Search Results (1,747)

Background: Anti-programmed death 1 receptor (PD-1) therapy is a promising treatment strategy for patients with unresectable advanced or recurrent gastric/gastroesophageal junction (G/GEJ) cancer. However, its response rate and survival benefits are still limited; an immunological analysis of the residual tumor after anti-PD-1 therapy would be important. Methods: We evaluated the clinical efficacy of tumor resection (TR) after chemotherapy or anti-PD-1 therapy in patients with unresectable advanced or recurrent G/GEJ cancer and analyzed the immune status of tumor microenvironment (TME) by immunohistochemistry using their surgically resected specimens. Results: Patients treated with TR after anti-PD-1 therapy had significantly longer survival compared to those treated with chemotherapy and anti-PD-1 therapy alone. Expression of human leukocyte antigen (HLA) class I and major histocompatibility complex (MHC) class II on tumor cells was markedly downregulated after anti-PD-1 therapy compared to chemotherapy. Furthermore, the downregulation of HLA class I may be associated with the activation of transforming growth factor-β signaling pathway in the TME. Conclusions: Immune escape from cytotoxic T lymphocytes may be induced in the TME in patients with unresectable advanced or recurrent G/GEJ cancer after anti-PD-1 therapy due to the downregulation of HLA class I and MHC class II expression on tumor cells. TR may be a promising treatment strategy for these patients when TR is feasible after anti-PD-1 therapy. Full article

Background/Objectives: Aberrant expression of high-mobility group protein B1 (HMGB1) has been linked to cancer development and progression. Methods: To better comprehend the role of HMGB1 expression in cancer, a tissue microarray containing 14,966 samples from 134 different tumor entities and 608 samples of 76 different normal tissue types was analyzed by immunohistochemistry. Results: Strong HMGB1 staining occurred in almost all normal cell types and in most cancers. Of 11,808 evaluable cancers, only 7.8% showed complete absence of HMGB1 staining (HMGB1 deficiency) while 9.9% showed 1+, 25.0% showed 2+, and 57.2% showed 3+ HMGB1 positivity. Absence of HMGB1 staining mostly occurred in pheochromocytoma (90.0%), seminoma (72.4%), gastrointestinal stromal tumor (28.6%), adrenal cortical carcinoma (25.0%), and Hodgkin’s lymphoma (25.0%). Low HMGB1 staining was linked to poor histologic grade (p < 0.0001), advanced pT stage (p < 0.0001), high UICC stage (p < 0.0001), and distant metastasis (p = 0.0413) in clear cell renal cell carcinoma, invasive tumor growth in urothelial carcinoma (pTa vs. pT2–4, p < 0.0001), mismatch repair deficiency (p = 0.0167) in colorectal cancers, and advanced pT stage in invasive breast carcinoma of no special type (p = 0.0038). Strong HMGB1 staining was linked to nodal metastases in high-grade serous ovarian carcinomas (p = 0.0213) and colorectal adenocarcinomas (p = 0.0137), as well as to poor histological grade in squamous cell carcinomas (p = 0.0010). Conclusions: HMGB1 deficiency and reduced HMGB1 expression occur in a broad range of different tumor entities. Low rather than strong HMGB1 staining is often linked to an aggressive tumor phenotype. Whether HMGB1 deficiency renders cells susceptible to specific drugs remains to be determined. Full article

