Molecular analysis of LEFTY expressing cells in early human embryoid bodies

¹ Department of Genetics, the Institute of Life Sciences, the Hebrew University, Jerusalem, Israel

^* To whom correspondence should be addressed. E-mail: nissimb{at}cc.huji.ac.il.

	Abstract

Human embryonic stem cells (HESCs) are self-renewing pluripotent cell lines that are derived from the inner cell mass of blastocyst stage embryos. These cells can produce terminally differentiated cells representing the three embryonic germ layers. We thus hypothesize that during the course of in vitro differentiation of HESCs, progenitor-like cells are transiently formed. We demonstrate that LEFTY proteins, which are known to play a major role during mouse gastrulation, are transiently expressed during HESCs differentiation. Moreover, LEFTY proteins seem to be exclusively expressed by a certain population of cells in the early human embryoid bodies (HEBs) that does not overlap with the ES cell marker, OCT4, expressing cell population. We also show that LEFTY expression is regulated at the cellular transcription level by molecular labeling of LEFTY positive cells. A DNA micro-array analysis of LEFTY over-expressing cells revealed a signature of cell surface markers such as CADHERIN 2 and 11. Expression of LEFTY controlled by NODAL appears to have a substantial role in mesodermal origin cell population establishment since inhibition of NODAL activity not only down-regulates expression of LEFTY A and LEFTY B but also BRACHYURY an early mesodermal marker. In addition, other mesodermal lineage related genes are down-regulated and this is accompanied by an up-regulation in ectoderm related genes. We propose that during the initial step of HESCs differentiation, mesoderm progenitor-like cells appear via activation of the NODAL pathway. Our analysis suggests that in vitro differentiation of HESCs can model early events in human development.

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