Search Results (2,253)

Background/Objectives: The tumor microenvironment (TME) of pancreatic ductal adenocarcinoma (PDAC) is characterized by an enriched stroma, hampering the effectiveness of therapy. This co-clinical study aimed to (1) provide insight into early post-treatment changes in the TME using multiparametric magnetic resonance imaging (mpMRI)-derived quantitative imaging biomarkers (QIBs) in a preclinical PDAC model treated with radiotherapy and correlate these QIBs with histology; (2) evaluate the feasibility of obtaining these QIBs in patients with PDAC using clinically approved mpMRI data acquisitions. Methods: Athymic mice (n = 12) at pre- and post-treatment as well as patients with PDAC (n = 11) at pre-treatment underwent mpMRI including diffusion-weighted (DW) and dynamic contrast-enhanced (DCE) data acquisition sequences. DW and DCE data were analyzed using monoexponential and extended Tofts models, respectively. DeepLIIF quantified the total percentage (%) of tumor cells in hematoxylin and eosin (H&E)-stained tissues from athymic mice. Spearman correlation and Wilcoxon signed rank tests were performed for statistical analysis. Results: In the preclinical PDAC model, mean pre- and post-treatment ADC and K^trans values differed significantly (p < 0.01), changing by 20.50% and 20.41%, respectively, and the median total tumor cells quantified by DeepLIIF was 24% (range: 15–53%). Post-treatment ADC values and relative change in v_e (rΔv_e) showed a significant negative correlation with total tumor cells (ρ = −0.77, p < 0.014 for ADC and ρ = −0.77, p = 0.009 for rΔv_e). In patients with PDAC, pre-treatment mean ADC and K^trans values were 1.76 × 10⁻³ (mm²/s) and 0.24 (min⁻¹), respectively. Conclusions: QIBs in both preclinical and clinical settings underscore their potential for future co-clinical research to evaluate emerging drug combinations targeting both tumor and stroma. Full article

Cervical cancer (CC) remains a significant public health problem. Despite the availability of standard treatment strategies, chemotherapy-resistant tumors persist, highlighting the need to explore new therapeutic approaches or adjuvant strategies. This underscores the importance of preclinical in vivo models. Conventional models, such as murine xenografts, patient-derived xenografts (PDXs), and patient-derived organoids (PDOs), provide valuable biological relevance but are often time-consuming, costly, and resource-intensive. In this context, the chick embryo chorioallantoic membrane (CAM) assay represents a rapid, low-cost, and technically accessible in vivo platform. The CAM is a non-innervated, highly vascularized extraembryonic structure that provides a suitable environment for tumor generation from xenografts. However, despite the broad use of the CAM assay for tumor xenografts, standardized and comparative methodological optimizations specifically addressing technical variables for cervical cancer tumor induction remain limited. Therefore, the aim of this study was to optimize the CAM assay for tumor generation using the HeLa and SiHa cell lines. The generated tumors are vascularized and exhibit Ki-67 expression. The CAM assay is an excellent short-term exploratory model based on developing chicken embryos for studying the developmental biology of cervical tumors, which would accelerate the preclinical investigation of new therapeutic molecules. Full article

Background: Clear cell renal cell carcinoma (ccRCC) is the most common subtype of kidney cancer and is marked by pronounced intra-tumoural heterogeneity that complicates therapeutic response. Patient-derived organoids offer a physiologically relevant model to capture this diversity and evaluate treatment effects. When integrated with spatial transcriptomics, they might enable the mapping of spatially resolved transcriptional and isoform-level changes within the tumour microenvironment. Methods: We established a robust workflow for generating patient-derived ccRCC organoids, that are not passaged and retain original cellular components. These retain key features of the original tumours, including cancer cell, stromal, and immune components. Results: Spatial transcriptomic profiling revealed multiple transcriptionally distinct regions within and across organoids, reflecting the intrinsic heterogeneity of ccRCC. Isoform-level analysis identified spatially variable expression of glutaminase (GLS) isoforms, with heterogeneous distributions of both the GAC and KGA variants. Treatment with NUC-7738, a phosphoramidate derivative of 3′-deoxyadenosine, induced marked transcriptional remodelling of organoids, including alterations in ribosomal and mitochondrial gene expression. Conclusions: This study demonstrates that combining long-read spatial transcriptomics with patient-derived organoid models provides a powerful and scalable approach for dissecting gene and isoform-level heterogeneity in ccRCC and for elucidating spatially resolved transcriptional responses to novel therapeutics. Full article

