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• W2032885228 abstract "RUNX1 is an essential gene for mouse hematopoietic development. Mouse knockout models for RUNX1 or its binding partner, core binding factor β (CBFβ), are embryonic lethal and lack definitive hematopoiesis. CBFβ-SMMHC (INV), encoded by the inv(16) chromosomal translocation in approximately 8% of acute myeloid leukemia (AML) cases, is a fusion protein comprising of CBFβ and the rod domain of smooth muscle myosin heavy chain (SMMHC). INV inhibits the expression of RUNX1-regulated genes, by sequestering RUNX1 and by directly repressing RUNX1- regulated genes. Recently, we reported that dominant inhibition of endogenous RUNX1 by lentivector-expressed INV inhibited proliferation of human and mouse myeloid progenitors. Conditional exogenous expression of RUNX1 using an estrogen receptor lentivector system in mouse primary HSPCs, resulted in increased proliferation [D'Costa, 2005]. Although RUNX1 is established as a key regulator of hematopoietic development in the mouse embryo, its role in human hematopoietic development has not been extensively studied for lack of an appropriate model. The availability of several human embryonic stem cell (HESC) lines provides a potential new system with demonstrated utility for the study of human embryonic hematopoiesis. We were interested to see if exogenous expression of RUNX1 would enhance human embryonic stem cell (HESC)-derived hematopoiesis, and conversely if dominant RUNX1 inhibitors AML1-ETO and INV would inhibit HESC-derived hematopoiesis. We transduced H1 HESC with dual promoter lentivectors expressing 4-hydroxy-tamoxifen (4HT)-regulated RUNX1-ER (EF.RUNX1- ER/PGK-GFP) or AML1-ETO-ER (EF.AML-ETO-ER/Ub-GFP) or control GFP (EF.GFP). Cells from these transduction cultures were FACS-sorted, GFP+ cells expanded by culture on PMEFs for 4 weeks and allowed to undergo embryoid body (EB) formation. EBs (cultured with or without 4HT) were harvested at selected time points, dissociated with trypsin/EDTA, counted, and immunostained for CD34 and CD45 and assayed in hematopoietic colony-forming assays. Induction of RUNX1 or AML-ETO expression in undifferentiated HESC by 4-HT did not cause any changes in proliferation or differentiation of the cells as assessed by staining for SSEA-4, Tra-160 and CD9. However, on EB formation, at the 3 time points tested, there were 2–3-fold greater percentages and numbers of transduced (GFP+) cells in the EF.RUNX1-ER/PGK.GFP lentivector-transduced cell culture induced with 4HT than in the EF.RUNX1-ER/PGK.GFP lentivector-transduced cell culture without 4HT. In contrast, no 4HT-dependent difference was seen in the EF.GFP control lentivector-transduced cell cultures. At these time points, 2–4% of the GFP+ cells in the EF.RUNX1-ER/PGK.GFP lentivector-transduced cell culture containing 4HT expressed CD34 and there were similar numbers of CD45+ cells, whereas there were lower percentages and total numbers of these potential hematopoietic cells in the control cultures. Thus, induction of RUNX1 boosted total cell numbers and the numbers of CD34+ and CD45+ cells in EBs at a time when EBs were previously shown [Zambidis, 2005] to contain developing hematopoietic progenitors. The Johns Hopkins University holds patents on CD34 monoclonal antibodies and inventions related to stem cells. Dr. Civin is entitled to a share of the sales royalty received by the University under licensing agreements between the University, Becton Dickinson Corporation and Baxter HealthCare Corporation. The terms of this arrangement are being managed by the Johns Hopkins University in accordance with its conflict of interest policies." @default.
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• W2032885228 date "2006-01-01" @default.
• W2032885228 modified "2023-10-14" @default.
• W2032885228 title "342. RUNX1 Increases Proliferation of Embryoid Bodies Derived from Human Embryonic Stem Cells" @default.
• W2032885228 doi "https://doi.org/10.1016/j.ymthe.2006.08.400" @default.
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