Stress In Colonic Cells Research Articles

Food grade titanium dioxide induced endoplasmic reticulum stress in colon cells: Comparison between normal and colorectal carcinoma cells

BackgroundFood-grade titanium dioxide (E171) has been under scrutiny in the last decade since its possible adverse effects; however, the cellular mechanisms underlying E171 toxicity have not been thoroughly described. AimWe aimed to compare the effects of E171 on endoplasmic reticulum (ER) homeostasis in normal and cancer colon cells. Experimental designWe exposed normal, carcinoma, and adenocarcinoma cells to 0.1, 1, 10, 50 and 100 μg/cm2 of E171 for 24, 48 and 72 h, and we evaluated ER stress, cell viability, titanium uptake, intracellular calcium concentration, and gene expression related to unfolded protein response (UPR) and chaperone pathways. ResultsCell viability decreased only after 72 h of exposure to 100 μg/cm2 of E171. Adenocarcinoma cells internalized higher titanium amounts than normal and carcinoma cells, but the effects in ER distribution, intracellular calcium concentration, and gene expression were similar among the three cell lines. The expression of UPR and chaperone pathways were downregulated at the lowest concentrations but upregulated at the highest concentrations in the three cell lines. ConclusionE171 induces ER stress through alterations in ER distribution, intracellular calcium, and UPR and chaperone protein pathways.

Inhibition of TRAP1 Accelerates the DNA Damage Response, Activation of the Heat Shock Response and Metabolic Reprogramming in Colon Cancer Cells.

Colorectal cancer (CRC) is one of the major causes of cancer-related mortality worldwide. The tumor microenvironment plays a significant role in CRC development, progression and metastasis. Oxidative stress in the colon is a major etiological factor impacting tumor progression. Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a mitochondrial member of the heat shock protein 90 (HSP90) family that is involved in modulating apoptosis in colon cancer cells under oxidative stress. We undertook this study to provide mechanistic insight into the role of TRAP1 under oxidative stress in colon cells. We first assessed the The Cancer Genome Atlas (TCGA) CRC gene expression dataset to evaluate the expression of TRAP1 and its association with oxidative stress and disease progression. We then treated colon HCT116 cells with hydrogen peroxide to induce oxidative stress and with the TRAP1 inhibitor gamitrinib-triphenylphosphonium (GTPP) to inhibit TRAP1. We examined the cellular proteomic landscape using liquid chromatography tandem mass spectrometry (LC-MS/MS) in this context compared to controls. We further examined the impact of treatment on DNA damage and cell survival. TRAP1 expression under oxidative stress is associated with the disease outcomes of colorectal cancer. TRAP1 inhibition under oxidative stress induced metabolic reprogramming and heat shock factor 1 (HSF1)-dependent transactivation. In addition, we also observed enhanced induction of DNA damage and cell death in the cells under oxidative stress and TRAP1 inhibition in comparison to single treatments and the nontreatment control. These findings provide new insights into TRAP1-driven metabolic reprogramming in response to oxidative stress.

Nanoparticles Induce Oxidative Stress in HT-29 Colon Adenocarcinoma Cell Line After 24 and 48 Hour Exposure

Nanoparticle research is currently an area of intense scientific research, due to a wide variety of potential applications in biomedical, optical, and electronic fields. Nanoparticles are of great scientific interest as they are effectively a bridge between bulk materials and atomic or molecular structures. Superoxide dismutase (SOD), Glutamine synthetase (GS), Catalase (CAT) are some of the defense mechanisms against cellular oxidative stress, especially against free oxygen radicals. In this study, we aimed to investigate that Ag, SiO₂ and ZnO nanoparticles affect cancer cell lines (HT-29) and relationship between SOD, GS and CAT. We investigated that alterations in gene expressions of SOD, GS and CAT caused by exposure to nanoparticles in HT-29 cells. The difference between the Ct values (ΔCt) of the gene of interest was calculated for each experimental sample. As a result of Ag, SiO₂ and ZnO nanoparticle application, there was a 2-fold increase in SOD and CAT expression in the first 24 hours compared to control. As a result of 48 hours of application, it was observed that Ag nanoparticles caused 4-fold increase in SOD and 6-fold statistically significant increase in CAT and GS expression of SiO₂ nanoparticles. Consequently, after 48 hours of nanoparticle application, SiO₂, CAT and GS expression were more effective than 24-hour application. Our results suggest that nanoparticles may cause increased oxidative stress in colon cells and may have therapeutic properties by affecting cancer cells in these aspects.

Translocator Protein-Mediated Stabilization of Mitochondrial Architecture during Inflammation Stress in Colonic Cells.

Chronic inflammation of the gastrointestinal tract increasing the risk of cancer has been described to be linked to the high expression of the mitochondrial translocator protein (18 kDa; TSPO). Accordingly, TSPO drug ligands have been shown to regulate cytokine production and to improve tissue reconstruction. We used HT-29 human colon carcinoma cells to evaluate the role of TSPO and its drug ligands in tumor necrosis factor (TNF)-induced inflammation. TNF-induced interleukin (IL)-8 expression, coupled to reactive oxygen species (ROS) production, was followed by TSPO overexpression. TNF also destabilized mitochondrial ultrastructure, inducing cell death by apoptosis. Treatment with the TSPO drug ligand PK 11195 maintained the mitochondrial ultrastructure, reducing IL-8 and ROS production and cell death. TSPO silencing and overexpression studies demonstrated that the presence of TSPO is essential to control IL-8 and ROS production, so as to maintain mitochondrial ultrastructure and to prevent cell death. Taken together, our data indicate that inflammation results in the disruption of mitochondrial complexes containing TSPO, leading to cell death and epithelia disruption. Significance: This work implicates TSPO in the maintenance of mitochondrial membrane integrity and in the control of mitochondrial ROS production, ultimately favoring tissue regeneration.

