@misc{10481/95670,
year = {2012},
month = {6},
url = {https://hdl.handle.net/10481/95670},
abstract = {Genetic manipulation of human embryonic stem cells (hESCs) is instrumental for tracing lineage commitment and to
studying human development. Here we used hematopoietic-specific Wiskott-Aldrich syndrome gene (WAS)-promoter
driven lentiviral vectors (LVs) to achieve highly specific gene expression in hESCs-derived hematopoietic cells. We first
demonstrated that endogenous WAS gene was not expressed in undifferentiated hESCs but was evident in hemogenic
progenitors (CD45-CD31+CD34+) and hematopoietic cells (CD45+). Accordingly, WAS-promoter driven LVs were unable to
express the eGFP transgene in undifferentiated hESCs. eGFP+ cells only appeared after embryoid body (EB) hematopoietic
differentiation. The phenotypic analysis of the eGFP+ cells showed marking of different subpopulations at different days of
differentiation. At days 10–15, AWE LVs tag hemogenic and hematopoietic progenitors cells (CD45-CD31+CD34dim and
CD45+CD31+CD34dim) emerging from hESCs and at day 22 its expression became restricted to mature hematopoietic cells
(CD45+CD33+). Surprisingly, at day 10 of differentiation, the AWE vector also marked CD45-CD31low/-CD34- cells,
a population that disappeared at later stages of differentiation. We showed that the eGFP+CD452CD31+ population
generate 5 times more CD45+ cells than the eGFP-CD45-CD31+ indicating that the AWE vector was identifying
a subpopulation inside the CD45-CD31+ cells with higher hemogenic capacity. We also showed generation of CD45+ cells
from the eGFP+CD45-CD31low/-CD34- population but not from the eGFP-CD45-CD31low/-CD34- cells. This is, to our
knowledge, the first report of a gene transfer vector which specifically labels hemogenic progenitors and hematopoietic
cells emerging from hESCs. We propose the use of WAS-promoter driven LVs as a novel tool to studying human
hematopoietic development.},
organization = {Fondo de Investigaciones Sanitarias (FIS)/Fondo de Desarrollo Regional (FEDER) (grant No. PS09/00340)},
organization = {Consejería de Innovación Ciencia y Empresa (grants No. P09-CTS-04532 and PAIDI-Bio-326) and Consejería de Salud (grant No. PI0001/2009) from Junta de Andalucia},
organization = {FEDER and Fondo de Cohesión Europeo (FSE) (Programa operative FEDER/FSE de Andalucía 2007–2013)},
organization = {Fundación Progreso y Salud (Consejería de Salud - Junta de Andalucía)},
organization = {P09-CTS-04532 and PI0001/2009 grants},
organization = {Miguel Servet (CP09/00228) and Sara Borrell contracts (Fondo de Investigaciones Sanitarias (FIS) - Institute of health Carlos III)},
organization = {Junta de Andalucía/FEDER (P08-CTS-3678) and (SAS-111244 and P10-CTS-6406)},
organization = {FIS/FEDER (PI10/00449) and (PI11/00119)},
organization = {MICINN (PLE-2009-0111)},
organization = {Foundation ‘‘Spanish Association Against Cancer’’/Junta Provincial de Albacete (CI110023)},
organization = {ISCIII/Miguel Servet Program (CP07/00059 and CP09/006)},
publisher = {Plos One},
title = {Specific Marking of hESCs-Derived Hematopoietic Lineage by WAS-Promoter Driven Lentiviral Vectors},
doi = {10.1371/journal.pone.0039091},
author = {Muñoz, Pilar and Toscano, Miguel G and Real Luna, Pedro José and Benabdellah, Karim and Cobo, Marien and Bueno, Clara and Ramos Mejía, Verónica and Menéndez, Pablo and Martín Molina, Francisco},
}