Search Results (1,153)

Introduction: We aimed to clarify the influence of the thrombus composition on ischemic stroke endovascular thrombectomy (EVT) efficiency by utilizing various staining methods for patients that presented with occlusions of the middle cerebral artery (MCA). Methods: Between September 2017 and May 2021, we analyzed thrombi retrieved during endovascular thrombectomy EVT in patients with acute ischemic stroke due to middle cerebral artery (MCA) occlusion. Patients with reperfusion failure, intracranial atherosclerotic occlusions, and inadequate staining were excluded. The thrombus composition was stratified using three staining techniques—Hematoxylin and Eosin (H&E), Martius Scarlet Blue (MSB) staining, and immunohistochemistry (IHC) for red blood cells (RBCs), white blood cells (WBCs), fibrin (Fibrin II), and platelets (CD41). Associations between EVT efficiency outcomes and the thrombus composition were evaluated. Results: During the study period, thrombus was available for analysis in 159 patients. A total of 59 patients were included in the main analysis. Increases in the trichotomized RBS tertiles were associated with decreases in the components of various platelet/other components but not for fibrin. A modified first pass effect (mFPE) of the modified Thrombolysis in Cerebral Infarction perfusion scale (mTICI) 2b or higher was associated with larger thrombus surface area (16.0 ± 11.6 vs. 47.4 ± 62.3 mm², p = 0.005), a higher MSB fibrin content (29.8 ± 10.7 vs. 21.3 ± 10.9%, p = 0.002), and IHC fibrin (28.5 ± 14.5 vs. 20.1 ± 11.4%, p = 0.008). There was a marginal association between the mTICI 2b mFPE and lower MSB platelet/other components (27.6 ± 20.9 vs. 34.4 ± 14.9%, p = 0.078). The discrepancy between MSB platelet/others and IHC platelets was greater in the mFPE (-) group, suggesting that components other than platelets may contribute to EVT resistance. A mFPE of mTICI 2c or higher was associated with greater thrombus surface area (17.8 ± 11.9 vs. 37.7 ± 55.0 mm², p = 0.015) and MSB fibrin (32.1 ± 10.3 vs. 22.8 ± 11.0%, p = 0.002). There was a marginal reverse association between the mTICI 2c mFPE and MSB RBCs (33.4 ± 20.2% vs. 41.5 ± 17.3%, p = 0.062). There was no significant association between final near-complete reperfusion and the thrombus composition. Conclusions: In patients presenting with occlusions of the MCA, a higher thrombus fibrin content is associated with better EVT efficiency. Both a higher MSB platelet/other components and RBC content may have a negative influence on EVT efficiency. These results may help identify preprocedural biomarkers beyond the conventional assessment of RBCs, WBCs, and fibrin compositions, which could guide decision-making during mechanical thrombectomy. Full article

Diabetic retinopathy (DR) is a microvascular disorder of the retina due to diabetes mellitus (DM). Natural products are widely used to prevent and treat various diseases caused by DM. This study aims to evaluate the effect of buah merah (Pandanus conoideus Lamk.) extract on retinal density and apoptosis in a diabetic rat model. A total of 30 male rats (Rattus norvegicus) weighing 120–150 g were induced with diabetes using alloxan and divided into five groups: group 1 (normal control), 2 (diabetic control), 3 (diabetes + 1 mL buah merah extract), 4 (1.5 mL), and 5 (2 mL). Buah merah extract, equivalent to 12 mg total carotenoids, 10 mg total tocopherols, 1.348 mg alpha-tocopherol, and 3.4 mg beta-carotene, was administered for 14 days. Retina was examined using hematoxylin and eosin (HE) staining for photoreceptor and retinal ganglion cell density, and immunohistochemistry (IHC) of Caspase-3 for apoptosis. The results showed that group 3 had photoreceptor and retinal ganglion cell densities close to normal, with photoreceptor density values of 722.52 ± 147.56 and ganglion 18.73 ± 5.61. The post hoc test confirmed a significant protective effect of buah merah extract in group 3 (p-value 0.014). However, buah merah extract was found to maintain photoreceptor and retinal ganglion cell density, but no significant inhibitory effect was observed on photoreceptor or retinal ganglion cell apoptosis. Further studies are needed to better understand the mechanism and potential therapeutic effect of buah merah extract. Full article

