Understanding the Contributions of the Myelinating Cells in Amyotrophic Lateral Sclerosis
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2025
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Melbourne Authors
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Florey Department of Neuroscience and Mental Health
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PhD thesis
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© 2025 Katherine Nola Lewis
Abstract
Amyotrophic lateral sclerosis (ALS), also known as motor neuron disease (MND), is a fatal neurodegenerative disease characterised by the degeneration of motor neurons in the central nervous system (CNS) and peripheral nervous system (PNS). From diagnosis, patient survival is around 2-5 years and currently there are limited treatment options which only extend survival by mere months. The need for more effective therapeutics is integral for improving these survival outcomes, which, in turn, requires a deeper understanding of what drives disease onset and progression.
One of the hallmark pathologies of ALS is the cytoplasmic accumulation of TDP-43, a protein produced by the TARDBP gene. In 97% of all ALS patients, TDP-43 becomes mislocalised from the cell nucleus, disrupts stress granule formation and autophagy, and forms cytoplasmic aggregates, resulting in cell death. It is not only the motor neurons that exhibit pathological TDP-43, but also the surrounding glia, including the myelinating cells. Myelin is the lipid-rich sheath that encases neuronal axons, enabling for the efficient transmission of action potentials while simultaneously providing the neuron with metabolic and trophic support. Oligodendrocytes are the myelin producing cells of the CNS, which arise through the differentiation of their progenitor: oligodendrocyte precursor cells (OPCs). Schwann cells are the myelinating cells of the PNS and perform integral functions such as modulating neuronal signalling, contributing to the immune response, and trafficking essential metabolic and trophic resources to the axon. Despite both oligodendroglia and Schwann cells being found to exhibit pathological TDP-43 proteins in ALS, however their direct contributions to disease phenotype remains understudied. Since myelination of both the CNS and PNS is a tightly regulated, evolutionarily conserved process, if disrupted in ALS, the myelinating cells could be contributing to ALS pathology. Understanding the role of the myelinating cells in ALS will allow for a greater comprehension of their contributions to pathology to determine if they are viable targets for therapeutic intervention. This PhD aimed to comprehensively investigate the contributions of the myelinating cells in ALS using a TDP-43Q331K (TDP-43) transgenic mouse model. This model employs the human TDP-43Q331K transgene to produce an overexpression of pathogenic TDP-43 protein. While the overall pathology is relatively mild, this mouse presents with motor behavioural phenotypes, neuromuscular junction (NMJ) pathology, and motor end-plate defects, all of which are considered in line with ALS patient phenotypes and was therefore an ideal model system to interrogate myelin perturbations in ALS.
Chapter 3 examined the proliferative and population dynamics of oligodendroglia and CNS myelin throughout the disease course in the lumbar spinal cord. In the ventral horn grey matter at disease end-stage, OPC proliferation and differentiation were significantly increased in the TDP-43 mice compared to their wildtype (WT) littermates. Despite an increase in OPC differentiation, the population of mature oligodendrocytes remained unchanged in the TDP-43 mice, in part due to increased oligodendroglial cell death. Myelin was examined using Spectral Confocal Reflectance Microscopy (SCoRe) and showed an increase in myelin reflectance, which was revealed via electron microscopy (EM) to be likely caused by degenerating, misfolded, and pathological myelin in the TDP-43 mice compared to WT. Thus, oligodendroglia and CNS myelin were significantly impacted in TDP-43 mice at end-stage, suggesting they may be contributing to disease and exacerbating motor phenotypes.
Chapter 4 explored the PNS myelin and Schwann cell behaviours in the TDP-43 mice. We also crossed a myelin protein zero (P0)-cre mouse strain with the floxxed TDP-43Q331K mice to selectively excise the human transgene from Schwann cells alone. Western Blot analyses of TDP-43 expression showed a ~50 % reduction in the TDP-43 protein in the sciatic nerves of P0-cre/TDP-43 mice compared to TDP-43 mice, with no change to TDP-43 protein expression in the spinal cord. Histological analyses of sciatic nerves of end-stage mice showed a change in the structure of the node of Ranvier and the flanking paranodes in the TDP-43 mice, which was not observed in the WT or P0-cre/TDP-43 mice. SCoRe imaging and EM showed an increase in myelin reflectance and myelin abnormalities, respectively, in sciatic nerves of TDP-43 mice, which were unchanged in the P0-cre/TDP-43 mice compared to WT. To assess the functional impact of these rescued pathologies, we conducted behavioural analyses, sciatic nerve electrophysiology, and retrograde axonal transport tracking, allowing us to assess neuronal function on a behavioural, circuit, and intracellular level. While deficits were identified in TDP-43 mice compared to WT, they were not rescued in the P0-cre/TDP-43 mice. Ultimately, despite a normalisation of missing paranodal proteins, node length, and PNS myelin phenotypes, rescuing Schwann cells alone is not sufficient to overcome the functional deficits in TDP-43Q331K mice. Thus, solely targeting Schwann cells therapeutically in ALS may not improve patient outcomes and investigation into the treatment of both Schwann cells and motor neurons may be of benefit.
We are the first to characterise the cellular behaviours of oligodendroglia, Schwann cells, and myelin, indeed identifying dysfunction, in the TDP-43Q331K mouse, bringing research one step closer to identifying if these cells present as a viable therapeutic target for ALS.
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Oligodendrocyte, Schwann Cell, Myelin, Amyotrophic Lateral Sclerosis (ALS), Motor Neuron Disease (MND), TDP-43