Periodic Reporting for period 4 - AxoMyoGlia (Spatio-functional cellular interplay in peripheral nerve diseases)
Okres sprawozdawczy: 2025-07-01 do 2025-12-31
Neuromuscular disorders represent prevalent neurological conditions characterized by compromised motor function. These disorders involve multiple interacting cell types within the neuromuscular unit, including motoneurons and their axons, glial cells, as well as muscle cells. Although these components form a highly interconnected system, the mechanisms governing their spatial and functional interactions in disease remain poorly understood. The AxoMyoGlia project aimed to elucidate how cellular interactions within the neuromuscular unit contribute to disease progression. Using peripheral neuropathies as a model system, the project investigated how glial dysfunction affects axonal integrity and how these effects propagate along the neuromuscular axis. A central goal was to understand how local alterations in glial cells influence not only axons but also distant compartments such as the spinal cord and muscle. To address this, we combined advanced transcriptomic and imaging technologies with genetic mouse models to generate a comprehensive, spatially resolved map of cellular interactions within the neuromuscular system. A key focus of the project was the spatial organization of disease processes. Our work has revealed that neuropathic alterations are not uniformly distributed along nerves, but instead show region-specific patterns, including distinct inflammatory responses and differential vulnerability of axonal subtypes. This spatial heterogeneity is closely linked to previously unrecognized diversity among glial cells, particularly myelinating Schwann cells, which exhibit distinct molecular and functional properties.
Overall, the objectives of AxoMyoGlia were to establish a molecular and morphological framework of cellular interactions in neuropathy, to define local and remote mechanisms of degeneration and regeneration, and to investigate cellular responses at the neuromuscular interface. By achieving these goals, the project sought to uncover fundamental mechanisms underlying neuromuscular disorders and to provide a basis for improved therapeutic strategies.
Overall, the objectives of AxoMyoGlia were to establish a molecular and morphological framework of cellular interactions in neuropathy, to define local and remote mechanisms of degeneration and regeneration, and to investigate cellular responses at the neuromuscular interface. By achieving these goals, the project sought to uncover fundamental mechanisms underlying neuromuscular disorders and to provide a basis for improved therapeutic strategies.
During the project, we established an integrated experimental and technological framework to investigate cellular interactions in the neuromuscular system across multiple spatial scales. A central achievement was the implementation of single-cell RNA sequencing using the iCell8 platform, enabling the combined analysis of transcriptomic profiles and morphological features of individual cells. Using this approach, we generated comprehensive datasets from peripheral nerves along the proximo-distal axis and identified previously unrecognized heterogeneity among myelinating Schwann cells. Distinct Schwann cell populations were found to exhibit specific molecular programs associated with axonal caliber and functional specialization, providing a mechanistic explanation for the selective vulnerability of axons in neuropathies. To complement these findings, we established spatial transcriptomics and advanced imaging approaches, including tissue clearing and volume electron microscopy, allowing the investigation of cellular interactions within intact tissue architecture. These analyses revealed pronounced spatial organization of neuropathic pathology, including region-specific inflammatory responses with increased immune cell infiltration in proximal nerve segments. In addition, we identified transcriptional and cellular alterations in the spinal cord, including activation and spatial reorganization of microglia around motoneurons, indicating a remote central nervous system response to peripheral nerve dysfunction. To dissect causal mechanisms, we developed novel mouse models enabling spatially restricted induction of glial dysfunction, including conditional overexpression of the neuropathy-associated gene Pmp22 in defined nerve compartments. These models allow direct investigation of how local perturbations propagate along the neuromuscular axis. In parallel, we established focal demyelination paradigms to study local and distant effects of myelin damage. Building on complementary work in the central nervous system, we demonstrated that persistent myelin ensheathment by dysfunctional glial cells can compromise axonal integrity, highlighting the importance of glial support functions for neuronal survival (Schäffner et al., Nature Neuroscience, 2023). Together, these findings establish a multi-scale framework linking glial heterogeneity, spatial organization of pathology, and axonal degeneration in neuromuscular disease.
The AxoMyoGlia project advanced the field beyond the current state of the art by introducing a spatially resolved and multi-scale perspective on axon-glia interactions in neuropathies. A key conceptual advance is the recognition that glial cells are not a uniform population, but instead comprise functionally distinct subtypes that differentially support axons and influence their vulnerability to disease. In addition, our findings demonstrate that neuropathic processes are spatially organized along the neuromuscular axis and involve coordinated responses between the peripheral and central nervous systems. The integration of single-cell transcriptomics, spatial transcriptomics, advanced imaging, and genetic models provides a powerful framework to link molecular signatures with structural and functional changes in disease. This approach enables the identification of cellular interactions and mechanisms that were previously inaccessible using conventional methods. Importantly, the discovery that impaired glial support functions can drive axonal degeneration, even in the presence of myelin, challenges existing concepts of demyelinating disease and opens new avenues for therapeutic intervention. Together with the identification of Schwann cell heterogeneity and spatially distinct inflammatory responses, these insights provide a refined understanding of disease mechanisms in peripheral neuropathies. In the future, we expect to further define the causal relationships between local glial dysfunction and system-wide neurodegeneration, and to identify novel targets for therapeutic strategies aimed at preserving axonal integrity and promoting regeneration in neuromuscular disorders.