MethodsX

ECG waveform data, which is increasingly required for artificial intelligence analyses, is often unavailable in raw data formats. Digitization can approximate original raw data from ECG images, but digitizers reconstruct ECG waveforms only approximately. Exact waveform reconstruction is needed to guarantee that any errors, even if small, do not unexpectedly propagate into downstream analyses. Portable document format (PDF) files containing vector graphics store waveforms derived directly from raw data. This improves reconstruction accuracy, but existing PDF reconstruction methods remain approximate because raw data values are rounded during PDF encoding/generation. Despite rounding, however, there is a 1:1 mapping between raw data and PDF-encoded values which can be reversed to recover the original raw data with no error. We developed open-source software in MATLAB and Python (http://github.com/BIVectors/pdf2ECG) for bit-exact waveform reconstruction of General Electric MUSE ECG PDFs based on double quantization of ECG waveform data. The software supports multiple PDF layouts and voltage gains, automatically flags waveform clipping under which exact reconstruction is not possible, and certifies if parameter values used for waveform reconstruction are consistent with parameter values used during PDF encoding. Accuracy was assessed by comparing PDF reconstructed waveforms to raw XML waveforms, and reconstruction parameters were perturbed to determine the accuracy of waveform certification. 1,632 PDF-XML pairs were assessed in 5 layouts, each at 3 gains. After software-identified waveform clipping was excluded, the software reconstructed 100% of ECG waveforms with zero error compared to ground-truth XML. Certification tests were extremely sensitive to minor perturbation in encoding parameters, and if passed, confirmed bit-exact waveform reconstruction without need for ground-truth comparisons. Although this method does not replace the need for general ECG image digitization, for MUSE generated PDFs this method is preferable to rasterization followed by image-based digitization.

Abstract Background: Conventional cuff based methods report single systolic and diastolic pressure values and depend either on Korotkoff sounds or on empirical oscillometric algorithms. Methods: We developed a patented cuff based method, termed plethysmometry, that simultaneously records cuff pressure and finger photoplethysmograms from the cuffed and contralateral arms. The contralateral photoplethysmogram serves as an internal physiological reference for pulse amplitude. Following an initial pressure ramp, 16 cuff pressure plateaus are applied. Absolute and normalized pulse amplitudes in the cuffed arm are related to cuff pressure to estimate lower and upper systolic and diastolic pressures. Mean arterial pressure is estimated independently from cuff pressure oscillation amplitude. The estimations are done by automated algorithms developed in MATLAB. Results: The method reports lower and upper systolic, diastolic and mean arterial pressure ranges (during the recording period), together with their average values. Average pulse pressure is also reported. Graphs are provided for reference to check the accuracy of reported pressures. Conclusion: Plethysmometry provides an objective digital record of the cuff pressure and pulse traces and estimates systolic and diastolic pressure ranges from the pulse response of the individual subject to graded cuff pressure. It also reports mean arterial pressure. Comparison of plethysmometric systolic and diastolic pressures with sphygmomanometrically derived pressures and diagnostic agreement are reported in a companion paper.

The chick embryo is a widely used vertebrate model for developmental biology, cardiovascular, and bioengineering research. We present an automated laser speckle contrast imaging (LSCI) platform for non-invasive, longitudinal, label-free imaging of the developing heart and extraembryonic vasculature of the chick embryo through the intact eggshell. The platform can detect blood vessels as small as 70 +/- 17 um in diameter and enable repeated imaging from day 0 to day 8 of incubation without disturbing normal embryonic development. It provides both structural (a map of growing extraembryonic blood vessels) and functional (blood flow dynamics and heart rate) information. Using this system, we generated the first continuous, non-invasive, time-lapse recordings of the early functioning chick heart and its extraembryonic vessels. Longitudinal imaging was performed on 15 embryos from different chicken breeds with eggshell colors white, brown and green. Analysis revealed slight variability in early cardiac and vascular morphogenesis across breeds and shell types and shows its potential for automated developmental staging. Finally, for the first time, the growing vasculature of the yolk sac and the chorioallantoic (CAM) membrane was imaged from day 4 to 8. The heart rate and growth of the extraembryonic blood vessels were quantified automatically during imaging.

