4-hydroxy-2-nonenal and Cell-Transformation--Neoplastic

Commensal bacteria and innate immunity play a major role in the development of colorectal cancer (CRC). We propose that selected commensals polarise colon macrophages to produce endogenous mutagens that initiate chromosomal instability (CIN), lead to expression of progenitor and tumour stem cell markers, and drive CRC through a bystander effect.. Primary murine colon epithelial cells were repetitively exposed to Enterococcus faecalis-infected macrophages, or purified trans-4-hydroxy-2-nonenal (4-HNE)-an endogenous mutagen and spindle poison produced by macrophages. CIN, gene expression, growth as allografts in immunodeficient mice were examined for clones and expression of markers confirmed using interleukin (IL) 10 knockout mice colonised by E. faecalis.. Primary colon epithelial cells exposed to polarised macrophages or 4-hydroxy-2-nonenal developed CIN and were transformed after 10 weekly treatments. In immunodeficient mice, 8 of 25 transformed clones grew as poorly differentiated carcinomas with 3 tumours invading skin and/or muscle. All tumours stained for cytokeratins confirming their epithelial cell origin. Gene expression profiling of clones showed alterations in 3 to 7 cancer driver genes per clone. Clones also strongly expressed stem/progenitor cell markers Ly6A and Ly6E. Although not differentially expressed in clones, murine allografts positively stained for the tumour stem cell marker doublecortin-like kinase 1. Doublecortin-like kinase 1 and Ly6A/E were expressed by epithelial cells in colon biopsies for areas of inflamed and dysplastic tissue from E. faecalis-colonised IL-10 knockout mice.. These results validate a novel mechanism for CRC that involves endogenous CIN and cellular transformation arising through a microbiome-driven bystander effect.

Topics: Aldehydes; Animals; Bystander Effect; Cell Line; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Enterococcus faecalis; Female; Gram-Positive Bacterial Infections; HCT116 Cells; Humans; In Situ Hybridization, Fluorescence; Intestinal Mucosa; Macrophages; Mice; Mice, Inbred NOD; Neoplasm Transplantation; Neoplastic Stem Cells

Studies of the decades-long latent stages of breast carcinogenesis have been limited to when hyperplastic lesions are already present. Investigations of earlier stages of breast cancer (BC) latency have been stymied by the lack of fiducial biomarkers needed to identify where in histologically normal tissues progression toward a BC might be taking place. Recent evidence suggests that a marker of chronic oxidative stress (OxS), protein adducts of 4-hydroxy-2-nonenal (4HNE), can meet this need. Specifically: (1) 4HNE immunopositive (4HNE+) mammary epithelial (ME) cells were found to be prevalent in normal (reduction mammoplasty) tissues of most women (including many teenagers) studied, representative of those living in the United States' high risk-posing environment and: (2) marked (> 1.5-fold) differences were identified between tissues of healthy young women with many vs. few 4HNE+ ME cells in the relative levels of transcripts for 42 of the 84 OxS-associated genes represented in SABioscience Oxidative-Stress/Oxidative-Defense PCR array. Herein we used synchrotron radiation-based Fourier-transform infrared (SR-FTIR) microspectroscopy to identify molecular changes associated with 4HNE adducts in basal and luminal ME cells in terminal ductal units (TDLU), which are the cells of origin of BC, and associated intralobular and interlobular stroma, known contributors to carcinogenesis. Multivariate analysis-derived wavenumbers differentiated 4HNE+ and 4HNE- cells in each of the anatomical compartments. Specifically, principal component and linear discriminant analyses of mid-infrared spectra obtained from these cells revealed unambiguous, statistically highly significant differences in the "biochemical fingerprint" of 4HNE+ vs. 4HNE- luminal and basal ME cells, as well as between associated intralobular and interlobular stroma. These findings demonstrate further SR-FTIR microspectroscopy's ability to identify molecular changes associated with altered physiological and/or pathophysiological states, in this case with a state of chronic OxS that provides a pro-carcinogenic microenvironment.