Background: RT-qPCR is the gold standard for quantitative gene expression analysis, but it requires homogenized tissue and thus loses spatial information. RNA in situ hybridization (ISH) preserves tissue localization but is technically challenging, especially in calcified tissues such as bone and teeth, where decalcification can damage RNA. RNAscope, an advanced ISH method with high sensitivity and specificity, has been applied successfully to bone, but its use in dental pulp remains largely unexplored despite the pulp’s crucial role in tooth function and health. Our goal was to identify the optimal decalcification process of mouse tooth samples for RNAscope ISH, which preserves RNA integrity in mouse tooth pulp. Methods: We tested five different decalcification procedures (EDTA, Plank-Rychlo solution, 5% formic acid, ACD decalcification buffer and Morse solution) on tooth samples from 3-month-old male C57BL/6J mice. Micro-CT and hematoxylin-eosin (HE) staining was performed to evaluate the decalcification, the quality and the microstructure of the sections. RNAscope ISH was used to examine mRNA integrity by analyzing the expression patterns of three housekeeping genes with different expression levels (low, medium and high). Results: All five decalcification methods demonstrated well-preserved tissue structure based on HE staining, but RNA integrity was only preserved in the case of mouse dental pulp using the ACD decalcification buffer and Morse’s solution. Conclusions: We successfully identified the optimal decalcification procedures preserving RNA integrity in mouse tooth samples, which may be useful for any target RNA examinations by RNAscope ISH in the future. Full article

Background/Objectives: This research aims to offer significant insights into the prospective application of bioactive hydrogels composed of alginate, gelatin, and grape skin extract from Serbia (GSE) for treating diabetic wounds, supporting the circular economy and environmental protection. Methods: An acute dermal irritation study was conducted according to OECD guidelines, revealing no visible signs of erythema or edema, confirming the hydrogel’s dermal safety. Afterwards, male Wistar rats were divided into four groups: untreated control (NC), silver sulfadiazine-treated (PC), hydrogel without extract (HG), and hydrogel with GSE (HG + GSE). Wound healing was assessed through a comprehensive approach that included macroscopic wound contraction; biochemical assessment of hydroxyproline content and oxidative stress markers (TBARS, SOD, CAT, GSH); quantification of inflammatory cytokines (TNF-α, IL-6); and histological examination of skin samples using hematoxylin–eosin (H&E) and Masson’s trichrome staining. Results: Daily HG+GSE application over 15 days accelerated wound closure, reaching 99.3% by day 15, surpassing PC (91.2%) and HG (87.7 ± 2.1%). Hydroxyproline levels followed a treatment-dependent pattern, with HG+GSE achieving the highest values throughout, reaching 6.78 ± 0.1 µg/mg dry tissue by day 15—more than double NC. The HG+GSE reduced lipid peroxidation while enhancing enzymatic and non-enzymatic antioxidant defenses and markedly lowered pro-inflammatory cytokine levels, indicating systemic anti-inflammatory activity. Histological analysis revealed faster re-epithelialization, increased collagen deposition, and more organized tissue architecture in the HG+GSE group. These outcomes are attributed to the sustained release of bioactive polyphenols such as naringin, caffeic acid, and epicatechin. Conclusions: Overall, this GSE-based hydrogel presents a multifunctional, biocompatible, sustainable, and effective strategy for diabetic wound care. Full article

Background/Objectives: Lung cancer (NSCLC), which accounts for approximately 85% of lung cancers, exhibits marked heterogeneity that complicates molecular characterization and treatment selection. Recent advances in deep learning (DL) have enabled the extraction of genomic-related morphological features directly from routine Hematoxylin and Eosin (H&E) histopathology, offering a potential complement to Next-Generation Sequencing (NGS) for precision oncology. This review aimed to evaluate how DL models have been applied to predict molecular alterations in NSCLC using H&E-stained slides. Methods: A systematic search following PRISMA 2020 guidelines was conducted across PubMed, Scopus, and Web of Science to identify studies published up to March 2025 that used DL models for mutation prediction in NSCLC. Eligible studies were screened, and data on model architectures, datasets, and performance metrics were extracted. Results: Sixteen studies met inclusion criteria. Most employed convolutional neural networks trained on publicly available datasets such as The Cancer Genome Atlas (TCGA) to infer key mutations including EGFR, KRAS, and TP53. Reported areas under the curve ranged from 0.65 to 0.95, demonstrating variable but promising predictive capability. Conclusions: DL-based histopathology shows strong potential for linking tissue morphology to molecular signatures in NSCLC. However, methodological heterogeneity, small sample sizes, and limited external validation constrain reproducibility and generalizability. Standardized protocols, larger multicenter cohorts, and transparent validation are needed before these models can be translated into clinical practice. Full article

