Effects of elafibranor on liver fibrosis and gut barrier function in a mouse model of alcohol-associated liver disease.

BACKGROUND: Alcohol-associated liver disease (ALD) is a leading cause of liver-related morbidity and mortality, but there are no therapeutic targets and modalities to prevent ALD-related liver fibrosis. Peroxisome proliferator activated receptor (PPAR) α and δ play a key role in lipid metabolism and intestinal barrier homeostasis, which are major contributors to the pathological progression of ALD. Meanwhile, elafibranor (EFN), which is a dual PPARα and PPARδ agonist, has reached a phase III clinical trial for the treatment of metabolic dysfunction-associated steatotic liver disease and primary biliary cholangitis. However, the benefits of EFN for ALD treatment is unknown. AIM: To evaluate the inhibitory effects of EFN on liver fibrosis and gut-intestinal barrier dysfunction in an ALD mouse model. METHODS: ALD-related liver fibrosis was induced in female C57BL/6J mice by feeding a 2.5% ethanol (EtOH)-containing Lieber-DeCarli liquid diet and intraperitoneally injecting carbon tetrachloride thrice weekly (1 mL/kg) for 8 weeks. EFN (3 and 10 mg/kg/day) was orally administered during the experimental period. Histological and molecular analyses were performed to assess the effect of EFN on steatohepatitis, fibrosis, and intestinal barrier integrity. The EFN effects on HepG2 lipotoxicity and Caco-2 barrier function were evaluated by cell-based assays. RESULTS: The hepatic steatosis, apoptosis, and fibrosis in the ALD mice model were significantly attenuated by EFN treatment. EFN promoted lipolysis and ß-oxidation and enhanced autophagic and antioxidant capacities in EtOH-stimulated HepG2 cells, primarily through PPARα activation. Moreover, EFN inhibited the Kupffer cell-mediated inflammatory response, with blunted hepatic exposure to lipopolysaccharide (LPS) and toll like receptor 4 (TLR4)/nuclear factor kappa B (NF-κB) signaling. EFN improved intestinal hyperpermeability by restoring tight junction proteins and autophagy and by inhibiting apoptosis and proinflammatory responses. The protective effect on intestinal barrier function in the EtOH-stimulated Caco-2 cells was predominantly mediated by PPARδ activation. CONCLUSION: EFN reduced ALD-related fibrosis by inhibiting lipid accumulation and apoptosis, enhancing hepatocyte autophagic and antioxidant capacities, and suppressing LPS/TLR4/NF-κB-mediated inflammatory responses by restoring intestinal barrier function.

Thiolated chitosan nanoparticles encapsulated nisin and selenium: antimicrobial/antibiofilm/anti-attachment/immunomodulatory multi-functional agent.

