Epithelial Colorectal Adenocarcinoma Research Articles

Effect of extracts from eggs of Helix aspersa maxima and Helix aspersa aspersa snails on Caco-2 colon cancer cells.

BackgroundColorectal cancer is the third most commonly diagnosed cancer. Natural compounds, administered together with conventional chemotherapeutic agent(s) and/or radiotherapy, may be a novel element in the combination therapy of this cancer. Considering the anticancer properties of compounds derived from different tissues of various snail species confirmed earlier, the purpose of the present research was to evaluate the effect of extracts from eggs of Helix aspera maxima and Helix aspersa aspersa snails, and fractions of extracts containing particles of different molecular weights on Caco-2 human epithelial colorectal adenocarcinoma cells.MethodsThe extracts and fractions were analyzed for antioxidant activity, phenols and total carbohydrates using colorimetric methods. Lipid peroxidation products and glutathione in eggs were also examined using these methods. Crude protein and fat in eggs were determined. Molecular weights of egg proteins and glycoproteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Astaxanthin, selected vitamins and amino acids in eggs were measured using liquid chromatography methods, and minerals by emission spectroscopy, mass spectrometry or X-ray fluorescence. The action of extracts on the cell viability was determined by the MTT (methylthiazolyldiphenyl-tetrazolium bromide) test, based on the mitochondrial oxidative activity, after 24 and 72 h of treatment. The influence of fractions on the cell viability was assayed after 24 h. The effect of extracts on the percentage of live and dead cells was evaluated by the trypan blue assay, in which live cells exclude trypan blue, while dead cells take up this dye, after 12, 24, 48 and 72 h of treatment. Their influence on the integrity of cell membranes was determined based on the activity of LDH (lactate dehydrogenase), released from damaged cells, after 24 and 72 h of treatment. Then, the effect of extracts on the content of lipid peroxidation products in cells was examined using colorimetric method, after 24 h of treatment. Their influence on types of cell death was determined by flow cytometry, after this time.ResultsThe extracts and their fractions containing molecules <3 kDa decreased the cell viability, after 24 h of treatment. The extracts reduced the percentage of live cells (also after 48 h), increased the degree of cell membrane damage and the amount of lipid peroxidation products, induced apoptosis and reduced necrosis.ConclusionsAntioxidants, phenols, lipid peroxidation products, anticancer peptides, restriction of methionine, appropriate ratio of essential amino acids to non-essential amino acids, vitamin D3, Ca, Mg, S, Cu, Mn, Zn, Se and other bioactive compounds comprised in the extracts and their additive and synergistic effects may have influenced Caco-2 cells. Natural extracts or the chemical compounds contained in them might be used in the combination therapy of colorectal cancer, which requires further research.

Folic Acid and Chitosan-Functionalized Gold Nanorods and Triangular Silver Nanoplates for the Delivery of Anticancer Agents.

BackgroundAdvances in the field of nanotechnology have shed light on the applications of nanoparticles for cancer treatment.MethodsFolic acid and chitosan-functionalized gold nanorods (FACS-R) and triangular silver nanoplates (FACS-T) were synthesized and their properties were elucidated by UV-visible spectrophotometry, Fourier-transform infrared spectroscopy, field emission transmission electron microscopy and high-resolution X-ray diffraction.ResultsThe average size of the FACS-R was determined to be a transverse length of 13.1 ± 1.8 nm and a longitudinal length of 47.2 ± 8.9 nm with an aspect ratio of 3.6. The average size of FACS-T was measured to be 31.8 ± 7.7 nm. Colloidal solutions of FACS-R and FACS-T were stable on the shelf at ambient temperature for 14 days in the dark. Anticancer agents were encapsulated in FACS-R and FACS-T. FACS-T showed a higher encapsulation efficiency with docetaxel, paclitaxel and diallyl disulfide than FACS-R. The cell viability on human gastric adenocarcinoma cells (AGS), human epithelial cervix adenocarcinoma cells (HeLa) and human colorectal adenocarcinoma cells (HT-29) after treatment with anticancer agent-encapsulated FACS-R and FACS-T was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Interestingly, paclitaxel-encapsulated FACS-R and FACS-T showed the highest percentages of early and late apoptosis on HeLa cells. A cell cycle analysis demonstrated increased G2/M arrest on HeLa cells with docetaxel and paclitaxel-encapsulated FACS-R and FACS-T. The FACS-T induced more G2/M arrest on HeLa cells than the FACS-R. To assess applications in near-infrared photothermal therapy (PTT), the cell viability on HeLa cells with the anticancer agent-encapsulated FACS-R and FACS-T was assessed in the presence or absence of 808 nm laser irradiation. The results showed that 808 nm laser irradiation significantly decreased cell viability.ConclusionCollectively, the triangular silver nanoplates were more effective than the gold nanorods for PTT. We believe that as-prepared nanoparticles have remarkable features and will become promising future nanomedicine.

