Search Results (71)

Osteoarthritis (OA) is a progressive joint disorder affecting over 250 million people globally and is characterized by chronic pain and disability. Among its key pathogenic mechanisms are mitochondrial dysfunction and elevated reactive oxygen species (ROS), often triggered by inflammatory mediators such as lipopolysaccharide (LPS). This study evaluates the protective effects of curcumin on mitochondrial function, autophagy, and apoptosis in an in vitro model of OA. Human bone marrow-derived mesenchymal stem cells (BMSCs) were differentiated into chondrocytes using MesenCult™-ACF medium. Differentiation was confirmed by histological staining for Type II Collagen, Alcian Blue, and Toluidine Blue. LPS was used to induce an OA-like inflammatory response. Mitochondrial membrane potential (ΔΨm) was assessed using Rhodamine 123 staining. Autophagy and apoptosis were evaluated using Acridine orange and propidium iodide staining, respectively. Western blotting was performed to analyze the expression of pro-caspase-3, Bcl-2, Beclin-1, LC3-I/II, and GAPDH. LPS significantly impaired mitochondrial function, limited autophagy, and enhanced apoptotic signaling (reduced pro-caspase-3). Curcumin (25 µM and 100 µM) restored ΔΨm, increased Beclin-1 and LC3-II, and maintained pro-caspase-3 expression, with Bcl-2 showing a non-monotonic response (higher at 25 µM than at 100 µM). Curcumin exerted cytoprotective effects in inflamed chondrocytes by stabilizing ΔΨm, promoting autophagy, and attenuating apoptotic activation, supporting its multi-target therapeutic potential in OA. Full article

Corilagin is a hydrolyzable ellagitannin and naturally occurring polyphenolic compound widely distributed in medicinal plants. It is also present in longan (Dimocarpus longan), known as lumyai in Thailand, a subtropical fruit extensively cultivated across China and Southeast Asia. Corilagin has been reported to exhibit strong antioxidant, anti-inflammatory, hepatoprotective, and anticancer activities through modulation of multiple cellular signaling pathways. However, despite these well-established pharmacological properties, its potential role in regulating bone marrow mesenchymal stem cell (BM-MSC) differentiation has not been fully explored in biomedical applications. In this study, we investigated the effects of corilagin on BM-MSC viability, protein-binding interactions, and lineage-specific differentiation toward osteogenic and chondrogenic pathways. Cytotoxicity assessment using human synovial SW-982 cells demonstrated that corilagin maintained cell viability at concentrations ranging from 1.56 to 50 µg/mL within 48 h, whereas prolonged exposure resulted in a time-dependent reduction in viability. In BM-MSCs, corilagin significantly enhanced osteogenic and chondrogenic differentiation in a dose-dependent manner, as evidenced by increased mineral deposition and cartilage matrix formation, as revealed by Alizarin Red S, Toluidine Blue, and Alcian Blue staining. Quantitative analyses further showed the upregulation of key lineage-specific genes, including Runx2 and osteopontin (OPN) for osteogenesis and Sox9 and aggrecan for chondrogenesis. Protein-binding assays confirmed the molecular interaction capacity of corilagin, supporting its biological activity. Overall, these findings demonstrate that corilagin promotes MSC-mediated osteogenic and chondrogenic differentiation while maintaining acceptable cytocompatibility, highlighting its potential as a natural small-molecule candidate for bone and cartilage tissue engineering and other biomedical fields with regenerative medicine applications. Full article

(1) Background: Intra-articular injection is a fundamental technique in preclinical research for evaluating therapeutics and inducing joint disease models in rodents. However, the absence of standardized and validated protocols compromises reproducibility and translational validity. (2) Methods: This study establishes and experimentally validates a refined protocol for precise intra-articular injection in the knee of adult male Wistar rats. The comprehensive procedure specifies anatomical landmarks (medial border of the patellar tendon), instrumentation (27 G needle, 100 µL Hamilton syringe), a maximum volume of 35 µL, and operative verification criteria based on tactile feedback. Experimental validation was performed by administering a suspension of wear particles (2.35 mg/mL) generated from tribocorrosion tests of CoCr surfaces biofunctionalized with graphene oxide-hyaluronic acid (GO-HA) into the left knee of five rats. (3) Results: Histological analysis using the cutting–grinding technique and Toluidine Blue staining confirmed the exclusive intra-articular localization of particles in all injected animals (5/5 success rate). Qualitative assessment revealed abundant particulate distribution within the synovial space, with numerous individual particles and multiple aggregates observed per high-power field, without evidence of extravasation in any case. (4) Conclusions: The protocol demonstrated high intra-operator repeatability and provides a reliable, ethically refined tool for precise intra-articular administration of nanomaterials and for generating robust joint disease models, thereby enhancing reproducibility and animal welfare in preclinical research. Full article

