Project description:We established a novel mouse model for postnatal erythropoietin (Epo)-deficiency anaemia, designated ISAM (inherited super anemic mouse), using a transgenic complementation rescue technique. To identify Epo-regulated genes in vivo, we examined the mRNA expression profile in the bone marrow of ISAM 6 hours after recombinant human EPO (rHuEPO) administration. Erythropoietin-induced gene expression in mouse bone marrow was measured at 6 hours after rHuEPO administration (3,000 U/kg). Three Epo-treated samples were analyzed, and two PBS-treated and one untreated samples were used as a control group.
Project description:This study was to develop a faithful pre-clinical MDS patient-derived xenotransplantation model in cytokine-humanized immunodeficient "MISTRG" mice. We performed targeted genome sequencing on bone marrow from patients with MDS or AML and from bone marrow of mice engrafted with the respective patient bone marrow. Cytokine-humanized immunodeficient mice faithfully replicate human MDS.
Project description:Diamond-Blackfan anemia (DBA) is characterized by anemia and cancer susceptibility, and is caused by mutations in ribosomal genes, including Rpl11. Here, we report that Rpl11-heterozygous embryos are not viable, and homozygous deletion of Rpl11 in adult mice results in death within a few weeks, accompanied by bone marrow aplasia and intestinal atrophy. Importantly, deletion of a single Rpl11 allele in adult mice results in anemia associated to decreased erythroid progenitors and defective erythroid maturation. These phenotypes are also present in mice transplanted with inducible heterozygous Rpl11 bone marrow, indicating a cell-autonomous role of RPL11 in erythropoiesis. Additionally, fibroblasts lacking one or both Rpl11 alleles show defective p53 activation upon ribosomal stress or DNA damage. Furthermore, fibroblasts and hematopoietic tissues from heterozygous Rpl11 mice present higher basal cMYC levels. Accordingly, heterozygous Rpl11 mice are highly susceptible to radiation-induced lymphomagenesis. We conclude that Rpl11-deficient mice recapitulate DBA disorder, including cancer predisposition. RNAseq profiles of bone marrow hematopoietic progenitors cells from WT (Rpl11+/+:: Tg.UbC-CreERT2) and LOX (Rpl11+/lox::Tb.Ub-CreERT2) mice, n=4 independent animals per genotype
Project description:CD70TG mice are a model for sterile chronic immune activation and develop Anemia of Inflammation, which is dependent on the production of Ifng by effector CD4 and CD8 T cells. We used microarrays to identify Ifng-dependent differentially expressed genes that could account for the erythropoietic defect. CD71+ cells were MACS-enriched from the bone marrow of WT, CD70TG, IFNg-/- and CD70TG*IFNg-/- male mice of 10-12 weeks of age (3 mice per genotype group).
Project description:CD70TG mice are a model for sterile chronic immune activation and develop Anemia of Inflammation, which is dependent on the production of Ifng by effector CD4 and CD8 T cells. We used microarrays to identify Ifng-dependent differentially expressed genes that could account for the erythropoietic defect. Overall design: CD71+ cells were MACS-enriched from the bone marrow of WT, CD70TG, IFNg-/- and CD70TG*IFNg-/- male mice of 10-12 weeks of age (3 mice per genotype group).
Project description:Since iron deficiency anemia (IDA) is one of the most common diseases in worldwide, it is an essential issue to prevent and to treat the IDA in public healthcare system. However, the precise adaptive responses and their mechanisms of the hematopoietic system induced by iron deficient state are not fully understood. In this study, low iron diet conditions which induce sever iron deficiency anemia in mice were established. Transcriptome analyses in erythroblasts under normal or iron deficient states were performed to describe the pathological details of IDA. Under iron deficient state, extensive gene expression changes and mitophagy disorder were induced during the terminal maturation of erythroblasts. These findings provide a new insight into pathophysiology and molecular biology of IDA and the function of iron as a coordinator of gene expression networks in erythrocyte maturation. Overall design: Subset II erythroblasts sorted from bone marrow of WT under normal diet or low iron diet after weaning for three weeks or seven weeks. Three biological replication was performed.
