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MoS2 nanosheets (NSs) are novel 2D nanomaterials (NMs) with potential uses in many areas, and therefore oral exposure route to MoS2 NSs is plausible. Currently, MoS2 NSs are considered as biocompatible NMs, but there is lacking of systemic investigations to study the interactions of MoS2 NSs with intestinal cells. In this study, we exposed the 3D Caco-2 spheroids to MoS2 NSs or MoS2 powders (denoted as MoS2-bulk), and investigated the potential adverse effects of MoS2-materials based on transcriptomics and lipidomics analysis. As expected, both MoS2 NSs and MoS2-bulk were dose-dependently internalized into 3D Caco-2 spheroids but did not induce cytotoxicity, membrane disruption or decrease of thiols. However, the Gene Ontology (GO) and Kyoto Encyclopedia of Gene and Genomes (KEGG) analysis indicated that nutrient absorption and metabolism was decreased. One of the most significantly decreased KEGG pathways is fat digestion and absorption (map04975), and Western blotting analysis further showed that fatty acid binding protein 1 and apolipoprotein A1, key proteins involved in fat digestion and absorption, were down-regulated by MoS2 NSs or MoS2-bulk. In addition, BODIPY 493/503 staining suggested that exposure to MoS2 NSs and MoS2-bulk decreased lipid levels in the spheroids. However, lipidomics data indicated that MoS2 materials only decreased 8 lipid classes, including lysophosphatidylcholine, lysodimethylphosphatidylethanolamine, N-acylethanolamine, ceramide phosphoethanolamines, gangliosides, lysosphingomyelin and sulfatide, whereas most of the lipid classes were indeed increased. In addition, MoS2 NSs was more potent to decrease the lipid classes compared with MoS2-bulk. Combined, the results from this study showed that MoS2 NSs and bulk materials were non-cytotoxic but altered lipid profiles in 3D Caco-2 spheroids.

Ankaferd Blood Stopper (ABS) is a medicinal plant extract that has anti-inflammatory effect. Inflammatory bowel disease is a pathological condition that directly affects colon health and increases the risk of colon cancer. Especially inflammation is an important factor in the formation and progression of this disease. The aim of the study was to investigate the protective effect of ABS on colonic inflammation. Caco-2 and RAW 264.7 cells were used as a model of in vitro colonic inflammation. RAW 264.7 cells were treated with lipopolysaccharide for 12 h to induce inflammation, and an inflammatory medium (IM) was obtained. Caco-2 cells were treated with 15 μL/mL ABS for 4 h, then incubated with IM. The cells also were incubated with 15 μL/mL ABS and IM together for 12 h. Tumor necrosis factor alpha (TNF-α) protein levels were targeted in testing inflammatory condition and cyclooxygenase-2 (COX-2) mRNA level was used as a marker gene to show the possible anti-inflammatory effect of ABS in Caco-2 cells. TNF-α level was 26.1-fold higher than the control group. IM caused 3.2-fold increase in COX-2 expression in Caco-2 cells. Pretreatment of Caco-2 cells with ABS resulted in 3.3-fold decrease in COX-2 mRNA levels relative to IM group. Furthermore, COX-2 mRNA level reduced 4.7-fold when ABS and conditional medium were given at the same time. ABS has suppressive effect on COX-2 mRNA expression in Caco-2 cells. These results suggest that ABS might have protective and therapeutic effect for colonic inflammation.

In Côte d'Ivoire, the market of aphrodisiacs of natural origin has been significantly growing over the last decades. The objective of this study was to evaluate the cytotoxicity of the most employed by Ivorian men, called Aphro. For that, Caco-2/TC7 enterocytes and Vero ATCC cells kidney epithelial cells grown in DMEM medium were exposed to Aphro at concentrations ranging from 216 to 864 µg/mL. Lactate dehydrogenase (LDH) released upon cell death was assayed after 1, 2 and 24h of exposure. In parallel, the effect of Apho was evaluated in differentiated confluent cells by measurements of the trans-epithelial electrical resistance. A time course and significant dose response related increase of LDH was measured in the culture medium of Caco-2/TC7 and Vero ATCC cells exposed to the aphrodisiac. The trans-epithelial electrical resistance of the Caco-2/TC7 cell monolayers also showed a significant decrease from 1400 Ω*cm2 at the onset of the experiment to 450 Ω*cm2 after 24 h of treatment. Cell detachment and structural disorganization were observed in both cell lines. This study reveals the strong cytotoxicity of Aphro extract on both cell lines.&nbsp

Hydroxyanthraquinones from plants have been used as both medicinal active ingredients and adulterants in slimming food supplements. Although sensible doses of certain natural hydroxyanthraquinones for laxative effects are generally safe in the short term, chronic intake has been related to tumorigenic, carcinogenic, and genotoxic effects. However, an increasing number of researchers are reporting the antiproliferative properties of the same ingredients in cancer cells, pointing towards a potential nutraceutical value for cancer prevention. Previous studies have evaluated anthraquinones’ anti-proliferative activity against various tumour cell lines and bioavailability in Caco-2 cells. However, there are scarce data about both their cytotoxicity in the later cell line and long-term stability. Therefore, this study will check the purity of several ‘aged’ samples using mutually complementary analytical techniques such as HPTLC and NMR assays as well as evaluate the anti-proliferative activity of the purest of these samples using the Caco-2 cell line. The chromatographic and spectroscopic analyses confirmed the long-term stability of those compounds, and their cytotoxic activity resulted in chrysazin (15 µg/mL) &gt, catenarin (27.29 µg/mL) &gt, rhein (49.55 µg/mL) &gt, helminthosporin (52.91 µg/mL) &gt, aloe-emodin (55.34 µg/mL). Our succinct review of the cytotoxicity of these compounds afforded two results: that this is the first clear report for catenarin being active in colon cancer cells and that this class of compounds needs to be better studied to clearly evaluate their benefit/risk profile in regard to both new chemo preventative nutraceuticals and anticancer therapies.

Cancer is the second death cause worldwide, with breast and colon cancer among the most prevalent types. Traditional treatment strategies have several side effects that inspire the development of novel anticancer agents derived from natural sources, like chalcone derivatives. For this investigation, twenty-three chalcones (4a-w) were synthesized and evaluated as antiproliferative agents against MCF-7 and Caco-2 cells, finding three and two compounds with similar or higher antiproliferative activity than daunorubicin, while only two chalcones showed better selectivity indexes than daunorubicin on MCF-7. From these results, we developed good-performance QSAR models (r > 0.850, q2>0.650), finding several structural features that could modify chalcone activity and selectivity. According to these models, chalcones 4w and 4t have high potency and selectivity against Caco-2 and MCF-7, respectively, which make them attractive candidates for hit-to-lead development of ROS-independent pro apoptotic agents.

Euglena gracilis, a type of microalgae, contains several nutrients and accumulates paramylon, a β-1,3-glucan. In recent studies, paramylon has shown to exhibit various activities including immunomoduratory and hepatoprotective effects. In the present study, using an in vitro cell culture system, we aimed to determine whether paramylon derived from the E. gracilis EOD-1 strain, which produces large amounts of paramylon, can augment SIRT1 expression in epidermal cells via activating gut–skin interactions. Results showed that paramylon augmented the expression of SIRT1 in Caco-2 cells, a human intestinal cell line. Furthermore, microarray analysis of Caco-2 cells treated with paramylon showed that paramylon activates epidermal cells through inducing the secretion of factors from intestinal cells. Then, we focused on skin cells as target cells of paramylon-activated intestinal cells. Results showed that secretory factors from Caco-2 cells treated with paramylon augmented the expression of SIRT1 in HaCaT cells, a human keratinocyte cell line, and that expression level of genes related to the growth and maintenance of epidermal cells were significantly changed in Caco-2 cells treated with paramylon as evidenced by microarray analysis. All these results suggest that paramylon can activate epidermal cells by inducing the production of secretory factors from intestinal cells.

