bladder Augmentation For Enterocystoplasty In Rats via Midline Laparotomy.
Studier-Fischer A, Maywald L, Grützner P … +10 more , Özdemir B, Haney CM, Knoedler S, Renner L, Carl N, Clanget T, Salg GA, Westhoff N, Michel MS, Kowalewski KF
J Vis Exp · 2026 Jun · PMID 42371952 · Publisher ↗
Bladder augmentation using intestinal segments remains a cornerstone reconstructive procedure for patients with low-capacity or poorly compliant bladders when conservative therapies fail. Despite its clinical relevance,...
Bladder augmentation using intestinal segments remains a cornerstone reconstructive procedure for patients with low-capacity or poorly compliant bladders when conservative therapies fail. Despite its clinical relevance, experimental progress and in-depth study are limited by the lack of standardized, reproducible protocols that reliably model surgical principles in a controlled preclinical setting. Here, we present a step-by-step rat enterocystoplasty protocol that uses a vascularized, detubularized ileal patch as a pedicled flap to augment the native bladder. The procedure includes midline laparotomy, controlled exposure, and stabilization of the bladder with holding sutures, urinary diversion via an exteriorized cannula tube Foley, isolation of a short ileal segment with an intact mesenteric pedicle, detubularization, and tension-free anastomosis with the bladder wall. Emphasis is placed on maintaining mesenteric perfusion, preventing pedicle torsion, ensuring watertight suturing, and securing catheter patency, which are critical for procedural success and reproducibility. Representative intraoperative validation using hyperspectral imaging (HSI) in eight non-survival rats demonstrated preserved tissue oxygenation and perfusion of the bowel segment after key operative steps, supporting tissue viability at the end of this procedure. Thus, this protocol provides a standardized experimental approach for reproducible surgical implementation of rat enterocystoplasty. Potential downstream applications of the model include the investigation of metabolic and electrolyte alterations, infection susceptibility, mucus production, and induction of dysplasia or malignancy, and remain to be addressed in future survival investigations.
Generating Tumor Organoid Cultures from the Lung Adenocarcinoma Cell Line.
Zhao X, He Y, Lin X … +1 more , Guo B
J Vis Exp · 2026 Jun · PMID 42371945 · Publisher ↗
Here, we describe a protocol for generating Cell Line-Derived Tumor Organoids (CDTOs) from the A549 human lung adenocarcinoma cell line. The protocol involves embedding 2D-expanded A549 cells in Matrigel and maintaining...
Here, we describe a protocol for generating Cell Line-Derived Tumor Organoids (CDTOs) from the A549 human lung adenocarcinoma cell line. The protocol involves embedding 2D-expanded A549 cells in Matrigel and maintaining them in 3D culture medium for long-term culture. Recommended seeding density of 500 cells/µL was determined to support consistent organoid formation. The resulting CDTOs were characterized by hematoxylin and eosin (H&E) staining and immunofluorescence (IF). The organoids maintained high expression of the lung adenocarcinoma markers Thyroid Transcription Factor 1 (TTF-1), adhesion protein E-Cadherin (ECAD), and cytoskeleton protein Keratin 7 (KRT7). Furthermore, tight junction protein Zona Occludens 1 (ZO-1) expression showed dysregulated polarity of tumor organoids. This protocol offers a technically straightforward, cost-effective, and purely tumorous organoid platform for lung adenocarcinoma research. Its simplicity and reproducibility also make it suitable for undergraduate laboratory teaching, where it can help students acquire fundamental 3D tumor organoid culture techniques within a limited lab schedule.
A Method for Adapting Euglena gracilis to Cassava-Based Substrates for Heterotrophic Cultivation.
Guaman Vargas A, Zhang Z, Liew A … +3 more , Hatvani Z, Noble A, Emery RJN
J Vis Exp · 2026 Jun · PMID 42371943 · Publisher ↗
Cassava processing generates nutrient-rich by-products that remain underutilized due to variable composition, cyanogenic compounds, and active microbial communities. This protocol presents a multistage adaptation strateg...
