Challenges in Defining a Reference Set of Differentially Expressed lncRNAs in Ulcerative Colitis by Meta-Analysis.

The study aimed to identify common differentially expressed lncRNAs from manually curated ulcerative colitis (UC) gene expression omnibus (GEO) datasets. Nine UC transcriptomic datasets of clearly annotated human colonic biopsies were included in the study. The datasets were manually curated to select active UC samples and controls. R packages geneknitR, gprofiler, clusterProfiler were used for gene symbol annotation. The R EdgeR package was used to analyze differential expression. This resulted in a total of nineteen lncRNAs that were differentially expressed in at least three datasets of the nine GEO datasets. Several of the differentially expressed lncRNAs found in UC were associated with promoting colorectal cancer (CRC) through regulating gene expression, epithelial to mesenchymal transition (EMT), cell cycle progression, and by promoting tumor proliferation, invasion, and migration. The expression of several lncRNAs varied between disease states and tissue locations within the same disease state. The identified differentially expressed lncRNAs may function as general markers for active UC independent of biopsy location, age, gender, or treatment, thereby representing a comparative resource for future comparisons using available GEO UC datasets.

DNA methylation fine-tunes pro-and anti-inflammatory signalling pathways in inactive ulcerative colitis tissue biopsies.

DNA methylation has been implied to play a role in the immune dysfunction associated with inflammatory bowel disease (IBD) and the disease development of ulcerative colitis (UC). Changes of the DNA methylation and correlated gene expression in patient samples with inactive UC might reveal possible regulatory features important for further treatment options for UC. Targeted bisulfite sequencing and whole transcriptome sequencing were performed on mucosal biopsies from patients with active UC (UC, n = 14), inactive UC (RM, n = 20), and non-IBD patients which served as controls (NN, n = 11). The differentially methylated regions (DMRs) were identified by DMRseq. Correlation analysis was performed between DMRs and their nearest differentially expressed genes (DEGs). Principal component analysis (PCA) was performed based on correlated DMR regulated genes. DMR regulated genes then were functional annotated. Cell-type deconvolutions were performed based on methylation levels. The comparisons revealed a total of 38 methylation-regulated genes in inactive UC that are potentially regulated by DMRs (correlation p value < 0.1). Several methylation-regulated genes could be identified in inactive UC participating in IL-10 and cytokine signalling pathways such as IL1B and STAT3. DNA methylation events in inactive UC seem to be fine-tuned by the balancing pro- and anti- inflammatory pathways to maintain a prevailed healing process to restore dynamic epithelium homeostasis.

Methylation-Regulated Long Non-Coding RNA Expression in Ulcerative Colitis.

Long non-coding RNAs (lncRNAs) have been shown to play a role in the pathogenesis of ulcerative colitis (UC). Although epigenetic processes such as DNA methylation and lncRNA expression are well studied in UC, the importance of the interplay between the two processes has not yet been fully explored. It is, therefore, believed that interactions between environmental factors and epigenetics contribute to disease development. Mucosal biopsies from 11 treatment-naïve UC patients and 13 normal controls were used in this study. From each individual sample, both whole-genome bisulfite sequencing data (WGBS) and lncRNA expression data were analyzed. Correlation analysis between lncRNA expression and upstream differentially methylated regions (DMRs) was used to identify lncRNAs that might be regulated by DMRs. Furthermore, proximal protein-coding genes associated with DMR-regulated lncRNAs were identified by correlating their expression. The study identified UC-associated lncRNAs such as MIR4435-2HG, ZFAS1, IL6-AS1, and Pvt1, which may be regulated by DMRs. Several genes that are involved in inflammatory immune responses were found downstream of DMR-regulated lncRNAs, including SERPINB1, CCL18, and SLC15A4. The interplay between lncRNA expression regulated by DNA methylation in UC might improve our understanding of UC pathogenesis.

Anti-apoptotic genes and non-coding RNAs are potential outcome predictors for ulcerative colitis.