Topics: Adult; Aldehydes; Biomarkers; Breast; Breast Neoplasms; Cell Transformation, Neoplastic; Epithelial Cells; Female; Humans; In Vitro Techniques; Oxidative Stress; Reference Values; Spectroscopy, Fourier Transform Infrared; Stromal Cells; Young Adult

The rate of onset of hepatocellular carcinoma (HCC) in patients with nonalcoholic steatohepatitis (NASH) has been reported recently to be comparable to that of patients with chronic hepatitis C. However, the precise mechanism contributing to carcinogenesis in the former remains unclear. Although increased oxidative stress is presumed to play a role in carcinogenesis in patients with NASH, this relationship remains to be directly proven. In this study, we investigated the involvement of oxidative DNA damage in hepatocarcinogenesis in patients with NASH.. Patients with nonalcoholic fatty liver disease who were treated at our university hospital were eligible for enrolment in the study(n = 49). The study cohort included 30 patients with NASH without HCC (NASH without HCC), six HCC patients with NASH (NASH-HCC), and 13 patients with simple steatosis. Quantitative immunohistochemistry with a KS-400 image analyzing system was used for 8-hydroxy-2'-deoxyguanosine (8-OHdG) detection.. The 8-OHdG content in the liver tissue of NASH-HCC patients was significantly different from that in the other patients. The median immunostaining intensity was 8.605 in the NASH-HCC cases, which was significantly higher than that in the cases of NASH without HCC (4.845; P = 0.003). Multivariate analysis using hepatic 8-OHdG content as a factor in addition to age and fasting blood sugar revealed a significant difference in clinicopathological factors between NASH-HCC and NASH without HCC cases. Old age (P = 0.015) and high relative immunostaining intensity for intrahepatic 8-OHdG (P = 0.037) were identified as independent factors.. 8-OHdG content in liver tissue may serve a marker of oxidative stress and could be a particularly useful predictor of hepatocarcinogenesis.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Aged; Aged, 80 and over; Aldehydes; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Deoxyguanosine; DNA Damage; DNA, Neoplasm; Fatty Liver; Female; Humans; Liver; Liver Neoplasms; Male; Middle Aged; Neoplasm Staging; Non-alcoholic Fatty Liver Disease; Oxidative Stress; Young Adult

Current knowledge of changes in the mammary epithelium relevant to breast carcinogenesis is limited to when histological changes are already present because of a lack of biomarkers needed to identify where such molecular changes might be ongoing at earlier during the of decades-long latent stages of breast carcinogenesis. Breast reduction tissues from young women and teenagers, representative of USA's high breast cancer incidence population, were studies using immunocytochemistry and targeted PCR arrays in order to learn whether a marker of chronic oxidative-stress [protein adducts of 4-hydroxy-2-nonenal (4HNE)] can identify where molecular changes relevant to carcinogenesis might be taking place prior to any histological changes. 4HNE-immunopositive (4HNE+) mammary epithelial cell-clusters were identified in breast tissue sections from most women and from many teenagers (ages 14-30 y) and, in tissues from women ages 17-27 y with many vs. few 4HNE+ cells, the expression of 30 of 84 oxidative-stress associated genes was decreased and only one was increased > 2-fold. This is in contrast to increased expression of many of these genes known to be elicited by acute oxidative-stress. The findings validate using 4HNE-adducts to identify where molecular changes of potential relevance to carcinogenesis are taking place in histologically normal mammary epithelium and highlight differences between responses to acute vs. chronic oxidative-stress. We posit that the altered gene expression in 4HNE+ tissues reflect adaptive responses to chronic oxidative-stress that enable some cells to evade mechanisms that have evolved to prevent propagation of cells with oxidatively-damaged DNA and to accrue heritable changes needed to establish a cancer.

Topics: Adolescent; Adult; Aldehydes; Cell Transformation, Neoplastic; Epithelial Cells; Epithelium; Female; Gene Expression Profiling; Genes, Essential; Humans; Mammary Glands, Human; Oxidative Stress; Reproducibility of Results; United States; Young Adult

To clarify the role of oxidative stress during breast carcinogenesis by studying the expression of 8-hydroxydeoxyguanosine (8-OHdG) (a marker of oxidative DNA damage) and 4-hydroxy-2-nonenal (HNE) (a marker of lipid peroxidation) during the different phases of breast carcinogenesis.. The study material consisted of a total of 219 patients: 31 with usual ductal hyperplasia (UDH), 25 with atypical ductal hyperplasia (ADH), 30 with ductal carcinoma in situ (DCIS) and 133 with invasive carcinoma. The expression of 8-OHdG and HNE were evaluated immunohistochemically. Both 8-OHdG (77.4%) and HNE (45.8%) expression was already seen in UDH lesions. Interestingly, the trend of these two immunostainings during breast carcinogenesis was diverse. 8-OHdG expression diminished significantly in invasive breast carcinomas compared to non-invasive lesions (P < 0.005 when set against non-invasive cohorts). Also within the same lesions, 8-OHdG expression was the most intensive in benign cells. Conversely, HNE immunostaining was strongest in invasive breast carcinomas (UDH versus invasive cohort, P = 0.015).. 4-hydroxy-2-nonenal as a marker of lipid peroxidation increases during breast carcinogenesis, reflecting the role of oxidative stress in the pathogenesis of breast cancer. However, 8-OHdG shows diminished levels in carcinomas, possibly resulting from the induction of DNA repair in these invasive lesions.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Adult; Aged; Aged, 80 and over; Aldehydes; Biomarkers, Tumor; Breast Neoplasms; Carcinoma, Ductal, Breast; Carcinoma, Intraductal, Noninfiltrating; Cell Transformation, Neoplastic; Deoxyguanosine; DNA Repair Enzymes; Female; Humans; Hyperplasia; Lipid Peroxidation; Middle Aged; Oxidative Stress; Retrospective Studies

