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Expression Patterns of Xenobiotic-Metabolizing Enzymes in Tumor and Adjacent Normal Mucosa Tissues among Patients with Colorectal Cancer: The ColoCare Study.
Beyerle, J, Holowatyj, AN, Haffa, M, Frei, E, Gigic, B, Schrotz-King, P, Boehm, J, Habermann, N, Stiborova, M, Scherer, D, et al
Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology. 2020;(2):460-469
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Abstract
BACKGROUND Xenobiotic-metabolizing enzymes (XME) play a critical role in the activation and detoxification of several carcinogens. However, the role of XMEs in colorectal carcinogenesis is unclear. METHODS We investigated the expression of XMEs in human colorectal tissues among patients with stage I-IV colorectal cancer (n = 71) from the ColoCare Study. Transcriptomic profiling using paired colorectal tumor and adjacent normal mucosa tissues of XMEs (GSTM1, GSTA1, UGT1A8, UGT1A10, CYP3A4, CYP2C9, GSTP1, and CYP2W1) by RNA microarray was compared using Wilcoxon rank-sum tests. We assessed associations between clinicopathologic, dietary, and lifestyle factors and XME expression with linear regression models. RESULTS GSTM1, GSTA1, UGT1A8, UGT1A10, and CYP3A4 were all statistically significantly downregulated in colorectal tumor relative to normal mucosa tissues (all P ≤ 0.03). Women had significantly higher expression of GSTM1 in normal tissues compared with men (β = 0.37, P = 0.02). By tumor site, CYP2C9 expression was lower in normal mucosa among patients with rectal cancer versus colon cancer cases (β = -0.21, P = 0.0005). Smokers demonstrated higher CYP2C9 expression levels in normal mucosa (β = 0.17, P = 0.02) when compared with nonsmokers. Individuals who used NSAIDs had higher GSTP1 tumor expression compared with non-NSAID users (β = 0.17, P = 0.03). Higher consumption of cooked vegetables (>1×/week) was associated with higher CYP3A4 expression in colorectal tumor tissues (β = 0.14, P = 0.007). CONCLUSIONS XMEs have lower expression in colorectal tumor relative to normal mucosa tissues and may modify colorectal carcinogenesis via associations with clinicopathologic, lifestyle, and dietary factors. IMPACT Better understanding into the role of drug-metabolizing enzymes in colorectal cancer may reveal biological differences that contribute to cancer development, as well as treatment response, leading to clinical implications in colorectal cancer prevention and management.
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BDNF Val66Met Polymorphism, the Allele-Specific Analysis by qRT-PCR - a Novel Protocol.
de Assis, GG, Hoffman, JR, Gasanov, EV
International journal of medical sciences. 2020;(18):3058-3064
Abstract
Background: Alteration in brain-derived neurotrophic factor (BDNF) production is a marker of neuropathological conditions, which has led to the investigation of Val66Met polymorphism occurring in the human BDNF gene (BDNF). Presently, there are no reported methods available for the analysis of Val66Met impact on human BDNF functioning. Purpose: To develop a qRT-PCR protocol for the allele-specific expression evaluation of the Val66Met polymorphism in BDNF. Methods: Using RNA extracted from muscle samples of 9 healthy volunteers (32.9 ± 10.3 y) at rest and following a maximal effort aerobic capacity exercise test, a protocol was developed for the detection of Val66/Met66 allele-specific BDNF expression in Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) - relative to housekeeping genes - and validated by absolute quantification in Droplet Digital Polymerase Chain Reaction (ddPCR). Results: Differences in the relative values of BDNF mRNA were confirmed by ddPCR analysis. HPRT1 and B2M were the most stable genes expressed in muscle tissue among different metabolic conditions, while GAPDH revealed to be metabolic responsive. Conclusion: Our qRT-PCR protocol successfully determines the allele-specific detection and changes in BDNF expression regarding the Val66Met polymorphism.
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Developmental programming in human umbilical cord vein endothelial cells following fetal growth restriction.