Background: Phosphaturic mesenchymal tumours secreting fibroblast growth factor 23 (hereinafter referred to as FGF23+ PMT) are rare neoplasms that can cause hypophosphataemic osteomalacia, owing to excessive FGF23 production. Mast cells (MCs) play a key role in tumour biology by modulating proliferative activity of atypical cells, resistance to innate and acquired immunity, angiogenesis, and metastatic behaviour. However, MCs associated with FGF23+ PMT have not previously been investigated. This study, to our knowledge, is the first to characterise features of the tumour microenvironment through spatial phenotyping of the immune and stromal landscape, together with histotopographic mapping of intercellular MC interactions with other subcellular populations in FGF23+ PMT. Methods: Histochemical staining (haematoxylin and eosin, toluidine blue, Giemsa solution, picro-Mallory protocol, silver impregnation), as well as monoplex and multiplex immunohistochemical staining with spatial phenotyping, were performed to detect atypical FGF23-secreting cells, immune cells (CD3, CD4, CD8, CD14, CD20, CD38, CD68, or CD163), stromal components (CD31, α-SMA, or vimentin), and specific MC proteases (tryptase, chymase, or carboxypeptidase A3). Bioinformatics analysis using artificial intelligence technologies was applied for spatial profiling of MC interactions with tumour, immunocompetent, and stromal cells in the tumour microenvironment. Results: Bioinformatic analysis of the entire tumour histological section, comprising over 70,000 cells stained using monoplex and multiplex immunohistochemical protocols, enabled identification of more than half of the cell population. The most abundant were CD14+ (30.7%), CD163+ (23.2%), and CD31+ (17.9%) cells. Tumour-associated MCs accounted for 0.7% of the total pool of immunopositive cells and included both mucosal and connective tissue subpopulations, predominantly of the tryptase + chymase-CPA3-specific protease phenotype. This pattern reflected combined multidirectional morphogenetic processes in the patient’s FGF23+ PMT. More than 50% of MCs were colocalized with neighbouring cells of the tumour microenvironment within 20 μm, most frequently with monocytes (CD14+CD68+), M2 macrophages (CD68+CD163+), and endothelial cells (CD31+). In contrast, colocalization with atypical FGF23-secreting cells was rare, indicating minimal direct effects on tumour cell activity. Interaction with T lymphocytes, including CD8+, was also infrequent, excluding their activation and the development of antitumour effects. Mapping of MC histotopography validated the hypothesis of their inductive role in monocyte differentiation into M2 macrophages and probable polarisation of macrophages from M1 into M2, thereby contributing to slow tumour growth. MCs were further involved in extracellular matrix remodelling and participated in the formation of pro-osteogenic niches within the FGF23+ PMT microenvironment, leading to pathological osteoid development. Conclusions: This study demonstrated active MC participation in the evolution of the FGF23+ PMT microenvironment. The findings may be applied in translational medicine to develop novel algorithms for personalised therapy in patients with FGF23-secreting tumours, offering an alternative when surgical removal of the tumour is not feasible. Full article