BACKGROUND: The increase in the resistance of bacterial strains to antibiotics has led to research into the bactericidal potential of non-antibiotic compounds. This study aimed to evaluate in vitro antibacterial/ antibiofilm properties of nisin and selenium encapsulated in thiolated chitosan nanoparticles (N/Se@TCsNPs) against prevalent enteric pathogens including standard isolates of Vibrio (V.) cholerae O1 El Tor ATCC 14,035, Campylobacter (C.) jejuni ATCC 29,428, Salmonella (S.) enterica subsp. enterica ATCC 19,430, Shigella (S.) dysenteriae PTCC 1188, Escherichia (E.) coli O157:H7 ATCC 25,922, Listeria (L.) monocytogenes ATCC 19,115, and Staphylococcus (S.) aureus ATCC 29,733. METHODS: The synthesis and comprehensive analysis of N/Se@TCsNPs have been completed. Antibacterial and antibiofilm capabilities of N/Se@TCsNPs were evaluated through broth microdilution and crystal violet assays. Furthermore, the study included examining the cytotoxic effects on Caco-2 cells and exploring the immunomodulatory effects of N/Se@TCsNPs. This included assessing the levels of both pro-inflammatory (IL-6 and TNFα) and anti-inflammatory (IL-10 and TGFß) cytokines and determining the gene expression of TLR2 and TLR4. RESULTS: The N/Se@TCsNPs showed an average diameter of 136.26 ± 43.17 nm and a zeta potential of 0.27 ± 0.07 mV. FTIR spectroscopy validated the structural features of N/Se@TCsNPs. Scanning electron microscopy (SEM) images confirmed their spherical shape and uniform distribution. Thermogravimetric Analysis (TGA)/Differential Scanning Calorimetry (DSC) tests demonstrated the thermal stability of N/Se@TCsNPs, showing minimal weight loss of 0.03%±0.06 up to 80 °C. The prepared N/Se@TCsNPs showed a thiol content of 512.66 ± 7.33 µmol/g (p < 0.05), an encapsulation efficiency (EE) of 69.83%±0.04 (p ≤ 0.001), and a drug release rate of 74.32%±3.45 at pH = 7.2 (p ≤ 0.004). The synthesized nanostructure demonstrated potent antibacterial activity against various isolates, with effective concentrations ranging from 1.5 ± 0.08 to 25 ± 4.04 mg/mL. The ability of N/Se@TCsNPs to reduce bacterial adhesion and internalization in Caco-2 cells underscored their antibiofilm properties (p ≤ 0.0001). Immunological studies indicated that treatment with N/Se@TCsNPs led to decreased levels of inflammatory cytokines IL-6 (14.33 ± 2.33 pg/mL) and TNFα (25 ± 0.5 pg/mL) (p ≤ 0.0001), alongside increased levels of anti-inflammatory cytokines IL-10 (46.00 ± 0.57 pg/mL) and TGFß (42.58 ± 2.10 pg/mL) in infected Caco-2 cells (p ≤ 0.0001). Moreover, N/Se@TCsNPs significantly reduced the expression of TLR2 (0.22 ± 0.09) and TLR4 (0.16 ± 0.05) (p < 0.0001). CONCLUSION: In conclusion, N/Se@TCsNPs exhibited significant antibacterial/antibiofilm/anti-attachment/immunomodulatory effectiveness against selected Gram-positive and Gram-negative enteric pathogens. However, additional ex-vivo and in-vivo investigations are needed to fully assess the performance of nanostructured N/Se@TCsNPs.

Ambient Particulate Matter Induces In Vitro Toxicity to Intestinal Epithelial Cells without Exacerbating Acute Colitis Induced by Dextran Sodium Sulfate or 2,4,6-Trinitrobenzenesulfonic Acid.

Inflammatory bowel disease (IBD) is an immunologically complex disorder involving genetic, microbial, and environmental risk factors. Its global burden has continued to rise since industrialization, with epidemiological studies suggesting that ambient particulate matter (PM) in air pollution could be a contributing factor. Prior animal studies have shown that oral PM10 exposure promotes intestinal inflammation in a genetic IBD model and that PM2.5 inhalation exposure can increase intestinal levels of pro-inflammatory cytokines. PM10 and PM2.5 include ultrafine particles (UFP), which have an aerodynamic diameter of <0.10 µm and biophysical and biochemical properties that promote toxicity. UFP inhalation, however, has not been previously studied in the context of murine models of IBD. Here, we demonstrated that ambient PM is toxic to cultured Caco-2 intestinal epithelial cells and examined whether UFP inhalation affected acute colitis induced by dextran sodium sulfate and 2,4,6-trinitrobenzenesulfonic acid. C57BL/6J mice were exposed to filtered air (FA) or various types of ambient PM reaerosolized in the ultrafine size range at ~300 µg/m3, 6 h/day, 3-5 days/week, starting 7-10 days before disease induction. No differences in weight change, clinical disease activity, or histology were observed between the PM and FA-exposed groups. In conclusion, UFP inhalation exposure did not exacerbate intestinal inflammation in acute, chemically-induced colitis models.

Intestinal Epithelial Co-Culture Sensitivity to Pro-Inflammatory Stimuli and Polyphenols Is Medium-Independent.