Due to the lack of clinical, immunologic, genetic, and laboratory markers to predict remission in ulcerative colitis (UC) without relapse, there is no clear recommendation regarding withdrawal of therapy. Therefore, this study was to investigate if transcriptional analysis together with Cox survival analysis might be able to reveal molecular markers that are specific for remission duration and outcome. Mucosal biopsies from patients in remission with active treatment-naïve UC and healthy control subjects underwent whole-transcriptome RNA-seq. Principal component analysis (PCA) and Cox proportional hazards regression analysis were applied to the remission data concerning duration and status of patients. A randomly chosen remission sample set was used for validation of the applied methods and results. The analyses distinguished two different UC remission patient groups with respect to remission duration and outcome (relapse). Both groups showed that altered states of UC with quiescent microscopic disease activity are still present. The patient group with the longest remission duration and no relapse revealed specific and increased expression of antiapoptotic factors belonging to the MTRNR2-like gene family and non-coding RNAs. In summary, the expression of anti-apoptotic factors and non-coding RNAs may contribute to personalized medicine approaches in UC by improving patient stratification for different treatment regimens.

Associations of breast cancer related exposures and gene expression profiles in normal breast tissue-The Norwegian Women and Cancer normal breast tissue study.

BACKGROUND: Normal breast tissue is utilized in tissue-based studies of breast carcinogenesis. While gene expression in breast tumor tissue is well explored, our knowledge of transcriptomic signatures in normal breast tissue is still incomplete. The aim of this study was to investigate variability of gene expression in a large sample of normal breast tissue biopsies, according to breast cancer related exposures (obesity, smoking, alcohol, hormone therapy, and parity). METHODS: We analyzed gene expression profiles from 311 normal breast tissue biopsies from cancer-free, post-menopausal women, using Illumina bead chip arrays. Principal component analysis and K-means clustering was used for initial analysis of the dataset. The association of exposures and covariates with gene expression was determined using linear models for microarrays. RESULTS: Heterogeneity of the breast tissue and cell composition had the strongest influence on gene expression profiles. After adjusting for cell composition, obesity, smoking, and alcohol showed the highest numbers of associated genes and pathways, whereas hormone therapy and parity were associated with negligible gene expression differences. CONCLUSION: Our results provide insight into associations between major exposures and gene expression profiles and provide an informative baseline for improved understanding of exposure-related molecular events in normal breast tissue of cancer-free, post-menopausal women.

Endogenous IL-1 receptor antagonist restricts healthy and malignant myeloproliferation.

Here we explored the role of interleukin-1ß (IL-1ß) repressor cytokine, IL-1 receptor antagonist (IL-1rn), in both healthy and abnormal hematopoiesis. Low IL-1RN is frequent in acute myeloid leukemia (AML) patients and represents a prognostic marker of reduced survival. Treatments with IL-1RN and the IL-1ß monoclonal antibody canakinumab reduce the expansion of leukemic cells, including CD34+ progenitors, in AML xenografts. In vivo deletion of IL-1rn induces hematopoietic stem cell (HSC) differentiation into the myeloid lineage and hampers B cell development via transcriptional activation of myeloid differentiation pathways dependent on NFκB. Low IL-1rn is present in an experimental model of pre-leukemic myelopoiesis, and IL-1rn deletion promotes myeloproliferation, which relies on the bone marrow hematopoietic and stromal compartments. Conversely, IL-1rn protects against pre-leukemic myelopoiesis. Our data reveal that HSC differentiation is controlled by balanced IL-1ß/IL-1rn levels under steady-state, and that loss of repression of IL-1ß signaling may underlie pre-leukemic lesion and AML progression.

Identifying predictive signalling networks for Vedolizumab response in ulcerative colitis.

