SCL Student

Mark Curtis

Research

Directed Differentiation of Human Embryonic Stem (hES) Cells to Neural Progenitors and Dopaminergic (DA) Neurons
Over the last decade extensive research has been invested in the controlled differentiation of hES cells to various cell types in the hopes of using these cells for transplantation therapy. One prominent neurodegenerative disease experienced by the human population is Parkinson’s disease (PD). PD is typified by the gradual degeneration of the DA neurons that populate the substantia nigra, a portion of the midbrain. In the future it is conceivable that DA neurons derived from hES cells could be used to remedy the symptoms associated with PD. At the current time however, several issues exist that prevent these differentiated cell populations from being clinically relevant. The formation of teratomas after in vivo transplantation remains a safety hazard. The presence of this tumor type following transplantation into 6-OHDA lesioned mice indicates that current protocols for the directed differentiation of hES cells to DA neurons produce heterogeneous populations of cells which likely include minorities of undifferentiated hES cells and neural progenitors (NPs).

Bone morphogenetic proteins (BMPs) have been implicated as key players within a pathway that controls hES cell fate decision. The presence of BMP activity induces differentiation of hES cells towards mesoderm and endoderm. Thus, inhibition of this group of proteins has presented itself as an efficient mechanism to induce the formation of neurectoderm and NPs. These NPs can subsequently be differentiated to the DA phenotype with the use of several patterning and signaling molecules, including Sonic Hedgehog (SHH) and Fibroblast Growth Factor 8 (FGF8).

My research focuses on developing an improved protocol for controlled neural induction from hES cells. This protocol will integrate previous methods that have been shown to successfully improve the yield of functional DA neurons from hES cells. It will include a three-tier differentiation regime wherein hES cells are first differentiated towards neurectoderm using a stromal cell coculture with PA6 cells. The second step will involve treatment of neurectoderm like cells with Noggin, a BMP antagonist, to induce the formation of neural progenitors. Finally, these cells will be differentiated towards the DA phenotype using SHH and FGF8. In addition to this differentiation protocol, an RNAi based treatment will be included wherein hES cells down regulated for the NANOG genes are subjected to the above regime. NANOG is a gene which aids in the maintenance of pluripotency of hES cells and it is believed that down-regulating this gene may further reduce the number of pluripotent intermediates present after the differentiation process.