Research on multimodal ultrasound parameters associated with postpartum pelvic floor dysfunction (PFD) across resting, maximal contraction, and Valsalva states remains limited. This retrospective study assessed these parameters in 172 patients examined between January 2022 and June 2025. After stratification into PFD and non-PFD groups, 1:1 propensity score matching (PSM) yielded 160 patients (80 per group) for analysis. Ultrasound parameters were evaluated across the three functional states. Logistic regression identified independent factors associated with PFD, and receiver operating characteristic (ROC) curve analysis assessed model performance. After PSM, baseline characteristics were comparable between groups (all p > 0.05). Compared with the non-PFD group, the PFD group had a larger resting levator ani hiatus area, a more inferior bladder neck position, and lower levator ani muscle thickness (all p < 0.001). During maximal contraction, the hiatus area reduction percentage and bladder neck elevation were lower. During the Valsalva maneuver, the hiatus area, bladder neck mobility, and most distal prolapse position were greater, and levator ani avulsion was more frequent (all p < 0.001). A larger resting levator ani hiatus area, lower resting muscle thickness, a lower hiatus area reduction percentage, greater bladder neck mobility during the Valsalva maneuver, and levator ani avulsion were independently associated with postpartum PFD (all p < 0.001). The five-parameter model had an area under the curve (AUC) of 0.870 (95% confidence interval, 0.815-0.926), with 91.3% sensitivity and 88.8% specificity. The model showed favorable internal fitting performance in the development dataset. Because independent validation was not performed, the findings should be considered hypothesis-generating; applications in clinical screening and individualized intervention remain unproven.
The rapid expansion of low-voltage (LV) networks and their integration with distributed energy resources requires intelligent and automated management solutions. Cloud-edge collaborative Internet of Things (IoT) platforms support real-time monitoring, control, and data acquisition. However, existing platforms generally lack workflow automation, visual process orchestration, user-guided decision support, and comprehensive validation. Consequently, they do not provide process-driven solutions for automated LV network acceptance testing. This study presents the design and evaluation of a Low-Voltage Acceptance Automation Platform based on a visualized process canvas. The proposed platform adopts a process-driven architecture in which acceptance workflows are visually created, managed, and executed. The visual process canvas transforms conventional static monitoring into dynamic workflow automation by enabling real-time workflow execution, validation, and decision-making. The framework incorporates the Open LV Network & Smart Meter dataset to support realistic modeling of electrical load behavior. The workflow includes IoT-based data acquisition, data preprocessing, feature engineering, workflow orchestration using the visual process canvas, machine learning-based validation, and real-time dashboard visualization. The proposed framework achieved a fault detection rate of 98.4%, a receiver operating characteristic area under the curve (ROC-AUC) of 0.968, and an operational decision latency of 31 ms, outperforming traditional cloud-centric and IoT-based baseline approaches by approximately 30%-40%.
Autophagy plays a complex role in pancreatic ductal adenocarcinoma (PDAC), contributing to tumor progression, stress adaptation, and therapy resistance. Accurate assessment of autophagosomal dynamics is therefore essential for studies of cancer biology. Among the available methods, immunoblotting of microtubule-associated protein light chain 3 (LC3) is widely used to monitor autophagosomal dynamics by distinguishing between the cytosolic (LC3-I) and lipidated, autophagosome-associated (LC3-II) forms. However, the low molecular weight of LC3 and the minimal difference in electrophoretic mobility between these isoforms present technical challenges that require careful optimization. Presented here is a reproducible protocol for the semi-quantitative analysis of LC3-II levels by Western blot in KPC-derived murine pancreatic cancer cells. The method incorporates optimized conditions for cell lysis, electrophoresis, protein transfer, and antibody-based detection to ensure reliable separation and detection of LC3 isoforms. In addition, a standardized workflow for densitometric analysis of LC3-II bands using Fiji (ImageJ) is provided. The sensitivity of the method is demonstrated through the detection of increased LC3-II levels under conditions of autophagy induction (gemcitabine treatment) and impaired autophagic flux (VMP1 knockdown). Critical technical parameters that influence data interpretation, including lysis buffer composition and antibody specificity, are also examined, together with common experimental pitfalls. Although LC3 immunoblotting alone is insufficient to fully define autophagic flux, when combined with complementary assays it provides a robust and accessible approach for monitoring autophagic activity. The protocol can be adapted to other experimental contexts, facilitating mechanistic studies of autophagy in cancer models.
