Search Results (916)

In vitro cartilage explant culture has been used to assess nutraceuticals on cartilage responses to inflammatory stimuli. However, applying extracts of nutraceuticals directly to cartilage explants does not account for effects of digestion and hepatic biotransformation, or selective exclusion of product metabolites from joint fluid by the synovial membrane. The current study produced a simulated biological extract of a common nutraceutical (glucosamine; G_sim) by exposing it to a simulated upper gastrointestinal tract digestion, hepatic biotransformation by liver microsomes, and purification to a molecular weight cut-off of 50 kDa. This extract was then used to condition cartilage explants cultured for 120 h in the presence or absence of an inflammatory stimulus (lipopolysaccharide). Media samples were analyzed for prostaglandin E₂ (PGE₂), glycosaminoglycan (GAG), and nitric oxide (NO). Tissue was digested and analyzed for GAG content and stained for viability. Conditioning of explants with G_sim significantly reduced media GAG in stimulated and unstimulated explants and reduced nitric oxide production in unstimulated explants. These data provide evidence for the value of glucosamine in protecting cartilage from deterioration following an inflammatory challenge, and the model improves applicability of these in vitro data to the in vivo setting. Full article

Alkyd resins are among the most common coatings used for exterior wood joinery. In Romania, solvent-borne alkyd coatings are widely used to finish wood. The study aims to compare the performance after 7 years of outdoor exposure of two types of alkyd coatings, a semi-transparent brown stain with micronized pigments (Alk1) and an opaque white enamel (Alk2), applied directly on wood or wood pre-treated with three types of resins: acryl-polyurethane (R1), epoxy (R2), and alkyd-polyurethane (R3). Fir (Abies alba) wood served as the substrate. Cracking, coating adhesion, and biological degradation were periodically assessed through visual inspection and microscopy. Additionally, a cross-cut test was performed, and the loss of coating on the directly exposed upper faces was measured using ImageJ. The results indicated that resin pretreatments somewhat reduced cracking but negatively affected coating adhesion after long-term exposure. All samples pretreated with resins and coated with Alk1 lost more than 50% (up to 78%) of the original finishing film by the end of the test. In comparison, coated control samples lost less than 50%. The Alk2 coating exhibited a film loss between 2% and 12%, compared to an average loss of 9% for the coated control. Overall, samples pretreated with alkyd-polyurethane resin (R3) and coated with alkyd enamel (Alk2) demonstrated the best performance in terms of cracking, adhesion, and discoloration. Full article

Bifora testiculata (L.) Spreng. (Apiaceae), an understudied species endemic to the Mediterranean and the only representative species of the genus Bifora in Sicily, was investigated for the first time for its essential oil (EO) chemical composition and cytotoxic properties. The EO was obtained via hydrodistillation and analyzed using GC-MS, revealing an aldehyde-rich profile (86.10%), dominated by trans-2-dodecenal (67.49%). Comparative analysis with previous studies on B. testiculata from Greece confirmed a similar aldehyde-rich profile, although minor compositional differences suggest potential chemotype variation. Given the biological relevance of trans-2-dodecenal and related aldehydes, further investigations into the cytotoxic properties of the EO of B. testiculata (Bt) and its main constituent against cancer cell lines were undertaken. Three human tumor cell lines (MDA-MB 231, A375, and CaCo2) and a human non-tumor cell line (HEK293) were subjected to viability tests using the MTT assay. The EO and trans-2-dodecenal exhibited remarkable cytotoxic activity against all cell lines, with IC₅₀ values ranging between 7.93 and 14.41 µg/mL for Bt and between 1.88 and 5.29 µg/mL for trans-2-dodecenal. AO/BE fluorescent staining and Hoechst nuclear staining showed the presence of apoptotic bodies in the treated cells. N-acetyl-L-cysteine was able to invert the effects of Bt and trans-2-dodecenal on cell lines, suggesting ROS involvement in cytotoxic activity. The results demonstrated that the Bt cytotoxic activity was mainly due to the presence of trans-2-dodecenal. 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

