Colon Adenocarcinoma SW480 Cells Research Articles

Interactions between genistein and Wnt pathway in colon adenocarcinoma and early embryos

The Wnt signaling pathway is one of the most ancient and pivotal signaling cascades, governing diverse processes in development and cancer regulation. Within the realm of cancer treatment, genistein emerges as a promising candidate due to its multifaceted modulation of various signaling pathways, including the Wnt pathway. Despite promising preclinical studies, the precise mechanisms underlying genistein's therapeutic effects via Wnt modulation remain elusive. In this study, we unveil novel insights into the therapeutic mechanisms of genistein by elucidating its inhibitory effects on Wnt signaling through macropinocytosis. Additionally, we demonstrate its capability to curtail cell growth, proliferation, and lysosomal activity in the SW480 colon adenocarcinoma cell model. Furthermore, our investigation extends to the embryonic context, where genistein influences gene regulatory networks governed by endogenous Wnt pathways. Our findings shed light on the intricate interplay between genistein, Wnt signaling, membrane trafficking, and gene regulation, paving the way for further exploration of genistein's therapeutic potential in cancer treatment strategies.

Abstract 3175: Dear ABBIE, edit that colon cancer

Abstract Genome editing has exciting potential for gene therapy of cancers. Colorectal cancer is the third most occurring cancer in both men and women. The current treatments for colon cancers include surgery, radiofrequency ablation, chemotherapies and immunotherapies. These either involve removal of intestinal tissue that can alter the quality of life of the patient, or require doses of drugs that generally result in various side effects. ABBIE (A Binding Based Integrase Enzyme) genome editing can insert operational gene sequences into a chosen site in the genomic DNA of cells via its targetability and integrase activity. The ABBIE system offers ease of use, faster time to a readout and decreased time per genome editing experiment. SW480 and Caco-2 colon adenocarcinoma cells were used with the ABBIE system to integrate a GFP donor sequence into a safe harbor locus of the genome. Integration is to be confirmed with fluorescence microscopy and DNA sequencing. The colon cancer cell line will then be edited with the ABBIE genome editing system to include a gene cassette that overexpresses caspase genes into the genomic DNA of these cells. ABBIE-Edited colon cancer cell lines are expected to exhibit increased apoptosis as compared to non-edited controls or controls edited with DNA that do not have the caspase genes. This strategy will help lead to novel and more targetable treatments for colon cancer. Citation Format: David C. Aguilar, Alfred Gallegos. Dear ABBIE, edit that colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3175.

Genistein Enhances TRAIL-Mediated Apoptosis Through the Inhibition of XIAP and DcR1 in Colon Carcinoma Cells Treated with 5-Fluorouracil.

Colorectal cancer is one of the most common cancers worldwide. However, surgical intervention and chemotherapy provide only limited benefits for the recovery and survival of patients. The anticarcinogenic effect of genistein has attracted attention because epidemiological studies have shown that soybean consumption is associated with a decrease in the incidence of cancer. There are limited studies on the effects of genistein in colorectal carcinoma cells. We aimed to investigate the cytotoxic, genotoxic, and apoptotic effects of genistein in SW480 and SW620 colon adenocarcinoma cells treated with 5-fluorouracil, the basis of chemotherapy, and the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) ligand, the mediator of apoptosis, both alone and in combination. Cytotoxicity and genotoxicity were determined by MTT and comet assays, respectively. The apoptotic effects were evaluated by reverse transcription-polymerase chain reaction assay, with the additional use of Annexin V FITC, mitochondrial membrane potential (MMP), caspase 3, 8, and 9 activity, and reactive oxygen species (ROS) assay kits. According to our findings, genistein, 5-fluorouracil, and TRAIL had synergistic apoptotic effects because of DR5 upregulation, ROS production, and DNA damage, which were mediated by increased caspase-8, and -9 activity and decreased MMP. The applied combinations of these compounds may contribute to the resistance problem that may occur in treating colorectal cancer, with a decrease in DcR1 and XIAP genes.

