Modeling early human neural development using iPS cells

Human induced Pluripotent Stem Cells (iPSCs) have long been known for their great potential in disease and development modeling as well as their possible use for cell transplantations. This thesis investigates the advantages and limitations of human iPSCs. We investigated the fundamentals of neural lineage specifications during neural induction. In addition to modeling human neural development, we assessed the therapeutic efficiency of iPSCs derived neural epithelial stem (NES) cell doses in a pre-clinical study of spinal cord injury (SCI). Modeling development and the generation of cells for transplantations is dependent on the generation of standardized and trustable cells. NES cells are a great tool for both fields. They provide unlimited supply of neural progenitor cells and are able to generate into all major neural lineages. This thesis consists of one protocol for the standardized generation of NES cells, two studies using these cells for pre-clinical cell transplantation studies, and one paper to investigate the role of an adhesion molecule during early neural development. All included papers provide deeper insights into development in health and disease.

In paper I we develop a protocol for the standardized and robust generation of NES cells. This neural induction protocol can be used to generate cells relevant for transplantations and for modeling human development in 2D.

In paper II we prove that the generation of big batches of transplantable cells is possible and has many advantages. The cells have robust viability and keep differentiation potential even after prolonged periods of freezing. The quality of the cells can be assed prior to transplantation.

In paper III off-the-shelf NES cell doses were transplanted into a rat SCI model. The cells were able to survive and differentiate into relevant neural cell types. Transplantation showed positive effects regarding regeneration tissue integrity.

In paper IV we identified changes in relevant signaling pathways and early development in cells with bi-allelic NRXNla deletion. We thereby identified a possible mechanism of action for this adhesion molecule early in development. Developmental changes included change in regionalization, switch in cell fate, further progression in EMT, and higher levels of TGFB and BMP signaling.

In total, this thesis provides methods and protocols for generation of possible cell transplantation products in the future. We also show that these cells can be used in disease modeling in 2D and to identify new roles of proteins beyond their known function.

List of scientific papers

I. Protocol for the derivation, culturing and differentiation of human iPS-cell-derived neuroepithelial stem cells to study neural differentiation in vitro. Javier Calvo-Garrido, Dania Winn, Camilla Maffezzini, Anna Wedell, Christoph Freyer, Anna Falk, Anna Wredenberg. STAR Protocols. (2021). https://doi.org/10.1016/j.xpro.2021.100528

II. Pre-clinical evaluation of clinically relevant iPSC derived neuroepithelial stem cells as an off-the-shelf cell therapy for spinal cord injury. Dania Winn, Elias Uhlin, Malin Kele, Ilse Eidhof, Anna Falk Frontiers. Pharmacology. (2024). https://doi.org/10.3389/fphar.2024.1390058

III. Multiple therapeutic effects of human neural stem cells derived from induced pluripotent stem cells in a rat model of post-traumatic syringomyelia. Tingting Xu, Xiaofei Li, Yuxi Guo, Elias Uhlin, Lena Holmberg, Sumonto Mitra, Dania Winn, Anna Falk, Eriks Sundström. eBioMedicine. (2022). https://doi.org/10.1016/j.ebiom.2022.103882

IV. Multi-Omic Profiling reveals the Impact of NRXNa Deletion on early Neural Development in an iPSC Model. Dania Winn, Alireza Ghahramani, Sarfraz Shafiq, Yan Jiang, Ilse Eidhof, Nathalie Bérubé, Anna Falk. [Manuscript]