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Fruits are essential components of the human diet, valued for their diverse bioactive compounds with potential health-promoting properties. This study focuses on three cold-hardy Actinidia species, namely A. arguta, A. kolomikta, and A. polygama, examining their polyphenolic content, antioxidant/antiradical activities, scavenging capacity and effects on intestinal cell viability (Caco-2 and HT29-MTX). A comprehensive profile of their phenolic compounds was identified, in descending order of total polyphenol content: A. kolomikta > A. arguta > A. polygama. Across species, 16 phenolic acids, 2 flavanols, 2 flavanones, 11 flavonols, and 3 flavones were quantified, with caffeine as a prominent compound. A. kolomikta achieved the highest antioxidant activity, with 'Vitakola' cultivar showing almost double the antioxidant activity compared to 'Tallinn' and 'Pozdni'. By contrast, A. arguta 'Geneva' and A. polygama 'Pomarancheva' exhibited significantly lower activity in both FRAP and DPPH assays. Notably, A. kolomikta cultivars showed distinct radical-scavenging capacities, particularly for superoxide, wherein 'Tallinn' and 'Pozdni' achieved the highest values. Cell viability tests on Caco-2 and HT29-MTX cells revealed a dose-dependent reduction in viability, notably stronger in Caco-2 cells. Overall, this study underscores the therapeutic potential of Actinidia species. [ABSTRACT FROM AUTHOR]

Background/Objectives: The proton-coupled amino acid transporter (PAT1) is an intestinal absorptive solute carrier responsible for the oral bioavailability of some GABA-mimetic drug substances such as vigabatrin and gaboxadol. In the present work, we investigate if non-steroidal anti-inflammatory drug substances (NSAIDs) interact with substrate transport via human (h)PAT1. Methods: The transport of substrates via hPAT1 was investigated in Caco-2 cells using radiolabeled substrate uptake and in X. laevis oocytes injected with hPAT1 cRNA, measuring induced currents using the two-electrode voltage clamp technique. The molecular interaction between NSAIDs and hPAT1 was investigated using an AlphaFold2 model and molecular docking. Results: NSAIDs such as ibuprofen, diclofenac, and flurbiprofen inhibited proline uptake via hPAT1, with IC50 values of 954 (logIC50 2.98 ± 0.1) µM, 272 (logIC50 2.43 ± 0.1) µM, and 280 (logIC50 2.45 ± 0.1) µM, respectively. Ibuprofen acted as a non-competitive inhibitor of hPAT1-mediated proline transport. In hPAT1-expressing oocytes, ibuprofen and diclofenac did not induce inward currents, and inhibited inward currents caused by proline. Molecular modeling pointed to a binding mode involving an allosteric site. Conclusions: NSAIDs interact with hPAT1 as non-translocated non-competitive inhibitors, and molecular modeling points to a binding mode involving an allosteric site distinct from the substrate binding site. The present findings could be used as a starting point for developing specific hPAT1 inhibitors. [ABSTRACT FROM AUTHOR]

This study aims to develop a self-nanoemulsifying drug delivery system (SNEDDS) to solve the limited oral bioavailability problem of apigenin, a bioactive flavonoid. Apigenin-loaded SNEDDS consisting of Gelucire 44/14, Tween 80, and PEG 400 in the mass ratios of 25:37.5:37.5 and 30:35:35 were prepared, and designated as GTP2575 and GTP3070, respectively. The physicochemical stability at 30 and 40 ºC for 6 months was evaluated and a good stability was found. The in vitro transport of apigenin across Caco-2 monolayers from the SNEDDS and the in vivo pharmacokinetics in rats were investigated and compared with apigenin intact form. The in vitro permeation results demonstrated an increased transcellular permeability compared to the apigenin coarse powder (p < 0.05), while there was comparable permeation of apigenin in GTP2575 and GTP3070 formulations, with the permeability constants (Papp) being 2.97 × 10-5 and 3.13 × 10-5, respectively (p > 0.05). The pharmacokinetic analysis in rats revealed that the pharmacokinetic parameters, such as Cmax, AUC0-24, and AUC0−∞, were significantly higher with apigenin-loaded SNEDDS than with apigenin coarse powder (p < 0.05). Apigenin's oral relative bioavailability increased by 3.8 and 3.3 times for GTP2575 and GTP3070, respectively, due to SNEDDS's effect on solubilization and transcellular permeability. The in vivo acute oral toxicity according to OECD 425 was evaluated and revealed low toxicity with an LD50 exceeding 2,000 mg/kg in all apigenin's formulations. These findings suggest that apigenin-loaded SNEDDS may represent a promising strategy for improving the oral delivery of apigenin. [ABSTRACT FROM AUTHOR]

This study examined the impact of Semen raphani on the absorption of ginsenosides from Panax ginseng C.A. Meyer (ginseng) using a Caco-2 cell model and Ultra-High-Performance Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry (UPLC-ESI-MS). Six primary ginsenosides (Rg1, Re, Rb1, Rb2, Rc, Rd) were quantified. Results showed that Semen Raphani increased the efflux rate of ginsenosides, particularly at higher concentrations, suggesting it inhibits their absorption. The research elucidates the intestinal absorption process of ginsenosides and the antagonistic mechanism of Semen Raphani against ginseng. [ABSTRACT FROM AUTHOR]