Cassava processing generates nutrient-rich by-products that remain underutilized due to variable composition, cyanogenic compounds, and active microbial communities. This protocol presents a multistage adaptation strategy for establishing a cassava-tolerant lineage of Euglena gracilis suitable for heterotrophic cultivation in cassava-derived substrates. The method begins with simultaneous exposure of Euglena cultures to autoclaved and non-autoclaved cassava media and to a garri concentration gradient of 10-30 g/L to assess tolerance to microbial complexity and substrate load. Cultures are then subjected to a higher concentration gradient of 30-50 g/L to evaluate performance with denser substrates. The resulting lineage is compared with a non-adapted wild-type strain across five cassava products; garri, dried pulp, pellets, peels, and animal feed to assess growth consistency in field-relevant materials. The protocol includes measurement of pH changes, cyanide levels using colorimetric strips, and preparation of dried biomass for protein analysis based on total nitrogen quantification. Together, these procedures provide a reproducible framework for adapting Euglena gracilis to cassava-derived substrates and for evaluating strain performance across diverse cassava products. This method enables the development of robust algal lineages for use in cassava-focused bioprocessing and waste-valorization applications.
Intracranial Breast Cancer Brain Metastases Mouse Model for Immune and Stromal Cells Characterization in Meninges and Olfactory Bulb.
Macela C, Carreira B, Ferreira-Silva M … +2 more , Florindo HF, Acúrcio RC
J Vis Exp · 2026 Jun · PMID 42371937 · Publisher ↗
The central nervous system (CNS) is surrounded by specialized immune interfaces, including the meninges and the olfactory bulb, which can serve both as permissive routes for metastatic colonization and as entry points fo...
The central nervous system (CNS) is surrounded by specialized immune interfaces, including the meninges and the olfactory bulb, which can serve both as permissive routes for metastatic colonization and as entry points for anti-tumor immune responses. Understanding how immune cell dynamics evolve within these structures during metastatic progression is essential for identifying mechanisms that either support or restrict tumor growth in the brain. This protocol details the establishment of a breast cancer brain metastases (BCBM) mouse model through intracranial injection of E0771 breast cancer cells into C57BL/6 mice. After tumor development, the meninges and olfactory bulb are carefully dissected, enzymatically digested, and processed into single-cell suspensions. These populations are then analyzed using multiparametric spectral flow cytometry to comprehensively characterize lymphoid, myeloid, and stromal cell subsets within CNS-associated immune niches. This workflow enables comparative analyses between tumor-bearing and non-tumor controls, identifying alterations within CNS immune interfaces that may underlie tumor immune evasion and represent potential therapeutic targets. Overall, the protocol provides a reproducible approach to study BCBM and interrogate immune regulation at CNS-peripheral immune system interfaces. It can be adapted for additional cancer cell lines, immunotherapies, or genetic mouse models to investigate immune resistance mechanisms and therapeutic response within the brain microenvironment.
Laparoscopic Ovum Pick-up For In Vivo Oocyte Retrieval In Alpacas.
Landinez-Aponte J, Joo M, Stott R … +4 more , Merrill N, Cisneros J, Liu Y, Wang Z
J Vis Exp · 2026 Jun · PMID 42371936 · Publisher ↗
Alpacas are increasingly incorporated into reproductive and biomedical research due to distinctive physiological and immunological characteristics that extend their relevance beyond agricultural applications. As camelids...