Gastrointestinal Stability and Cytotoxicity of Bacteriocins From Gram-Positive and Gram-Negative Bacteria: A Comparative in vitro Study.

Bacteriocins are receiving increased attention as potent candidates in food preservation and medicine. Although the inhibitory activity of bacteriocins has been studied widely, little is known about their gastrointestinal stability and toxicity toward normal human cell lines. The aim of this study was to evaluate the gastrointestinal stability and activity of microcin J25, pediocin PA-1, bactofencin A and nisin using in vitro models. In addition cytotoxicity and hemolytic activity of these bacteriocins were investigated on human epithelial colorectal adenocarcinoma cells (Caco-2) and rat erythrocytes, respectively. Pediocin PA-1, bactofencin A, and nisin were observed to lose their stability while passing through the gastrointestinal tract, while microcin J25 is only partially degraded. Besides, selected bacteriocins were not toxic to Caco-2 cells, and integrity of cell membrane was observed to remain unaffected in presence of these bacteriocins at concentrations up to 400 μg/mL. In hemolysis study, pediocin PA-1, bactofencin A, and nisin were observed to lyse rat erythrocytes at concentrations higher than 50 μg/mL, while microcin J25 showed no effect on these cells. According to data indicating gastrointestinal degradation and the absence of toxicity of pediocin PA-1, bactofencin A, and microcin J25 they could potentially be used in food or clinical applications.

Antitumor Activity of Zinc Nanoparticles Synthesized with Berberine on Human Epithelial Colorectal Adenocarcinoma (Caco-2) Cells through Acting on Cox-2/NF-kB and p53 Pathways

Antitumor Activity of Zinc Nanoparticles Synthesized with Berberine on Human Epithelial Colorectal Adenocarcinoma (Caco-2) Cells through Acting on Cox-2/NF-kB and p53 Pathways

Design, synthesis, anticancer activity and molecular docking analysis of novel dinitrophenylpyrazole bearing 1,2,3-triazoles

• Novel dinitrophenylpyrazole bearing 1,2,3-triazol molecular hybrids were reported. • Its structure elucidation was established via different spectroscopic techniques. • Synthesized compounds were evaluated anti-cancer activity ( in vitro ) against three human cell lines. • Compounds 9e, 9f , and 9 h showed the high potency growth inhibitory activity against the HeLa cell line. • Binding interaction were also validated with molecular docking studies. A novel series of dinitrophenylpyrazole bearing triazole scaffolds were designed and synthesized with appreciable yields. The structures of all the target molecules ( 9a–j ) were confirmed and characterized by 1 H NMR, 13 C NMR, FT-IR and HR-MS spectral techniques. These novel molecules were tested for their anti-cancer activity ( in vitro) using three tumor cell lines indicating breast adenocarcinoma (MCF-7), cervical carcinoma (HeLa), and human epithelial colorectal adenocarcinoma (Caco-2) via the MTT approach. Compounds 9e, 9f, and 9 h showed the high potency growth inhibitory activity against the HeLa cell line (IC 50 = 4.0 μM, 5.0 μM and 6.0 μM) higher than that of Combretastatin-A4 (IC 50 = 9.0 μM) as reference drug. Besides, compound 9 h was found to be highly potent against MCF-7 cell line IC 50 value of 8.0 μM. Additionally, in this manuscript, the structure-activity relationship (SAR) analysis is reported. Molecular docking studies also confirm the experimental findings and explains the most probable interaction pattern.