Infants rely on their developing immune system and the protective components of breast milk to defend against bacterial and viral pathogens, as well as immune disorders such as food allergies, prior to the introduction of solid foods. When breastfeeding is not feasible, fortified infant formula will most frequently be offered, usually based on a cow's milk-based substitute. The current study aimed to explore the immunomodulatory effects of combinations of commercially available human milk oligosaccharides (HMOs). An in vitro co-culture model of Caco-2 intestinal epithelial cells and THP-1 macrophages was established to replicate the hallmarks of intestinal inflammation and to evaluate the direct effects of different synthetic HMO combinations. Notably, a blend of the most prevalent fucosylated and sialylated HMOs, 2′-fucosyllactose (2′-FL) and 6′-siallylactose (6′-SL), respectively, resulted in decreased pro-inflammatory cytokine levels. These effects were dependent on the HMO concentration and on the HMO ratio resembling those in breastmilk. Interestingly, adding additional HMO structures did not enhance the anti-inflammatory effects. This research highlights the importance of carefully selecting HMO combinations in nutritional products, particularly for infant milk formulations, to effectively mimic the benefits associated with breastmilk. [ABSTRACT FROM AUTHOR]

Nonsteroidal anti-inflammatory drugs (NSAIDs) can induce serious adverse effects in gastrointestinal (GI) mucosa, increasing intestinal permeability and leading to mitochondrial dysfunction, oxidative stress, apoptosis and inflammation. As proton pump inhibitors are effective in protecting against NSAID-induced gastropathy but not NSAID-induced enteropathy, current research is focused on natural products as protective substances for therapy and prevention of intestinal injury. Herein, through the use of an in vitro model based on intestinal epithelial cell (Caco-2) damage caused by indomethacin (INDO), we examined the protective activity of a commercially available standardized extract (OFI+OE) from Opuntia ficus-indica (L.) Mill. cladodes and Olea europaea L. leaves. Pre-treatment with OFI+OE prevented INDO-induced intestinal epithelial barrier damage, as demonstrated by TEER measurement, fluorescein permeability, and tight junction protein expression. The extract showed positive effects against INDO-induced oxidative stress and correlated activation of apoptosis, decreasing pro-apoptotic markers BAX and Caspase-3 and increasing anti-apoptotic factor Bcl-2. Moreover, the extract inhibited the NF-κB pathway and pro-inflammatory cascade. In conclusion, these data support the use of OFI+OE extract as a natural strategy for therapy and prevention of intestinal mucosal damage, demonstrating its beneficial effects against INDO-induced intestinal damage, through modulation of oxidative, apoptotic, and inflammatory pathways. [ABSTRACT FROM AUTHOR]

Drug discovery is associated with high levels of compound elimination in all stages of development. The current practices for the pharmacokinetic testing of intestinal absorption combine Transwell® inserts with the Caco-2 cell line and are associated with a wide range of limitations. The improvement of pharmacokinetic research relies on the development of more advanced in vitro intestinal constructs that better represent human native tissue and its response to drugs, providing greater predictive accuracy. Here, we present a humanized, bioengineered intestinal construct that recapitulates aspects of intestinal microanatomy. We present improved histotypic characteristics reminiscent of the human intestine, such as a reduction in transepithelial electrical resistance (TEER) and the formation of a robust basement membrane, which are contributed to in-part by a strong stromal foundation. We explore the link between stromal–epithelial crosstalk, paracrine communication, and the role of the keratinocyte growth factor (KGF) as a soluble mediator, underpinning the tissue-specific role of fibroblast subpopulations. Permeability studies adapted to a 96-well format allow for high throughput screening and demonstrate the role of the stromal compartment and tissue architecture on permeability and functionality, which is thought to be one of many factors responsible for unexpected drug outcomes using current approaches for pharmacokinetic testing. [ABSTRACT FROM AUTHOR]

Disorders in the composition of the microbiota are often associated with pathological processes in the body. The use of new biotherapeutic drugs containing bacteria from normal microflora to correct these disorders may contribute to more effective treatment. The Bacteroidaceae family dominating in microflora is a candidate for inclusion in bacterial preparations. The adhesion properties of five bacterial strains from the Bacteroidaceae family were analyzed using the Caco-2 cell line and their effect on the transepithelial electrical resistance parameters (TEER) was assessed to determine the extent of their impact on the cell monolayer integrity. This approach allowed the selection of strain B. intestinalis 181, which was characterized by a moderate adhesion activity combined with a relatively low effect on the TEER parameter. [ABSTRACT FROM AUTHOR]

Background: Interleukin (IL)-10, an anti-inflammatory cytokine, suppresses the release of inflammatory cytokines, such as IL-6, and inhibits the decline in skin barrier function. Mice orally administered test substances by gavage are subjected to physical stress, which results in the secretion of stress hormones. Stress hormones cause inflammation and reduce skin barrier function. Objective: The effects of heat-killed Lactiplantibacillus (Lpb.) plantarum N1487-7 (N1487-7-HK) on IL-10 production in Caco-2 cells and macrophages, and skin barrier function in physically stressed mice was investigated. Methods: Lpb. plantarum N1487-7 was cultured in fermented barley extract medium at 30 °C for 22 h with shaking (100 rpm). The washed cells were killed by heating at 110 °C for 10 minutes. The cells were then dried in a centrifugal evaporator and ground to obtain the cell powder. The powder was administered to Caco-2 cells and macrophages, and IL-10 mRNA and IL-10 protein expressions were measured by real-time quantitative polymerase chain reaction and Western blotting, respectively. N1487-7-HK cells or distilled water were orally administered daily by gavage to Hos:HR-1 mice for 5 weeks (the N1487-7-HK group or the control group). During weeks 3 and 5 only, dextran sulfate sodium (DSS) was mixed into the drinking water of these groups to induce mild colitis. The normal group was not orally administered anything by gavage and was not given DSS. Serum IL-10 and the trans epidermal water loss (TEWL) of mice were measured. Results: N1487-7-HK promoted the expression of IL-10 mRNA and IL-10 protein in both Caco-2 cells and macrophages. In mice subjected to physical stress, serum IL-10 levels in the N1487-7-HK group were significantly increased compared to the control group. The transepidermal water loss (TEWL) in the control group was notably higher than that in the normal group during Weeks 1 and 2. In contrast, the N1487-7-HK group showed no significant difference in TEWL compared to the normal group. The N1487-7-HK group showed a significantly lower TEWL at Week 4 compared to Week 0. Conclusions: N1487-7-HK promoted IL-10 production from intestinal epithelial cells and macrophages. Intake of N1487-7-HK inhibited the decline in skin barrier function in mice subjected to physical stress. [ABSTRACT FROM AUTHOR]

