Micro-RNA Profiling Reveals the Key Role of miR-206 in the Hematopoietic Potential of Human Embryonic and Induced Pluripotent Stem Cells

• Published in: Blood 2018-11, Vol.132 (Supplement 1), p.1275-1275
• Main Authors: Flamant, Stephane, Chomel, Jean-Claude, Desterke, Christophe, Feraud, Olivier, Gobbo, Emilie, Mitjavila-Garcia, Maria Teresa, Foudi, Adlen, Griscelli, Frank, Turhan, Ali G, Bennaceur-Griscelli, Annelise
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2018-99-110729

Although human pluripotent stem cells (hPSCs) can theoretically be differentiated into any cell type, their ability to generate hematopoietic cells shows a major variability from one cell line to another. The reasons of this variable differentiation potential, which is constant and reproducible in a given hPSC line, are not clearly established. In order to study this phenomenon, we comparatively studied 4 human embryonic stem cell lines (hESC) and 11 human induced pluripotent stem cell (hiPSC) lines using transcriptome assays. These cell lines exhibited a significant variability to generate in vitro hematopoiesis as evaluated by day-16 embryoid body (EB) formation followed by clonogenic (CFC) assays. Four out of 11 iPSC lines (PB6, PB9, PB12.1, and PB14.3) were found to lack any hematopoietic differentiation ability whereas 7 cell lines showed variable hematopoietic potential. Among hESC lines, H9 and CL0 had low H1 and SA01 exhibited high hematopoietic potential using the above assays. Among hESC and hIPSC displaying hematopoietic potential, two sub-groups were further defined based on their hematopoietic CFC efficiency: a group of poor (generation of less than 100 CFC/105 cells, PB4 / PB10 /H9 /CL01), and high hematopoietic competency (more than 120 CFC/105 cells, PB3/ PB6.1 /PB7 /PB13 /PB17 /SA01/H1). Using global miRNome analysis performed at the pluripotency stage, the expression of 754 individual miRNAs was analyzed from 15 hPSC lines in order to explore a potential predictive marker between both sub-groups of pluripotent cells according to their hematopoietic potency. Using this approach, 27 miRNAs out of 754 appeared differentially expressed allowing the identification of a miRNA signature associated with hematopoietic-competency. The hematopoietic competency was associated with down-regulation of miR-206, miR-135b, miR-105, miR-492, miR-622 and upregulation of miR-520a, miR-296, miR-122, miR-515, miR-335. Amongst these, miR-206 harbored the most significant variation (0.04-Fold change). To explore the role of miRNA-206 in this phenomenon, we have generated a miR-206-eFGP-Puro lentiviral vector which was transfected in hESC line H1 followed by puromycin selection. As a control, H1 cell line was transfected with a Arabidopsis thaliana microRNA sequence (ath-miR-159a), which has no specific targets in mammalian cells. The correct expression of the transgenes were evaluated by flow cytometry (using GFP) and q-RT-PCR for miR-206 expression. The hematop