GSX1 Promotes Neural Stem Cell Proliferation and Differentiation In Vitro

• Main Author: Skinner, Ryan
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2021
• Subjects: Bioengineering
• ISBN: 9798802705698

Spinal cord injury affects over 300,000 people in the U.S. and the prevalence increases each year by 17,000 cases. SCI causes the formation of a complex injury environment due to the initial impact and secondary signaling cascades. Currently, there are no cures for SCI and treatments focus on decreasing inflammation and using surgical decompression to limit the spread of injury. The loss of neuron function in the spinal cord causes paralysis and highlights neurogenesis as a key factor in promoting recovery. Endogenous cell responses during SCI have suggested various cell populations as potential targets for new therapeutics. The recent advances in gene therapy are promising for introducing transcription factors into the cells after SCI to promote growth and recovery. Transcription factors Gsx1 and Nkx6.1 have been used in previous studies by our lab due to their role in spinal cord development. Lentivirus mediated Gsx1 expression in mouse models of SCI has been shown to promote functional recovery, while Nkx6.1 did not promote functional recovery. In this thesis work, a neural stem cell culture model was used to further determine the effect of Gsx1 on a neural stem cell population. Gsx1 transduction was performed using both the conventional and a TetON inducible lentivirus vector in mouse and human stem cells in vitro. Lentivirus mediated transduction of Gsx1 increased the expression of proliferation marker Ki67 in both mouse and human neural stem cells while Nkx6.1 did not significantly alter proliferation in either cell line 48 hours after doxycycline induction. In addition, an increased percentage of Map2 positive neurons and a decreased percentage of GFAP positive astrocytes was observed 14 days after induction in these cultured neural stem cell lines. The inducible lentivirus allowed for easier transduction and selection of cells in culture and created stable transduced cell lines for future studies. This study supports the role of Gsx1 in promoting neuronal cell fate and the potential for Gsx1 gene therapy for spinal cord injury.