BackgroundTrunk flexor and extensor measurement tools are useful for measuring progress throughout the identification, treatment and management of spinal pathologies. Handheld dynamometry is popular amongst clinicians due to its convenience. However, poor standardisation of the testing position can lead to suboptimal intra-rater reliability, which consequently reduces the validity of the results. The VALD ForceFrame Strength Testing System is a portable and standardized hybrid strength-assessment system that quantifies strength and power outputs. ObjectiveThis study aims to assess the test-retest reliability of isometric trunk flexion and extension strength using the ForceFrame testing technology. MethodsA total of 29 healthy participants undertook two testing sessions, each requiring them to perform maximal isometric trunk flexion and extension using the ForceFrame. Test re-test reliability was assessed using the Intraclass Correlation Coefficient (ICC), and the standard error of measure (SEM) was used to calculate the minimum detectable change. ResultsExcellent reliability was observed for both trunk flexion (ICC 0.92; 95% CI 0.83-0.96) and extension (ICC 0.93; 95% CI 0.86-0.97) tests, and the minimum detectable changes observed were 71.35N and 101.20N, respectively. ConclusionsThe results from this study suggest that the VALD force frame offers clinicians a reliable method of assessing change in trunk flexor and extensor strength.

Abstract Background Advanced in vitro models of the human heart have become promising tools for studying cardiac diseases and drug discovery. Three-dimensional (3D) engineered heart tissues (EHTs) provide more adult cardiac tissue-like structural organization compared to traditional two-dimensional (2D) in vitro models with straightforward contractile readouts. However, the assessment of the architecture of EHTs, especially the quantification of cellular and tissue structural components, has remained poorly studied. Here we demonstrate the use of light-sheet microscopy for structural imaging of intact EHTs and provide quantification of the EHT cellular and subcellular structures. Methods We developed an in-house inverted selective plane illumination microscopy (iSPIM) platform and combined it with iterative indirect immunofluorescence (IT-IF) staining for better signal acquisition. We developed an image-processing and analysis pipeline dedicated to volumetric EHT analysis. Results With this integrated platform, we achieved volumetric imaging of intact EHTs and quantification of 3D tissue architecture, revealing aligned cardiomyocytes within the imaged volume and increasing cardiac nuclei content towards the core of the tissue. The developed methods allowed the detection of heterogeneities between tissue sections, demonstrating the sensitivity required for future disease modeling or maturation studies. Conclusions The developed integrated platform opens new possibilities for volumetric analysis and characterization of 3D tissue structure using iSPIM, providing new insights into EHT architecture and the use of EHTs as disease modeling tools.

Background: Temporomandibular disorders (TMDs) are common conditions in the general population. Several therapeutic approaches are available, but there is no consensus on the most effective, particularly in physiotherapy. Objectives: To assess the efficacy of manual therapy (MT) and therapeutic exercise (TE) in reducing pain in patients with TMDs. Methods: Four databases (MEDLINE, PEDro, EMBASE, CINAHL) were searched up to March 2025. We included randomized controlled trials (RCTs) evaluating the effects of MT and/or TE in individuals with TMDs, compared to any other rehabilitation treatment, with pain intensity as the primary outcome. The methodological quality was assessed using the PEDro scale. A network meta-analysis was performed to compare the effectiveness of different rehabilitative approaches. Results: 25 RCTs for a total of 1,540 people were included. The main comparisons were: MT and/or TE vs no treatment or placebo, occlusal splint, and instrumental physiotherapy. In most studies, MT and TE were more effective than controls in reducing pain. However, the network meta-analysis indicated that multimodal treatments were the most effective. The methodological quality of the included studies ranged from 3 to 9 points. Conclusion: MT and TE are effective strategies for reducing pain in subjects with TMDs and should be used as first-line interventions before more invasive treatments; the combination of dry needling and manual therapy appears to be the most effective approach. Nonetheless, the variability in study quality suggests that further high-quality research is needed to confirm these findings.

