Search Results (2,167)

Bacteriophage therapy can successfully provide additional treatment to control Salmonella infection, but low gastric pH limits its oral application. The present study aimed to develop an improved encapsulation formulation with enhanced acid protection for oral delivery of Salmonella phages using polymers. This was achieved by encapsulating a phage cocktail containing three different bacteriophages against Salmonella sp. in alginate beads incorporating polyvinyl alcohol (PVA), PVP-K30, and calcium carbonate as viscosity modifiers and acid protection enhancers. Further, the beads were coated with poly-L-lysine to improve the stability and tested for their efficacy for improved phage viability under in vitro acidic conditions for subsequent use in oral delivery. Moist beads were slimy, and semi-dried beads presented a coarse surface as observed using FE-SEM. In vitro studies revealed that the free phage cocktail exhibited complete inactivation when exposed to acidic pH 2.5 after 15 min incubation. In contrast, the encapsulated phage cocktail showed a decrease of only 1.66 log units in viability when incubated for 90 min at pH 2.5. Furthermore, oral delivery of the encapsulated phage cocktail in the poultry model significantly reduced bacterial load in infected birds’ intestines. Full article

Preeclampsia (PE) is a hypertensive disorder impacting 5–7% of pregnancies globally. With no causative treatment available, diagnosed patients have limited therapeutic options, putting them at risk for pregnancy complications. The induction of oxidative stress by ROS—one of the major contributors in PE pathogenesis—causes downstream signaling and production of anti-angiogenic factors, such as sFLT1 and sEng. The anti-angiogenic factors may cause endothelial and trophoblast dysfunction, contributing to the development of hypertension, proteinuria, and in severe cases, eclampsia. To target placental oxidative stress, we developed and evaluated an organofluorine hydrazone antioxidant, HY-12, in vitro. Human trophoblast (HTR8/SVneo) cells were incubated with hydrogen peroxide to induce oxidative stress and act as a model of PE. The goal of the study was to assess the efficacy of HY-12 and its ability to reduce cell injury, mitochondrial stress, and anti-angiogenic response. In our human trophoblast-based assays, pre-treatment with HY-12 reduced mitochondrial-derived ROS production in cells exposed to hydrogen peroxide, proving its ability to alleviate the oxidative stress associated with the pathogenesis of PE. HY-12 reduced HIF1A expression and sFLT1 protein expression in H₂O₂-exposed HTR8 cells. Furthermore, HY-12 improved the activity of the mitochondrial electron chain enzyme cytochrome C oxidase (COX) in the hydrogen-peroxide-treated HTR8/SVneo cells, which is a promising attribute of the compound. In reducing placental trophoblast oxidative stress, HY-12 shows promise as a potential treatment of preeclampsia. In vivo studies are warranted to further determine the efficacy of this compound. Full article

Rheumatoid arthritis (RA) is an autoimmune disease affecting the synovium. Mitochondrial dysfunction is considered a critical factor in the pathogenesis of RA. The aim of the study was to determine the effect of methotrexate and tofacitinib on mitochondrial function and oxidative stress in an in vitro study on the model synovial cell line SW982. TNF-alpha-stimulated SW982 cells, as well as control untreated cells, were incubated with methotrexate and tofacitinib. A metabolic test was performed to assess mitochondrial function. The oxidative stress generated after the application of the therapeutics was determined by a chromatographic analysis. The results obtained showed an increase in ATP levels (p < 0.0001) and a decrease in proton leak (p < 0.0003) after treatment with tofacitinib. The opposite trend was observed—reduced ATP production (p < 0.0096) and increased levels of proton leak (p < 0.0001)—after treatment with methotrexate. A two-fold increase in 8-ISOPGF2A was measured in comparison to TNF-alpha-stimulated and untreated cells. The dynamics of mitochondrial activity and oxidative stress were monitored in a certified RA model cell line after the administration of two different therapeutics. Methotrexate was found to induce mitochondrial dysfunction and oxidative stress in vitro, while tofacitinib partially improved mitochondrial parameters. Full article