F₄-Neuroprostanes (F₄-NeuroPs), oxidative metabolites of docosahexaenoic acid, act as bioactive lipid mediators enhancing sperm motility and induce capacitation-like changes in vitro. Their biological action is proposed to involve sperm ion channels, in particular ryanodine receptors (RyRs), which regulate intracellular calcium homeostasis. We evaluated the effects of dantrolene, a RyR inhibitor, on motility and vitality of a selected spermatozoa at different concentrations (10, 30, 50, 100 μM). Then sperm motility, acrosome integrity, and RyR localization following co-incubation with dantrolene (D50 or D100 μM) and 4-/10-F_4t-NeuroPs (7 ng) were investigated. Acrosomal status was assessed using Pisum sativum agglutinin (PSA) staining and RyR localization by immunofluorescence. D50 was identified as the minimum effective dose to induce significant reductions in sperm motility. F₄-NeuroPs significantly increased rapid progressive motility versus controls. Co-incubation with F₄-NeuroPs + D50 reduced rapid motility and increased in situ and circular movement. The acrosome staining appeared altered or absent to different percentages, and RyR localization was also seen in the midpiece. These findings suggested that F₄-NeuroPs enhance sperm motility via RyR-mediated pathways, as confirmed by dantrolene inhibition. Accordingly, our results underscore the physiological relevance of RyRs in sperm function and suggest new insights into lipid-based mechanisms regulating sperm motility. Full article

Triple-negative breast cancer (TNBC) remains a major clinical challenge due to its aggressive behavior and lack of targeted therapies. Accurate early prediction of response to neoadjuvant chemotherapy (NACT) is essential for guiding personalized treatment strategies and improving patient outcomes. In this study, we present an attention-based multiple instance learning (MIL) framework designed to predict pathologic complete response (pCR) directly from pre-treatment hematoxylin and eosin (H&E)-stained biopsy slides. The model was trained on a retrospective in-house cohort of 174 TNBC patients and externally validated on an independent cohort (n = 30). It achieved a mean area under the curve (AUC) of 0.85 during five-fold cross-validation and 0.78 on external testing, demonstrating robust predictive performance and generalizability. To enhance model interpretability, attention maps were spatially co-registered with multiplex immunohistochemistry (mIHC) data stained for PD-L1, CD8+ T cells, and CD163+ macrophages. The attention regions exhibited moderate spatial overlap with immune-enriched areas, with mean Intersection over Union (IoU) scores of 0.47 for PD-L1, 0.45 for CD8+ T cells, and 0.46 for CD163+ macrophages. The presence of these biomarkers in high-attention regions supports their biological relevance to NACT response in TNBC. This not only improves model interpretability but may also inform future efforts to identify clinically actionable histological biomarkers directly from H&E-stained biopsy slides, further supporting the utility of this approach for accurate NACT response prediction and advancing precision oncology in TNBC. Full article

The present investigation was conducted to observe the effects of different energy densities of a low-level red laser (LLRL) on human embryonic stem cell-derived mesenchymal stem cells (hESC-MSCs). hESC-MSCs were cultured and irradiated with a LLRL from 0.5 to 5.0 J/cm² at a wavelength of 635 nm. Biological parameters such as proliferation, viability, and migration were observed after 72 h of LLRL irradiation. Compared with the control, LLRL irradiation significantly increased the proliferation and viability of hESC-MSCs from 0.5 to 2.5 J/cm² (p < 0.001, p < 0.05). LLRL irradiation from 0.5 to 3.0 J/cm² significantly increased the migration of hESC-MSCs (p < 0.01). These results revealed that LLRL irradiation at lower energy densities significantly increased the proliferation, viability, and migration of hESC-MSCs. However, higher energy densities were ineffective; this was also true when we examined osteogenic differentiation, as low energy densities of LLRL had a positive effect on differentiation, whereas higher energy densities had a negative effect on alkaline phosphatase activity, Alizarin Red staining and gene expression analysis. In addition, not all stem cell markers were affected by the laser, and a slight decrease in the expression of CD146, which is a stemness marker, was detected, indicating improved differentiation. These findings indicate that low energy densities of LLRL irradiation have positive effects on the proliferation, migration, and differentiation of hESC-MSCs. However, higher energy densities showed inhibitory effects. Full article

