A Novel Population of Human Cardiac Resident Mesenchymal Stem Cells
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ABSTRACT: We describe a novel population of human adult cardiac resident stem cells (CRSCs), which are positive for W8B2 antigen, originating from human adult atrial appendages. W8B2+ CRSCs exhibit a spindle-shaped morphology, are clonogenic and able to self-renew. W8B2+ CRSCs show high expression of mesenchymal but not hematopoietic nor endothelial markers. W8B2+ CRSCs expressed GATA4, HAND2, and TBX5, but not C-KIT, SCA-1, NKX2.5, PDGFR?, ISL1 or Wilm’s tumor gene-1 (WT1). W8B2+ CRSCs can differentiate into the cardiovascular lineages and secrete various cytokines. Comparative RNA sequencing was performed using W8B2+ cell from human atrial appendages (passage 2 from 3 different patients), c-Kit+ cell from human atrial appendages (passage 2 from 3 different patients) and W8B2+ cell from bone marrow (passage 3 from 2 different patients, from PromoCell, Heidelberg, Germany).
Project description:Next Generation RNA Sequencing was carried out on human paired left and right atrial appendages from patients with and without Atrial Fibrillation. EdgeR software was used to show a total of 247 genes were found to have significant differential expression between left and right atria.
Project description:Malignant mesothelioma (MM) is an asbestos-related malignancy. Discrimination between MM and reactive mesothelial hyperplasia (RM) is often difficult. MM cells have a broad histological spectrum, and consist mainly of epithelioid, sarcomatoid, and biphasic cell types. The prognosis of MM is generally poor, but better prognosis has been reported with the epithelioid type of MM than the non-epithelioid type. We applied a genome-wide analysis to the identification of new markers that may aid in differentiating the epithelioid type of MM from other histological types and from RM cells. Array-based comparative genomic hybridization analysis was performed on malignant mesothelioma (MM) primary cell cultures, reactive mesothelial hyperplasia (RM) primary cell cultures; early passage of in vitro primary cell cultures to minimize acquisition of additional genomic changes. If available, matched peripheral blood was applied to analysis.
Project description:The DKAT cell line is a novel model of triple-negative breast cancer that was isolated from the pleural effusion of a 35 year-old caucasian woman with triple-negative breast cancer. When cultured in serum-free media (MEGM, Lonza) this cell line exhibits an epithelial morphology and gene expression pattern. However, when cultured in the presence of serum (SCGM, Lonza) it undergoes a reversible EMT. We used microarrays to look at gene expression changes in the DKAT cell line when cultured in Mammary Epithelial Growth Media (MEGM, where DKAT cells have an epithelial morphology) vs Stromal Cell Growth Media (SCGM, where DKAT cells have a mesenchymal morphology). DKAT breast cancer cells (passage 10) grown in MEGM (Lonza) were split and cultured in T75 flasks in either in MEGM or SCGM for 14 days. RNA was isolated using Qiagen RNeasy kit and hybridization on Affymetrix microarrays was performed. Three separate cDNA reactions were performed and these were run as technical replicates.
Project description:We compare the transcriptome of embryonic stem cells (ESCs), adult stem cells with apparent greater differentiation potential such as multipotent adult progenitor cells (MAPCs), mesenchymal stem cells (MSCs) and neurospheres (NS). Mouse and rat MAPCs were used in this study and two different array platforms (Affymetrix and NIA) were used for mouse samples. Experiment Overall Design: Three mRNA samples (biological replicates) per cell type taken at different passage number were compared. Cell types include mouse ESCs, mouse MSCs and three clones isolated using mouse MAPC culture condition: mMAPC-1, mMAPC-2 and mClone-3. mMAPC-1 and mClone-3 were obtained from same bone marrow isolation, mMAPC-2 was obtained in a different bone marrow isolation. Two clones derived from same rat bone marrow using rat MAPC culture conditions were compared. Three mRNA samples (biological replicates) per clone were taken at different passage numbers
Project description:Transcription profiling of human mesenchymal cells (MSCs) after ex vivo expansion under culture conditions supporting extensive cell proliferation. Highly efficient cell expansion within short time (~40 days) and comparison of gene expression profiles from MSCs after primary seeding, defined as early passage versus MSCs after a minimum of 17 (max 34) population doublings, defined as late passage. Combined data of passage 1 and 2 was compared to passage 0 (zero) as reference
Project description:Mesenchymal stromal cells (MSC) were isolated from human bone marrow. Here, we have compared gene expression profiles of MSC at early and late passages. Long-term culture associated gene expression changes were then correlated with DNA-methylation profiles. The goal of this study was to determine if senescence-associated DNA-methylation (SA-DNAm) changes are reflected by differential gene expression. Overall, genes with SA-DNAm changes (particularly with SA-hypomethylation) were detected at low level and seemed to be scarcely expressed at early and late passages. MSC were isolated from three different donors and culture expanded until replicative senescence. Gene expression profiles were compared at early and late passage using GeneChip Humang Gene 1.0 ST arrays (Affymetrix). Six hybridizations are included in this series.