Terstappen, F, Calis, JJA, Paauw, ND, Joles, JA, van Rijn, BB, Mokry, M, Plösch, T, Lely, AT
Clinical epigenetics. 2020;(1):185
Abstract
BACKGROUND Fetal growth restriction (FGR) is associated with an increased susceptibility for various noncommunicable diseases in adulthood, including cardiovascular and renal disease. During FGR, reduced uteroplacental blood flow, oxygen and nutrient supply to the fetus are hypothesized to detrimentally influence cardiovascular and renal programming. This study examined whether developmental programming profiles, especially related to the cardiovascular and renal system, differ in human umbilical vein endothelial cells (HUVECs) collected from pregnancies complicated by placental insufficiency-induced FGR compared to normal growth pregnancies. Our approach, involving transcriptomic profiling by RNA-sequencing and gene set enrichment analysis focused on cardiovascular and renal gene sets and targeted DNA methylation assays, contributes to the identification of targets underlying long-term cardiovascular and renal diseases. RESULTS Gene set enrichment analysis showed several downregulated gene sets, most of them involved in immune or inflammatory pathways or cell cycle pathways. seven of the 22 significantly upregulated gene sets related to kidney development and four gene sets involved with cardiovascular health and function were downregulated in FGR (n = 11) versus control (n = 8). Transcriptomic profiling by RNA-sequencing revealed downregulated expression of LGALS1, FPR3 and NRM and upregulation of lincRNA RP5-855F14.1 in FGR compared to controls. DNA methylation was similar for LGALS1 between study groups, but relative hypomethylation of FPR3 and hypermethylation of NRM were present in FGR, especially in male offspring. Absolute differences in methylation were, however, small. CONCLUSION This study showed upregulation of gene sets related to renal development in HUVECs collected from pregnancies complicated by FGR compared to control donors. The differentially expressed gene sets related to cardiovascular function and health might be in line with the downregulated expression of NRM and upregulated expression of lincRNA RP5-855F14.1 in FGR samples; NRM is involved in cardiac remodeling, and lincRNAs are correlated with cardiovascular diseases. Future studies should elucidate whether the downregulated LGALS1 and FPR3 expressions in FGR are angiogenesis-modulating regulators leading to placental insufficiency-induced FGR or whether the expression of these genes can be used as a biomarker for increased cardiovascular risk. Altered DNA methylation might partly underlie FPR3 and NRM differential gene expression differences in a sex-dependent manner.
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Gene Expression Profiling of the Pancreas in Patients Undergoing Total Pancreatectomy With Islet Autotransplant Suggests Unique Features of Alcoholic, Idiopathic, and Hereditary Pancreatitis.
Blobner, BM, Bellin, MD, Beilman, GJ, Shelton, CA, Park, HJ, Whitcomb, DC
Pancreas. 2020;(8):1037-1043
Abstract
OBJECTIVES To determine if RNA collected from pancreatic tissue, without the use of RNAlater, is useful for RNA sequencing (RNA-seq) despite degradation, and if so, then, via RNA-seq analysis, how does gene expression vary between pancreatitis etiologies. METHODS Data were assessed in 2 dimensions, based on RNA-seq signal shape assessed by RSeQC v.2.6.4 and RNA expression after accounting for different degrees of degradation. RESULTS Six measures of RNA characteristics (median RNA fragment size, reads per million kilobases saturation, transcript integrity number, distribution of hexamers, percentage of nucleotides that are guanine or cytosine, and duplicated reads) were significantly different between hereditary pancreatitis and idiopathic pancreatitis. Differential expression analysis revealed that 150 genes were differentially expressed between hereditary and idiopathic etiologies, 197 genes were differentially expressed between alcoholic and idiopathic etiologies, and 200 genes were differentially expressed between alcoholic and hereditary etiologies. We then determined that many enriched pathways between hereditary and idiopathic etiologies are related to the matrisome, and many of the enriched pathways between alcoholic and idiopathic etiology or hereditary etiology are related to ion transport. CONCLUSIONS We found distinct RNA-seq signals between different pancreatitis etiologies in both of the dimensions in critical pathways for pancreas biology.
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Concordance between gene expression in peripheral whole blood and colonic tissue in children with inflammatory bowel disease.