Objective To determine the comparative effectiveness of different digital health interventions for adults with low back pain. Design Systematic review and network meta-analysis of randomised controlled trials. Data sources PubMed, Embase, the Cochrane Central Register of Controlled Trials (CENTRAL), CINAHL, APA PsycINFO, ClinicalTrials.gov, and the World Health Organization International Clinical Trials Registry Platform (WHO ICTRP), from inception to 19 March 2026 (updated to 8 August 2026). Study selection Randomised controlled trials enrolling adults with low back pain. Interventions were classified as telemedicine, web-based applications (Web-App), mobile applications (Mobile-App), virtual reality (VR), exergaming, biofeedback devices, face-to-face care, and control. Paired reviewers independently screened records, extracted data, and assessed risk of bias using the Cochrane Risk of Bias 2 tool. Methods Frequentist random-effects network meta-analyses synthesised pain intensity and physical function (primary outcomes), and pain-related fear avoidance, health-related quality of life, self-efficacy, anxiety, and depression (secondary outcomes), assessed at post-intervention (from the end of treatment to < 2 months post-intervention), short-term ([&ge;]2 to <6 months), mid-term ([&ge;]6 to <12 months), and long-term ([&ge;]12 months) follow-up. Effects were reported as standardised mean differences with 95% confidence intervals. Transitivity was examined across potential effect modifiers, and Confidence in Network Meta-Analysis (CINeMA) informed confidence ratings. Results Ninety-two reports representing 84 trials involving 11 649 participants were included. At post-intervention, compared with control, VR showed the largest estimated reduction in pain intensity (SMD -1.43, 95% CI -1.87 to -0.98; moderate confidence) and improvement in physical function (-0.86, -1.45 to -0.26; very low confidence). VR was also associated with lower pain-related fear avoidance (-1.14, -1.68 to -0.61) and better health-related quality of life (0.97, 0.57 to 1.36) (both moderate confidence). Exergaming, Mobile-App, Web-App and telemedicine were also associated with lower pain intensity, while Mobile-App, Web-App, telemedicine, and biofeedback devices were associated with better physical function. For self-efficacy, exergaming (0.77, 0.05 to 1.49) showed favourable post-intervention effects, although confidence in this estimate was very low. No intervention showed a clear effect on anxiety, with confidence ranging from very low to moderate across comparisons, whereas telemedicine was associated with lower depression (-0.22, -0.44 to 0.00; low confidence). Networks became progressively sparse at later follow-up, and evidence of sustained effects was inconsistent across interventions and outcomes. Global inconsistency was detected in the post-intervention pain-intensity network, with local inconsistency identified for selected exergaming comparisons, and substantial heterogeneity was present in several networks. Conclusions Among adults with low back pain, VR showed the largest estimated post-intervention effects versus control for pain intensity, physical function, pain-related fear avoidance, and health-related quality of life, although comparative effects differed across outcomes. Self-efficacy showed favourable estimates for selected interventions, whereas evidence for anxiety and depression was limited. Evidence beyond treatment completion was increasingly sparse, and confidence in many network estimates was low or very low; firm comparative recommendations therefore remain premature. Systematic review registration PROSPERO CRD420261339841

Aim. To evaluate the accuracy, run-to-run consistency, and agreement of large language models on the Ukrainian Integrated Licensing Examination Krok 2 for the Specialty of Physical Therapy. Materials and Methods. Five cloud-based LLMs (ChatGPT-5.5, Claude 5 Sonnet, Gemini 3.5 Flash, Grok 4, DeepSeek-V3) were evaluated using 150 Ukrainian-language multiple-choice questions from the official Krok 2 database. Each model completed two independent runs with identical inputs. Accuracy (with 95% confidence intervals, Wilson method), run-to-run consistency, and agreement were assessed. Statistical analysis included Cohen's {kappa}, Cochran's Q and pairwise McNemar tests with Holm-Bonferroni correction. Results. All models achieved >80% accuracy in both runs, exceeding the passing threshold of [&ge;]64%. In the first run, accuracy ranged from 80.00% (DeepSeek-V3) to 93.33% (Gemini 3.5 Flash); in the second run, from 82.67% (Claude 5 Sonnet) to 88.67% (Gemini 3.5 Flash). In the first run significant differences were observed between Gemini 3.5 Flash and Grok 4 (93.33% vs 84.00%; p=0.0022) and DeepSeek-V3 (93.33% vs 80.00%; p=0.0001). There were no statistically significant differences between models in the second run. R2R consistency exceeded 90% for all models with the highest in ChatGPT-5.5 (98.67%) and lowest in DeepSeek-V3 (90.00%). Cohen's {kappa} ranged from 0.64 to 0.94, indicating substantial to almost perfect agreement. The stability of LLMs' response correctness across two runs ranged from 77.33% (DeepSeek-V3) to 88.00% (Gemini 3.5 Flash). Conclusions. All LLMs demonstrated high performance on Ukrainian-language Krok 2 examination tasks. Gemini 3.5 Flash showed the highest accuracy in both runs and response correctness stability while ChatGPT-5.5 exhibited the greatest response consistency and agreement. Although the accuracy and stability of LLM responses may be interrelated, they represent distinct aspects of model performance. This highlights the need for a comprehensive approach to evaluating LLMs. The findings indicate the potential of LLMs as an auxiliary tool in the professional practice of physical therapy specialists.

