Download 2014-10-Chandrasekaran-Heidelberg-Abstract

IMC
October 9-11, 2014
PATIENT SPECIFIC INDUCED PLURIPOTENT STEM CELLS
AND THEIR NEURONAL DIFFERENTIATION
Abinaya Chandrasekaran1 – Lone Neelchen Roesingh1,2 – Csilla Nemes1– Eszter
Varga11,3– Istvan Bock1,3– Shuling Zhou1,4– Anna Ochalek Dorota3 – Viktor Szegedi1–
Hasan Avci1– András Dinnyés1,3,5 and Julianna Kobolák1
1 Biotalentum Ltd; Gödöllő, Hungary
2 University of Geneva; Dept Pathology and Immunology, Geneva, Switzerland
3 Molecular Animal Biotechnology Lab; Szent Istvan University, Godollo, Hungary
4 Institute for Veterinary Science, University of Copenhagen, Denmark
5 Departments of Equine Sciences and Farm Animal Health, Faculty of Veterinary Medicine; Utrecht University, The Netherlands
Keywords: Induced Pluripotent stem cells; neural stem cells; SMAD inhibition; Alzheimer’s disease.
Patient-derived induced pluripotent stem cells (iPSCs) are providing a useful tool to model
the pathology of certain diseases, especially when the affected tissue is hard to study, like in
the CNS. Pluripotent cells, carrying the genotype of a given patient, can be differentiated in
vitro to neural precursor cells (NPCs) which are then used to obtain neuronal and glial cells
in culture. These cells offer a valuable platform which allows the investigation of the
patient-specific pathomechanism of a disease. Furthermore, NPCs and their derivates can
be used for in vitro drug testing assays. Finally, these cells serve as an unlimited source of
immune compatible cells for cell replacement therapies to treat neurodegenerative disease
such as Parkinson’s or Alzheimer’s disease.
Our aim is to develop a highly efficient and reproducible assay to generate neurons and
glial cells from healthy and diseased iPSCs. Here we compared the usage of various small
molecules and recombinant proteins in a step-wise manner, which selectively induces
neural differentiation of iPSCs into NPCs and, later, into different neural linages. The
treatment induced neural rosette formation followed by the occurrence of neuroepithelial
cells (NEPs) resulting in a homogenous population of NPCs. At the NPC stage all small
molecules were withdrawn allowing for the terminal differentiation of mature post-mitotic
neurons. Neuronal lineage was confirmed by immunocytochemistry (ICC) and RT-qPCR
for neural and neuronal specific genes. Furthermore, 2D and 3D neuronal cultures were
investigated in electrophysiological measurements. We have established a highly
reproducible and efficient procedure to generate patient-specific neuronal cells form iPSCs.
References
1. S.Yamanaka et al. “Modeling Alzheimer’s Disease with iPSCs Reveals
Stress Phenotypes Associated with Intracellular Ab and Differential Drug
Responsiveness”,Cell Stem Cell., Elsevier Inc., pp.487-496, 2013.
2. Isaac S. Kohane et al. “iPSC-derived neurons as a higher-throughput readout for autism:
promises and pitfalls”, Trends in Molecular Medicine., pp.1-14, 2013.
This work was supported by grants from EU FP7 projects (STEMMAD, PIAPP-GA-2012-324451; STEMCAM,
PIAP-GA-2009-25118;
Anistem,
PIAPP-GA-2011-286264;
EpiHealth,
HEALTH-2012-F2-278418;
EpiHealthNet, PITN-GA-2012-317146; D-BOARD, FP7-HEALTH-2012-INNOVATION-1-305815).