The generation of patient-specific induced pluripotent stem cells (iPSCs) from amniotic fluid cells (AFCs) carrying defined chromosomal aneuploidies provides a powerful platform for modeling genetic disorders. However, establishing a reliable and reproducible reprogramming pipeline for aneuploid AFCs remains technically challenging due to the intrinsic genomic instability and variable proliferative capacity of these cells. Here, we present a comprehensive, non-integrating method for generating aneuploid human iPSCs from primary AFCs using episomal plasmid electroporation. This protocol details the complete workflow, encompassing cell thawing and expansion with a gradual media adaptation strategy, optimized plasmid delivery via electroporation system, sequential post-electroporation culture with mesenchymal-to-epithelial transition (MET)-directed media changes, and mechanical colony picking based on defined morphological criteria. We further describe validation procedures, including immunofluorescence staining for core pluripotency markers, G-banding karyotype analysis to confirm aneuploid karyotype maintenance, and PCR-based episomal vector clearance verification. This feeder-free, integration-free protocol yields aneuploid iPSC lines suitable for disease modeling, drug screening, and studies of chromosome biology.
Pearl powder, nacre, and nacre-derived water-soluble matrix (WSM) are natural mineral-organic materials with growing relevance for bone-regenerative research. Their biological activity appears to depend not only on calcium carbonate content but also on soluble matrix components that can influence osteoblast behavior, mineral deposition, and cellular stress responses. This narrative review compares pearl-specific osteogenic studies in cell, scaffold, and defect-repair models with selected comparator nanoparticle studies used to frame unresolved safety questions. The pearl-specific literature generally supports osteogenic activity, particularly in preosteoblast or osteoblast-lineage models and WSM-containing materials, but the evidence remains heterogeneous because species source, extraction method, particle size, release behavior, dose, and endpoint selection differ across studies. Evidence linking nano-pearl powder to autophagy-associated osteogenic differentiation is promising, especially in MC3T3-E1 models, but the existing pearl-specific literature provides limited direct evidence for high-dose toxicity, impaired autophagic flux, immunogenicity, heavy-metal-related risk, degradation-matched repair, or chronic biosafety. Future work should integrate material characterization, dose-response testing, oxidative and mitochondrial readouts, autophagic flux analysis, and osteogenic endpoints into a single experimental design. Such studies will be needed to define whether WSM-based or nano-pearl formulations can provide a reproducible and safe osteogenic window for bone-regenerative biomaterial development.
Branched actin polymerization mediated by the actin-related protein 2/3 (Arp2/3) complex provides the primary pushing forces for a variety of cellular processes, including cell migration, endocytosis, and phagocytosis. Myosin-I motors, which frequently colocalize with branched actin networks at the cell leading edge, have also been shown to participate in these processes and are thought to regulate actin network organization and mechanical output. However, the molecular mechanisms by which myosin-I interacts with the Arp2/3 complex to modulate branched actin assembly and force generation remain largely unknown. Here, we describe a highly tunable in vitro actin comet-tail bead motility assay that reconstitutes the interplay among myosin-I, actin, and the Arp2/3 complex at the cell leading edge on the surface of micron-sized beads. This method is adapted from well-established actin comet-tail assays by co-immobilizing myosin-I with nucleation-promoting factors (NPFs) on bead surfaces, thereby creating a membrane-like actin assembly interface. The assay enables visualization and quantitative analysis of actin network assembly, network density, bead motility, and growth efficiency. It also provides indirect, qualitative readouts of myosin-I-mediated force enhancement in branched actin networks. This protocol includes bead functionalization, reaction assembly, fluorescence imaging, quantitative image analysis, and troubleshooting strategies providing a reproducible platform for studying myosin-I-regulated actin assembly at membrane-like interfaces.