Background/Objectives: This study reports the synthesis of TiO₂ nanoparticles, their functionalization with folic acid (FA), and the subsequent loading with zinc phthalocyanine (ZnPc) to develop photosensitizers for photodynamic therapy (PDT) targeting glioma cells. Methods: TiO₂, TiO₂-FA, and TiO₂-FA-ZnPc nanoparticles were synthesized via a sol–gel process involving the hydrolysis and condensation of titanium (IV) isopropoxide. FA and ZnPc were incorporated in vitro during the synthesis. The resulting materials were characterized by transmission and scanning electron microscopy (TEM and SEM), X-ray diffraction (XRD), Raman and UV–Vis spectroscopy, thermogravimetric analysis (TGA), and nitrogen adsorption–desorption measurements. Reactive oxygen species (ROS) generation was evaluated in vitro using the 1,3-diphenylisobenzofuran (DPBF) probe. A 40 ppm solution of each TiO₂ system was irradiated with UV light, and the degradation of DPBF was monitored. Biological assays were conducted to assess the viability of human glioblastoma cells (LN18 and U251) incubated with the TiO₂-based materials, with and without UV exposure. Human fibroblast cells (BJ) were used to evaluate biocompatibility. Results: All TiO₂-based materials retained key characteristics, including high surface area (~600–700 m²/g), mesoporous structure (pore diameter ~4–5 nm), mixed anatase–amorphous morphology, and a bandgap of approximately 3.46 eV. The UV–Vis spectrum of TiO₂-FA-ZnPc displayed additional absorption bands in the visible region (600–700 nm), consistent with ZnPc incorporation. Upon UV irradiation, the DPBF absorbance at 410 nm decreased over time, indicating ROS generation and resulting in complete degradation within 10 min (TiO₂), 12 min (TiO₂-FA), and 14 min (TiO₂-FA-ZnPc). BJ cells exhibited good biocompatibility at all concentrations. LN18 and U251 cells showed no cytotoxicity below 100 μg/mL unless exposed to UV light. Conclusions: The synthesized TiO₂-based systems demonstrate good biocompatibility and significant phototoxicity under UV irradiation, highlighting their strong potential for application in photodynamic therapy. Full article

Direct-fed microbials (DFM) have emerged as a promising dietary strategy for enteric methane abatement. However, it is unclear whether in vitro studies trialing DFM should use ruminal fluid previously adapted to the DFM of interest or if the DFM can be directly added to an unadapted inoculum. Ten lactating, multiparous, rumen cannulated Holstein-Friesian cows were randomly allocated to one of two groups: 1) adapted, basal diet plus 4 g/d of a blend of Bacillus subtilis and Bacillus licheniformis (1.6 × 10⁹ CFU/g each), delivered via the rumen canula; and 2) naive, basal diet only. Ruminal fluid from both groups was incubated in an in vitro 24-h batch culture system with two rates of Bacillus spp. and three feed substrates (hay, pasture, cereal grain), resulting in 12 treatments. Methane production was 16% greater, and total volatile fatty acid concentration was 7% greater in incubations using adapted ruminal fluid compared to those using naive ruminal fluid; however, neither parameter was affected when Bacillus spp. was added to the batch incubation system. Future in vitro studies evaluating DFM should consider including a period of in vivo adaptation to mimic their potential impact under in vivo feeding conditions. Full article

Skin aging is commonly characterized by increased wrinkles, loss of elasticity, and hyperpigmentation, significantly affecting personal appearance and quality of life. Although botulinum toxin type A (BTX-A) has been widely applied in cosmetic anti-wrinkle treatments, its intrinsic cytotoxicity limits broader clinical applications. In this study, we developed a novel exosome-based BTX-A composite delivery system designed to synergize the anti-aging properties of exosomes with the wrinkle-reducing effects of BTX-A while reducing toxicity. Human adipose-derived mesenchymal stem cells were genetically modified via lentiviral transduction to overexpress Synaptic Vesicle Glycoprotein 2C (SV2C), the receptor of BTX-A, thereby producing SV2C-enriched functionalized exosomes (EXO^SV2C). These exosomes (2.0 × 10⁷ particles/mL) were incubated with BTX-A (3 U/mL) to generate the EXO^SV2C-BTX-A complex. In vitro, EXO^SV2C-BTX-A significantly promoted the proliferation and migration of human dermal fibroblasts and effectively alleviated D-galactose (D-gal)-induced cellular senescence and collagen type I loss. These effects were superior to those observed with either BTX-A or exosomes alone. In vivo, intradermal injection of EXO^SV2C-BTX-A for 28 days markedly suppressed D-gal-induced skin aging in 8-week-old male KM mice, as evidenced by reduced malondialdehyde levels in dermal tissue, enhanced collagen type I expression, and preserved skin structure. Notably, the composite exhibited significantly lower toxicity compared to free BTX-A. Collectively, these findings highlight EXO^SV2C-BTX-A as a promising exosome-mediated BTX-A delivery platform with enhanced anti-aging efficacy and improved biocompatibility, offering a potential therapeutic strategy for skin rejuvenation. Full article

