Search Results (74)

Background/Objectives: The accumulation of Nε-carboxymethyllysine (CML), a major advanced glycation end product (AGE), has been implicated in neuronal dysfunction by promoting oxidative stress, endoplasmic reticulum (ER) stress, and dysregulation of amyloid-β (Aβ) metabolism. This study evaluated the neuroprotective properties of coixol, a naturally occurring polyphenolic compound derived from the outer layers of Coix lacryma-jobi L. var. ma-yuen, in a CML-induced injury model using IMR-32 human neuronal-like cells. Methods: Cells were pretreated with coixol (1 μmol/L), N-acetyl-L-cysteine (NALC, 1 mmol/L), or 4-phenylbutyric acid (4-PBA, 200 μmol/L) for 1 h prior to CML (100 μmol/L) exposure for 24 h. Cell viability was determined by colorimetric analysis of 3-(4,5-dimethyl-2-yl)-2,5-diphenyltetrazolium bromide, while intracellular reactive oxygen species (ROS) generation was quantified using a fluorescence-based oxidative stress probe. Activities of key antioxidant enzymes and caspase-3 were determined using commercial assay kits. The expression of Aβ isoforms, amyloidogenic enzymes, ER stress markers, and apoptosis-related signaling proteins was quantified through validated immunoassays. Results: Coixol pretreatment significantly enhanced cell viability by attenuating ROS accumulation and restoring antioxidant enzyme activities. Concurrently, coixol suppressed ER stress signaling via downregulation of the protein kinase R-like ER kinase/C/EBP homologous protein axis and modulated apoptosis by increasing B-cell lymphoma (Bcl)-2, reducing Bcl-2-associated X protein expression, and inhibiting caspase-3 activation and DNA fragmentation. Furthermore, coixol regulated Aβ metabolism by inhibiting the expression of β-site amyloid precursor protein-cleaving enzyme 1 and presenilin 1, while restoring insulin-degrading enzyme and neprilysin levels, leading to reduced accumulation of Aβ40 and Aβ42. Conclusions: Compared to NALC and 4-PBA, coixol demonstrated comparable or superior modulation across multiple pathological pathways. These findings highlight coixol’s potential as a neuroprotective candidate in AGE-associated neurodegenerative conditions. Full article

This study introduces a novel nucleic acid testing (NAT) protocol that integrates rapid deoxyribonucleic acid (DNA) extraction, isothermal amplification, and visual detection to enable efficient analysis of opportunistic pathogens. Polyethylenimine-functionalized iron oxide (PEI-Fe₃O₄) nanoparticles were prepared by combining PEI, acting as a stabilizing agent, with iron salt, which was utilized as the metal ion precursor by the ultrasonication-assisted co-precipitation method, and characterized for structural, optical, and magnetic properties. PEI-Fe₃O₄ exhibited cationic and anionic behavior in response to pH variations, enhancing adaptability for DNA binding and release. PEI-Fe₃O₄ enabled efficient extraction of E. faecium DNA within 10 min at 40 °C, yielding 17.4 ng/µL and achieving an extraction efficiency of ~59% compared to a commercial kit (29.5 ng/µL). The extracted DNA was efficiently amplified by loop-mediated isothermal amplification (LAMP) at 65 °C for 45 min. Pyrogallol-rich poly(tannic acid)-stabilized gold nanoparticles (PTA-AuNPs) served as colorimetric probes for direct visual detection of the DNA amplified using LAMP. The magnetic-nanogold (PEI-Fe₃O₄/PTA-AuNPs) hybrid system achieved a limit of quantification of 1 fg/µL. To facilitate field deployment, smartphone-based RGB analysis enabled quantitative and equipment-free readouts. Overall, the PEI-Fe₃O₄/PTA-AuNPs hybrid system used in NAT offers a rapid, cost-effective, and portable solution for DNA detection, making the system suitable for microbial monitoring. Full article