Freeze-Drying Ethylcellulose Microparticles Loaded with Etoposide for In Vitro Fast Dissolution and In Vitro Cytotoxicity against Cancer Cell Types, MCF-7 and Caco-2

The aim of this study was to improve the solubility of etoposide–ethylcellulose (ET–ETO) microparticles using the freeze-drying technique. Ethylcellulose (EC) microparticles loaded with etoposide (ETO) were prepared with different drug–polymer molar ratios of 1:1, 1:3, 1:6, and 1:20 by the solvent evaporation method. The size of the prepared microparticles was 0.088 µm. The results showed that the amount of ETO encapsulated into the microparticles was 387.3, 365.0, 350.0, and 250 µg/50 mg microparticles for microparticles with drug–polymer ratios of 1:1, 1:3, 1:6, and 1:20, respectively. The FT-IR spectra showed no chemical interaction between ETO and the polymer in the solid state. The results obtained from the dissolution experiment showed that the freeze-dried microparticles were stable in 0.1 N HCl (gastric pH) for 2 h. At pH 7.4, the ETO release was 60 to 70% within the first 15 min and approximately 100% within 30 min. Results from the application of different dissolution models showed that the equations that best fit the dissolution data for the ET–ETO microparticles at pH 7.4 were the Higuchi and Peppas model equations. The in vitro cytotoxicity assay of free ETO and freeze-dried microspheres prepared in this study with a drug–polymer ratio of 1:1 was performed in two mammalian cancer cell lines, MCF-7 (for bone cancer of the mammary organ) and Caco-2 (for mammalian epithelial colorectal adenocarcinoma). The results showed that the half-maximal inhibitory concentrations (IC50 values) for ETO and freeze-dried ET–ETO microparticles were 18.6 µM and 27.1 µM, respectively. In conclusion, freeze-dried ET–ETO is a promising formulation for developing a fast-dissolving form of ETO with a significant antiproliferative activity against the tested cell lines used in this study. It is a promising formulation for local duodenal area targeting.

Synthesis, X-ray crystal structures and anticancer studies of four Pd(II) dithiocarbamate complexes

Bis(piperidinyldithiocarbamato)palladium(II) ([Pd(Pipdtc) 2 ]), bis(1-phenylpiperazinyldithiocarbamato)palladium(II) [Pd(1-PhPzdtc) 2 ], bis(phenyldithiocarbamato)palladium(II) ([Pd(Phdtc) 2 ]) and bis(4-benzylpiperidinyl)palladium(II) ([Pd(4-BenzPipdtc) 2 ]) were prepared and characterized by elemental analysis, spectroscopic techniques and single crystal X-ray crystallography. Molecular structures of the Pd(II) complexes revealed square planar PdS 4 geometry in which each Pd(II) is coordinated to two chelating dithiocarbamato anions. [Pd(Pipdtc) 2 ] and [Pd(1-PhPzdtc) 2 ] crystallized in monoclinic P2 1/c space group while [Pd(Phdtc) 2 ] and [Pd(4-BenzPipdtc) 2 ] crystallized in triclinic P-1 space group. Cytotoxic studies of the dithiocarbamate ligands and corresponding Pd(II) dithiocarbamate complexes were tested against three human cell lines: Cervical cancer, breast adenocarcinoma, and epithelial colorectal adenocarcinoma at four different concentrations. All the compounds displayed high to moderate potency against the cell lines. [Pd(Pipdtc) 2 ] and [Pd(Phdtc) 2 ] are very potent against the three cell lines and are more active than the corresponding dithiocarbamate ligands whereas the anticancer potency of 1-PhPzdtc-Na and 4-BenzPipdtc-Na are generally higher than the corresponding Pd(II) complexes.

