THE QUEST FOR PORCINE PLURIPOTENT STEM CELLS

Abstract

The isolation of embryonic stem cells (ESC) and embryonic germ cells (EGC) from early embryos is a major milestone in modern science and holds a great potential for human medicine. In 2007, Shinia Yamanaka and co-workers reprogrammed somatic cells to pluripotency by induced expression of pluripotency transcription factors. These so-called induced pluripotent stem cells (iPSC) are equivalent to ESC in terms of pluripotency and have the same potential for use in regenerative therapies. However, before the use of pluripotent cells or their derivatives in humans, potential therapies need to be tested in suitable animal models to ensure their safety. In this respect, the domestic pig is particularly suited for the testing of stem cell-based therapies intended for humans, since in general physiology and metabolism are similar in human and pigs. Since the isolation of the different types of pluripotent cells in human and mouse, there have been reports of derivation of ESC-like and EGC-like cell lines from porcine embryos. Despite the significant progress that has been reported in these studies, none of the described porcine cell lines have fulfilled all of the criteria for pluripotency, such as long-term maintenance and the ability to differentiate into all of the cells in the organism, including the germ line. This has prevented the use of these cells in the genetic engineering of livestock as well as their therapeutic application in animal models for human diseases. The derivation of the first porcine cell lines with iPSC characteristics (Ezashi et al. 2009 PNAS 27, 10 993–10 998) has provided a viable alternative to the ESC/EGC, and some major successes have been already achieved. The majority of the putative iPSC described in the literature have demonstrated pluripotent characteristics such as expression of various pluripotency markers and an ability to differentiate into the three primary germ layers in vivo by forming teratomas in immunodeficient mice. One group has reported the derivation of iPSC lines that have been capable to generate chimeras with germline contribution (West et al. 2011 Stem Cells 29, 1640–1643), which is the first fully confirmed report of successfully produced porcine germ line chimera to date. Additionally, the differentiation of putative iPSC into rod photoreceptors and their integration into the retinas of recipient pigs has been reported (Zhou et al. 2011 Stem Cells 29, 972–980). Despite these major achievements, some challenges remain to be overcome in order to make porcine iPSC more widely applicable in disease models and in the transgenic technology. Due to some variations in the morphological and molecular characteristics of the reported putative iPSC lines, it needs to be determined which markers are the hallmarks of truly pluripotent porcine iPSC. Second, it is still not clear which are the optimal culture conditions for derivation and long-term culture of these cells. Since the culture conditions used today have been proven ineffective to maintain pluripotency in porcine ESC and EGC, the question remains whether the continuous expression of the transgenes is an important factor in the long-term culture of iPSC. Finally, it needs to be determined whether putative porcine iPSC derived from cell types other than multipotent stem cells (such as mesenchymal stem cells used by West et al., 2011) possess full pluripotency, which should be demonstrated by germ line chimera production via blastocyst injection or tetraploid complementation.