BACKGROUND: In ulcerative colitis (UC), the molecular mechanisms that drive disease development and patient response to therapy are not well understood. A significant proportion of patients with UC fail to respond adequately to biologic therapy. Therefore, there is an unmet need for biomarkers that can predict patients' responsiveness to the available UC therapies as well as ascertain the most effective individualised therapy. Our study focused on identifying predictive signalling pathways that predict anti-integrin therapy response in patients with UC. METHODS: We retrieved and pre-processed two publicly accessible gene expression datasets (GSE73661 and GSE72819) of UC patients treated with anti-integrin therapies: (1) 12 non-IBD controls and 41 UC patients treated with Vedolizumab therapy, and (2) 70 samples with 58 non-responder and 12 responder UC patient samples treated with Etrolizumab therapy without non-IBD controls. We used a diffusion-based signalling model which is mainly focused on the T-cell receptor signalling network. The diffusion model uses network connectivity between receptors and transcription factors. RESULTS: The network diffusion scores were able to separate VDZ responder and non-responder patients before treatment better than the original gene expression. On both anti-integrin treatment datasets, the diffusion model demonstrated high predictive performance for discriminating responders from non-responders in comparison with 'nnet'. We have found 48 receptor-TF pairs identified as the best predictors for VDZ therapy response with AUC ≥ 0.76. Among these receptor-TF predictors pairs, FFAR2-NRF1, FFAR2-RELB, FFAR2-EGR1, and FFAR2-NFKB1 are the top best predictors. For Etrolizumab, we have identified 40 best receptor-TF pairs and CD40-NFKB2 as the best predictor receptor-TF pair (AUC = 0.72). We also identified subnetworks that highlight the network interactions, connecting receptors and transcription factors involved in cytokine and fatty acid signalling. The findings suggest that anti-integrin therapy responses in cytokine and fatty acid signalling can stratify UC patient subgroups. CONCLUSIONS: We identified signalling pathways that may predict the efficacy of anti-integrin therapy in UC patients and personalised therapy alternatives. Our results may lead to the advancement of a promising clinical decision-making tool for the stratification of UC patients.

Novel long non-coding RNAs of relevance for ulcerative colitis pathogenesis.

BACKGROUND AND AIMS: The study aimed to identify yet unknown and uncharacterized long non-coding RNAs (lncRNAs) in treatment-naïve ulcerative colitis (UC), and to define their possible roles in UC pathogenesis. For that purpose, accurate quantification methods for lncRNA transcript detection, multiple and "stringent" strategies were applied. New insights in the regulation of functional genes and pathways of relevance for UC through expression of lncRNAs are expected. METHODS: The study was based on sequencing data derived from a data set consisting of treatment-naïve UC patients (n = 14) and control subjects (n = 16). Two complementary aligners were used to identify lncRNAs. Several different steps were used to validate differential expression including plotting the reads over the annotation for manual inspection. To help determine potential lncRNA involvement in biological processes, KEGG pathway enrichment was done on protein-coding genes which co-expressed with the lncRNAs. RESULTS: A total of 99 lncRNAs were identified in UC. The lncRNAs which were not previously characterized (n = 15) in UC or other autoimmune diseases were selected for down-stream analysis. In total, 602 protein-coding genes correlated with the uncharacterized lncRNAs. KEGG pathway enrichment analysis revealed involvement of lncRNAs in two significantly enriched pathways, lipid and atherosclerosis, and T-cell receptor signaling. CONCLUSION: This study identified a set of 15 yet uncharacterized lncRNAs which may be of importance for UC pathogenesis. These lncRNAs may serve as potential diagnostic biomarkers and might be of use for the development of UC treatment strategies in the future.

Profiling the T Cell Receptor Alpha/Delta Locus in Salmonids.

In jawed vertebrates, two major T cell populations have been characterized. They are defined as α/ß or γ/δ T cells, based on the expressed T cell receptor. Salmonids (family Salmonidae) include two key teleost species for aquaculture, rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar) which constitute important models for fish immunology and important targets for vaccine development. The growing interest to decipher the dynamics of adaptive immune responses against pathogens or vaccines has resulted in recent efforts to sequence the immunoglobulin (IG) or antibodies and T cell receptor (TR) repertoire in these species. In this context, establishing a comprehensive and coherent locus annotation is the fundamental basis for the analysis of high-throughput repertoire sequencing data. We therefore decided to revisit the description and annotation of TRA/TRD locus in Atlantic salmon and two strains of rainbow trout (Swanson and Arlee) using the now available high-quality genome assemblies. Phylogenetic analysis of functional TRA/TRD V genes from these three genomes led to the definition of 25 subgroups shared by both species, some with particular feature. A total of 128 TRAJ genes were identified in Salmo, the majority with a close counterpart in Oncorhynchus. Analysis of expressed TRA repertoire indicates that most TRAV gene subgroups are expressed at mucosal and systemic level. The present work on TRA/TRD locus annotation along with the analysis of TRA repertoire sequencing data show the feasibility and advantages of a common salmonid TRA/TRD nomenclature that allows an accurate annotation and analysis of high-throughput sequencing results, across salmonid T cell subsets.