Palmer, NP, Silvester, JA, Lee, JJ, Beam, AL, Fried, I, Valtchinov, VI, Rahimov, F, Kong, SW, Ghodoussipour, S, Hood, HC, et al
PloS one. 2019;(10):e0222952
Abstract
BACKGROUND Presenting features of inflammatory bowel disease (IBD) are non-specific. We hypothesized that mRNA profiles could (1) identify genes and pathways involved in disease pathogenesis; (2) identify a molecular signature that differentiates IBD from other conditions; (3) provide insight into systemic and colon-specific dysregulation through study of the concordance of the gene expression. METHODS Children (8-18 years) were prospectively recruited at the time of diagnostic colonoscopy for possible IBD. We used transcriptome-wide mRNA profiling to study gene expression in colon biopsies and paired whole blood samples. Using blood mRNA measurements, we fit a regression model for disease state prediction that was validated in an independent test set of adult subjects (GSE3365). RESULTS Ninety-eight children were recruited [39 Crohn's disease, 18 ulcerative colitis, 2 IBDU, 39 non-IBD]. There were 1,118 significantly differentially (IBD vs non-IBD) expressed genes in colon tissue, and 880 in blood. The direction of relative change in expression was concordant for 106/112 genes differentially expressed in both tissue types. The regression model from the blood mRNA measurements distinguished IBD vs non-IBD disease status in the independent test set with 80% accuracy using only 6 genes. The overlap of 5 immune and metabolic pathways in the two tissue types was significant (p<0.001). CONCLUSIONS Blood and colon tissue from patients with IBD share a common transcriptional profile dominated by immune and metabolic pathways. Our results suggest that peripheral blood expression levels of as few as 6 genes (IL7R, UBB, TXNIP, S100A8, ALAS2, and SLC2A3) may distinguish patients with IBD from non-IBD.
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Anti-Inflammatory and Tissue Regenerative Effects of Topical Treatment with Ozonated Olive Oil/Vitamin E Acetate in Balanitis Xerotica Obliterans.
Currò, M, Russo, T, Ferlazzo, N, Caccamo, D, Antonuccio, P, Arena, S, Parisi, S, Perrone, P, Ientile, R, Romeo, C, et al
Molecules (Basel, Switzerland). 2018;(3)
Abstract
Balanitis xerotica obliterans (BXO) is a chronic inflammatory skin disorder, considered the male genital variant of lichen sclerosus. Anti-inflammatory drugs are commonly used in BXO. We evaluated the effects of an innovative formulation of ozonated olive oil with vitamin E acetate (OZOILE®) on the inflammatory status and tissue remodeling in male children with BXO. The mRNA transcripts of proteins involved either in inflammation or in dynamics of tissue regeneration were analyzed by quantitative real-time PCR, in foreskins affected by BXO removed from patients untreated or treated with OZOILE® cream for 7 days before circumcision. We found a significant reduction in mRNA levels of IL-1β, TNF-α, INF-γ, transglutaminase 2 and NOS2 in foreskins treated with OZOILE® in comparison to untreated ones (p < 0.001). No significant differences were observed in NF-κB activation in the specimens obtained from treated and untreated patients. Hence, OZOILE® treatment up-regulated hypoxia-inducible factor (HIF)-1alpha, vascular endothelial growth factor (VEGF) and E-cadherin gene expression (p < 0.001). The treatment with OZOILE® showed effective results in children affected by BXO by reducing the inflammatory process and stimulating mechanisms for tissue regeneration of the foreskin. A randomized clinical trial on a large number of children affected by BXO might be useful to verify the efficacy of topical treatment with OZOILE®.
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Differential Gene Expression in Prostate Tissue According to Ejaculation Frequency.
Sinnott, JA, Brumberg, K, Wilson, KM, Ebot, EM, Giovannucci, EL, Mucci, LA, Rider, JR
European urology. 2018;(5):545-548
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Abstract
UNLABELLED In a prospective study of 31 925 men with 18 yr of follow-up, higher ejaculation frequency (EF) throughout adulthood was associated with lower rates of prostate cancer. To further explore this association, we evaluated whole transcriptome gene expression in the prostate tissue from study participants who developed prostate cancer between 1992 and 2004 (n=157 tumor tissue, n=85 adjacent normal). We tested for trends in gene expression according to the level of EF as self-reported in 1992 for ages 20-29 yr, 40-49 yr, and the year prior to the questionnaire, 1991. There were no associations between EF and gene expression in areas of tumor after accounting for multiple testing. In contrast, in the adjacent normal tissue, 409 genes and six pathways were differentially expressed at a false discovery rate ≤0.2 across categories of EF in 1991. These results suggest that ejaculation affects the expression of genes in the normal prostate tissue. The identified genes and pathways provide potential biological links between EF and prostate tumorigenesis. PATIENT SUMMARY To explore previous findings that men who ejaculate more frequently have lower risk of prostate cancer, we evaluated molecular alterations in the prostate tissue according to each man's frequency of ejaculation prior to diagnosis. We identified biological processes that could link ejaculation frequency and prostate cancer.