As food transits the gastrointestinal tract, food structures are disrupted and nutrients are absorbed across the gut barrier. In the past decade, great efforts have focused on the creation of a consensus gastrointestinal digestion protocol (i.e., INFOGEST method) to mimic digestion in the upper gut. However, to better determine the fate of food components, it is also critical to mimic food absorption in vitro. This is usually performed by treating polarized epithelial cells (i.e., differentiated Caco-2 monolayers) with food digesta. This food digesta contains digestive enzymes and bile salts, and if following the INFOGEST protocol, at concentrations that although physiologically relevant are harmful to cells. The lack of a harmonized protocol on how to prepare the food digesta samples for downstream Caco-2 studies creates challenges in comparing inter laboratory results. This article aims to critically review the current detoxification practices, highlight potential routes and their limitations, and recommend common approaches to ensure food digesta is biocompatible with Caco-2 monolayers. Our ultimate aim is to agree a harmonized consensus protocol or framework for in vitro studies focused on the absorption of food components across the intestinal barrier. [ABSTRACT FROM AUTHOR]

The food coloring agent E171 raises many questions concerning its negative impact on human health because of the fact that it contains nanoparticle fractions (NPs, diameter < 100 nm). Numerous studies showed its influence on organisms, including the ability to disrupt the intestinal barrier. In the present study, we verified the potential toxicity and pro-inflammatory activity of three different E171 samples (containing NPs fractions) and one TiO2 NPs sample (60–600 µg mL−1) towards Caco-2 colon cancer cells. The experiments revealed no significant changes in terms of the vitality of Caco-2 cells after 24 h of exposure (XTT test). However, after 72 h, a decrease in the proliferation of Caco-2 cells caused by three TiO2 substances was observed. Moreover, deterioration of the metabolic activity of Caco-2 cells (ATP test) by all analyzed substances at 300 and 600 µg mL−1 was seen. While a 24-h exposure to each tested substance resulted in a negligible release of LDH, a prolonged exposure (72 h) indicated an elevated release of LDH, suggesting potential toxicity. All TiO2 samples induced the elevated release of two primary proinflammatory cytokines, i.e., IL-1β and TNF-α, in a dose-independent manner. The discrepancies in the results come from the differences in the share of individual sizes in four TiO2 products. [ABSTRACT FROM AUTHOR]

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Aim: Colorectal cancer (CRC) impacts almost one million people worldwide each year, with 500,000 deaths annually. In our country, CRC ranks as the third most prevalent cancer in women and the fourth in men. Melatonin, a hormone with well-established antioxidant and anti-cancer properties, is produced by the pineal gland. Agomelatine, used to antidepressant therapies, also acts on melatonin receptors and may offer therapeutic benefits in cancer treatment. This study investigates the effects of melatonin and agomelatine on the viability of human colorectal cancer cell lines (HCT-116 and Caco-2). Materials and Methods: HCT-116 and Caco-2 cell lines were treated with melatonin and Agomelatine, both dissolved in 96% ethanol, at doses of 0.1, 1, 5, and 10 mM for a duration of 24 hours. Cell viability was measured using 3-(4,5-dimetiltiazol-2-il)-2,5- difeniltetrazolyum bromide (MTT) assay. Statistical analyses were performed using IBM SPSS Statistics 24.0, applying Bonferroni correction and the Mann-Whitney U-test, with significance set at p<0.05. The half-maximal inhibitory concentration (LogIC50) was calculated from the MTT assay results. Results: All tested concentrations of melatonin and agomelatine significantly reduced cell viability in both HCT-116 and Caco-2 cell lines (p<0.05). Conclusion: The study concludes that agomelatine and melatonin have substantial cytotoxic and anti-tumor properties against human colorectal cancer cells. [ABSTRACT FROM AUTHOR]

Dengue virus (DENV), from the Flaviviridae family, is the causative agent of dengue fever and poses a significant global health challenge. The virus primarily affects the vascular system and liver; however, a growing body of evidence suggests its involvement in the gastrointestinal (GI) tract, contributing to clinical symptoms such as abdominal pain, vomiting, and diarrhea. However, the mechanisms underlying DENV infection in the digestive system remain largely unexplored. Prior research has detected viral RNA in the GI tissue of infected animals; however, whether the dengue virus can directly infect human enterocytes remains unclear. In this study, we examine the infectivity of human intestinal cell lines to the dengue virus and their subsequent response. We report that the Caco-2 cell line, a model of human enterocytes, is susceptible to infection and capable of producing viruses. Notably, differentiated Caco-2 cells exhibited a lower infection rate yet a higher level of virus production than their undifferentiated counterparts. These findings suggest that human intestinal cells are a viable target for the dengue virus, potentially elucidating the GI symptoms observed in dengue fever and offering a new perspective on the pathogenetic mechanisms of the virus. [ABSTRACT FROM AUTHOR]

