Figure 6
Generation of iPSCs from BM samples from a patient in the chronic phase of CML. (A) Flow cytometric analysis of hESC-specific marker expression in CML iPSC15 and CML iPSC17. (B) Bright-field image demonstrating typical hESC morphology of CML iPSCs growing on MEFs. (C) Representative immunofluorescent staining of CML iPSCs with REX1 antibody. Bar indicates 50 μm. (D) Representative H&E staining of teratoma generated from CMLiPS15 showing derivatives of 3 germ layers as indicated in each of the panels. (E) Flow cytometric demonstration of differentiation of CMLiPS15 into blood cells in OP9 coculture. (F) Colony-forming unit assay from blood progenitor cells differentiated from line CML iPSC15. BFU-E indicates burst-forming unit-erythroid; CFU-GEMM; colony-forming unit-granulocyte, erythrocyte, monocyte, and megakaryocyte; CFU-M, colony-forming unit-macrophage; CFU-GM, colony-forming unit-granulocyte and monocyte. (G) CML iPSC lines 15 and 17 are free of transgene and vector sequence; E indicates the episomal fraction and G the genomic fraction of DNA; BM, human BM genomic DNA; P1, human BM mononuclear cells transfected with identical constructs. The T series of primers are transgene specific. ACTB indicates human actin primers that were used to check the DNA quality. (H) CML iPSCs express pluripotent genes, but not the corresponding transgenes. P1 indicates human BM mononuclear cells transfected with identical constructs. The hESC line H1 is the positive control and the Philadelphia chromosome-positive line K562 is used as the negative control for pluripotency, but as a positive control for the BCR-ABL fusion gene.

Generation of iPSCs from BM samples from a patient in the chronic phase of CML. (A) Flow cytometric analysis of hESC-specific marker expression in CML iPSC15 and CML iPSC17. (B) Bright-field image demonstrating typical hESC morphology of CML iPSCs growing on MEFs. (C) Representative immunofluorescent staining of CML iPSCs with REX1 antibody. Bar indicates 50 μm. (D) Representative H&E staining of teratoma generated from CMLiPS15 showing derivatives of 3 germ layers as indicated in each of the panels. (E) Flow cytometric demonstration of differentiation of CMLiPS15 into blood cells in OP9 coculture. (F) Colony-forming unit assay from blood progenitor cells differentiated from line CML iPSC15. BFU-E indicates burst-forming unit-erythroid; CFU-GEMM; colony-forming unit-granulocyte, erythrocyte, monocyte, and megakaryocyte; CFU-M, colony-forming unit-macrophage; CFU-GM, colony-forming unit-granulocyte and monocyte. (G) CML iPSC lines 15 and 17 are free of transgene and vector sequence; E indicates the episomal fraction and G the genomic fraction of DNA; BM, human BM genomic DNA; P1, human BM mononuclear cells transfected with identical constructs. The T series of primers are transgene specific. ACTB indicates human actin primers that were used to check the DNA quality. (H) CML iPSCs express pluripotent genes, but not the corresponding transgenes. P1 indicates human BM mononuclear cells transfected with identical constructs. The hESC line H1 is the positive control and the Philadelphia chromosome-positive line K562 is used as the negative control for pluripotency, but as a positive control for the BCR-ABL fusion gene.

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