Background : The aim of this study is to improve our understanding of the mechanisms underlying the role of sepsis in the limb muscles of ICU patients with acute quadriplegic myopathy (AQM) by using a unique porcine ICU model, i.e., 5-day longitudinal experiments where animals are sedated, mechanically ventilated and exposed to factor triggering AQM that is endotoxin-induced sepsis.
Role of sepsis in the development of limb muscle weakness in a porcine intensive care unit model.
Sex, Specimen part, Disease
View SamplesBackground: The aim of this study is to improve our understanding of the mechanisms underlying the sparing of masticatory muscles relative to limb muscles in ICU patients with acute quadriplegic myopathy (AQM) by using a unique porcine ICU model, i.e., 5-day longitudinal experiments where animals are sedated, mechanically ventilated and exposed to factors triggering AQM, such as muscle unloading, endotoxin-induced sepsis, and systemic exposure to CS and NMBA.
Mechanisms underlying the sparing of masticatory versus limb muscle function in an experimental critical illness model.
Sex, Specimen part, Disease, Disease stage, Treatment, Time
View SamplesThe present work aimed to identify reference genes for RT-qPCR studies of hypoxia in cervical cancer. From 422 candidate reference genes selected from the literature, we used Illumina array-based expression profiles to identify 182 genes not affected by hypoxia treatment in eight cervical cancer cell lines or correlated with the hypoxia-associated dynamic contrast-enhanced magnetic resonance imaging parameter ABrix in 42 patients. Among these genes, we selected nine candidates (CHCHD1, GNB2L1, IPO8, LASP1, RPL27A, RPS12, SOD1, SRSF9, TMBIM6) that were not associated with tumor volume, stage, lymph node involvement or disease progression in array data of 150 patients, for further testing by RT-qPCR. geNorm and NormFinder analyses of RT-qPCR data of 74 patients identified CHCHD1, SRSF9 and TMBIM6 as the most suitable set of reference genes, with stable expression both overall and across patient subgroups with different hypoxia status (ABrix) and clinical parameters. The suitability of the three candidates as reference genes were validated in studies of the hypoxia-induced genes DDIT3, ERO1A, and STC2. After normalizing with CHCHD1, SRSF9 and TMBIM6, the RT-qPCR data of these genes showed a significant correlation with Illumina expression (P<0.001, n=74) and ABrix (P<0.05, n=32), and the STC2 data were associated with clinical outcome, in accordance with the Illumina data. Thus, CHCHD1, SRSF9 and TMBIM6 seem to be suitable reference genes for studying hypoxia-related gene expression in cervical cancer samples by RT-qPCR. STC2 might be a useful prognostic hypoxia biomarker in cervical cancer that warrants further investigation.
Identification and Validation of Reference Genes for RT-qPCR Studies of Hypoxia in Squamous Cervical Cancer Patients.
Specimen part, Cell line
View SamplesThe aryl hydrocarbon receptor (AHR) functions in higher organisims in development, metabolism and toxic responses. Its Caenorhabditis elegans (C. elegans) ortholog, AHR-1, facilitates neuronal development, growth and movement. We investigated the effect of AHR mutation on the transcriptional profile of L4 stage C. elegans using RNA-seq and quantitative real-time PCR in order to understand better AHR-1 function at the genomic level. Illumina HiSeq 2000 sequencing yielded 51.1, 61.2 and 54.0 million reads from wild-type controls, ahr-1(ia03) and ahr-1(ju145) mutants, respectively, providing detection of over 18,000 transcripts in each sample. Fourteen transcripts were over-expressed and 125 under-expressed in both ahr-1 mutants when compared to wild-type. Under-expressed genes included soluble guanylate cyclase (gcy) family genes, some of which were previously demonstrated to be regulated by AHR-1. A neuropeptide-like protein gene, nlp-20, and an F-box domain protein gene fbxa-192 and its pseudogenes fbxa-191 and fbxa-193 were also under-expressed. Conserved xenobiotic response elements were identified in the 5'' flanking regions of some but not all of the gcy, nlp-20 and fbxa genes. These results extend previous studies demonstrating control of gcy family gene expression by AHR-1, and furthermore suggest a role of AHR-1 in regulation of a neuropeptide gene as well as pseudogenes. Overall design: One sample was created from each of the following strains: wild-type N2, ahr-1(ia03) mutant and ahr-1(ju145) mutant. In data analysis, each mutant sample was individually compared to the wild-type sample to find differentially expressed genes.
Transcriptional profiling reveals differential expression of a neuropeptide-like protein and pseudogenes in aryl hydrocarbon receptor-1 mutant Caenorhabditis elegans.
Subject
View SamplesDeficiency of the micronutrient zinc is a widespread condition in agricultural soils, generating a negative impact on crop quality and yield. Nevertheless, there is insufficient knowledge on the regulatory and molecular mechanisms underlying the plant response to inadequate zinc nutrition.
Transcriptomic profiling of Arabidopsis gene expression in response to varying micronutrient zinc supply.