Elevated expression of CXCL3 in colon cancer promotes malignant behaviors of tumor cells in an ERK-dependent manner

BackgroundCXC chemokine ligand 3 (CXCL3) is a member of CXC-type chemokine family that is identified as a major regulator in immune and inflammation responses. Recently, numerous evidence indicated that CXCL3 is broadly expressed in various human tumor types, and it is also known to play a critical role in mediating tumor development and progression. However, the expression profile of CXCL3 and the exact molecular mechanism behind the role of CXCL3 in colon adenocarcinoma (COAD) has not been fully elucidated.MethodsThe expression and clinical significance of CXCL3 mRNA and protein in the tissues from COAD patients were estimated using bioinformatics and immunohistochemistry assays. The expression and roles of exogenous administration or overexpression of CXCL3 in HT-29 and SW480 COAD cells were determined using enzyme-linked immunosorbent assay(ELISA), Cell Counting Kit-8 (CCK-8) and Transwell assays. Mechanically, CXCL3-induced malignant behaviors were elucidated using western blotting assay and extracellular signal-regulated protein kinase 1/2 (ERk1/2) inhibitor PD98059.ResultsThe cancer genome atlas (TCGA)-COAD data analysis revealed that CXCL3 mRNA is highly expressed and has high clinical diagnostic accuracy in COAD. Increased expression of CXCL3 mRNA was associated with patient’s clinical stage, race, gender, age, histological subtype, nodal mestastasis and tumor protein 53 (TP53) mutation status. Similarly, immunohistochemistry assay also exhibited that CXCL3 protein in COAD tissues was significantly up-regulated. Gene expression associated assay implied that CXC chemokine ligand 1 (CXCL1) and CXC chemokine ligand 2 (CXCL2) were markedly correlated with CXCL3 in COAD. Protein-protein interaction (PPI) analysis revealed that cyclin B1 (CCNB1), mitotic arrest deficient 2 like 1 (MAD2L1), H2A family member Z (H2AFZ) and CXCL2 may be the important protein molecules involved in CXCL3-related tumor biology. Gene set enrichment analysis (GSEA) analysis revealed that CXCL3 was mainly enriched in the cell cycle, DNA replication, NOD-like receptors, NOTCH and transforming growth factor-β (TGF-β) Signal pathways. In vitro, exogenous administration or overexpression of CXCL3 resulted in increased malignant behaviors of HT-29 and SW480 cells, and down-regulation of CXCL3 expression inhibited the malignant behaviors of these tumor cells. In addition, overexpression of CXCL3 affected the expression of genes related to extracellular signal regulated kinase (ERK) pathway, including ERK1/2, p-ERK, B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax) and Cyclin D1. Finally, CXCL3-induced malignant behaviors in HT-29 and SW480 cells were obviously attenuated following treatment with ERK inhibitor PD98059.ConclusionCXCL3 is upregulated in COAD and plays a crucial role in the control of malignant behaviors of tumor cells, which indicated its involvement in the pathogenesis of COAD.

Comparison of Phytochemical Composition and Untargeted Metabolomic Analysis of an Extract from Cnidoscolus aconitifolius (Mill.) I. I. Johnst and Porophyllum ruderale (Jacq.) Cass. and Biological Cytotoxic and Antiproliferative Activity In Vitro.

Cnidoscolus aconitifolius (CA) and Porophyllum ruderale (PR) are representative edible plants that are a traditional food source in Mexico. This research aimed to analyze the phytochemical composition and untargeted metabolomics analysis of CA and PR and evaluate their antiproliferative effect in vitro. The phytochemical composition (UPLC-DAD-QToF/MS-ESI) identified up to 38 polyphenols and selected organic acids that were clustered by the untargeted metabolomics in functional activities linked to indolizidines, pyridines, and organic acids. Compared with PR, CA displayed a higher reduction in the metabolic activity of human SW480 colon adenocarcinoma cells (LC50: 10.65 mg/mL), and both extracts increased the total apoptotic cells and arrested cell cycle at G0/G1 phase. PR increased mRNA Apc gene expression, whereas both extracts reduced mRNA Kras expression. Rutin/epigallocatechin gallate displayed the highest affinity to APC and K-RAS proteins in silico. Further research is needed to experiment on other cell lines. Results suggested that CA and PR are polyphenol-rich plant sources exhibiting antiproliferative effects in vitro.