Platelet-rich plasma (PRP) has become an increasingly valuable biologic approach for personalized regenerative medicine because of its potent anti-inflammatory/healing effects. It is thought to be an excellent source of growth factors that can promote tissue healing and lessen fibrosis. Although this treatment has demonstrated effectiveness in numerous disease areas, its impact on pulmonary fibrosis (PF) caused by silver nanoparticles (AgNPs) via its antiapoptotic effects remains to be explored. AgNPs were synthesized biologically by Bacillus megaterium ATCC 55000. AgNP characterization was carried out via UV–Vis spectroscopy, X-ray diffraction (XRD), dynamic light scattering (DLS), transmission electron microscopy (TEM), and scanning electron microscopy (SEM) imaging to reveal monodispersed spheres with a mean diameter of 45.17 nm. A total of 48 male Wistar rats divided into six groups, with 8 rats per group, were used in the current study on the basis of sample size and power. The groups used were the PRP donor, control, AgNP, AgNP + PRP, AgNP + dexamethasone (Dexa) rat groups, and a recovery group. Body weights, hydroxyproline (HP) levels, and CASP3 and TWIST1 gene expression levels were assessed. H&E and Sirius Red staining were performed. Immunohistochemical studies for inducible nitric oxide synthase (iNOS) and cluster of differentiation 68 (CD68) with histomorphometry were conducted. A significant reduction in body weight (BWt) was noted in the AgNP group compared with the AgNP + PRP group (p < 0.001). HP, CASP3, and TWIST1 expression levels were significantly increased by AgNPs but decreased upon PRP (p < 0.001) treatment. Compared with those in the control group, the adverse effects of AgNPs included PF, lung alveolar collapse, thickening of the interalveolar septa, widespread lymphocytic infiltration, increased alveolar macrophage CD68 expression, and iNOS positivity in the cells lining the alveoli. This work revealed that PRP treatment markedly improved the histopathological and immunohistochemical findings observed in the AgNP group in a manner comparable to that of the Dexa. In conclusion, these results demonstrated the therapeutic potential of PRP in a PF rat model induced via AgNPs. This study revealed that PRP treatment significantly improved the histopathological and immunohistochemical alterations observed in the AgNP-induced group, with effects comparable to those of the Dexa. In conclusion, these findings highlight the therapeutic potential of PRP in a rat model of AgNP-induced PF. Full article

Bestrophinopathies are a group of inherited retinal diseases caused by mutations in the BEST1 gene. The protein encoded by this gene, bestorphin-1 (hBest1), is a calcium-dependent transmembrane channel localized on the basolateral membrane of retinal pigment epithelial (RPE) cells. We have already demonstrated the surface behavior and organization of recombinant hBest1 and its interactions with membrane lipids such as 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), sphingomyelin (SM) and cholesterol (Chol) in models of biological membranes, which affect the hBest1 structure–function relationship. The main aim of our current investigation is to integrate pure hBest1 protein into lipid bilayer nanostructures. We synthesized and characterized various hBest1-containing nanostructures based on 1,2-Dipalmitoylphosphatidylcholine (DPPC), SM, glycerol monooleate (GMO) and Chol in different ratios and determined their cytotoxicity and incorporation into cell membranes and/or cells by immunofluorescence staining. Our results show that these newly designed nanoparticles are not cytotoxic and that their incorporation into MDCK II cell membranes (used as a model system) may provide a mechanism that could be applied to RPE cells expressing mutated hBest1 in order to restore their ion transport functions, affected by mutated and malfunctioning hBest1 molecules. Full article

