Principles of stem cell biology and cancer: future applications and therapeutics. Edited by T. Regad, T. J. Sayers and R. C. Rees. John Wiley & Sons (2015)
1.1. Derivation of human embryonic stem cells from the ICM
1.1.1. Early development of the ICM: the cells of origin for hESCs
The mammalian zygote (fertilized ovum) is defined as being totipotent, as it is capable of developing into a new offspring and the placenta required for full gestation. The zygote initially undergoes cleavage-stage cell division, forming cells (early blastomeres) that remain totipotent. With further development to the preimplantation blastocyst stage, a primary cell differentiation results in outside trophectoderm cells (TE) and an inside aggregate of inner cell mass (ICM) cells. The TE forms placental tissue and membranes, while the ICM forms the foetus and extra-embryonic membranes. Therefore, ICM cells are defined as being pluripotent, forming all cells of the developing offspring other than the complete placenta (unless genetically manipulated). Embryonic stem cells (ESCs) are derived in vitro from ICM cells, which adapt to specific conducive conditions that enable indefinite cell proliferation (self-renewal) without further differentiation and thereby confer a pluripotent capacity. This in vitro pluripotent state is due principally to the induction and maintenance of expression of key ‘gate-keeper’ genes, including Oct4, Nanog and Sox2, which then regulate one another (Silva & Smith, 2008). The capacity for self-renewal is sustained by high telomerase activity, which protects chromosome telomeres from degradation during mitosis (Blasco, 2007).
Mammalian ESCs were first derived in the mouse (mESC) (Evans and Kaufman, 1981; Martin, 1981). When mESCs are integrated into an embryo and returned to a recipient, they can contribute to all cell lineages, including germ cells. Their utility soon became invaluable for many transgenic procedures. Successful derivation of human (hESC) lines was reported by Thomson et al. (1998), who essentially followed the same procedure as used for the mouse. ICMs isolated from preimplantation human blastocysts were plated on to mitotically inactivated mouse embryonic feeders in culture medium with basic fibroblast growth factor (bFGF) and foetal calf serum (FCS). This culture medium was also supplemented with leukaemia inhibitory factor (LIF), a cytokine necessary to maintain mESCs (Smith et al., 1988), although (as is now known) not necessary for standard hESC derivation. Human ESCs display (or lose on differentiation) plasma membrane expression of stage-specific embryonic antigens (SSEAs) that correlate with the preimplantation morphological development of human embryos (Henderson et al., 2002) and form teratomas (benign tumours) in immune-deficient mice that can contain cell phenotypes from the three major cell lineages (endoderm, mesoderm and ectoderm), as well as trophoblast. The differentiation of trophoblast cells indicates that hESCs are not entirely equivalent to mESCs, as usually defined, but align with slightly later LIF-independent mouse epiblast pluripotent stem cells, which have the propensity to differentiate to trophoblast in vitro (Brons et al., 2007).