Search Results (526)

Neurodegenerative diseases (NDDs), including Alzheimer’s disease (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), are becoming more prevalent and still lack effective disease-modifying therapies (DMTs). However, translational efficiency remains critically low. For example, a ClinicalTrials.gov analysis of AD programs (2002–2012) estimated ~99.6% attrition, while PD programs (1999–2019) achieved an overall success rate of ~14.9%. In vitro platforms are assessed, ranging from immortalized neuronal lines and primary cultures to human-induced pluripotent stem cell (iPSC)-derived neurons/glia, neuron–glia co-cultures (including neuroinflammation paradigms), 3D spheroids, organoids, and blood–brain barrier (BBB)-on-chip systems. Complementary in vivo toxin, pharmacological, and genetic models are discussed for systems-level validation and central nervous system (CNS) exposure realism. The therapeutic synthesis focuses on AD, covering symptomatic drugs, anti-amyloid immunotherapies, tau-directed approaches, and repurposed drug classes that target metabolism, neuroinflammation, and network dysfunction. This review links experimental models to translational decision-making, focusing primarily on AD and providing a brief comparative context from other NDDs. It also covers emerging targeted protein degradation (PROTACs). Key priorities include neuroimmune/neurovascular human models, biomarker-anchored adaptive trials, mechanism-guided combination DMTs, and CNS PK/PD-driven development for brain-directed degraders. Full article

The recreation of the tumor microenvironment remains a significant challenge in the development of experimental cancer models. The present study constitutes an investigation into the interconnection between tumor, endothelial and stromal cells in heterotypic breast cancer spheroids. The generation of models was achieved through the utilization of MCF7, MDA-MB-231, and SK-BR-3 tumor cell lines, in conjunction with endothelial TIME-RFP cells and either cancer-associated (BrC4f) or normal (BN120f) fibroblasts, within ultra-low attachment plates. It was established that stromal cells, most notably fibroblasts, were conducive to the aggregation of tumor cells into spheroids and the formation of pseudovessels in close proximity to fibroblast bands. In contrast to the more aggressive tumor models MDA-MB-231 and SK-BR-3, microenvironment cells do not influence the migration ability of MCF7 tumor cells. Heterotypic spheroids incorporating CAFs demonstrated a more aggressive and immunosuppressive phenotype. Multiplex immunoassay analysis of cytokines, followed by STRING cluster analysis, was used to identify key processes including angiogenesis, invasion, stem cell maintenance, and immunosuppression. Furthermore, a cluster of cytokines (LIF, SDF-1, HGF, SCGFb) was identified as potentially involved in the regulation of PD-L1 expression by tumor cells. This finding reveals a potential mechanism of immune evasion and suggests new avenues for therapeutic investigation. Full article

SA 178 grade C carbon steel is a material commonly used in boiler tubes. Boilers are crucial in the energy industry; however, their service life degrades over time. If a boiler malfunctions, processing operations must be halted, resulting in financial losses for the company. The aim of this study is to examine the effect of microstructural evolution, especially the transformation of lamellar pearlite into spheroidized pearlite, on the service life degradation of boiler tubes. Understanding these changes is essential for preventing catastrophic system failures. The methodology involves the use of Small-Angle X-ray Scattering (SAXS) supported by metallographic analysis, Scanning Electron Microscopy (SEM), Energy-Dispersive X-ray (EDX) Spectroscopy, and mechanical testing. The SAXS results indicate that the microstructure of SA 178, which initially consisted of ferrite and lamellar pearlite, gradually transforms into spheroidized pearlite. These microstructural changes lead to reductions in tensile strength from 523 MPa for 0% spheroidization to 335 MPa for 100% spheroidization, as well as a reduction in hardness from 175 HV to 89 HV, ultimately decreasing the service life of the boiler tube. Full article