Despite the latest pursuit to discover novel phenolic compounds in hempseed and its biological properties, it remains a mystery whether they can be absorbed in the human body. Extrusion treatment and extraction of the free phenolic fraction significantly (p < 0.05) improved human Caco-2 intestinal cell absorption for hempseed hull lignanamides including cannabisin B (Papp value of 1.35 × 10-5 ± 1.0 × 10-6) as compared to the bound fraction of raw hull (Papp value of 2.82 × 10-6 ± 5.2 × 10-7). Co-supplementation of the flavonoid naringenin (20 μM) further improved these absorption rates. Higher cellular antioxidant activity was observed in the free extraction fractions. Treatment with the free phenolic fraction of extruded hempseed hull (100 mg/mL) alleviated tert-butyl hydroperoxide's (25 μM) negative effects on cell viability, intracellular malondialdehyde levels, apoptosis induction, glutathione S-transferase, and glutathione levels. RNA-sequencing with the limma method unveiled a total of 2795 differentially expressed genes in 21 day-old Caco-2 intestinal cells, suggesting the changes in cell metabolism after exposure to extruded hempseed hull extract. This study could promote the utilization of extrusion technology to improve the absorption and antioxidant capacities of bioactive phenolics in plant food processing byproducts.

The integrity and permeability of the intestinal epithelial barrier are important indicators of intestinal health. Impaired intestinal epithelial barrier function and increased intestinal permeability are closely linked to the onset and progression of various intestinal diseases. Sinapic acid (SA) is a phenolic acid that has anti-inflammatory, antihyperglycemic, and antioxidant activities; meanwhile, it is also effective in the protection of inflammatory bowel disease (IBD), but the specific mechanisms remain unclear. Here, we evaluated the anti-inflammatory of SA and investigated its potential therapeutic activity in LPS-induced intestinal epithelial barrier and tight junction (TJ) protein dysfunction. SA improved cell viability; attenuated epithelial permeability; restored the protein and mRNA expression of claudin-1, ZO-1, and occludin; and reversed the redistribution of the ZO-1 and claudin-1 proteins in LPS-treated Caco-2 cells. Moreover, SA reduced the inflammatory response by downregulating the activation of the TLR4/NF-κB pathway and attenuated LPS-induced intestinal barrier dysfunction by decreasing the activation of the MLCK/MLC pathway. This study demonstrated that SA has strong anti-inflammatory activity and can alleviate the occurrence of high intercellular permeability in Caco-2 cells exposed to LPS.

In this study, pyrralylisoleucine (Pyrr-Ile) and pyrralylalanine (Pyrr-Ala), two dipeptide-bound pyrralines with different C-termini were synthesized as the representatives of dietary advanced glycation end products (dAGEs). The structures of Pyrr-Ile and Pyrr-Ala were characterized by high-resolution mass spectrometry, nuclear magnetic resonance, and Fourier transform infrared spectroscopy. Then, the transport of Pyrr-Ile and Pyrr-Ala across intestinal epithelial cells was investigated using Caco-2 cell monolayers, and their interaction with aminopeptidase N (APN) was analyzed. The results showed that the apparent permeability coefficient (Papp) of Pyrr-Ala was (14.1 ± 2.26) × 10-7 cm·s-1 calculated by free pyrraline, while the Papp values of Pyrr-Ile were (32.4 ± 5.35) × 10-7 and (19.1 ± 1.46) × 10-7 cm·s-1 when they were, respectively, calculated according to their dipeptide-bound or free form. Both Pyrr-Ala and Pyrr-Ile were potential substrates of APN, and their hydrolysis by APN may make the intact transmembrane transport of Pyrr-Ala and Pyrr-Ile more difficult, especially for Pyrr-Ala. Besides, the occurrence of product inhibition in hydrolysis of Pyrr-Ile was possible. Pyrr-Ile and Pyrr-Ala were different in Papp values and transport forms, which suggested that the C-terminus may play an important role in their transport across the Caco-2 cell monolayers. In addition, the results highlight the intact transmembrane transport of dipeptide-bound pyrraline.

OBJECTIVES: Cancer diseases have been linked to a huge number of causes that led to deaths in this century along with cardiovascular and lung diseases. Most death-leading types of cancer are colon, lung, breast, and prostate cancers. Due to the remarkable properties of gold (Au) nanocarrier, they are used to deliver and improve tamoxifen (Tam) citrate activity in Caco-2 and MCF-7 cells. MATERIALS AND METHODS: In this study, preparation of Au nanoparticles (NPs), zeta-potential and size, high resolution transient electron microscopy (HRTEM), high-performance liquid chromatography, ultraviolet-visible spectra, fluorescence microscopy, fourier infrared spectroscopy, and real-time cellular analysis xCELLigence technology were investigated. RESULTS: The zeta-average size of the Tam- β-cyclodextrin (β-CD)-hyaluronic acid (HA)-chitosan (Chi)-Au nanocomposite is 82.02 nm with a negative zeta potential of -23.6. Furthermore, HRTEM images showed that, successful formulation of polymer shell around Au core and the Au NP shape is mostly spherical, triangle and irregular. Furthermore, the fluorescence microscope image showed proper cellular uptake of the Tam-β- CD-HA-Chi-Au nanocomposite in MCF-7 and Caco-2 cells. Additionally, Tam-β-CD-HA-Chi-Au nanocomposite significantly improved the cytotoxic activity of Tam citrate on Caco-2 cells. IC50 value of Tam reduced from 8.55 µM to 5.32 µM, after 48 h of incubation time (p value

Lactoferrin (LF) is a bioactive glycoprotein in human milk and has positive effects on neonates. The LF knockout mouse model was generated as a mother mouse that provided LF-free milk. The intestinal development of suckling neonates drinking normal milk and LF-free milk was studied. The results showed that the intestinal density, maturity, and barrier integrity of mice drinking LF-free milk were lower than those of mice drinking normal milk. Therefore, the importance of adding lactoferrin to the human formula is considered. Human lactoferrin (HLF), bovine lactoferrin (BLF), and recombinant HLF (RHLF) were used to compare their functional impact on Caco-2 cell lines. Cell proliferation, differentiation, the establishment of the intestinal barrier, and protective effects on lipopolysaccharide injury were detected. Our results showed that RHLF exhibited more similar functions to HLF than BLF and showed the combined advantages of HLF and BLF in promoting the establishment of the intestinal barrier. This study emphasizes the important role of LF in neonatal intestinal development and provides a theoretical basis for the availability of RHLF.