The complexification of in vitro models requires the compatibility of cells with the same medium. Since immune cells are the most sensitive to growth conditions, growing intestinal epithelial cells in their usual medium seems to be necessary. This work was aimed at comparing the sensitivity of these epithelial cells to pro-inflammatory stimuli but also to dietary polyphenols in both DMEM and RPMI-1640 media. Co-cultures of Caco-2 and HT29-MTX cells were grown for 21 days in the two media before their stimulation with a cocktail of TNF-α (20 ng/mL), IL-1ß (1 ng/mL), and IFN-γ (10 ng/mL) or with LPS (10 ng/mL) from E. coli (O111:B4). The role of catechins (15 µM), a dietary polyphenol, was evaluated after its incubation with the cells before their stimulation for 6 h. The RPMI-1640 medium did not alter the intensity of the inflammatory response observed with the cytokines. By contrast, LPS failed to stimulate the co-culture in inserts regardless of the medium used. Lastly, catechins were unable to prevent the pro-inflammatory response observed with the cytokines in the two media. The preservation of the response of this model of intestinal epithelium in RPMI-1640 medium is promising when considering its complexification to evaluate the complex cellular crosstalk leading to intestinal homeostasis.

The Use of Polysaccharide AOP30 from the Rhizome of Alpinia officinarum Hance to Alleviate Lipopolysaccharide-Induced Intestinal Epithelial Barrier Dysfunction and Inflammation via the TLR4/NfκB Signaling Pathway in Caco-2 Cell Monolayers.

Alpinia officinarum Hance is rich in carbohydrates and is flavored by natives. The polysaccharide fraction 30 is purified from the rhizome of A. officinarum Hance (AOP30) and shows excellent immunoregulatory ability when administered to regulate immunity. However, the effect of AOP30 on the intestinal epithelial barrier is not well understood. Therefore, the aim of this study is to investigate the protective effect of AOP30 on the intestinal epithelial barrier using a lipopolysaccharide (LPS)-induced intestinal epithelial barrier dysfunction model and further explore its underlying mechanisms. Cytotoxicity, transepithelial electrical resistance (TEER) values, and Fluorescein isothiocyanate (FITC)-dextran flux are measured. Simultaneously, the protein and mRNA levels of tight junction (TJ) proteins, including zonula occludens-1 (ZO-1), Occludin, and Claudin-1, are determined using Western blotting and reverse-transcription quantitative polymerase chain reaction methods, respectively. The results indicate that AOP30 restores the LPS-induced decrease in the TEER value and cell viability. Furthermore, it increases the mRNA and protein expression of ZO-1, Occludin, and Claudin-1. Notably, ZO-1 is the primary tight junction protein altered in response to LPS-induced intestinal epithelial dysfunction. Additionally, AOP30 downregulates the production of TNFα via the Toll-like receptor 4 (TLR4)/NF-κB signaling pathway. Collectively, the findings of this study indicate that AOP30 can be developed as a functional food ingredient or natural therapeutic agent for addressing intestinal epithelial barrier dysfunction. It sheds light on the role of AOP30 in improving intestinal epithelial function.

Pasteurized form of a potential probiotic lactobacillus brevis IBRC-M10790 exerts anti-inflammatory effects on inflammatory bowel disease in vitro.

BACKGROUND: Inflammatory bowel disease (IBD) is a chronic, relapsing inflammatory disorder of the gastrointestinal system. So far, no treatment has been identified that can completely cure IBD. Lactobacillus brevis is hypothesized to be beneficial in preventing inflammation. This study aimed to evaluate the potential probiotic effects of live and pasteurized L. brevis IBRC-M10790 on the in vitro cell co-culture model of IBD. METHODS: An in vitro intestinal model was established using a transwell co-culture system of Caco-2 intestinal epithelial cells and RAW264.7 macrophages. Inflammatory conditions were induced in RAW264.7 cells using lipopolysaccharide. The effects of live and pasteurized L. brevis IBRC-M10790 on inflammatory mediators and epithelial barrier markers were investigated. RESULTS: L. brevis IBRC-M10790 was able to significantly decrease the proinflammatory cytokines (IL-6, IL-1ß, and TNF-α) and increase the anti-inflammatory cytokine (IL-10) in the in vitro co-culture system. In addition, L. brevis increased adherens and tight junction (TJ) markers (ZO-1, E-cadherin, and Occludin) in Caco-2 intestinal epithelial cells. Based on the results, pasteurized L. brevis showed a higher protective effect than live L. brevis. CONCLUSIONS: Our findings suggest that live and pasteurized forms of L. brevis possess probiotic properties and can mitigate inflammatory conditions in IBD.

Pigmented corn as a gluten-free source of polyphenols with anti-inflammatory and antioxidant properties in CaCo-2 cells.