Reconstruction of extensive bone defects due to age, trauma, or post-illness conditions remains challenging. Biomimetic scaffolds with osteogenic capabilities have been proposed as an alternative to the classical autograft and allograft implants. Three-dimensional scaffolds were obtained based on Ce-doped mesoporous bioactive glass (MBG) and Spongia agaricina (SA) as sacrificial templates functionalized with vitamin D₃. The study aimed to investigate the effect of vitamin D₃ functionalization on the optimal variant of a 3D scaffold doped with 3 mol% ceria, selected in our previous work based on its biological and physicochemical properties. Scanning electron microscopy (SEM) images of the non-functionalized/functionalized scaffolds revealed a porous structure with interconnected pores ranging from 100 to 350 μm. Fourier transform infrared spectroscopy (FTIR) and SEM analysis confirmed the surface functionalization. Cytotoxicity evaluation showed that all investigated scaffolds do not exhibit cytotoxicity and genotoxicity toward the Saos-2 osteosarcoma cell line. Moreover, the study demonstrated that functionalization with vitamin D₃ enhanced osteogenic activity in dental pulp stem cells (DPSCs) by increasing calcium deposition and osteocalcin secretion, as determined by Alizarin red stain and a colorimetric ELISA kit, as a result of its synergistic action with cerium ions. The results showed that the Ce-doped MBG scaffold functionalized with vitamin D₃ had the potential for applications in bone regeneration. Full article

Lipopolysaccharide (LPS)-induced inflammation impairs sperm function; however, its impact on ejaculated rabbit sperm remains unexplored. This dose-response study aims to determine the LPS concentration that negatively affects sperm motility in vitro, while also providing the first identification of TLR4 localization on rabbit spermatozoa. Additionally, it evaluates malondialdehyde (MDA) levels in seminal plasma as an indicator of oxidative stress. Sperm motility was analyzed using computer-assisted sperm analysis (CASA) after incubation with increasing LPS concentrations (0, 50, 100, 200, 400, 600, and 800 µg/mL) at multiple time points (0, 1, 2, and 4 h). LPS doses ≥ 400 µg/mL significantly reduced progressive and non-progressive motility, as well as curvilinear velocity (all p < 0.001), while increasing the proportion of static spermatozoa (p < 0.05). Receiver operating characteristic (ROC) analysis identified 300 µg/mL as the threshold dose for motility decline. Immunofluorescence revealed TLR4 localization in the midpiece of sperm tails, with weak labeling in control samples and a marked increase after 4 h of incubation with 400 μg/mL LPS. MDA levels were assessed using the thiobarbituric acid reactive substances (TBARS) assay with a colorimetric kit, showing no significant effect of LPS treatment. No correlation was found between MDA and other semen parameters. ccThese findings identify TLR4 on rabbit sperm for the first time and establish a threshold LPS dose for future in vitro studies. Full article

Background/Objectives: The present study aimed to test the efficacy of the chair-side rapid salivary C-reactive protein assay kit in differentiating oral potentially malignant disorders (OPMDs) and oral cancer from normal mucosa using whole salivary samples. Methods: In this study, unstimulated saliva samples of cases (OPMDs and oral cancer) and controls (systemically healthy subjects) were used to detect CRP levels using a novel colorimetric, quantitative rapid assay kit. Kruskal–Wallis ANOVA with a post hoc Dunn’s test were applied to determine the difference in the mean and SD values between the case and control groups. ROC analysis was performed to identify the positive and negative likelihood ratios. Results: The mean calculated salivary CRP level in the oral cancer group was 4.21 ng/mL, in the OPMD group it was 2.51 ng/mL and in the healthy controls it was 0.7 ng/mL. Post hoc tests showed that the salivary CRP levels were significantly higher in the oral cancer and OPMD groups than in the controls. Conclusions: The present study showed that the novel rapid salivary test kit could significantly differentiate between the salivary CRP values of cases and controls but there was no significant difference between the OPMD and malignancy groups. It also showed higher sensitivity values, confirming the efficacy of this kit as a screening tool. Full article

