Defects in in vitro generated stem cells

There are two methods for reprogramming mature cells to pluripotent stem cells, which can give rise to all cells of the body. The first direct comparison of the methods reveals that both can cause subtle molecular defects.

Pluripotent stem cells hold promise for disease modelling and therapeutics, because they have the potential to differentiate into almost all cell lineages. In particular, there is much interest in patient-derived pluripotent stem cells, which are genetically matched to the patient’s own cells, minimizing the risk of rejection by the immune system.

In the past decade, two cell-reprogramming methods have been successfully used to generate patient-derived pluripotent stem cells: (1) cloning and (2) direct reprogramming of differentiated cells to induced pluripotent stem cells, through the addition of a defined transcription-factor cocktail.

However, the molecular differences between cells derived using each method remain unclear.

Derivation of induced pluripotent stem (iPS) cells is an appealing technology, because iPS cells can be reproducibly derived from patient samples. But comparison of iPS cells with the pluripotent embryonic stem (ES) cells generated during normal embryogenesis shows that human iPS cells are not completely reprogrammed, and reveals epigenetic differences between the two cell types (epigenetic marks are lingering genomic modifications that affect gene expression without changing DNA sequence).

Cloning — also called somatic cell nuclear transfer (SCNT) — involves transfer of the nuclear material from a mature donor cell into an egg from which the nucleus has been removed. Pluripotent cells, called nuclear transfer ES (NT ES) cells, which are genetically matched to the donor, then arise as the egg begins to develop into an embryo. Generation of patient-specific NT ES cells from adult human cells is now feasible. Although SCNT does not involve introducing transcription factors that have the potential to cause cancer (which is a problem with iPS cell generation), the protocol is technically difficult.

Figure 1: Comparing techniques for generating stem cells.

Comparing techniques for generating stem cells.

Three techniques can be used to generate pluripotent stem cells in vitro:

a) Induced pluripotent stem (iPS) cells are generated from mature cells, which can be directly converted by the addition of a transcription-factor cocktail.

b) In somatic cell nuclear transfer (SCNT), the nucleus is removed from an egg and replaced with the nucleus from a mature donor cell. As this hybrid cell develops into an embryo, pluripotent stem cells called nuclear transfer embryonic stem (NT ES) cells can be extracted from a region called the inner cell mass (ICM).

c) Embryos derived from in vitro fertilization (IVF) give rise to IVF ES cells that can be extracted from the ICM.

Incomplete demethylation patterns correlated with abnormal gene transcription were observed in iPS cells. NT ES cells were more similar to IVF ES cells, although some transcriptional alterations were apparent in both reprogrammed cell types.

There is an abundance of factors that can be used to reprogram cells and expand them in vitro, and each can influence the epigenetic and functional properties of reprogrammed cells in distinct ways. This complexity disrupts simplistic attempts to define and obtain ‘perfect’ stem cells. (Vladislav KrupalnikJacob H. HannaNature, 511,160 – 162 (10 July 2014)) The Research article subject to this review: Ma et al., Abnormalities in human pluripotent cells due to reprogramming mechanisms, Nature, 511,177–183(10 July 2014)doi:10.1038/nature13551


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