Imidacloprid is the most widely used pesticide of the neonicotinoid class. Neonicotinoid toxicities against various insects are well known. Nevertheless, there are rising evidences that neonicotinoids exert cytotoxic effects on different non-target organisms including mammals, fish, birds etc. Besides, depending on pesticide application, the exposed plants absorb some part of used neonicotinoids and their residues are detected in agricultural products worldwide. Thus, the continuous consumption of fruits and vegetables contaminated with neonicotinoids is a high risk factor for humans despite the low doses. Intestine epithelial cells are the first targets of the neonicotinoid cytotoxicity in humans because of its direct way of administration. The epithelial cells provide the barrier function of the intestinal system via specialized intercellular adhesion. The effects of imidacloprid on the intestine barrier function and inflammatory cytokines production are still unknown. In the present study, we exposed the human Caucasian colon adenocarcinoma (Caco-2) epithelial cells to low doses (0.10–0.75 µg/mL) of imidacloprid in order to assess the expression of tight and adherens junctions proteins, occludin and E-cadherin, and production of proinflammatory cytokine TNF α and iNOS. Imidacloprid induced dose-dependent decline in both occludin and E-cadherin levels. By contrast, TNF-α and iNOS contents were upregulated in imidacloprid-exposed Caco-2 cells. Decrease in tight and adherens junctions proteins indicates that the barrier function of intestine epithelial cells could be damaged by imidacloprid administration. In addition, TNF-α and iNOS upregulation indicates that imidacloprid is potent to activate proinflammatory response in enterocytes. Thus, imidacloprid can affect intestine barrier function through the increase of proinflammatory cytokine production and decrease in adhesiveness of enterocytes. The further assessment of the role of adhesion proteins and inflammatory cytokines in neonicotinoid pesticide cytotoxicity as it affects enterocyte barrier function is required to highlight the risk factor of use of neonicotinoids.

Background and Objective: Phenol red, the pH indicator in cell culture media, influences the expression of cytochrome P450s (CYPs) in cell lines. This study aimed to examine how phenol red modified CYP induction by benzo[a]pyrene and dexamethasone in human hepatocarcinoma (HepG2), colorectal adenocarcinoma (Caco-2) and choriocarcinoma (BeWo) cells. Materials and Methods: The cells (1×105 cells/well in a 24-well plate) were incubated with benzo[a]pyrene (0.1, 1 and 10 μM) or dexamethasone (1, 5 and 10 μM) in either phenol red or phenol red-free media for 24 hrs. The mRNA expression of CYPs was determined by Real-Time Polymerase Chain Reaction (RT/qPCR). Results: Phenol red enhanced expression of benzo[a]pyrene-induced CYP1A2 inHepG2 and BeWo cells and suppressed benzo[a]pyrene-induced CYP2A6 expression in HepG2 and Caco-2 cells, benzo[a]pyrene induced CYP2B6 expression in HepG2 cells and benzo[a]pyrene- and dexamethasone-induced CYP3A4 expression in HepG2 and Caco-2 cells. The expression of CYP3A5 was affected differently in HepG2 and Caco-2 cell lines. Phenol red enhanced benzo[a]pyrene- and dexamethasone-induced CYP3A5 expression in Caco-2 cells but suppressed benzo[a]pyrene- and dexamethasone-induced CYP3A5 expression in HepG2 cells. Conclusion: Phenol red differentially influenced expression of benzo[a]pyrene- and dexamethasone-induced CYP1A2, CYP2A6, CYP2B6, CYP3A4 and CYP3A5 mRNAs in HepG2, Caco-2 and BeWo cells. Therefore, the inclusion of phenol red in cell culture media is of concern in studies of drug and xenobiotic metabolism via CYPs in human cell line models.

Self-microemulsifying drug delivery systems (SMEDDSs) have recently returned to the limelight of academia and industry due to their enormous potential in oral delivery of biomacromolecules. However, information on gastrointestinal lipolysis and trans-epithelial transport of SMEDDS is rare. Aggregation-caused quenching (ACQ) fluorescent probes are utilized to visualize the in vivo behaviors of SMEDDSs, because the released probes during lipolysis are quenched upon contacting water. Two SMEDDSs composed of medium chain triglyceride and different ratios of Tween-80 and PEG-400 are set as models, meanwhile Neoral® was used as a control. The SMEDDS droplets reside in the digestive tract for as long as 24 h and obey first order kinetic law of lipolysis. The increased chain length of the triglyceride decreases the lipolysis of the SMEDDSs. Ex vivo imaging of main tissues and histological examination confirm the trans-epithelial transportation of the SMEDDS droplets. Approximately 2%–4% of the given SMEDDSs are transported via the lymph route following epithelial uptake, while liver is the main termination. Caco-2 cell lines confirm the cellular uptake and trans-epithelial transport. In conclusion, a fraction of SMEDDSs can survive the lipolysis in the gastrointestinal tract, permeate across the epithelia, translocate via the lymph, and accumulate mainly in the liver.

This study aimed to demonstrate usefulness of the fluorophore-labeled bile acid derivative, N-(24-[7-(4-N,N-dimethylaminosulfonyl-2,1,3-benzoxadiazole)]amino-3α,7α,12α-trihydroxy-27-nor-5β-cholestan-26-oyl)-2′-aminoethane sulfonate (tauro-nor-THCA-24-DBD) as a substrate of apical sodium-dependent bile acid transporter (ASBT, SLC10A2), which is expressed at distal ileum for reabsorption of bile acids and to find a novel fluorescence-based method to evaluate ASBT activity. In HPLC analysis, chromatogram of tauro-nor-THCA-24-DBD showed double peaks: R- and S-isomers of the compound. When ASBT was expressed in Xenopus laevis oocytes, their uptakes were higher than those by control oocytes, demonstrating both are transported by ASBT. Therefore, results were analyzed separately as peak 1, peak 2 and sum of them. Concentration dependent uptake of tauro-nor-THCA-24-DBD in ASBT-expressing oocytes was saturable with Km 122 μM and Vmax 1.49 pmol/oocyte/30 min for peak 1, 30.7 μM and 1.34 pmol/oocyte/30 min for peak 2, and 40.6 μM and 2.36 pmol/oocyte/30 min for sum, respectively. These uptakes were decreased in the presence of taurocholic acid and in the Na+ free condition. Furthermore, in Caco-2 cells, tauro-nor-THCA-24-DBD uptake was also Na+-dependent and saturable. Additionally, these uptakes were decreased by elobixibat, a selective ASBT inhibitor. Accordingly, it was concluded that tauro-nor-THCA-24-DBD is a substrate of ASBT and useful to evaluate the intestinal ASBT transport activity.

Arabinoxylans (AX) are gelling polysaccharides with potential applications as colon-targeted biomaterials. Nevertheless, the fermentation of highly cross-linked AX particles (AXP) by colonic bacteria and the effect of its fermentation supernatants on the proliferation of human colon cancer cells have not been investigated so far. In this study, electrosprayed AXP were fermented by Bifidobacterium longum , Bifidobacterium adolescentis , and Bacteroides ovatus . The effect of AXP fermentation supernatant (AXP-fs) on the inhibition of the human colon cancer cell line Caco-2 proliferation was investigated. AXP presented a mean diameter of 533 µm, a spherical shape, and a cross-linking content (dimers and trimers of ferulic acid) of 1.65 µg/mg polysaccharide. After 48 h of bacteria exposure, AXP were only partly fermented, probably due to polymeric network steric hindrance that limits the access of bacterial enzymes to the polysaccharide target sites. AXP partial fermentation was evidenced by a moderate short-chain fatty acid production (SCFA) (23 mM) and a collapsed and disintegrated microstructure revealed by scanning electron microscopy. AXP-fs exerted slight inhibition of Caco-2 cell proliferation (11%), which could be attributed to the SCFA generated during partly polysaccharide fermentation. These findings indicate that electrosprayed AXP are a slow-fermentable biomaterial presenting slight anti-cancer properties and potential application in colon cancer prevention.