Yeasts of the genus Candida (C.) and the bacterium Staphylococcus aureus (S. aureus) are among the most common pathogens responsible for infections that are difficult to treat, including those resistant to standard therapy. In recent decades, this has become an increasing clinical problem. In response to the limitations of traditional procedures, antimicrobial photodynamic therapy (aPDT), which combines light, a photosensitizer, and oxygen, is gaining growing interest. The aim of this study was to evaluate the in vitro effectiveness of aPDT using a 635 nm diode laser in combination with toluidine blue O (TBO) against Candida spp. and S. aureus. Reference strains of C. albicans, C. glabrata, C. krusei, and S. aureus were subjected to aPDT. In phase I of this study, the optimal TBO incubation time was assessed with constant laser parameters. In phase II, the impact of the physical parameters of the laser, irradiation time, and output power, was analyzed, with the TBO incubation time set based on the phase I results, to evaluate the degree of microbial reduction (CFU/mL). Statistical analyses were then conducted to assess significance. TBO-mediated aPDT significantly reduced microbial viability, depending on incubation time and laser settings. The minimal effective incubation times were 10 min for Candida spp. and 5 min for S. aureus. The highest pathogen inactivation efficacy was observed at an output power of 400 mW and an irradiation time of 120 s. The use of the photosensitizer or laser alone did not result in significant antimicrobial effects. TBO-mediated aPDT may serve as an effective complement to conventional antimicrobial therapy and, in selected cases (e.g., drug resistance), has the potential to partially or fully replace it. The observed minimal effective incubation times provide a practical baseline, but further statistical comparisons are required to determine whether these durations are truly optimal. Full article

This study explores the visible-light-driven photolysis of Ferrioxalate complexes for the degradation of Toluidine Blue (TB), a persistent phenothiazine dye, using a 1 L recirculating batch-loop photoreactor. The reactor system incorporated two tubular photochemical units (35 cm × 3 cm each) in series: the first equipped with an immersed blue fluorescent lamp (12 W, 30 cm-tube), and the second with dual external blue LED lamps (18 W total, 30 cm) encasing a double-walled glass cell. Continuous flow between the units was maintained via a peristaltic pump. Experimental investigations were used to evaluate the effects of key parameters such as Fe(III) and oxalate concentrations, initial TB load, pH, light source, flow rate, ligand type, dissolved gas type, external H₂O₂ addition, and the presence of various inorganic ions. The results demonstrate efficient dye degradation, with ~75% TB removal within 1 h under combined fluorescent and LED irradiation, where each reactor contributing comparably. The optimal performance was achieved at pH 4, with a 10 oxalate-to-Fe(III) molar ratio (1 mM:0.1 mM) and a flow rate of 25 mL s⁻¹. Among various ligands tested (oxalate, acetate, citrate, EDTA), oxalate proved to be the most effective. The presence and type of anions significantly influenced degradation efficiency due to their potential scavenging effects. Although the process achieved high dye removal, TOC analysis indicated only moderate mineralization, suggesting the accumulation of non-colored intermediates. External H₂O₂ addition moderately improved TOC removal, likely due to enhanced hydroxyl radical generation via the Fenton mechanism. These findings highlight the promise of Ferrioxalate-based photochemical systems under visible light for dye removal, while also emphasizing the need for further research into by-product identification, mineralization enhancement, and toxicity reduction to ensure safe effluent discharge. Full article

Background/Objectives: Pompia is an ancient, endemic citrus ecotype native to Sardinia (Italy), characterized by distinctive morphology and high content of bioactive compounds. Despite increasing interest, several aspects of this fruit, including its histological characteristics, remain poorly understood. This study aims to address this gap by investigating the anatomical features and spatial distribution of secretory cavities involved in essential oil (EO) production and accumulation, while also evaluating the EO’s chemical profile and associated biological activity. Methods: Pompia peel (flavedo and albedo) was subjected to histological analysis through fixation, dehydration, resin inclusion and sectioning. Sections were stained with 0.05% toluidine blue and observed under a light microscope to measure different parameters of secretory cavities. Essential oil (EO) was obtained from Pompia peel by hydrodistillation and characterized by gas chromatography–mass spectrometry (GC–MS) analysis. The biological activity of Pompia EO was assessed in vitro using NIH/3T3 fibroblasts, where wound-healing was evaluated by scratch assay and anti-senescence effects by β-galactosidase and γH2AX activity. Results: Microscopic analysis of the peel revealed pronounced variability in depth and size of the secretory cavities, along with the presence of lenticel-like structures in the epidermis. GC–MS analysis showed that Pompia EO is dominated by limonene (89%), with minor compounds including myrcene, geranial and neral. In vitro biological assays demonstrated that the EO promotes cell migration in a wound-healing model at concentrations ≥ 12.5 µg/mL and reduces markers of cellular senescence, including β-galactosidase activity and γH2AX foci, in etoposide-induced senescent fibroblasts. Conclusions: Overall, this study provides the first histological characterization of Pompia peel and confirms the bioactive potential of its EO. These findings support future applications in skin regeneration and anti-aging strategies and contribute to the valorization of this underexplored Citrus ecotype. Full article