Fish are widely recognized as effective bioindicators in ecotoxicological studies due to their repeated exposure to aquatic pollutants that accumulate in metabolically active organs, often leading to histopathological changes. In aquaculture, cage-farmed fish experience continuous environmental and culture-related stress, which can affect renal integrity. The kidney, a central osmoregulatory organ, is particularly sensitive to such conditions. Renal tissues were collected from different growth stages of cage-farmed rainbow trout. Hematoxylin and eosin staining was performed to detect morphological alterations, while transmission electron microscopy was used to assess cellular damage at the ultrastructural level. The expression of fibronectin and caspase-3, markers of extracellular matrix remodeling and apoptosis, respectively, was also evaluated. TEM examination showed pronounced alterations in both the glomeruli and renal tubules, accompanied by increased expression of fibronectin and caspase-3, indicating ongoing tissue remodeling and cellular stress. This study demonstrates that cage-farmed rainbow trout exhibit progressive ultrastructural kidney alterations that appear to be associated with environmental confinement, nutritional practices, and prophylactic treatments. These conditions collectively contribute to renal stress and the onset of nephropathic changes in aquaculture settings. Further research should focus on molecular marker expression to better understand renal adaptation and injury progression under intensive farming conditions. Full article

Background/Objectives: Regenerating critical-sized bone defects is a significant clinical challenge. Autogenous bone grafts are the gold standard but have limitations, including donor site morbidity. As an alternative, this study introduces a novel biocomposite combining an ethyl cyanoacrylate (ECA) polymer with Hancornia speciosa (Hs) latex. The ECA acts as a scaffold and delivery vehicle for the latex, which contains phytochemicals with known angiogenic properties. Methods: We created 5 mm critical-sized calvarial defects in 36 Wistar rats, which were divided into four experimental groups. Bone regeneration was evaluated at 30, 60, and 90 days using micro-computed tomography (micro-CT) for morphometric analysis and hematoxylin and eosin staining for histology. Results: The composite-treated group (Hs+ECA) showed significantly higher bone volume (57.2; IQR: 56.7–61.2) than the control (53.9; IQR: 49.4–56.4) and ECA-only (48.4; IQR: 47.2–59.9) groups at 90 days (p < 0.05). By day 60, the bone volume in the Hs+ECA group was statistically similar (p > 0.05) to that of the autogenous bone group. Histological analysis revealed an organized repair process with neoangiogenesis observed only in the Hs+ECA group, confirming the material’s strong bioactivity. Conclusions: The Hs+ECA composite is a promising biomaterial that acts as an effective delivery system for the bioactive components of the latex. The induced angiogenesis was critical to its regenerative success. This cost-effective material warrants further investigation for clinical applications in regenerative dentistry. Full article