Reduction of intestinal glucose absorption might result from either delayed carbohydrate digestion or blockage of glucose transporters. Previously, oxyresveratrol was shown to inhibit α-glucosidase, but its effect on glucose transporters has not been explored. The present study aimed to assess oxyresveratrol-induced inhibition of the facilitative glucose transporter 2 and the active sodium-dependent glucose transporter 1. An aqueous extract of Artocarpus lacucha, Puag Haad, which is oxyresveratrol-enriched, was also investigated. Glucose transport was measured by uptake into Caco-2 cells through either glucose transporter 2 or sodium-dependent glucose transporter 1 according to the culture conditions. Oxyresveratrol (40 to 800 µM) dose-dependently reduced glucose transport, which appeared to inhibit both glucose transporter 2 and sodium-dependent glucose transporter 1. Puag Haad at similar concentrations also inhibited these transporters but with greater efficacy. Oxyresveratrol and Puag Haad could help reduce postprandial hyperglycemic peaks, which are considered to be most damaging in diabetics.

The intestinal barrier is vital for preventing inflammatory bowel disease (IBD). This study aimed to investigate the potential mechanism behind the protective effects of B. dentium N8 on the intestinal barrier using the lipopolysaccharide (LPS)-induced Caco-2 cells model. Our probiotic validation results showed that B. dentium N8 had a higher adhesion ability and a more substantial inhibition effect on Escherichia coli ATCC 25922 adhesion to HT-29 cells. Regarding the epithelial integrity, B. dentium N8 significantly increased the trans-epithelial electrical resistance (TEER) value and decreased the paracellular permeability of Caco-2 cells stimulated by lipopolysaccharide (LPS). In addition, B. dentium N8 significantly increased ZO-1, occludin, and claudin-1 mRNA expression. B. dentium N8 downregulated the mRNA expression level of TLR4 and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6). Furthermore, B. dentium N8 had a better protective effect on the intestinal barrier than that of E7. Comparative genomics of B. dentium N8 and E7 showed B. dentium N8 had the specific genes encoding for adhesion ability and immune system regulation. The findings provide the theoretical basis for B. dentium N8 possessing a protective effect on the intestinal barrier, which indicate that it could be used as a novel therapy for IBD.

Graphical abstract, Highlights • For the first time, phytochemical analysis and antioxidant activity of D. calcarata water and methanol extracts were elucidated. • The D. calcarata water extract demonstrated the highest total phenolic, tannin and flavonoid contents and the best antioxidant activity. • This work also deciphered the anticancer activities and molecular mechanism of D. calcarata extracts against HT-29 and Caco-2 cells. • The Caco-2 cells were significantly more sensitive to D. calcarata ME as compare to HT-29 cells, which were more sensitive to D. calcarata WE. • Methanol extract upregulated p53 expression in both cell lines and selectively induced the expression of STAT5B mRNA., Colorectal cancer is the fourth leading cause of oncological-related deaths and the third most diagnosed malignancy, worldwide. The emergence of chemoresistance is a fundamental drawback of colorectal cancer therapies and there is an urgent need for novel plant-derived therapeutics. In this regard, other compounds are needed to improve the efficacy of treatment against colorectal cancer. Medicinal plants have been effectively used by traditional doctors for decades to treat various ailments with little to no side effects. Drimia calcarata (D. calcarata) is one of the plants used by Pedi people in South Africa to treat a plethora of ailments. However, the anticancer therapeutic use of D. calcarata is less understood. Thus, this study was aimed at evaluating the potential anticancer activities of D. calcarata extracts against human colorectal cancer cells. The phytochemical analysis and antioxidant activity were analysed using LC–MS, DPPH, and FRAP. The inhibitory effects and IC50 values of D. calcarata extracts were determined using the MTT assay. Induction of cellular apoptosis was assessed using fluorescence microscopy, the Muse® Cell Analyser, and gene expression analysis by Polymerase Chain Reaction (PCR). Water extract (WE) demonstrated high phenolic, tannin, and flavonoid contents than the methanol extract (ME). LC–MS data demonstrated strong differences between the ME and WE. Moreover, WE showed the best antioxidant activity than ME. The MTT data showed that both ME and WE had no significant activity against human embryonic kidney Hek 293 cell line that served as non-cancer control cells. Caco-2 cells demonstrated high sensitivity to the ME and demonstrated resistance toward the WE, while HT-29 cells exhibited sensitivity to both D. calcarata extracts. The expression of apoptosis regulatory genes assessed by PCR revealed an upregulation of p53 by ME, accompanied by downregulation of Bcl-2 and high expression of Bax after treatment with curcumin. The Bax gene was undetected in HT-29 cells. The methanol extract induced mitochondrial-mediated apoptosis in colorectal Caco-2 and HT-29 cells and WE induced the extrinsic apoptotic pathway in HT-29 cells. ME downregulated STAT1, 3, and 5B in HT-29 cells. The D. calcarata bulb extracts, therefore, contain potential anticancer agents that can be further targeted for cancer therapeutics.

Background: The mucus integrity and abnormal inflammatory response are the crucial biomarker of inflammatory bowel disease (IBD). Velvet antler (VA) has been used as traditional Chinese medicines for many years. Anti-inflammatory property was demonstrated via suppression of cyclooxygenase-2 and cytokines protein expression. And it has further proved to promote wound healing in streptozotocin-induced diabetic rats model. The aforementioned functionalities of VA extracts may be associated with the treatment of IBD. Thus, the aim of present study was to evaluate the effect of velvet antler water extracts form Formosan Sambar deer (Rusa unicolor swinhoei, SVAE) and red deer (Cervus elaphus, RVAE) on the barrier function and to investigate the possible mechanism using in vitro model. Methods: Human colonic epithelial cell models (Caco-2) were co-cultured with various concentrations of both SVAE and RVAE (250-500 µg mL-1) in dextran sulfate sodium (DSS)-induced colitis model. Trans-epithelial electrical resistance (TEER) value and the macromolecule permeability of Fluorescein isothiocyanate (FITC)-labeled dextran were measured to evaluate the integrity of monolayer of Caco-2. Western blotting was performed for analysis of protein expressions of occludin, Zonula occludens-1 (ZO-1), claudin-1, claudin-2 and myosin light chain kinase (MLCK). The cytotoxicity was conducted by MTT assay. Results: Results indicated that both SVAE and RVAE could enhance integrity of monolayer in dextran sulfate sodium (DSS)-induced colonic epithelial cell model (Caco-2) through reducing the decline of trans-epithelial electrical resistance (TEER) and macromolecule permeability at the concentration of 250 μg mL-1. RVAE significantly increased the expression of tight junction proteins (occludin and ZO-1) while SVAE significantly reduced the activity of MLCK (P < 0.05.). Elevated C-C chemokine ligand 20 (CCL20) production suggested that both SVAE and RVAE could enhance the repair of epithelial cell. Besides, MTT assay revealed that both extracts showed no cytotoxicity. Conclusion: Thus, SVAE and RVAE supplementation may attenuate barrier damage by enhancing the occludin and ZO-1 protein expression, decreasing MLCK expression, promoting the CCL20 production. In the future, animal study is needed for further confirmation.