The insulin-like growth factor (IGF) system regulates implantation, placental development, and angiogenesis in eutherian mammals. However, little is known about the changes in this system in equine placenta (chorioallantois; CA) and the endometrium (EN) during pregnancy, or the relationship to vascular endothelial growth factor (VEGF) expression. The current study investigated the expression of the IGF system components, namely the ligands (IGF1 and IGF2), their receptors (IGF1R, IGF2R, and INSR), and their binding proteins (IGFBPs and IGF2BPs) in equine CA at 45 days, 4, 6, 10, and 11 months of gestational age (GA) and immediately postpartum (PP), and in equine EN at 4, 6, 10, and 11 months GA. IGF1 immunolocalization and serum concentrations were also evaluated across gestation. IGF1 mRNA expression in CA increased from day 45 to peak at 6 months and then gradually declined to reach a nadir in PP samples. This profile correlated positively with the VEGF expression profile (r = 0.62, p = 0.001). In contrast, IGF2 expression in CA was not correlated with VEGF (p = 0.14). Interestingly, IGF2 mRNA was more abundant in equine CA than IGF1 (p < 0.05) throughout gestation. Among the IGFBPs investigated in CA, the expression of IGFBP2 and IGF2BP2 was highly abundant (p < 0.05) at day 45 compared to other GAs. Conversely, mRNA expression for IGFBP3 and IGFBP5 was more abundant (p < 0.05) in PP than at all investigated GAs. Immunohistochemistry revealed that IGF1 is localized in the equine chorionic epithelium (cytoplasm and nucleus). IGF1 serum concentrations peaked at 9 months and declined to their lowest levels PP. In conclusion, this study demonstrates a positive correlation between IGF1 and VEGF expression in equine CA during gestation, suggesting that the IGF system plays a crucial role in placental angiogenesis by regulating VEGF. Full article

To determine the association between FGFR4 (rs351855 and rs7708357) gene variants, serum levels, and immunohistochemical markers (Ki-67 and p53) in pituitary adenoma (PA), a case-control study was conducted involving 300 subjects divided into two groups: the control group (n = 200) and a group of PA (n = 100). The genotyping of FGFR4 rs351855 and rs7708357 was carried out using the real-time polymerase chain reaction (RT-PCR) method. The serum FGFR4 levels were measured using the ELISA method. Immunohistochemical analysis (Ki-67 and p53) was conducted. Statistical analysis of the data was performed using IBM SPSS Statistics 30.0 software. There were no statistically significant differences after analyzing the genotypes and alleles of FGFR4 rs351855 and rs7708357 in patients with PA and control groups (all p > 0.05). After evaluating the distribution of genotypes and alleles of FGFR4 rs351855 and rs7708357 in micro/macro, invasiveness, activity, and recurrence of PA and the control groups, the analysis showed no statistically significant differences between the groups (p > 0.05). Similarly, no significant differences in FGFR4 levels were observed between PA patients and control group (median (IQR): 3642.41 (1755.08) pg/mL vs. 3126.24 (1334.15) pg/mL, p = 0.121). Immunohistochemistry for Ki-67 revealed a labeling index (LI) of <1% in 25.5% of patients with PA, an LI of 1% in 10.9%, and an LI of >1% in 63.6% of patients. Further analyses showed no statistically significant associations with tumor size, invasiveness, activity, or recurrence. Immunohistochemistry for p53 revealed that macroadenomas had a significantly higher p53 H-score compared to microadenomas (median (IQR): 30.33 (28.68) vs. 18.34 (17.65), p = 0.005). Additionally, a moderate, statistically significant positive correlation between the Ki-67 LI and the p53 expression was found (Spearman’s ρ = 0.443, p = 0.003, n = 43). FGFR4 variants and serum protein levels were not significantly associated with PA risk or tumor features. Conversely, immunohistochemical markers Ki-67 and p53 were more informative, with higher p53 expression in macroadenomas and a moderate positive correlation between Ki-67 and p53, highlighting their potential relevance in tumor growth assessment. Full article

Glioblastoma (GBM) is the most aggressive primary brain tumor, characterized by rapid progression, profound heterogeneity, and resistance to conventional therapies. This review provides an integrated overview of GBM’s pathophysiology, highlighting key mechanisms such as neuroinflammation, genetic alterations (e.g., EGFR, PDGFRA), the tumor microenvironment, microbiome interactions, and molecular dysregulations involving gangliosides and sphingolipids. Current diagnostic strategies, including imaging, histopathology, immunohistochemistry, and emerging liquid biopsy techniques, are explored for their role in improving early detection and monitoring. Treatment remains challenging, with standard therapies—surgery, radiotherapy, and temozolomide—offering limited survival benefits. Innovative therapies are increasingly being explored and implemented, including immune checkpoint inhibitors, CAR-T cell therapy, dendritic and peptide vaccines, and oncolytic virotherapy. Advances in nanotechnology and personalized medicine, such as individualized multimodal immunotherapy and NanoTherm therapy, are also discussed as strategies to overcome the blood–brain barrier and tumor heterogeneity. Additionally, stem cell-based approaches show promise in targeted drug delivery and immune modulation. Non-conventional strategies such as ketogenic diets and palliative care are also evaluated for their adjunctive potential. While novel therapies hold promise, GBM’s complexity demands continued interdisciplinary research to improve prognosis, treatment response, and patient quality of life. This review underscores the urgent need for personalized, multimodal strategies in combating this devastating malignancy. Full article