Background/Objectives: Hip osteoarthritis (HOA) is a progressive joint disease characterized by cartilage loss, subchondral bone changes, and synovial inflammation. While tumor necrosis factor receptor 1 (TNFR1), interleukin-6 (IL-6), and transforming growth factor-beta 1 (TGF-β1) are recognized as key mediators of joint pathology, their compartment-specific expression in the human hip synovium remains insufficiently characterized. Therefore, we aimed to investigate their localization and expression in the intimal and subintimal compartments of synovial tissue in patients with HOA compared to controls (CTRL). Methods: Synovial membrane samples were obtained from 19 patients with primary HOA undergoing total hip arthroplasty and 10 CTRL subjects undergoing arthroplasty for acute femoral neck fracture without HOA. Specimens were processed for hematoxylin and eosin (H&E) and immunofluorescence staining. Expression of TNFR1, IL-6, and TGF-β1 was quantified in the intima and subintima using ImageJ analysis. Group differences were assessed using two-way Analysis of variance (ANOVA) with Tukey’s test when assumptions were met; for heteroscedastic outcomes we applied Brown–Forsythe ANOVA with Dunnett’s T3 multiple comparisons. Results: Histological analysis confirmed synovitis in HOA samples, with intimal hyperplasia and mononuclear infiltration. IL-6 was significantly upregulated in the intima of HOA synovium compared with CTRLs, while subintimal expression remained unchanged. In contrast, TGF-β1 expression was reduced in the HOA intima, eliminating the normal intima–subintima gradient. For TNFR1, the within-HOA contrast (int > sub) was significant, whereas the intimal HOA vs. CTRL comparison showed a non-significant trend. Transcriptomic analysis supported IL-6 upregulation, while TNFR1 and TGF-β1 did not reach statistical significance at the mRNA level in an orthogonal, non-hip (knee-predominant) dataset. Conclusions: These findings demonstrate compartment-specific cytokine dysregulation in HOA, with increased intimal TNFR1 and IL-6 alongside reduced intimal TGF-β1. The synovial lining emerges as a dominant site of inflammatory signaling, underscoring its importance in disease progression. Full article

Objective: This study aimed to evaluate peri-implant tissue changes during early osseointegration using a combined approach of digital radiographic analysis, RGB pseudocolorization, and histomorphometry in a sheep tibia model. Materials and Methods: Thirty titanium implants were placed in the tibiae of six adult sheep and evaluated at 14 and 28 days post-implantation. Digital periapical radiographs were acquired, grayscale values and RGB channel intensities were measured using Fiji/ImageJ, and compared with histological parameters (bone tissue, collagen, and medullary spaces) quantified from picrosirius–hematoxylin-stained sections. Manual overlay of radiographic and histological images was performed to ensure spatial correspondence of regions of interest. Statistical analyses assessed differences over time and correlations between image data and histological composition. Results: Radiographic grayscale values and histologically measured bone and collagen increased significantly from 14 to 28 days (p < 0.01), while medullary spaces decreased (p < 0.001), indicating progressive bone formation and matrix maturation. RGB analysis revealed significant increases in green channel intensity and decreases in red channel intensity (p < 0.05), while the blue channel remained stable. At 14 days, strong correlations were observed between blue channel intensity and bone tissue (r = 0.81; p = 0.015), and between green channel intensity and collagen (r = 0.98; p < 0.001). Visual overlays demonstrated alignment between radiographic high-density zones and histologically dense bone regions. Conclusions: RGB pseudocolorized radiographic analysis, correlated with histological findings, offers a non-invasive and reproducible method for early detection of peri-implant tissue maturation. This feasibility correlation study provides a foundation for future investigations integrating imaging, histology, and biomechanical testing. Full article

Mastitis is a common mammary gland disease in mammals that severely impairs lactation function, with Staphylococcus aureus (S. aureus) being the primary pathogenic bacterium. However, the molecular mechanisms underlying S. aureus-induced mastitis in sheep remain incompletely elucidated. This study employed RNA sequencing (RNA-SEq) technology to systematically analyze the dynamic transcriptomic characteristics of mammary tissue in small-tailed sheep (SHT) after S. aureus infection, aiming to clarify the molecular regulatory mechanism of the host immune response and its relationship with the occurrence of mastitis. Twelve lactating STH were selected to establish an S. aureus-induced mastitis model. Blood, milk, and tissue samples were collected at 0, 24, 48, and 72 h post-infection (hpi). The infected sheep exhibited typical mastitis symptoms, including exacerbated breast swelling, reduced milk yield, elevated udder temperature, and darker, more viscous milk. Hematoxylin–eosin (HE) staining revealed significant pathological changes over time, such as stromal hyperplasia, extensive inflammatory cell infiltration, severe necrosis and sloughing of mammary epithelial cells, and compromised tissue integrity. RNA-Seq analysis identified 1299 differentially expressed genes (DEGs), among which 75 core genes maintained stable expression throughout the infection time (24 hpi, 48 hpi, and 72 hpi). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses indicated that these DEGs were associated with metabolic processes, protein binding, Toll-like receptor signaling, and the NF-κB pathway. The PPI network analysis identified core hub genes including PTK2B, STAT3, and JAK1/3, providing critical evidence for therapeutic target screening. Furthermore, qPCR verification indicated that the expressions of innate immune receptors TLR2, TLR4, TLR7, and TLR10, as well as pro-inflammatory factors IL-1β, IL-16, TNF-α, type I interferon (IFN-α), and nuclear transcription factor NF-κB were significantly upregulated in a time-dependent manner (p < 0.05). In conclusion, this study delineated the dynamic response of ovine mammary tissue to S. aureus infection, systematically elucidated temporal gene expression patterns, and revealed the molecular mechanisms underlying the tissue’s initial defense against inflammatory challenges. Full article

