Down syndrome (DS) is the most frequent cause of human congenital mental retardation. Cognitive deficits in DS result from perturbations of normal cellular processes both during development and in adult tissues, but the mechanisms underlying DS etiology remain poorly understood. To assess the ability of induced pluripotent stem cells (iPSCs) to model DS phenotypes, as a prototypical complex human disease, we generated bona fide DS and wild-type (WT) nonviral iPSCs by episomal reprogramming. DS iPSCs selectively overexpressed chromosome 21 genes, consistent with gene dosage, which was associated with deregulation of thousands of genes throughout the genome. DS and WT iPSCs were neurally converted at >95% efficiency and had remarkably similar lineage potency, differentiation kinetics, proliferation, and axon extension at early time points. However, at later time points DS cultures showed a twofold bias toward glial lineages. Moreover, DS neural cultures were up to two times more sensitive to oxidative stress-induced apoptosis, and this could be prevented by the antioxidant N-acetylcysteine. Our results reveal a striking complexity in the genetic alterations caused by trisomy 21 that are likely to underlie DS developmental phenotypes, and indicate a central role for defective early glial development in establishing developmental defects in DS brains. Furthermore, oxidative stress sensitivity is likely to contribute to the accelerated neurodegeneration seen in DS, and we provide proof of concept for screening corrective therapeutics using DS iPSCs and their derivatives. Nonviral DS iPSCs can therefore model features of complex human disease in vitro and provide a renewable and ethically unencumbered discovery platform.

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This study compared the developmental potential of bovine nuclear transfer embryos with varying amounts of cytoplasm. Embryos formed from single cytoplasts fused to blastomeres by a single electrical pulse or from double cytoplasts using a double electrical pulse resulted in reconstituted embryos containing 75% and 150% of the original oocyte volume. No differences in fusion, cleavage, or development rates to blastocysts were observed between the groups. Mean cell numbers 2 days after fusion were significantly lower in single‐cytoplast clones. Cell numbers of resulting blastocysts were likewise significantly lower in single‐cytoplast clones. Embryos formed by fusion of blastomeres with single cytoplasts using a single electrical pulse or from double cytoplasts using either a single or a double pulse resulted in reconstituted embryos containing 50%, 100% and 100% of the original oocyte volume. Again, no differences in fusion or cleavage rates were observed between groups, but the development to blastocysts at day 7 was significantly higher in double cytoplasts constructed with one fusion pulse than in single cytoplasts (P< 0.05). Mean cell numbers 2 days after fusion were significantly lower in single‐cytoplast clones (P< 0.05), but at the blastocyst stage, no statistically significant differences in cell numbers were observed. The results of this study show that cytoplasmic volume plays a role in the development of nuclear transfer embryos. When using crude enucleation methods such as oocyte bisection, normal cytoplasmic volumes can be achieved by fusing double cytoplasts with embryonic blastomeres. Mol. Reprod. Dev. 50:185–191, 1998. © 1998 Wiley‐Liss, Inc.

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