Background/Objectives: Obesity-associated hyperleptinemia has been linked to breast cancer (BC) progression via mechanisms that remain incompletely understood. This study explores the role of leptin and its receptor (LEPR) in facilitating BC cell proliferation, migration, epithelial–mesenchymal transition (EMT), and STAT3 signaling pathway activation. Methods: We analyzed gene expression and survival data from TCGA BRCA dataset. MCF-7 and MDA-MB-231 BC cells were exposed to leptin at 10 ng/mL (lean-associated levels) and 100 ng/mL (elevated levels linked to obesity). MTT assays, colony formation tests, wound-healing and tumor spheroid dissemination experiments evaluated cell proliferation and migration. Immunofluorescence and Western blot analysis assessed changes in EMT markers and cytoskeletal alterations, while Western blotting and qPCR assessed STAT3 and NCOA1 expression and activation levels. Results: Elevated LEPR expression was linked with unfavorable prognosis in BC patients. Higher doses of leptin (100 ng/mL) significantly enhanced cellular proliferation rates and migratory capabilities, in both cell lines, and promoted EMT characteristics marked by downregulated E-cadherin and cytoskeleton structural changes. Whereas heightened JAK2/STAT3 signaling correlated with elevated leptin dosages, STAT3 inhibition using AG490 reversed leptin-induced migration while reinstating E-cadherin levels to baseline. Furthermore, leptin upregulated NCOA1, an essential STAT3 coactivator, facilitating increased expression of Cyclin D1 and VEGF target genes. Clinical positive relationships were seen between LEP/LEPR expressions and NCOA1 levels and between NCOA1 and various gene signatures related to STAT3/P-STAT3 within BC specimens. Conclusions: Obesity-associated hyperleptinemia enhances aggressiveness in BC through a mechanism involving LEPR-mediated activation pathways encompassing NCOA1/STAT3, which drive proliferation, migration, and EMT. This assigns a potential therapeutic utility for obesity-related advancements found within BC pathology. Full article

Background: Three-dimensional (3D) cancer models are currently essential tools in high-throughput screening (HTS), serving as a bridge between in vitro and in vivo approaches during drug development. Even though spheroids offer many advantages over 2D cultures, analyzing 3D cultures with heterogeneous morphology remains challenging due to the lack of standardized visualization techniques and multiparameter analysis. Methods: In this work, an optimized CellProfiler pipeline and a Python algorithm for weighting morphological features are used to visualize, extract, and analyze morphological data from spheroids derived from colorectal and pancreatic cancer cell lines with diverse morphologies (HCT116, LoVo, PANC-1, and CFPAC-1). Results: We developed a feature weighting process that combines multiple morphological parameters into a single metric using principal component analysis (PCA). There is a strong correlation between this process and a standard Alamar Blue proliferation assay (r = 0.89, ρ = 0.91, p < 0.001). Using this method, we were able to ascertain the IC₅₀ values of substances that did not produce results in cell lines with heterogeneous morphology (LoVo and CFPAC-1) using a standard proliferation assay. Conclusions: By removing the need for tracer dyes, the resulting methodology may lower costs while accelerating preclinical drug development through informative multiparameter analysis of compound efficacy. Full article

Ductile iron—also known as spheroidal graphite iron (SGI)—is a versatile material that exhibits a wide range of applications. In addition to the matrix structure itself, graphite morphology and material defects, such as graphite degenerations, also have a decisive influence on its mechanical properties. Missing or incomplete classifications of these deviating graphite morphologies in common standards, alongside insufficient knowledge regarding their effects, leads to a more complicated lifetime assessment and often results in rejection of SGI components. Therefore, relevant material parameters were derived from experimental quasi-static and fatigue investigations on SGI materials with different graphite degenerations and correlated with microstructural parameters quantified by an optimized digital image analysis method. While a linear correlation between the amount and mechanical properties was identified for spiky graphite, for chunky graphite, the presence in general can lead to a detrimental reduction in mechanical properties. Full article

Extracellular vesicles (EVs) derived from stem cells have emerged as promising mediators of osteogenesis, suggesting cell-free alternatives for bone tissue engineering and regenerative medicine. This review provides a comprehensive analysis of the main stem cell sources used for EV production, including bone marrow mesenchymal stem cells (BM-MSCs), adipose-derived stem cells (ADSCs), umbilical cord MSCs (UC-MSCs), induced pluripotent stem cells (iPSCs), and alternative stromal populations. Particular attention is given to the ways in which different conditioning and differentiation strategies, such as osteogenic induction, hypoxia, and mechanical stimulation, modulate EV cargo composition and enhance their therapeutic potential. We further discuss the in vitro models employed to evaluate EV-mediated bone regeneration, ranging from 2D cultures to complex 3D spheroids, scaffold-based systems, and bone organoids. Overall, this review emphasizes the current challenges related to standardization, scalable production, and clinical translation. It also outlines future directions, including bioengineering approaches, advanced preclinical models, and the integration of multi-omics approaches and artificial intelligence to optimize EV-based therapies. By integrating current knowledge, this work aims to guide researchers toward more consistent and physiologically relevant strategies to harness EVs for effective bone regeneration. Finally, this work uniquely integrates a comparative analysis of EVs from multiple stem cell sources with engineering strategies and emerging clinical perspectives, thereby providing an updated and translational framework for their application in bone regeneration. Full article