Background: Long COVID can result in persistent functional limitations, pain, and fatigue after acute COVID-19. Evidence describing these outcomes among survivors in Bangladesh remains limited. This study assessed functional impairment, pain, and fatigue and explored factors associated with these outcomes among long COVID survivors in Bangladesh. Methods: This multisite cross-sectional study included 240 adults with a history of COVID-19. Functional status was assessed using the Post-COVID-19 Functional Status Scale. Pain and fatigue were assessed using the Brief Pain Inventory and Brief Fatigue Inventory, respectively. Descriptive statistics, Spearman correlation, and regression analyses were used to examine functional impairment and factors associated with pain and fatigue outcomes. Results: Functional limitations were common, with moderate limitation particularly frequent in basic activities of daily living, while slight limitation predominated across several other domains. Pain and fatigue measures showed significant correlations. Older age was associated with greater impairment in basic activities of daily living. Booster vaccination was associated with lower pain and fatigue scores, although the booster subgroup was small (n = 17). Tobacco non-use was also associated with lower scores across several pain and fatigue domains. Physiotherapy-related variables showed associations with some functional and symptom outcomes; however, these findings may reflect confounding by indication because participants with greater impairment or more complex symptoms may have been more likely to receive rehabilitation. Conclusions: Long COVID survivors in Bangladesh experience substantial and multidimensional functional, pain, and fatigue burdens. The findings support a comprehensive assessment that considers functional status, pain, fatigue, and participation together. Longitudinal studies with larger samples and standardized rehabilitation protocols are needed to clarify symptom trajectories, identify potentially modifiable factors, and evaluate the effectiveness of individualized rehabilitation interventions.

Objective: To examine changes in function and pain interference among patients utilizing a digital exercise therapy application (DETA) for knee and shoulder pain. Methods: We conducted a retrospective analysis of data from patients utilizing a DETA for non-operative knee and shoulder pain. Primary outcomes for the study were 8-week changes in physical function, measured using the PROMIS-Physical Function (knee pain) and PROMIS-Upper Extremity Function (shoulder pain); and pain interference, measured using the PROMIS-Pain Interference. Data on patient characteristics was used to examine outcomes among specific patient subgroups. Results: We observed a 3.5 point (SD: 7.8) increase in function and a 3.9 point (SD: 7.5) decrease in pain interference overall (p<0.001), with changes in function and pain interference surpassing the minimal clinically important difference for patients with knee and shoulder pain. Patients with knee pain experienced greater improvements in function than those with shoulder pain (p=0.02), as did patients who report moderate-high levels of exertion during exercise compared to those reporting low levels of exertion (p=0.05). We saw greater reductions in pain interference among those with higher education (p=0.03), those with more acute symptoms (p=0.01) and those reporting moderate-high levels of exercise exertion (p=0.02). Conclusion: We observed a clinically meaningful improvement in pain interference and function among patients utilizing a DETA for knee and shoulder pain. Patients with knee pain, higher levels of education, more acute symptoms, and those who reported higher levels of exertion with exercise experienced the greatest changes in function and pain. Impact Statement: This study indicates that self-guided interventions, such as those delivered through a DETA, may be beneficial for patients with knee and shoulder pain. These results may also inform the development of patient profiles that can be used to identify patients most appropriate to receive care through a mobile application.