Previously, we have shown that overexpression of 4-hydroxy-2-nonenal (HNE)-detoxifying enzyme glutathione S-transferase A4-4 (hGSTA4-4) in human lens epithelial cells (HLE B-3) leads to pro-carcinogenic phenotypic transformation of these cells [R. Sharma, et al. Eur. J. Biochem. 271 (2004) 1960-1701]. We now demonstrate that hGSTA4-transfection also causes a profound change in the expression of genes involved in cell adhesion, cell cycle control, proliferation, cell growth, and apoptosis, which is consistent with phenotypic changes of the transformed cells. The expression of p53, p21, p16, fibronectin 1, laminin gamma1, connexin 43, Fas, integrin alpha6, TGFalpha, and c-jun was down-regulated, while the expression of protein kinase C beta II (PKCbetaII), c-myc, cyclin-dependent kinase 2 (CDK2), and TGFbeta was up-regulated in transfected cells. These results demonstrate that HNE serves as a crucial signaling molecule and, by modulating the expression of genes, can influence cellular functions.

Topics: Aldehydes; Cell Line; Cell Transformation, Neoplastic; Epithelium, Corneal; Gene Expression Profiling; Gene Expression Regulation; Glutathione Transferase; Humans; Recombinant Proteins; Signal Transduction; Transfection

Trans-4-hydroxy-2-nonenal (4-HNE), a major electrophilic by-product of lipid peroxidation, is able to interact with DNA to form exocyclic guanine adducts. 4-HNE is a mutagen and a significant amount of 4-HNE-guanine adduct has been detected in normal cells. Recently, it has been reported that exposure of the wild-type p53 human lymphoblastoid cell line to 4-HNE causes a high frequency of G to T transversion mutations at the third base of codon 249 (-AGG*-) in the p53 gene, a mutational hotspot in human cancers, particularly hepatocellular carcinoma. These findings raise a possibility that 4-HNE could be an important etiological agent for human cancers that have a mutation at codon 249 of the p53 gene. However, to date, the sequence specificity of 4-HNE-DNA binding remains unclear due to the lack of methodology. To address this question, we have developed a method, using UvrABC nuclease, a nucleotide excision repair enzyme complex isolated from Escherichia coli, to map the distribution of 4-HNE-DNA adducts in human p53 gene at the nucleotide sequence level. We found that 4-HNE-DNA adducts are preferentially formed at the third base of codon 249 in the p53 gene. The preferential binding of 4-HNE was also observed at codon 174, which has the same sequence and the same nearest neighbor sequences (-GAGG*C-) as codon 249. These results suggest that 4-HNE may be an important etiological agent for human cancers that have a mutation at codon 249 of the p53 gene.

Topics: Aldehydes; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Codon; CpG Islands; DNA Adducts; DNA Damage; DNA Methylation; DNA, Bacterial; DNA, Neoplasm; DNA, Superhelical; Endodeoxyribonucleases; Escherichia coli Proteins; Genes, p53; Humans; Lipid Peroxidation; Liver Neoplasms; Neoplasms; Plasmids

The susceptibility of rat liver tissue to oxidative stress during its neoplastic transformation was analyzed by both qualitative and quantitative measurements of the carbonyl products of lipid peroxidation. Diethylnitrosamine was used as initiating agent of hepatocarcinogenesis and lipid peroxidation levels were monitored in the homogenates from normal liver, hyperplastic nodules and tumour, incubated in the presence or in the absence of ascorbate or adenosine diphosphate-iron complex. While the basal levels of lipid peroxidation in the three experimental conditions were found to be quite similar, in the presence of the pro-oxidant stimulus a remarkable reduction in aldehyde production was shown not only by the hepatoma tissue but also by the preneoplastic nodules.

Topics: Aldehydes; Animals; Arachidonic Acid; Arachidonic Acids; Cell Transformation, Neoplastic; Chromatography, High Pressure Liquid; Diethylnitrosamine; Fatty Acids; Hyperplasia; Lipid Peroxides; Liver; Liver Neoplasms; Liver Neoplasms, Experimental; Male; Malondialdehyde; Oxidation-Reduction; Phosphatidylethanolamines; Rats; Rats, Inbred Strains