A high number of varieties from corn (Zea mays L.) have been consumed for long time all over the world, however pigmented varieties are recently gaining renewed attention due to their beneficial effects and polyphenolic content. The natural lack of gluten makes corn suitable for consumption by celiac population, who need to control their inflammatory state through an appropriate gluten-free diet. The biological effects of polyphenols from pigmented corn are poorly investigated in the context of celiac disease. In this work, we analyzed through HPLC-DAD the phenolic composition of two Italian purple and red varieties ("Scagliolo Rosso" and "Rostrato di Rovetta", respectively) comparing their effects in human intestinal epithelial cells (CaCo-2 cells). The possible impact of gastro-intestinal digestion following oral consumption was assessed as well. The phenolic profile showed the presence of phenolic acids in both varieties, while anthocyanins were identified in Scagliolo Rosso only. After simulated digestion, the level of polyphenols did not significantly change and paralleled with an increased scavenging activity. In CaCo-2 cells, stimulated by a proinflammatory cocktail containing gliadin-derived peptides (IL-1ß, IFN-γ, digested gliadin), pigmented corn extracts inhibited the release of CXCL-10 and sICAM-1, with mechanisms partially ascribed to NF-κB impairment. At the same concentration (200 µg/mL), ROS production and catalase depletion were reverted through Nrf-2-independent mechanisms. Our data suggest that polyphenols from pigmented corns might help in controlling the inflammatory and oxidative state of people with celiac disease at intestinal level, at concentrations potentially achievable through a gluten-free diet.

Ca2+-nano starch-lutein endowed 3D printed surimi with antioxidation and mutual reinforcing transmembrane transport mechanisms via hepg2 and caco-2 cells model.

To better enhance printing effects meanwhile casting functionality, antioxidation and absorption of bioactive component in printed Ca2+-nano starch (NS)-lutein (L)-surimi were investigated. Results shown that Ca2+-NS-L promoted surimi printability due to enhanced gel strength and denser structure. Mixing Ca2+-NS-L endowed printed surimi with antioxidation (DPPH, ABTS, hydroxyl radical, Fe2+ reduction were 42 %, 79 %, 65 %, 0.104 mg·mL-1, respectively) due to the ability of lutein with more -OH groups and conjugate bonds to capture free radicals. It also manifested in cellular antioxidation that Ca2+-NS-L-surimi regulated the level of Nrf2 to protect gene expression of antioxidases (SOD, CAT, GSH-Px increased by 30-180 %, compared to damaged cells) through keap1-Nrf2-ARE pathway. Additionally, lutein absorption and transportation of Ca2+-NS-L-surimi increased by 20 %, compared to NS-L. Possibly, combination of samples and membrane was facilitated by surface hydrophobic, promoting endocytosis. Meanwhile, digestive surimi (peptides) with acidic-alkaline amino acids and negative charges made samples be attracted and moved in bypass parts under electrostatic traction and repulsion (electrostatic domain) to promote transport process. Also, Ca2+ facilitated CaM expression in membrane and formed Ca2+ channel by combining with CaM to accelerate entry of samples into cells. Conclusively, Ca2+-NS-L both strengthened printability of surimi and antioxidation, promoting application of printed functional surimi.

Chitosan as an Antimicrobial, Anti-Insect, and Growth-Promoting Agent for Potato (Solanum tuberosum L.) Plants.

A growing trend in plant protection is replacing chemical preparations with environmentally friendly biological compositions. Chitosan, due to its biocompatibility, biodegradability, and bioactivity, is an effective agent against plant diseases. The purpose of the study was to evaluate chitosan as a potential biopesticide for potato plants. Three variants of chitosan were tested: high (310-375 kDa, >75% deacetylated), medium (190-310 kDa, 75-85% deacetylated), and low (50-190 kDa, 75-85% deacetylated) molecular weight. The chitosan variants were dissolved in lactic and succinic acids and tested for antibacterial and antifungal properties against eight strains of mould and two strains of bacteria responsible for potato diseases. The possible cytotoxicity of chitosan was evaluated against different cell lines: insect Sf-9, human keratinocyte HaCaT, and human colon carcinoma Caco-2. The bioprotective activities of the chitosan were also evaluated in situ on potato tubers. Chitosan inhibited the growth of almost all the selected phytopathogens. The most active was medium molecular chitosan in lactic acid. This formula was characterized by low toxicity towards human cells and high toxicity towards Sf-9 cells. It was also found to have positive effects on the growth of stems and roots, gas exchange, and chlorophyll index in potato plants. Selected chitosan formulation was proposed as a functional biopesticide for potato protection against phytopathogens.

