Search Results (171)

Extracellular vesicles (EVs), nanoscale membrane-enclosed particles, are natural carriers of proteins and nucleic acids. Microalgae are widely used as a source of bioactive substances in the food and cosmetic industries and definitely have a potential to be used as the producers of EVs for biomedical applications. In this study, the extracellular vesicles isolated from the culture medium of two unicellular microalgae, Chlamydomonas reinhardtii (Chlamy-EVs) and Parachlorella kessleri (Chlore-EVs), were characterized by atomic force microscopy (AFM), cryo-electronic microscopy (cryo-EM), and nanoparticle tracking analysis (NTA). The biocompatibility with human cells in vitro (HEK-293T, DF-2 and A172) and biodistribution in mouse organs and tissues in vivo were tested for both microalgal EVs. An exogenous therapeutic protein, human heat shock protein 70 (HSP70), was successfully loaded to Chlamy- and Chlore-EVs, and its efficient delivery to human glioma and colon carcinoma cell lines has been confirmed. Additionally, in order to search for potential therapeutic biomolecules within the EVs, their proteomes have been characterized. A total of 105 proteins were identified for Chlamy-EVs and 33 for Chlore-EVs. The presence of superoxide dismutase and catalase in the Chlamy-EV constituents allows for considering them as antioxidant agents. The effective delivery of exogenous cargo to human cells and the possibility of the particle yield optimization by varying the microalgae growth conditions make them favorable producers of EVs for biotechnology and biomedical application. Full article

Lou Gehrig’s disease, also known as ALS, is a progressive neurodegenerative condition that weakens muscles and can lead to paralysis as it progresses. For patients with severe paralysis, eye-tracking devices such as eye mouse enable communication. However, the equipment is expensive, and the calibration process is very difficult and frustrating for patients to use. To alleviate this problem, we propose a simple and efficient method to type texts intuitively with graphical guidance on the screen. Specifically, the method detects patients’ eye blinks in video frames to navigate through three sequential steps, narrowing down the choices from 9 letters, to 3 letters, and finally to a single letter (from a 26-letter alphabet). In this way, a patient is able to rapidly type a letter of the alphabet by blinking a minimum of three times and a maximum of nine times. The proposed method integrates an API of large language model (LLM) to further accelerate text input and correct sentences in terms of typographical errors, spacing, and upper/lower case. Experiments on ten participants demonstrate that the proposed method significantly outperforms three state-of-the-art methods in both typing speed and typing accuracy, without requiring any calibration process. Full article

Lactoferrin (LF), a multifunctional glycoprotein found abundantly in bovine colostrum, is known for its regenerative and anti-inflammatory properties. In this study, we investigated the wound healing and immunomodulatory effects of colostrum-derived exosome-encapsulated lactoferrin (EV-exoLF) on dermal fibroblasts. EV-exoLF was isolated and characterized via nanoparticle tracking analysis and flow cytometry. Functional assays demonstrated that EV-exoLF significantly promoted fibroblast viability and migration in both mouse NIH/3T3 and human HS-68 cell lines. Furthermore, EV-exoLF reduced the expression of pro-inflammatory cytokines (IL-1 and IL-6) and phosphorylated JNK in lipopolysaccharide (LPS)-treated fibroblasts. These findings suggest that EV-exoLF not only enhances fibroblast-mediated wound closure but also mitigates inflammation, highlighting its therapeutic potential in skin regeneration. Colostrum-derived exosomal lactoferrin may serve as a promising natural, cell-free strategy for managing inflammatory skin conditions and improving wound healing outcomes. Full article