Background/Objectives: Degenerative processes of the hip joint increasingly affect not only the articular cartilage but also periarticular structures such as the joint capsule and acetabular labrum. This study aimed to investigate the structural and molecular changes occurring in these tissues during advanced hip osteoarthritis. Methods: A combined analysis using immunohistochemistry (IHC), scanning electron microscopy (SEM), and micro-computed tomography (microCT) was conducted on tissue samples from patients undergoing total hip arthroplasty and from controls with morphologically normal joints. Markers associated with proliferation (Ki67), inflammation (CD68), angiogenesis (CD31, ERG), chondrogenesis (SOX9), and lubrication (Lubricin) were evaluated. Results: The pathological group showed increased expression of Ki67, CD68, CD31, ERG, and SOX9, with a notable decrease in Lubricin. SEM analysis revealed ultrastructural disorganization, collagen fragmentation, and neovascular remodeling in degenerative samples. A significant correlation between structural damage and molecular expression was identified. Conclusions: These results suggest that joint capsule and acetabular labrum degeneration are interconnected and reflect a broader pathophysiological continuum, supporting the use of integrated IHC and SEM profiling for early detection and targeted intervention in hip joint disease. Full article

Diffuse axonal brain injury (DAI) is a common, debilitating consequence of traumatic brain injury, yet its detection and severity grading remain challenging in clinical and experimental settings. This study evaluated the sensitivity of diffusion tensor imaging (DTI), histology, and neurological severity scoring (NSS) in assessing injury severity in a rat model of isolated DAI. A rotational injury model induced mild, moderate, or severe DAI in male and female rats. Neurological deficits were assessed 48 h after injury via NSS. Magnetic resonance imaging, including DTI metrics, such as fractional anisotropy (FA), relative anisotropy (RA), axial diffusivity (AD), mean diffusivity (MD), and radial diffusivity (RD), was performed prior to tissue collection. Histological analysis used beta amyloid precursor protein immunohistochemistry. Sensitivity and variability of each method were compared across brain regions and the whole brain. Histology was the most sensitive method, requiring very small groups to detect differences. Anisotropy-based MRI metrics, especially whole-brain FA and RA, showed strong correlations with histology and NSS and demonstrated high sensitivity with low variability. NSS identified injury but required larger group sizes. Diffusivity-based MRI metrics, particularly RD, were less sensitive and more variable. Whole-brain FA and RA were the most sensitive MRI measures of DAI severity and were comparable to histology in moderate and severe groups. These findings support combining NSS and anisotropy-based DTI for non-terminal DAI assessment in preclinical studies. Full article

Background/Objectives: Eph receptor A5 (EphA5) is a receptor tyrosine kinase that is implicated in multiple malignancies, although its role in endometrial cancer (EC) remains unclear. The aim of this study was to investigate the clinicopathological significance of EphA5 expression in EC and explore its association with proliferative and metabolic markers. Methods: We retrospectively analyzed 75 EC tissue samples from treatment-naïve patients by using immunohistochemistry and H-score quantification. Associations between EphA5 expression and clinicopathological parameters were assessed through logistic regression analysis. Kaplan–Meier analysis was used to evaluate survival outcomes. Correlation analysis, stratified according to cancer stage, was used to explore biomarker interactions. Results: High EphA5 expression levels were significantly associated with elevated Ki-67 expression (adjusted odds ratio (aOR): 1.08 per 1-point H-score increase, p = 0.024) and decreased pAMPK expression (aOR: 0.89 per 1-point H-score increase, p = 0.024), indicating its involvement in proliferative and metabolic pathways. Paradoxically, patients with high EphA5 levels had significantly better overall survival probabilities (H-score > 105, log-rank p = 0.007). Stage-specific analyses suggested that EphA5 levels correlated with proliferation in early-stage disease and epithelial–mesenchymal transition in advanced stages. Conclusions: EphA5 may act as a context-dependent biomarker in EC. Despite its positive correlation with proliferation and negative association with metabolic stress signaling, high EphA5 expression levels were predictive of a favorable prognosis. Full article