DNA hypo-methylation facilitates anti-inflammatory responses in severe ulcerative colitis.

Severe ulcerative colitis (UC) is a potentially life-threatening disease with a potential colorectal cancer (CRC) risk. The aim of this study was to explore the relationship between transcriptomic and genome-wide DNA methylation profiles in a well-stratified, treatment-naïve severe UC patient population in order to define specific epigenetic changes that could be responsible for the grade of disease severity. Mucosal biopsies from treatment-naïve severe UC patients (n = 8), treatment-naïve mild UC (n = 8), and healthy controls (n = 8) underwent both whole transcriptome RNA-Seq and genome-wide DNA bisulfite- sequencing, and principal component analysis (PCA), cell deconvolutions and diverse statistical methods were applied to obtain a dataset of significantly differentially expressed genes (DEGs) with correlation to DNA methylation for severe UC. DNA hypo-methylation correlated with approximately 80% of all DEGs in severe UC when compared to mild UC. Enriched pathways of annotated hypo-methylated genes revealed neutrophil degranulation, and immuno-regulatory interactions of the lymphoid system. Specifically, hypo-methylated anti-inflammatory genes found for severe UC were IL10, SIGLEC5, CD86, CLMP and members of inflammasomes NLRP3 and NLRC4. Hypo-methylation of anti-inflammatory genes during severe UC implies an interplay between the epithelium and lamina propria in order to mitigate inflammation in the gut. The specifically DNA hypo-methylated genes found for severe UC can potentially be useful biomarkers for determining disease severity and in the development of new targeted treatment strategies for severe UC patients.

Transcriptional Signatures That Define Ulcerative Colitis in Remission.

BACKGROUND: This study addresses whether existing specific transcriptional profiles can improve and support the current status of the definition of ulcerative colitis (UC) remission apart from the existing endoscopic, histologic, and laboratory scoring systems. For that purpose, a well-stratified UC patient population in remission was compared to active UC and control patients and was investigated by applying the next-generation technology RNA-Seq. METHODS: Mucosal biopsies from patients in remission (n = 14), patients with active UC (n = 14), and healthy control patientss (n = 16) underwent whole-transcriptome RNA-Seq. Principal component analysis, cell deconvolution methods, gene profile enrichment, and pathway enrichment methods were applied to define a specific transcriptional signature of UC in remission. RESULTS: Analyses revealed specific transcriptional signatures for UC in remission with increased expression of genes involved in O-glycosylation (MUC17, MUC3A, MUC5AC, MUC12, SPON1, B3GNT3), ephrin-mediated repulsion of cells (EFNB2E, EFNA3, EPHA10, EPHA1), GAP junction trafficking (TUBA1C, TUBA4A, TUBB4B, GJB3, CLTB), and decreased expression of several toll-like receptors (TLR1, TLR3, TLR5, TLR6). CONCLUSIONS: This study reveals specific transcriptional signatures for remission. Partial restoration and improvement of homeostasis in the epithelial mucus layer and revival of immunological functions were observed. A clear role for bacterial gut flora composition can be implied. The results can be useful for the development of treatment strategies for UC in remission and may be useful targets for further investigations aiming to predict the outcome of UC in the future.

Identifying anti-TNF response biomarkers in ulcerative colitis using a diffusion-based signalling model.

