Project description:Genome wide DNA methylation profiling of leukemia stem, blast cells obtained from 15 AML patients and of normal hematopoietic stem/progenitor cells from 5 normal bone marrow. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles across approximately 450,000 CpGs in the samples. Samples included 20 leukemia stem cells, 24 blast cells and 30 normal hematopoietic stem and progenitor cells (6 different types from 5 normal bone marrows).
Project description:This is a mathematical model describing the hematopoietic lineages with leukemia lineages, as controlled by end-product negative feedback inhibition. Variables include hematopoietic stem cells, progenitor cells, terminally differentiated HSCs, leukemia stem cells, and terminally differentiated leukemia stem cells.
Project description:The comparative characterization of hematopoietic stem cells from healthy stem cell donors and patients with acute myeloid leukemia on a proteome level has the potential to reveal differentially regulated proteins which might be candidates for specific immunotherapy target molecules. Exemplarily, we analyzed the proteome of the cytosolic and the membrane fraction of CD34 and CD123 co-expressing FACS-sorted leukemic progenitors from five patients with acute myeloid leukemia employing mass spectrometry. As a reference, CD34+CD123+ normal hematopoietic progenitor cells from five healthy stem cell donors were analyzed. In this TMT 10-plex labeling based approach 2068 proteins were identified with 256 proteins differentially regulated in one or both cellular compartments. This study demonstrates the feasibility of a mass spectrometry based proteomic approach to detect differentially expressed proteins in two compartment fractions of leukemic stem cells as compared to their healthy stem cell counterparts.
Project description:The formation of hematopoietic cells relies on the chromatin remodeling activities of ISWI ATPase SMARCA5 (SNF2H) and its complexes. The Smarca5 null and conditional alleles have been used to study its functions in embryonic and organ development in mice. These mouse model phenotypes vary from embryonic lethality of constitutive knockout to less severe phenotypes observed in tissue-specific Smarca5 deletions, e.g., in the hematopoietic system. Here we show that, in a gene dosage-dependent manner, the hypomorphic allele of SMARCA5 (S5tg) can rescue not only the developmental arrest in hematopoiesis in the hCD2iCre model but also the lethal phenotypes associated with constitutive Smarca5 deletion or Vav1iCre-driven conditional knockout in hematopoietic progenitor cells. Interestingly, the latter model also provided evidence for the role of SMARCA5 expression level in hematopoietic stem cells, as the Vav1iCre S5tg animals accumulate stem and progenitor cells. Furthermore, their hematopoietic stem cells exhibited impaired lymphoid lineage entry and differentiation. This observation contrasts with the myeloid lineage which is developing without significant disturbances. Our findings indicate that animals with low expression of SMARCA5 exhibit normal embryonic development with altered lymphoid entry within the hematopoietic stem cell compartment.
Project description:<p>Methionine cycle plays critical roles in cell fate determination by shaping epigenetic landscape, yet its function in human erythropoiesis remains undefined. Here, we show that disruption of methionine metabolism by compromising key enzyme adenosylhomocysteinase (AHCY) reshapes H3K4me3 landscape, causing erythroid cell fate reprogramming. AHCY deficiency severely impaired erythroid differentiation and expansion, leading to the generation of non-erythroid lineage hematopoietic cells, including stem/progenitor cells and immune cells, as evidenced by single-cell RNA sequencing, Pseudo temporal analysis delineated a precise dedifferentiation trajectory, revealing erythroblasts transitioning back to MEPs and HSCs. Moreover, human hematopoietic system could be reconstituted in the immunodeficient NCG-X mice by transplanting AHCY deficient erythroblasts. Mechanistically, AHCY deficiency reduced global H3K4me3 levels and altered its genomic distribution, resulting in the upregulated expression of non-erythroid transcription factors and downregulated expression of erythrocyte lineage-specific transcription factors. Integrated single-cell analyses identified transitional states with diminished AHCY in the erythroblasts of acute myeloid leukemia (AML) patient. Further flow cytometry confirmed the reduced H3K4me3 level in patient derived erythroid cells. Erythroblast isolated from AML patients with reduced H3K4me3 exhibited dedifferentiation potential into progenitor-like states. Our findings reveal a metabolic-epigenetic axis governing cell fate reprogramming in human erythropoiesis and provide insights into leukemia associated anemia.</p>
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.