Biologically based detoxification strategies are increasingly being explored as alternatives to conventional methods for the removal of toxic contaminants in food products. Among these, aflatoxin B1 (AFB1) is one of the most potent mycotoxins due to its high toxicity, genotoxicity, and persistence in the human body once ingested. In this study, the detoxification potential of bacterial strains belonging to the genera Lactobacillus/Pediococcus (n = 10) and Bacillus (n = 10) was evaluated using extracts from naturally contaminated corn flour. Detoxification was assessed after incubation for 12, 24, and 48 h in specific culture media. AFB1 quantification and metabolite profiling were performed at each time point using Quadrupole Time-of-Flight Mass Spectrometry (LC-QTOF-MS). The highest detoxification rates were observed with Lactobacillus curvatus 14 (L. curvatus 14) (41.1 ± 19.3%) and Pediococcus pentosaceus 4 (P. pentosaceus 4) (25.4 ± 11.3%) after 48 h, and Bacillus firmus 6 (B. firmus 6) (25.1 ± 12.9%) after 24 h. An in vitro digestion model was also applied to assess detoxification under gastrointestinal conditions. Results showed substantial AFB1 reduction at the colonic stage, reaching 72.26 ± 7.54% for P. pentosaceus 4 and 69.67 ± 9.70% for L. curvatus 14. These findings underscore the potential application of Lactobacillus, Pediococcus, and Bacillus strains in biological detoxification strategies to reduce dietary exposure to AFB1. Full article

Background/Objectives: Bisphenols (BPs) and especially bisphenol S (BPS), an analog of bisphenol A (BPA), are widely used and induce oxidative stress, resulting in the inhibition of cell proliferation and induction of apoptosis which all are crucial for reproduction, the progression of pregnancy, and fertility. The present study integrates trophoblastic cells as an in vitro model to provide evidence and investigate the molecular interactions regarding placenta-related pregnancy complications after cytotoxic exposure to BPS. Methods: Human endometrial epithelial adenocarcinoma Ishikawa cell lines and trophoblastic cells were cultured. Cells obtained from the cultures were divided into plates and incubated for 24 h with different concentrations of bisphenol S (BPS). Cell viability was measured using the Countess Automated Cell Counter and the viability of Ishikawa cells was assessed after 48 h and for trophoblasts after 24 h. The effect of siRNA on NANOG expression was evaluated using qRT-PCR. Quantification of DNMT and NANOG was performed by qPCR and the G6PD gene was used as an internal control. Results: Real-time PCR results showed that the expression of the DNMT1 gene varies depending on the concentration of BPS in trophoblastic cells. In Ishikawa cell lines, real-time PCR results showed that DNMT1 gene expression was higher due to cell increase, but the measured fold change did not differ significantly. Data analysis indicated a statistically significant difference between CpDNMT1 in trophoblasts with and without BPS, where higher values were observed in the case of BPS presence (p = 0.019). The largest difference was observed between CpDNMT1 trophoblasts without BPS and CpDNMT1 Ishikawa with BPS (p < 0.001). Silencing the NANOG gene resulted in a reduced expression of DNMT1, while the G6PD gene was still detected. Conclusions: The results of this study highlight the cytotoxic effects of BPS and consequently its effect on trophoblast viability. The results of NANOG-DNMT1 gene expression related to BPS exposure reinforces our understanding of EDC-induced placental dysfunction. Full article

This study evaluated the effects of replacing soybean meal (SBM) with winged bean tuber (Psophocarpus tetragonolobus) fermented using ruminal Candida tropicalis KKU20 on gas kinetics, ruminal fermentation, and degradability using the in vitro gas production technique. A 3 × 4 factorial arrangement in a completely randomized design was used. Factor A included three roughage-to-concentrate (R:C) ratios: 60:40, 50:50, and 40:60. Factor B consisted of four levels of SBM replacement with yeast-fermented winged bean tuber (YFWBT): 0%, 33%, 66%, and 100%. Fermentation with C. tropicalis KKU20 increased the crude protein content of winged bean tuber by 13.32%. No significant interaction was found between the R:C ratio and YFWBT level for cumulative gas production at 24, 48, or 96 h (p > 0.05). Cumulative gas production at 96 h was not affected by either factor. However, at 24 and 48 h, gas production increased with higher proportions of concentrate (p < 0.05). Both the R:C ratio and YFWBT level significantly influenced pH and ammonia–nitrogen (NH₃-N) concentrations (p < 0.01). After 24 h, NH₃-N ranged from 7.66 to 13.76 mg/dL, rising to 16.44–16.63 mg/dL after 48 h. A significant interaction (p < 0.01) was observed for in vitro dry matter degradability (IVDMD) and in vitro organic matter degradability (IVOMD). Increasing concentrate levels along with YFWBT inclusion improved degradability at both incubation times. The highest IVDMD (64.49%) and IVOMD (65.81%) were recorded at 48 h in the 40:60 R:C ratio with 33% YFWBT. At 48 h, a significant interaction effect (p < 0.05) was also found for total volatile fatty acid (VFA) and propionic acid (C3) concentrations. Total VFA peaked in the 40:60 group with 33% YFWBT (104.31 mM), while the highest C3 concentration (26.22%) was observed in the same R:C group with 66% YFWBT. At 24 h, total VFA was significantly affected by the R:C ratio (p < 0.05), with the lowest values in the 60:40 group and increasing in response to higher concentrate and YFWBT levels. Incorporating YFWBT at 33% in diets with an R:C ratio of 40:60 optimized degradability, indicating its potential as a sustainable alternative to SBM in ruminant nutrition. Full article