Alpacas are increasingly incorporated into reproductive and biomedical research due to distinctive physiological and immunological characteristics that extend their relevance beyond agricultural applications. As camelids, alpacas exhibit induced ovulation, prolonged follicular dynamics, asymmetric uterine physiology, and a unique immune system characterized by heavy-chain-only antibodies. These features support their use in comparative and translational research, particularly in reproductive biology and genetic engineering. Laparoscopic ovum pick-up (LOPU) is a minimally invasive approach for oocyte recovery in species with complex reproductive anatomy, allowing direct ovarian visualization, precise follicular aspiration, and reduced tissue trauma compared with transvaginal techniques. The objective of this study was to develop and standardize a reproducible in vivo oocyte retrieval protocol in alpacas using LOPU. Eight adult female alpacas (3-8 years of age) were enrolled in this study and subjected to repeated LOPU sessions conducted once per month throughout the study period. Alpacas were evaluated by transrectal ultrasonography, and animals presenting ovarian follicles ≥7 mm received 50 µg gonadotropin-releasing hormone intramuscularly. Ovarian superstimulation was induced using a total dose of 200 mg follicle-stimulating hormone administered in a decreasing twice-daily regimen over four days. LOPU was performed 12-14 h after the final hormone administration under general anesthesia induced with ketamine, xylazine, and butorphanol and maintained with isoflurane. A three-port laparoscopic approach with carbon dioxide insufflation was used. Follicles were aspirated using a 20-gauge needle connected to a regulated vacuum system (25-30 mmHg; 13-15 mL/min). Oocytes were recovered in TCM-199 medium and evaluated under a stereomicroscope. Alpacas exhibited a mean of 14.88 ± 3.85 follicles, with 10.94 ± 3.62 follicles aspirated per session and 6.68 ± 2.21 oocytes recovered, corresponding to a recovery rate of 61.30 ± 8.21%. These findings suggest that LOPU may represent a viable approach for in vivo oocyte retrieval in alpacas, with potential applications in reproductive biotechnology workflows.
Hybrid CNN-ViT Modeling For Predicting Functional Outcome After Ischemic Stroke: A Retrospective Study.
Wang Y, Peng Z, Mi R … +3 more , Chen X, Song Y, Wu R
J Vis Exp · 2026 Jun · PMID 42371934 · Publisher ↗
Acute ischemic stroke (AIS) is a major cause of death and long-term disability, and early prediction of functional recovery is important for guiding clinical decision-making and rehabilitation planning. Functional outcom...
Acute ischemic stroke (AIS) is a major cause of death and long-term disability, and early prediction of functional recovery is important for guiding clinical decision-making and rehabilitation planning. Functional outcome at 90 days is commonly assessed using the modified Rankin Scale (mRS), but predicting long-term outcome from early clinical and imaging information remains challenging. We hypothesized that integrating multiparametric magnetic resonance imaging (MRI) with structured clinical variables using a hybrid convolutional neural network (CNN)-Vision Transformer (ViT) architecture would improve prediction of 90-day functional outcomes compared with single-modality models. A retrospective cohort of 300 AIS patients who underwent multiparametric MRI was analyzed and divided into a training-validation cohort (n = 250) and an internal independent test cohort (n = 50). An additional external test cohort of 37 AIS patients was included to assess model generalizability. A hybrid CNN-ViT model was developed to extract multiparametric MRI features, and imaging and clinical predictions were integrated using stacked logistic regression. Model performance was evaluated using the area under the receiver operating characteristic curve (AUC), sensitivity, specificity, and accuracy. Among the evaluated clinical models, the support vector machine achieved the highest internal test AUC (0.878). The imaging model achieved an AUC of 0.782. The multimodal fusion model achieved the best overall internal performance, with an AUC of 0.885, sensitivity of 0.920, specificity of 0.760, and accuracy of 0.840. Similar performance trends were observed in the external test cohort. These findings suggest that stacked fusion of multiparametric MRI and clinical predictions may improve 90-day functional outcome prediction after AIS. However, larger multicenter validation studies are required before clinical implementation.
Effectiveness of Clinical Pathway-Based Rehabilitation Nursing on Swallowing Function in Stroke Patients With Dysphagia: A Retrospective Cohort Study.
Chai T, Luo J, Wang J … +2 more , Mei N, Sun Y
J Vis Exp · 2026 Jun · PMID 42371933 · Publisher ↗
Dysphagia after stroke increases the risk of aspiration pneumonia, malnutrition, and prolonged hospitalization. Standardized nursing pathways may improve care delivery, but real-world evidence remains limited. This retro...