Project description:Pharmacological and gene ablation studies have demonstrated a crucial role of the cardiac natriuretic peptides (NP) hormones ANF and BNP in the maintenance of cardiovascular homeostasis. In addition, hypertension and chronic congestive heart failure are clinical entities that may be regarded as states of relative NP deficiency. Hence the study of the function of the endocrine heart is highly relevant. To identify genes that are related to the endocrine function of the heart we have conducted differential gene expression studies of the rat atria and ventricles using oligonucleotide arrays. Experiment Overall Design: The atrial appendages and the ventricular free walls were obtained from thirteen normal male Sprague Dawley rats. The total RNA was obtained from four pools of atrial and four pools of ventricular tissues. Four biological replicates for each muscle type were generated, i.e. 4 atrial replicates and 4 ventricular replicates.
Project description:Wounds, especially non healing wounds are characterised by elevated tissue lactate concentrations. Lactate is known for being able to stimulate collagen synthesis and vessel growth. Lately it has been shown that lactate, in vivo, plays an important role in homing of stem cells. With this work we aimed to show the influence of lactate on the gene expressionprofil of human mesenchymal stem cells (hMSC). hMSCs were obtained from bone marrow and characterised with fluorescence-activated cell sorting (FACS) analysis. Subsequently the hMSCs were treated with either 0, 5, 10 and 15 mM lactate (pH 7,4) for 24 hours. RNA Isolation from stimulated hMSCs and controls was performed. The Microarray analysis was performed using Affymetrix HuGene 1.0 ST Gene Chip. Selected targets were subsequently analysed using quantitative real time PCR (RTq-PCR). We were able to show that lactate in moderate concentrations of 5 respectively 10 mM leads to an anti-imflammatory, anti-apoptotic but growth and proliferation promoting gene expression after 24 h. In contrast, high latate concentrations of 15 mM leads to the opposed effect, namely promoting inflammation and apoptosis. Hypoxia induced genes did not not show any significant regulation. Contrary to expectation, we were not able to show any significant regulation of glycolysis associate candiadtes. We were able to show that lactate alters gene expression but does not change the cell phenotype, which might be helpful for further investigations of new treatment strategies for chronic non-healing wounds as well as tumor-therapy and neuronal plasticity. Timecourse experiment with hMSCs treated with 5, 10, 15 mM lactic acid for 1, 3, 7 days. hMSCs without lactate served as controls *** CEL files lost due to hard disk crash
Project description:It is now well established that bone marrow (BM) constitutes a microenvironment required for differentiation. Bone marrow mesenchymal stromal cells (BM-MSCs) strongly support MM cell growth, by producing a high level of Interleukin-6 (IL-6), a major MM cell growth factor. BM-MSCs also support osteoclastogenesis and angiogenesis. Previous studies have suggested that the direct (VLA-4, VCAM-1, CD44, VLA-5, LFA-1, syndecan-1,M-bM-^@M-&) and indirect interactions (soluble factors) between MM plasma cells and BM-MSCs result in constitutive abnormalities in BM-MSCs. In particular, MM BM-MSCs express less CD106 and fibronectin and more DKK1, IL-1M-NM-2 and TNF-M-NM-1 as compared with normal BM-MSCs. In order to gain a global view of the differences between BM-MSCs from MM patients and healthy donors, we used gene expression profiling to identify genes associated to the transformation of MM BM-MSCs. BM-MSCs were isolated from 3 healthy donors and 4 untreated multiple myeloma patients. Total RNA from BM-MSCs was exctracted and hybridyzed on Affymetrix GeneChipM-BM-. Human Genome U133 Plus 2.0 Array. Amplification, hybridization and scanning were done according to standard Affymetrix protocols (www.affymetrix.com). CEL files were normalized with RMA method.