Age, Specimen part
View SamplesEmerging biomarkers based on medical images and molecular characterization of tumor biopsies open up for combining the two disciplines and exploiting their synergy in treatment planning. We compared pretreatment classification of cervical cancer patients by two previously validated imaging- and gene-based hypoxia biomarkers, evaluated the influence of intratumor heterogeneity, and investigated the benefit of combining them in prediction of treatment failure. The imaging-based biomarker was hypoxic fraction, determined from diagnostic dynamic contrast enhanced (DCE)-MR images. The gene-based biomarker was a hypoxia gene expression signature determined from tumor biopsies. Paired data were available for 118 patients. Intratumor heterogeneity was assessed by variance analysis of MR images and multiple biopsies from the same tumor. The two biomarkers were combined using a dimension-reduction procedure. The biomarkers classified 75% of the tumors with the same hypoxia status. Both intratumor heterogeneity and distribution pattern of hypoxia from imaging were unrelated to inconsistent classification by the two biomarkers, and the hypoxia status of the slice covering the biopsy region was representative of the whole tumor. Hypoxia by genes was independent on tumor cell fraction and showed minor heterogeneity across multiple biopsies in 9 tumors. This suggested that the two biomarkers could contain complementary biological information. Combination of the biomarkers into a composite score led to improved prediction of treatment failure (HR:7.3) compared to imaging (HR:3.8) and genes (HR:3.0) and prognostic impact in multivariate analysis with clinical variables. In conclusion, combining imaging- and gene-based biomarkers enables more precise and informative assessment of hypoxia-related treatment resistance in cervical cancer, independent of intratumor heterogeneity.
Combining imaging- and gene-based hypoxia biomarkers in cervical cancer improves prediction of chemoradiotherapy failure independent of intratumour heterogeneity.
Specimen part
View SamplesThis SuperSeries is composed of the SubSeries listed below.
The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells.
Specimen part
View SamplesActivation of CD4 T cells is a reaction to challenges such as microbial pathogens, cancer and toxins that defines adaptive immune responses. The roles of T cell receptor crosslinking, intracellular signaling, and transcription factor activation are well described, but the importance of post-transcriptional regulation by RNA-binding proteins (RBPs) has not been considered in depth. We describe a new model expanding and activating primary human CD4 T cells and applied this to characterizing activation-induced assembly of splicing factors centered on U2AF2. We immunoprecipitated U2AF2 to identify what mRNA transcripts were bound as a function of activation by TCR crosslinking and costimulation. In parallel, mass spectrometry revealed the proteins incorporated into the U2AF2-centered RNA/protein interactome. Molecules that retained interaction with the U2AF2 complex after RNAse treatment were designated as central interactome members (CIMs). Mass spectrometry also identified a second class of activation-induced proteins, peripheral interactome members (PIMs), that bound to the same transcripts but were not in physical association with U2AF2 or its partners. siRNA knockdown of two CIMs and two PIMs caused changes in activation marker expression, cytokine secretion, and gene expression that were unique to each protein and mapped to pathways associated with key aspects of T cell activation. While knocking down the PIM, SYNCRIP, impacts a limited but immunologically important set of U2AF2-bound transcripts, knockdown of U2AF1 significantly impairs assembly of the majority of protein and mRNA components in the activation-induced interactome. These results demonstrated that CIMs and PIMs, either directly or indirectly through RNA, assembled into activation-induced U2AF2 complexes and play roles in post-transcriptional regulation of genes related to cytokine secretion. These data suggest an additional layer of regulation mediated by the activation-induced assembly of RNA splicing interactomes that is important for understanding T cell activation.
The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells.
Specimen part
View SamplesActivation of CD4 T cells is a reaction to challenges such as microbial pathogens, cancer and toxins that defines adaptive immune responses. The roles of T cell receptor crosslinking, intracellular signaling, and transcription factor activation are well described, but the importance of post-transcriptional regulation by RNA-binding proteins (RBPs) has not been considered in depth. We describe a new model expanding and activating primary human CD4 T cells and applied this to characterizing activation-induced assembly of splicing factors centered on U2AF2. We immunoprecipitated U2AF2 to identify what mRNA transcripts were bound as a function of activation by TCR crosslinking and costimulation. In parallel, mass spectrometry revealed the proteins incorporated into the U2AF2-centered RNA/protein interactome. Molecules that retained interaction with the U2AF2 complex after RNAse treatment were designated as central interactome members (CIMs). Mass spectrometry also identified a second class of activation-induced proteins, peripheral interactome members (PIMs), that bound to the same transcripts but were not in physical association with U2AF2 or its partners. siRNA knockdown of two CIMs and two PIMs caused changes in activation marker expression, cytokine secretion, and gene expression that were unique to each protein and mapped to pathways associated with key aspects of T cell activation. While knocking down the PIM, SYNCRIP, impacts a limited but immunologically important set of U2AF2-bound transcripts, knockdown of U2AF1 significantly impairs assembly of the majority of protein and mRNA components in the activation-induced interactome. These results demonstrated that CIMs and PIMs, either directly or indirectly through RNA, assembled into activation-induced U2AF2 complexes and play roles in post-transcriptional regulation of genes related to cytokine secretion. These data suggest an additional layer of regulation mediated by the activation-induced assembly of RNA splicing interactomes that is important for understanding T cell activation.
The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells.
Specimen part
View SamplesNegative-pressure wound therapy (NPWT) is widely used to improve skin wound healing and to accelerate wound bed preparation. Although NPWT has been extensively studied as a treatment for deep wounds, its effect on epithelialization of superficial dermal wounds remains unclear. To clarify the effect of NPWT on reepithelialization, we applied NPWT on split- thickness skin graft donor sites from the first postoperative day (POD) to the seventh POD. Six patients took part in the study and two samples were obtained from each. The first biopsy sample was taken at elective surgery before split-thickness skin grafting and the second one during reepithelialization on the seventh POD. In all 12 samples (eight from four NPWT patients, and four from two control patients) were collected for this study. From each sample, we carried out a comprehensive genome-wide microarray analysis. Data from patients receiving NPWT were compared groupwise with data from those not receiving NPWT.
Gene expression profiling of negative-pressure-treated skin graft donor site wounds.
Treatment, Subject
View Samples