Iron uptake in the intestinal epithelium is associated with transport of ferrous iron via the DMT1 transporter (SLC11a2; NRAMP2). In later years, uptake of iron from complex sources, such as nanoparticles, has been found to be mediated through endocytosis. Here we propose that iron from the simple salt ferrous fumarate, a common iron supplement, can be absorbed by clathrin-mediated endocytosis. We used siRNA to silence DMT1 transporter expression, pharmacological inhibition of endocytosis, and Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) to show that iron uptake from ferrous fumarate can be mediated by both transport via DMT1 and by clathrin-dependent endocytosis in Hutu-80 cells. Iron uptake (ferritin L) from ferrous fumarate (0.5 mM, 24 h) in DMT1 silenced cells was significantly decreased (60% ± 11%) in comparison to iron controls while a 1-h dose of ferrous fumarate (0.5 mM) significantly decreased ferritin L formation in the presence of the clathrin inhibitor chlorpromazine (61% ± 10%, in post-confluent cells and 37% ± 9% in non-confluent cells). A pilot showed a similar trend for Ferritin (H) levels (confluent cells) and for total cellular iron load (non-confluent cells). ToF-SIMS analysis revealed diminished membrane-associated iron load in endocytosis-inhibited ferrous fumarate treated cells. The reported results support a clathrin-mediated endocytosis mechanism for uptake of iron from ferrous fumarate in addition to iron uptake by DMT1. More studies are needed to understand what determines which uptake mechanism are employed and to which extent.

IntroductionThe potential for drug-induced gastrointestinal (GI) toxicity is significant, since the GI tract is one of the first barriers which come in to contact with oral drugs. In pharmaceutical research, the complex behavior of human intestinal cells is traditionally investigated using 2D cultures, in which one cell type grows under static conditions. With the development of advanced microphysiological systems (MPS) more in vivo like conditions can be generated which increase the physiological nature and also the predictive validity of these models. Caco-2 cells are known for their capability to build tight junctions. These connections are responsible for the maintenance of intestinal homeostasis and can be used as a specific safety endpoint, by measuring the Trans Epithelial Electrical Resistance (TEER), for the investigation of drug-induced GI toxicity. Compared to a widely used Caco-2 cell 2D Transwell model, the advanced MPS model (Mimetas OrganoPlate®) allows for the recapitulation of the enterocyte cell layer of the intestinal barrier as the Caco-2 cells grow in a tubular structure through which the medium continuously flows.MethodsThe OrganoPlate® intestinal model was qualified to be implemented as a routine test system for the early prediction of drug-induced GI toxicity based on the measurement of the tightness of the cell layer by measuring changes in the TEER. For this qualification 23 well known compounds as well as a positive, negative and solvent control were selected. The compounds were selected based on their known effect on the GI system (chemotherapeutics, tight junction disruptor, liver toxins, controls, NSAIDs and a mixed group of drugs).ResultsThe TEER values were measured 4 h and 24 h after treatment. In parallel the cell viability was determined after 24 h to be able to distinguish between an unspecific cytotoxic effect or direct tight junction damage. Overall, from the 23 tested compounds, 15 showed the expected outcome, i.e., the compound led to a decrease of the TEER for the positive control compounds, or the TEER value remained stable after treatment with non-GI-toxic compounds.ConclusionIn summary, this MPS model allowed the recapitulation of the human intestinal GI barrier and will enable a faster and more robust assessment of drug-induced damage in the GI tract.

While many types of diabetes medications are currently available, orally administered formulations of glucagon-like peptide-1 (GLP-1) receptor agonists have recently been launched. Therefore, gastrointestinal epithelial cells will be increasingly exposed to GLP-1 receptor agonists; however, their effects on these cells remain unclear. The present study attempted to clarify the effects of GLP-1 receptor agonists on intestinal epithelial barrier functions. Semaglutide (5, 50, and 500 ng/mL) and dulaglutide (15, 150, and 1500 ng/mL) were selected as GLP-1 receptor agonists and applied to the Caco-2 cell line. Changes in mRNA and protein expression levels of epithelial cell barrier regulators due to exposure to GLP-1 receptor agonists were examined by real-time RT-PCR and Western blotting. Neither semaglutide nor dulaglutide changed the growth rate or ratio of Caco-2 cells. Furthermore, they did not significantly affect the mRNA expression levels of membrane proteins involved in epithelial cell barrier functions. However, dulaglutide increased the protein expression levels of these membrane proteins in a concentration-dependent manner, whereas semaglutide did not. Only dulaglutide enhanced epithelial cell barrier functions. Since various gastrointestinal symptoms develop in patients with diabetes and epithelial cell barrier functions may be compromised, medicines that promote barrier function, such as dulaglutide, may effectively attenuate these changes. However, their mechanisms of action remain unknown; therefore, further studies are warranted.