Optical mapping is a fundamental tool for cardiac electrophysiology research. Processing of these signals can be non-trivial, requiring multiple pre- and post-processing steps to limit interpretation errors. Existing software packages have features siloed across platforms and require users to combine several packages for in-depth analysis. To address this, we developed CADENCE, an open-source MATLAB-based application combining feature breadth (e.g., SVD denoising, conduction velocity, alternans, annotated rotors) and depth (e.g., 15 independent alternans features across both voltage and calcium modalities) in an end-to-end pipeline including data conversion, signal conditioning, feature extraction, and signal analysis. We demonstrate CADENCE's capabilities across datasets spanning four species (rat, rabbit, pig, and human), three tissue preparations (slices, wedge, and whole heart), and three camera configurations (single, dual, and panoramic). CADENCE was validated against three blinded expert observers (15 hearts) and demonstrated agreement comparable to inter-observer variability (SD ratios 1.07/0.95/0.89 for APD80, activation, CaTD80). Dominant frequency was recovered with a median error of 0.03% across 84 of 88 paced recordings with 1:1 capture. CADENCE's code is publicly available and distributed as a compiled standalone application that the end-user can install and run without a MATLAB license to encourage adoption and standardization across laboratories.

Albumin binding can substantially alter the unbound concentration and apparent cellular potency of small-molecule anticancer agents. BAY-1816032 is a selective BUB1 kinase inhibitor with activity in malignant pleural mesothelioma, but its interaction with serum albumin has not been defined. This study combined intrinsic bovine serum albumin (BSA) fluorescence measurements, molecular docking, and cell-viability assays to examine whether albumin association influences BAY-1816032 activity. BAY-1816032 produced pronounced quenching of BSA fluorescence at low drug-to-protein molar ratios, and the quenching response was greater at mildly acidic pH than at mildly basic pH. Docking to BSA (PDB 4F5S) placed the top-ranked pose in Sudlow site I (subdomain IIA), with a Vina score of -7.8 kcal/mol and predicted hydrogen-bond contacts with Tyr149, Arg256, and Asp450. In NCI-H28, NCI-H2052, NCI-H2452, and AB12 mesothelioma cells, supplementation of 10% fetal bovine serum medium with 2% (w/v; 20 mg/mL) BSA shifted BAY-1816032 concentration-response curves to the right. Relative to 10% serum alone, IC50 values increased from 2.4 to 11.1 uM in H28, 3.6 to 22.0 uM in H2052, 2.0 to 4.9 uM in H2452, and 2.4 to 11.1 uM in AB12. These data support an albumin-dependent reduction in the freely available drug fraction and identify site I as the leading computational binding-site hypothesis. Because fluorescence-intensity and docking data do not independently establish a dissociation constant or quenching mechanism, direct unbound-fraction measurements and competitive displacement experiments are required for quantitative confirmation.

BACKGROUND: The protein 4.1a/4.1b ratio is an intrinsic molecular marker of mean red blood cell (RBC) age resulting from progressive deamidation of protein 4.1b during RBC circulation. Despite its potential for estimating mean RBC age from a single blood sample, its application has been limited by labor-intensive RBC membrane preparation, and its interindividual variability has not been characterized in a large human population. METHODS: We developed a low-volume method for direct RBC membrane isolation from whole blood using saponin permeabilization and differential centrifugation without prior RBC washing. Membrane proteins were separated by SDS-PAGE, and protein 4.1a/4.1b ratios were determined by densitometric analysis with Gaussian peak fitting. Assay conditions, sample-volume requirements, membrane protein recovery, and analytical reproducibility were evaluated. The method was subsequently applied to 114 anonymized residual samples from routine diagnostic testing selected for complete blood count parameters within the corresponding laboratory reference intervals. RESULTS: The optimized procedure used 50 uL of whole blood, 0.06% saponin, and centrifugation at 9000 g, although protein 4.1a/4.1b remained measurable from starting volumes as low as 5 uL. Mean within-subject CVs were 3.90% for electrophoretic/densitometric determination and 5.39% for membrane protein recovery; whole-procedure CVs in 3 subjects were 1.40% to 3.50%. In the population analysis, the mean protein 4.1a/4.1b ratio was 1.428 (SD 0.146), with an observed range of 1.052 to1.799 and a central 95% interval of 1.151 to 1.695. Ratios did not differ significantly between females and males (P=0.246) or among females aged 18 to 50 years, females aged >50 years, and males (P=0.151). The ratio showed modest positive correlations with chronological age in females (Spearman p=0.26, P=0.047) and males (p=0.33, P=0.015), while substantial interindividual variability remained across the age range. CONCLUSIONS: Direct isolation of RBC membranes from small volumes of whole blood enables reproducible determination of the protein 4.1a/4.1b ratio using a simplified workflow. Application of the assay to a comparatively large human population revealed substantial interindividual variation in this molecular marker of mean RBC age that was not explained by sex or chronological age alone. The method provides a practical approach for investigating biological variation in mean RBC age and its potential contribution to laboratory measurements influenced by RBC survival.