Background/Objectives: Metabolic reprogramming is an essential feature of tumors. Mitochondrial sirtuins SIRT3 and SIRT5 differently regulate glutamine metabolism with SIRT5 inhibiting glutaminase (GLS) and SIRT3 increasing glutamate dehydrogenase (GDH). Considering the important and interconnected role of glutamine, SIRT3 and SIRT5 for cancer growth and progression, our hypothesis is that a simultaneous modulation of SIRT3 and SIRT5 could represent a valid anti-tumoral strategy. Methods: wt and GLS1-silenced triple negative breast cancer spheroids were treated with 3-TYP, a selective SIRT3 inhibitor, and with MC3138, a new selective SIRT5 activator, both alone and in combination. The effects of such treatments on hypoxia, autophagy and mitophagy markers were determined by immunofluorescence and Western blot. Mitochondria morphology was studied by transmission electron microscopy (TEM) and mitochondrial ROS production by confocal analysis. Results: We observed that 3-TYP+MC3138 treatment decreased the size of spheroids by affecting HIF-1α, c-Myc, glutamine transporter SLC1A5 and autophagy (LC3II) and mitophagy (BNIP3) markers. Moreover, such treatments altered the morphology and conformation of the mitochondria. Finally, we also documented an increase in mitochondria reactive oxygen species (mtROS). Conclusions: The combined inhibition of SIRT3 and activation of SIRT5 greatly reduces the size of spheroids through the inhibition of hypoxic response, which is then followed by the alteration of the autophagic and mitophagic process and the toxic accumulation of mitochondrial ROS, representing a new anti-tumoral strategy. Full article

Myopathy encompasses a group of diseases characterized by abnormalities in both muscle function and structure. However, the underlying regulatory mechanisms of newly formed myofiber development remain poorly defined. No promising therapeutic approach has been developed, but numerous medication options are available to alleviate symptoms. Our previous studies demonstrated that adenosine kinase (ADK) is critical in regulating adenosine metabolism, pathological angiogenesis, pathological vascular remodeling, and vascular inflammatory diseases. Adenosine dynamically distributes between extracellular and intracellular, and adenosine concentration regulates ADK expression. However, the mechanism by which adenosine triggers an ADK-dependent intracellular signaling pathway to regulate skeletal muscle regeneration is not well defined. This study aimed to evaluate whether the adenosine-induced intracellular signaling pathway is involved in regulating myopathy, and how it regulates the development of newly formed myofibers. In this study, an intramuscular injection of cardiotoxin was used to induce a skeletal muscle injury model; satellite cells and C2C12 cells were employed. Whether adenosine regulates satellite cell activity, new myofiber formation and differentiation, as well as fusion of myofibers, were determined by H&E staining, BrdU incorporation assay, and spheroid sprouting assay. Interaction between ADK and PFKFB3 was evaluated by IF staining, PPI network analysis, molecular docking simulation, and CO-immunoprecipitation assay. The results demonstrated that adenosine dynamically distributes between extracellular and intracellular through concentrative nucleoside transports or equilibrative nucleoside transporters, and it rapidly induces an ADK-dependent intracellular signaling pathway, which interacts with PFKFB3-mediated glycolytic metabolism to promote satellite cell activity, new myofiber formation, differentiation, and fusion, and eventually enhances skeletal muscle regeneration after injury stress. The remarkable endogenous regeneration capacity of skeletal muscle, which is regulated by adenosine-triggered intracellular signaling, presents a promising therapeutic strategy for treating muscle trauma and muscular dystrophies. Full article