Excess dietary fat, and associated bile acids, can impair intestinal barrier integrity, produce intestinal or systemic inflammation and promote tumorigenesis. Dietary polyphenols in foods such as berries display antioxidant and other protective effects in many biological systems, but little is known about their protective effects on intestinal epithelial cells exposed to dietary fat. In a Caco-2 cell model of dietary fat-induced intestinal epithelial cell cytotoxicity, oxidative stress and barrier impairment, we investigated the relative protection afforded by an anthocyanin-rich bilberry extract (ARBE) or resveratrol. Exposure of the cells to mixed micelles (MM) of fatty acids and bile acids for 24 h markedly increased intracellular reactive oxygen species (ROS) and mitochondrial superoxide generation, decreased cell viability, increased expression of TNF-α mRNA and disrupted differentiated monolayer integrity. Starting prior to exposure to MM, treatments with ARBE or resveratrol, at polyphenol concentrations from 1.25-20 μM, strongly attenuated MM-induced intracellular ROS generation, and ARBE but not resveratrol decreased mitochondrial superoxide generation. Both ARBE and resveratrol inhibited the MM-induced expression of TNF-α mRNA. In assessments of differentiated monolayer integrity by transepithelial electrical resistance (TEER) and paracellular permeability, resveratrol protection was apparent at 3 h of MM exposure, but less at 6 h and absent by 9 h. In contrast, ARBE largely reversed MM-induced impairment by 9 h, with TEER values reaching 82% of control and the MM-induced paracellular permeability reduced by 78%. While they appeared to act differently, the results suggest that dietary sources of anthocyanins and resveratrol can help confer resistance of intestinal epithelial cells to oxidative stress and inflammation, and resultant barrier dysfunction and tumorigenesis, induced by high dietary fat common in the Western diet.

The current COVID-19 pandemic requires urgent development of effective therapeutics. 5-amino levulinic acid (5-ALA) is a naturally synthesized amino acid and has been used for multiple purposes including as an anticancer therapy and as a dietary supplement due to its high bioavailability. In this study, we demonstrated that 5-ALA treatment potently inhibited infection of SARS-CoV-2, a causative agent of COVID-19, in cell culture. The antiviral effects could be detected in both human and non-human cells, without significant cytotoxicity. Therefore, 5-ALA is worth to be further investigated as an antiviral drug candidate for COVID-19.

Background: Flavonoid compounds are one kind of active ingredients isolated from a traditional Chinese herb Zizyphi spinosae semen (ZSS). Studies have shown that ZSS flavonoids have significant antioxidant effects. Methods: In this study, the Caco-2 cell monolayer model was constructed to investigate the intestinal absorption characteristics and mechanism of Isovitexin (IV), Swertisin (ST), Isovitexin-2''-O-β-D-glucopyranoside (IVG), Spinosin (S), 6'''-p-coumaroylspinosin (6-CS) and 6'''-feruloylspinosin (6-FS). Results: The results of the bidirectional transport assay showed that the six flavonoids have good intestinal absorption in a near-neutral and 37°C environment, and the absorbability in descending order was 6-FS>6- CS>IVG>S>IV>ST. The results of carrier inhibition experiments and transport kinetics indicated that the absorption mechanism of six flavonoids was energy-dependent monocarboxylate transporter (MCT)-mediated active transport. In particular, the para-cellular pathway also participated in the transport of IV, ST, IVG and S. Furthermore, the efflux process of six flavonoids was mediated by P-glycoprotein (P-gp) and multidrug resistance protein (MRP), which may result in a decrease of bioavailability. Conclusion: Our findings provide significant information for revealing the relationship between the intestinal absorption mechanism of flavonoids and its structure as well as laying a basis for the research of flavonoid preparations.

Pterospartum tridentatum is an edible endemic plant common in Portugal. Its flowers are used in culinary and are part of the popular medicine owing to its therapeutic properties. In this study, P. tridentatum flower infusion and hydroethanolic extracts were characterized concerning their phenolic composition and biological potential. By high-performance liquid chromatography method coupled to a diode array detector analysis were detected 13 phenolics. Genistein was the major one. Concerning the biological potential, the hydroethanolic extract was the most active against 2,2-diphenyl-1-picrylhydrazyl● and also as α-glucosidase inhibitor, while the infusion proved to be a remarkable free radical scavenger. Concerning human epithelial colorectal adenocarcinoma (Caco-2) cells, it was observed that both extracts displayed dose-dependent cytotoxicity on the viability of Caco-2 cells, presenting cytotoxic selectivity for these cancer cells when compared to the NHDF normal cell line. Additionally, they also showed protective effects against oxidative stress induced by tert-butyl hydroperoxide on Caco-2 cells. The obtained results suggest that these extracts may be interesting to enrich nutraceutical, pharmaceutical and food industries; however, more studies need to be done for their inclusion in pharmaceutical preparations and/or food additives. PRACTICAL APPLICATION: Pterospartum tridentatum is an endemic plant commonly used in folk medicine due to its depurative and hypoglycaemic properties. For this reason, we decided to determine the phenolic content of infusion and hydroethanolic extracts of P. tridentatum and their biological potential. The obtained results proved that P. tridentatum extracts are a rich source of bioactive compounds and possess great antioxidant and antidiabetic activities, ability to protect human erythrocytes against oxidative damage, inhibiting hemolysis, hemoglobin oxidation, and lipid peroxidation, and to interfere with Caco-2 cells growth and to protect these cells when subject to tert-butyl hydroperoxide oxidative stress conditions.

Intestinal epithelial cells form a barrier between the intestinal lumen and host connective tissues and play an important role in maintaining intestinal nutrient homeostasis. This study investigated effects of Allomyrina dichotoma (rhinoceros beetle) larval extract (ADLE) on the intestinal barrier damage and explored mechanisms for reversing intestinal barrier dysfunction in lipopolysaccharide (LPS)-stimulated Caco-2, human intestinal epithelial cells. LPS reduced intestinal epithelial barrier function by increasing transepithelial electrical resistance, and this effect was significantly attenuated by ADLE treatment. ADLE also significantly countered the inhibition of tight junction-related protein expression in both LPS-induced Caco-2 cells and intestine from HFD-induced mice. Moreover, ADLE significantly decreased expression and production of inflammatory factors, such as iNOS, cox-2, nitric oxide, and cytokines induced by LPS stimulus. Reduction in phosphorylation of adenosine monophosphate-activated protein kinase was averted by ADLE treatment in LPS treated INS-1 cells. Finally, reactive oxygen stress level was decreased and ATP production was increased by ADLE treatment. ADLE appears to display gut health-promoting effects by reducing inflammation and inducing tight junction proteins in Caco-2 cells. Therefore, ADLE might be useful for preventing or treating intestine cell damage in inflammatory bowel disease.

The present study evaluated the protective effect of Lactobacillus acidophilus against the toxicity of the Beauvericin on the Caco-2 cell line. After culturing Caco-2 cells and applying different concentrations of Beauvericin and L. acidophilus individually and in combination, cell viability was assessed by enzyme-linked immunosorbent assay at different times. The results indicate the potential risk of Beauvericin to human health and the interventional role of L. acidophilus, which improved cell viability in the presence of Beauvericin.