Background: Endometriosis is an estrogen-driven, inflammatory disorder affecting ~10% of menstruators, causing severe pain. Current treatments that reduce estrogen or inflammation have inconsistent efficacy and poor tolerability. Therapies identified via drug-repositioning methods, which target patient-specific pathways, offer a promising alternative. Simvastatin and primaquine were identified as potential treatments, strongly reversing endometriosis-associated gene expression pathways, and behavioral testing in an animal model showed that these drugs diminish endometriosis-associated pain. This paper examines their effects in human assembloids to provide proof-of-concept validation and insights into their mechanisms of action relevant to endometriosis. Methods: We established human endometrial assembloids using immortalized epithelial (12Z) and stromal fibroblast (iEc) cell lines, as well as primary tissue from patients. Assembloids were generated in 96-well agarose molds with both cell types. We tested ibuprofen, fenoprofen, primaquine, and simvastatin. Following a 24-hour exposure, assembloids and monolayer cultures were harvested and stranded mRNA-seq libraries were sequenced on an Illumina NovaSeqX Plus System. Results: Gene set enrichment analysis showed key pathway reversals: Primaquine reversed the chemical carcinogenesis ROS pathway in assembloids. Simvastatin reversed cytokine-cytokine receptor interaction in both epithelial and stromal cell lines, and the cytoskeleton in muscle cells pathway in stromal cell lines. Fenoprofen reversed the calcium signaling pathway in stromal cell lines. Conclusion: Simvastatin induced the most significant gene expression changes, notably reversing cytokine-cytokine receptor interactions in both cell lines, mirroring findings from our rat model. Despite the small sample size limitation, these experiments highlight the promise of assembloid models to test therapeutic candidates for endometriosis.

Context: Cytological specimens allow minimally invasive, repeated sampling for diagnosis and molecular testing. However, cytology biobanking protocols have primarily been optimized for morphology, and the effects of preanalytical handling and storage on intracellular proteins remain unclear. Objective: To assess how fixation, staining, storage temperature, and duration affect intracellular protein preservation in cytological specimens and to identify conditions suitable for in situ proximity ligation assay (isPLA). Design: Human cancer cell lines were processed under different preanalytical conditions. Intracellular proteins and protein protein interactions were assessed by isPLA, fluorescence microscopy, and quantitative image analysis. Slides were stored at room temperature or minus 20 Celsius, with or without MayGrunwaldGiemsa (MGG) staining. Results: Storage temperature had the greatest effect on protein signal. Unstained slides stored at minus 20 Celsius retained strong signals for up to 24 months, whereas room temperature storage reduced detectable protein interactions by approximately 95%. MGG staining further reduced signal intensity during prolonged storage. Nevertheless, stained slides remained suitable for isPLA when staining, microscopic evaluation, destaining, and antibody labeling were completed on the same day. Prolonged room temperature storage of stained slides caused loss of antigenicity, increased autofluorescence, and false negative findings. Conclusions: Low temperature storage and standardized preanalytical handling preserve intracellular protein signals in cytological specimens. These conditions support reliable single cell protein profiling of archived samples and their use in biomarker research and translational pathology.