Dysphagia after stroke increases the risk of aspiration pneumonia, malnutrition, and prolonged hospitalization. Standardized nursing pathways may improve care delivery, but real-world evidence remains limited. This retrospective cohort study, reported in accordance with the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement, included 210 adult stroke patients with dysphagia treated at Suzhou Hospital, Affiliated Hospital of Medical School, Nanjing University, from January 2020 to May 2025. Patients received either clinical pathway-based rehabilitation nursing (n = 106) or conventional care (n = 104). Primary outcomes were improvement in swallowing function at discharge and aspiration pneumonia. Secondary outcomes included nutritional status, functional recovery, ICU stay, overall complications, and length of hospital stay. Group differences were assessed using chi-square tests, independent-samples t tests, or Mann-Whitney U tests, and multivariable logistic regression was used to identify independent factors associated with swallowing improvement. Compared with conventional care, pathway-based care was associated with a higher rate of swallowing improvement (72.6% vs. 51.9%, P < 0.001), a lower incidence of aspiration pneumonia (7.5% vs. 15.4%, P = 0.015), higher functional recovery at discharge (Barthel Index ≥70: 68.9% vs. 49.0%, P < 0.001), a shorter ICU stay (1.7 ± 1.0 vs. 2.5 ± 1.2 days, P = 0.002), a lower overall complication rate (21.7% vs. 31.7%, P = 0.029), and a shorter hospital stay (35.7 ± 7.8 vs. 47.9 ± 10.1 days, P < 0.001). In multivariable analysis, clinical pathway-based nursing remained independently associated with swallowing recovery (adjusted OR = 2.12, 95% CI, 1.38-3.27; P < 0.001). Clinical pathway-based rehabilitation nursing was associated with improved swallowing outcomes and fewer in-hospital complications in this retrospective cohort, supporting further prospective multicenter evaluation and integration into standardized stroke rehabilitation practice.
Bawei Aiguning Granules For Lung Cancer Bone Metastases: A Randomized, Double-blind, Placebo-Controlled Trial Protocol With A Representative Case.
Zhao P, Hong Y, Chang J … +12 more , Guo S, Yan B, Zhou C, Wang X, Zhao F, Tong J, Zhang X, Jiao L, Gong Y, Wang Y, Sun X, Yang Y
J Vis Exp · 2026 Jun · PMID 42371923 · Publisher ↗
Bone metastases (BM) frequently lead to skeletal-related events (SREs) and significant pain in lung cancer patients, requiring more effective supportive management strategies. We designed a prospective, multicenter, rand...
Bone metastases (BM) frequently lead to skeletal-related events (SREs) and significant pain in lung cancer patients, requiring more effective supportive management strategies. We designed a prospective, multicenter, randomized, double-blind, placebo-controlled trial to evaluate Bawei Aiguning Granules (BWAGN) in combination with standard bone-targeted therapy. A total of 156 patients will be randomized across six centers. The primary endpoint is the 1-year incidence of SREs. Secondary endpoints include time to first SRE, pain assessment, quality of life, survival, Traditional Chinese Medicine symptom score, and bone turnover markers. Safety will be assessed according to Common Terminology Criteria for Adverse Events v5.0. Alongside the trial protocol, this manuscript presents currently available descriptive observations from a representative participant. During follow-up, no SREs were documented, and serial Electron Computed Tomography (ECT) examinations showed no obvious new imaging abnormalities suggestive of bone progression or SRE-related change. Functional status, symptom burden, and tumor markers showed favorable changes over time, while routine safety assessments remained generally stable, with no adverse events or serious adverse events recorded. These observations are descriptive only and should be interpreted with caution. Definitive conclusions regarding efficacy and safety will require completion of the protocol-defined follow-up and formal analysis of the full study cohort.
Establishment of A Mouse Sepsis-Induced Immunosuppression Model By Intranasal Instillation of Bacteria.
Cao K, Shi L, Feng X … +8 more , Li Q, Fan J, Wang R, Gan L, Du J, Jiang J, Wang Z, Zeng L
J Vis Exp · 2026 Jun · PMID 42371921 · Publisher ↗
Sepsis is a life-threatening organ failure caused by the host's inappropriate response to infection. Pneumonia is the primary cause of sepsis. Animal models are important tools for studying the pathogenesis of sepsis and...