Project description:Somatic mutation in the X-linked phosphatidylinositol glycan class A (PIG-A) gene causes glycosylphosphatidylinositol (GPI) anchor deficiency in humans with Paroxysmal Nocturnal Hemoglobinuria (PNH). Clinically, patients with PNH have intravascular hemolysis, venous thrombosis and bone marrow failure. We produced a conditional Pig-a knock-out mouse model specifically inactivating the Pig-a gene in hematopoietic cells to study the role of PIG-A deficiency in PNH pathophysiology. We used Affymetrix Mouse Genome 430 2.0 chips to investigate the gene expression pattern in the mouse model of targeted Pig-a deletion. Keywords: Gene expression comparison between untransplanted and transplanted GPI-deficient bone marrow cells Overall design: We performed microarray analysis on 3 pools of sorted GPI-deficient (GPI-) and GPI normal (GPI+) bone marrow cells derived from the same Pig-a knock-out animals, and identified 1275/669 genes of the 45,101 transcripts potentially available for screening using 2-fold change, <10% false discovery rate (FDR) and percentage of present calls as selection criteria. The major representative genes belong to the category of immune response. We tested whether the molecular differences between GPI- and GPI+ bone marrow cells could be preserved when GPI- cells were transplanted in lethally-irradiated wild type mice (C57BL/6). Microarray analysis was performed using sorted bone marrow cells from 4 animals transplanted with GPI-deficient bone marrow cells (T-GPI-) from Pig-a knock-out donors, 3 animals transplanted with GPI-normal bone marrow cells from C57BL/6 donors (T-GPI+), and GPI-normal bone marrow cells from 4 wild-type C57BL/6 mice (WT-GPI+). We found 296 probesets with 2-fold change cutoff, of which T-GPI- cells had 145 up-regulated genes in comparison to WT-GPI+ cells, and had 123 genes differentially expressed when compared to T-GPI+ cells. The gene expression of the GPI-deficient cells was very similar between the two sets of microarray experiments affirming the maintenance of the phenotype before and after bone marrow cell transplantation.
Project description:RPS19 mutations are the most common cause of the human disorder Diamond Blackfan Anemia. The R62W mutation was hypothesized to act in a dominant negative fashion and mice expressing RPS19R62W have many of the characteristics of Diamond Blackfan Anemia. Diamond-Blackfan Anemia (DBA), is an inherited erythroblastopenia associated with mutations in at least 8 different ribosomal protein genes. Mutations in the gene encoding Ribosomal Protein S19 (RPS19) have been identified in ~25% of DBA families. Most of these mutations disrupt either the translation or stability of the RPS19 protein and are predicted to cause DBA by haploinsufficiency. However, approximately ~30% of RPS19 mutations are missense mutations that do not alter the stability of the RPS19 protein and are hypothesized to act by a dominant negative mechanism. To formally test this hypothesis, we generated a transgenic mouse model expressing an RPS19 mutation in which an Arginine residue is replaced with a Tryptophan residue at codon 62 (RPS19R62W). Constitutive expression of RPS19R62W in developing mice was lethal. Conditional expression of RPS19R62W resulted in growth retardation, a mild anemia with reduced numbers of erythroid progenitors and significant inhibition of terminal erythroid maturation, similar to DBA. RNA profiling demonstrated over 700 dysregulated genes belonging to the same pathways that are disrupted in RNA profiles of DBA patient cells. Overall design: The samples compared are RNA extracted from CD71 positive erythroblasts sorted from the bone marrow of wild type mice and mice expressing a mutant RPS19 (RPS19R62W) transgene
Project description:Transcriptome profile of highly purified multipotential (P), erythroid (E), and myeloid (M) bone marrow progenitors from three RPS19 mutated Diamond-Blackfan anemia and six control human subjects. Two group comparison of sex and age matched subjects. Bone marrow progenitors, gene expression profiling, Diamond-Blackfan anemia, RPS19