Mouse embryonic stem cells can differentiate in vitro into spontaneously contracting cardiomyocytes. The main objective of this study was to investigate cardiogenesis in cultures of differentiating embryonic stem cells (ESCs) and to determine how closely it mimics in vivo cardiac development. We identified and isolated a population of cardiac progenitor cells (CPCs) through the use of a reporter DNA construct that allowed the expression of a selectable marker under the control of the Nkx2.5 enhancer. We proceeded to characterize these CPCs by examining their capacity to differentiate into cardiomyocytes and to proliferate. We then performed a large-scale temporal microarray expression analysis in order to identify genes that are uniquely upregulated or downregulated in the CPC population. We determined that the transcriptional profile of the mESC derived CPCs was consistent with pathways known to be active during embryonic cardiac development. We conclude that in vitro differentiation of mESCs recapitulates the early steps of mouse cardiac development.
Mouse ES cell-derived cardiac precursor cells are multipotent and facilitate identification of novel cardiac genes.
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View SamplesThe mechanism of CD4(+) T cell depletion during chronic human immunodeficiency virus type 1 (HIV-1) infection remains unknown. Many studies suggest a significant role for chronic CD4(+) T cell activation. We assumed that the pathogenic process of excessive CD4(+) T cell activation would be reflected in the transcriptional profiles of activated CD4(+) T cells. Here we demonstrate that the transcriptional programs of in vivo activated CD4(+) T cells from untreated HIV(+) individuals are clearly different from those activated CD4(+) T cells from HIV(-) individuals. We observed a dramatic up-regulation of cell cycle-associated and interferon-stimulated transcripts in activated CD4(+) T cells of untreated HIV(+) individuals. Furthermore, we find an enrichment of proliferative and Type I interferon-responsive transcription factor binding sites in the promoters of genes that are differentially expressed in activated CD4(+) T cells of untreated HIV(+) individuals compared to HIV(-) individuals. We confirm these findings by examination of in vivo activated CD4(+) T cells. Taken together, these results suggest that activated CD4(+) T cells from untreated HIV(+) individuals are in a hyper-proliferative state that is modulated by Type I interferons. From these results, we propose a new model for CD4(+) T cell depletion during chronic HIV-1 infection.
Chronic CD4+ T-cell activation and depletion in human immunodeficiency virus type 1 infection: type I interferon-mediated disruption of T-cell dynamics.
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View SamplesTo define the molecular abnormalities at the stem cell level in polycythemia vera (PV), we examined global gene expression in circulating CD34+ cells from 19 JAK2 V617F-positive PV patients and 6 normal individuals using Affymetrix oligonucleotide microarray technology. We observed that CD34+ cell gene expression not only differed between the PV patients and the normal controls but also between men and women PV patients. Based on these gender-specific differences in gene expression, we were able to identify 102 genes differentially regulated concordantly by both men and women, which likely represent a core set of genes whose dysregulation is involved in the pathogenesis of PV. Gene expression was verified by Q-PCR of patient CD34+ cell RNA. Using the 102 gene set and unsupervised hierarchical clustering, the 19 PV patients could be separated in two groups that differed significantly with respect to hemoglobin level, thrombosis frequency, splenomegaly, splenectomy or chemotherapy exposure, leukemic transformation and overall survival. These results were confirmed using top scoring pairs, which identified a different set of 29 genes that independently segregated the 19 patients into the same two clinical groups: those with an aggressive form of the disease (7 patients), and those with an indolent form (12 patients).
Two clinical phenotypes in polycythemia vera.
Sex, Disease
View SamplesA model system of Potyvirus turnip mosaic virus and Arabidopsis was used in this experiment. GFP-tagged virus supplied a visualized marker for us to localize the viral infection foci and its expansion on leaf under UV light. Initially, we dissect an individual infection focus and its adjacent region into four parts and define those four parts as zone 0, 1, 2, and 3, which represented different viral infection stages respectively. Corresponding fours parts were also dissected from control plant treated with turnip leaf sap only. This process was replicated three times totally.