Hydrothermal Processing and In Vitro Simulated Human Digestion Affects the Bioaccessibility and Bioactivity of Phenolic Compounds in African Pumpkin (Momordica balsamina) Leaves.

The African pumpkin (Momordica balsamina) contains bioactive phenolic compounds that may assist in reducing oxidative stress in the human body. The leaves are mainly consumed after boiling in water for a specific time; this hydrothermal process and conditions of the gastrointestinal tract may affect the presence and bioactivity of phenolics either positively or negatively. In this study, the effects of hydrothermal processing (boiling) and in vitro simulated human digestion on the phenolic composition, bioaccessibility and bioactivity in African pumpkin were investigated in comparison with those of spinach (Spinacia oleracea). A high-resolution ultra-performance liquid chromatography, coupled with diode array detection, quadrupole time-of-flight and mass spectrometer (UPLC-DAD-QTOF-MS) was used to profile phenolic metabolites. Metabolites such as 3-caffeoylquinic acid, 5-caffeoylquinic acid, 3,4-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid were highly concentrated in the boiled vegetable extracts compared to the raw undigested and all digested samples. The majority of African pumpkin and spinach extracts (non-digested and digested) protected Deoxyribonucleic acid (DNA), (mouse fibroblast) L929 and human epithelial colorectal adenocarcinoma (Caco-2) cells from 2,2′-Azobis(2-methylpropionamidine) dihydrochloride (AAPH)-induced oxidative damage. From these results, the consumption of boiled African pumpkin leaves, as well as spinach, could be encouraged, as bioactive metabolites present may reduce oxidative stress in the body.

Adherent cell thawing by infrared radiation

Cryopreservation of adherent cells is crucial for commercial cell therapy technology, including effective distribution and storage. Fast thawing has been shown to increase cell recovery in vitrified samples. Previously, radiofrequency (RF) has been investigated as a heating source on large samples, either with or without magnetic particles. Also, laser heating with the aid of dye or nanoparticles has been utilized on sub–millimeter samples successfully. For slow freezing cryopreservation methods, the influence of rate of thawing on viability is less clear. Cryopreservation of surface adhered cells result in many cases in detachment from the surface. We illustrate how intense infrared radiation from a focused halogen illuminator accelerates thawing. We show that two epithelial cell lines, retinal pigment epithelium cells and heterogeneous human epithelial colorectal adenocarcinoma cells, can be effectively cryopreserved and recovered using a combination of slow freezing and fast thawing under infrared illumination. We were able to successfully thaw samples, of 2–4 mm thick, including the media, on the order of a second, providing a heating rate of thousands of Kelvin per minute. Under optimal conditions, we observed higher post–thawing cell viability rates and higher cell adhesion with infrared thawing than with water bath thawing. We suggest that bulk warming with infrared radiation has an advantage over surface warming of surface–attached cells, as it alleviates cell stress during the process of thawing. These findings will pave the way for novel approaches to treating substrate–adhered cells and 3D scaffolds with cells and organoids. This technology may serve as a crucial component in lab–on–chip systems for medical testing and therapeutic use.

Cytotoxic steroidal glycosides from Polygonatum odoratum (Mill.) Druce

Twenty-five steroidal glycosides including eight undescribed compounds which were named as polygonatumosides H–O, were isolated from the 70 % EtOH extract of rhizomes of Polygonatum odoratum (Mill.) Druce (Asparagaceae). Their structures were elucidated by extensive spectroscopic analyses and chemical methods. The isolated compounds were evaluated cytotoxicity against three human cancer cell lines: human non-small cell lung cancer (A549), human epithelial colorectal adenocarcinoma (Caco2), and human hepatocellular carcinoma (HepG2) cells. Five compounds showed cytotoxicity against these cell lines with IC50 values in the range of 1.7–30.8 μM.

