Duchenne muscular dystrophy (DMD) is an X-linked muscular disorder characterized by the lack of dystrophin (Dp427), a cytoskeletal protein expressed by skeletal muscles and, at lower levels, by other cells, including neurons of both central and peripheral nervous systems. Cells other than muscles may also express shorter dystrophin isoforms. Clinically, DMD is characterized by progressive muscular degeneration, which results in premature death; however, DMD patients may also exhibit neurological and autonomic disorders of various degrees. Neural alterations are established during embryonic development, suggesting a prominent role for dystrophins in nervous system differentiation. In adults, Dp427 is mainly localized at specific post-synaptic sites, where it contributes to the stabilization of neurotransmitter receptors subtypes (i.e. GABA). Oligondendrocytes and Schwann cells (SCs) also express Dp427, along with Dp140 and Dp116 isoforms, respectively. Both types of glial cells are responsible for axon myelination and intense glia-neuron cross-talk, crucial for axon growth and activity. SCs are also important for axon regeneration and maintenance of neuromuscular junctions that motor neurons (MNs) establish on skeletal muscles. To date, it is unknown whether MNs are directly affected by DMD. This project focuses on aspects of MNs survival, differentiation and activity, using adult and 5 day-old (P5) wild-type and mdx mice, a DMD animal model. At this aim, expression and localization of specific myelin proteins, as well as histological/morphometric parameters will be evaluated in the sciatic nerve. Expression and localization of GABA receptor, important in axon-SC crosstalk, will be also analyzed. This is particularly intriguing when considering that Dp427 is involved in brain GABAA receptor stabilization and activity. Comprehend whether DMD is responsible for MN functional alteration is a crucial aspect for the clinical research aiming at muscle regeneration.
This project will take into considerations aspects related to the DMD so far not investigated. Specifically, we will focus on the neural part of the disease, e.g. the large MNs, which synapse onto skeletal muscles, and their myelinating SCs. The importance of this issue stems from the definition itself of the pathology, considered a neuromuscular disease. Nevertheless, and with a good reason, most of the studies are focused on the muscular aspect, which is what inevitably leads to early death of the young patients. Loss of muscles fibers, however, implies that those MNs, which were making synapses on them, are also affected, as evinced by the disorganization of the NMJ. But the question remains on whether MNs are affected by the lack of Dp427 independently from muscles, whether their development and process of myelination is altered compared to normal individuals, and whether their axonal regenerative capabilities are preserved or reduced. Unaltered regenerative properties are essential for the correct reinnervation of muscle fibers, and recovery of damaged muscle fibers is the focus of the clinical research on DMD. In this year of project a few important information will be collected on both MN and SC, and on the cross talk they establish during axonal growth and physiological remodeling. The animal used will be adult (6-7- weeks) and early post-natal (P5) dystrophic mdx mice and their corresponding wild type. At P5, MN are still maturing and periaxonal myelinating SCs are completing their differentiation, while muscle fibers are still not heavily damaged by the lack of Dp427, a process that occurs by 6-7 weeks. This date is, therefore, ad hoc to study MN axon and myelinating SCs in their differentiation process. The information acquired will be fundamental ground for future studies more focused on the capability of MNs to react to peripheral axonal damage, regenerate their axons and establish new NMJs on recovered muscle fibers.
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