Identification of potential bioactive compounds of Passiflora edulis leaf extract against colon adenocarcinoma cells

Plant bioactive compounds such as flavonoids and triterpenes can affect lipid metabolism. Here, we report the cytotoxic and lipid-lowering activities of the ethanolic extract of P. edulis leaves on human colon adenocarcinoma SW480 cells, also the molecular interactions of bioactive compounds present in P. edulis extract on ACC and HMGCR enzymes. The extract reduced cell viability and decreased intracellular triglyceride content by up to 35% and 28% at 24 and 48 h, respectively; whereas the effect was evident on cholesterol only at 24 h. In-silico analysis revealed that luteolin, chlorogenic acid, moupinamide, isoorientin, glucosyl passionflower, cyclopasifloic acid E and saponarin had optimal molecular coupling on Acetyl-CoA Carboxylase 1 and 2 as well as 3-hydroxy-3-methyl-glutaryl-CoA reductase, with possible inhibitory effects. These results show the ability of ethanolic extract to reduce intracellular levels of cholesterol and triglycerides in SW480 cells, which attracts attention for the treatment of colorectal cancer.

Antiproliferative potential of Andean Berry (Vaccinium meridionaleSwartz) juice in combination with Aspirin in human SW480 colon adenocarcinoma cells

Andean Berry (Vaccinium meridionale Sw.) is a South American fruit rich in phytochemicals with promising anti-cancer properties as co-adjuvants to nonsteroidal anti-inflammatory drugs such as Aspirin. This study aimed to evaluate the antiproliferative potential of Andean Berry Juice (ABJ) in combination with Aspirin in human SW480 colon adenocarcinoma cells. ABJ primarily contained 3,4-dihydroxybenzoic and chlorogenic acids. The combined treatment of ABJ (IC50 : 30.0±0.11%) and Aspirin (IC50 : 20.0±0.57) exhibited a higher (p<.01) antiproliferative effect than each counterpart. Moreover the same mixture displayed a lower reduced glutathione/oxidized glutathione ratio (GSH/GSSG) than the untreated cells. ABJ-Aspirin combination induced late apoptosis stage without stimulating mitochondrial depolarization and prompted phosphatidylserine relocalization. These results emphasize the antiproliferative potential of bioactive compounds from ABJ and Aspirin combinations. PRACTICAL APPLICATIONS: Natural products such as Andean Berry (V. meridionale Sw.) juice (ABJ) contains antioxidant polyphenols that could reduce the need to use non-steroidal anti-inflammatory drugs, currently employed in cancer treatment, to prevent its side effects. The high abundance of polyphenols from this underutilized berry could stimulate the standardization of its production and industrial exploitation to be transformed into suitable food products delivering natural bioactive compounds with potential anti-cancer effects in vitro.

Prognostic value and oncogenic effect of increased RBM8A expression in colon adenocarcinoma.

RNA Binding Motif Protein 8A (RBM8A) is a component of the spliceosome, which couples pre- and post-mRNA splicing events. Dysregulated expression of RBM8A has been recently discovered in hepatocellular carcinoma and gastric cancer. This study aimed to investigate the expression pattern of RBM8A protein in colon adenocarcinoma and to explore its correlation with patients' clinicopathological characteristics as well as clinical outcomes. Accordingly, we initially found that RBM8A was significantly higher in colon adenocarcinoma tissues compared to adjacent tissues on both mRNA and protein levels. Meanwhile, RBM8A protein was positively correlated with tumor size, depth of invasion, and lymph node metastasis. Furthermore, patients with increased RBM8A expression or positive lymph nodes exhibited a poorer overall survival. Consistently, immunoblotting data showed that RBM8A was higher in SW480 and SW620 colon adenocarcinoma cell lines compared to nontumorous cells. Finally, in vitro and in vivo assays elucidated the role of RBM8A on promoting colon cancer growth by using knockdown strategy. Taken together, our study demonstrated that RBM8A may act as a proto-oncogene, which could be a promising biomarker and therapeutic target in the survival prediction and treatment of colon adenocarcinoma.

Experimental and theoretical characterization of the strong effects on DNA stability caused by half-sandwich Ru(II) and Ir(III) bearing thiabendazole complexes.