Assessment of autonomic dysfunction is crucial for clinical management of PD and MSA. Autonomic dysfunction can provide important clues for its diagnosis and clinical management. Furthermore, it often progresses over time and significantly affects patients' quality of life. Therefore, it is essential to develop bedside clinical assessment protocols for autonomic dysfunction. This paper provides a step-by-step description to demonstrate the evaluation method. In both outpatient and bedside settings, autonomic function should be systematically evaluated: beginning with a medical interview, followed by a physical examination and, when necessary, using simple instrumental tests. Assessment should emphasize circulatory, sudomotor, urinary, and bowel dysfunction. This paper will explain each of the following items. Circulatory assessment: medical interview, evaluation of cold-discolored extremities, capillary refill, 10-second cold water stress load test, active standing test, the coefficient of variation of R-R intervals, and overnight oximetry provides further detailed evaluation in MSA. Sudomotor symptoms: medical interview, assessment of sweating at rest and under stress using palpation, visual inspection, and the spoon test, and finger wrinkling after immersion in warm water. Urinary assessment: urination frequency, sensation of incomplete bladder emptying, voiding diaries, and the measurement of post-void residual (PVR) urine volume. Bowel assessment: medical interview, abdominal percussion, and rectal ultrasonography. Bedside assessment is influenced by the examiner's experience, medication, and the testing environment. Considering the limitations of bedside assessments, more comprehensive diagnostic evaluations should be conducted when necessary. We also describe the characteristics of autonomic dysfunction in PD and MSA. Appropriate management of each type of autonomic dysfunction is also essential.
Perineal recurrence of urethral squamous cell carcinoma (SCC) after radical penectomy is a rare and technically demanding entity, and non-surgical modalities such as radiotherapy or systemic chemotherapy may not achieve optimal local control in this setting, necessitating aggressive surgical resection combined with complex reconstructive techniques. We report the multidisciplinary management of an extensive perineal recurrence in a 56-year-old male with a history of primary urethral SCC (pT2 pN0 G2, first diagnosed in 2013) treated by radical penectomy and bilateral inguinal lymphadenectomy, who presented in April 2021 with perineal tumor recurrence. After an endoscopic and diagnostic evaluation, a perineal resection was performed, resulting in rpT4 G2 with positive surgical margins cranially toward the bladder and ventrally toward the symphysis. Complete tumor resection was deemed unachievable in that conventional way. After further consultation, the patient underwent robot-assisted radical Cystoprostatovesicourethrectomy with en bloc perineal tumor excision, bilateral pelvic and salvage inguinal lymphadenectomy with ICG-guided lymphangiography, prophylactic appendectomy, urinary and stool diversion via ileal conduit and colostomy. Histopathology confirmed R0 resection. Three weeks after the initial ablative surgery, the resulting large perineal defect, extending from the symphysis anteriorly to the rectum posteriorly, was closed with a left pedicled gracilis muscle flap. The 5-year follow-up was oncologically and functionally uneventful (last CT 03/2026). This case underscores the importance of interdisciplinary management involving urology, visceral surgery, reconstructive surgery, and radiology. It also highlights the benefits of integrating minimally invasive robotic techniques.
