Increasing fetal hemoglobin (HbF) levels in adult red blood cells provides clinical benefit to patients with sickle cell disease and some forms of beta-thalassemia. To identify potentially druggable HbF regulators in adult human erythroid cells, we employed a protein kinase-domain focused CRISPR/Cas9-based genetic screen with a newly optimized sgRNA scaffold. The screen uncovered the heme-regulated inhibitor HRI (also known as EIF2AK1), an erythroid-specific kinase that controls protein translation, as an HbF repressor. HRI depletion markedly increased HbF production in a specific manner and reduced sickling in cultured erythroid cells. Diminished expression of the HbF repressor BCL11A accounted in large part for the effects of HRI depletion. Taken together, these results suggest HRI as a potential therapeutic target for hemoglobinopathies. Overall design: A CRISPR-screen reveals HRI kinase as a fetal hemoglobin repressor and further validated in HUDEP2 and CD34+ derived primary erythroid cultures.
Domain-focused CRISPR screen identifies HRI as a fetal hemoglobin regulator in human erythroid cells.
Specimen part, Disease, Subject
View SamplesFour conditions of cultured CD8+ T cells were analyzed with Affymetrix HG-U133-Plus-2.0 microarrays.
Detection of self-reactive CD8⁺ T cells with an anergic phenotype in healthy individuals.
Specimen part
View SamplesGene expression study of human and Chimpanzee iPS cell.
New type of Sendai virus vector provides transgene-free iPS cells derived from chimpanzee blood.
Specimen part
View SamplesMice with MUTYH-null allele (Mutyh+/-, Mutyh-/-) were fed a high-fat/high-cholesterol (HFHC) diet or HFHC + high iron diet. The incidence of liver tumors and histological features of the liver were compared.
MUTYH is associated with hepatocarcinogenesis in a non-alcoholic steatohepatitis mouse model.
Specimen part, Disease, Treatment
View SamplesAnalysis of Foxp3(+)epigenetics(-) T cells, Foxp3(-)epigenetics(+) T cells, and Foxp3(+)epigenetics(+) T cells. Results indicate regulatory T cell (Treg) ontogenesis requires two independent processes, expression of the transcription factor Foxp3 and establishment of Treg epigenetic programs induced by T cell receptor (TCR) stimulation.
T cell receptor stimulation-induced epigenetic changes and Foxp3 expression are independent and complementary events required for Treg cell development.
Specimen part
View SamplesABSTRACT Background: Viral myocarditis is a life-threatening illness that may lead to heart failure or cardiac arrhythmias. This study examined whether human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) could be used to model the pathogenic processes of coxsackievirus-induced viral myocarditis and to screen antiviral therapeutics for efficacy. Methods and Results: Human iPSC-CMs were infected with a luciferase-expressing mutant of the coxsackievirus B3 strain (CVB3-Luc). Brightfield microscopy, immunofluorescence, and calcium imaging were used to characterize virally infected hiPSC-CMs. Viral proliferation on hiPSC-CMs was subsequently quantified using bioluminescence imaging. For drug screening, select antiviral compounds including interferon beta 1 (IFN1), ribavirin, pyrrolidine dithiocarbamate (PDTC), and fluoxetine were tested for their capacity to abrogate CVB3-Luc proliferation in hiPSC-CMs in vitro. The ability of some of these compounds to reduce CVB3-Luc proliferation in hiPSC-CMs was consistent with the reported drug effects in previous studies. Finally, mechanistic analyses via gene expression profiling of hiPSC-CMs infected with CVB3-Luc revealed an activation of viral RNA and protein clearance pathways within these hiPSC-CMs after IFN1 treatment. Conclusions: This study demonstrates that hiPSC-CMs express the coxsackievirus and adenovirus receptor, are susceptible to coxsackievirus infection, and can be used to confirm antiviral drug efficacy. Our results suggest that the hiPSC-CM/CVB3-Luc assay is a sensitive platform that could be used to screen novel antiviral therapeutics for their effectiveness in a high-throughput fashion.
Human induced pluripotent stem cell-derived cardiomyocytes as an in vitro model for coxsackievirus B3-induced myocarditis and antiviral drug screening platform.
Specimen part, Treatment
View SamplesTo evaluate gene expression changes in mixed tissue samples used as process controls in male Sprague Dawley rats over time.
Assessment of repeated microarray experiments using mixed tissue RNA reference samples.
No sample metadata fields
View SamplesThe female germline undergoes a unique line of differentiation processes that endows totipotency to the egg. During these processes, biologically significant events such as meiosis and oocyte growth are controlled in an orderly manner, with any disorder causing infertility and developmental arrest of the next generation. Reconstitution in vitro of the entire process of oogenesis from pluripotent stem cells is a key achievement in stem cell biology and regenerative medicine, but a robust and reproducible culture system has not been established. Here, we report successful reconstitution in vitro throughout the entire process of oogenesis from pluripotent stem cells, yielding in vitro-produced eggs that gave rise to healthy pups. Moreover, the pluripotent stem lines were re-derived from the in vitro-generated eggs, thereby reconstituting one generation on a dish. This culture system will provide a unique platform for elucidating the molecular mechanisms underlying totipotency and could open an avenue to producing vast numbers of eggs in vitro. Overall design: The transcriptomes of ES cells (ESCs), primordial germ cell-like cells at day 6 of differentiation (PGCLCs_d6), PGCLCs aggregated with gonadal somatic cells (PGCLCs_agg3), in vitro-produced primary oocytes in secondary follicles (vitro_2nd_fol_oocyte) and MII oocytes (vitro MII oocytes) are determined by RNA-seq analysis. For comparison, the transcriptomes of E12.5 PGCs (vivo_E12.5_PGCs), P8 primary oocytes (vivo_2nd_fol._oocyte) and MII oocytes (vivo_MII_oocyte) in vivo are also determined. Biologically triplicated (rep1-3) or duplicated (rep1-2) samples are sequnced simultaneously.
Mechanical stress accompanied with nuclear rotation is involved in the dormant state of mouse oocytes.
Specimen part, Cell line, Subject
View SamplesAnalysis of umbilical vein endothelial cells (HUVEC) treated with Egr-3 siRNA under the VEGF treatment for 0,1, and 4 h. Egr-3, a member of early growth response family, is immediately and dramatically induced by VEGF in HUVEC, which regulates expression of many genes related to endothelial activation.
Vascular endothelial growth factor activation of endothelial cells is mediated by early growth response-3.
No sample metadata fields
View SamplesRhizoctonia solani is an economically important soil-borne necrotrophic fungal pathogen, with a broad host range and for which little effective resistance exists in crop plants. Arabidopsis is resistant to the R. solani AG8 isolate but susceptible to R. solani AG2-1. Affymetrix microarray analysis was performed to determine genes that are affected in common and specifically by AG8 and AG2-1.
Genetic and genomic analysis of Rhizoctonia solani interactions with Arabidopsis; evidence of resistance mediated through NADPH oxidases.
Age, Specimen part, Treatment
View Samples