Chronic Kidney Disease (CKD) is a disorder that affects over 10% of the global population, and that would benefit from innovative methodologies that would provide early detection. Since it has been reported that there are high levels of urease in CKD patients’ saliva, this sample is a promising non-invasive alternative to blood for CKD detection and monitoring. This work introduces a novel 3D µPAD for quantifying urease activity in saliva in a range of 0.041–0.750 U/mL, with limits of detection and quantification of 0.012 and 0.041 U/mL, respectively. The device uses the urease in the sample to convert urea into ammonia, causing a colorimetric change in the bromothymol blue. The accuracy of the developed device was evaluated by comparing the measurements of several saliva samples (#13) obtained with the μPAD and with a commercially available kit. Stability studies were also performed to assess its functionality as a point-of-care methodology, and the device was stable for 4 months when stored in a vacuum. After the sample placement, it could be scanned within 40 min without providing significantly different results. The developed device quantifies urease activity in saliva within 30 min, providing a simple, portable, lab-free alternative to existing methodologies. Full article

Cold atmospheric plasma (CAP) has antimicrobial properties and is also known to stimulate the immune system. These properties could be useful for the development of a novel therapeutic or preventive strategy against respiratory infections in the upper respiratory tract (URT) such as ventilator-associated pneumonia (VAP) without inducing an immune overreaction. This study investigated the cellular responses of polymorphonuclear neutrophils (PMNs) after exposure to CAP in a three-dimensional (3D) model of the URT. In vitro experiments were conducted using PMNs isolated from human blood to assess cell migration, intracellular production of reactive oxygen species (ROS), NETosis, surface marker expression (CD11b, CD62L, and CD66b), and cell death with live cell imaging and flow cytometry. CAP was applied for 5 min using two distinct modalities: pressurized air plasma with a plasma intensive care (PIC) device and nebulized air plasma (NP) with a new humidity resistent surface microdischarge (SMD) plasma source, both developed by Terraplasma Medical GmbH. There were no significant signs of cell damage or overstimulation with either device under the conditions tested. However, the NP device caused milder effects on PMN functionality compared to the PIC device, but also demonstrated reduced antibacterial efficacy and reactive oxygen/nitrogen species (RONS) production, as analyzed with colorimetric/fluorimetric assay kits. These findings highlight a trade-off between the two CAP modalities, each with distinct advantages and limitations. Further studies are necessary to investigate these effects in the clinical setting and evaluate the long-term safety and efficacy of CAP treatment in the URT. Full article

Wheat embryo albumin (WEA), rich in amino acids with a good balanced proportion, demonstrates plentiful biological activities. The effects of WEA and its peptide with the best antioxidant ability (F3) as a post-workout and pre-workout energy supplement on alleviating acute exercise fatigue were investigated. Under two experimental cases, the exhaustion-to-death swimming time and exhaustion swimming time were determined. Fatigue-related biochemical indexes including lactate dehydrogenase (LDH), the level of blood urea nitrogen (BUN), alanine transaminase (ALT), aspartate transaminase (AST), liver glycogen (LG), and muscle glycogen (MG) were measured with commercial kits. Antioxidant capacity in vivo was analyzed by determining the content of malondialdehyde (MDA), the level of glutathione (GSH), and the activity of superoxide dismutase (SOD) based on colorimetric methods. The results indicated that administration of WEA and F3 post-workout or pre-workout significantly prolonged exhaustive swimming time (p < 0.05) and increased the levels of glycogen in the liver and muscle of mice (p < 0.05). Meanwhile, WEA and F3 significantly reduced the activities of ALT, AST, and LDH and the level of BUN compared with the ones of mice in an exercise fatigue model (p < 0.05). Additionally, in comparison with the model group, supplements of WEA and F3 obviously decreased the content of MDA while enhancing the activity of SOD and the level of GSH both in the liver and muscle of mice. These results demonstrated that WEA and F3 can mitigate exercise fatigue and are conducive to recovery from fatigue in exhausted mice. It suggests that WEA and its peptide F3 could be a promising energy supplementary material against fatigue caused by continuous or high-intensity exercise. Full article

