Cell Line THP-1 Research Articles

Polysaccharides and Atherosclerosis: Focus on the Role of Key Genes of Cholesterol Metabolism in Macrophages

Based on nutrigenomics approaches, the mechanisms of action of glucan on the accumulation and efflux of cholesterol Cholesterol from macrophages derived from the THP-1 cell line were studied. At the first stage, it was established that glucan does not have the ability to inhibit the accumulation of Cholesterol in the macrophages caused by the action of modified low-density lipoproteins (mLDL) on them. However, glucan accelerates the efflux of Cholesterol from the macrophages loaded with it, and this effect is commensurate with the action of verapamil and alpinetin. At the second stage of the work, transcriptomic analysis made it possible to detect differentially expressed genes by observing the effect of glucan on both accumulation and efflux of Cholesterol from the macrophages. In mLDL-induced Cholesterol accumulation, the presence of glucan in the cell model did not change the overall pattern of gene expression, at the same time, monitoring Cholesterol efflux revealed the ability of glucan to increase (LDLR, INSIG1, SCARB1, STARD3, ABCA1) or decrease (FADS1, SR-BI, DSC1) the expression of a set of genes different from the key Cholesterol accumulation genes. Evaluation of the biological activity of genes, associated with accumulation Cholesterol, indicates their role in immune and inflammatory processes, while genes with other functions, including genes known as regulators of Cholesterol metabolism, are involved to a greater extent in Сholesterol efflux under the action of glucan. A conclusion was made that in relation to the pathology of atherosclerosis, it is necessary to consider glucan not so much as a prophylactic, but as a therapeutic agent.

Designing sustainable antioxidant and anti-inflammatory ingredients from seaweeds for functional gummies

Atlantic wakame (AW) and sea lettuce (SL) are edible seaweeds commonly produced in Asia and Europe, typically consumed as fresh or dried biomass. Because of the short shelf life, developing new post-harvest treatments, such as the extraction of bioactive compounds, is essential for establishing reliable macroalgae supply chains for food applications. This study aims to develop a sustainable and functional seaweed-based food ingredient rich in bioactive compounds for functional food applications. The results indicated that ultrasound-assisted extraction using 70% ethanol for 10 minutes was optimal for both SL (410±23 mg of gallic acid equivalent [GAE] per kg) and AW (666±63 mg GAE/kg). Both extracts exhibited chemical antioxidant activity and inhibited the protection of human plasma against chemically induced oxidation. SL extract did not exhibit free radical scavenging activity (DPPH). Importantly, no cytotoxicity (IC50>250 µg/mL) was observed in human-derived THP-1, HUVEC, HT-29, and HepG2 cell lines, ensuring the relative safety of these extracts for potential food technology applications. AW and SL extracts (100 µg/mL) reduced intracellular reactive oxygen species generation in lipopolysaccharide (LPS)-challenged THP-1 cells by 66% and 30%, respectively, highlighting the higher antioxidant activity of AW extract. AW and SL extracts did not modulate TNF-α and IL-6 secretion to show anti-inflammatory activity. Gummies infused with AW and SL extracts had a higher total phenolic content than commercial products, but their antioxidant activity was lower. Sensory evaluation revealed that gummies containing seaweed extracts received lower scores for colour, taste, overall impression, and purchase intention than those without extracts.

5'-Noraristeromycin Repurposing: Well-known S-Adenosyl-L-homocysteine Hydrolase Inhibitor As a Potential Drug Against Leukemia

5ˊ-Noraristeromycin as a racemic mixture of enantiomers was found to exhibit a pronounced cytotoxic effect on leukemia cells; IC50 for the Jurkat, K562, and THP-1 cell lines was 7.3, 1.3, and 3.7 μM, respectively. The general toxicity of 5'-noraristeromycin was studied in experiments on white mice upon single-dose intragastric administration; toxicometric parameters were determined, and the clinical and pathomorphological presentation of acute intoxication was studied. LD50 of the substance was shown to be 63.2 (52.7÷75.8) mg/kg; LD16, 44.7 mg/kg, and LD84, 89.4 mg/kg. Administration of the substance at a dose within the studied dose range is accompanied by systemic damage to the internal organs and tissues of the experimental animals.