Background/Objectives: Diatomaceous earth (DE), a natural substance rich in amorphous silica and recognized as a food additive, is gaining attention as a dietary silicon supplement. However, its bioavailability and impact on lipid digestion and absorption remain poorly characterized. This study aimed to investigate silicon bioavailability after short-term DE supplementation and its effects on postprandial glycemia and triglyceridemia, the expression of lipid metabolism-related proteins, and the modulation of the intestinal mucosal barrier. Methods: Female Wistar rats received daily oral supplementation of DE (equivalent to 2 or 4 mg silicon/kg body weight) for one week. Silicon digestibility, excretion, and hepatic accumulation were quantified. Postprandial glycemia and triglyceridemia were monitored. Lipid profile was analyzed by HPSEC in gastric and intestinal contents. Jejunal morphology and mucin-secreting cells were assessed histologically. Lipid metabolism markers were evaluated by immunohistochemistry and Western blot in both intestinal and hepatic tissues. Results: DE supplementation enhanced silicon absorption and increased hepatic levels. Fecal output and moisture content were also elevated, especially at the higher dose. DE significantly reduced postprandial triglyceridemia and consequently increased luminal triglyceride retention. These changes were associated with decreased jejunal levels of IFABP, ACAT2, and MTP, as well as reduced hepatic levels of MTP and LDLr, alongside increased levels of ABCG5/G8 and LXRα/β, indicating a partial blockage of lipid absorption and enhanced cholesterol efflux. The effects on the intestinal barrier were evidenced by villi shortening and an increase in mucin-producing cells. Conclusion: Food-grade DE is a bioavailable source of silicon with hypolipidemic potential, mainly by reducing intestinal lipid absorption. This is supported by lower postprandial triglycerides, increased luminal lipid retention, and decreased expression of lipid transport proteins. The study in healthy female rats underscores the importance of sex-specific responses and supports DE as a dietary strategy to improve lipid metabolism. Full article

Gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs) are rare tumors with different clinical and biological characteristics. Ki-67 staining and mitotic counts are the most commonly used prognostic markers, but these methods are time-consuming and lack reproducibility, highlighting the need for innovative approaches that improve histological evaluation and prognosis. In our previous study, we observed that the microRNA (miRNA) expression profile of GEP-NENs correlates with the three grades of GEP-NENs. This study aimed to characterize a group of miRNAs that discriminate well-differentiated GEP-NENs grading 1 (G1) and grading (G2). Fifty formalin-fixed and paraffin-embedded tissue specimens from well-differentiated GEP-NENs G1 and G2 tissues were used for this study. The expression levels of 21 miRNAs were examined using qRT-PCR, while FGFR2 and FGF1 protein expression were evaluated through immunohistochemistry (IHC). We identified four miRNAs (hsa-miR-133, hsa-miR-150-5p, hsa-miR-143-3p and hsa-miR-378a-3p) that are downregulated in G2 GEP-NENs compared to G1. Bioinformatic analysis revealed that these miRNAs play a key role in modulating the FGF/FGFR signaling pathway. Consistent with this observation, we found that fibroblast growth factor receptor 2 (FGFR2) expression is markedly higher in G2 NENs patients, whereas its expression remains low in G1 NENs. Our findings highlight the potential use of miRNAs to confirm the histological evaluation of GEP-NENs by employing them as biomarkers for improving histological evaluation and tumor classification. Full article