Peptide-mediated targeting to colorectal cancer can increase selectivity and specificity of this cancer diagnosis acting as biomarkers. The present work aimed to select peptides using the phage display technique and associate the peptides with polymeric nanospheres in order to evaluate their cytotoxicity and selectivity during cell interaction with Caco-2 human colon tumor cell line. Two peptides identified by phage display (peptide-1 and peptide-2) were synthesized and exhibited purity higher than 84%. Poly(lactic acid)-block-polyethylene glycol nanospheres were prepared by nanoprecipitation and double emulsion methods in order to load the two peptides. Nanoparticles ranged in size from 114 to 150 nm and peptide encapsulation efficiency varied from 16 to 32%, depending on the methodology. No cytotoxic activity was observed towards Caco-2 tumor cell line, either free or loaded peptides in concentrations up to 3 μM at incubation times of 6 and 24 h, indicating safety as biomarkers. Fluorescein isothiocyanate-labeled peptides allowed evaluating selective interactions with Caco-2 cells, where peptide-1 entrapped in nanospheres showed greater intensity of co-localized cell fluorescence, in comparison to peptide-2. Peptide-1 loaded in nanospheres revealed promising to be investigated in further studies of selectivity with other human colon rectal cells as a potential biomarker.Graphical abstract.

Gynostemma pentaphyllum (the immortal herb) has been an important component of Chinese Traditional Medicine for millennia. Recent clinical studies have revealed that the plant exhibits numerous beneficial biological activities, making it of interest to the pharmaceutical industry. An extract of the herb contains over 200 individual secondary metabolites including flavonol glycosides and dammarane saponins. To focus attention on the compounds most likely to be responsible for the biological activities, this study predicts the potential oral bioavailability of nine dammarane saponins and five flavonol glycosides from G. pentaphyllum using the Caco-2 cell monolayer permeability model. Two flavonoids, 8 and 9, and four saponins, 10, 11, 12, and 14, exhibited high permeability across the monolayers. The results indicated that a higher degree of glycosylation-facilitated permeability, suggestive of active transport. This study demonstrates the utility of the Caco-2 permeability assay as a method of identifying possible bioavailable compounds from medicinal herbal extracts.

Rhubarb-Aconite Decoction (RAD), a famous Chinese medicine prescription, has been widely used for treating intestinal injury. However, the effect of RAD on intestinal epithelial cells is unclear. The aim of this study was to investigate the effects of RAD drug-containing serum on the oxidative stress injury and inflammatory response induced by endotoxin (ET) in Caco-2 cells in vitro. Lipid peroxide malondialdehyde (MDA), lactate dehydrogenase (LDH), caspase-11, tumor necrosis factor-α(TNF-α), interleukin-3(IL-3), and cytokeratin (CK)18, adenosine triphosphate (ATP) activity, and intracellular free calcium ion levels were measured. The results showed that ET triggered the activation of caspase-11 and the massive release of TNF-α, increased the inhibitory rate of cell growth, MDA, and LDH expressions in Caco-2 cells. Moreover, RAD drug-containing serum could inhibit caspase-11 activation, decrease the release of TNF-α and IL-3, reduce intracellular free calcium ion, and enhance CK 18 expression and ATP activity. These novel findings demonstrated that ET-induced oxidative stress injury and inflammatory response of Caco-2 cells were improved by RAD drug-containing serum, indicating that RAD may be a good choice for the treatment of intestinal injury.

The mechanisms of cellular absorption and transport underlying the differences between flavonoid aglycones and glycosides and the effect of the structural feature are not well established. In this study, aglycone, mono-, and diglycosides of quercetin and cyanidin were selected to examine the effects of the structural feature on the bioavailability of flavonoids using hexose transporters SGLT1 and GLUT2 in a Caco-2 BBe1 cell model. Cellular uptake and transport of all glycosides were significantly different. The glycosides also significantly inhibited cellular uptake of d-glucose, indicating the involvement of the two hexose transporters SGLT1 and GLUT2 in the absorption, and the potential of the glycosides in lowering the blood glucose level. The in silico prediction model also supported these observations. The absorption of glycosides, especially diglycosides but not the aglycones, was significantly blocked by SGLT1 and GLUT2 inhibitors (phloridzin and phloretin) and further validated in SGLT1 knockdown Caco-2 BBe1 cells.

Wutou-Gancao herb-pair is extensively used to attenuate the toxicity and enhance the efficacy of aconite. In this study, potential synergic mechanism of the herb pair was investigated by utilizing multiple approaches. In silico and in vitro Caco-2 cell models were applied to study the potential binding mode of bioactive ingredients existing in liquorice with P-glycoprotein (P-gp), as well as the inhibition effects on P-gp. Additionally, anti-inflammatory activity of aconitine (AC) combined with active ingredients of liquorice, as well as pharmacokinetic patterns of AC after co-administration was investigated. Anti-inflammatory effect of AC (1 mg/kg) in rats was enhanced in combination with bioactive ingredients of liquorice (10 mg/kg). In the meanwhile, the exposure of AC in vivo was altered, in terms of Cmax and AUC. For instance, the Cmax and AUC were increased to 1.9 and 1.3 folds, respectively, when used in combination with liquiritigenin. The in silico study revealed the potential binding mode with outward facing conformation of P-gp. The resulting data obtained from transport of rhodamine-123 (Rh-123) across Caco-2 cell monolayer further indicated that the function of P-gp was inhibited by chemicals in liquorice. The synergic effect was therefore proposed to be attributed to inhibition of P-gp by liquorice since AC has been demonstrated to be the substrate of P-gp. The resuls revealed that potential synergic mechanism of Wutou-Gancao herb-pair by inhibiting function of key efflux transporter P-gp to enhance the exposure of AC in systematic circulation, and further the anti-inflammatory effect, which helps clarify the compatibility rationale of these two herbs., Graphical abstract Image 1, Highlights • The binding of active ingredients of liquorice with P-gp was revealed by docking. • The permeation of Rho 123 was inhibited by active ingredients of liquorice. • The PK behavior of aconitine was affected by active ingredients of liquorice. • The efficacy of aconitine were enhanced by active ingredients of liquorice.

Background: Caco-2 cells monolayer is one of invitro models to evaluate the translocation capacity of Lactobacillus spp probioticstrains. The translocation is influenced by mucosa permeability of enterocytes as shown by increasing transepithelial resistance (TER) and formation of tight junction proteins. The pore size of the supported permeable membrane used in in vitro assay was one of the crucial factors in performing bacterial translocation assay. Almost no study has been conducted using Caco-2 cells monolayer grown on 8-μm pore size polycarbonate membrane for evaluating probiotics translocation. Therefore this study aimed to determine whether the Caco-2 cells monolayer model was suitable as an in vitro translocation model.Methods: Caco-2 cells monolayer was seeded onto 8-μm collagen-coated polycarbonate membrane insert Transwell®. Differentiation of Caco-2 cells was detected by measuring the TER, while the ZO-1 protein (the tight junction proteins) was detected by immunofluorescence. H2O2 was used as a tight junction disruptive agent. Data were analyzed using SPSS version 23 software to compare the mean of TER measurement between untreated and H2O2-treated Caco-2 cells monolayer.Results: The result showed that the TER of Caco-2 cells monolayer was gradually increasing until day 14, reaching more than 800 ohm.cm2. Furthermore, the ZO-1 protein was successfully detected, indicated the tight junction formation. TER value of H2O2-treated cells showed significantly lower than that of untreated cells (PLactobacillus rhamnosus FBB81 was used for validating the translocation. There was no translocation observed; however, translocation was observed in H2O2-treated cells.Conclusion: Altogether suggests that Caco-2 cells grown on 8 μm-pore size permeable filters could be considered as a suitable in vitro model for probiotics strains translocation.