The addition of antioxidants to pharmaceutical products is a potential approach to inhibit nitrosamine formation, particularly in solid oral dosage forms like tablets and capsules. The objective was to assess the effect of ten antioxidants on the permeability of four Biopharmaceutics Classification System (BCS) Class III drugs. Bi-directional drug permeability studies in the absence and presence of antioxidants were performed in vitro across MDCK-II monolayers. No antioxidant increased drug permeability, while the positive control sodium lauryl sulfate always increased drug permeability. Results support that any of the ten antioxidants, up to at least 10 mg, can be added to a solid oral dosage form without modulating passive drug intestinal permeability. Additional considerations are also discussed. [ABSTRACT FROM AUTHOR]

The Caco-2 cell culture model was envisioned by Dr. Ronald T. Borchardt and established by the brilliant work of Dr. Ismael Hidalgo. As a practitioner of the Caco-2 cell culture model, I witnessed its initial explosive adoption by big pharma and academic research institutions, followed by regulatory agencies, and is now a well-established model. Through the tremendous efforts of many researchers in both industrial and academic organizations, fully characterized and developed Caco-2 cell culture model is now used in absorption screening, phase I and phase II metabolism, efflux transporter study, etc. Major challenges associated with the Caco-2 model include lack of mucous membranes, insufficient cytochrome P450 expression, inadequate expression of many nutrient transporters under standard culture conditions, etc. Despite this significant insufficiency, it is still much better characterized than any other cell culture models of the intestine and will remain an excellent tool for pharmaceutical sciences in decades to come. [ABSTRACT FROM AUTHOR]

Current drug development tends towards complex chemical molecules, referred to as "beyond rule of five" (bRo5) compounds, which often exhibit challenging physicochemical properties. Measuring Caco-2 permeability of those compounds is difficult due to technical limitations, including poor recovery and detection sensitivity. We implemented a novel assay, with optimized incubation and analytics, to measure permeability close to equilibrium. In this setup an appropriate characterization of permeability for bRo5 compounds is achievable. This equilibrated Caco-2 assay was verified with respect to data validity, compound recovery, and in vitro to in vivo correlation for human absorption. Compared to a standard assay, it demonstrated comparable performance in predicting the human fraction absorbed (fa) for reference compounds. The equilibrated assay also successfully characterized the permeability of more than 90% of the compounds analyzed, the majority of which were bRo5 (68%). These compounds could not be measured using the standard assay. Permeability and efflux ratio (ER) were highly predictive for in vivo absorption for a large set of internal bRo5 compounds. Reference cut-offs enabled the correct classification of high, moderate, and low absorption. This optimized equilibrated Caco-2 assay closes the gap for a high-throughput cellular permeability method in the bRo5 chemical space. [ABSTRACT FROM AUTHOR]

Compounds based on thiazolidinones have drawn particular attention in the field of medicinal chemistry as potential sources of novel drug-like molecules. In this study thiazolidin-4-one derivatives (3–17) were prepared by refluxing a mixture of β-aroylacrylic acid (1) and aryl thiosemicarbazone (2) and characterized using IR and 1H NMR spectroscopy. In silico, physicochemical descriptors and pharmacokinetic properties were assessed using the SwissADME web tool. Moreover, docking studies of synthesized compounds were performed in order to explore their binding to cyclooxygenase-2 (COX-2) (PDB code: 5IKT) and p53 (PDB code: 6MXZ). Some synthesized compounds were evaluated for their in vitro cytotoxic activity against colon cancer (Caco-2) cell line. Among the tested compounds, compound (16) showed a higher effect with (IC50 of 70 µg/ml). The synthesized compounds revealed reduced Caspase-3 and p53 gene expression, suggesting the potential antineoplastic effects of these compounds. [ABSTRACT FROM AUTHOR]

The psychedelic beverage ayahuasca is originally obtained by Banisteriopsis caapi (B. caapi) (BC) and Psychotria viridis (P. viridis) (PV). However, sometimes these plant species are replaced by others that mimic the original effects, such as Mimosa hostilis (M. hostilis) (MH) and Peganum harmala (P. harmala) (PH). Its worldwide consumption and the number of studies on its potential therapeutic effects has increased. This study aimed to evaluate the anticancer properties of ayahuasca in human colorectal adenocarcinoma cells. Thus, the maximum inhibitory concentration (IC50) of decoctions of MH, PH, and a mixture of these (MHPH) was determined. The activities of caspases 3 and 9 were evaluated, and the cell proliferation index was determined through immunocytochemical analysis (Ki-67). Two fluorescent probes were used to evaluate the production of oxidative stress and the activity of the antioxidant enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx) was also evaluated. It was demonstrated that exposure to the extracts significantly induced apoptosis in Caco-2 cells, while decreasing cell proliferation. MH and MHPH samples significantly reduced oxidative stress and significantly increased glutathione peroxidase activity. No significant differences were found in SOD activity. Overall, it was demonstrated that the decoctions have a potential anticancer activity in Caco-2 cells. [ABSTRACT FROM AUTHOR]

Upon dietary administration, probiotic microorganisms can reach as live cells the human gut, where they interact with the microbiota and host cells, thereby exerting a beneficial impact on host functions, mainly through immune-modulatory activities. Recently, attention has been drawn by postbiotics, i.e. non-viable probiotic microbes, including their metabolic products, which possess biological activities that benefit the host. Lactiplantibacillus plantarum is a bacterial species that comprises recognised probiotic strains. In this study, we investigated in vitro the probiotic (and postbiotic) potential of seven L. plantarum strains, including five newly isolated from plant-related niches. The strains were shown to possess some basic probiotic attributes, including tolerance to the gastrointestinal environment, adhesion to the intestinal epithelium and safety. Besides, their cell-free culture supernatants modulated cytokine patterns in human macrophages in vitro, promoting TNF-α gene transcription and secretion, while attenuating the transcriptional activation and secretion of both TNF-α and IL-8 in response to a pro-inflammatory signal, and enhancing the production of IL-10. Some strains induced a high IL-10/IL-12 ratio that may correlate to an anti-inflammatory capacity in vivo. Overall, the investigated strains are good probiotic candidates, whose postbiotic fraction exhibits immunomodulatory properties that need further in vivo studies. The main novelty of this work consists in the polyphasic characterisation of candidate beneficial L. plantarum strains obtained from relatively atypical plant-associated niches, by an approach that explores both probiotic and postbiotic potentials, in particular studying the effect of microbial culture-conditioned media on cytokine pattern, analysed at both transcriptional and secretion level in human macrophages. [ABSTRACT FROM AUTHOR]