Loquat fruit is valued for its pleasant taste and favorable ripening period. However, its delicate texture and high perishability make it highly vulnerable to damage during packaging, so the fruit is usually packed by hand. Developing a fruit-sizing machine could increase commercial market opportunities. Automated mass detection reduces manual sorting errors and labor requirements. Overall, it enhances grading accuracy, speed, and uniformity in loquat processing. It also helps distinguish between ripe, underripe, and overripe fruits through subtle mass differences. Mass modeling has proven to be an effective baseline approach for the development and optimization of grading machines, and its efficiency has been demonstrated across different fruit types. Here, we present a comparative analysis of various models for mass modeling of six international and Italian loquat varieties (“Algerie,” “Peluche,” “Golden Nugget,” “Virticchiara,” “Nespolone di Trabia,” and “Claudia”) cultivated in southern Italy. On fifty fruits per variety, singular mass and spatial diameters [longitudinal (DL), maximum transverse (DT1), and minimum transverse (DT2) were measured. Linear and non-linear regression analyses, including quadratic, polynomial, and cubic models, were applied to both the complete dataset and individual varieties. A set of predictors was used, including DL (length), DT1 (width), and DT2 (thickness), ellipsoid and oblate spheroid volume. Model performance was evaluated based on higher R² values, and lower RMSE and MBE values. The best general model was obtained using an ellipsoidal volume (R² = 0.97, RMSE = 2.76). Both linear and cubic models demonstrated high suitability across all varieties, with ellipsoidal volume emerging as the most effective predictor. Conversely, (DL) based models were the least suitable, yielding the lowest (R² = 0.41) values in “Virticchiara.” The developed general and specific-variety models and equations provide a solid foundation for establishing high-performance systems for mass and size estimation, which can be effectively integrated into a fruit sizer machine. Full article

Cancer-associated fibroblasts (CAFs) are critical components of the tumor microenvironment that promote cancer progression and immune evasion. Adipocyte enhancer-binding protein 1 gene (AEBP1), which encodes aortic carboxypeptidase-like protein (ACLP), has been implicated in tissue remodeling and fibrosis, yet its role in CAF biology across cancers remains poorly understood. Here, we performed a pan-cancer transcriptomic analysis using The Cancer Genome Atlas (TCGA) and found that AEBP1 expression strongly correlates with expression of collagen family genes in the majority of solid tumors. Integration of single-cell RNA-sequencing datasets from breast and pancreatic cancers revealed that AEBP1 is predominantly expressed in CAFs, where it is co-expressed with collagens and CAF marker genes. Functional experiments using three-dimensional (3D) spheroids composed of oral squamous cell carcinoma (OSCC)-derived CAFs showed that AEBP1 knockdown significantly reduced spheroid stiffness without altering their morphology or size, indicating that ACLP contributes to the mechanical properties of tumor tissues. Together with earlier findings linking AEBP1/ACLP to reduced CD8⁺ T-cell infiltration, our results suggest that stromal AEBP1/ACLP enhances both extracellular matrix stiffness and immune suppression and highlights AEBP1/ACLP as a potential therapeutic target through which to remodel the tumor microenvironment and improve anti-tumor immunity. Full article

Prostaglandin E₂ (PGE₂) has been implicated in multiple biological processes during pregnancy in ruminants. However, the regulatory effects of PGE₂ on endometrial function during the diestrus period and its underlying molecular mechanisms remain poorly understood. Herein, PGE₂ treatment promoted the accumulation of lipid droplets and induced cytoskeletal reorganization in bEECs. As a well-established inducer of lipid droplet formation, oleic acid (OA) treatment significantly increased the number of lipid droplets in bEECs, altered the distribution of F-actin and disrupted the expression patterns of key adhesion-related proteins. Transcriptomic analysis revealed that the PPAR signaling pathway was the key pathway that responded to PGE₂ treatment in bEECs, and its downstream target gene FABP3 was markedly up-regulated. Knockdown of FABP3 led to a reduced number of BTC spheroids and down-regulation of adhesion-related proteins in bEECs while increasing the density of microvilli and up-regulating the expression of epithelial markers. Prostaglandin E receptor 4 (PTGER4) was the primary receptor that responded to PGE₂ treatment, and PTGER4 knockdown or pharmacological inhibition with GW-627368 suppressed FABP3 expression in bEECs. Moreover, uterine samples from dairy cows at different stages of the estrous cycle showed that FABP3 expression was significantly elevated in the endometrium tissue during mid-diestrus compared to metestrus, with predominant localization in the luminal and superficial glandular epithelium. Collectively, these findings indicate that FABP3 regulates lipid droplet accumulation and adhesion ability in bEECs via the PGE₂/PTGER4/PPAR signaling axis, providing new insights into the metabolic regulation of endometrial receptivity in ruminants. Full article