COPD is a progressive respiratory disorder characterized by persistent airflow limitation and chronic inflammation, yet the role of N4-acetylcytidine (ac4C) RNA modification in its pathogenesis remains largely unexplored. This study aimed to systematically screen for ac4C-related genes (ac4C-RGs) from a published database and investigate their regulatory networks in COPD, thereby identifying potential biomarkers for further mechanistic studies without assuming a direct regulatory relationship between any specific gene and ac4C modification. Differentially expressed genes (DEGs) were identified from transcriptomic profiles, and weighted gene co-expression network analysis (WGCNA) was applied to uncover key co-expression modules. Cross-analysis among DEGs, significant modules, and ac4C-RGs was conducted. Key genes were screened using LASSO regression, XGBoost, and random forest algorithms, followed by logistic regression‑based diagnostic model construction. Model performance was evaluated by receiver operating characteristic (ROC) curve analysis, area under the curve (AUC) with 95% confidence intervals, calibration curve assessment, and decision curve analysis (DCA). A total of 160 overlapping genes were identified, and six hub genes (PTRF, PRKCDBP, UPP1, TOR3A, FAM168B, and B4GALT2) were consistently selected by all three machine learning algorithms. The diagnostic model demonstrated good discriminative performance, with AUCs of 0.766, 0.759, and 0.723 in the training, internal test, and external validation sets, respectively. Regulatory network analysis suggested potential ceRNA axes and transcription factor interactions, while immune infiltration profiling revealed significant correlations between key genes and multiple immune cell subsets. Drug-gene interaction analysis and molecular docking indicated that fluorouracil, capecitabine, and 5-benzylacyclouridine may exhibit favorable predicted binding affinities with UPP1. In conclusion, PTRF, PRKCDBP, UPP1, TOR3A, FAM168B, and B4GALT2 were identified as potential ac4C-related biomarkers in COPD, potentially involved in immune and metabolic regulation, providing a foundation for future functional investigations and therapeutic exploration.
This paper presents a systematic optical range extension method for laser tracker calibration in a limited indoor space. The method is based on optical path folding using pyramid prisms and is designed to quantitatively reduce Abbe error. Both approaches realize dual-range measurement of laser trackers within confined indoor baseline space. Experimental results reveal that calibration tests carried out over an 80 m distance with dual pyramid prisms yield a maximum indicated error of -36.0 µm for the laser tracker, whereas the configuration adopting a single pyramid prism delivers a maximum indicated error of merely -2.7 µm. When the two measurement systems share a pyramid prism, the shortened Abbe arm effectively diminishes Abbe error. Additionally, the optical symmetry between the incident and emergent light beams further suppresses such error, leading to a lower indicated measurement error. Relevant experimental data confirm that the optical path layout using a single pyramid prism offers superior measurement precision. The experimental data show that the optical path experiment using a single pyramid prism has higher measurement accuracy. We hope the related results could serve as a reference for multiple range-extension measurements with the laser tracker.
Oocytes are densely packed with mitochondria, the energy-producing organelles that contain their own genome, mitochondrial DNA (mtDNA). Each cell contains multiple copies of mtDNA, with copy number varying among tissue types. Oocytes possess the highest mtDNA copy number, containing hundreds of thousands of mtDNA molecules per cell. Because mitochondria are inherited exclusively through the maternal lineage, accurate detection of mtDNA variants is essential for studies of inheritance, aging, and disease. The presence of multiple mtDNA copies allows wild-type and mutant molecules to coexist within the same cell, a condition known as heteroplasmy, in which low-frequency and de novo variants may occur at frequencies below 1%. Conventional next-generation sequencing (NGS) lacks sufficient accuracy to reliably distinguish these rare variants from errors introduced during library preparation and sequencing. Here, we present a protocol for enriching mtDNA from single human oocytes using Exonuclease V to remove linear DNA, followed by duplex sequencing library preparation for highly accurate mtDNA analysis. This workflow enables error-corrected sequencing of individual oocytes, facilitating reliable detection of low-frequency mtDNA variants and analysis of heteroplasmy and de novo mutagenesis. The protocol provides a reproducible approach for investigating mitochondrial genome variation in single oocytes using Illumina-compatible sequencing platforms.