Enhancing forage protein is key to sustainable ruminant nutrition. The nutritive value of Hedysarum coronarium L. was investigated by studying different preservation systems (fresh vs. dehydrated pellet vs. hay) (Exp. 1) and morphological fractions (flowers vs. leaves vs. stems) (Exp. 2). For the fresh and pelleted systems, two cuts were used. Proximately, total polyphenols and condensed tannins were detected. In vitro fermentation characteristics were studied by incubating samples with buffered sheep rumen fluid, estimating methane production by volatile fatty acids. Fresh and pelleted sulla were more nutritionally advantageous than hay, in terms of metabolizable energy and protein-to-fiber ratio. Pelleting at the beginning of flowering proved to be a suitable forage for sheep feeding due to protein (15.1% DM), metabolizable energy (9.64 MJ/kg DM), structural carbohydrates (39.5% DM), and total polyphenols (13.5 GAE g/kg DM) content. This cut showed an in vitro fermentation rate (9.86 mL/h), organic matter degradability (55.7%), and volatile fatty acids (87.3 mmoL/g) that were higher (p < 0.05) than hay. Flowers and leaves showed higher (p < 0.05) levels of secondary metabolites than stems as well as lower methane production. These results suggest the potential influence of these compounds in reducing rumen emissions. Dehydration and pelleting resulted in an effective preservation method for maintaining nutrients in sulla forage. Full article

Previous in vitro studies have shown the therapeutic potential of bee venom (BV) against different types of glioblastoma cells. Our aim was to evaluate the cytotoxic effect of BV on glioma in the zebrafish model. First, safe concentrations of BV and melittin were determined by determining the LD₅₀ for each substance. Two human glioma cell lines, 8MGBA and LN-229, were used in this study. After staining the tested cells for visualization under UV light, they were then implanted into 2-day-old zebrafish embryos. Zebrafish were incubated for 3 days with crude BV and melittin at concentrations of 1.5 and 2.5 µg/mL vs. control group. Tumor growth was assessed with a stereo microscope. We found differential proliferative responses of two human glioma lines in a zebrafish model. The 8MGBA cell line, but not LN-229, showed proliferative potential when implanted into 2-day-old zebrafish embryos. This study showed a dose-dependent cytotoxic effect only for BV against 8MGBA cells. The observed cytotoxic effect is not dependent on the presence of the peptide melittin—the main BV component with the greatest cytotoxic potential. Simultaneously, a slight increase in LN-229 cell proliferation was observed after 3 days of incubation with melittin at a concentration of 2.5 µg/mL. This indicates that any consideration of bee venom as a therapeutic substance must take into account the type of glioblastoma. Full article

While the Ankom RF system facilitates efficient high-throughput in vitro fermentation studies, its high cost and limited flexibility constrain its broader applicability. To address these limitations, we developed and validated a low-cost, modular gas monitoring system (FerME), assembled from commercially available components. To evaluate its performance and reproducibility relative to the Ankom RF system (Ankom Technology, Macedon, NY, USA), in vitro rumen fermentation experiments were conducted under strictly controlled and identical conditions. Whole rumen contents were collected approximately 2 h post-feeding from individual mid- or late-lactation dairy cows and immediately transported to the laboratory. Each fermenter received 50 mL of processed rumen fluid, 100 mL of anaerobically prepared artificial saliva buffer, and 1.2 g of the donor cow’s diet. Bottles were sealed with the respective system’s pressure sensors, flushed with CO₂, and incubated in a 50 L water bath maintained at 39 °C. FerME (New Taipei City, Taiwan) and Ankom RF fermenters were placed side-by-side to ensure uniform thermal conditions. To assess the effect of filter bag use, an additional trial employed Ankom F57 filter bags (Ankom Technology, Macedon, NY, USA; 25 μm pore size). Trial 1 revealed no significant differences in cumulative gas production, volatile fatty acids (VFAs), NH₃-N, or pH between systems (p > 0.05). However, the use of filter bags reduced gas output and increased propionate concentrations (p < 0.05). Trial 2, which employed filter bags in both systems, confirmed comparable results, with the FerME system demonstrating improved precision (CV: 4.8% vs. 13.2%). Gas composition (CH₄ + CO₂: 76–82%) and fermentation parameters remained consistent across systems (p > 0.05). Importantly, with 12 pressure sensors, the total cost of FerME was about half that of the Ankom RF system. Collectively, these findings demonstrate that FerME is a reliable, low-cost alternative for real-time rumen fermentation monitoring and could be suitable for studies in animal nutrition, methane mitigation, and related applications. Full article