Probiotic evaluation, adherence capability and safety assessment of Lactococcus lactis strain isolated from an important herb "Murraya koenigii".

Lactic acid bacteria (LAB) isolated from medicinal herb Murraya koenigii, commonly known as curry leaf, which promotes the growth and maintenance of gut microbiota, were studied for their probiotic potential. The key objective of this research was to isolate and evaluate probiotic characteristics, test adherence capabilities, and confirm their safety. Lactococcus lactis (MKL8), isolated from Murraya koenigii, was subjected to in vitro analysis to assess its resistance to the gastric environment, ability to adhere Caco-2 cells, anti-microbial activity, hydrophobicity, auto-aggregation, and safety profiling through MTT assay and hemolytic. MKL8 exhibited growth at 0.5% phenol concentrations (> 80%) and was able to survive in conditions with high bile concentrations (> 79%) and a relatively low pH (72%-91%). It shows high tolerance to high osmotic conditions (> 73%) and simulated gastric juice (> 72%). Additionally, MKL8 demonstrated strong hydrophobicity (85%), auto-aggregation (87.3%-91.7%), and adherence to Caco-2 cells. Moreover, it had an inhibitory effect against pathogens too. By performing the hemolytic and MTT assays, the non-toxicity of MKL8 isolate was examined, and it exhibited no harmful characteristics. Considering MKL8's resistance to gastrointestinal tract conditions, high surface hydrophobicity, non-toxicity, and ability to inhibit the tested pathogens, it can be concluded that MKL8 demonstrated promising probiotic properties and has potential for use in the food industry.

Amifostine loaded lipid-calcium carbonate nanoparticles as an oral drug delivery system for radiation protection.

Amifostine (AMF) as the first-line radiation protection drug, usually suffered from low compliance and short half-life upon clinical applications. The development of oral drug delivery system (DDS) for AMF is a promising solution. However, the inherent shortages of AMF present significant challenges in the design of suitable oral DDS. Here in this study, we utilized the ability of calcium ions to bind with AMF and prepared AMF loaded calcium carbonate (CC) core, CC/AMF, using phase transferred coprecipitation method. We further modified the CC/AMF using phospholipids to prepare AMF loaded lipid-calcium carbonate (LCC) hybrid nanoparticles (LCC/AMF) via a thin-film dispersion method. LCC/AMF combines the oral advantages of lipid nanoparticles with the drug-loading capabilities of CC, which was shown as uniform nano-sized formulation with decent stability in aqueous solution. With favorable intestinal transport and absorption effects, it effectively enhances the in vivo radiation protection efficacy of AMF through oral administration. More importantly, we further investigated the cellular accumulation profile and intracellular transport mechanism of LCC/AMF using MDCK and Caco-2 cell lines as models. This research not only alters the current administration method of AMF to enhance its convenience and compliance, but also provides insights and guidance for the development of more suitable oral DDS for AMF in the future.

Digested casein phosphopeptides impact intestinal calcium transport in vitro.

Calcium is the most abundant mineral in the human body and is involved in critical physiological and cellular processes. It is essential for the development, maintenance, and integrity of bone tissue throughout life. Identifying new natural food-grade chelating agents to improve calcium uptake is of increasing interest. Casein phosphopeptides (CPPs), highly phosphorylated peptides obtained after enzymatic hydrolysis of caseins, represent promising calcium-chelating candidates. The aim of this study was to investigate, using cell culture models, the ability of a digested milk matrix enriched in CPPs to regulate calcium transport through the intestinal barrier and elucidate the involved mechanisms. To this end, a CPP-preparation underwent in vitro static digestion and was subsequently incubated with an intestinal barrier model to monitor calcium uptake and transport. Our results demonstrated that the digested CPP preparation enhanced the trans-epithelial calcium transport via paracellular pathways and that CPPs, identified by peptidomics, crossed the intestinal barrier in the same time.