Considering the increase in the production and use of nanomaterials (NM) in food/feed and food contact materials, novel strategies for efficient and sustainable hazard characterization, especially in the early stages of NM development, have been proposed. Some of these strategies encompass the utilization of in vitro simulated digestion prior to cytotoxic and genotoxic assessment. This entails exposing NM to fluids that replicate the three successive phases of digestion: oral, gastric, and intestinal. Subsequently, the resulting digestion products are added to models of intestinal cells to conduct toxicological assays, analyzing multiple endpoints. Nonetheless, exposure of intestinal cells to the digested products may induce cytotoxicity effects, thereby posing a challenge to this strategy. The aim of this work was to describe the challenges encountered with the in vitro digestion INFOGEST 2.0 protocol when using the digestion product in toxicological studies of NM, and the adjustments implemented to enable its use in subsequent in vitro biological assays with intestinal cell models. The adaptation of the digestion fluids, in particular the reduction of the final bile concentration, resulted in a reduced toxic impact of digestion products. [ABSTRACT FROM AUTHOR]

Colorectal cancer is known for its substantial impact on global morbidity and mortality, with higher prevalence in developed regions. This study delves into the potential treatment advantages of resveratrol (RSV) in addressing colorectal cancer. Apoptosis and gene expression associated with apoptotic factors were explored using Caco-2 cells, a pertinent model for colorectal adenocarcinoma. The effect of RSV on Caco-2 cell viability was investigated using MTT assay and neutral red uptake assay. The level of generated ROS was high in cells exposed to RSV. Likewise, the enzyme superoxide dismutase, responsible for converting ROS into hydrogen peroxide, was concurrently elevated. The effect of RSV on DNA damage was examined through the TUNEL assay. The gene expression analyses for pro-apoptotic elements were studied using qRT-PCR. Furthermore, the impact of RSV on the migration of Caco-2 cells was conducted through a wound-healing assay. Our results reveal RSV’s cytotoxicity on Caco-2 cells, showing dose-dependent inhibition of viability, indicating its promise as a treatment agent. The induction of cell death by apoptosis is substantiated by DNA damage. Notably, the upregulated expression of caspase-3, Bax, and p53 genes suggests RSV’s potential to modulate key apoptosis-related elements. In addition, RSV displayed an inhibitory effect on cellular migration, a significant (p < 0.05 and p < 0.01) in cancer metastasis. These findings underscore RSV’s potential to be a multifaceted therapeutic agent targeting apoptosis and metastatic processes in colorectal cancer.

Nonsteroidal anti-inflammatory drugs (NSAIDs) can induce serious adverse effects in gastrointestinal (GI) mucosa, increasing intestinal permeability and leading to mitochondrial dysfunction, oxidative stress, apoptosis and inflammation. As proton pump inhibitors are effective in protecting against NSAID-induced gastropathy but not NSAID-induced enteropathy, current research is focused on natural products as protective substances for therapy and prevention of intestinal injury. Herein, through the use of an in vitro model based on intestinal epithelial cell (Caco-2) damage caused by indomethacin (INDO), we examined the protective activity of a commercially available standardized extract (OFI+OE) from Opuntia ficus-indica (L.) Mill. cladodes and Olea europaea L. leaves. Pre-treatment with OFI+OE prevented INDO-induced intestinal epithelial barrier damage, as demonstrated by TEER measurement, fluorescein permeability, and tight junction protein expression. The extract showed positive effects against INDO-induced oxidative stress and correlated activation of apoptosis, decreasing pro-apoptotic markers BAX and Caspase-3 and increasing anti-apoptotic factor Bcl-2. Moreover, the extract inhibited the NF-κB pathway and pro-inflammatory cascade. In conclusion, these data support the use of OFI+OE extract as a natural strategy for therapy and prevention of intestinal mucosal damage, demonstrating its beneficial effects against INDO-induced intestinal damage, through modulation of oxidative, apoptotic, and inflammatory pathways.

A total of 55 food and clinical S. Schwarzengrund isolates were assayed for plasmid content, among which an IncFIB-IncFIC(FII) fusion plasmid, conferring streptomycin resistance, was detected in 17 isolates. Among the 17 isolates, 9 were food isolates primarily collected from poultry meat, and 8 clinical isolates collected from stool, urine, and gallbladder. SNP--based phylogenetic analyses showed that the isolates carrying the fusion plasmid formed a subclade indicating the plasmid was acquired and is now maintained by the lineage. Phylogenetic analysis of the plasmid suggested it is derived from avian pathogenic plasmids and might confer an adaptive advantage to the S. Schwarzengrund isolates within birds. IncFIB-IncFIC(FII) fusion plasmids from all food and three clinical isolates were self-conjugative and successfully transferred into E. coli J53 by conjugation. Food and clinical isolates had similar virulome profiles and were able to invade human Caco-2 cells. However, the IncFIB-IncFIC(FII) plasmid did not significantly add to their invasion and persistence potential in human Caco-2 cells. [ABSTRACT FROM AUTHOR]