Background: Localized or segmental overgrowth is a rare condition that may co-occur with vascular malformations. These diseases generally are caused by somatic mutations in the PI3K/AKT/mTOR pathway. Most lesions are progressive and may cause symptoms including pain, functional impairment, and disfigurement. Treatment options are limited, especially for the overgrowth component. Objective: To evaluate the efficacy and safety of the mTOR inhibitor sirolimus in patients with overgrowth with or without a vascular malformation over a 6-months treatment period. Methods: We conducted a single-arm, open-label, phase II study including 18 patients (12 pediatric, six adult) with overgrowth, all of whom initiated oral sirolimus. 16/18 (88.9%) participants received oral sirolimus for 6 months, with dosing adjusted to achieve target blood trough levels of 3-8 ng/mL. Primary outcome was radiological response at six months assessed by the Response Evaluation Criteria in Solid Tumors (RECIST) 1.1. Secondary outcome parameters were evaluated at three, six and nine months and included size and visual appearance of the overgrowth, quality of life (QoL), pain intensity, performance status, and safety. Results: No patient achieved complete or partial radiological response. 17 of 18 patients (94.4%) maintained stable disease at month six; one was not evaluable. QoL measured with WHOQOL-BREF improved modestly in patients aged >= 18 years (n=5, mean difference to baseline +9.3 (95%-CI: -0.9 to 19.5) in physical and +9.2 (95%-CI: 0.7 to 17.7) in psychological domains at month 6). In pediatric participants, no relevant improvement in KINDL total scores was observed over the study period. No significant changes occurred in pain scores. Performance status measured by the Lansky/Karnofsky scales showed minor improvement (mean +3.5 points (95%-CI: 1.0 to 6.1)). All patients experienced at least one adverse event, but no serious adverse events or CTCAE v5.0 grade [&ge;]3 toxicities occurred. Sirolimus was well tolerated. Six patients decided to resume sirolimus after study completion. Conclusions: Sirolimus induced minor lesion shrinkage in a subset of patients, although this did not meet the predefined radiological RECIST endpoint. However, sirolimus can lead to functional improvement in some adults, which may indicate that the primary radiological outcome did not capture patient-perceived benefit.

BackgroundNeuropathic pain is a common and debilitating condition with persistently poor translation from preclinical research to effective clinical therapies. Preclinical systematic reviews and meta-analyses (SRMAs) can provide more robust estimates of intervention effects and help identify sources of bias and heterogeneity, but their use is limited by the time and effort required to synthesise rapidly expanding literatures. Automation may help overcome these barriers. MethodsWe developed Neuropathic Pain - Systematic Online Living Evidence Summaries (NP-SOLES), a machine-assisted approach used in this study to support systematic identification of animal studies relevant to neuropathic pain. Using NP-SOLES, we conducted a pilot systematic review and meta-analysis of controlled animal studies assessing the effects of citrus flavonoids on thermal and mechanical pain-related outcomes in streptozotocin-induced models of painful diabetic neuropathy. ResultsIn this pilot application, machine-assisted evidence synthesis supported reduced manual screening burden, illustrating the potential of such approaches to facilitate timely preclinical SRMAs. Fourteen studies met inclusion criteria for meta-analysis on thermal pain-related outcomes. Pooled estimates suggested a reduction in thermal pain-related behaviours following flavonoid treatment; however, substantial heterogeneity and widespread methodological limitations were observed. Nine studies were eligible for qualitative analysis on mechanical pain-related outcomes, which showed an overall reduction in pain-related behaviours following treatment but had similar methodological limitations. Reporting of key measures to reduce risk of bias, including randomisation and blinding, were consistently poor, and subgroup analyses were constrained by sparse data. ConclusionsAvailable evidence from preclinical animal studies suggests a potential analgesic effect of citrus flavonoids on thermal and mechanical pain-related outcomes in diabetic neuropathy models, but confidence in this estimate is limited by significant risks of bias. More rigorously designed and transparently reported preclinical studies are required before clinical translation can be justified.