Potential bioaccessibility and bioavailability of polyphenols and functional properties of tiger nut beverage and its by-product during in vitro digestion.

"Horchata de chufa" is a beverage produced from tiger nut tubers, which yields a high amount of by-product. This study explored the functional properties of the Spanish tiger nut beverage (TNB) and its by-product (TNBP) together with the bioaccessibility and bioavailability of polyphenols in vitro. TNB and TNBP were characterized for polyphenols via LC/MS/MS and underwent in vitro digestion (INFOGEST). The total antioxidant capacity (TAC) of all bioaccessible fractions and digestion residues was assessed. Intestinal bioaccessible fractions were tested for the ability to inhibit the activity of digestive enzymes (α-amylase, α-glucosidase, and lipase) and the content of polyphenols, whose bioavailability was assessed in a Caco-2 cell model. Thirteen polyphenols were quantified and found to be more abundant in TNB (603 ± 1.4 µg g-1 DW) than in TNBP (187 ± 1.0 µg g-1 DW). Polyphenol bioaccessibility was higher for TNBP than that for TNB (57% vs. 27%), and despite a similar TAC of the intestinal bioaccessible fractions (10.2 ± 0.1 µmoL vs. 9.2 ± 0.03 µmoL eq. Trolox per g DW for TNB and TNBP, respectively), the different patterns of polyphenols released upon digestion suggested the higher ability of TNBP fraction to inhibit α-glucosidase and lipase. TNBP digestion residue showed higher TAC than TNB. Moreover, TNB polyphenols exhibited over 80% bioavailability, whereas TNBP polyphenols' bioavailability ranged from 62% to 84%. Overall, the findings demonstrated that TNBP maintains a high nutritional value, thus suggesting its possible reuse in innovative, healthy, and sustainable foods.

AMPK Deficiency Increases DNA Methylation and Aggravates Colorectal Tumorigenesis in AOM/DSS Mice.

The incidence of colorectal cancer (CRC) is closely linked to metabolic diseases. Accumulating evidence suggests the regulatory role of AMP-activated protein kinase (AMPK) in cancer metabolic reprogramming. In this study, wild-type and AMPK knockout mice were subjected to azoxymethane-induced and dextran sulfate sodium (AOM/DSS)-promoted colitis-associated CRC induction. A stable AMPK-deficient Caco-2 cell line was also established for the mechanistic studies. The data showed that AMPK deficiency accelerated CRC development, characterized by increased tumor number, tumor size, and hyperplasia in AOM/DSS-treated mice. The aggravated colorectal tumorigenesis resulting from AMPK ablation was associated with reduced α-ketoglutarate production and ten-eleven translocation hydroxylase 2 (TET2) transcription, correlated with the reduced mismatch repair protein mutL homolog 1 (MLH1) protein. Furthermore, in AMPK-deficient Caco-2 cells, the mRNA expression of mismatch repair and tumor suppressor genes, intracellular α-ketoglutarate, and the protein level of TET2 were also downregulated. AMPK deficiency also increased hypermethylation in the CpG islands of Mlh1 in both colonic tissues and Caco-2 cells. In conclusion, AMPK deficiency leads to reduced α-ketoglutarate concentration and elevates the suppressive epigenetic modifications of tumor suppressor genes in gut epithelial cells, thereby increasing the risk of colorectal tumorigenesis. Given the modifiable nature of AMPK activity, it holds promise as a prospective molecular target for the prevention and treatment of CRC.

Flavonoids-Enriched Vegetal Extract Prevents the Activation of NFκB Downstream Mechanisms in a Bowel Disease In Vitro Model.

Inflammatory bowel disease (IBD) incidence has increased in the last decades due to changes in dietary habits. IBDs are characterized by intestinal epithelial barrier disruption, increased inflammatory mediator production and excessive tissue injury. Since the current treatments are not sufficient to achieve and maintain remission, complementary and alternative medicine (CAM) becomes a primary practice as a co-adjuvant for the therapy. Thus, the intake of functional food enriched in vegetal extracts represents a promising nutritional strategy. This study evaluates the anti-inflammatory effects of artichoke, caihua and fenugreek vegetal extract original blend (ACFB) in an in vitro model of gut barrier mimicking the early acute phases of the disease. Caco2 cells cultured on transwell supports were treated with digested ACFB before exposure to pro-inflammatory cytokines. The pre-treatment counteracts the increase in barrier permeability induced by the inflammatory stimulus, as demonstrated by the evaluation of TEER and CLDN-2 parameters. In parallel, ACFB reduces p65NF-κB pro-inflammatory pathway activation that results in the decrement of COX-2 expression as PGE2 and IL-8 secretion. ACFB properties might be due to the synergistic effects of different flavonoids, indicating it as a valid candidate for new formulation in the prevention/mitigation of non-communicable diseases.