Clear cell renal cell carcinoma (ccRCC) is the most common subtype of kidney cancer and is characterized by substantial clinical heterogeneity, highlighting the need for reliable prognostic biomarkers. This study evaluated the expression pattern, prognostic relevance, and immune-related associations of ARHGAP22 in ccRCC using transcriptomic and clinical data from The Cancer Genome Atlas Kidney Renal Clear Cell Carcinoma (TCGA-KIRC) cohort, together with external validation data and protein-expression information from the Human Protein Atlas (HPA). ARHGAP22 expression was compared between tumor and adjacent normal tissues, and its associations with overall survival, clinicopathological characteristics, tumor microenvironment scores, and estimated immune-cell fractions were assessed. Co-expression and functional-enrichment analyses were also performed to characterize potential biological associations. ARHGAP22 was significantly upregulated in ccRCC tissues at the transcriptomic level, with corresponding differences observed in immunohistochemical images. High ARHGAP22 expression was associated with shorter overall survival, advanced clinicopathological features, and higher ImmuneScore, StromalScore, and ESTIMATEScore values. CIBERSORT-based analysis showed that the high-expression group had higher estimated fractions of M2 macrophages and regulatory T cells and lower estimated fractions of naïve B cells, resting mast cells, and activated dendritic cells after false discovery rate correction. Functional-enrichment analyses linked ARHGAP22-associated genes to immune-related processes, cell migration, and chemokine- and cytokine-mediated signaling pathways. These findings suggest that ARHGAP22 may represent a potential prognostic and immune-related biomarker in ccRCC, although further independent clinical and experimental validation is required.
Circadian rhythms are endogenous oscillations of approximately 24 h that regulate a wide range of cellular and physiological processes, including gene expression, metabolism, and behavior. These rhythms arise from interconnected transcriptional-translational feedback loops that respond to temporal and environmental cues. Because circadian regulation is highly dynamic, even minor experimental variations can influence phase, amplitude, and rhythmicity, making standardized experimental workflows essential for generating reliable and reproducible results. The goal of the present protocol is to provide a practical and reproducible workflow for synchronizing cultured cells, performing time-course sampling, and analyzing circadian clock gene expression under standard laboratory conditions. The protocol describes serum shock-based synchronization, staggered sample collection over 24-72 h to avoid overnight sampling, ribonucleic acid extraction, complementary deoxyribonucleic acid synthesis, quantitative real-time polymerase chain reaction, and circadian rhythm analysis using appropriate statistical approaches. The workflow also highlights critical experimental considerations, including synchronization conditions, sample quality assessment, reference gene selection, and data analysis, to improve reproducibility across experiments. This method provides an accessible approach for investigating molecular circadian mechanisms and evaluating rhythmic gene expression in cultured cells, facilitating studies of circadian regulation in physiological and disease-related experimental models.
Volleyball spiking is a complex explosive skill that requires coordinated force generation and transfer across the upper limbs, trunk, and lower limbs. Neuromuscular electrical stimulation (NMES) has been proposed as a preconditioning strategy for enhancing neuromuscular activation; however, its acute effects on phase-specific volleyball spike kinematics remain unclear. This study investigated the acute effects of NMES priming on spike performance and three-dimensional kinematics in male university volleyball players. Thirty participants were randomly assigned to an NMES conditioning group (n = 15) or a control group (n = 15). All participants completed a baseline spike assessment and a second assessment 72 h later. Immediately before the second assessment, the NMES group received 30 min of stimulation applied to the upper-limb muscles, including the deltoid, biceps brachii, and triceps brachii; the external oblique as a core muscle; and the vastus lateralis as a lower-limb muscle. The control group completed the same time-matched assessment without NMES. Three-dimensional kinematic data were collected using a synchronized four-camera three-dimensional motion-analysis system. The primary analysis compared performance and kinematic variables between the groups at the second assessment. Compared with the control group, the NMES group demonstrated greater ball velocity, smaller knee and ankle angles at bilateral-foot contact, greater hip, knee, and ankle angles at bilateral take-off, and higher thigh, calf, and foot segmental velocities. The NMES group also showed a smaller shoulder-hip separation angle at bilateral take-off and ball contact. These findings indicate that acute NMES priming applied to upper-limb, core, and lower-limb muscles may influence phase-specific joint configurations, lower-limb segmental velocities, and ball velocity during volleyball spiking. NMES may therefore have potential as a pre-performance neuromuscular activation strategy, although further studies using concealed allocation, blinded assessment, and direct group-by-time analyses are required.