The synthesis and characterization of two half-sandwich complexes of Ru(II) and Ir(III) with thiabendazole as ancillary ligand and their DNA binding ability were investigated using experimental and computational methods. 1H NMR and acid-base studies have shown that aquo-complexes are the reactive species. Kinetic studies show that both complexes bind covalently to DNA through the metal site and non covalently through the ancillary ligand. Thermal stability studies, viscosity, circular dichroism measurements and quantum chemical calculations have shown that the covalent binding causes breaking of the H-bonding between base pairs, bringing about DNA denaturation and compaction. Additionally, molecular dynamics (MD) simulations and quantum mechanics/molecular mechanics (QM/MM) calculations shed light into the binding features of the Ru(II) and Ir(III) complexes and their respective enantiomers toward double-helical DNA, highlighting the important role played by the NˆN ancillary ligand once the complexes are covalently linked to DNA. Moreover, metal quantification in the nucleus of SW480 colon adenocarcinoma cells were carried out by inductively coupled plasma-mass spectrometry (ICP-MS), both complexes are more internalized than cisplatin after 4h of exposition. However, in spite of the dramatic changes in the helicity of the DNA secondary structure induced by these complexes and their nuclear localization, antiproliferative studies have revealed that both, Ru(II) and Ir(III) complexes, cannot be considered cytotoxic. This unexpected behavior can be justified by the fast formation of aquo-complexes, which may react with components of the cell culture medium or the cytoplasm compartment in such a way that they may become deactivated before reaching DNA.

3D h9e peptide hydrogel: An advanced three-dimensional cell culture system for anticancer prescreening of chemopreventive phenolic agents

Traditional 2D monolayer cell culture model may overestimate chemopreventive agent's response due to lacking physiological relevance in three-dimensional microenvironment. This study was aimed to apply a novel 3D h9e peptide hydrogel cell culture system to evaluate the anticancer efficacy of chemopreventive phenolic acid on hepatocarcinoma HepG2 and colon adenocarcinoma SW480 cells. Both cell lines grew better in this 3D system with better cell growth and longer exponential phase than that in 2D model. Chlorogenic acid (CGA), known as a chemopreventive phenolic acid, at 0–40 μM for 72 h inhibited cell growth but not viability in both HepG2 and SW480 cells. The inhibition was much less potent in 3D system with an IC50 value of 58.0 ± 15.8 or 285.6 ± 75.4 μM when compared with 2D model with IC50 of 5.3 ± 0.3 or 12.0 ± 2.5 μM for HepG2 or SW480, respectively. Furthermore, the recovery of cells grown in 3D system after post-CGA appeared faster than 2D model. Taken together, an advanced 3D model has been established with favoring cell growth and less susceptible to inhibitory treatments in contrast to 2D model, thus predict closely to in vivo situation and may bridge the gap of in vitro to in vivo for prescreening chemopreventive agents for cancer prevention.

Gene expression profile of cancer stem‑like cells in the SW480 colon adenocarcinoma cell line.

Cancer stem cells (CSCs) serve an important role in tumorigenesis, tolerance to treatment, relapse and metastasis. Although the signaling pathways involved in cancer are well known, the molecular mechanisms underlying CSC actions require further investigation. In the present study, a population of colon CSCs with a cluster of differentiation 133 (CD133) surface‑expression phenotype from the human SW480 colon adenocarcinoma cell line were isolated by flow cytometry. The CD133+ cells exhibited increased tumor sphere‑forming efficiency in vitro and increased tumorigenic potential in vivo. Furthermore, the gene expression profile of colon CSCs was investigated using gene chip technology. The results demonstrated differential gene expression between the CD133+ and CD133‑ subpopulations, including in relation to a number of important genes with functions in transcription control, cell cycle, karyomitosis and protein phosphorylation, including cyclin dependent kinase inhibitor 1A, cyclin B1, checkpoint kinase 1, cyclin dependent kinase 1 and transforming growth factor β receptor 2. Pathway analysis revealed that the mitogen‑activated protein kinase, p53 and calcium signaling pathways, as well as other important signaling pathways, were differentially activated between CD133+ and CD133‑ cells. To the best of our knowledge, these results provide the first evidence for the gene expression profile of colon CSCs in the SW480 colon adenocarcinoma cell line. Although further studies will be required to identify the functional roles of these genes in the CSC phenotype, these observations provide the basis for future studies to elucidate the pathogenesis of colorectal cancer and to develop novel tumor‑targeting therapies.