Motivation: Resistance to anti-TNF therapy in subgroups of ulcerative colitis (UC) patients is a major challenge and incurs significant treatment costs. Identification of patients at risk of nonresponse to anti-TNF is of major clinical importance. To date, no quantitative computational framework exists to develop a complex biomarker for the prognosis of UC treatment. Modelling patient-wise receptor to transcription factor (TF) network connectivity may enable personalized treatment. Results: We present an approach for quantitative diffusion analysis between receptors and TFs using gene expression data. Key TFs were identified using pandaR. Network connectivities between immune-specific receptor-TF pairs were quantified using network diffusion in UC patients and controls. The patient-specific network could be considered a complex biomarker that separates anti-TNF treatment-resistant and responder patients both in the gene expression dataset used for model development and separate independent test datasets. The model was further validated in rheumatoid arthritis where it successfully discriminated resistant and responder patients to tocilizumab treatment. Our model may contribute to prognostic biomarkers that may identify treatment-resistant and responder subpopulations of UC patients. Availability and implementation: Software is available at https://github.com/Amy3100/receptor2tfDiffusion. Supplementary information: Supplementary data are available at Bioinformatics Advances online.

Genome-wide DNA Methylation in Treatment-naïve Ulcerative Colitis.

BACKGROUND AND AIMS: The aim of this study was to investigate the genome-wide DNA methylation status in treatment-naïve ulcerative colitis [UC], and to explore the relationship between DNA methylation patterns and gene expression levels in tissue biopsies from a well-stratified treatment-naïve UC patient group. METHODS: Mucosal biopsies from treatment-naïve patients [n = 10], and a healthy control group [n = 11] underwent genome-wide DNA bisulfite sequencing. Principal component analysis [PCA] and diverse statistical methods were applied to obtain a dataset of differentially methylated genes. DNA methylation annotation was investigated using the UCSC Genome Browser. Gene set enrichments were obtained using the Kyoto Encyclopaedia of Genes and Genomes [KEGG] and PANTHER. RESULTS: Of all significantly differentially expressed genes [DEGs], 25% correlated with DNA methylation patterns; 30% of these genes were methylated at CpG sites near their transcription start site [TSS]. Hyper-methylation was observed for genes involved in homeostasis and defence, whereas hypo-methylation was observed for genes playing a role in immune response [i.e. chemokines and interleukins]. Of the differentially DNA methylated genes, 25 were identified as inflammatory bowel disease [IBD] susceptibility genes. Four genes [DEFFA6, REG1B, BTNL3, OLFM4] showed DNA methylation in the absence of known CpG islands. CONCLUSIONS: Genome-wide DNA methylation analysis revealed distinctive functional patterns for hyper-and hypo-methylation in treatment-naïve UC. These distinct patterns could be of importance in the development and pathogenesis of UC. Further investigation of DNA methylation patterns may be useful in the development of the targeting of epigenetic processes, and may allow new treatment and target strategies for UC patients.

Transcriptomic Landscape of Treatment-Naïve Ulcerative Colitis.

BACKGROUND AND AIMS: Ulcerative colitis [UC] is a chronic inflammatory disease that effects the gastrointestinal tract and is considered one of the most prominent and common forms of inflammatory bowel disease [IBD]. This study aimed to define and describe the entire transcriptomic landscape in a well-stratified, treatment-naïve UC patient population compared with control patients by using next-generation technology, RNA-Seq. METHODS: Mucosal biopsies from treatment-naïve UC patients [n = 14], and healthy controls [n = 16] underwent RNA-Seq. Principal component analysis [PCA], cell deconvolution methods, and diverse statistical methods were applied to obtain and characterise a dataset of significantly differentially expressed genes [DEGs]. RESULTS: Analyses revealed 1480 significantly DEGs in treatment-naïve UC when compared with controls. Cell populations of monocytes, T cells, neutrophils, B cells/ lymphoid cells, and myeloid cells were increased during inflammation, whereas the fraction of epithelial cells were reduced in UC, which is reflected by the DEGs; 79 DEGs were identified as IBD susceptibility genes, and 58 DEGs were expressed in a gender-specific manner. MUC5B, REG3A, DEFA5, and IL33 might be considered as colorectal cancer [CRC] risk factors following UC in males. AQP9 together with CLDN2 may have a role regulating tissue-specific physiological properties in tight junctions in UC. An additional functional role for AQP9 in the synthesis and/or the function of mucus can be implied. CONCLUSIONS: This study reveals new potential players in UC pathogenesis in general, and provides evidence for a gender-dependent pathogenesis for UC. These results can be useful for the development of personalised treatment strategies for UC in the future.

The impact of partial hepatectomy on oxidative state in the liver remnant - An in vivo swine model.