Current histological staining methods for intestinal tissue analysis face limitations in simultaneously visualizing multiple tissue components, often requiring multiple sequential stains that increase processing time and tissue consumption. This proof-of-concept study aimed to define and develop a pentachromatic staining method for enhanced visualization of gastrointestinal tissue architecture. We developed the Pentachroma O-H method, an original protocol using readily available histological reagents (Alcian Blue pH 2.5, Weigert’s resorcin–fuchsin, Mayer’s hematoxylin, and Van Gieson’s solution) applied in an optimized sequence. The protocol was tested on healthy human ileum tissue obtained from surgical specimens as proof of concept. Thirty serial sections were stained with Pentachroma O-H and compared to adjacent sections stained with conventional hematoxylin–eosin (H&E) to document the emerging morphological characteristics of this original stain. Pentachroma O-H achieved distinct five-color differentiation in approximately 45 min: acidic mucins appeared turquoise–blue, collagen fibers red, elastic fibers black–purple, smooth muscle and erythrocyte cytoplasm yellow, and nuclei blue–black. The method clearly delineated intestinal architecture, including mucosal goblet cells, muscularis mucosae, connective tissue vasculature (parietal smooth muscle and elastic laminae), fibers, and cellular components, as well as lymphoid tissue aggregates and infiltrates, with improved contrast compared to H&E. All tissue components were simultaneously visualized in single sections with excellent morphological preservation. This first description of Pentachroma O-H demonstrates its capability to provide comprehensive ileum tissue visualization equivalent to multiple traditional special stains in a single, efficient protocol, offering significant potential advantages for gastrointestinal pathology assessment and warranting future validation studies across diverse tissue types and pathological conditions. Full article

Flap viability remains a major challenge in reconstructive surgery due to ischemia–reperfusion injury, excessive inflammation, and impaired tissue regeneration. Boron, a trace element with pro-healing and anti-inflammatory properties, has shown therapeutic promise in various wound models; however, its role in flap healing remains unclear. In this study, we aimed to evaluate the therapeutic potential of sodium pentaborate pentahydrate (SPP)-containing hydrogel, a boron compound we developed, for enhancing flap survival and tissue repair. A dorsal random-pattern flap model was established in male Wistar rats, which were treated topically with an SPP-containing formulation twice daily for seven days. Histological changes were evaluated using hematoxylin–eosin and Masson’s trichrome staining, and proteomic alterations were analyzed using label-free nanoLC-MS/MS followed by bioinformatics analysis. The treatment significantly improved flap survival (p < 0.0001), enhanced granulation tissue formation, promoted organized collagen deposition, and reduced inflammatory infiltration. Proteomic profiling identified 179 differentially expressed proteins, with 14 upregulated and 165 downregulated. Upregulated proteins were enriched in pathways related to complement activation, antioxidant defense, and extracellular matrix remodeling, whereas downregulated proteins were associated with immune overactivation, cellular stress, and senescence, indicating a shift toward regulated inflammation and tissue homeostasis. To our knowledge, this is the first study to demonstrate that an SPP-containing hydrogel promotes flap healing by supporting vascularization, modulating immune responses, and enhancing extracellular matrix remodeling. These findings highlight SPP as a promising therapeutic strategy for improving flap viability in reconstructive surgery. Full article