Induction of the PERK-eIF2𝛼-ATF4 pathway in M1 macrophages under endoplasmic reticulum stress

Translation inhibition can activate two cell death pathways. The first pathway is activated by translational aberrations, the second by endoplasmic reticulum (ER) stress. In this work, the effect of ribosome-inactivating protein type II (RIP-II) viscumin on M1 macrophages derived from the THP-1 cell line was investigated. The number of modified ribosomes was evaluated by real-time PCR. Transcriptome analysis revealed that viscumin induces the ER stress activated by the PERK sensor.

Preparation and Complex Characterisation of Stabilised Gold Nanoparticles: Biodistribution and Application for High Resolution In Vivo Imaging

The Turkevich method was optimized to prepare gold nanoparticles (AuNP) stabilized by polyethyleneglycol (PEG) for µCT. Using various independent modalities, we thoroughly characterized the optimized PEG-AuNPs. Here, we show that PEG-AuNPs are retained in the blood and provide a high contrast in the high-resolution µCT imaging of blood vessels and inner organs. The biodistribution is characterized by prolonged circulation in the blood and accumulation in the liver, spleen and skin. The accumulation of AuNP in the skin resulted in the blue discoloration of eyes and the whole skin. In vitro experiments using a leukemic monocyte THP-1 cell line model expressing high levels of NLRP3 demonstrated that the NLRP3inflammasome was not activated by PEG AuNP. Over 9 months, the mice were scanned by µCT and were in good health. Scans in mice using PEG-stabilized AuNPs in this study were sharper, with a higher contrast, when compared to a commercial contrasting agent at the same dose. The PEG-AuNPs were morphologically and chemically stable for at least two years when stored in the refrigerator.

Hydrolytic instability of laser-ablatively deposited CaSi2 coatings in air and neutral water affects the behavior of bone healing-related cell types

Calcium silicide (CaSi2) instability in humid air or pH-neutral water remains unknown, although the topochemical formation of silicene from CaSi2 in various aqueous phases is a well-known process. Here we report on laser ablation of CaSi2 in the vacuum and ethanol, characterize coatings deposited on titanium substrates with electron microscopy, X-ray photoelectron, Raman, and infrared spectroscopy, and examine the instability of the deposited coatings in humid air and neutral water. We further investigate the behavior of human mesenchymal stromal cells (hMSCs), vascular cells (HUVECS, human umbilical vein endothelial cells), and macrophages (derived from THP-1 cell line) in contact with the deposited coatings submerged in cell culture medium. The observed results indicate that the CaSi2 coatings undergo topochemical conversion to silicene, which is accompanied by hydrolytic reactions leading to inorganic Ca compounds and SiO2, and that the response of all cell types in the hydrolyzed CaSi2 surface domain is negatively affected by the nature of the hydrolytic products. In contrast, cells showed a tendency for enhanced biocompatibility towards the CaSi2 particles ablated in the vacuum. The results suggest that coating approaches can significantly influence cell behavior outcomes.

1-Piperidine Propionic Acid Protects from Septic Shock Through Protease Receptor 2 Inhibition.

Sepsis is a complex disorder caused by a dysregulated host response to infection, with high levels of morbidity and mortality. Treatment aimed to modulate immune response and maintain vascular function is still one of the major clinical challenges. This study was designed to test the effect of the small molecule 1-Piperidine Propionic Acid (1-PPA) as molecular targeted agent to block protease-activated receptor 2 (PAR2), one of the major modulators of inflammatory response in LPS-induced experimental endotoxemia. In the THP-1 cell line, LPS-induced cytokine expression was inhibited by 1-PPA in a dose-dependent manner. In LPS-injected mice, treatment with 1-PPA was effective in reducing mortality and sepsis-related symptoms and improved cardiac function parameters. After 6 h from LPS injection, a significant decrease in IL-6, IL-1β, and IL-10 was observed in the lung tissue of 1-PPA-treated mice, compared to controls. In these mice, a significant decrease in vasoactive molecules, especially kininogen-1, was also observed, mainly in the liver. Histopathological analysis confirmed typical features of sepsis in different organs and these findings were markedly reduced in mice treated with 1-PPA. These data demonstrate the effectiveness of 1-PPA in protecting the whole organism from sepsis-induced damage.