Chagas disease, caused by Trypanosoma cruzi, affects a significant proportion of patients who develop digestive and cardiac complications, including megaviscera. This pathogenesis has been associated with autoimmune mechanisms mediated by molecular mimicry. In this study, an in silico evaluation of the potential structural basis of cross-reactivity of β-tubulin 1.9 of T. cruzi and the human β-4A tubulin isoform 3 was conducted. Using bioinformatics tools, homologous regions were identified and potentially immunogenic epitopes were predicted, considering their structural modeling and molecular docking. The proteins shared 87% sequence identity and 95% similarity, with an almost identical structural overlap, RMSD 0.291 Å. Three epitopes, VPFPRLHFF, NDLVSEYQQYQDATI, and GQSGAGNNWAKGHYTEGAELIDS, exhibited high predicted antigenicity, with the 9-mer and 16-mer peptides displaying structurally compatible docking poses within the binding grooves of MHC class I and class II molecules, respectively, while B-cell epitope potential was inferred from sequence-based property predictions. Normal mode analysis, used as an exploratory approach, suggested comparable flexibility profiles for the parasitic- and human-derived peptide–MHC complexes. These findings provide an exploratory structural framework supporting a potential role of β-tubulin epitopes in molecular mimicry processes implicated in the development of chagasic megaviscera. Full article

Late-onset sporadic Alzheimer’s disease (LOAD) is the most common form of dementia. The disease is characterized by progressive loss of memory and behavioral changes followed by neurodegeneration of all cortical areas. While the contribution of genetic and environmental factors is important, advanced aging remains the most important disease risk factor. Because LOAD does not naturally occur in most animal species, except humans, studies have traditionally relied on the use of transgenic mouse models recapitulating early-onset familial Alzheimer’s disease (EOAD). Hence, the development of more representative LOAD models through reprograming of patient-derived cells into neuronal, glial, and immune cells became a necessity to better understand the disease’s origin and pathophysiology. Herein, and focusing on neurons, we review current work in the field and compare results obtained with two different reprograming methods to generate LOAD patient’s neuronal cells: the induced pluripotent stem cell and induced neuron technologies. We also evaluate if these models can faithfully mimic cellular and molecular pathologies observed in LOAD patients’ brains. Full article

Objective: The aim of this study was to assess the association between the Lung Immune Prognostic Index (LIPI) measured at diagnosis and both event-free survival (EFS) and overall survival (OS) in patients with stage III non-small cell lung cancer (NSCLC). Methods: This retrospective cohort included patients diagnosed with stage III NSCLC between September 2022 and July 2024, all of whom had a minimum follow-up duration of six months. LIPI was calculated using the derived neutrophil-to-lymphocyte ratio (dNLR) and lactate dehydrogenase levels at diagnosis. Clinical, demographic, and treatment-related data were systematically collected. Survival outcomes were estimated using the Kaplan–Meier approach, while factors associated with prognosis were examined through Cox proportional hazards models. Results: The study population consisted of 68 patients, predominantly male (86.8%), with a mean age of 63.4 ± 8.7 years. According to the Lung Immune Prognostic Index classification, 29 patients (42.6%) were categorized as having a good score, 27 (39.7%) as intermediate, and 12 (17.6%) as poor. During a median follow-up of 15.4 months, a total of 40 progressions and 22 deaths occurred. Median EFS was 17.7, 9.4, and 5.8 months for good, intermediate, and poor LIPI groups, respectively (p < 0.001). Median OS was 25.7 months in the good LIPI group, was not reached in the intermediate group due to insufficient events, and was 6.7 months in the poor group (p < 0.001). In multivariate Cox analysis, poor LIPI was independently associated with inferior survival (EFS: HR = 2.87, 95% CI: 1.85–4.46, p < 0.001; OS: HR = 2.59, 95% CI: 1.40–4.78, p = 0.002). Conclusions: LIPI calculated at diagnosis is an independent prognostic factor for both EFS and OS in stage III NSCLC. Validation in larger, prospective cohorts is warranted to further define its prognostic role in stage III NSCLC. Full article