Previous work showed that the bioavailability of fluoride in dark tea was lower than NaF solution. However, limited information is available indicating the effects of tea components on the fluoride bioavailability. In this study, the effects of the components in tea on the bioavailability of fluoride were evaluated in the Caco‐2 cell line model. Additionally, the mechanism of effect of aluminium on fluoride transport was investigated. The result showed that 10–100 μm of epigallocatechin gallate (EGCG) could not influence fluoride transport. Al³⁺ significantly decreased fluoride transport in both apical‐basolateral and basolateral‐apical directions. Moreover, aluminium could form different forms of aluminium fluoride complexes, which were transported through Caco‐2 cells by different pathways. F⁻ transport was mainly dependent on the paracellular pathway and active transport involving Cl⁻ channels. The paracellular pathway played a predominant role in transport of AlF₃. The paracellular pathway and active transport both participated in AlF₂⁺ transport.

Breast cancer resistance protein (BCRP) is expressed on the apical membrane of small intestinal epithelial cells and functions as an efflux pump with broad substrate recognition. Therefore, quantitative evaluation of the contribution of BCRP to the intestinal permeability of new chemical entities is very important in drug research and development. In this study, we assessed the BCRP-mediated efflux of several model drugs in Caco-2 cells using WK-X-34 as a dual inhibitor of P-glycoprotein (P-gp) and BCRP and LY335979 as a selective inhibitor of P-gp. The permeability of daidzein was high with an apparent permeability coefficient for apical-to-basal transport (PAB) of 20.3 × 10−6 cm/s. In addition, its efflux ratio (ER) was 1.55, indicating that the contribution of BCRP to its transport is minimal. Estrone-3-sulfate and ciprofloxacin showed relatively higher ER values (>2.0), whereas their BCRP-related absorptive quotient (AQBCRP) was 0.21 and 0.3, respectively. These results indicate that BCRP does not play a major role in regulating the permeability of estrone-3-sulfate and ciprofloxacin in Caco-2 cells. Nitrofurantoin showed a PAB of 1.8 × 10−6 cm/s, and its ER was 7.6. However, the AQBCRP was 0.37, suggesting minimal contribution of BCRP to nitrofurantoin transport in Caco-2 cells. In contrast, topotecan, SN-38, and sulfasalazine had low PAB values (0.81, 1.13, and 0.19 × 10−6 cm/s, respectively), and each AQBCRP was above 0.6, indicating that BCRP significantly contributes to the transport of these compounds in Caco-2 cells. In conclusion, Caco-2 cells are useful to accurately estimate the contribution of BCRP to intestinal drug absorption. SIGNIFICANCE STATEMENT We performed an in vitro assessment of the contribution of breast cancer resistance protein (BCRP) to the transport of BCRP and/or P-glycoprotein (P-gp) substrates across Caco-2 cell monolayers using absorptive quotient, which has been proposed to represent the contribution of drug efflux transporters to the net efflux. The present study demonstrates that the combined use of a BCRP/P-gp dual inhibitor and a P-gp selective inhibitor is useful to estimate the impact of BCRP and P-gp on the permeability of tested compounds in Caco-2 cells.

Activation of the transient receptor potential (TRP) channel TRPA1 by cinnamaldehyde has been shown to stimulate serotonin release in enterochromaffin QGP-1 cells. However, the impact of cinnamaldehyde on serotonin release in enterocytes is less well understood. In addition, since the neurotransmitter serotonin plays a regulatory role in a large variety of gastrointestinal and metabolic functions, it is of interest to study which structural characteristics determine cinnamaldehyde-induced serotonin release by enterocytes. Thus, the present study analyzed serotonin release in differentiated Caco-2 cells as a model for enterocytes in comparison to enterochromaffin QGP-1 cells after stimulation with cinnamaldehyde and 17 naturally occurring structurally related compounds by means of a serotonin ELISA. Stimulation with cinnamaldehyde induced a dose-dependent increase in serotonin release starting from 0.5 mM in both cell lines, with a larger effect size in Caco-2 enterocytes compared to that in QGP-1 enterochromaffin cells. Serotonin release in Caco-2 cells induced by additional 17 structurally related compounds correlated with serotonin release in QGP-1 cells, showing the highest effects for coniferylaldehyde with a 15.84 ± 3.23-fold increase in Caco-2 cells, followed by the parent compound cinnamaldehyde (13.45 ± 2.15), cinnamyl alcohol (6.68 ± 1.08), and α-methyl-cinnamaldehyde (6.59 ± 0.93). Analysis of structural and molecular characteristics that modulate serotonin release in Caco-2 enterocytes revealed that the ability of a compound to activate TRPA1, demonstrated by means of HEK293 cells transiently expressing hTRPA1, is a decisive factor to stimulate serotonin release in Caco-2 enterocytes, preferring small, electrophilic compounds with a lower polar surface area. In addition, blocking of TRPA1 using 30 μM AP-18 significantly reduced the cinnamaldehyde-induced serotonin release by 30.0 ± 5.24%, confirming a TRPA1-dependent component in serotonin release by Caco-2 cells.

In previous studies we demonstrated that zinc stimulates iron uptake in intestinal Caco-2 cells via Zinc-PI3K-IRP2-DMT1 axis. In the current study we investigated the effect of zinc on basolateral iron release and characterized the associated mechanisms. In Caco-2 cells grown on permeable supports, zinc induced iron transport and expression of DMT1, HEPH mRNA and protein, but not that of FPN1. LY294002, an inhibitor of PI3K, inhibited the zinc-induced iron transport, DMT1, HEPH mRNA and protein expression. In addition, LY294002 also inhibited the basal expression of HEPH and FPN1 resulting in blockade of iron egress from cells. In addition, siRNA-silencing of HEPH led to inhibition of both zinc-induced and basal iron transport. Conversely, TPEN, a chelator of zinc, inhibited iron uptake, DMT1, HEPH and FPN1 mRNA and protein expression. These results suggest that intestinal cell zinc status is a critical determinant of iron absorption and effects are mediated via activation of PI3K. Further, PI3K pathway appears to selectively modulate the expression of iron transporters and iron absorption, therefore this might serve as a therapeutic target in iron overload disorders.

Acetylated compounds prepared from naphthoquinones have been reported as antitumoral prodrugs. Exploring the synthetic versatility of the naphthoquinone and triazolic nuclei, herein we report a simple and efficient synthetic route to prepare a series of sixteen prodrugs prototype of 1,2,3-triazoles-naphthoquinodoic acetyl derivatives. The compounds 10a-10h and 11a-11h were obtained by oxidative cycloaddition reaction between lawsone and 4-vinyl-1H-1,2,3-triazoles promoted by ceric ammonium nitrate (CAN) in alkaline medium followed by reductive acetylation of the quinones in excess of metallic zinc and acetic anhydride in yields up to > 98%. All derivatives revealed to be hemocompatible and the compound 11e exhibited the most promising profile against Caco-2 cells showing the higher selectivity index. Molecular docking suggests that these compounds could exert their cytotoxic activity through inhibition of one topoisomerase II isoform, at least.