Sepsis is a life-threatening organ failure caused by the host's inappropriate response to infection. Pneumonia is the primary cause of sepsis. Animal models are important tools for studying the pathogenesis of sepsis and evaluating new treatment strategies. Previous animal models of sepsis lacked natural infection pathways, making it difficult to accurately reflect the clinical infection process. This study introduced a mouse sepsis immunosuppression model established by intranasal instillation of Klebsiella pneumoniae (KP) to simulate the natural respiratory infection pathway. After modeling with different concentrations of KP, the inflammation of the model was evaluated at 12 h for cytokine interleukin-6 (IL-6) levels, and organ damage was evaluated at 24 h for alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatine kinase (CK), and blood urea nitrogen (BUN) levels. The survival rate of the model was analyzed after 7 days. The percentage of lymphocytes and white blood cells, the proportion of major histocompatibility complex class II (MHC-II) expression in monocytes, and the mean fluorescence intensity (MFI) of IL-10 and TGF-β1 in CD4 T cells were observed to evaluate the immune suppression in mice. After 7 days of modeling, a secondary infection was performed by intranasal instillation of Pseudomonas aeruginosa (PA). The bacterial load in bronchoalveolar lavage fluid (BALF) 24 h after secondary infection and the mortality rate within 14 days were assessed to further evaluate the model's immune suppression. Nasal instillation of high-concentration Klebsiella pneumoniae can effectively induce an immune suppression. Its purpose is to simulate sepsis induced by a natural respiratory infection pathway and to provide a standard model for sepsis research in animals.
A Protocol For Designing And Evaluating Mixed Reality Cultural Experiences In Museums And Heritage Sites Using Spatial Narratives And Metrics.
Zhang M, Liu X, Wang D … +1 more , Lu S
J Vis Exp · 2026 Jun · PMID 42371914 · Publisher ↗
Mixed reality (MR) applications are increasingly used in museums and heritage sites; however, their deployment often lacks methodological coherence, with spatial narrative, interaction, and evaluation treated as separate...
Mixed reality (MR) applications are increasingly used in museums and heritage sites; however, their deployment often lacks methodological coherence, with spatial narrative, interaction, and evaluation treated as separate elements. This protocol presents a standardized, field-deployable methodology that distinguishes itself from prior evaluation frameworks by integrating dynamically adaptive spatial narratives with a hybrid assessment model. The approach unifies spatial narrative sequencing, guided interactive interfaces, and multimodal user experience metrics-specifically triangulating subjective psychometric evaluations with objective behavioral telemetry (e.g., spatial dwell times and interaction logs). The protocol describes a two-site, three-arm concurrent control trial adaptable to indoor museums and semi-open archaeological venues. Participants are assigned to one of three conditions: a traditional digital guide, a MR static narrative, or a MR adaptive narrative. A standardized six-zone route is implemented, with assessments conducted at baseline, immediately after the visit, and at a 14-day follow-up. Outcome measures include spatial presence, usability, narrative engagement, perceived authenticity, and knowledge acquisition. The protocol further incorporates structured behavioral logging, predefined technical thresholds, and systematic cybersickness monitoring to ensure safety and feasibility under routine visitor conditions. This framework provides researchers and cultural institutions with a reproducible methodology to coordinate spatial storytelling and interface control, enabling rigorous evaluation of MR interventions in heritage contexts.
Image Caption Generation using Deep Learning Approaches.
Singh AP, Manoria M, Joshi S
J Vis Exp · 2026 Jun · PMID 42371865 · Publisher ↗
Image caption generation is an endeavor to provide a meaningful textual description that involves an image. Extracted information is relevant to the activities present in the images. ResNet (Residual Network) is well kno...