Endothelial cells (ECs) regulate vascular homeostasis, and their dysfunction is a key driver of many cardiovascular and cerebrovascular diseases. Human-induced pluripotent stem cell-derived endothelial cells (hiPSC-ECs) provide access to patient-specific vascular cells that can be directed toward arterial, venous, or organotypic phenotypes, enabling personalized in vitro modeling of endothelial pathology. In this review, we discuss how patient-specific iPSC-ECs are used as predictive and personalized two- and three-dimensional models to dissect disease mechanisms and prioritize targeted therapies. We highlight some limitations of this methodology and outline future directions for integrating iPSC-EC-based assays into individualized treatment algorithms. Full article

Background: Ovarian cancer remains the deadliest gynecologic malignancy, with its progression closely tied to age-associated remodeling of the tumor immune microenvironment. The laying hen serves as a valuable spontaneous model for human ovarian cancer. Its single-cell analyses may provide valuable insights into the immune-related axis linking ovarian aging to carcinogenesis. Methods: This study applied single-cell RNA sequencing to profile ovaries from three laying hen groups, including 35-week-old normal ovaries (A35w), 110-week-old normal ovaries (B110w), and 110-week-old ovarian cancer tissues (C110w). Key analyses had UCell-based scoring of senescence-related pathways and cancer hallmarks, differential expression analysis for overlapping dysregulated genes, LASSO regression-based prognostic model construction, and assessment of chemotherapy sensitivity and immune infiltration. Results: A comprehensive cellular landscape of chicken ovaries was established, identifying major immune populations including B cells, CD4+ T cells, CD8+ T cells, macrophages, and plasma cells. Senescence-related pathways and cancer hallmarks showed progressive activation in immune cells from A35w to B110w to C110w. A total of 216 genes commonly dysregulated in aging and carcinogenesis, reveal core links between immune dysfunction and malignant transformation. The 20-gene prognostic model derived from these genes stratified human ovarian cancer patients into high-risk and low-risk groups with significant overall survival differences, exhibited robust predictive performance across TCGA, GSE32063, and GSE140082. The model also predicted the differential chemotherapy sensitivity in high-risk and low-risk patients and correlated with specific immune infiltration patterns in the tumor microenvironment. Conclusions: Notably, this is the first single-cell RNA sequencing study of chicken ovarian cancer, and we constructed the 20-gene prognostic model for human ovarian cancer using 216 genes that change significantly in immune cells during both ovarian aging and carcinogenesis. This work provides support to establish the hen as a potential preclinical animal model and a translational tool to guide personalized therapy. Full article

Background/Objectives: Head and neck squamous cell carcinoma (HNSCC) is a biologically heterogeneous malignancy with poor outcomes in advanced disease. Increasing evidence indicates that the tumor microenvironment, particularly cancer-associated fibroblasts (CAFs), plays an important role in tumor progression and immune regulation. However, the diversity of CAF subsets and their clinical relevance in HNSCC remain incompletely understood. This study aimed to characterize CAF heterogeneity and assess the prognostic significance of CAF subset-specific transcriptional programs. Methods: Single-cell RNA sequencing data from HNSCC tumors were analyzed to identify CAF subsets based on differentially expressed genes. CAF subset-specific gene signatures were used to construct prognostic risk models for overall survival (OS) and progression-free survival (PFS) in The Cancer Genome Atlas HNSCC cohort, with validation in an independent dataset. CAF-driven prognostic groups were defined, and their immune landscapes and biological pathways were evaluated. Bulk RNA sequencing of primary CAF cultures was performed for validation. Results: Six CAF subsets were identified, including myofibroblastic (myCAF), inflammatory (iCAF), antigen-presenting, and extracellular matrix-related CAFs. Risk scores derived from inflammatory CAF subsets consistently predicted shorter OS across independent cohorts, whereas PFS prediction showed greater cohort dependency. CAF-based stratification identified patient subgroups with distinct immune profiles and pathway enrichment patterns. These results were supported by validation analyses and by bulk RNA sequencing of primary CAFs, demonstrating preservation of myCAF- and iCAF-like transcriptional programs ex vivo. Conclusions: CAF heterogeneity has important prognostic and immunological implications in HNSCC. Inflammatory CAF-related transcriptional programs represent robust markers of patient survival and may complement tumor-intrinsic biomarkers. Full article