This study aimed to retrieve, appraise, and synthesize the best available evidence for life-sustaining treatment (LST) decision-making assessment in patients with advanced cancer and to explore its applicability within the Chinese clinical and cultural context. Systematic searches were conducted in BMJ Best Practice, UpToDate, DynaMed, JBI, GIN, NICE, SIGN, NCCN, Cochrane Library, MEDLINE, CINAHL, Embase, China National Knowledge Infrastructure, Wanfang Data, and SinoMed from March 3, 2019, to March 3, 2024, with additional retrospective searches. Eligible evidence sources included clinical decision-support resources, guidelines, expert consensus documents, evidence summaries, systematic reviews, and meta-analyses. Two reviewers independently screened the literature, appraised methodological quality, and extracted evidence, with disagreements resolved through consensus and, when necessary, adjudication by a senior third reviewer. Eight documents were included: two guidelines, one evidence summary, three clinical decision-support resources, and two systematic reviews. Nine evidence items were generated and assessed using the FAME framework, covering patient identification, prognosis and palliative care needs, family-system assessment, decision-making capacity, structured assessment tools, communication about LST preferences, documentation, and dynamic reassessment near the end of life. Evidence-based LST decision-making assessment may help reduce overtreatment and support high-quality end-of-life care for patients with advanced cancer. By translating the summarized evidence into a culturally adapted workflow, this study provides a practical reference for integrating patient preferences, family participation, and clinical assessment in Chinese palliative care practice.
Impedance-based technologies provide a robust and objective means to quantify cytopathic effects (CPE) in cell culture monolayers, overcoming the limitations of traditional endpoint or visually subjective assays. Cytolytic viruses typically induce pronounced morphological changes and cell death in infected cultures, making impedance measurements, reflecting changes in cell adhesion, morphology, and viability, a sensitive and dynamic indicator of viral infection and progression. As a result, impedance-based readouts offer a powerful and efficient approach for evaluating the protective effects of antiviral interventions. This study presents a real-time, label-free screening platform that leverages advanced impedance measurement systems to monitor virus-induced CPE continuously. This approach enables rapid, quantitative assessment of antiviral efficacy for both small-molecule drugs and monoclonal antibodies, without additional labeling or staining steps. The protocol details the complete assay workflow, from cell seeding and viral infection to data acquisition and analysis. Furthermore, the system-integrated software, specifically tailored for virology applications, streamlines data processing and interpretation, facilitating the determination of monoclonal antibody neutralizing potency and antiviral compound activity.
Deep vein thrombosis (DVT) is a common complication after orthopedic surgery. This prospective observational study evaluated whether a single preoperative baseline measurement of the venous circumference-squared-to-area ratio (C2/A), alone and with D-dimer, was associated with DVT detected during the first 7 postoperative days. One hundred fifty adults undergoing orthopedic surgery were enrolled; 37 developed DVT and 113 did not. Preoperative circumference (C) and area (A) were measured separately at end-expiration with the ultrasound system's built-in measurement package during the same acquisition cycle. Investigators calculated C2/A as C2 divided by A. To apply this rule consistently to the retained participant-level dataset, all revision analyses used C2/A recalculated from the retained C and A fields. Postoperative ultrasound was performed at 12-h intervals for 7 days under a prespecified intensified study-surveillance schedule used only for outcome ascertainment. The areas under the receiver operating characteristic curves were 0.893 for common femoral vein C2/A, 0.817 for superficial femoral vein C2/A, 0.906 for popliteal vein C2/A, and 0.834 for D-dimer. The originally specified superficial femoral vein C2/A plus D-dimer model had an apparent area under the curve of 0.887 and a stratified 5-fold cross-validated out-of-fold area under the curve of 0.877. A 5-variable exploratory model had an out-of-fold area under the curve of 0.938. Using cohort-derived thresholds of 16.26 for superficial femoral vein C2/A and 2.15 mg/L FEU for D-dimer, DVT occurred in 0 of 78 patients with neither marker elevated, 15 of 50 with one elevated, and 22 of 22 with both elevated. Preoperative venous C2/A and D-dimer were associated with early postoperative DVT, but the thresholds and models are exploratory and require external validation.