Image caption generation is an endeavor to provide a meaningful textual description that involves an image. Extracted information is relevant to the activities present in the images. ResNet (Residual Network) is well known for its ability to classify images, having developed deep hierarchical representations. The intention of this paper is to use ResNet with various smart filters to classify images more deeply, enabling the generation of genuine and meaningful descriptions that are highly precise with respect to the reference captions. Here, the work uses a smart filtering technique to enhance images, a CNN to encode features, model training, and thereafter an RNN (Recurrent Neural Network) to decode the features. ResNet is a very effective model for computer vision tasks, especially object classification and semantic analysis. ResNet is well known for residual connections, which are also known as skipping connections that solve the vanishing gradient problem, which is a crucial problem in deep learning. Here, the MSCOCO (Microsoft Common Object in Context) benchmark is used to train the model, which is a large dataset with reference annotations useful for various computer vision tasks. ResNet helps enhance generalization capability, which is particularly useful for diverse images. As per the results obtained, BLUE scores are B1: 0.579, B2: 0.404, B3: 0.279, B4: 0.191; METEOR: 0.195; ROUGE: 0.396; and CIDEr: 0.6.
Research Progress In Acupuncture For Parkinson's Disease: Insights Into The Mitochondrial Ferroptosis Pathway.
Yi L, Dong YB, Ran CC … +5 more , Chen Y, Yu CX, Mao K, Zhou Y, Tu H
J Vis Exp · 2026 Jun · PMID 42371862 · Publisher ↗
The pathogenesis of Parkinson's disease (PD) is highly complex, with mitochondrial ferroptosis-a novel regulated cell death modality-playing a critical role in dopaminergic neuronal degeneration. This review systematical...
The pathogenesis of Parkinson's disease (PD) is highly complex, with mitochondrial ferroptosis-a novel regulated cell death modality-playing a critical role in dopaminergic neuronal degeneration. This review systematically elucidates the potential mechanisms of acupuncture in treating PD by modulating mitochondrial ferroptosis pathways, alongside analyzing the specificity of acupoint prescriptions and stimulation parameters. Databases, including PubMed and China National Knowledge Infrastructure (CNKI), were systematically searched for literature regarding acupuncture interventions in PD models and ferroptosis mechanisms. The analysis focused on acupuncture's regulation of mitochondrial iron overload, the System Xc⁻/glutathione (GSH)/glutathione peroxidase 4 (GPX4) antioxidant axis, and the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. Acupuncture is proposed to mitigate mitochondrial ferroptosis through multidimensional synergistic mechanisms. First, it remodels iron metabolism by inhibiting transferrin receptor 1 (TFR1)/divalent metal transporter 1 (DMT1)-mediated iron uptake and upregulating ferritin heavy chain 1 (FTH1) to sequester iron harmlessly, thereby reducing the mitochondrial labile iron pool. Second, it may help restore antioxidant defenses by activating Nrf2 nuclear translocation, upregulating GPX4, and restoring the System Xc⁻/GSH pathway to scavenge lipid peroxides. Third, regarding intervention specificity, the Fengfu (GV16) and Taichong (LR3) combination primarily modulates neuroinflammation, whereas Baihui (GV20) penetration upregulates neurotrophic factors. Additionally, 2 Hz low-frequency electroacupuncture emerges as the optimal parameter for neuroprotection, while acupuncture also exerts systemic effects via synchronous brain-gut regulation. Acupuncture maintains mitochondrial homeostasis and inhibits neuronal ferroptosis by restoring the iron-lipid-antioxidant triangular balance. Future multi-omics and clinical translational research are required to optimize parameters and provide robust evidence for neuroprotective PD therapies.
Quantifying Embryonic Lethality Rate in Caenorhabditis elegans.
Nourie A, Tam G, Ponomarova O
J Vis Exp · 2026 Jun · PMID 42371859 · Publisher ↗
Embryonic lethality is a cornerstone phenotype used in Caenorhabditis elegans research to characterize developmental defects arising from factors such as toxic exposures and genetic mutations. Therefore, accurately quant...
