Postdoctoral Research Fellow The University of Queensland Brisbane, Queensland, Australia
Abstract: Traumatic spinal cord injury (SCI) is a devastating condition which causes irreversible damage to neural circuits and pathways. Therapeutic strategies involving neural stem cell transplants hold significant promise to replace lost tissue, restore continuity across the lesion and, in doing so, promote functional recovery from SCI. Propriospinal interneurons may be ideal for transplantation purposes as these cells naturally connect distant segments of the spinal cord through both ascending and descending projections, and they are also thought to have crucial roles in motor control. This study explored the feasibility of generating V2a propriospinal interneurons from mouse-induced pluripotent stem cells (iPSCs) and subsequently transplanting these cells syngeneically into SCI mice. To guide iPSCs towards a Chx10+ V2a interneuron fate, a 6-day differentiation protocol was employed, that included the precise temporal addition of specific morphogens: retinoic acid, sonic hedgehog, and a notch signalling inhibitor. A constitutive tdTomato expressing Chx10- Blasticidin S deaminase (BSD) selectable cell line was then generated for purification and identification of Chx10+ transplants. Following differentiation, a significant downregulation of the pluripotency gene Oct4 was observed by day 4, along with the induction of the neural stem cell markers Nestin and NCAM. Expression analysis of the transcription factor Chx10, and also Tuj1 as a more general neuronal lineage marker, confirmed that V2a interneurons can indeed be generated from these mouse iPSCs (~17% efficiency). The transplantation of purified V2a interneurons into SCI mice resulted in their survival, extensive neurite outgrowth, and integration within the injured spinal cord. Importantly, these cellular changes corresponded with significant functional recovery at 1 month post-transplantation. Future experiments will focus on assessing the longer-term therapeutic potential of these cells as neuronal relays, with the ultimate goal of restoring function across the site of SCI.
Funding Source: Perry Cross Spinal Research Foundation