Plasma-derived fibrin hydrogels are widely used in tissue engineering because of their excellent biological properties. Specifically, human plasma-derived fibrin hydrogels serve as 3D matrices for autologous skin graft production, skeletal muscle repair, and bone regeneration. Nevertheless, for advanced applications such as in vitro skin equivalents and engineered grafts, the intrinsic limitations of native fibrin hydrogels in terms of long-term mechanical stability and resistance to degradation need to be addressed to enhance the usefulness and application of these hydrogels in tissue engineering. In this study, we chemically modified plasma-derived fibrin by incorporating succinimidyl alginate (SA), a version of alginate chemically modified to introduce reactive succinimidyl groups. These NHS ester groups (N-hydroxysuccinimide esters), attached to the alginate backbone, are highly reactive toward the primary amine groups present in plasma proteins such as fibrinogen. When mixed with plasma, the NHS groups covalently bond to the amine groups in fibrin, forming stable amide linkages that reinforce the fibrin network during hydrogel formation. This chemical modification improved mechanical properties, reduces contraction, and enhanced the stability of the resulting hydrogels. Hydrogels were prepared with a final fibrinogen concentration of 1.2 mg/mL and SA concentrations of 0.5, 1, 2, and 3 mg/mL. The objective was to evaluate whether this modification could create a more stable matrix suitable for supporting skin tissue development. The mechanical and microstructure properties of these new hydrogels were evaluated, as were their biocompatibility and potential to create 3D skin models in vitro. Dermo-epidermal skin cultures with primary human fibroblast and keratinocyte cells on these matrices showed improved dermal stability and better tissue structure, particularly SA concentrations of 0.5 and 1 mg/mL, as confirmed by H&E (Hematoxylin and Eosin) staining and immunostaining assays. Overall, these results suggest that SA-functionalized fibrin hydrogels are promising candidates for creating more stable in vitro skin models and engineered skin grafts, as well as for other types of engineered tissues, potentially. Full article

4-Aminodiphenylamine (4-ADPA) is a common additive in rubber tires, known for its antioxidant properties. It plays a crucial role in enhancing tire durability by preventing issues such as drying, cracking, and degradation from prolonged exposure to environmental factors like heat, oxygen, and ozone. However, despite its advantages in extending tire lifespan, the use of 4-ADPA raises significant environmental concerns. As tires wear down, microscopic tire wear particles (TWPs) containing 4-ADPA are released into the environment with substantial leaching, contaminating the waterways. The 4-ADPA leachates pollute and pose a threat to aquatic ecosystems, affecting various forms of marine life. The current study investigates the ecotoxicological effects of 4-ADPA on the aquatic plant Lemna minor (L. minor), focusing on its impact on relative growth and physiological biomarkers. Several parameters were assessed to evaluate ecotoxicity, including frond morphology, fresh biomass, total frond number, chlorophyll content, and starch accumulation. L. minor was grown for 7 and 14 days under controlled laboratory conditions using Hoagland media with varying concentrations of 4-ADPA (10–100 μg/L), while a control group was maintained in media without 4-ADPA. The results indicate that exposure to 4-ADPA led to a dose-dependent reduction in fresh biomass, total frond number, and chlorophyll levels. Lugol’s staining revealed increased starch accumulation in the fronds after exposure to 4-ADPA. The biological effects observed in L. minor following exposure to 4-ADPA, even at environmentally relevant concentrations, demonstrate a significant ecotoxicological impact on aquatic ecosystems. Further research involving additional species and investigating the mechanisms behind 4-ADPA toxicity is recommended to better understand its long-term consequences. Full article

Marine organism-associated microbes are an important source of structurally diverse and biologically active secondary metabolites exhibiting antimicrobial, anticancer, and anti-inflammatory activities. In this study, we investigated Penicillium brevicompactum MSW10-1, isolated from Hydractinia echinata, a marine invertebrate adapted to extreme intertidal and subtidal environments with variable temperature, salinity, and oxygen conditions. Through a combination of LC/MS-guided chemical analysis and chromatographic purification, eight secondary metabolites were isolated, including brevicolactones A (1) and B (2). The absolute chemical structures of 1 and 2 were determined based on NMR spectroscopic experiments, HR-ESIMS data, and quantum chemical ECD calculations. The isolated compounds (1–8) were evaluated for their ability to inhibit hepatic lipogenesis, a key process in lipid metabolism that is dysregulated in metabolic-dysfunction-associated steatotic liver disease. Furthermore, the inhibitory effects of the isolated compounds on lipid accumulation were further evaluated in primary mouse hepatocytes, using Oil Red O staining. These findings suggested that the isolated compounds may serve as promising candidates for the treatment of metabolic liver diseases associated with lipid dysregulation. Full article

