Development of Neuroregenerative Gene Therapy to Reverse Glial Scar Tissue Back to Neuron-Enriched Tissue

Abstract

Injuries in the central nervous system (CNS) often causes neuronal loss and glial scar formation. We have recently demonstrated NeuroD1-mediated direct conversion of reactive glial cells into functional neurons in adult mouse brains. Here, we further investigate whether such direct glia-to-neuron conversion technology can reverse glial scar back to neural tissue in a severe stab injury model of the mouse cortex. Using an adeno-associated virus (AAV)-based gene therapy approach, we ectopically expressed a single neural transcription factor NeuroD1 in reactive astrocytes in the injured areas. We discovered that the reactive astrocytes were efficiently converted into neurons both before and after glial scar formation, and the remaining astrocytes proliferated to repopulate themselves. The astrocyte-converted neurons were highly functional, capable of firing action potentials and establishing synaptic connections with other neurons. Unexpectedly, the expression of NeuroD1 in reactive astrocytes resulted in a significant reduction of toxic A1 astrocytes, together with a significant decrease of reactive microglia and neuroinflammation. Furthermore, accompanying the regeneration of new neurons and repopulation of new astrocytes, new blood vessels emerged and blood-brain-barrier (BBB) was restored. These results demonstrate an innovative neuroregenerative gene therapy that can directly reverse glial scar back to neural tissue, opening a new avenue for brain repair after injury.

Regeneration of Functional Neurons After Spinal Cord Injury via in situ NeuroD1-Mediated Astrocyte-to-Neuron Conversion

“…Following our earlier report of efficient conversion of reactive astrocytes into functional neurons by NeuroD1 through retroviral infection (Guo et al, 2014), intravenous injection of AAV9 expressing NeuroD1 has been reported to infect a small but significant number of resting astrocytes in the striatum and convert them into neurons (Brulet et al, 2017). On the other hand, our intracranial injection of AAV9 expressing NeuroD1 in stab injury model (Zhang et al, 2020) and ischemic injury model (Chen et al, 2020) have both resulted in high conversion efficiency, suggesting that reactive astrocytes after injury are more likely converted into neurons than the resting astrocytes. Consistent with this hypothesis, our results in this study also support that reactive astrocytes in injured spinal cord can be effectively converted into neurons with ∼95% efficiency, regardless of retrovirus or AAV delivery system.…”

“…The high efficiency of astrocyte-to-neuron conversion is the major reason why we target astrocytes among glial cells for in vivo conversion. Importantly, we have demonstrated in the mouse cortex that after astrocyte-to-neuron conversion, the remaining astrocytes can proliferate and replenish themselves (Zhang et al, 2020). Unlike killing reactive astrocytes which has been reported to make injury worse (Anderson et al, 2016), we demonstrated that converting reactive astrocytes into functional new neurons significantly ameliorated the glial scar in the brain, leading to a reversal of glial scar back to neural tissue (Zhang et al, 2020).…”

“…Importantly, we have demonstrated in the mouse cortex that after astrocyte-to-neuron conversion, the remaining astrocytes can proliferate and replenish themselves (Zhang et al, 2020). Unlike killing reactive astrocytes which has been reported to make injury worse (Anderson et al, 2016), we demonstrated that converting reactive astrocytes into functional new neurons significantly ameliorated the glial scar in the brain, leading to a reversal of glial scar back to neural tissue (Zhang et al, 2020). Another practical reason for targeting astrocytes is that among glial cells including astrocytes, NG2 cells (OPCs), and microglia, AAV preferentially infect astrocytes, with much less infection rate on NG2 cells or microglia.…”

“…We previously demonstrated that expressing NeuroD1 in reactive astrocytes after brain injury can directly convert astrocytes into neurons (Guo et al, 2014;Chen et al, 2020;Zhang et al, 2020). In this study, we investigated whether such in vivo direct conversion technology can regenerate functional new neurons in injured spinal cord.…”

“…Our group has previously demonstrated that the neurogenic transcription factor NeuroD1 can reprogram reactive astrocytes into functional neurons in the stab-injured brain and in a mouse model for Alzheimer's disease (Guo et al, 2014). Following studies demonstrated that NeuroD1-mediated in vivo astrocyteto-neuron conversion can reverse glial scar back to neural tissue (Zhang et al, 2020) and repair the damaged motor cortex after ischemic stroke (Chen et al, 2020). More recently, by combining NeuroD1 and Dlx2 together, we have demonstrated that astrocytes in the striatum of R6/2 mouse model for Huntington's disease can be converted into GABAergic neurons (Wu et al, 2020).…”