Bioaccessibility during In Vitro Digestion and Antiproliferative Effect of Bioactive Compounds from Andean Berry (Vaccinium meridionale Swartz) Juice

Berry consumption is associated with colorectal-cancer chemoprevention, but digestive conditions can affect this property. The bioaccessibility and apparent permeability coefficients of bioactive compounds from Andean Berry Juice (ABJ) after in vitro gastrointestinal digestion and colonic fermentation were analyzed. The antiproliferative effect of the fermented nondigestible fraction was evaluated against SW480 colon-adenocarcinoma cells. Gallic acid displayed the highest bioaccessibility in the mouth, stomach, small intestine, and colon. However, chlorogenic acid exhibited the highest apparent permeability coefficients (up to 1.98 × 10-4 cm/s). The colonic-fermentation fraction showed an increase of ≥50% antiproliferative activity against SW480 cells (19.32%, v/v), equivalent to those of gallic acid (13.04 μg/g), chlorogenic acid (7.07 μg/g), caffeic acid (0.40 μg/g), ellagic acid (7.32 μg/g), rutin (6.50 μg/g), raffinose (0.14 mg/g), stachyose (0.70 mg/g), and xylose (9.41 mg/g). Bioactive compounds from ABJ are bioaccessible through the gastrointestinal tract and colon fermentation, resulting in antiproliferative activity.

Simultaneous stimulation with tumor necrosis factor-α and transforming growth factor-β1 induces epithelial-mesenchymal transition in colon cancer cells via the NF-κB pathway.

Epithelial-mesenchymal transition (EMT) is critical in the progression of numerous types of carcinoma, and endows invasive and metastatic properties upon cancer cells. The tumor microenvironment facilitates tumor metastasis to distant organs. Various signaling pathways contribute to this process. In the present study, SW480 colon adenocarcinoma cells were treated with transforming growth factor-β1 (TGF-β1; 10 ng/ml) and tumor necrosis factor-α (TNF-α; 20 ng/ml), alone or in combination, for 72 h, and EMT was assessed using immunofluorescence, western blot analysis and migration assays. The functions of p38 mitogen-activated protein kinase, extracellular signal-regulated kinase (ERK) and nuclear factor-κB (NF-κB) pathways in EMT were examined. It was demonstrated that the cooperation of TGF-β1 and TNF-α signaling promoted the morphological conversion of the SW480 cells from an epithelial to a mesenchymal phenotype. Furthermore, simultaneous exposure to TNF-α and TGF-β1 downregulated the expression of E-cadherin (an epithelial marker) and increased the expression of N-cadherin and vimentin (mesenchymal markers). Additionally, the migratory capacity of the SW480 cells increased. The inhibition of p38 and ERK signaling exhibited no effect on EMT, whereas the inhibition of inhibitor of NF-κB kinase subunit β blocked the EMT induced by TGF-β1 and TNF-α. In conclusion, the results of the present study demonstrated that TNF-α and TGF-β1 synergistically promoted EMT in SW480 cells via the NF-κB pathway, independent of p38 activation and ERK1/2 signaling. These results suggest a novel function of TGF-β1 and TNF-α during EMT in colon carcinoma and, thus, provide insights into potential therapeutic interventions.

English

Vaccium meridionale Swartz, commonly Andean Berry, has a high content of several phytochemicals, such as anthocyanins, phenolic acids, and other flavonoids. However, in spite of its antioxidant capacity, there is little information about its anticarcinogenic properties. This study evaluated the antiproliferative and pro-apoptotic activity of Andean Berry Juice (ABJ) on human colon adenocarcinoma SW480 cells. The antiproliferative activity of ABJ was evaluated on SW480 cells using the Sulphorodamine B assay.The effect on cell viability, cytotoxicity and activation of caspase-3 was analyzed using The ApoTox-Glo™ Triplex Assay. Specific apoptotic biomarkers cleaved PARP, total Bcl-2-associated death promote (BAD), phosphorylated BAD, total p53, and phosphorylated p53 were also analyzed. To determine the intracellular redox-state, the Glutathione Assay Kit and 2′-7′-Dichlorodihydrofluorescein diacetate (DCFH-DA) were used, respectively. The antiproliferative assay showed a IC50 value of 8% v/v ABJ, the caspase 3 activity was increased in time-dependent manner in SW480 treated cells, the proapoptotic proteins (cleaved caspase 3, cleaved PARP, P53 and total BAD) were increased by 1.6 to 2.0 fold. In addition, the ABJ-treated SW480 cells increased significantly the production of intracellular reactive oxygen species (ROS), parallel with reduction in the intracellular content of glutathione (GSH) and consequently a decrease of GSH/ oxidized glutathione (GSSG) ratio. In conclusion, the ABJ was able to inhibit SW480 cells proliferation involving apoptotic mechanisms through the perturbation of intracellular oxidative state. Key words: Vaccinum, colon cancer, anthocyanins, apoptosis, oxidative stress.

