Epigenome-wide DNA methylation profiling in cord blood CD4+ T cells from children treated prenatally with Lactobacillus reuteri and/or omega-3 fatty acids
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ABSTRACT: In this study, we have examined the effect of combined prenatal Lactobacillus reuteri and omega-3 fatty acid supplementation on DNA methylation patterns in neonatal immune cells from children with a high propensity of developing allergies. We addressed this matter by assessing epigenome-wide DNA methylation patterns in CD4+ T helper cells obtained at birth, using the Illumina MethylationEPIC 850K array. Samples were retrieved from all four arms of the on-going PRObiotics and OMega-3 (PROOM-3, ClinicalTrials.gov-ID: NCT01542970) allergy prevention trial, and treatment-induced differential methylation as well as network modules were assessed.
Project description:The purpose of this experiment was to determine the transcriptional differences between neural progenitor cells from neonatal lung injury mice vs. control mice, as well as neonatal lung injury mice treated with umbilical-cord mesenchymal stromal cell extracellular vesicles vs. neonatal lung injury mice treated with placebo (PBS). Neural progenitor cells were isolated from the subventricular zone and hippocampus and cultured for 2 consecutive neurosphere assays. RNA was then extracted from the cells, and the microarray labelling, hybridization, and scanning was conducted by the Génome Québec Innovation Centre (Montréal, Canada).
Project description:We are studying the role of PAI-1 in hypergastrinemic transgenic mice (PAI-1 knockout, HK-ATPase beta knockout and corresponding control strains). As a model of hypergastrinaemia, HK-ATPase beta knockout mice are preferred instead of giving a high dose of proton pump inhibitors daily. We have successfully crossed these two strains and generated double knockout mice (PAI-1--/HK-ATPase--) and 3 different control groups (PAI-1++/HK-ATPase++, PAI-1--/HK-ATPase++ and PAI-1++/HK-ATPase--) (9 animals in each group). The gastric phenotype of all mice has been examined by morphologic and molecular analysis after 12 months. Gastric biopsies and blood samples have been taken for histology, large scale gene expression analysis, verification of mRNA and protein expression and gastrin measurements. Global gene expression analysis has been done on gastric mucosal samples from all groups of mice. The differences between wild type mice and the transgenic animals are expected to provide novel hypotheses on the role of PAI-1 in gastrin-induced changes in the gastric mucosa.
Project description:Fibrosis is defined as an abnormal matrix remodeling and loss of tissue homeostasis due to excessive synthesis and accumulation of extracellular matrix proteins in tissues. At present, there is no effective therapy for organ fibrosis. Previous studies demonstrated that aged plasminogen activator inhibitor-1(PAI-1) knockout mice develop spontaneously cardiac-selective fibrosis without affecting any other organs including kidney. Therefore, the PAI-1 knockout model of cardiac fibrosis provides an excellent opportunity to find the igniter(s) of cardiac fibrosis and its status in unaffected organs. We hypothesized that differential expressions of profibrotic and antifibrotic genes in PAI-1 knockout hearts and unaffected organs lead to cardiac selective fibrosis. In order to address this prediction, we have used a genome-wide gene expression profiling of transcripts derived from aged PAI-1 knockout hearts and kidneys. The variations of global gene expression profiling were compared within four groups: wildtype heart vs. knockout heart; wildtype kidney vs. knockout kidney; knockout heart vs. knockout kidney and wildtype heart vs. wildtype kidney. Analysis of illumina-based microarray data revealed that several genes involved in different biological processes such as immune system processing, response to stress, cytokine signaling, cell proliferation, adhesion, migration, matrix organization and transcriptional regulation were affected in hearts and kidneys by the absence of PAI-1, a potent inhibitor of urokinase- and tissue-type plasminogen activator. Importantly, the expressions of a number of genes, involved in profibrotic pathways were upregulated or downregulated in PAI-1 knockout hearts compared to wildtype hearts and PAI-1 knockout kidneys. To our knowledge, this is the first comprehensive report on the influence of PAI-1 on global gene expression profiling in the heart and kidney and its implication in several biological processes including fibrogenesis. Total RNA was extracted from hearts and kidneys derived from three PAI-1 knockout (12- month old) and three wild-type mice (12-month old) using RNeasy Fibrous Tissue Mini Kit (Qiagen, Valencia, CA) following the manufacturer’s instructions. The quality of RNA (RNA Integrity, RIN) in all 12 samples (3 wildtype hearts; 3 PAI-1 KO hearts; 3 wildtype kidneys; and 3 PAI-1 KO kidneys) was checked using the bioanalyzer. We have used a genome-wide gene expression profiling of transcripts derived from aged PAI-1 knockout hearts and kidneys. The variations of global gene expression profiling were compared within four groups: wildtype heart vs. knockout heart; wildtype kidney vs. knockout kidney; knockout heart vs. knockout kidney and wildtype heart vs. wildtype kidney.