Synergistic cytotoxicity of bromoacetic acid and three emerging bromophenolic disinfection byproducts against human intestinal and neuronal cells

Halogenated disinfection byproducts (halo-DBPs) are drinking water contaminants of great public health concern. Nine haloaliphatic DBPs have been regulated by the U.S. Environmental Protection Agency and various halophenolic compounds have been identified as emerging DBPs. In this study, we evaluated the cytotoxic interactions of the regulated bromoacetic acid and three emerging bromophenolic DBPs, i.e., 2,4,6-tribromophenol, 3,5-dibromo-4-hydroxybenzoic acid, and 3,5-dibromo-4-hydroxybenzaldehyde. Cytotoxicity was measured for each DBP individually as well as each of their mixtures using in vitro human epithelial colorectal adenocarcinoma (Caco-2) and neuroblastoma (SH-SY5Y) cells. Concentration addition (CA) model and isobolographic analysis were employed to characterize the interactions among the DBPs. Our results show that the cytotoxicity of four bromo-DBPs against both cell-types followed the descending rank order of bromoacetic acid > 2,4,6-tribromophenol > 3,5-dibromo-4-hydroxybenzaldehyde > 3,5-dibromo-4-hydroxybenzoic acid. Compared with the toxicity data in literature, our finding that bromoacetic acid showed higher cytotoxicity than bromophenolic DBPs was consistent with the results from Chinese hamster ovary cells (a commonly used in vitro model of DBP toxicological studies); but different from the results obtained from in vivo biological models. Significantly, with CA model prediction, we found that mixtures of four bromo-DBPs exhibited synergistic cytotoxic effects on both human cell types. Isobolographic analysis of binary DBP mixtures revealed that, for Caco-2 cells, bromoacetic acid, 2,4,6-tribromophenol, and 3,5-dibromo-4-hydroxybenzoic acid induced synergism; for SH-SY5Y cells, bromoacetic acid induced synergism with all three bromophenolic DBPs. The production of reactive oxidative species (ROS) induced by DBP mixtures could be an important reason for the synergistic cytotoxicity.

Valorisation of Prunus avium L. By-Products: Phenolic Composition and Effect on Caco-2 Cells Viability.

Prunus avium L. by-products, such as stems, leaves, and flowers, are used in folk medicine to prevent and treat some diseases. However, their phenolic composition and in vitro bioactivities against tumor cells are poorly known. In this work, we compared the phenolic profile and the biological potential of aqueous infusions and hydroethanolic extracts of P. avium leaves, stems, and flowers from Saco cultivar, collected from the Fundão region (Portugal). Among the fifty-two phenolic compounds tentatively identified by HPLC-DAD-ESI/MSn, the hydroxycinnamic acids were the most abundant. Both extracts of stems revealed a higher activity against DPPH•. Meanwhile, hydroethanolic extracts from stems and flowers and aqueous infusions of flowers were the most effective in inhibiting the growth of the human epithelial colorectal adenocarcinoma (Caco-2) cells at concentrations above 200 μg/mL. More detailed knowledge about the phenolic composition and health-promoting properties of Portuguese P. avium by-products allows for increasing the biological and commercial value of these bio-wastes, which may have a positive impact on food and pharmaceutical industries, as on the valorization of the local economy.

Cytotoxic Effect of Silver Nanoparticles on Colorectal Adenocarcinoma Cell Line

Silver nanoparticles (Ag-NPs) with antimicrobial activity are by far the most commercialized nano-compound. However, there is limited information about antitumor potential. Therefore, the present study was aimed to observe the effect of small-dimensional Ag-NPs (20Nm) on human epithelial colorectal adeno-carcinoma (Caco-2) cells. Caco-2 cells were exposed to Ag-NPs with different concentrations (1–15μg/mL) for 24 and 48 hours and cytotoxicity was analyzed by MTT assay and IC50 was calculated. In addition, flow cytometry was used to determine the effect of Ag-NPs on the cell cycle progression and on apoptosis. An MTT assay revealed that following exposure to Ag-NPs the viability of Caco-2 cells decreased, and the viability decreased further with increasing exposure time. The IC50 was 7.85 µg/ml and 4.92 µg/ml at 24 and 48 h respectively after exposure. The flow cytometry technique revealed that Ag-NPs induced a significant increase in the percentage of cells in G1 phase of the cell cycle and of apoptotic cells. In conclusion, Ag-NPs exhibited dose- and time-dependent cytotoxicity, and these results have signposted that Ag-NPs were promising nanoparticles which could be a potential candidate in treating colorectal cancer.