Background/Objectives: Endometrial cancer (EC) is the most prevalent gynecological malignancy, with increasing incidence and mortality. HOXA5, a developmental transcription factor, has been linked to prognosis in various cancers, but its role in EC remains unclear. This study aimed to evaluate the prognostic potential of HOXA5 in EC and to explore its association with common tumor-related proteins. Methods: We analyzed 75 EC tissue samples using immunohistochemistry to evaluate HOXA5 expression and its association with clinicopathological features and tumor-related biomarkers, including Ki-67, CD31, and fibronectin. Statistical analyses included logistic regression and Kaplan–Meier survival analysis. Results: High HOXA5 expression was significantly associated with elevated Ki-67 levels (p = 0.001) but paradoxically correlated with improved overall survival (p = 0.026). CD31 and fibronectin levels were significantly lower in the high-HOXA5 group (p = 0.007 and p = 0.001, respectively), suggesting reduced angiogenic and invasive potential. However, neither marker remained significant in multivariable analysis. Conclusions: HOXA5 may exert a dual role in EC by promoting proliferation while limiting tumor progression via suppression of angiogenesis and matrix remodeling. It holds potential as a prognostic biomarker and therapeutic target. Full article

Intestinal alkaline phosphatase (IAP) is a brush border enzyme secreted by enterocytes, playing a crucial role in maintaining gut mucosal defense. This study investigated the expression dynamics of IAP in the small intestine of pigs challenged with Escherichia coli (E. coli) K88, compared to healthy controls. Five-week-old pigs (n = 8) were orally administered E. coli K88 at a concentration of 2 × 10⁸ CFU/mL, with a dose of 2 mL per pig at 0 and 24 h. Five days post-challenge, tissue samples from the duodenum, jejunum, and ileum were collected for mucosal morphometric analysis and evaluation of IAP expression via immunohistochemistry, Western blotting, and real-time PCR. The results revealed the presence of IAP on the apical surface of villi throughout the small intestine, along with significantly upregulated IAP expression in E. coli-challenged pigs compared to controls. These findings suggest that Gram-negative bacteria such as E. coli can induce IAP expression, likely through lipopolysaccharide (LPS) stimulation, thereby enhancing its enzymatic activity as part of the intestinal defense mechanism. This study provides insight into the protective role of IAP and highlights its potential as a biomarker for assessing gut health and diagnosing enteric infections in animals. Full article

Background: Gastric cancer (GC) is a malignant tumor of the gastrointestinal tract, characterized by high mortality rates and responsible for about one million new cases each year globally. Surgery is the main treatment, but achieving radical resection remains a relevant intraoperative challenge. Fluorescence-guided surgery offers clinicians greater capabilities for real-time detection of tumor nodules and visualization of tumor margins. In this field, the main challenge remains the development of fluorescent dyes that can selectively target tumor tissues. Methods: we examined the expression of the most suitable GC markers, including carcinoembryonic antigen cell adhesion molecule-5 (CEACAM5) and Claudin-4 (CLDN4), in GC cell lines. To further evaluate their expression, we performed immunohistochemistry (IHC) on tumor and healthy tissue samples from 30 GC patients who underwent partial gastrectomy at the Digestive System Surgery Unit, AOU Careggi, Florence. Additionally, we validated anti-CEACAM5 expression on patient-derived organoids. Furthermore, we developed a fluorescent molecule targeting CEACAM5 on the surface of GC cells and assessed its binding properties on patient tissue slices and fragments. Results: in this work, we first identified CEACAM5 as an optimal GC biomarker, and then we developed a fluorescent antibody specific for CEACAM5. We also evaluated its binding specificity for GC cell lines and patient-derived tumor tissue, achieving an optimal ability to discriminate tumor tissue from healthy mucosa. Conclusions: Overall, our results support the development of our fluorescent antibody as a promising tumor-specific imaging agent that, after further in vivo validation, could improve the accuracy of complete tumor resection. Full article