Background/Objectives: High-fat-diet (HFD) consumption drives chronic liver injury via gut dysbiosis and metabolic disturban. Apple cider vinegar, rich in polyphenols and organic acids, shows potential in metabolic regulation. This study aimed to investigate whether apple cider vinegar powder (ACVP) alleviates HFD-induced liver injury by modulating the gut–liver axis. Methods: For 12 weeks, C57BL/6 J mice received daily ACVP gavage while being fed a HFD. A series of biological assessments were conducted, including systemic metabolic evaluations (body weight, serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST), and lipid/glucose levels), hepatic steatosis (hematoxylin and eosin (H&E) staining), intestinal microbiome characterization (16S rRNA gene genomic analysis), and comprehensive metabolite profiling of cecal contents (non-targeted metabolomics). Pearson correlation networks integrated multi-omics data. Results: ACVP attenuated HFD-induced weight gain by 26.3%, hepatomegaly and dyslipidemia, as well as reduced hepatic lipid vacuoles and serum ALT (48%)/AST (21.5%). ACVP restored gut microbiota diversity, enriching Muribaculaceae. Cecal metabolomics identified 38 HFD-perturbed metabolites reversed by ACVP, including indolelactate, hyocholate, and taurocholic acid. the Kyoto encyclopedia of genes and genomes (KEGG) analysis revealed ACVP-mediated recovery of linoleic acid metabolism. Correlation networks linked Akkermansia to anti-inflammatory metabolites (e.g., trans-ferulic), while Desulfobacterota correlated with pro-inflammatory oxylipins (e.g., 12,13-dihydroxy-9Z-octadecenoic acid (DHOME)). Conclusions: ACVP mitigates HFD-induced liver injury by remodeling gut microbiota, restoring microbial metabolites, and enhancing gut–liver crosstalk. Full article

Colorectal cancer remains a leading malignancy. As the aberrant activation of Wnt/β-catenin signaling causes colorectal cancer, Wnt/β-catenin signaling inhibitors are potential candidates for colorectal cancer treatment. Our drug screening platform identified ursolic acid (UA), a triterpenoid with various biological activities, as a potential anticancer drug because it inhibits the T-cell factor (TCF)/β-catenin-mediated transcriptional activity. Here, we discovered that UA inhibited Wnt signaling by reducing the Wnt reporter activity and Wnt target gene expression, leading to a delay in cell cycle progression and the suppression of cell proliferation. Stepwise epistatic analyses suggested that UA functions on β-catenin protein stability in Wnt signaling. Further studies revealed that UA reduced β-catenin protein levels by Western blotting and immunofluorescent staining and induced autophagy by microtubule-associated protein 1 light chain 3 beta (LC3B) punctate staining. The cotreatment with UA and the autophagy inhibitors chloroquine and wortmannin recovered the β-catenin protein levels. Therefore, UA was confirmed to induce β-catenin degradation by the autophagy–lysosomal degradation system through inhibition in the phosphatidylinositol 3-kinase (PI3K)/Ak strain transforming (protein kinase B; AKT)/mammalian target of rapamycin (mTOR) signaling pathway. Our results not only highlight the potential of UA in Wnt-driven colorectal cancer therapy but also provide a workable Wnt signaling termination approach for the treatment of other Wnt-related diseases. Full article

Trans fats are linked to numerous chronic diseases and cellular dysfunction; however, Spain has not implemented effective regulatory measures to restrict their presence in food products. This study addressed these gaps by analyzing trans fat content in commercial pastries sold in Spain and their biological impacts on mesenchymal stem cells, further examining its compliance with international guidelines. Also, a novel and scalable method for extracting fatty acids from pastry samples was developed and applied, enabling precise analysis using gas chromatography alongside sensory property assessments. The findings revealed significant variability in TFA levels across samples. To assess the biological implications of these TFAs, mesenchymal stem cells (MSCs) were cultured to perform dose–response experiments using two selected pastry samples with the highest TFA content. Cellular adhesion, cytotoxicity, and proliferation were evaluated through MTT assays, bright-field, and fluorescence staining using FITC and DAPI markers. Results demonstrated dose-dependent impacts of TFAs on MSC viability, including reduced adhesion and proliferation alongside increased cytotoxicity. This study underlines the need for stricter regulatory frameworks to monitor TFA levels worldwide, including in Spain’s food industry. Additionally, it highlights the potential health risks associated with excessive TFA consumption, particularly concerning cellular health and growth mechanisms, which provide insights into its potential bioaccumulation implications. These findings provide a foundation for further research into dietary guidelines and industrial practices aimed at minimizing TFA exposure while promoting public health safety. Full article