Tracking T cells in vivo using MRI is a major challenge due to the difficulty of labeling these non-phagocytic cells with a sufficient contrast agent to generate a detectable signal change. In this study, we explored CD4-Superparamagnetic iron oxide nanoparticles (SPION), which is commonly used in magnetic cell sorting, as a potential receptor-mediated, specific CD4⁺ T cell MRI labeling agent. We optimized the labeling protocol for maximal CD4⁺ cell labeling and viability. Cell health was confirmed with trypan blue assay, and labeling efficacy was confirmed with Prussian blue staining, transmission electron microscopy, and MRI of labeled cell pellets. Key cell functionality was assessed by flow cytometry. Next, CD4-SPION-labeled T cells or unlabeled T cells were delivered via intravenous injection in naïve mice. Liver MRIs pre-, 24 h, and 72 h post-T cell injection were performed to determine in vivo tracking ability. Our results show that CD4-SPION induces significant attenuation of T2 signals in a concentration-dependent manner, confirming their potential as an effective MRI contrast agent. In vitro, analyses showed that CD4⁺ T cells were able to uptake CD4-SPION without affecting cellular activity and key functions, as evidenced by Prussian blue staining and flow cytometric analysis of IL-2 receptor and the IL-7 receptor α-chains, CD69 upregulation, and IFN-γ secretion. In vivo, systemically distributed CD4-SPION-labeled T cells could be tracked in the liver at 24 and 72 h after injection, contrary to controls. Histological staining of tissue sections validated the findings. Our results showed that SPION CD4⁺ T cell sorting coupled with longitudinal MR imaging is a valid method to track CD4⁺ T cells in vivo. This safe, specific, and sensitive approach will facilitate the use of SPION as an MRI contrast agent in clinical practice, allowing for non-invasive tracking of adoptive cell therapies in multiple disease conditions. Full article

Extracellular vesicles (EVs) are lipid membrane-enclosed particles released by all cells and can be isolated from various sources, even from solid tissues. This study focuses on isolating and characterizing EVs from mouse lymph nodes (LNs). Male C57BL/6 mice were injected with complete Freund’s adjuvant, with or without ovalbumin. Inguinal and popliteal LNs were incised 9 days after immunization, and EV isolation was carried out using a combination of differential centrifugation and size-exclusion chromatography. The characteristic morphology of small and large EVs was confirmed by transmission electron microscopy. Particle size distribution and concentration were determined by nanoparticle tracking analysis, while protein and lipid contents were measured by bicinchoninic acid assay, and sulfo-phospho-vanillin assays, respectively, to calculate the protein-to-lipid ratio. Immune and EV markers were analyzed by using flow cytometry and Western blot assay, revealing significant changes between immunized mice compared to controls. This study establishes a novel protocol for isolating and characterizing EVs from LNs and highlights the impact of immunization on EV properties, offering insights into their roles in immune processes. Full article

Echocardiography is a cornerstone technique for evaluating cardiac function in preclinical research using murine models. This review provides a comprehensive overview of the echocardiographic approaches employed to assess ventricular function in mouse models of heart disease, highlighting methodological principles, technical challenges, and the translational relevance of findings. Various echocardiographic modalities enable the precise evaluation of systolic and diastolic function. This article emphasizes standardization in image acquisition and analysis to minimize inter-operator variability and ensure reproducibility. It details echocardiographic parameters and strain imaging across commonly used mouse models of non-ischemic dilated cardiomyopathy, diabetic cardiomyopathy, hypertensive heart disease, and ischemic heart disease. Furthermore, it explores the advantages and limitations of anesthesia, probe positioning, and physiological monitoring during imaging. The integration of advanced imaging technologies such as Speckle-Tracking Echocardiography (STE), Three-Dimensional (3-D), and Four-Dimensional (4-D) echocardiography is discussed as a promising avenue for enhancing data quality and improving the translational potential of preclinical cardiac studies. Full article