BACKGROUND: Previous studies on oxidative state after partial hepatectomy (PHx) report conflicting data on levels of glutathione (GSH) and are mainly presented in rodent models by methodology less sensitive than the present technologies. The current swine model presents GSH levels and the following genetic response post-PHx, utilizing an analytical platform more sensitive and precise than earlier available. METHOD: Twelve pigs were randomized to a PHx- and a control group (n=6 in each). The PHx group had a 60% hepatectomy. Serial in vivo liver biopsies during 12h of anaesthesia post-PHx were analyzed for GSH by liquid chromatography mass spectrometry (LC-MS/MS). Transcriptional alterations of genes (GS, GCLM, GCLC, GR, HGF, NFE2L2, TGFß1) regulating GSH synthesis were measured by real-time PCR. RESULTS: No difference was detected between the GSH levels in the PHx- and the control group during the experiment (P=0.247). Still, decreased gene expression of GS (P=0.026) and NFE2L2 (P=0.014) the first nine hours, and a decrease of TGFß1 (P=0.029) the first seven hours post-PHx was seen in the liver remnant. CONCLUSION: The results show that the liver has an extended capacity to maintain GSH homeostasis during major stress and parenchymal loss, even at the early onset of such trauma. This observation was not explained by increased expression of key genes in GSH pathways. Consequently, the results indicate an inherent compensatory capacity to maintain GSH homeostasis in the reduced organ.

Prognostic value of the MicroRNA regulators Dicer and Drosha in non-small-cell lung cancer: co-expression of Drosha and miR-126 predicts poor survival.

BACKGROUND: Dicer and Drosha are important enzymes for processing microRNAs. Recent studies have exhibited possible links between expression of different miRNAs, levels of miRNA processing enzymes, and cancer prognosis. We have investigated the prognostic impact of Dicer and Drosha and their correlation with miR-126 expression in a large cohort of non-small cell lung cancer (NSCLC) patients. We aimed to find patient groups within the cohort that might have an advantage of receiving adjunctive therapies. METHODS: Dicer expression in the cytoplasm and Drosha expression in the nucleus were evaluated by manual immunohistochemistry of tissue microarrays (TMAs), including tumor tissue samples from 335 patients with resected stages I to IIIA NSCLC. In addition, in situ hybridizations of TMAs for visualization of miR-126 were performed. Kaplan-Meier analysis was performed, and the log-rank test via SPSS v.22 was used for estimating significance levels. RESULTS: In patients with normal performance status (ECOG = 0, n = 197), high Dicer expression entailed a significantly better prognosis than low Dicer expression (P = 0.024). Dicer had no significant prognostic value in patients with reduced performance status (ECOG = 1-2, n = 138). High Drosha expression was significantly correlated with high levels of the microRNA 126 (miR-126) (P = 0.004). Drosha/miR-126 co-expression had a significant negative impact on the disease-specific survival (DSS) rate (P < 0.001). Multivariate analyses revealed that the interaction Dicer*Histology (P = 0.049) and Drosha/miR-126 co-expression (P = 0.033) were independent prognostic factors. CONCLUSIONS: In NSCLC patients with normal performance status, Dicer is a positive prognostic factor. The importance of Drosha as a prognostic factor in our material seems to be related to miR-126 and possibly other microRNAs.

Plasma fatty acid ratios affect blood gene expression profiles--a cross-sectional study of the Norwegian Women and Cancer Post-Genome Cohort.