Effect of Akkermansia muciniphila, Faecalibacterium prausnitzii, and Their Extracellular Vesicles on the Serotonin System in Intestinal Epithelial Cells.

The gastrointestinal (GI) tract is an essential reservoir of serotonin or 5-hydroxytryptamine (5-HT), which possesses a set of bacterial species communities. Intestinal microbiota has the ability to modulate the host's serotonin system. In this regard, we evaluated the effect of Akkermansia muciniphila and Faecalibacterium prausnitzii along with their extracellular vesicles (EVs) on serotonin system-related genes in human epithelial colorectal adenocarcinoma (Caco-2) cells. The differentiated Caco-2 cells were treated with A. muciniphila and F. prausnitzii with the multiplicity of infection ratio of 1 and 10 and the EV concentration of 1μg/mL and 50μg/mL, respectively. After 24h, the serotonin level was quantified using an ELISA kit and also the gene expression of serotonin system-related genes was examined using the quantitative real-time PCR method. According to the results, treatment with A. muciniphila and F. prausnitzii-derived EVs increased the serotonin level, while none of the bacteria could affect the serotonin level in the Caco-2 cells. Both bacteria had significant effects on the mRNA expression of serotonin system-related genes in the Caco-2 cells. Moreover, we observed that A. muciniphila and F. prausnitzii-derived EVs could impact the expression of major genes involved in the serotonin system. Our findings showed that A. muciniphila and F. prausnitzii along with their EVs could modulate serotonin system-related genes; hence, they may be useful in microbiota modulation therapies to maintain the homeostasis of the serotonin system.

The Interaction of the Flavonoid Fisetin with Human Glutathione Transferase A1-1.

Glutathione transferases (GSTs) are a family of Phase II detoxification enzymes that are involved in the development of the multidrug resistance (MDR) mechanism in cancer cells and therefore affect the clinical outcome of cancer chemotherapy. The discovery of nontoxic natural compounds as inhibitors for GSTs is a promising approach for chemosensitizing and reversing MDR. Fisetin (7,3′,4′-flavon-3-ol) is a plant flavonol present in many plants and fruits. In the present work, the interaction of fisetin with human glutathione transferase A1-1 (hGSTA1-1) was investigated. Kinetic analysis revealed that fisetin is a reversible inhibitor for hGSTA1-1 with IC50 1.2 ± 0.1 μΜ. It functions as a mixed-type inhibitor toward glutathione (GSH) and as a noncompetitive inhibitor toward the electrophile substrate 1-chloro-2,4-dinitrobenzene (CDNB). In silico molecular modeling and docking predicted that fisetin binds at a distinct location, in the solvent channel of the enzyme, and occupies the entrance of the substrate-binding sites. Treatment of proliferating human epithelial colorectal adenocarcinoma cells (CaCo-2) with fisetin causes a reduction in the expression of hGSTA1-1 at the mRNA and protein levels. In addition, fisetin inhibits GST activity in CaCo-2 cell crude extract with an IC50 (2.5 ± 0.1 μΜ), comparable to that measured using purified recombinant hGSTA1-1. These actions of fisetin can provide a synergistic role toward the suppression and chemosensitization of cancer cells. The results of the present study provide insights into the development of safe and effective GST-targeted cancer chemosensitizers.