Background/Objectives: Cell-based therapies have become increasingly important in the treatment of cancers and inflammatory diseases; however, therapies utilizing monocyte–macrophage lineage cells remain relatively underexplored. Non-invasive cell tracking allows a better understanding of the fate of such cells, which is essential for leveraging their therapeutic potential. Here, we employed a Zirconium-89 (⁸⁹Zr)-oxine cell labeling method to compare the trafficking of monocytes and macrophages in vivo. Methods: Mouse bone marrow-derived monocytes and macrophages were each labeled with ⁸⁹Zr-oxine and evaluated for their viability, radioactivity retention, chemotaxis, and phagocytic function in vitro. Labeled cells were intravenously administered to healthy mice and to murine models of granuloma and syngeneic tumors. Cell migration was monitored using microPET/CT, while cell recruitment to the lesions was further assessed via ex vivo biodistribution and flow cytometry. Results: Labeled cells exhibited similar survival and proliferation to unlabeled cells for up to 7 days in culture. While both maintained phagocytic function, monocytes showed higher CCL2-driven chemotaxis compared to macrophages. ⁸⁹Zr-oxine PET revealed initial cell accumulation in the lungs, followed by their homing to the liver and spleen within 2–24 h, persisting through the 5-day observation period. Notably, monocytes trafficked to the liver and spleen more rapidly than macrophages. In both inflammation and cancer models, monocytes demonstrated higher accumulation at the lesion sites compared to macrophages. Conclusions: This study demonstrates the usefulness of ⁸⁹Zr-oxine PET in tracking monocyte–macrophage lineage cells, highlighting their distinct migration patterns and providing insights that could advance monocyte-centered cell therapies. Full article

Although prior studies have identified cumulus cells (CCs)-expressed genes and miRNAs that regulate cumulus expansion and/or CC apoptosis and may serve as markers for selecting competent oocytes and embryos, there remains an urgent need to identify CCs-expressed genes and miRNAs whose expression levels are directly correlated with oocyte developmental potential (DP). In this study, we first established CC models from mouse cumulus-oocyte complexes (COCs) that exhibited significantly different DP following in vitro or in vivo maturation. Subsequently, we performed mRNA/miRNA sequencing and functional analyses using these in vitro and in vivo CC models. We identified and validated Spp1, Fn1, Sdc1, and Ngf as DP-beneficial genes; Fos and Jun as DP-detrimental genes; and miR-7686-5p, miR-133a-3p, novel-miR-239, novel-miR-193, and miR-339-5p as DP-detrimental miRNAs. Finally, by employing a well-in-well activation/embryo culture system that enables tracking the COC origin of CCs and embryos, we further validated Spp1 and Ngf as DP-beneficial genes, Jun as the DP-detrimental gene, and miR-7686-5p, novel-miR-239, and miR-339-5p as DP-detrimental miRNAs. In conclusion, we identified and validated new sets of CCs-expressed genes and miRNAs whose expression levels were directly correlated with oocyte DP. These newly identified genes and miRNAs may serve as potential biomarkers for selecting competent oocytes and embryos. Full article

Spinocerebellar ataxia type 1 (SCA1) is a neurodegenerative disorder that predominantly affects the Purkinje cells (PCs) of the cerebellum, leading to cerebellar degeneration, motor dysfunction, and cognitive impairment. Sphingosine-1-phosphate (S1P) signaling, known to modulate neuroinflammation, has been identified as a potential therapeutic target in SCA1. To investigate the therapeutic efficacy of the S1P modulator fingolimod, we treated a mouse model for SCA1, ATXN1[82Q]/+ mice during three different periods with fingolimod and assessed the effects. Potential therapeutic effects were monitored by tracking locomotion during the treatment period and examining PC morphology, connectivity, and markers for neuroinflammation post-mortem. Fingolimod treatment reduced astrocyte and microglial activation during all three treatment periods. We found no effect on calbindin levels or the thickness of the molecular layer, but fingolimod did improve the extent of the synaptic input of climbing fibers to PCs. While fingolimod improved important aspects of cellular pathology, we could only detect signs of improvement in the locomotion phenotype when treatment started at a later stage of the disease. In conclusion, fingolimod is able to mitigate neuroinflammation, preserve aspects of PC function in SCA1, and remediate part of the ataxia phenotype when treatment is appropriately timed. Although behavioral benefits were limited, targeting S1P pathways represents a potential therapeutic strategy for SCA1. Further studies are needed to optimize treatment regimens and assess long-term outcomes. Full article