This study focuses on a typical chemically contaminated site in the southeastern coastal region of China, investigating the natural attenuation mechanisms of benzene and o-toluidine in groundwater through high-throughput sequencing, physicochemical analyses, and stable isotope techniques. The results demonstrate significant heterogeneity in the spatial distribution and degradation processes of pollutants within the contaminated zones (W27, W28, W31). Environmental factors such as HCO₃⁻, SO₄²⁻, and ORP predominantly influence the microbial community structure and functional distribution. Stable isotope data reveal that δD and δ¹³C enrichment effects are most pronounced in the deep layer (W28_40m), indicating active pollutant degradation, while degradation in the deeper layers of W27 and W31 is constrained by anaerobic conditions and reduced microbial activity. The combined analysis of hydrogen and carbon isotopes elucidates the degradation pathways and dynamic processes of pollutants within the contaminated zones, providing quantitative evidence for natural attenuation mechanisms and scientific support for optimizing site remediation strategies. Full article

Detecting multiple tumor markers is of great importance. It helps in early cancer detection, accurate diagnosis, and monitoring treatment. In this work, gold nanoparticles–toluidine blue–graphene oxide (AuNPs-TB–GO) and gold nanoparticles–carboxyl ferrocene–tungsten disulfide (AuNPs–FMC–WS₂) nanocomposites were prepared for labeling Carcinoembryonic antigen (CEA) antibody and Carbohydrate antigen 72–4 (CA72-4) antibody, respectively, and used as two kinds of probes with different electrochemical signals. With the excellent magnetic performance of biotin immune magnetic beads (IMBs), the biofunctional IMBs were firmly deposited on the magnetic glassy carbon electrode (MGCE) surface by applying a constant magnetic field, and then the CEA and CA72-4 antibody were immobilized on the IMBs by the avidin–biotin conjugation. The assay was based on the change in the detection peak current. Under the optimum experimental conditions, the linear range of detection of CEA is of the two-component immunosensor is from 0.01 to 120 ng/mL, with a low detection limit of 0.003 ng/mL, and the linear range of detection of CA72-4 is from 0.05 to 35 U/mL, with a detection limit of 0.016 U/mL. The results showed that the proposed immunosensor enabled simultaneous monitoring of CEA and CA72-4 and exhibited good reproducibility, excellent high selectivity, and sensitivity. In particular, the proposed multiplexed immunoassay approach does not require sophisticated fabrication and is well-suited for high-throughput biosensing and application to other areas. Full article

In this work, three methods for the synthesis of composites based on poly(ortho-toluidine) (POT) and WS₂ are reported: (a) the solid-state interaction (SSI) of POT with WS₂ nanoparticles (NPs); (b) the in situ chemical polymerization (ICP) of ortho-toluidine (OT); and (c) the electrochemical polymerization (ECP) of OT. The preparation of WS₂ sheets was performed by the ball milling of the WS₂ NPs followed by ultrasonication in the solvent N,N’-dimethyl formamide. During the synthesis of the POT/WS₂ composites by SSI and ICP, an additional exfoliation of the WS₂ NPs was reported. In this work, we demonstrated the following: (a) the ICP method leads to POT/WS₂ composites, which contain repeating units of POT in the leucoemeraldine salt (LS) state, while (b) the ECP method leads to POT/WS₂ composites, which contain repeating units of POT in the emeraldine salt (ES) state. Capacitances equal to 123.5, 465.76, and 751.6 mF cm⁻² in the cases of POT-ES/WS₂ composites, synthesized by SSI, ICP, and ECP, respectively, were reported. Full article