Human malignancies are one of the major health-related issues throughout the world and are anticipated to rise in the future. Despite huge investments made in anticancer drug development, limited success has been obtained and the average number of FDA approvals per year is declining. So, an increasing interest in drug repurposing exists. Metformin (MET) and aspirin (ASP) possess anticancer properties. This work aims to test the effect of these two drugs in combination on colorectal cancer (CRC) cells in vitro. The effects of MET and/or ASP on cell proliferation, viability, migratory ability, anchorage-independent growth ability (colony formation), and nutrient uptake were determined in two (HT-29 and Caco-2) human CRC cell lines. Individually, MET and ASP possessed antiproliferative, cytotoxic, and antimigratory effects and reduced colony formation in HT-29 cells (BRAF- and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit α (PI3KCA)-mutant), although MET did not affect either 3H-deoxy-D-glucose or 14C-butyrate uptake and lactate production, and ASP caused only a small decrease in 14C-butyrate uptake. Moreover, in these cells, the combination of MET and ASP resulted in a tendency to an increase in the cytotoxic effect and in a potentiation of the inhibitory effect on colony formation, although no additive antiproliferative and antimigratory effects, and no effect on nutrient uptake and lactate production were observed. In contrast, MET and ASP, both individually and in combination, were almost devoid of effects on Caco-2 cells (BRAF- and PI3KCA-wild type). We suggest that inhibition of PI3K is the common mechanism involved in the anti-CRC effect of both MET, ASP and their combination and, therefore, that the combination of MET + ASP may especially benefit PI3KCA-mutant CRC cases, which currently have a poor prognostic. [ABSTRACT FROM AUTHOR]

Adhesion G protein-coupled receptors (aGPCRs) play an important role in neurodevelopment, immune defence and cancer; however, their role throughout viral infections is mostly unexplored. We have been searching for specific aGPCRs involved in SARS-CoV-2 infection of mammalian cells. In the present study, we infected human epithelial cell lines derived from lung adenocarcinoma (Calu-3) and colorectal carcinoma (Caco-2) with SARS-CoV-2 in order to analyse changes in the level of mRNA encoding individual aGPCRs at 6 and 12 h post infection. Based on significantly altered mRNA levels, we identified four aGPCR candidates—ADGRB3/BAI3, ADGRD1/GPR133, ADGRG7/GPR128 and ADGRV1/GPR98. Of these receptors, ADGRD1/GPR133 and ADGRG7/GPR128 showed the largest increase in mRNA levels in SARS-CoV-2-infected Calu-3 cells, whereas no increase was observed with heat-inactivated SARS-CoV-2 and virus-cleared conditioned media. Next, using specific siRNA, we downregulated the aGPCR candidates and analysed SARS-CoV-2 entry, replication and infectivity in both cell lines. We observed a significant decrease in the amount of SARS-CoV-2 newly released into the culture media by cells with downregulated ADGRD1/GPR133 and ADGRG7/GPR128. In addition, using a plaque assay, we observed a reduction in SARS-CoV-2 infectivity in Calu-3 cells. In summary, our data suggest that selected aGPCRs might play a role during SARS-CoV-2 infection of mammalian cells. [ABSTRACT FROM AUTHOR]

Objective: Previous research has partially revealed distinct gut microbiota in ankylosing spondylitis (AS). In this study, we performed non-targeted fecal metabolomics in AS in order to discover the microbiome--metabolome interface in AS. Based on prospective cohort studies, we further explored the impact of the tumor necrosis factor inhibitor (TNFi) on the gut microbiota and metabolites in AS. Methods: To further understand the gut microbiota and metabolites in AS, along with the influence of TNFi, we initiated a prospective cohort study. Fecal samples were collected from 29 patients with AS before and after TNFi therapy and 31 healthy controls. Metagenomic and metabolomic experiments were performed on the fecal samples; moreover, validation experiments were conducted based on the association between the microbiota and metabolites. Results: A total of 7,703 species were annotated using the metagenomic sequencing system and by profiling the microbial community taxonomic composition, while 50,046 metabolites were identified using metabolite profiling. Differential microbials and metabolites were discovered between patients with AS and healthy controls. Moreover, TNFi was confirmed to partially restore the gut microbiota and the metabolites. Multi-omics analysis of the microbiota and metabolites was performed to determine the associations between the differential microbes and metabolites, identifying compounds such as oxypurinol and biotin, which were correlated with the inhibition of the pathogenic bacteria Ruminococcus gnavus and the promotion of the probiotic bacteria Bacteroides uniformis. Through experimental studies, the relationship between microbes and metabolites was further confirmed, and the impact of these two types of microbes on the enterocytes and the inflammatory cytokine interleukin-18 (IL-18) was explored. Conclusion: In summary, multi-omics exploration elucidated the impact of TNFi on the gut microbiota and metabolites and proposed a novel therapeutic perspective: supplementation of compounds to inhibit potential pathogenic bacteria and to promote potential probiotics, therefore controlling inflammation in AS. [ABSTRACT FROM AUTHOR]

The antitumor activity of different ent-kaurane diterpenes has been extensively studied. Several investigations have demonstrated the excellent antitumor activity of synthetic derivatives of the diterpene atractyligenin. In this research, a series of new synthetic amides and their 15,19-di-oxo analogues obtained from atractyligenin by modifying the C-2, C-15, and C-19 positions were designed in order to dispose of a set of derivatives with different substitutions at the amidic nitrogen. Using different concentrations of the obtained compounds (10–300 μM) a reduction in cell viability of HCT116 colon cancer cells was observed at 48 h of treatment. All the di-oxidized compounds were more effective than their alcoholic precursors. The di-oxidized compounds had already reduced the viability of two colon cancer cells (HCT116 and Caco-2) at 24 h when used at low doses (2.5–15 μM), while they turned out to be poorly effective in differentiated Caco-2 cells, a model of polarized enterocytes. The data reported here provide evidence that di-oxidized compounds induced apoptotic cell death, as demonstrated by the appearance of condensed and fragmented DNA in treated cells, as well as the activation of caspase-3 and fragmentation of its target PARP-1. [ABSTRACT FROM AUTHOR]