LncRNA BRE-AS1 regulates the JAK2/STAT3-mediated inflammatory activation via the miR-30b-5p/SOC3 axis in THP-1 cells

Long non-coding RNAs (lncRNAs) have emerged as pivotal regulators in numerous biological processes, including macrophage-mediated inflammatory responses, which play a critical role in the progress of diverse diseases. This study focuses on the regulatory function of lncRNA brain and reproductive organ-expressed protein (BRE) antisense RNA 1 (BRE-AS1) in modulating the inflammatory activation of monocytes/macrophages. Employing the THP-1 cell line as a model, we demonstrate that lipopolysaccharide (LPS) treatment significantly upregulates BRE-AS1 expression. Notably, specific knockdown of BRE-AS1 via siRNA transfection enhances LPS-induced expression of interleukin (IL)-6 and IL-1β, while not affecting tumor necrosis factor (TNF)-α levels. This selective augmentation of pro-inflammatory cytokine production coincides with increased phosphorylation of Janus kinase (JAK)2 and signal transducer and activator of transcription (STAT)3. Furthermore, BRE-AS1 suppression results in the downregulation of suppressor of cytokine signaling (SOCS)3, an established inhibitor of the JAK2/STAT3 pathway. Bioinformatics analysis identified binding sites for miR-30b-5p on both BRE-AS1 and SOCS3 mRNA. Intervention with a miR-30b-5p inhibitor and a synthetic RNA fragment that represents the miR-30b-5p binding site on BRE-AS1 attenuates the pro-inflammatory effects of BRE-AS1 knockdown. Conversely, a miR-30b-5p mimic replicated the BRE-AS1 attenuation outcomes. Our findings elucidate the role of lncRNA BRE-AS1 in modulating inflammatory activation in THP-1 cells via the miR-30b-5p/SOCS3/JAK2/STAT3 signaling pathway, proposing that manipulation of macrophage BRE-AS1 activity may offer a novel therapeutic avenue in diseases characterized by macrophage-driven pathogenesis.

Impact of Senescent Cell-Derived Extracellular Vesicles on Innate Immune Cell Function.

Extracellular vesicles (EVs) are components of the senescence-associated secretory phenotype (SASP) that influence cellular functions via their cargo. Here, the interaction between EVs derived from senescent (SEVs) and non-senescent (N-SEVs) fibroblasts and the immune system is investigated. Via endocytosis, SEVs are phagocytosed by monocytes, neutrophils, and B cells. Studies with the monocytic THP-1 cell line find that pretreatment with SEVs results in a 32% (p < 0.0001) and 66% (p < 0.0001) increase in lipopolysaccharide (LPS)-induced tumor necrosis factor-alpha (TNF-α) production when compared to vehicle control or N-SEVs respectively. Interestingly, relative to vehicle control, THP-1 cells exposed to N-SEVs exhibit a 20% decrease in TNF-α secretion (p < 0.05). RNA sequencing reveals significant differences in gene expression in THP-1 cells treated with SEVs or N-SEVs, with vesicle-mediated transport and cell cycle regulation pathways featuring predominantly with N-SEV treatment, while pathways relating to SLITS/ROBO signaling, cell metabolism, and cell cycle regulation are enriched in THP-1 cells treated with SEVs. Proteomic analysis also reveals significant differences between SEV and N-SEV cargo. These results demonstrate that phagocytes and B cells uptake SEVs and drive monocytes toward a more proinflammatory phenotype upon LPS stimulation. SEVs may therefore contribute to the more proinflammatory immune response seen with aging.