Probiotic Lactobacillus fermentum TSF331, Lactobacillus reuteri TSR332, and Lactobacillus plantarum TSP05 improved liver function and uric acid management-A pilot study.

Metabolic-associated fatty liver disease (MAFLD) is predominantly associated with metabolic disturbances representing aberrant liver function and increased uric acid (UA) levels. Growing evidences have suggested a close relationship between metabolic disturbances and the gut microbiota. A placebo-controlled, double-blinded, randomized clinical trial was therefore conducted to explore the impacts of daily supplements with various combinations of the probiotics, Lactobacillus fermentum TSF331, Lactobacillus reuteri TSR332, and Lactobacillus plantarum TSP05 with a focus on liver function and serum UA levels. Test subjects with abnormal levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and UA were recruited and randomly allocated into six groups. Eighty-two participants successfully completed the 60-day intervention without any dropouts or occurrence of adverse events. The serum AST, ALT, and UA levels were significantly reduced in all treatment groups (P < 0.05). The fecal microbiota analysis revealed the intervention led to an increase in the population of commensal bacteria and a decrease in pathobiont bacteria, especially Bilophila wadsworthia. The in vitro study indicated the probiotic treatments reduced lipid accumulation and inflammatory factor expressions in HepG2 cells, and also promoted UA excretion in Caco-2 cells. The supplementation of multi-strain probiotics (TSF331, TSR332, and TSP05) together can improve liver function and UA management and may have good potential in treating asymptomatic MAFLD. Trial registration. The trial was registered in the US Library of Medicine (clinicaltrials.gov) with the number NCT06183801 on December 28, 2023.

Attenuated replication and damaging effects of SARS-CoV-2 Omicron variants in an intestinal epithelial barrier model.

Many COVID-19 patients suffer from gastrointestinal symptoms and impaired intestinal barrier function is thought to play a key role in Long COVID. Despite its importance, the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on intestinal epithelia is poorly understood. To address this, we established an intestinal barrier model integrating epithelial Caco-2 cells, mucus-secreting HT29 cells and Raji cells. This gut epithelial model allows efficient differentiation of Caco-2 cells into microfold-like cells, faithfully mimics intestinal barrier function, and is highly permissive to SARS-CoV-2 infection. Early strains of SARS-CoV-2 and the Delta variant replicated with high efficiency, severely disrupted barrier function, and depleted tight junction proteins, such as claudin-1, occludin, and ZO-1. In comparison, Omicron subvariants also depleted ZO-1 from tight junctions but had fewer damaging effects on mucosal integrity and barrier function. Remdesivir, the fusion inhibitor EK1 and the transmembrane serine protease 2 inhibitor Camostat inhibited SARS-CoV-2 replication and thus epithelial barrier damage, while the Cathepsin inhibitor E64d was ineffective. Our results support that SARS-CoV-2 disrupts intestinal barrier function but further suggest that circulating Omicron variants are less damaging than earlier viral strains.

Lactobacillus acidophilus inhibits the TNF-α-induced increase in intestinal epithelial tight junction permeability via a TLR-2 and PI3K-dependent inhibition of NF-κB activation.