Cervical cancer remains a significant global health burden, with chemoradioresistance representing a major obstacle to successful treatment. To elucidate the mechanisms underlying this resistance, we established a unique pair of isogenic primary cervical cancer cell lines, AdMer35 and AdMer43, obtained from a patient with squamous cell carcinoma of the cervix before and after radiation therapy. The aim of our study was to characterize the transcriptomic and cellular heterogeneity of these cells. We conducted an in-depth comparative analysis using single-cell RNA sequencing. Analysis of this paired, patient-derived isogenic model suggests that chemoradioresistance can arise through coordinated multilevel cellular adaptations. Resistant AdMer43 cells demonstrated transcriptional reprogramming, with the upregulation of embryonic stemness factors (HOX, POU5F1, SOX2), a shift in extracellular matrix from fibrillar to non-fibrillar collagens, and activation of inflammatory pathways. We identified and characterized critical cell-state dynamics: resistant cells exhibited a remodeled ecosystem with a metabolically reprogrammed senescent-like cell population showing an enhanced pro-tumorigenic communication via EREG, SEMA3C, BMP, and WNT pathways. Furthermore, we identified a progenitor-like cell population with a minimal CNV burden, potentially serving as a reservoir for tumor persistence. These findings offer novel insights for developing targeted strategies to eliminate resistant cell pools and improve cervical cancer outcomes. Full article

Polydopamine (PDA) surface coatings are widely used in biomedical engineering to enhance cell–substrate interactions; however, their effects on cancer-cell behavior remain unclear. In this study, we investigated how PDA-coated two-dimensional (2D) culture surfaces influence oncogenic traits of human prostate cancer (PC) cells in vitro. Using LNCaP, DU145, and PC3 cell lines, we found that PDA-coated substrates markedly increased the adhesion, migration, invasion, proliferation, and colony formation in a dose- and time-dependent manner. PDA exposure also induced epithelial–mesenchymal transition (EMT), upregulated cancer stem cell markers (CD44, CD117, CD133, Sox2, Oct4, and Nanog), and elevated expression of metastasis- and chemoresistance-associated molecules (MMP-2, MMP-9, MDR1, and MRP1). Mechanistically, PDA coatings enhanced integrin α₂β₁-associated cell adhesion, accompanied by increased focal adhesion kinase (FAK) phosphorylation and downstream activation of JNK signaling. Pharmacological inhibition of integrin α₂β₁ (BTT-3033), FAK (PF573228) and JNK (SP600125) effectively abrogated PDA-induced malignant phenotypes and restored chemosensitivity to cabazitaxel, cisplatin, docetaxel, curcumin, and enzalutamide. Collectively, these findings identify PDA-coated surfaces as a simple, efficient, and reductionist in vitro platform for studying adhesion-mediated signaling and phenotypic plasticity in PC cells, while acknowledging that further validation in three-dimensional (3D) and patient-derived models will be required to establish in vivo relevance. Full article

Bone defects in the maxillofacial region severely impair patient function and esthetics. Free autologous bone grafting remains the gold-standard treatment; however, surgical intervention at donor sites limits clinical applicability. Treatment using artificial materials also presents challenges, including insufficient bone regeneration and poor biocompatibility. Bio three-dimensional (3D) printing, which enables the fabrication of scaffold-free 3D constructs from cellular spheroids has emerged as a promising regenerative approach. This study investigated the osteogenic potential of scaffold-free constructs composed of human dental pulp stem cell (DPSC) spheroids in a rat mandibular defect model. DPSCs isolated from extracted human teeth were used to generate spheroids, which were assembled into 3D constructs using a Bio 3D printer. The spheroids exhibited higher mRNA expression of stem cells and early osteogenic markers than monolayer cultures. The constructs were transplanted into mandibular defects of immunodeficient rats, and bone regeneration was assessed eight weeks post-transplantation. Radiographic and micro-Computed Tomography analyses revealed significantly greater bone volume and mineral density in the 3D construct group. Histological and immunohistochemical examinations confirmed newly formed bone containing osteogenic cells derived from the transplanted DPSCs. These findings indicate that Bio 3D-printed, scaffold-free DPSC constructs promote mandibular bone regeneration and may provide a novel strategy for maxillofacial reconstruction. Full article