The field of microglial research has evolved throughout the years. Microglia, the immune cells of the central nervous system, are recognized as highly heterogeneous and dynamic cells, known for modifying their structure and function based on the local context. Investigating microglial density, spatial distribution, and morphological states is important for uncovering their distinct functional states in health and pathology. However, quantifying these features accurately is often a methodological challenge. Fully automated computational approaches often fail to capture subtle biological nuances and complex structural variations. Also, the high diversity of image sets makes it difficult to maintain consistent reliability. In contrast, entirely manual quantification is labor-intensive and prone to observer bias. To bridge this gap, we propose a semi-automated framework using the open-source software CellProfiler. Our workflow is divided into two distinct pipelines designed to combine automation of batch analysis with targeted user oversight, allowing for manual intervention when necessary to ensure maximum accuracy. Both pipelines are capable of recognizing microglial soma and tracing their processes. In the density workflow, it automatically calculates cell density and provides spatial distribution measurements, such as closest-neighbor distance and spacing index. For morphological profiling, it yields extensive structural data, including area, perimeter, convex area, form factor, and various shape descriptors. Furthermore, we demonstrate how researchers can optimize the pipeline settings to accommodate varied image datasets and experimental conditions. We hope this open-source framework can standardize microglial density, distribution, and morphological analysis and reduce systematic bias, providing researchers with a robust tool to better characterize their heterogeneity.

Background: System One Model is designed for structured decision-making and can provide probabilistic outputs, but their performance in domain-specific physical therapy tasks has not been established. Objective: To evaluate the performance and potential utility of Jev, a System One Model, for professional knowledge-based selection tasks in physical therapy by comparing it with GPT-4o. Methods: A total of 380 multiple-choice questions from the 2024 and 2025 Korean Physical Therapist Licensing Examinations were independently submitted to Jev and GPT-4o. The outcomes included answer accuracy, subgroup performance, output validity, API response latency, token usage, estimated API cost, and Jev's prediction uncertainty. A retrospective probability-based model-cascading simulation was also performed. Results: Jev correctly answered 282 questions (74.2%), whereas GPT-4o answered 326 (85.8%), a difference of -11.6 percentage points. Jev produced valid structured responses for all questions, whereas GPT-4o produced three invalid responses. Jev's selected-answer probabilities discriminated correct from incorrect answers (AUROC = 0.866), and accuracy reached 99.3% when probabilities were greater than or equal to 0.90. The median API response latency was 0.94 s for Jev and 1.06 s for GPT-4o, with estimated costs of US$0.008 and US$0.161, respectively. At a retrospective cascading threshold of 0.70, accuracy was 86.6% with 175 simulated GPT-4o calls. Conclusion: Jev showed lower overall accuracy than GPT-4o but provided reliable structured outputs, informative prediction probabilities, lower latency, and lower estimated cost. Its probabilistic outputs warrant further prospective evaluation for selective model escalation and decision-support workflows in physical therapy.

Objective: The aim of the current study was to evaluate the feasibility of a multi-channel distributed wearable ultrasound (D-WUS) system capable of simultaneously probing dynamic muscle activity across multiple anatomical sites to assess neuromuscular coordination during physical activity. Methods: Twenty NCAA Division I women athletes (8 with prior knee injury) performed countermovement jumps (CMJs) on dual force plates while wearing a four-channel D-WUS system targeting the vastus lateralis (VL) and vastus medialis oblique (VMO) in both limbs. Real-time M-mode ultrasound data were processed using a composite factorization approach integrating nonnegative matrix factorization and principal component analysis (CF-PCA) to extract dominant muscle-contraction patterns. Correlation and timing metrics between VL and VMO were compared across injury status. Results: CMJ performance metrics (jump height, RSImod) did not differ between groups (p > 0.05). However, D-WUS revealed significant differences in muscle coordination in the right limb: VL-VMO correlation (p = 0.029) and timing symmetry (p = 0.019) were reduced in participants with prior knee injury; left-limb differences were not significant. Right-limb timing symmetry was negatively correlated with SMFA score (rho = -0.48, p = 0.033). Functional survey scores (SMFA) also differed significantly (p = 0.001), while VISA-P scores did not. Conclusion: These preliminary findings provide feasibility-level evidence that distributed wearable ultrasound can detect limb-specific neuromuscular coordination differences not reflected in force-plate biomechanics, motivating larger prospective studies to establish reliability and clinical utility. Significance: With further validation, this approach may offer a scalable, objective means of monitoring muscle contraction during rehabilitation.