Project description:Pancreatic neuroendocrine tumors (PanNETs) account for 1-3% of all neoplasms of the pancreas. Epigenetic changes seem to be the main drivers of PanNET tumorigenesis. Indeed, the most frequent mutated genes in PanNETs are DAXX (Death domain associated protein 6), ATRX (Transcriptional Regulator ATRX) and MEN1 (menin 1), which are all involved in epigenetic regulation. The objective of the project is to study the epigenetic changes undergoing in PanNETs. Specifically, we aimed at investigating whether do exist biologically important epigenetic differences in PanNET, useful for a clinically relevant classification.
Project description:Pancreatic neuroendocrine tumors (PanNETs) account for 1-3% of all neoplasms of the pancreas. Epigenetic changes seem to be the main drivers of PanNET tumorigenesis. Indeed, the most frequent mutated genes in PanNETs are DAXX (Death domain associated protein 6), ATRX (Transcriptional Regulator ATRX) and MEN1 (menin 1), which are all involved in epigenetic regulation. The objective of the project is to study the epigenetic changes undergoing in PanNETs. Specifically, we aimed at investigating whether do exist biologically important epigenetic differences in PanNET, useful for a clinically relevant classification.
Project description:Pancreatic neuroendocrine tumors (PanNETs) account for 1-3% of all neoplasms of the pancreas. Epigenetic changes seem to be the main drivers of PanNET tumorigenesis. Indeed, the most frequent mutated genes in PanNETs are DAXX (Death domain associated protein 6), ATRX (Transcriptional Regulator ATRX) and MEN1 (menin 1), which are all involved in epigenetic regulation. The objective of the project is to study the epigenetic changes undergoing in PanNETs. Specifically, we aimed at investigating whether do exist biologically important epigenetic differences in PanNET, useful for a clinically relevant classification.
Project description:Cord blood NK cells were cultured with or without IL-15 overnight and then RNA was isolated using an RNeasy Mini kit (Qiagen). Samples were processed using GeneChip Whole Transcript Sense Target Labeling assays using the Ambion WT Expression kit and Affymetrix GeneChip WT Terminal Labeling and Controls kit (Affymetrix). The resulting ssDNAs were hybridized to the Human Gene 2.0 ST Array (Affymetrix) and transcript levels compared between IL-15 treated and untreated using GeneSpring software.
Project description:Cord blood stem cells (CBSC) and resting or activated NK cells were co-cultured for 4 h and then CBSC were re-isolated by sorting using a FACSAria cell sorter (BD Biosciences). RNA was isolated from sorted CBSC using an RNeasy Micro kit (Qiagen). Samples were processed using GeneChip Whole Transcript Sense Target Labeling assays using the Ambion WT Expression kit and Affymetrix GeneChip WT Terminal Labeling and Controls kit (Affymetrix). The resulting ssDNAs were hybridized to the Human Gene 2.0 ST Array (Affymetrix) and transcript levels compared to CBSC alone using GeneSpring software.