Colorectal cancer (CRC) is one of the most common causes of cancer-related deaths globally, making a timely and reliable diagnosis essential. Manual histopathology assessment, though clinically standard, is prone to observer variability, while existing computational approaches often trade accuracy for interpretability, limiting their clinical utility. This paper introduces a deep learning framework that couples the YOLOv8 architecture for multiclass lesion classification with EigenCAM for transparent model explanations. The pipeline integrates three core stages: (i) acquisition and preprocessing of 5000 hematoxylin-and-eosin-stained slides from the University Medical Center Mannheim, categorized into eight tissue types; (ii) comparative evaluation of five YOLOv8 variants (Nano, Small, Medium, Large, XLarge); and (iii) interpretability through EigenCAM visualizations to highlight discriminative regions driving predictions. Extensive statistical validation (including box plots, empirical cumulative distribution functions, Bland–Altman plots, and pair plots) demonstrated the robustness and reliability of the framework. The YOLOv8 XLarge model achieved 99.38% training accuracy and 96.62% testing accuracy, outperforming recent CNN- and Transformer-based systems (≤95%). This framework establishes a clinically dependable foundation for AI-assisted CRC diagnosis by uniting high precision with visual interpretability. It represents a significant step toward real-world deployment in pathology workflows. Full article

Helicobacter pylori is a gastric pathogen that induces chronic gastritis, which may progress to neutrophilic activity, glandular atrophy, intestinal metaplasia, and gastric carcinoma. The aim of this study was to evaluate H. pylori-induced tissue damage. A total of 602 gastric biopsy samples were collected, categorized, and analyzed using hematoxylin and eosin and Giemsa staining, followed by molecular confirmation through PCR targeting the species-specific 16S rRNA gene. H. pylori density and histopathological features were evaluated and graded according to the updated Sydney classification system. H. pylori was detected in 55% (n = 334) of cases, and the antrum (50.83%, p < 0.00001) was the predominant site. A slightly higher prevalence was observed in females, accounting for 56.9% compared to males at 43.1%, which was attributed to sociocultural exposure differences. Individuals aged 11–40 years accounted for 58.3% (n = 195), highlighting early-age acquisition of infection. H. pylori infection was significantly linked to moderate-to-severe inflammation (63.2%, p < 0.00001) and neutrophilic activity (53.3%, p < 0.00001). Intestinal metaplasia and atrophy were infrequent, present in 0.6% (95% CI, 0.02, p = 0.149) and 0.9% (95% CI, 0.05, p = 0.430) of individuals. H. pylori infection causes chronic inflammation and neutrophilic infiltration of the stomach mucosa. Early identification and histopathological examination are essential in assessing H. pylori-related gastric pathology. Full article

Background/Objectives: Chronic colitis presents a growing global health burden with rising incidence. This study investigated the ameliorative effect of Dendrobium officinale polysaccharide (DOP) against dextran sulfate sodium (DSS)-induced chronic colitis in mice, specifically examining its dual modulation of gut microbiota and metabolic pathways. Methods: DOP was extracted and purified from Dendrobium officinale stems and leaves. A chronic colitis model was established in male C57BL/6J mice via DSS induction. Eighty-four mice were randomized into seven groups: control, model, low/high-dose leaf-DOP, low/high-dose stem-DOP, and sulfasalazine positive control. We assessed body weight, disease activity index (DAI), colon length, splenic/thymic indices, inflammatory cytokines, and histopathology (Hematoxylin and Eosin/Alcian blue staining), with tight junction protein and tumor necrosis factor-alpha (TNF-α) expression quantified via immunofluorescence. 16S rRNA sequencing and untargeted metabolomics evaluated microbial and metabolic shifts. Results: DOP significantly attenuated colitis severity, restored colon histoarchitecture, elevated goblet cell counts, upregulated zonula occludens-1 (ZO-1) and occludin expression, and suppressed TNF-α. Crucially, DOP remodeled dysbiosis by enriching beneficial taxa (e.g., Candidatus_Saccharimonas, Lachnoclostridium) while reducing pathogens (Mucispirillum). Metabolomics further elucidated DOP-mediated regulation of purine and nicotinate/nicotinamide metabolism—pathways mechanistically linked to its anti-inflammatory and barrier-repair effects. Conclusions: DOP effectively alleviates symptoms of DSS-induced chronic colitis in mice, protects intestinal barrier integrity, and achieves therapeutic potential through simultaneous regulation of the gut microbiome and metabolome. Full article