Hyperlipidemia is the root cause of numerous chronic conditions, leading to high mortality rates around the globe. Spirulina (Arthrospira platensis) microalgae serve as a promising reservoir of bioactive compounds with diverse pharmacological properties. The current study examined the nutritional profile of spirulina powder in relation to strict glycemic control, specifically focusing on its potential to lower lipid levels. In an in vivo investigation, normal healthy male Wistar albino rats (n = 60) were divided into two groups: a negative control group (NC) of ten rats and a high-fat diet group (n = 50) that were fed a cholesterol-rich diet until their cholesterol levels reached or exceeded 250 mg/dL. Subsequently, the hypercholesterolemic rats were then randomly allocated to several treatment groups: a positive control (PC); a standard treatment diet (STD) involving fenofibrate at a dose of 20 mg/kg body weight; and three experimental groups (T1, T2, and T3) that received spirulina powder supplementation at doses of 300, 600, and 900 mg per kg body weight, respectively, for the period of 12 weeks. Blood samples were analyzed for oxidative stress biomarkers, insulin levels, lipid profiles, liver function, and expression of gene levels in the diabetogenic pathway. The study utilized spectrophotometric colorimetric methods to identify oxidative stress biomarkers, serum kit methods to measure lipid profiles and liver enzymes, and the assessment of qPCR for mRNA quantity. According to the research findings, spirulina powder has certain noteworthy features. It had the greatest quantity of chlorogenic acid (4052.90 µg/g) among seven phenolics and two flavonoid compounds obtained by HPLC-UV analysis. Furthermore, the proximate analysis demonstrated that spirulina is high in protein (16.45 ± 0.8%) and has a significant energy yield of 269.51 K-calories per 100 g. A maximal spirulina dose of 900 mg/kg/wt significantly lowered oxidative stress, cholesterol, triglyceride, low-density lipoproteins (LDL), and insulin levels (p ≤ 0.05). In contrast, high-density lipoprotein (HDL) and total antioxidant capacity (TAC) levels increased significantly (p ≤ 0.05) compared to all other groups, except the NC group. The study provides remarkable proof about the pharmacological impact of spirulina powders. Significant reductions (p ≤ 0.05) in liver enzymes {alanine aminotransferase (ALT) and aspartate aminotransferase (AST)} were observed across all treatment groups, with the exception of the NC, compared to the positive control. The treatment groups had significantly greater gene expression levels of INS-1, PDX-1, IGF-1, and GLUT-2 than the positive control group (p ≤ 0.05). These findings highlight spirulina’s potential as a long-term regulator of hyperglycemia in rat models with induced hyperlipidemia, owing to its phenolic bioactive components that serve as antioxidants. Full article

Objective: The aim of this study was to analyze the effects of different particle sizes of Tetrastigma hemsleyanum Diels et Gilg (TDG) powders on physical properties, dissolution, in vitro antioxidant activity, and in vivo hepatoprotective properties. Methods: The particle size of TDG coarse powders (TDG-CP), TDG fine powders (TDG-FP), and TDG micro powders (TDG-MP) were measured by a laser particle size analyzer. The physical properties were measured according to the latest version of the Chinese Pharmacopoeia (Committee Chinese Pharmacopoeia 2020). The content of the total flavonoids, total polysaccharides, kaempferol-3-O-rutinoside, and rutin of TDG powders were determined using the NaNO₂-Al (NO₃)₃ colorimetric method, the sulphate-phenol colorimetric method, and HPLC, respectively. In vitro dissolution and antioxidant activity were determined by the paddle method in phosphate buffer (pH 6.8) and the DPPH radical scavenging method, respectively. In addition, the liver tissue pathology was evaluated by hematoxylin and eosin staining (H&E), and the AST and ALT activities were measured by automatic biochemical analyzer. The superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) activities were measured by using commercial analysis kits. Results: As the particle size decreases, the fluidity of TDG powders decreased and the porosity increased. In addition, there were no significant differences in physical properties between low temperature pulverized powders and room temperature pulverized powders. The final dissolution rates of the four bioactive ingredients in TDG-MP were found to be 85.06%, 85.61%, 83.88%, and 83.26%, respectively, whereas in TDG-CP, the dissolution rates were significantly lower at 18.79%, 17.96%, 22.46%, and 24.35%. The EC₅₀ values of TDG-CP, TDG-FP, and TDG-MP on DPPH scavenging activity were 0.82, 0.31, and 0.10 mg/mL, respectively. The AST and ALT activities of the TDG-FP group and the TDG-MP group were significantly decreased and the SOD, CAT, and GSH activities were significantly increased when compared with that of the model group. The inflammatory cell infiltration and vacuolar degeneration of liver cells in the TDG-FP group and the TDG-MP group were significantly improved. Conclusions: The particle size of TDG powders had a significant effect on the physical properties and in vivo bioactivity. TDG pulverized to a fine particle size or smaller is a promising approach for clinical applications with improved physicochemical and biological properties. Full article