Treatment of polyethylene microplastics degraded by ultraviolet light irradiation causes lysosome-deregulated cell death

BackgroundMicroplastics (MPs), plastic particles < 5 mm in size, are prevalent in the environment, and human exposure to them is inevitable. To assess the potential risk of MPs on human health, it is essential to consider the physicochemical properties of environmental MPs, including polymer types, size, shape, and surface chemical modifications. Notably, environmental MPs undergo degradation due to external factors such as ultraviolet (UV) rays and waves, leading to changes in their surface characteristics. However, limited knowledge exists regarding the health effects of MPs, with a specific focus on their surface degradation. This study concentrates on cytotoxic MPs with surface degradation through UV irradiation, aiming to identify the mechanisms underlying their cell toxicity.ResultsPolyethylene (PE) and surface-degraded PE achieved through UV light irradiation were employed as model MPs in this study. We explored the impact of PE and degraded PE on cell death in murine macrophage cell line RAW264.7 cells and human monocyte cell line THP-1 cells. Flow cytometric analysis revealed that degraded PE induced programmed cell death without activating caspase 3, while non-degraded PE did not trigger programmed cell death. These findings suggest that degraded PE might induce programmed cell death through mechanisms other than caspase-driven apoptosis. To understand the mechanisms of cell death, we investigated how cells responded to degraded PE-induced cellular stress. Immunofluorescence and western blotting analyses demonstrated that degraded PE induced autophagosome formation and increased p62 expression, indicating inhibited autophagy flux after exposure to degraded PE. Furthermore, degraded PE exposure led to a decrease in acidic lysosomes, indicating lysosomal dysregulation. These results imply that degraded PE induces lysosomal dysfunction, subsequently causing autophagy dysregulation and cell death.ConclusionsThis study unveils that UV-induced degradation of PE results in cell death attributed to lysosomal dysfunction. The findings presented herein provide novel insights into the effects of surface-degraded MPs on biological responses.

Assessing the Toxicity of Metal- and Carbon-Based Nanomaterials In Vitro: Impact on Respiratory, Intestinal, Skin, and Immune Cell Lines.

The field of nanotechnology has experienced exponential growth, with the unique properties of nanomaterials (NMs) being employed to enhance a wide range of products across diverse industrial sectors. This study examines the toxicity of metal- and carbon-based NMs, with a particular focus on titanium dioxide (TiO2), zinc oxide (ZnO), silica (SiO2), cerium oxide (CeO2), silver (Ag), and multi-walled carbon nanotubes (MWCNTs). The potential health risks associated with increased human exposure to these NMs and their effect on the respiratory, gastrointestinal, dermal, and immune systems were evaluated using in vitro assays. Physicochemical characterisation of the NMs was carried out, and in vitro assays were performed to assess the cytotoxicity, genotoxicity, reactive oxygen species (ROS) production, apoptosis/necrosis, and inflammation in cell lines representative of the systems evaluated (3T3, Caco-2, HepG2, A549, and THP-1 cell lines). The results obtained show that 3T3 and A549 cells exhibit high cytotoxicity and ROS production after exposure to ZnO NMs. Caco-2 and HepG2 cell lines show cytotoxicity when exposed to ZnO and Ag NMs and oxidative stress induced by SiO2 and MWCNTs. THP-1 cell line shows increased cytotoxicity and a pro-inflammatory response upon exposure to SiO2. This study emphasises the importance of conducting comprehensive toxicological assessments of NMs given their physicochemical interactions with biological systems. Therefore, it is of key importance to develop robust and specific methodologies for the assessment of their potential health risks.

An integrated In-Silico-In-Chemico-In-Vitro (iSiCiV) Approach to identify biomarkers to predict the skin sensitisation potential of phytochemicals