Spatial analysis of arabidopsis thaliana gene expression in response to Turnip mosaic virus infection.
Age, Specimen part
View SamplesEctopic Myc Expression in P493-6 B-cells at three levels:
Induction of ectopic Myc target gene JAG2 augments hypoxic growth and tumorigenesis in a human B-cell model.
Specimen part, Cell line
View SamplesReactivation of latent HCMV is a significant infectious complication of organ transplantation, and current therapies target viral replication once reactivation of transcriptionally silent, latent virus has already occurred. The specific molecular pathways that activate viral gene expression are not well understood. Our studies aim to identify these factors, with the goal of developing novel therapies that prevent transcriptional reactivation in transplant recipients. MCMV is a valuable model for studying latency and reactivation of CMV induced by organ transplantation. We previously demonstrated that transplantation of MCMV-latently infected kidneys into allogeneic recipients induces transcriptional reactivation of immediate early (IE) gene expression within 48 hr.
Transplant-induced reactivation of murine cytomegalovirus immediate early gene expression is associated with recruitment of NF-κB and AP-1 to the major immediate early promoter.
Age, Specimen part
View SamplesA new site of methylation was identified on histone H3 K42 in Saccharomyces cerevisiae. Mutations were engineered at this site to mimic either a constitutively modified state, K42L, or a constitutively unmodified state, K42Q in addition to an alanine substitution. K42A. The effects of these mutations on global transcription was monitored in yeast cells whose sole source of histone H3 was from a plasmid expressing these mutant proteins, and compared to that of an isogenic strain expressing the wild-type histone H3 protein from the same plasmid.
An evolutionarily 'young' lysine residue in histone H3 attenuates transcriptional output in Saccharomyces cerevisiae.
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View SamplesThe emergence of immune resistance variants during immunotherapy is poorly understood. We generated a highly immune resistant cell line (P3) from a susceptible cell line (P0) by subjecting it to 3 rounds of in vivo immune selection. Subsequently, microarray analysis of P0 and P3 was performed to identify genes that may contribute to the increase in immune resistance.
Ectopic expression of vascular cell adhesion molecule-1 as a new mechanism for tumor immune evasion.
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View SamplesIn this experiment, we analyzed the gene expression profile of WT and TBK1-deficient splenic DCs by RNA sequencing based on three independent samples. The results revealed significant alterations in the expression of a number of genes, most notably the enhanced expression of a large subset of IFN-responsive genes in the TBK1-deficient DCs. Overall design: Fresh splenic DC mRNA profiles of 8-week old wild type (WT) and TBK1-DKO mice were generated by deep sequencing, in triplicate
The kinase TBK1 functions in dendritic cells to regulate T cell homeostasis, autoimmunity, and antitumor immunity.
Specimen part, Cell line, Subject
View SamplesWe identified a rare subset of autoreactive lymphocytes with a hybrid phenotype of T and B cells including coexpression of TCR and BCR and key lineage markers of both cell types (hereafter referred to as dual expressers or DEs). To investigate the dual phenotype of DEs at single cell resolution, we examined their transcriptomes using single cell RNA sequencing (scRNA-seq). We sorted individual DEs, Bcon and Tcon cells from PBMCs of one type I diabetes patient and analyzed the transcriptomes of 34 DEs, 20 Bcon , and 23 Tcon using the plate-based SMART-seq2 protocol (Tirosh and Suva, 2018; Tirosh et al., 2016). Our results show that DEs have uniquely expressed genes along with genes encoding lineage markers of T and B cells. Overall design: Examination of the transcriptomes of three cell types, Des (Dual Expressors), Bcon (Conventional B) and Tcon (Conventional T) cells from the PBMCs of one type I diabetes patient
A Public BCR Present in a Unique Dual-Receptor-Expressing Lymphocyte from Type 1 Diabetes Patients Encodes a Potent T Cell Autoantigen.
Specimen part, Disease, Subject
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