Objectives: The objective of this study is to evaluate the wound healing potential of Eurasian propolis by analyzing the phytochemical profile and the biological effects of three representative propolis samples. Methods: Specific colorimetric assays were used to estimate the total phenolic and flavonoid contents and the triterpenoids content. Some of the main components of Eurasian propolis (pinocembrin, pinobanksin, CAPE, chrysin and galangin) were analyzed using HPLC-DAD. Scavenging activity and total antioxidant capacity were assessed through DPPH and ORAC assays, respectively. Human keratinocyte, fibroblast, and monocytic cell lines were used for the biological in vitro analyses. The direct wound healing properties were tested through scratching assays and ELISA kits for the assessment of the production of growth factors (FGF-7, Latency Associated Peptide-LAP), while the indirect effects were evaluated through the estimation of the levels of MMP9, IL-1β, IL-8, and TNF-α using ELISA kits together with a cell-free test on the inhibition capacity on collagenases. Network Pharmacology analysis was employed to further explore possible mechanisms of the action of propolis on the healing process. Results: The analyses confirmed the high phenolic content of Eurasian propolis (142.50–211.30 mg GAE/g), dominated by flavonoids (95.50–196.80 mg Galangin Equivalents/g), and terpenes (431.50–650.00 mg β-sitosterol Equivalents/g), while also verifying the significant antioxidant (4.9–8.9 mM/g Trolox Equivalents) and antiradical (DPPH IC₅₀ 26.1–54.4 μg/mL) activities. The samples showed indirect wound healing properties by mitigating inflammation and remodeling (reduced IL-1β and MMP9) and potentially modulating the immune response (upregulated IL-8). In vitro studies confirmed these effects, demonstrating decreased MMP9 production and collagenase inhibition when cells were co-treated with propolis and a stressor. Propolis also suppressed IL-1β release in fibroblasts, although its impact on TNF-α was inconclusive. Notably, co-treatment upregulated IL-8 in monocytes, suggesting a potential immunomodulatory role. Conclusions: Eurasian propolis may not directly stimulate cell proliferation during wound healing. Its anti-inflammatory and immunomodulatory properties could indicate an indirect contribution in helping the process. Full article

This work reported gold nanoparticles (AuNPs)-based colorimetric immunoassay with the Cu-based metal–organic framework (MOF) to load pyrroloquinoline quinone (PQQ) for the catalytic oxidation of cysteine. In this method, both Cu²⁺ and PQQ in the MOF could promote the oxidation of inducer cysteine by redox cycling, thus limiting the cysteine-induced aggregation of AuNPs and achieving dual signal amplification. Specifically, the recombinant carcinoembryonic antigen (CEA) targets were anchored on the MOF through the metal coordination interactions between the hexahistidine (His₆) tag in CEA and the unsaturated Cu²⁺ sites in MOF. The CEA/PQQ-loaded MOF could be captured by the antibody-coated ELISA plate to catalyze the oxidation of cysteine. However, once the target CEA in the samples bound to the antibody immobilized on the plate surface, the attachment of CEA/PQQ-loaded MOF would be limited. Cysteine remaining in the solution would trigger the aggregation of AuNPs and cause a color change from red to blue. The target concentration was positively related to the aggregation and color change of AuNPs. The signal-on competitive plasmonic immunoassay exhibited a low detection limit with a linear range of 0.01–1 ng/mL. Note that most of the proteins in commercial ELISA kits are recombinant with a His₆ tag in the N- or C-terminal, so the work could provide a sensitive plasmonic platform for the detection of biomarkers. Full article