T cells play a key role in cancer immunotherapy, and understanding their interactions with tumors is essential for developing novel immunotherapies. This protocol describes the step-by-step workflow for establishing colorectal cancer patient-derived organoids (PDOs) from our biobank and developing autologous PDO-T cell co-culture systems. Using Annexin V NIR and cleaved caspase-3 staining, we standardized flow cytometry protocols for surface and intracellular staining to assess T cell reactivity and cytotoxicity, as well as tumor organoid killing. In addition to flow cytometry-based analysis, the protocol includes methodological approaches for functional assays, such as ELISpot assays to quantify granzyme B and perforin secretion, and live-cell imaging to monitor T cell-mediated tumor killing over time via Annexin V-based apoptosis detection. This workflow further details the setup of co-culture conditions, including the incorporation of target inhibitors and immune checkpoint inhibitors in both monotherapy and combination treatment settings. Guidelines are provided for selecting and applying key immunological and tumor-associated markers, including CD137 for T cell reactivity, CD107a for T cell degranulation, caspase-3 for tumor apoptosis, and Ki67 for tumor proliferation. Collectively, this paper provides a comprehensive and reproducible framework for colorectal cancer organoid establishment, autologous T cell culture and expansion, co-culture setup, therapeutic perturbations, and downstream analyses using flow cytometry, ELISpot, confocal microscopy, and live-cell imaging.
Perihilar cholangiocarcinoma is frequently diagnosed at an advanced stage and is often unresectable. Malignant biliary obstruction is a major cause of morbidity and commonly requires palliative biliary drainage. Endoscopic retrograde cholangiopancreatography (ERCP) with stent placement is widely used to relieve obstruction; however, tumor ingrowth and overgrowth frequently result in recurrent biliary stenosis and reduced long-term stent patency. Endoluminal radiofrequency ablation (eRFA) has emerged as an adjunctive treatment to improve local tumor control and maintain biliary drainage. Nevertheless, conventional eRFA is generally performed under fluoroscopic guidance alone, which may limit accurate delineation of tumor extent and precise energy delivery. This video article presents a single-case technical demonstration of a stepwise ERCP-based protocol integrating peroral cholangioscopy (POCS)-guided biliary mapping, targeted tissue sampling, precise eRFA, and self-expandable metal stent (SEMS) placement for unresectable perihilar cholangiocarcinoma. Direct visualization with POCS enables assessment of tumor morphology, identification of tumor boundaries, selective cannulation of involved biliary branches, and targeted biopsy acquisition. Information obtained during biliary mapping is subsequently used to define target ablation segments and guide controlled energy delivery. Following eRFA, SEMS are deployed to restore and maintain biliary drainage. This protocol provides a practical framework for visualization-guided endoscopic management of complex hilar biliary strictures and highlights key procedural steps that may improve treatment precision and procedural reproducibility in selected patients with unresectable perihilar cholangiocarcinoma.
From a neuropathological perspective, the most well-known pathological change in Alzheimer's disease (AD) to date is primarily tau protein hyperphosphorylation, which leads to the formation of intracellular neurofibrillary tangles (NFTs) and amyloid protein deposition-namely, the accumulation of amyloid protein (Aβ) and other protein aggregates outside cells, resulting in neurofibrillary tangles. The glymphatic system, driven by arterial pulsation, clears waste, including amyloid-β, via perivascular spaces (PVS). In AD, vascular dysfunction and slow-wave sleep speculation have been separately observed, and both are hypothesized to impair glymphatic clearance. However, causal links among these factors remain unproven, and the proposed self‑reinforcing cycle-where vascular failure may aggravate protein deposition and further vascular damage-is speculative. This study utilized two-dimensional cine phase-contrast MRI (PC-MRI) and four-dimensional flow MRI (4D-Flow) to quantify hydrodynamic parameters, including cerebrospinal fluid (CSF) flow in the cerebral aqueduct (CA) and blood flow in the internal carotid artery (ICA). Altered CSF-vascular association was observed in AD, suggesting possible glymphatic involvement. The integration of CSF and vascular flow metrics provides a promising biomarker framework for early AD detection and monitoring. PC-MRI and 4D-Flow reveal altered CSF-vascular association in Alzheimer's disease, reflecting potential glymphatic alteration and offering a non-invasive, quantitative method for potential adjunctive biomarker and disease monitoring.