The stability of tea phenolic compounds is influenced by pH value and digestive processes. However, the complex mixture of constituents in tea may modulate the stability of these compounds during digestion. In this study, tea infusions obtained from green, black, and Oolong tea leaves were exposed to in vitro simulated gastrointestinal digestion, and the stability of ( +)-catechin, caffeine, (−)-epicatechin, epigallocatechin-3-gallate (EGCG), and gallic acid was compared to that of isolated compounds. Changes in antioxidant activity were also evaluated by means of DPPH assay and in a H2O2-induced in vitro oxidative stress model, using Caco-2 cells. The stability of teas antioxidant constituents was different when using teas extract, compared to the reference compound alone, with the total phenolic content being more stable in extracts containing them in higher amount. EGCG degradation correlated well with changes in the DPPH inhibition assay, confirming its pivotal role in the antioxidant activity of tea. Differently, the antioxidant effect in the in vitro cell-based model was much more related to the initial total phenolic content of the extracts, with green tea being more effective than black tea and Oolong tea. Moreover, the antioxidant activity of teas was strongly affected by gastrointestinal digestion. Taken together, these findings suggest a protective role of teas phytocomplex against gastrointestinal digestion of antioxidant constituents. In conclusion, the effect of gastrointestinal digestion on the antioxidant activity of tea should be taken into account, as this may be different from one extract to another and information on the stability of active constituents cannot be extrapolated from data obtained using single compounds.

Scope: Fructose exacerbates post-prandial hypertriacylglycerolaemia. This may be partly due to increased enterocyte de novo lipogenesis (DNL). It is unknown whether this is concentration-dependent or whether fructose has a greater effect on lipid synthesis than glucose. The dose-dependent effects of fructose and glucose on DNL and de novo triacylglycerol (TAG)-glycerol synthesis were investigated in an enterocyte model, Caco-2 cells. Methods and results: Caco-2 cells were treated for 96h with 5mM, 25mM or 50mM fructose or glucose, or 12.5mM fructose/12.5mM glucose mix. DNL was measured following addition of [13C2]-acetate to apical media. In separate experiments, [13C6]-fructose and [13C6]-glucose were used to measure DNL and de novo TAG-glycerol synthesis. DNL from [13C2]-acetate was detected following all treatments, with greater amounts in intracellular than secreted (media) samples (all P

This work describes an untargeted analytical approach for the screening, identification, and characterization of the trans-epithelial transport of green tea (Camellia sinensis) catechin extracts with in vitro inhibitory effect against the SARS-CoV-2 papain-like protease (PLpro) activity. After specific catechin extraction, a chromatographic separation obtained six fractions were carried out. The fractions were assessed in vitro against the PLpro target. Fraction 5 showed the highest inhibitory activity against the SARS-CoV-2 PLpro (IC50 of 0.125 μg mL−1). The untargeted characterization revealed that (−)-epicatechin-3-gallate (ECG) was the most abundant compound in the fraction and the primary molecule absorbed by differentiated Caco-2 cells. Results indicated that fraction 5 was approximately 10 times more active than ECG (IC50 value equal to 11.62 ± 0.47 μg mL−1) to inhibit the PLpro target. Overall, our findings highlight the synergistic effects of the various components of the crude extract compared to isolated ECG.

To unearth suitable complexes that are capable of inhibiting the growth of MDA-MB-468 and Caco-2 cells, 2,2′-bipyrimidine-based luminescent Ru(II)/Ir(III)–arene monometallic and homo- and hetero-bimetallic complexes were synthesized. The complex [(η6-p-cymene)(η5-Cp*)RuIIIrIIICl2(K2-N,N-bipyrimidine)](PF6)2 [LRuIr] exhibited the best potency in both cells along with good GSH stability and strong binding efficacy with the biomolecules. The apoptotic event occurred in MDA-MB-468 cancer cells via cell cycle arrest.

This study evaluates in vitro cytotoxic and antiproliferative activity on human colon tumor cell line Caco-2 (ATCC-HTB-37) of a standardized (5 mg GAE/mL) ethanolic extract from Stokesia laevis (Slae26), of five polyphenols compounds (reference substances, ref.), namely luteolin-7-O-glucoside, luteolin-8-C-glucoside, caffeic acid, gentisic acid, and p-aminobenzoic acid (PABA), as well as of Slae26 combinations with the five reference substances, 1:1 mass rate (GAE, ref.). Cell viability studies (MTS test) have revealed IC50 values of 36 μg GAE/mL in the case of Slae26 ethanolic extract, while Slae26 combinations with the five phenolics indicated IC50 values around 5 μg GAE/mL. In silico docking studies on the molecular targets human tankyrase 1 (TNKS1) and human tankyrase 2 (TNKS2) in complex with their native ligands, Co-crystallized 3J5A and Co-crystallized FLN, indicated score values of −104.15 and −76.97, respectively; in the series of the reference compounds studied, luteolin-7-O-glucoside was revealed with the best score values on both molecular targets (−80.49 and −85.17), together signifying real antiproliferative potential against human colon cancer of Slae26, of luteolin-7-O-glucoside, and of Slae26 combinations with all food-related bioactive compounds tested.

Gastrointestinal cancers, particularly colorectal cancer, are mainly influenced by the dietary factor. A diet rich in fruits and vegetables can help to reduce the incidence of colorectal cancer thanks to the phenolic compounds, which possess antimutagenic and anticarcinogenic properties. Polyphenols, alongside their well-known antioxidant properties, also show a pro-oxidative potential, which makes it possible to sensitize tumor cells to oxidative stress. HO-1 combined with antioxidant activity, when overexpressed in cancer cells, is involved in tumor progression, and its inhibition is considered a feasible therapeutic strategy in cancer treatment. In this study, the effects of protocatechuic acid (PCA) on the viability of colon cancer cells (CaCo-2), annexin V, LDH release, reactive oxygen species levels, total thiol content, HO-1, γ-glutamylcysteine synthetase, and p21 expression were evaluated. PCA induced, in a dose-dependent manner, a significantly reduced cell viability of CaCo-2 by oxidative/antioxidant imbalance. The phenolic acid induced modifications in levels of HO-1, non-proteic thiol groups, γ-glutamylcysteine synthetase, reactive oxygen species, and p21. PCA induced a pro-oxidant effect in cancer cells, and the in vitro pro-apoptotic effect on CaCo-2 cells is mediated by the modulation of redox balance and the inhibition of the HO-1 system that led to the activation of p21. Our results suggest that PCA may represent a useful tool in prevention and/or therapy of colon cancer.

International audience; Adhesion to the digestive mucosa is considered a key factor for bacterial persistence within the gut. In this study, we show that Ruminococcus gnavus E1 can express the radA gene, which encodes an adhesin of the MSCRAMMs family, only when it colonizes the gut. The RadA N-terminal region contains an all-β bacterial Ig-like domain known to interact with collagens. We observed that it preferentially binds human immunoglobulins (IgA and IgG) and intestinal mucins. Using deglycosylated substrates, we also showed that the RadA N-terminal region recognizes two different types of motifs, the protein backbone of human IgG and the glycan structure of mucins. Finally, competition assays with lectins and free monosaccharides identified Galactose and N-Acetyl-Galactosamine motifs as specific targets for the binding of RadA to mucins and the surface of human epithelial cells.