Negative pressure wound therapy (NPWT) is widely recognized for its efficacy in treating diabetic wounds, but the mechanisms involved in the wound healing process remain unclear. By examining changes in blood cytokine levels as molecular signaling precursors, we aim to provide a comprehensive cytokine profile to support adjunctive therapy research and clinical applications. A diabetic mouse wound model was established to compare cytokine profiles between NPWT-treated and standard dressing groups, identifying key signaling candidates that may facilitate wound healing. By integrating normal mouse data with large-scale cytokine analysis, we developed a time-stratified NPWT approach to track acute-phase cytokine fluctuations in diabetic conditions. NPWT did not significantly enhance coagulation-related cytokine expression but effectively reduced inflammation, albeit with a delayed regulatory effect compared to wild-type mice. A one-sided binomial test revealed that NPWT advanced the cytokine expression peak from 16 to 2 h, partially restoring the early healing pattern seen in normal mice and suggesting its potential role in modulating early-stage wound repair. These findings provide novel insights into early cytokine regulation during wound healing and highlight the potential of NPWT to inform therapeutic strategies. This refined monitoring approach may contribute to improved clinical decision-making and support enhanced wound management in diabetic patients. Full article

Influenza (flu) is a highly prevalent respiratory illness caused by influenza viruses, representing a significant global health burden due to its substantial morbidity and mortality rate. Vaccination remains the most effective strategy for influenza prevention, and well-characterized animal models of influenza infection serve as essential tools for evaluating vaccine protective efficacy. However, animal models utilizing live influenza virus strains pose significant biosafety concerns, and many such strains are not readily available for research. To address these challenges, we established a novel visual mouse infection model using an HIV-based vector system. This model employs influenza pseudoviruses carrying a luciferase reporter gene, enabling real-time monitoring of viral load and in vivo tracking of viral distribution during infection. Using this infection model, we assessed the in vivo protective efficacy of an influenza vaccine and cross-validated the pseudovirus-based evaluation results against a live virus-infected mouse model. Our study thus establishes a safer and more convenient platform for evaluating influenza vaccine efficacy, including the assessment of broad-spectrum neutralization capacity. Full article

Background/Objectives: ⁸⁹Zr-oxine is an ex vivo cell labeling agent that enables cells to be tracked in vivo by positron emission tomography (PET) over a period of up to two weeks. To better understand where ⁸⁹Zr-oxine binds within cellular components, factors affecting labeling and intracellular distribution of ⁸⁹Zr were examined. Methods: Mouse primary T cells, natural killer cells, dendritic cells, and monocytes, and cell lines EL4 (mouse lymphoma), DC2.4 (mouse dendritic cell), Kit225K6 (human T cell leukemia) and MC38 (mouse colon adenocarcinoma) were labeled with ⁸⁹Zr-oxine or ¹¹¹In-oxine and protein binding within the cellular compartments, the labeling thresholds, and radioactivity retention were subsequently determined. Results: Cell incorporation of ⁸⁹Zr-oxine (27.8–71.8 kBq/10⁶ cells) positively correlated with cellular size and protein mass. Most (>97%) ⁸⁹Zr was protein-bound and primarily localized in the cytoplasm, membrane, and nuclear fractions (>81%) with distribution patterns varying by cell type. By contrast, ¹¹¹In-oxine showed lower protein-binding activity of approximately 59–65%, with 62–65% of ¹¹¹In localized in the cytoplasm. Autoradiography of electrophoresed subcellular fractionated cell samples indicated stable binding by ⁸⁹Zr-oxine to proteins in all subcellular fractions but unstable protein binding by ¹¹¹In. Saturation studies showed that ⁸⁹Zr-oxine labeling was saturable, and further labeling reduced cellular retention. Biodistribution of dendritic cells labeled with either ⁸⁹Zr-oxine or ¹¹¹In-oxine indicated greater retention of ⁸⁹Zr in the labeled cells in vivo than ¹¹¹In. Conclusions: ⁸⁹Zr-oxine stably binds many intracellular proteins and shows much higher and more stable protein binding than ¹¹¹In-oxine. Intracellular protein binding of ⁸⁹Zr accounts for the ability of ⁸⁹Zr-oxine labeling to successfully track cells in vivo long-term on PET. Full article