Background: Defective intestinal epithelial tight junction (TJ), characterized by an increase in intestinal TJ permeability, has been shown to play a critical role in the pathogenesis of inflammatory bowel disease (IBD). Tumor necrosis factor-α (TNF-α) is a key pro-inflammatory cytokine involved in the immunopathology of IBD and has been shown to cause an increase in intestinal epithelial TJ permeability. Although TNF-α antibodies and other biologics have been advanced for use in IBD treatment, these therapies are associated with severe side effects and have limited efficacy, and there is an urgent need for therapies with benign profiles and high therapeutic efficacy. Probiotic bacteria have beneficial effects and are generally safe and represent an important class of potential therapeutic agents in IBD. Lactobacillus acidophilus (LA) is one of the most used probiotics for wide-ranging health benefits, including in gastrointestinal, metabolic, and inflammatory disorders. A specific strain of LA, LA1, was recently demonstrated to have protective and therapeutic effects on the intestinal epithelial TJ barrier. However, the mechanisms of actions of LA1 remain largely unknown. Methods: The primary aim of this study was to investigate microbial-epithelial interactions and novel signaling pathways that regulate the effect of LA1 on TNF-α-induced increase in intestinal epithelial TJ permeability, using cell culture and animal model systems. Results and Conclusion: Pre-treatment of filter-grown Caco-2 monolayers with LA1 prevented the TNF-α-induced increase in intestinal epithelial TJ permeability by inhibiting TNF-α-induced activation of NF-κB p50/p65 and myosin light chain kinase (MLCK) gene and kinase activity in a TLR-2-dependent manner. LA1 produced a TLR-2- and MyD88-dependent activation of NF-κB p50/p65 in immune cells; however, LA1, in intestinal cells, inhibited the NF-κB p50/p65 activation in a TLR-2-dependent but MyD88-independent manner. In addition, LA1 inhibition of NF-κB p50/p65 and MLCK gene was mediated by TLR-2 pathway activation of phosphatidylinositol 3-kinase (PI3K) and IKK-α phosphorylation. Our results demonstrated novel intracellular signaling pathways by which LA1/TLR-2 suppresses the TNF-α pathway activation of NF-κB p50/p65 in intestinal epithelial cells and protects against the TNF-α-induced increase in intestinal epithelial TJ permeability.

Short Caco-2 model for evaluation of drug permeability: A sodium valerate-assisted approach.

The human colorectal adenocarcinoma cell line Caco-2, widely used for studying intestinal drug permeability, is typically grown on permeable filter supports and matures in 21 days with frequent media changes. The process is labor-intensive, prone to contamination, and has low throughput, contributing to the overall high utilization cost. Efforts to establish a low-cost, high-throughput, and short-duration model have encountered obstacles, such as weaker tight junctions causing monolayer leaks, incomplete differentiation resulting in low transporter expression, intricate and challenging protocols, and cytotoxicity, limiting the usability. Hence, this study aimed to develop a low-cost, efficient, and short-duration model by addressing the aforementioned concerns by customizing the media and finding a safe differentiation inducer. We generated a new rapid model using sodium valerate, which demonstrated sufficient transporter activity, improved monolayer integrity, and higher levels of differentiation markers than the 21-day model. Furthermore, this model exhibited consistent and reliable results when used to evaluate drug permeability over multiple days of repeated use. This study demonstrates the potential of a sodium valerate-assisted abbreviated model for drug permeability assessment with economic and practical advantages.

A Comparative Study of the Stability, Transport, and Structure-Activity Relationship of Round Scad Derived Peptides with Antineuroinflammatory Ability.

Our previous study identified round scad neuroprotective peptides with different characteristics. However, the intrinsic relationship between their structure and bioactivity, as well as their bioavailability, remains unclear. The aim of this study is to elucidate the bioavailability of these peptides and their structure-activity relationship against neuroinflammation. Results showed that the SR and WCP peptides were resistant to gastrointestinal digestion. Additionally, peptides SR, WCP, and WCPF could transport Caco-2 monolayers as intact peptides. The permeability coefficients (Papp) of SR, WCP, and WCPF in Caco-2 monolayer were (1.53 ± 0.01) × 10-5, (2.12 ± 0.01) × 10-5, and (8.86 ± 0.03) × 10-7 cm/s, respectively. Peptides SR, WCP, and WCPF, as promising inhibitors of JAK2 and STAT3, could attenuate the levels of pro-inflammatory cytokines and regulate the NFκB and JAK2/STAT3 signaling pathway in LPS-treated BV-2 cells. WCPF exerted the highest anti-inflammatory activity. Moreover, bioinformatics, molecular docking, and quantum chemistry studies indicated that the bioactivity of SR was attributed to Arg, whereas those of WCP and WCPF were attributed to Trp. This study supports the application of round-scad peptides and deepens the understanding of the structure-activity relationship of neuroprotective peptides.