BackgroundSkin sensitisation is one of the important regulatory endpoints used to determine whether a given test chemical can elicit an allergic response in susceptible individuals. The in-vitro approaches for predicting skin sensitisation potential for chemicals have been well-developed and widely accepted. Continuous efforts are being made to identify novel solutions to overcome the limitations of the existing approaches. PurposeThe herbal preparations used in dermal therapeutics are frequently found to elicit skin sensitisation, requiring caution. The herbal preparations tend to contain more than one phytochemical. The expected active ingredient may not possess skin sensitisation. However, the other phytochemical traces could pose synergistic and potentiating effects towards enhanced/suppressed skin sensitisation. In this regard, isolation, purification, yield, and in-vitro / in-vivo evaluation of skin sensitisation for all the traces would be time, cost, and animal-consuming. Hence, the regulatory agencies appreciate using computational tools to substantiate in-vitro methods. The regulators have also endorsed using in-silico approaches like quantitative structure-activity relationships (QSAR) and read-across. These approaches are limited to structure-based predictions and in combination with system-level approaches, which comprehensively facilitate a better understanding of complex biological systems. Study designIn the present study, 603 phytochemicals were screened in-silico using OECD QSAR Toolbox. The limitations of the QSAR Toolbox in predicting skin sensitisers were identified. MethodThe systems biology-based approach to complement the QSAR-based prediction was developed, and 41 biomarkers were identified as a determinant of skin sensitisation. The molecules attributed to these biomarkers were predicted as skin sensitisers. ResultsAs a result, 31 phytochemicals were predicted as skin sensitisers. Acrolein, eugenol, formaldehyde, and glycerol were selected for experimental validation of the in-silico predictions using direct peptide reactivity assay (DPRA), KeratinoSens™, and h-CLAT assays representing each key event in skin sensitisation. Further, identified biomarkers were validated in THP-1 and KeratinoSens cell lines using qRT-PCR. Among all genes, IL-8 and CCL4 were upregulated by 6.4 and 5.6 folds, respectively, in THP-1, while AKR1C2 and AKR1C3 were upregulated by 7.8 and 11.1 folds, respectively, in KeratinoSens. IL-8 level was analysed by ELISA and found to be in line with qRT-PCR results. ConclusionThe study proposes a possible strategy based on the “iSiCiV” approach to enhance the prediction of the dermal sensitisation potential of a test compound. The approach offers a robust platform and highlights an opportunity to shift the paradigm of skin sensitisation assay with a better understanding of toxicology mechanisms.

Possible effects of ancestry-related oxysterol-binding protein-like 10 genetic polymorphisms on dengue virus replication and anti-dengue immune response

PurposeOxysterol-binding protein-like 10 (OSBPL10) gene has been associated with reduced susceptibility to severe dengue in individuals of African descent. The aim of this study was to determine the possible effect of OSBPL10 on dengue virus (DENV) replication as well as the impact of African and European haplotypes of six OSBPL10 small nuclear polymorphisms (SNPs) on dengue multiplication and innate immune response. MethodsWe conducted gene knockdown experiments targeting OSBPL10 in THP-1 and Huh-7D12 cell lines, followed by a DENV-2 replication assay. Extracellular viral load was determined using qRT-PCR. To investigate the impact of SNPs haplotypes on viral replication and gene expression we cultured peripheral blood mononuclear cells (PBMC) from individuals with homozygous African and European haplotypes of OSBPL10 with DENV-2. Individual genotyping was performed using High Resolution Melt (HRM) analysis. The level of viral replication was assessed through plaque assay, while RT-PCR was employed to determine the expression levels of RXR-α, IFN-γ, IL-10 and IL-8 genes. ResultsIn vitro OSBPL10 knockdown significantly reduced DENV-2 replication. Individuals carrying European haplotypes showed higher DENV titers along with elevated levels of RXR-α and IL-8 mRNA compared to those carrying African haplotypes, who exhibited lower viral loads alongside increased IFN-γ and IL-10 expression. ConclusionsOur findings further explore the role of OSBPL10 in DENV multiplication, immune response to infection. The European haplotypes of OSBPL10 appear to increase DENV replication and promote RXR-α and IL-8 mRNA expression which correlates with the suppressive effect of these mediators on type I IFN, promoting viral replication and a deficient antiviral response. In contrast, the African haplotype showed a reduction in DENV replication and enhanced IFN-γ and IL-10 mRNA expression, which could be related to the better management of dengue infection and the low frequency of severe disease in this ethnic groupe.