A SARS-CoV-2 cytopathicity dataset generated by high-content screening of a large drug repurposing collection

SARS-CoV-2 is a novel coronavirus responsible for the COVID-19 pandemic, in which acute respiratory infections are associated with high socio-economic burden. We applied high-content screening to a well-defined collection of 5632 compounds including 3488 that have undergone previous clinical investigations across 600 indications. The compounds were screened by microscopy for their ability to inhibit SARS-CoV-2 cytopathicity in the human epithelial colorectal adenocarcinoma cell line, Caco-2. The primary screen identified 258 hits that inhibited cytopathicity by more than 75%, most of which were not previously known to be active against SARS-CoV-2 in vitro. These compounds were tested in an eight-point dose response screen using the same image-based cytopathicity readout. For the 67 most active molecules, cytotoxicity data were generated to confirm activity against SARS-CoV-2. We verified the ability of known inhibitors camostat, nafamostat, lopinavir, mefloquine, papaverine and cetylpyridinium to reduce the cytopathic effects of SARS-CoV-2, providing confidence in the validity of the assay. The high-content screening data are suitable for reanalysis across numerous drug classes and indications and may yield additional insights into SARS-CoV-2 mechanisms and potential therapeutic strategies.

Oligosaccharides increase the genotoxic effect of colibactin produced by pks+ Escherichia coli strains

BackgroundColibactin is a genotoxin that induces DNA double-strand breaks that may lead to carcinogenesis and is produced by Escherichia coli strains harboring the pks island. Human and animal studies have shown that colibactin-producing gut bacteria promote carcinogenesis and enhance the progression of colorectal cancer through cellular senescence and chromosomal abnormalities. In this study, we investigated the impact of prebiotics on the genotoxicity of colibactin-producing E. coli strains Nissle 1917 and NC101.MethodsBacteria were grown in medium supplemented with 20, 30 and 40 mg/mL of prebiotics inulin or galacto-oligosaccharide, and with or without 5 μM, 25 μM and 125 μM of ferrous sulfate. Colibactin expression was assessed by luciferase reporter assay for the clbA gene, essential for colibactin production, in E. coli Nissle 1917 and by RT-PCR in E. coli NC101. The human epithelial colorectal adenocarcinoma cell line, Caco-2, was used to assess colibactin-induced megalocytosis by methylene blue binding assay and genotoxicity by γ-H2AX immunofluorescence analysis.ResultsInulin and galacto-oligosaccharide enhanced the expression of clbA in pks+ E. coli. However, the addition of 125 μM of ferrous sulfate inhibited the expression of clbA triggered by oligosaccharides. In the presence of either oligosaccharide, E. coli NC101 increased dysplasia and DNA double-strand breaks in Caco-2 cells compared to untreated cells.ConclusionOur results suggest that, in vitro, prebiotic oligosaccharides exacerbate DNA damage induced by colibactin-producing bacteria. Further studies are necessary to establish whether oligosaccharide supplementation may lead to increased colorectal tumorigenesis in animal models colonized with pks+ E. coli.

Effects of Akkermansia muciniphila and Faecalibacterium prausnitzii on serotonin transporter expression in intestinal epithelial cells.

The highest level of peripheral serotonin in the body can be found in the gastrointestinal (GI) tract as its reservoir. There is complete interaction between human gastrointestinal microbiota and serotonin system. Serotonin in the GI is transferred by serotonin transporters (SERTs), which play a crucial role in the bioavailability of serotonin in the GI. SERT impairment is associated with the pathology of GI disorders. It is known that intestinal microbiota can regulate the SERT function. Therefore, it may be useful to regulate of SERT expression by modulation of microbiota and improvement of intestinal motility and GI sensation. In this study, we aimed to evaluate the effects of two next-generation probiotics, including Akkermansia muciniphila and Faecalibacterium prausnitzii, and their supernatants on SERT gene expression in human epithelial colorectal adenocarcinoma cells (Caco-2). The Caco-2 cells were treated with multiplicity of infection (MOI) ratio of 100 of A. muciniphila and F. prausnitzii, as well as their supernatants. After 24 h, SERT gene expression was examined by quantitative real-time polymerase chain reaction (qRT-PCR) assay. A. muciniphila up-regulated the SERT mRNA level by 3.01 folds, compared to the control group. F. prausnitzii, similar to A. muciniphila, increased the expression of SERT gene in Caco-2 cells by 3.43 folds (P < 0.001). Moreover, the supernatants of A. muciniphila and F. prausnitzii significantly up-regulated the expression of SERT gene in the cell line by 2.4 and 5.7 folds, respectively, compared to the control group (P < 0.001). The present results showed that A. muciniphila and F. prausnitzii, as well as their supernatants, increased the expression of SERT gene in Caco-2 cells. Therefore, they might be helpful in the microbiota-modulating treatment of inflammatory bowel diseases.