With the development of high-performance computers, cloud storage, and advanced sensors, people’s ability to gather complex learning data has greatly improved. However, analyzing these data remains a significant challenge. Especially for spatiotemporal learning data such as eye-tracking and mouse movement, understanding and analyzing these data to identify the learning insights behind them is a difficult task. We propose a visualization platform called “MultiScaleAnalyzer”, which employs hierarchical structure to illustrate spatiotemporal learning data in multiple views. From high-level overviews to detailed analyses, “MultiScaleAnalyzer” provides varying resolutions of data tailored to educators’ need. To demonstrate the platform’s effectiveness, we applied “MultiScaleAnalyzer” to a mathematical word problem-solving dataset, showcasing how the visualization platform facilitates the exploration of student problem-solving patterns and strategies. Full article

Background/Objectives: Glioblastoma (GBM) is an aggressive and lethal primary brain tumor with a poor prognosis, with a 5-year survival rate of approximately 5%. Despite advances in oncologic treatments, including surgery, radiotherapy, and chemotherapy, survival outcomes have remained stagnant, largely due to the failure of conventional therapies to address the tumor’s inherent heterogeneity. Radiomics, a rapidly emerging field, provides an opportunity to extract features from MRI scans, offering new insights into tumor biology and treatment response. This study evaluates the potential of delta radiomics, the study of changes in radiomic features over time in response to treatment or disease progression, exploring the potential of delta radiomics to track temporal radiation changes in tumor morphology and microstructure. Methods: A cohort of 50 female CD1 nude mice was injected intracranially with G7 glioblastoma cells and divided into irradiated (IR) and non-irradiated (non-IR) groups. MRI scans were performed at baseline (week 11) and post-radiation (weeks 12 and 14), and radiomic features, including shape, histogram, and texture parameters, were extracted and analyzed to capture radiation-induced changes. The most robust features were those identified through intra-observer reproducibility assessment, ensuring reliability in feature selection. A machine learning model was developed to classify irradiated tumors based on delta radiomic features, and statistical analyses were conducted to evaluate feature feasibility, stability, and predictive performance. Results: Our findings demonstrate that delta radiomics effectively captured significant temporal variations in tumor characteristics. Delta radiomics features exhibited distinct patterns across different time points in the IR group, enabling machine learning models to achieve a high accuracy. Conclusions: Delta radiomics offers a robust, non-invasive method for monitoring the treatment of glioblastoma (GBM) following radiation therapy. Future research should prioritize the application of MRI delta radiomics to effectively capture short-term changes resulting from intratumoral radiation effects. This advancement has the potential to significantly enhance treatment monitoring and facilitate the development of personalized therapeutic strategies. Full article

Hypertrophic cardiomyopathy (HCM) is often characterized by augmented cardiac contractility, which frequently remains undetectable in its early stages. Emerging evidence suggests that hypercontractility is linked to mitochondrial defects that develop early in HCM progression. However, imaging markers for identifying these early alterations in myocardial function are lacking. We used cardiac magnetic resonance feature tracking (CMR-FT) to assess myocardial strain in a Mybpc3-knockin (KI) mouse model that mimicked human HCM. While homozygous (HOM) mice exhibited cardiac hypertrophy, heterozygous (HET) mice represented an early, asymptomatic stage of HCM. To explore mitochondrial contributions to hypercontractility, we evaluated mitochondrial integrity via scanning electron microscopy (SEM) and correlated these findings with strain abnormalities. Young HET female, but not male mice exhibited significant torsion abnormalities (p = 0.02), reduced left ventricular global longitudinal strain (LVGLS, p = 0.009), and impaired right ventricular global longitudinal strain (RVGLS, p = 0.035) compared to the controls. Strain abnormalities correlated strongly with mitochondrial morphological alterations, including changes in volume and area distribution (R > 0.7). Abnormal myocardial strain patterns, including torsion and GLS, serve as early markers of HCM and are closely associated with underlying mitochondrial dysfunction. The HET Mybpc3-KI HCM model provides important insights into the initial stages of HCM progression, highlighting strain abnormalities and sex-specific differences to enhance early diagnosis and therapeutic strategies. Full article