Embryonic lethality is a cornerstone phenotype used in Caenorhabditis elegans research to characterize developmental defects arising from factors such as toxic exposures and genetic mutations. Therefore, accurately quantifying the penetrance of embryonic lethality is critical for understanding the mechanisms and impact of these perturbations on animal development. Although automated scoring platforms exist, these approaches often require substantial financial investment and specialized hardware or software. This article describes a low-tech, low-cost assay that quantifies embryonic lethality by measuring the percentage of embryos that successfully fail to hatch. Notably, this version of the assay eliminates the need for a worm pick, making it exceptionally suitable for beginners. An additional advantage of this approach is that each technical replicate assays progeny from multiple animals, avoiding the potential biases of single-worm assays. This protocol includes a discussion of critical parameters, such as the minimum number of progeny required to accurately capture phenotype penetrance and the rationale for the methodology. Finally, we provide guidance on adapting the assay to assess the effects of gene knockdown via RNA interference, pharmacological treatments, or nutritional supplementation, adding a versatile and accessible tool to the toolkit of C. elegans researchers.
Standardized Preparation of Isolated Rabbit Duodenal Smooth Muscle For In Vitro Pharmacological Assessment of Contractile Activity.
Peng X, Luo Z, Li M … +4 more , Zhang R, Bao Y, Zeng N, Liu R
J Vis Exp · 2026 Jun · PMID 42371857 · Publisher ↗
The coordinated contractile activity of gastrointestinal smooth muscle forms the physiological basis for maintaining normal digestion, absorption, and transport functions, with its dysfunction closely associated with mot...
The coordinated contractile activity of gastrointestinal smooth muscle forms the physiological basis for maintaining normal digestion, absorption, and transport functions, with its dysfunction closely associated with motility disorders. The isolated organ perfusion technique eliminates complex in vivo interferences-such as neural, endocrine, and hemodynamic factors-serving as a classic model for pharmacological studies. To this end, this study details a stepwise protocol for the standardized preparation of rabbit isolated duodenal smooth muscle. The procedural workflow highlights the critical steps of preparing solutions, euthanizing the rabbit, performing a precise aseptic mid-abdominal laparotomy for rapid tissue excision, suspending the smooth muscle strips, and calibrating transducers for real-time tension recording. Based on this technical platform, we further validated the method by evaluating the effects of Huoxiang Zhengqi Oral Liquid (HXZQ-OL) on spontaneous contractions and acetylcholine/barium chloride-induced tetanic contractions. In conclusion, this protocol yields a robust and highly reproducible methodological framework, providing a broadly applicable in vitro screening platform for the pharmacological assessment of therapeutic agents targeting gastrointestinal motility.
Buckwheat: A Sustainable Alternative Crop Under Changing Climate.
Rashid M, Showkat M, Mir R … +13 more , Bashir S, Farooq J, Jaan T, Verma A, Magray M, Bhat B, Malik H, Manikandan K, Sofi P, Sofi N, Ali S, Rakwal R, Zargar SM
J Vis Exp · 2026 Jun · PMID 42371856 · Publisher ↗
Buckwheat (Fagopyrum spp.), a pseudocereal crop with high nutritional and ecological values, is gaining global attention as a climate-smart crop suitable for sustainable agriculture. Its short life cycle, adaptability to...
Buckwheat (Fagopyrum spp.), a pseudocereal crop with high nutritional and ecological values, is gaining global attention as a climate-smart crop suitable for sustainable agriculture. Its short life cycle, adaptability to marginal environments, and natural resilience to various abiotic and biotic stresses make it a promising crop for future cropping systems. Buckwheat, being a fast-growing and short-duration crop, fits seamlessly into crop rotation schedules and intercropping systems, especially in temperate and high-altitude regions. Recent literature clearly demonstrates the ability of buckwheat to withstand a wide range of stressors. Consequently, there is a great need to report the scientific findings backing the stress tolerance through underpinning physiological, molecular, and genetic insights behind its resilience. Additionally, buckwheat is reported to improve soil health, supporting biodiversity, and contributing to low-input farming systems. Current research gaps and strategic directions for breeding and biotechnological interventions to enhance buckwheat's resilience and productivity under changing climatic conditions have been highlighted in this study.
About
- Frequency
- Sun
- Papers found
- 200
- RSS feed
- Subscribe