Project description:Shatavari is a herbal dietary supplement that may increase skeletal muscle strength in younger and older adults. Shatavari contains compounds with both estradiol-like and antioxidant properties, which could enhance muscle function. Postmenopausal women may derive the greatest benefit, as estrogen deficiency adversely impacts skeletal muscle function. However, mechanistic insights are limited and the effects of shatavari on muscle function require further characterization. In this randomised, double-blind trial, 17 young (23 ±5yr) and 22 older (63±5yr) women completed an eight-week leg resistance training programme. They consumed either a placebo or shatavari (1000mg/d, equivalent to 26,500 mg/d fresh weight) supplement throughout. Pre and post training, measures of leg strength, neuromuscular function and vastus lateralis (VL) biopsies were obtained. Tandem-mass-tagged VL proteomic analyses were performed. Additionally, resistance training (RT) is the gold standard intervention for ameliorating sarcopenia. Outstanding mechanistic questions remain regarding the malleability of the molecular determinants of skeletal muscle function in older age. Discovery proteomics can expand such knowledge. We further aimed to compare the effect of RT on the skeletal muscle proteome and neuromuscular function (NMF) in older and younger women.
Project description:Background and Aims Small intestinal neuroendocrine tumours (SINETs) are the commonest malignancy of the small intestine; however underlying pathogenic mechanisms remain poorly characterised. Whole genome and exome sequencing has demonstrated that SINETs are mutationally quiet with the most frequent known mutation in the cyclin dependent kinase inhibitor 1B gene (CDKN1B) occurring in only ~8% of tumours, suggesting that alternative mechanisms may drive tumourigenesis. The aim of this study is to perform genome-wide molecular profiling of SINETs in order to identify pathogenic drivers based on molecular profiling. This study represents the largest unbiased integrated genomic, epigenomic, and transcriptomic analysis undertaken in this tumour type. Methods Here we present data from integrated molecular analysis of SINETs (n=97) including whole exome or targeted CDKN1B sequencing (n=29), HumanMethylation450 BeadChip (Illumina) array profiling (n=69), methylated DNA immunoprecipitation sequencing (n=16), copy number variance analysis (n=47) and Whole Genome-DASL (Illumina) expression array profiling (n=43). Results Based on molecular profiling SINETs can be classified in to three groups which demonstrate significantly different progression-free survival after resection of primary tumour (not reached at 10 years vs 56 months vs 21 months, p=0.04). Epimutations were found at a recurrence rate of up to 85% and 21 epigenetically dysregulated genes were identified, including CDX1 (86%), CELSR3 (84%), FBP1 (84%) and GIPR (74%). Conclusions This is the first comprehensive integrated molecular analysis of SINETs. We have demonstrated that these tumours are highly epigenetically dysregulated. Furthermore, we have identified novel molecular subtypes with significant impact on progression free survival. Background and Aims Small intestinal neuroendocrine tumours (SINETs) are the commonest malignancy of the small intestine; however underlying pathogenic mechanisms remain poorly characterised. Whole genome and exome sequencing has demonstrated that SINETs are mutationally quiet with the most frequent known mutation in the cyclin dependent kinase inhibitor 1B gene (CDKN1B) occurring in only ~8% of tumours, suggesting that alternative mechanisms may drive tumourigenesis. The aim of this study is to perform genome-wide molecular profiling of SINETs in order to identify pathogenic drivers based on molecular profiling. This study represents the largest unbiased integrated genomic, epigenomic, and transcriptomic analysis undertaken in this tumour type. Methods Here we present data from integrated molecular analysis of SINETs (n=97) including whole exome or targeted CDKN1B sequencing (n=29), HumanMethylation450 BeadChip (Illumina) array profiling (n=69), methylated DNA immunoprecipitation sequencing (n=16), copy number variance analysis (n=47) and Whole Genome-DASL (Illumina) expression array profiling (n=43). Results Based on molecular profiling SINETs can be classified in to three groups which demonstrate significantly different progression-free survival after resection of primary tumour (not reached at 10 years vs 56 months vs 21 months, p=0.04). Epimutations were found at a recurrence rate of up to 85% and 21 epigenetically dysregulated genes were identified, including CDX1 (86%), CELSR3 (84%), FBP1 (84%) and GIPR (74%). Conclusions This is the first comprehensive integrated molecular analysis of SINETs. We have demonstrated that these tumours are highly epigenetically dysregulated. Furthermore, we have identified novel molecular subtypes with significant impact on progression free survival. This study included 97 tumour samples from 85 individuals, this included both primary and metastatic tumour samples. 25 normal small intestinal samples were analysed.