Background: Osteoarthritis (OA) is a chronic condition characterized by joint pain and disability, driven by excessive oxidative stress and inflammatory cytokine production in chondrocytes, resulting in cell death and cartilage matrix breakdown. Our previous study showed that in monosodium iodoacetate (MIA)-induced OA rats, oral administration of heat-killed Lactobacillus delbrueckii subsp. lactis 557 (LDL557) could significantly decrease OA progression. Methods: Accordingly, we designed an in vitro cell culture study aimed at investigating the effects of heat-killed LDL557 extracts on chondrocytes using SW1353 cells (a human chondrosarcoma cell line) challenged with 5 μM MIA to mimic OA conditions. Results: The results showed that the 10 μg/mL LDL557 extracts protected SW1353 cells from MIA-induced death and reduced extracellular matrix (ECM) loss, as evaluated by toluidine blue O staining and extracellular matrix component synthesis with RT-qPCR measurement. This was achieved by decreasing the expression of MIA-induced pro-inflammatory cytokines, including IL-1β, IL-6, and TNF-α, while slightly increasing the MIA-suppressed expression of the anti-inflammatory cytokine IL-10, which were evidenced by RT-qPCR analysis. Moreover, the RT-qPCR evaluation also indicated that the LDL557 extracts slightly reduced the expression of COX-2 compared with the control, while it did not reduce the MIA-increased expression of microsomal prostaglandin E synthase-1 (mPGES-1). In addition, the LDL557 extracts influenced neither the matrix-degrading protease expressions measured via RT-qPCR nor the oxidative stress measured via fluorescence flow cytometry in the cells with or without the MIA challenge. Conclusions: This study demonstrates that LDL557 extracts may protect chondrocytes from OA damage by reducing inflammation-related factors and thus mitigating cartilage matrix loss, suggesting LDL557 extracts are attractive alternatives for OA applications. Full article

Within the area of study of neurodegenerative diseases, particularly Alzheimer’s disease (AD), this research focused on the synthesis and evaluation of novel tetrahydroquinoline (THQ) derivatives with potential antioxidant activity. The toluidine N-propargylation synthesis protocol was optimized, achieving a significant increase in yield by using sodium carbonate and reaction temperature variation. Subsequently, four THQ compounds with alkene variation were successfully synthesized, including some which had not been previously reported in the literature. The synthesized compounds were characterized by nuclear magnetic resonance (NMR), mass spectrometry, and infrared spectroscopy (IR), which confirmed their structures and purity. In silico analyses performed with SwissADME and OSIRIS Property Explorer indicated that most of the compounds exhibited excellent drug-like characteristics and favorable pharmacokinetic profiles. Antioxidant evaluation was performed using DPPH and ABTS assays, in which all compounds demonstrated excellent antioxidant capacity, with EC50 values below 10 μg/mL in the ABTS assay, significantly outperforming the ascorbic acid control (EC50 = 35 μg/mL). The results suggest that the predominant radical-scavenging mechanism is single electron transfer (SET). This study provides a solid foundation for further investigations into the potential of THQs’ derivatives as antioxidants, and potential cholinesterase inhibitors in the context of neurodegenerative diseases such as Alzheimer’s. As a future projection, an enzymatic evaluation, including regarding mechanisms of action and the exploration of a hybrid synthesis of THQ/triazole, is proposed based on these promising results. Full article

Objectives: Vitamin B complexes are frequently used in clinical practice for peripheral nerve trauma. However, there is a lack of scientific data on their effectiveness. This study aims to investigate the impact of the vitamin B complex on nerve recovery in a rat model of peripheral nerve paralysis. Materials and Methods: Sixty male Wistar Albino rats were divided into six groups. Models of nerve injury, including blunt trauma, nerve incision, and autograft, were performed on all rats approximately 1 cm distal to the sciatic notch. B-complex vitamins were injected intraperitoneally at 0.2 mL/day to the treatment groups. The control groups were given 0.2 mL/day saline. After 1 month, the study was terminated, electromyography (EMG) was performed to measure the conduction velocity, and nerve tissue was taken from the repair line. The sciatic function indexes (SFIs) were calculated and analyzed. The histopathological samples were stained with hematoxylin and eosin and Toluidine blue and examined with a light microscope. Pathologically, myelination, fibrosis, edema, and mast cell densities in the nervous tissue were evaluated. Results: The vitamin B treatment groups demonstrated significant improvements in SFI compared to the control groups, indicating functional improvement in nerve damage (p < 0.05). In the nerve graft group, the vitamin B group showed a shorter latency, higher velocity, and larger peak-to-peak compared to the controls (p < 0.05). In the nerve transection group, the vitamin B group had better latency, velocity, and peak-to-peak values than the controls (p < 0.05). In the crush injury group, the vitamin B group exhibited an improved latency, velocity, and peak-to-peak compared to the controls (p < 0.05). Better myelination, less fibrosis, edema, and mast cells were also in the vitamin B group (p < 0.05). Conclusions: Vitamin B treatment significantly improves nerve healing and function in peripheral nerve injuries. It enhances nerve conduction, reduces fibrosis, and promotes myelination, indicating its therapeutic potential in nerve regeneration. Full article