High blood concentrations of n-6 fatty acids (FAs) relative to n-3 FAs may lead to a "physiological switch" towards permanent low-grade inflammation, potentially influencing the onset of cardiovascular and inflammatory diseases, as well as cancer. To explore the potential effects of FA ratios prior to disease onset, we measured blood gene expression profiles and plasma FA ratios (linoleic acid/alpha-linolenic acid, LA/ALA; arachidonic acid/eicosapentaenoic acid, AA/EPA; and total n-6/n-3) in a cross-section of middle-aged Norwegian women (n = 227). After arranging samples from the highest values to the lowest for all three FA ratios (LA/ALA, AA/EPA and total n-6/n-3), the highest and lowest deciles of samples were compared. Differences in gene expression profiles were assessed by single-gene and pathway-level analyses. The LA/ALA ratio had the largest impact on gene expression profiles, with 135 differentially expressed genes, followed by the total n-6/n-3 ratio (125 genes) and the AA/EPA ratio (72 genes). All FA ratios were associated with genes related to immune processes, with a tendency for increased pro-inflammatory signaling in the highest FA ratio deciles. Lipid metabolism related to peroxisome proliferator-activated receptor γ (PPARγ) signaling was modified, with possible implications for foam cell formation and development of cardiovascular diseases. We identified higher expression levels of several autophagy marker genes, mainly in the lowest LA/ALA decile. This finding may point to the regulation of autophagy as a novel aspect of FA biology which warrants further study. Lastly, all FA ratios were associated with gene sets that included targets of specific microRNAs, and gene sets containing common promoter motifs that did not match any known transcription factors. We conclude that plasma FA ratios are associated with differences in blood gene expression profiles in this free-living population, and that affected genes and pathways may influence the onset and progression of disease.

Cancer-associated fibroblasts from human NSCLC survive ablative doses of radiation but their invasive capacity is reduced.

BACKGROUND: Cancer-Associated Fibroblasts (CAFs) are significant components of solid malignancies and play central roles in cancer sustainability, invasion and metastasis. In this study we have investigated the invasive capacity and matrix remodelling properties of human lung CAFs after exposure to ablative doses of ionizing radiation (AIR), equivalent to single fractions delivered by stereotactic ablative radiotherapy (SART) for medically inoperable stage-I/II non-small-cell lung cancers. METHODS: CAFs were isolated from lung tumour specimens from 16 donors. Initially, intrinsic radiosensitivity was evaluated by checking viability and extent of DNA-damage response (DDR) at different radiation doses. The migrative and invasive capacities of CAFs were thereafter determined after a sub-lethal single radiation dose of 18 Gy. To ascertain the mechanisms behind the altered invasive capacity of cells, expression of matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs) were measured in the conditioned media several days post-irradiation, along with expression of cell surface integrins and dynamics of focal contacts by vinculin-staining. RESULTS: Exposing CAFs to 1 × 18 Gy resulted in a potent induction of multiple nuclear DDR foci (> 9/cell) with little resolution after 120 h, induced premature cellular senescence and inhibition of the proliferative, migrative and invasive capacity. AIR promoted MMP-3 and inhibited MMP-1 appearance to some extent, but did not affect expression of other major MMPs. Furthermore, surface expression of integrins α2, ß1 and α5 was consistently enhanced, and a dramatic augmentation and redistribution of focal contacts was observed. CONCLUSIONS: Our data indicate that ablative doses of radiation exert advantageous inhibitory effects on the proliferative, migratory and invasive capacity of lung CAFs. The reduced motility of irradiated CAFs might be a consequence of stabilized focal contacts via integrins.

Deciphering normal blood gene expression variation--The NOWAC postgenome study.

There is growing evidence that gene expression profiling of peripheral blood cells is a valuable tool for assessing gene signatures related to exposure, drug-response, or disease. However, the true promise of this approach can not be estimated until the scientific community has robust baseline data describing variation in gene expression patterns in normal individuals. Using a large representative sample set of postmenopausal women (N = 286) in the Norwegian Women and Cancer (NOWAC) postgenome study, we investigated variability of whole blood gene expression in the general population. In particular, we examined changes in blood gene expression caused by technical variability, normal inter-individual differences, and exposure variables at proportions and levels relevant to real-life situations. We observe that the overall changes in gene expression are subtle, implying the need for careful analytic approaches of the data. In particular, technical variability may not be ignored and subsequent adjustments must be considered in any analysis. Many new candidate genes were identified that are differentially expressed according to inter-individual (i.e. fasting, BMI) and exposure (i.e. smoking) factors, thus establishing that these effects are mirrored in blood. By focusing on the biological implications instead of directly comparing gene lists from several related studies in the literature, our analytic approach was able to identify significant similarities and effects consistent across these reports. This establishes the feasibility of blood gene expression profiling, if they are predicated upon careful experimental design and analysis in order to minimize confounding signals, artifacts of sample preparation and processing, and inter-individual differences.