Gunshot wound morphology and gunshot residues (GSRs) evaluation have been poorly investigated in veterinary forensic pathology. The aims of the present study were to assess the gunshot wound morphology in animals and evaluate the detectability of lead deriving from GSRs using colorimetric techniques. To these aims, cadavers were divided into four different groups. Group A comprised eight animals who died from firearm-related injuries, while groups B and C included dog limbs shot using different shooting ranges; group D comprised dog limbs stabbed with a screwdriver. Morphological analysis was performed on all entry gunshot wounds. Lead residues were investigated using a Bullet Hole Testing Kit (BTK) and Rhodizonate Sodium histochemical staining (NaR-s). Gunshot wounds in group A showed an abrasion ring associated with hemorrhages and tissue necrosis. Groups B and C showed injuries related to the shooting range. NaR-s showed positive results in both animals that died from gunshot wounds and experimentally shot limbs. However, the number of positive cases and the pattern of lead distribution varied with the shooting range. Positive results by BTK were limited to close-contact shots in group B limbs. Our results suggest that both pathological examination and NaR tests represent valid tools for investigating gunshot wounds in veterinary pathology. Full article

Background: The application of normo- and hyperbaric O₂ is a common therapy option in various disease patterns. Thereby, the applied O₂ affects the whole body, including the blood and its components. This study investigates influences of pressure and oxygen fraction on human blood plasma, nutrient media, and the functions of neutrophil granulocytes (PMNs). Methods: Neutrophil migration, reactive oxygen species (ROS) production, and NETosis were examined by live cell imaging. The treatment of various matrices (Roswell Park Memorial Institute 1640 medium, Dulbecco’s Modified Eagle’s Medium, H₂O, human plasma, and isolated PMNs) with hyperbaric oxygen (HBO) was performed. In addition, the expression of different neutrophil surface epitopes (CD11b, CD62L, CD66b) and the oxidative burst were investigated by flow cytometry (FACS). The application of cold atmospheric plasma (CAP) to normoxic and normobaric culture media served as a positive control. Soluble reaction products such as H₂O₂, reactive nitrogen species (RNS: NO₂⁻ and NO₃⁻), and ROS-dependent dihydrorhodamine oxidation were quantified by fluoro- and colorimetric assay kits. Results: Under normobaric normoxia, PMNs migrate slower and shorter in comparison with normobaric hyper- or hypoxic conditions and hyperbaric hyperoxia. The pressure component has less effect on the migration behavior of PMNs than the O₂ concentration. Individual PMN cells produce prolonged ROS at normoxic conditions. PMNs showed increased expression of CD11b in normobaric normoxia, lower expression of CD62L in normobaric normoxia, and lower expression of CD66b after HBO and CAP treatment. Treatment with CAP increased the amount of ROS and RNS in common culture media. Conclusions: Hyperbaric and normobaric O₂ influences neutrophil functionality and surface epitopes in a measurable way, which may have an impact on disorders with neutrophil involvement. In the context of hyperbaric experiments, especially high amounts of H₂O₂ in RPMI after hyperbaric oxygen should be taken into account. Therefore, our data support a critical indication for the use of normobaric and hyperbaric oxygen and conscientious and careful handling of oxygen in everyday clinical practice. Full article

Contamination of trace levels of volatile organic compounds (VOCs) in enclosed spaces is not usually a significant cause for concern; however, it can be relevant in the case of canine scent detection training as a canine’s superior sense of smell makes them highly likely to detect low levels of contamination, contributing to inefficient training. Thus, herein, we address the need for a simple, low-cost, robust, vapochromic sensor to determine the cross-contamination of VOCs within closed containers, such as canine training aid kits. This study focuses on the development of a vapor sensor, which produces a rapid colorimetric change when a target chemical vapor is present. A pH indicator is used as the colorimetric dye and its incorporation into a sol–gel matrix on a paper substrate is confirmed via SEM characterization. The sensor’s stability and performance is tested against exposure to various levels of sunlight and temperature. The design allows the sensor to present a clear and unambiguous visible response to the release of the volatile target within a closed container. It can be readily incorporated into existing training kits and functions as a straightforward reminder of when training aids need to be changed or a new containment system should be considered. Full article