The liver is the central organ in metabolism; however, modeling hepatic diseases remains limited by current experimental models. Animal models frequently fail to predict human liver physiology, while primary hepatocytes rapidly dedifferentiate in culture. We performed comprehensive transcriptomic profiling of induced pluripotent stem cells (iPSCs) differentiation into hepatocyte-like cells (HLCs) under two-dimensional (2D) and three-dimensional (3D) culture conditions. RNA sequencing analysis revealed the sequential activation of lineage-specific markers across major developmental stages: definitive endoderm (FOXA2, SOX17, CXCR4, CER1, GATA4), posterior foregut (PROX1, GATA6), and hepatoblasts (HNF4A, AFP). Comparative analysis demonstrated a markedly enhanced hepatic gene expression of 3D organoids, as demonstrated by a 33-fold increase in HNF4A expression and elevated levels of mature hepatocyte markers, including ALB, SERPINA1, and UGT2B15. However, the 3D cultures retained fetal characteristics (290-fold higher AFP expression) and exhibited significantly impaired metabolic function, with CYP3A4 expression levels reduced by 2000-fold compared to the adult human liver. This partial maturation was further supported by a moderate correlation with adult liver tissue (ρ = 0.57). We demonstrated high reproducibility across five biologically distinct iPSCs lines, including those derived from patients with rare monogenic disorders. The establishment of quantitative benchmarks provides a crucial tool for standardizing in vitro liver models. Furthermore, we delineate the specific limitations of the current model, highlighting the need for further protocol optimization to enhance metabolic maturation and P450 enzyme activity. Functional validation of metabolic activity (CYP enzyme assays, albumin secretion) was not performed; therefore, conclusions regarding hepatocyte functionality are based on transcriptomic evidence. Full article

Background/Objectives: Glucose Transporter 1 Deficiency Syndrome (GLUT1-DS) is a neurodevelopmental disorder caused by mutations in the gene encoding glucose transporter 1 (GLUT1), which leads to impaired glucose transport into the brain and is characterized by drug-resistant epilepsy. Limited glucose supply disrupts neuronal and astrocytic energy homeostasis, but how hypometabolism translates into network hyperexcitability remains poorly understood. Here, we used induced pluripotent stem cells (iPSCs)-derived brain organoids to examine how reduced metabolic substrate availability shapes epileptiform dynamics in human neuronal circuits from GLUT1-DS. Methods: Brain organoids were generated from a healthy donor or a GLUT1-DS patient and interfaced with multielectrode arrays (MEA) for recording of neuronal activity. A unified Python (v3.10)-based analytical pipeline was developed to quantify spikes, bursts, and power spectral density (PSD) across frequency bands of neuronal activity. Organoids were challenged with reduced glucose, pentylenetetrazol (PTZ), potassium chloride (KCl), and tetrodotoxin (TTX) to assess metabolic and pharmacological modulation of excitability. Results: GLUT1-DS organoids exhibited elevated baseline hyperexcitability compared to healthy control, characterized by increased spike rates, prolonged bursts, increased spikes per burst, and elevated PSD. Reduced glucose availability further amplified these features selectively in GLUT1-DS. Conclusions: Human brain organoids reproduce the pathological coupling between hypometabolism and hyperexcitability in GLUT1-DS. Our platform provides a mechanistic model and quantification tool for evaluating metabolic and anti-epileptic therapeutic strategies. Full article

Human adult cardiomyocytes (CMs) have limited regenerative capacity, posing a significant challenge in restoring cardiac function following substantial CM loss due to an acute ischemic event or chronic hemodynamic overload. Nearly half of patients show no improvement in left ventricular ejection fraction during recovery from acute myocardial infarction. At baseline, both humans and mice exhibit low but continuous cell turnover originating from the existing CMs. Moreover, myocardial infarction can induce endogenous CM cell cycling. Consequently, research has focused on identifying drivers of CM rejuvenation and proliferation from pre-existing CMs. High-throughput screening has facilitated the discovery of novel pro-proliferative targets through small molecules, microRNAs, and pathway-specific interventions. More recently, omics-based approaches such as single-nucleus RNA sequencing and spatial transcriptomics have expanded our understanding of cardiac cellular heterogeneity. The big-data strategies provide critical insights into why only a subset of CMs re-enter the cell cycle while most remain quiescent. In this review, we compare several high-throughput screening strategies used to identify novel targets for CM proliferation. We also summarize the benefits and limitations of various screening models—including zebrafish embryos, rodent CMs, human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), and cardiac organoids—underscoring the importance of integrating multiple systems to uncover new regenerative mechanisms. Further work is needed to identify translatable and safe targets capable of inducing functional CM expansion in clinical settings. By integrating high-throughput screening findings with insights into CM heterogeneity, this review provides a comprehensive framework for advancing cardiac regeneration research and guiding future therapeutic development. Full article