Pasteurization Inactivates SARS-CoV-2-Spiked Breast Milk.

* Abbreviations: Caco-2 — : human epithelial colorectal adenocarcinoma SARS-CoV-2 — : severe acute respiratory syndrome coronavirus 2 TCID50 — : tissue culture infectious dose 50 Vero E6 — : Cercopithecus aethiops –derived epithelial kidney The current coronavirus disease 2019 pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), raises unprecedented questions regarding virus transmission and risks for pregnant or breastfeeding women. We and others described that SARS-CoV-2 RNA is detectable in the breast milk of infected mothers.1–3 In two cases in which the mother continued breastfeeding, the newborns also tested positive for SARS-CoV-2.1,2 However, the origin of the infections of the newborns remained unclear and raised concerns of possible virus transmission via breast milk. The safety and feasibility of breastfeeding is of high importance because breast milk contains nutrients, hormones, and immunoprotective entities that are essential for the development, health, and protection of the neonate. So far, the World Health Organization recommends continued breastfeeding after maternal SARS-CoV-2 infection while taking measures to maintain strict hygiene.4 We here explored the inactivation of SARS-CoV-2 spiked into human milk … Address correspondence to Janis A. Muller or Stephanie Pfaender, Abteilung fur Molekulare und Medizinische Virologie, Ruhr-Universitat Bochum, Universitatsstrasse 150, 44801 Bochum, Germany. E-mail: janis.mueller{at}uni-ulm.de or stephanie.pfaender{at}ruhr-uni-bochum.de

An in vitro selective inhibitory effect of silver(i) aminoacidates against bacteria and intestinal cell lines and elucidation of the mechanism of action by means of DNA binding properties, DNA cleavage and cell cycle arrest.

Novel silver(i) aminoacidate complexes {[Ag(HVal)(H2O)(NO3)]}n (AgVal) and {[Ag3(HAsp)2(NO3)]}n·nH2O (AgAsp) were prepared, investigated and fully characterized by vibrational spectroscopy (mid-IR), elemental analysis, thermogravimetric analysis, X-ray crystallography and mass spectrometry. Their stability in D2O and PBS buffer was verified by time-dependent 1H and 13C NMR measurements. Their in vitro antibacterial activity (against pathogenic Staphylococcus aureus CCM4223, Escherichia coli CCM4787) and that against probiotic bacteria Lactobacillus plantarum CCM7102 and Lactobacillus reuteri (L26) were determined and potential dosing concentration was evaluated. The cytotoxicity of both the complexes against intestinal porcine epithelial (IPEC-1) and human epithelial colorectal adenocarcinoma (CaCo-2) cell lines was determined using the colorimetric MTT assay and against human metastatic melanoma (A2058), human pancreatic adenocarcinoma (PaTu 8902), human cervical adenocarcinoma (HeLa), human colorectal carcinoma (HCT116), human leukaemic T cell lymphoma (Jurkat), and human dermal fibroblasts (HDF) using colorimetric MTS assay. The selectivity index (SI) was identified for intestinal cancer (CaCo-2) and healthy (IPEC-1) cells. The mechanism of action of AgVal and AgAsp was further elucidated and discussed by the study of their binding affinity toward the CT DNA, the ability to cleave the supercoiled form of pUC19 DNA and the ability to influence numbers of cells within each cell cycle.