A pancreatic cancer organoid incorporating macrophages reveals the correlation between the diversity of tumor-associated macrophages and cancer cell survival

Pancreatic ductal adenocarcinoma (PDAC) is a progressive cancer with a poor prognosis. It contains a complex tumor microenvironment (TME) that includes various stromal cell types. Comprehending cellular communications within the TME is difficult due to a lack of research models that can recapitulate human PDAC-TME. Previously, we recapitulated, in part, the PDAC-TME containing a diversity of cancer-associated fibroblasts (CAFs) in vitro. This was done by establishing a PDAC organoid by co-culturing patient-derived cancer cells with human induced pluripotent stem cell (hiPSC)-derived mesenchymal and endothelial cells, which was designated the fused pancreatic cancer organoid (FPCO). We further incorporated macrophages derived from the THP-1 cell line, which are the source of tumor-associated macrophages (TAMs), a major TME component, into FPCO, which was designated M0-FPCO. Bulk RNA sequencing (RNAseq) analysis revealed that macrophages in M0-FPCO (FPCO-Mac) lost their pro-inflammatory features but acquired pro-angiogenic features. Consistently, the formation of an endothelial cell network was enhanced in M0-FPCO. Single-cell RNA-seq (scRNA-seq) analysis revealed that M0-FPCO contained five TAM subpopulations similar to the corresponding TAM in human PDAC tissue in the integrated analysis, including SPP1+-TAM, which has been correlated with tumor angiogenesis and cell proliferation. Focusing on PDAC cells, we found that they could survive longer within the organoid in the presence of TAM. Consistent with the prolonged proliferation and survival of PDAC cells, PDAC subclusters were characterized by proliferative features, such as increased M0-FPCO. Therefore, by establishing a PDAC organoid with macrophages, we recapitulated the diversity of TAMs and identified the role of TAM in endothelial network formation as well as in the modulation of PDAC cell properties. SignificancePDAC organoids, including macrophages using hiPSC, showed that PDAC-TAM has angiogenic features and contributes to PDAC cell survival.

Preliminary Assessment of the In Vitro Immunological Activity of Native Lactobacillus Strains from the Ecuadorian Amazon

The isolation and characterization of new Lactobacillus strains from fermented traditional foods is a global trend as it enhances the potential for discovering novel probiotic foods. The fermented cocoa (Theobroma cacao) mucilage is an intriguing substrate for isolating lactic acid bacteria involved in fermentation. This study represents a preliminary investigation into the isolation, quantification, characterization, and immunological activity of Lactobacillus strains derived from cocoa beans sampled from two farmer associations (Kallari and Wiñak) in the Ecuadorian Amazon region. A mother culture was prepared using fermented cocoa pulp, cultivated on selective MRS media. After growth, the isolates were morphologically characterized. A significantly higher bacterial concentration was recorded in Kallari Association samples if compared with Wiñac. A total of 25 strains were isolated, eight of which were rod-shaped and positive to catalase tests and were characterized as Lactobacillus. In vitro, immunological activity was performed on differentiated THP-1 cell lines. Cells were treated with bacterial concentrates, and immunological activity was determined through interleukin-10 expression. Results indicated that W6 strain showed the highest immunological activity. These results indicated that Lactobacillus strains isolated from fermented cocoa pulp in the Ecuadorian Amazon show promise as a new source of probiotics. Keywords: cocoa; lactic acid bacteria; biological activity; isolation; cocoa fermentation; probiotics; cocoa pulp

Towards characterization of cell culture conditions for reliable proteomic analysis: in vitro studies on A549, differentiated THP-1, and NR8383 cell lines

Proteomic investigations result in high dimensional datasets, but integration or comparison of different studies is hampered by high variances due to different experimental setups. In addition, cell culture conditions can have a huge impact on the outcome. This study systematically investigates the impact of experimental parameters on the proteomic profiles of commonly used cell lines—A549, differentiated THP-1 macrophage-like cells, and NR8383—for toxicity studies. The work focuses on analyzing the influence at the proteome level of cell culture setup involving different vessels, cell passage numbers, and post-differentiation harvesting time, aiming to improve the reliability of proteomic analyses for hazard assessment. Mass-spectrometry-based proteomics was utilized for accurate protein quantification by means of a label-free approach. Our results showed that significant proteome variations occur when cells are cultivated under different setups. Further analysis of these variations revealed their association to specific cellular pathways related to protein misfolding, oxidative stress, and proteasome activity. Conversely, the influence of cell passage numbers on the proteome is minor, suggesting a reliable range for conducting reproducible biological replicates. Notable, substantial proteome alterations occur over-time post-differentiation of dTHP-1 cells, particularly impacting pathways crucial for macrophage function. This finding is key for the interpretation of experimental results. These results highlight the need for standardized culture conditions in proteomic-based evaluations of treatment effects to ensure reliable results, a prerequisite for achieving regulatory acceptance of proteomics data.