Laser microfabrication of a microheater chip for cell culture outside a cell incubator

Microfluidic chips have demonstrated their significant application potentials in microbiological processing and chemical reactions, with the goal of developing monolithic and compact chip-sized multifunctional systems. Heat generation and thermal control are critical in some of the biochemical processes. The paper presents a laser direct-write technique for rapid prototyping and manufacturing of microheater chips and its applicability for lab-on-a-chip cell culture outside a cell incubator. The aim of the microheater is to take the role of conventional incubators for cell culture for facilitating microscopic observation and/or other online monitoring activities during cell culture and provides portability of cell culture operation. Microheaters (5mm×5mm) have been successfully fabricated on soda-lime glass substrates covered with aluminium layer of thickness 120nm. Experimental results show that the microheaters exhibit good performance in temperature rise and decay characteristics, with localized heating at targeted spatial domains. These microheaters were suitable for a maximum long-term operation temperature of 120°C and validated for operation at 37°C for 48h. Results demonstrated that the microheaters are suitable for the culture of immortalised cell lines. The growth and viability of SW480 colon adenocarcinoma cells cultured the developed microheater chip were comparable to the results obtained in a conventional cell incubator.

Multifunctional microspherical magnetic and pH responsive carriers for combination anticancer therapy engineered by droplet-based microfluidics.

pH stimuli responsive drug delivery platforms that can target specific locations along the gastrointestinal tract hold great promise for colorectal cancer therapy. Herein, we present a facile approach to produce microfluidic engineered pH-sensitive magnetic microspherical carriers containing multifunctional therapeutic payloads for synergistic treatment of colorectal cancer. Chemotherapeutics, 5 fluorouracil (5FU) and curcumin (CUR), were chosen due to their synergistic effect for colorectal cancer treatment and prevention. Drugs were loaded onto naturally derived porous silicon nanoparticles (SiNPs) and magnetic bacterial iron oxide nanowires (BacNWs), which acted as drug nanocontainers and magnetic elements, respectively. Drug loaded SiNPs and BacNWs were then encapsulated into polymeric microspheres using droplet-based microfluidics. To ensure controlled drug delivery into the desired site of action (colon and rectum), the microspheres were fabricated using hypromellose acetate succinate polymers, which are insoluble in the acidic medium of the stomach (i.e. pH 1.2) but soluble at basic pH (colon and rectum). Our results confirmed that the microspheres exhibit a narrow size distribution (CV > 5%) with precise size control. Moreover, in vitro dissolution and drug release data confirmed their pH-responsive properties. Motivated by these results, we explored the biocompatibility of microspheres using human RAW 264.7 macrophages. The results revealed the safety of drug free microspheres up to 1000 μg mL-1. Finally, the synergistic action of 5FU and CUR loaded microspheres was investigated on SW480 colon adenocarcinoma cells.

Epigenetic silencing of serine protease HTRA1 drives polyploidy.

BackgroundIncreased numbers and improperly positioned centrosomes, aneuploidy or polyploidy, and chromosomal instability are frequently observed characteristics of cancer cells. While some aspects of these events and the checkpoint mechanisms are well studied, not all players have yet been identified. As the role of proteases other than the proteasome in tumorigenesis is an insufficiently addressed question, we investigated the epigenetic control of the widely conserved protease HTRA1 and the phenotypes of deregulation.MethodsMouse embryonal fibroblasts and HCT116 and SW480 cells were used to study the mechanism of epigenetic silencing of HTRA1. In addition, using cell biological and genetic methods, the phenotypes of downregulation of HTRA1 expression were investigated.ResultsHTRA1 is epigenetically silenced in HCT116 colon carcinoma cells via the epigenetic adaptor protein MBD2. On the cellular level, HTRA1 depletion causes multiple phenotypes including acceleration of cell growth, centrosome amplification and polyploidy in SW480 colon adenocarcinoma cells as well as in primary mouse embryonic fibroblasts (MEFs).ConclusionsDownregulation of HTRA1 causes a number of phenotypes that are hallmarks of cancer cells suggesting that the methylation state of the HtrA1 promoter may be used as a biomarker for tumour cells or cells at risk of transformation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2425-8) contains supplementary material, which is available to authorized users.

Wnt/β-Catenin Signaling Regulates the Expression of the Ammonium Permease Gene RHBG in Human Cancer Cells.