Natural or synthetic scaffolds are essential for developing three-dimensional (3D) cell culture models, as they provide structural stability and accurately replicate the cellular microenvironment. When integrated into optimized setups, scaffold-supported cellular aggregates, such as spheroids, can be non-destructively characterized and monitored using 3T Magnetic Resonance Imaging (MRI). However, a significant technical limitation is the presence of MR artifacts generated by scaffolds, which can severely obscure the visualization of the embedded spheroids. This study systematically evaluated the suitability of various scaffolds and matrices (including Matrigel^®, fibrin glue, and several hydrogels) for MRI and MR spectroscopy (MRS). The materials were investigated both native and seeded with chondrosarcoma cells (SW1353). Our findings revealed considerable variability in MR compatibility across different materials. Specifically, fibrin glue proved unsuitable for MR applications due to substantial artifact generation that interfered with the visualization of cellular components. Furthermore, the results emphasize the importance of the observation period, as material degradation processes can introduce confounding factors in longitudinal MR studies. The choice of scaffold material is paramount for the successful analysis of 3D cell models via MRI. Careful selection is required, as the materials’ properties and temporal stability directly impact the interpretability of the acquired data. Full article

Bioengineered dermal–epidermal composites (DECs) have demonstrated promise initiating skin regeneration and hair follicle neogenesis after injury. DECs in our work comprise a collagen matrix embedded with human dermal papilla cells (HDPCs) overlaid with human keratinocytes. HDPCs, as three-dimensional spheroids, enhance hair follicle formation, working in tandem with keratinocytes. Herein, 3D printed stamped PDMS microwell arrays were used as a strategy for spatially patterning dermal papilla spheroids in the dermal components of the DEC. DECs were transferred to cell culture media for 5 days followed by air–liquid interface culture for 2 days. Spheroid diameter, cell viability, and qPCR gene expression analyses were conducted. DECs were surgically grafted on immunocompromised mice, and healing was followed for 10 weeks. HDPCs cultured in the microwell arrays formed patterned viable spheroids and successfully transferred to the collagen dermal matrix. RNA analysis using qPCR showed upregulation of key HDPC markers (VCAN and BMP6) in DC microwell patterned HDPC spheroids compared to monolayers. This work represents a novel 3D printing strategy optimizing designing patterned HDPC spheroids in the extracellular matrix to regenerate functional human skin instead of scar tissue after injury. Full article

Salivary gland carcinomas (SGCs) are aggressive malignancies with limited treatment options, primarily due to the complexity of the tumor microenvironment (TME). Cancer-associated fibroblasts (CAFs) remodel the extracellular matrix (ECM), enhance cancer cell stemness, and drive drug resistance. This study introduces a decellularized CAF-derived spheroid system as a biomimetic platform to study tumor–stromal interactions in SGC. Multicellular spheroids were generated by co-culturing Medical Research Council cell strain 5 (MRC-5) fibroblasts (fetal lung-derived) with A253 salivary gland cancer cells, producing distinct spatial architecture, with fibroblasts at the core and cancer cells at the periphery. Compared with A253-only spheroids, A253/MRC-5 spheroids exhibited enhanced proliferation and elevated expression of stemness markers (aldehyde dehydrogenase 1 [ALDH1], CD133, cytokeratin 19 [CK19]). MRC-5 spheroids displayed robust ECM and growth factor expression that persisted after decellularization. Decellularized spheroids retained biological activity, enabling A253 cells to develop invasive phenotypes, metabolic reprogramming, and stemness-associated signatures. Transcriptomic analysis revealed a transition from proliferative pathways to stress-adaptive survival programs, mirroring in vivo tumor behavior. Moreover, A253 cells cultured with decellularized fibroblast spheroids exhibited altered cisplatin sensitivity, highlighting the critical role of stromal ECM in therapeutic response. In conclusion, this study establishes decellularized CAF spheroids as a simplified yet biologically relevant TME-mimetic platform. By recapitulating tumor–stromal crosstalk without live co-culture, this system provides a powerful tool for mechanistic studies of salivary gland cancer, preclinical drug screening, and development of stroma-targeted therapies. Full article