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Removing Batch Effects from Longitudinal Gene Expression - Quantile Normalization Plus ComBat as Best Approach for Microarray Transcriptome Data.
Müller, C, Schillert, A, Röthemeier, C, Trégouët, DA, Proust, C, Binder, H, Pfeiffer, N, Beutel, M, Lackner, KJ, Schnabel, RB, et al
PloS one. 2016;(6):e0156594
Abstract
Technical variation plays an important role in microarray-based gene expression studies, and batch effects explain a large proportion of this noise. It is therefore mandatory to eliminate technical variation while maintaining biological variability. Several strategies have been proposed for the removal of batch effects, although they have not been evaluated in large-scale longitudinal gene expression data. In this study, we aimed at identifying a suitable method for batch effect removal in a large study of microarray-based longitudinal gene expression. Monocytic gene expression was measured in 1092 participants of the Gutenberg Health Study at baseline and 5-year follow up. Replicates of selected samples were measured at both time points to identify technical variability. Deming regression, Passing-Bablok regression, linear mixed models, non-linear models as well as ReplicateRUV and ComBat were applied to eliminate batch effects between replicates. In a second step, quantile normalization prior to batch effect correction was performed for each method. Technical variation between batches was evaluated by principal component analysis. Associations between body mass index and transcriptomes were calculated before and after batch removal. Results from association analyses were compared to evaluate maintenance of biological variability. Quantile normalization, separately performed in each batch, combined with ComBat successfully reduced batch effects and maintained biological variability. ReplicateRUV performed perfectly in the replicate data subset of the study, but failed when applied to all samples. All other methods did not substantially reduce batch effects in the replicate data subset. Quantile normalization plus ComBat appears to be a valuable approach for batch correction in longitudinal gene expression data.
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Differential systemic gene expression profile in patients with diabetic macular edema: responders versus nonresponders to standard treatment.
Dabir, SS, Das, D, Nallathambi, J, Mangalesh, S, Yadav, NK, Schouten, JS
Indian journal of ophthalmology. 2014;(1):66-73
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Abstract
INTRODUCTION Diabetic macular edema (DME) is a vision-threatening complication of diabetic retinopathy. The current practice of management is a" trial and error "method of using intravitreal antivascular endothelial growth factor (VEGF)'' or steroids to treat the patient and watch the response. However, if the patient's genetic profile helps us choose appropriate medicine, it would help customize treatment option for each patient. This forms the basis of our study. MATERIALS AND METHODS A case-control, prospective, observational series, where DME patients were treated with bevacizumab and subclassified as treatment naοve, treatment responders, and treatment nonresponders. Blood samples of 20 subjects were studied, with five patients in each of the groups (nondiabetic- group 1, treatment naοve- group 2, treatment responder- group 3, and treatment nonresponder-group 4). Whole blood RNA extraction followed by labeling, amplification and hybridization was done, and microarray data analyzed. Genes were classified based on functional category and pathways. RESULTS The total number of genes upregulated among all three experimental groups was 5, whereas 105 genes were downregulated. There were no common genes upregulated between the responders and nonresponders. There was only one gene upregulated between the diabetic and diabetic responders posttreatment. There were 19 genes upregulated and 8 genes downregulated in the inflammatory pathway in group 2 versus group 1. There were no downregulated genes detected in vascular angiogenesis and transcription group. There were identical numbers of genes up- and downregulated in the inflammatory pathway. Seventeen genes were upreguated and 11 genes downregulated in receptor activity, which remained the predominant group in the group classification. DISCUSSION In summary, this study would provide an insight into the probable signaling mechanisms for disease pathogenesis as well as progression. This type of study eventually would aid in developing or improvising existing treatment modules with a rational approach towards personalized medicine, in future addressing the differential responses to treatment.