Ammonium is a metabolic waste product mainly detoxified by the liver. Hepatic dysfunction can lead to cytotoxic accumulation of circulating ammonium and to subsequent encephalopathy. Transmembrane ammonium transport is a widely spread process ensured by the highly conserved proteins of the Mep-Amt-Rh superfamily, including the mammalian Rhesus (Rh) factors. The regulatory mechanisms involved in the control of RH genes expression remain poorly studied. Here we addressed the expression regulation of one of these factors, RHBG. We identify HepG2 hepatocellular carcinoma cells and SW480 colon adenocarcinoma cells as expressing RHBG and show that its expression relies on β-catenin signaling. siRNA-mediated β-catenin knockdown resulted in significant reduction of RHBG mRNA in both cell lines. Pharmaceutical inhibition of the TCF4/β-catenin interaction or knockdown of the transcription factor TCF4 also downregulated RHBG expression. We identify a minimal RHBG regulatory sequence displaying a promoter activity and show that β-catenin and TCF4 bind to this fragment in vivo. We finally characterize the role of potential TCF4 binding sites in RHBG regulation. Taken together, our results indicate RHBG expression as a direct target of β-catenin regulation, a pathway frequently deregulated in many cancers and associated with tumorigenesis.

English

Tea is a beverage made from leaves with high contents of polyphenolic substances that vary based on the process they are subjected to. In this study, the apical and young leaves from Vaccinium meridionale (named mortino) were processed to obtain two kinds of tea: green and black tea. This was done in order to compare their antioxidant activity, content of secondary metabolites at different temperatures of extraction and their antiproliferative effect against SW480 colon cancer cells. Results showed that at 40°C, the green tea infusion presented higher antioxidant activity than the black tea infusion, based on their evaluation using Trolox equivalent antioxidant capacity (TEAC)-diphenylpicrylhydrazyl (DPPH) and oxygen radical absorbance capacity (ORAC) techniques. The green tea also had maximum contents of epicatechin, caffeine, ferulic, chlorogenic and ascorbic acid than the black tea. The total contents of phenols, including hydroxycinnamic acids (caffeic and p-coumaric acid) presented similar results, in both types of tea at the same temperature, as well as the total contents of flavonoids and catechin. When temperatures increased, the extraction of bioactive compounds was more efficient in the black tea infusion than the green tea. This situation led to the increased growth rate per the temperature of the total content of phenols, among which chlorogenic, caffeic and p-coumaric acid were prominent, as well as the corresponding non-polyphenolic substances such as ascorbic acid. The latter may be responsible for the increased antioxidant activity as the temperature increased in the extraction. This antioxidant activity was observed in the black tea from mortino leaves, using TEAC-DPPH, fluorescence recovery after photobleaching (FRAP) and ORAC assays. Both types of teas had a dose-dependent antiproliferative effect against SW480 colon adenocarcinoma cells. The IC50 of the green and black tea was 26.3 and 36 µg/ml, respectively. These findings suggest that a tea prepared from mortino leaves may be a promising source of antioxidant and bioactive compounds against colon cancer cells. Key words: Antioxidant, antiproliferative, mortino, polyphenols.

Endogenous enzyme-hydrolyzed fruit of Cirsium brachycephalum: Optimal source of the antiproliferative lignan trachelogenin regulating the Wnt/β-Catenin signaling pathway in the SW480 colon adenocarcinoma cell line

The molecular constituents of Cirsium brachycephalum fruits were identified, quantified and isolated for the first time. The lignan glycoside tracheloside was the main compound, which was transformed quantitatively into its aglycone trachelogenin by endogenous enzymatic treatment of the fruit. Following this transformation by high performance liquid chromatography (HPLC) hyphenated with UV and mass spectrometry (MS) detections on a quantitative basis, the enzyme-hydrolyzed fruit was found to be the richest raw material containing trachelogenin (17.2mg/g) reported to date. Thus, the enzyme-hydrolyzed fruit was used to isolate trachelogenin using preparative HPLC in order to (1) unambiguously confirm its identity by gas chromatography–MS, nuclear magnetic resonance spectroscopy and optical rotation, and (2) investigate its in vitro antiproliferative activities against the SW480 colon adenocarcinoma cell line. Trachelogenin significantly affected the phosphorylation of key proteins such as β-Catenin, c-Myc and GSK3 in the β-Catenin signaling pathway in a concentration-dependent manner. These changes account for the antiproliferative effects of trachelogenin.