The kinetics of insulin aggregation and fibril formation were studied in vitro using Scanning Electron Microscopy (SEM) and Fourier Transform Infrared (FTIR) spectroscopy. Our investigation centered on the protein’s morphological and structural changes to better understand the transient molecular configurations that occur during the lag phase. SEM images showed that, already at early incubation stages, a network of disordered pseudo-filaments, ranging in length between 200 and 500 nanometers, develops on the surface of large aggregates. At later stages, fibrils catalyzed by protein aggregates were observed. Principal Component Analysis (PCA) of the FTIR data identified signatures of intramolecular β-sheet secondary structures forming during the lag phase and at the onset of the exponential growth phase. These absorption bands are linked to secondary nucleation mechanisms due to their transient nature. This interpretation is further supported by a chemical equilibrium model, which yielded a reliable secondary nucleation rate constant, K₂, on the order of 10⁴ M⁻² s⁻¹. Full article

The aim of this work is to evaluate the effectiveness of antimicrobial photodynamic therapy (aPDT) against oral MDR (multi-drug-resistant) Candida spp. infections related to orthodontic treatment with palatal expanders through in vitro study. Methods: PDT protocol: Curcumin + H₂O₂ was used as a photosensitizer activated by a 460 nm diode LED lamp, with an 8 mm blunt tip for 2 min in each spot of interest. In vitro simulation: A palatal expander sterile device was inserted into a custom-designed orthodontic bioreactor, realized with 10 mL of Sabouraud dextrose broth plus 10% human saliva and infected with an MDR C. albicans clinical isolate CA95 strain to reproduce an oral palatal expander infection. After 48 h of incubation at 37 °C, the device was treated with the PDT protocol. Two samples before and 5 min after the PDT process were taken and used to contaminate a Petri dish with a Sabouraud field to evaluate Candida spp. CFUs (colony-forming units). Results: A nearly 99% reduction in C. albicans colonies in the palatal expander biofilm was found after PDT. Conclusion: The data showed the effectiveness of using aPDT to treat palatal infection; however, specific patient oral micro-environment reproduction (Ph values, salivary flow, mucosal adhesion of photosensitizer) must be further analyzed. Full article

Background/Objectives: Tamoxifen, a selective estrogen receptor modulator widely used as an adjunct in the treatment of breast cancer, has known effects on bone metabolism, although its impact on osseointegration and cellular responses during early bone healing remains unclear. Understanding these effects is essential given the increasing use of dental implants in cancer survivors. The study aimed to observe the influence of tamoxifen on human osteosarcoma (SAOS-2) cells lines, as well on the osseointegration of titanium implants in ovariectomized female rats. Methods: SAOS-2 cells were incubated with Dulbecco’s modified growth medium. Six titanium (Ti) disks were used at each time point. The samples were divided into groups with the presence (TAM, n = 36) or not (CTR, n = 36) of tamoxifen in a concentration of 2 μM. In vivo, 72 animals were divided in groups with bilateral ovariectomy or SHAM and tamoxifen administration or not (15 mg/kg). Cell viability, mineralization rate, and collagen synthesis were assessed, as well as bone/implant contact (BIC) and bone ingrowth (BIN). Results: Tamoxifen caused a decrease in SAOS-2 viability, although an increase in the mineralization rate was observed. In vivo, the TAM groups presented higher BIC and BIN when compared to their control, but a lower percentage of mature collagen cells. Conclusions: Based on our findings, in vitro, the therapy with TAM slightly reduced the viability of SAOS-2 cells while significantly increasing the mineralization rate. In vivo, the therapy positively influenced BIC and BIN during the osseointegration phase. Full article