This study aimed to investigate the tyrosine kinase inhibitor Nintedanib and its potential role in reversing epithelial–mesenchymal transition (EMT) induced by transforming growth factor beta 2 (TGF-β2) in retinal pigment epithelial (RPE) cells, along with its therapeutic potential using a mouse model of subretinal fibrosis. We hypothesized that the blockade of angiogenesis promoting and fibrosis inducing signaling using the receptor tyrosine kinase inhibitor Nintedanib (OfevTM) can prevent or reverse EMT both in vitro and in our in vivo model of subretinal fibrosis. Primary human retinal pigment epithelial cells (phRPE) and adult retinal pigment epithelial cell line (ARPE-19) cells were treated with TGF-β210 ng/mL for two days followed by four days of Nintedanib (1 µM) incubation. Epithelial and mesenchymal phenotypes were assessed by morphological examination, quantitative real-time polymerase chain reaction(qPCR) (ZO-1, Acta2, FN, and Vim), and immunocytochemistry (ZO-1, vimentin, fibronectin, and αSMA). Metabolites were measured using luciferase-based assays. Extracellular acidification and oxygen consumption rates were measured using the Seahorse XF system. Metabolic-related genes (GLUT1, HK2, PFKFB3, CS, LDHA, LDHB) were evaluated by qPCR. A model of subretinal fibrosis using the two-stage laser-induced method in C57BL/6J mice assessed Nintedanib’s therapeutic potential. Fibro-vascular lesions were examined 10 days later via fluorescence angiography and immunohistochemistry. Both primary and ARPE-19 RPE stimulated with TGF-β2 upregulated expression of fibronectin, αSMA, and vimentin, and downregulation of ZO-1, consistent with morphological changes (i.e., elongation). Glucose consumption, lactate production, and glycolytic reserve were significantly increased in TGF-β2-treated cells, with upregulation of glycolysis-related genes (GLUT1, HK2, PFKFB3, CS). Nintedanib treatment reversed TGF-β2-induced EMT signatures, down-regulated glycolytic-related genes, and normalized glycolysis. Nintedanib intravitreal injection significantly reduced collagen-1+ fibrotic lesion size and Isolectin B4+ neovascularization and reduced vascular leakage in the two-stage laser-induced model of subretinal fibrosis. Nintedanib can induce Mesenchymal-to-Epithelial Transition (MET) in RPE cells and reduce subretinal fibrosis through metabolic reprogramming. Nintedanib can therefore potentially be repurposed to treat retinal fibrosis. Full article

Extracellular vesicles (EVs) have emerged as promising acellular tools for modulating immune responses for tissue engineering applications. This study explores the potential of human fibroblast-derived EVs delivered within a three-dimensional (3D) injectable scaffold composed of polycaprolactone (PCL) nanofibers and collagen (PNCOL) to reprogram macrophage behavior and support scaffold integrity under inflammatory conditions. EVs were successfully isolated from human fibroblasts using ultracentrifugation and characterized for purity, size distribution and surface markers (CD63 and CD9). Macrophage-laden PNCOL scaffolds were prepared under three conditions: macrophage-only (MP), fibroblast co-encapsulated (F-MP), and EV-encapsulated (EV-MP) groups. Structural integrity was assessed via scanning electron microscopy and Masson’s trichrome staining, while immunomodulatory effects were evaluated through metabolic assays, gene expression profiling, and immunohistochemistry for macrophage polarization markers (CD80, CD206). When co-encapsulated with pro-inflammatory (M1) macrophages in PNCOL scaffolds, fibroblast-derived EVs preserved scaffold structure and significantly enhanced macrophage metabolic activity compared to the control (MP) and other experimental group (F-MP). The gene expression and immunohistochemistry data demonstrated substantial upregulation of anti-inflammatory markers (TGF-β, CD163, and CCL18) and surface protein CD206, indicating a phenotypic shift toward M2-like macrophages for EV-encapsulated scaffolds relative to the other groups. The findings of this study demonstrate that fibroblast-derived EVs integrated into injectable PCL–collagen scaffolds offer a viable, cell-free approach to modulate inflammation, preserve scaffold structure, and support regenerative healing. This strategy holds significant promise for advancing immuno-instructive platforms in regenerative medicine, particularly in settings where conventional cell therapies face limitations in survival, cost, or safety. Full article