Within-host infection dynamics of Omicron dramatically differs from previous variants of SARS-CoV-2. However, little is still known about which parameters of virus-cell interplay contribute to the observed attenuated replication and pathogenicity of Omicron. Mathematical models, often expressed as systems of differential equations, are frequently employed to study the infection dynamics of various viruses. Adopting such models for results of in vitro experiments can be beneficial in a number of aspects, such as model simplification (e.g., the absence of adaptive immune response and innate immunity cells), better measurement accuracy, and the possibility to measure additional data types in comparison with in vivo case. In this study, we consider a refinement of our previously developed and validated model based on a system of integro-differential equations. We fit the model to the experimental data of Omicron and Delta infections in Caco-2 (human intestinal epithelium model) and Calu-3 (lung epithelium model) cell lines. The data include known information on initial conditions, infectious virus titers, and intracellular viral RNA measurements at several time points post-infection. The model accurately explains the experimental data for both variants in both cell lines using only three variant- and cell-line-specific parameters. Namely, the cell entry rate is significantly lower for Omicron, and Omicron triggers a stronger cytokine production rate (i.e., innate immune response) in infected cells, ultimately making uninfected cells resistant to the virus. Notably, differences in only a single parameter (e.g., cell entry rate) are insufficient to obtain a reliable model fit for the experimental data. [ABSTRACT FROM AUTHOR]

β-Cryptoxanthin is a type of carotenoid that contributes to the maintenance and improvement of human health. However, it is sensitive to heat during cooking and food processing. Furthermore, like other carotenoids, β-cryptoxanthin is also highly hydrophobic and does not readily solubilize in the gastrointestinal tract. This property results in its low bioaccessibility, and considerable knowledge has been accumulated on improving the bioavailability of β-cryptoxanthin. In this study, we evaluated the effects of different processing methods on the residual amount of β-cryptoxanthin in kumquats. The results showed that preparation of kumquat paste contained more β-cryptoxanthin than its puree. Next, when the paste was mixed with fermented Aspergillus oryzae milk, the β-cryptoxanthin content did not change. A pasteurization study of the kumquat paste mixed with the fermented milk showed that β-cryptoxanthin was more stable at 65 °C than at 85 °C. In addition, to improve the bioaccessibility of β-cryptoxanthin, a mixture of the fermented milk and kumquat-derived β-cryptoxanthin was evaluated using a human intestinal epithelial Caco-2 cell model. The results showed that β-cryptoxanthin in the mixture permeated the membrane 14 times more than β-cryptoxanthin alone. [ABSTRACT FROM AUTHOR]

Genistein was esterified with nine acyl chlorides. The resulting esters were characterized using HPLC-MS and 1H and 13C NMR. The antioxidant activity of genistein esters was assessed in both chemical models and Caco-2 cells. All genistein esters exhibited lower 1,1-Diphenyl-2-picrylhydrazyl radical 2,2-Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl (DPPH) and 2, 2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activities compared to genistein. Seven genistein monoesters exhibited stronger cellular antioxidant activity (CAA) than genistein (CAA unit = 1.1), with genistein-7-O-octanoate (G-C8:0) (CAA unit = 25.9) being the strongest. Further mechanism study revealed that G-C8:0 treatment significantly enhanced the activities of antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-PX), along with upregulating the protein expression of nuclear factor E2-related factor (Nrf2) and its downstream antioxidases, including SOD, Heme oxygenase 1 (HO-1), and GSH-PX. Overall, these findings indicate that esterification is a promising method for augmenting the cellular antioxidant activity of parent isoflavones.

Cancer is the leading cause of death worldwide, with breast and colorectal cancers being the two most common cancer forms.The present work was designed to investigate the probable oncolytic effect of lentogenic and velogenic Newcastle disease viruses on MCF-7 and Caco-2 cell lines compared to the commonly used chemotherapies as an in vitro preliminary study to further prelude an in vivo study.The cytotoxic effects ofNewcastle disease virus strains NDV/chicken/Egypt/Giza/2015 (velogenic NDV genotype VIID) and Lasota strains, as well as the commonly used chemotherapies (Paclitaxel or Doxorubicin) were investigated on MCF-7 and Caco-2 cell lines at different concentrations. Both the human colorectal adenocarcinoma (Caco-2) and the Michigan Cancer Foundation-7 (MCF-7) human breast cancer cell lines were inoculated with NDV VIID and LaSota at concentrations of 10-2, 10-3, 10-4, and 10-5, Paclitaxel (for MCF-7) at concentrations of 0.5, 1, and 2 μM, and Doxorubicin (for Caco-2) at 0.1, 1 and 10 μM in four replicates each. The cytotoxic effect was performed using a neutral red assay for both virus strains and in combination with chemotherapeutic agents. The present study clarified that both VIID and LaSota strains of NDV, particularly at titers of 10-3 and 10-4 TCID50/ml, respectively, displayed a significant (P ≤ 0.05) cytotoxic effect on both MCF-7 and Caco-2 cell lines. Moreover, the combined treatment of the TCID50 (Tissue Culture Infective Dose 50) doses of both NDV strains and the tested chemotherapies showed a more significant (P ≤ 0.05) cytotoxic effect than the sole use of each. Depending on the results, we can conclude that this study opens the way for further in vivo studies aiming to provide more safe treatment for human cancers, save human lives, and avoid dramatic ends.