Cytokine secretion by in vitro cultures of lung epithelial cells, differentiated macrophages and differentiated dendritic cells incubated with pneumococci and pneumococcal extracellular vesicles.

Streptococcus pneumoniae is an important human pathogen that can colonize the respiratory tract of healthy individuals. The respiratory tract mucosa is thus the first barrier for this pathogen. In this study, we have tested three models of the respiratory epithelium with immune cells: (i) monolayer of A549 human lung epithelial cells, (ii) A549 + macrophages differentiated from the human monocytic THP-1 cell line (dMφ) and (iii) A549 + dMφ + dendritic cells differentiated from THP-1 (dDC) using a two-chamber system. Pneumococcal strains Rx1 (non-encapsulated) and BHN418 (serotype 6B) were incubated with the cells and secretion of IL-6, IL-8, IL-1β, TNF-α and IL-10 was evaluated. Overall, the models using co-cultures of A549 + dMφ and A549 + dMφ + dDC elicited higher levels of pro-inflammatory cytokines and the non-encapsulated strain elicited an earlier cytokine response. BHN418 pspA (pneumococcal surface protein A) and pspC (pneumococcal surface protein C) knockouts elicited similar cytokine secretion in the co-culture models, whereas BHN18 ply (pneumolysin) knockout induced much lower levels. The results are in accordance with the activation of the inflammasome by Ply. Finally, we evaluated pneumococcal extracellular vesicles (pEVs) in the co-culture models and observed secretion of pro-inflammatory cytokines in the absence of cytotoxicity. Since pEVs are being studied as vaccine candidate against pneumococcal infections, the co-cultures of A549 + dMφ and A549 + dMφ + dDC are simple models that could be used to evaluate pEV vaccine batches.

Investigating the Impact of IL-6 and CXCL8 on Neurodegeneration and Cognitive Decline in Alzheimer's Disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder primarily affecting the elderly, characterized by severe cognitive impairment and memory loss. Emerging evidence suggests that neuroinflammation plays a significant role in AD pathogenesis, with cytokines like interleukin-6 (IL-6) and C-X-C motif chemokine ligand 8 (CXCL8) contributing to the disease progression. We utilized GEO datasets to identify IL-6 and CXCL8 as pivotal inflammatory markers in AD. In vitro experiments were conducted using SK-N-BE(2)-M17 and THP-1 cell lines treated with IL-6 and CXCL8 to model AD. Additionally, in vivo tests on Amyloid Precursor Protein/Presenilin 1 (APP/PS1) AD mouse models were performed to assess the impact of these cytokines on cognitive functions and brain pathology. The results indicated a significant decrease in cell viability, increased apoptosis, and elevated inflammatory factor secretion following IL-6 and CXCL8 treatment in vitro. In vivo, AD mouse models treated with these cytokines exhibited exacerbated emotional distress, decreased social interaction, impaired cognitive functions, and increased amyloid protein deposition in neural tissues. The study highlights the detrimental effects of IL-6 and CXCL8 on neuronal health and cognitive functions in AD. These findings suggest that targeting these cytokines could offer potential therapeutic interventions for improving patient outcomes in Alzheimer's disease.

Impact of Spent Mushroom Substrate Combined with Hydroponic Leafy Vegetable Roots on Pleurotus citrinopileatus Productivity and Fruit Bodies Biological Properties.

