Understanding Pathogenesis of Lissencephaly with Patient-Derived Induced Pluripotent Stem Cells. M. Bershteyn1, A. Kriegstein1, A. Wynshaw-Boris2 1) Edyth and Eli Broad Institute of Regeneration Medi, University of California, San-Francisco School of Medicine, San Francisco, CA; 2) Department of Genetics and Genome Sciences, Case Western Reserve University, Cleveland OH, USA.

   Normal development of the cerebral cortex requires a complex series of cellular events, including specification, proliferation, migration and differentiation, to establish the proper structure and function. Mutations that disrupt these key developmental processes give rise to cortical malformations. Miller Dieker Syndrome (MDS) is a genetic developmental disorder characterized by severe cortical malformations including reduced brain size (microcephaly) and nearly absent cortical folding (lissencephaly), with devastating neurological consequences such as mental retardation and intractable epilepsy. MDS is caused by heterozygous deletions of human band 17p13.3, harboring several dozen genes, including PAFAH1B1. Analyses of Pafah1b1 mutant mice revealed defects in neuronal migration, which is considered to be the main cellular deficiency in lissencephaly. However, it is unknown whether induction, proliferation or differentiation of neural stem cells are also disrupted in MDS, and the roles of most of the other deleted genes from locus 17p13.3 in cortical development or MDS pathogenesis have not been examined. To study the cellular and molecular mechanisms of MDS pathogenesis, we generated induced pluripotent stem cells (iPSCs) from MDS patients. We found that MDS iPSCs can proliferate, self-renew and generate all three germ layers in vitro and in vivo. Moreover, using 2-D and 3-D in vitro differentiation methods, we observed efficient induction of neuroepithelial stem cells and radial glia, exhibiting characteristic mitotic behaviors such as interkinetic nuclear migration and mitotic somal translocation. These results suggest that haploinsufficiency for locus 17p13.3 does not grossly impair these key features of cortical development. However, upon high-density culture conditions, MDS neural progenitors exhibited increased apoptosis. Our results so far suggest that reduced viability of neural stem cells may be a primary factor in MDS pathogenesis that precedes any defects in neuronal migration. Additional studies are underway to characterize other aspects of human cortical development such as proliferation of various types of progenitors, migration and differentiation.

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