As it has been revealed that the activation of human immune cells through the activity of intestinal microorganisms such as pro- and prebiotics plays a vital role, controlling the proliferation of beneficial bacteria and suppressing harmful bacteria in the intestine has become essential. The importance of probiotics, especially for skin health and the immune system, has led to the emergence of products in various forms, including probiotics, prebiotics, and parabiotics. In particular, atopic dermatitis (AD) produces hypersensitive immunosuppressive substances by promoting the differentiation and activity of immune regulatory T cells. As a result, it has been in the Th1 and Th2 immune balance through a mechanism that suppresses skin inflammation or allergic immune responses caused by bacteria. Furthermore, an immune mechanism has recently emerged that simultaneously controls the expression of IL-17 produced by Th17. Therefore, the anti-atopic effect was investigated by administering doses of anti-atopic candidate substances (Lactobacilus sakei CVL-001, Lactobacilus casei MCL, and Lactobacilus sakei CVL-001 Lactobacilus casei MCL mixed at a ratio of 4:3) in an atopy model using 2,4-dinitrochlorobenzene and observing symptom changes for 2 weeks to confirm the effect of pro-, para-, and mixed biotics on AD. First, the body weight and feed intake of the experimental animals were investigated, and total IgG and IgM were confirmed through blood biochemical tests. Afterward, histopathological staining was performed using H&E staining, Toluidine blue staining, Filaggrin staining, and CD8 antibody staining. In the treatment group, the hyperproliferation of the epidermal layer, the inflammatory cell infiltration of the dermal layer, the expression of CD8, the expression of filaggrin, and the secretion of mast cells were confirmed to be significantly reduced. Lastly, small intestine villi were observed through a scanning microscope, and scoring evaluation was performed through skin damage. Through these results, it was confirmed that AD was reduced when treated with pro-, para-, and mixed biotics containing probiotics and parabiotics. Full article

5-fluorouracil (5-FU), a chemotherapeutic agent against oral squamous cell carcinoma (OSCC), is limited by poor pharmacokinetics and toxicity. The pH-sensitive zeolite imidazolate framework-8 (ZIF-8) may increase the selectivity and length of 5-FU released into the acidic tumor microenvironment. This study examined the in vitro 5-FU absorption and release profiles of ZIF-8, and then progressed to cytotoxicity assays using the OSCC primary cell line SCC7. The 5-FU loading capacity of ZIF-8 was calculated with UV-vis spectroscopy (λ = 260 nm). 5-FU release was quantified by submerging 5-FU@ZIF-8 in pH 7.4 and 5.5 acetate buffer over 48 h. For the cytotoxicity assays, 5-FU, ZIF-8, and 5-FU@ZIF-8 were added to SCC7 cultures at 25, 50, and 100 μg/mL. Cell viability was assessed through toluidine blue staining and further quantified through transcriptomic RNA sequencing. ZIF-8 stabilized at a maximum absorption of 2.71 ± 0.22 mg 5-FU, and released 0.66 mg more 5-FU at pH 5.5 than 7.4 for at least 72 h. The cytotoxicity assays showed that 5-FU@ZIF-8 had a synergistic inhibitory effect at 50 μg/mL. The RNA sequencing analysis further revealed the molecular targets of 5-FU@ZIF-8 in SCC7. 5-FU@ZIF-8 may release 5-FU based on the pH of the surrounding microenvironments and synergistically inhibit OSCC. Full article