Agricultural activities produce large quantities of organic byproducts and waste rich in lignocellulosic materials, which are not sufficiently utilized. In this study, alternative agricultural waste products, namely, spent mushroom substrate (SMS) from the cultivation of edible Pleurotus ostreatus mushrooms and the roots of leafy vegetables from hydroponic cultivation (HRL), were evaluated for their potential to be used as substrates for the cultivation of Pleurotus citrinopileatus and their effects on the quality, the nutritional value, the chemical properties (lipid, protein, carbohydrate, ash, fatty acid and carbohydrate composition) and the bioactive content (total phenolic compounds and antioxidant activity) of produced mushrooms. SMS and HRL (in different ratios with and without additives) and wheat straw with additives (WS-control) were used. During incubation, the linear growth rate of the mycelium (Kr, mm/day) was measured and used for screening. Mushroom cultivation took place in bags, where several characteristics were examined: earliness (duration between the day of substrate inoculation and the day of first harvest) and biological efficiency (B.E. %, the ratio of the weight of fresh mushrooms produced per dry weight of the substrate × 100). Furthermore, this study aimed to investigate the effect of the protein extract (PE) and carbohydrate extract (CE) of P. citrinopileatus after in vitro digestion (fraction less than 3kDa: PE-DP-3; digestate fraction: CE-D, respectively) on the expression of antioxidant-related genes in the THP-1 cell line. The results showed that mushrooms grown on SMS 50%-HRL 40% had the fastest growth (6.1 mm/d) and the highest protein and lipid contents (34.7% d.w.; 5.1% d.w.). The highest B.E. (73.5%), total carbohydrate (65.7%) and total phenolic compound (60.2 mg GAE/g d.w.) values were recorded on the control substrate. Antioxidant activity was observed in all extracts; the total flavonoid content was low in the samples, and the maximum total triterpene value was detected in SMS 80%-HRL 20% (9.8 mg UA/g d.w.). In all mushrooms, linoleic acid (C18:2) was the main fatty acid (above 60%), and fructose was the dominant individual saccharide. In the investigation of the regulation pathway, NFE2L2 gene expression was upregulated only in the SMS 60%-HRL 40% intervention during incubation with CE-D samples. Additionally, the transcription levels of antioxidant-related genes, SOD1, CAT, HMOX1 and GSR, were increased in the SMS 60-30% intervention. Compared to WS, the alternative substrates are observed to trigger a pathway concerning CE that may resist oxidative stress. This study supports the utilization of agricultural byproducts through sustainable and environmentally friendly practices while simultaneously producing high-value-added products such as mushrooms. Therefore, alternative substrates, particularly those containing HRL, could serve as natural sources of antioxidant potential.

Effects of Three Retrograde Filling Materials on Production of Inflammatory Cytokines and Resorbing Mediators

: Root-end filling materials are typically used following endodontic surgeries, and due to their different chemical compositions, they have varying effects on the survival and differentiation of tissues around the root apex. The purpose of this in vitro study was to evaluate the effects of three retrograde filling materials—ProRoot MTA, Super ethoxy benzoic acid (EBA), and Geristore—on the production of pro-inflammatory and resorptive cytokines (RANK, RANK-L, OPG, IL-1β, and TNF-α). After mixing and setting, the experimental materials were exposed to UV rays and then applied to the prepared cells. This study used osteoblast (MG-63) and THP-1 monocyte cell lines, as well as peripheral blood monocytes (PBM). The evaluation times for RANK-L and OPG were 24 and 48 hours, while IL-1β and tumor necrosis factor (TNF-α) were evaluated at 24 hours. The RANK levels were measured using flow cytometry, and monocyte survival was assessed at different times using the MTT assay. The results were analyzed using Minitab statistical software, which indicated that the type of material significantly affected cytokine production. Exposure of monocytes to EBA resulted in a decrease in TNF-α and IL-1β secretion compared to the other groups. However, in the THP-1 cell line, there was no significant difference in the release of pro-inflammatory cytokines between the groups. Conversely, Geristore induced the highest level of RANK-L and the lowest level of OPG in cultures with the MG-63 cell line. According to the MTT assay, the viability order of the materials from early to late was Geristore &lt; EBA &lt; MTA.