Abstract B55: Elevation of oxidative expression proteins in colorectal adenomatous polyps

Abstract The optimal management of patients with colorectal adenomatous polyps depends on the accuracy of appropriate staging strategies because patients with similar colorectal adenocarcinoma architecture display heterogeneity in the course and outcome of the disease. Chronic oxidative stress may play a critical role in the pathogenesis of colorectal cancers, given that a major promutagenic threat faced by colonic epithelial cells is that of chronic oxidative stress. Although reactive oxygen species arise in most cells as a result of normal metabolic processes, additional sources of oxidative stress in colonic cells may include enzymes such as cyclooxygenase 2 (COX-2) and ATPase 6 and 8. High levels of COX-2 and ATPase protein expression have been detected in human colorectal polyps, which are lesions that can progress to colorectal carcinomas. This study applied spectrophotometric, dinitrophenylhydrazone (DNPH) assay, two dimensional gel electrophoresis, and Western blot analyses to assess the levels of oxidatively modified carbonyl proteins in 37 pairs of normal/surrounding and colorectal adenomatous polyps and cancer tissues. Our data showed that the levels of reactive protein carbonyl groups increased as level of malignancy increased. The level of carbonyl protein was 58% and 26% less in N/S tissues as compare to tumor tissue in adenocarcinoma, and colorectal adenomatous polyps tissues respectably. The differences between the level of carbonyl potion in N/S and tumor tissues was more pronounced in African American patient than in Caucasians. Also the levels of carbonyl proteins were notably higher in tissue contained higher number of tumor cells in different tissues blocks obtained from same patient. Our results may suggest that oxidative stress is involved in oxidatively modification of carbonyl proteins, leading to increased aggressiveness of colorectal polyps. This study was supported by NIGMS grant # GM099663- awarded to Dr. Felix O. Aikhionbare Citation Format: LaShanale M. Wallace, Sharifeh Mehrabi, Shakeria Cohen, Felix O. Aikhionbare. Elevation of oxidative expression proteins in colorectal adenomatous polyps. [abstract]. In: Proceedings of the Seventh AACR Conference on The Science of Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; Nov 9-12, 2014; San Antonio, TX. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2015;24(10 Suppl):Abstract nr B55.

Application of confocal laser scanning microscopy to detect oxidative stress in human colon cells

Introduction Excess of intracellular reactive oxygen species in relation to antioxidative systems results in an oxidative environment which may modulate gene expression or damage cellular molecules. These events are expected to greatly contribute to processes of carcinogenesis. Only few studies are available on the oxidative/reductive conditions in the colon, an important tumour target tissue. It was the objective of this work to further develop methods to assess intracellular oxidative stress within human colon cells as a tool to study such associations in nutritional toxicology.Methods We have measured H2O2-induced oxidative stress in different colon cell lines, in freshly isolated human colon crypts, and, for comparative purposes, in NIH3T3 mouse embryo fibroblasts. Detection was performed by loading the cells with the fluorigenic peroxide-sensitive dye 6-carboxy-2′,7′-dichlorodihydrofluorescein diacetate (diacetoxymethyl ester), followed by in vitro treatment with H2O2 and fluorescence detection with confocal laser scanning microscopy (CLSM). Using the microgel electrophoresis (“Comet”) Assay, we also examined HT29 stem and clone 19A cells and freshly isolated primary colon cells for their relative sensitivity toward H2O2-induced DNA damage and for steady-state levels of endogenous oxidative DNA damage.Results A dose-response relationship was found for the H2O2-induced dye decomposition in NIH3T3 cells (7.8–125 μM H2O2) whereas no effect occurred in the human colon tumour cell lines HT29 stem and HT29 clone 19A (62–1000 μM H2O2). Fluorescence was significantly increased at 62 μM H2O2 in the human colon adenocarcinoma cell line Caco-2. In isolated human colon crypts, the lower crypt cells (targets of colon cancer) were more sensitive towards H2O2 than the more differentiated upper crypt cells. In contrast to the CLSM results, oxidative DNA damage was detected in both cell lines using the Comet Assay. Endogenous oxidative DNA damage was highest in HT29 clone 19A, followed by the primary colon cells and HT29 stem cells.Conclusions Oxidative stress in colon cells leads to damage of macromolecules which is sensitively detected in the Comet Assay. The lacking response of the CLSM-approach in colon tumour cells is probably due to intrinsic modes of protective activities of these cells. In general, however, the CLSM method is a sensitive technique to detect very low concentrations of H2O2-induced oxidative stress in NIH3T3 cells. Moreover, by using colon crypts it provides the unique possibility of assessing cell specific levels of oxidative stress in explanted human tissues. Our results demonstrate that the actual target cells of colon cancer induction are indeed susceptible to the oxidative activity of H2O2.