Pilot studies for the treatment of sarcopenia and myotonic dystrophy: a multimodal approach

Sarcopenia is a condition of age-related muscle atrophy representing a risk factor for frailty, loss of independence and physical disability in the elderly. Myotonic dystrophy (DM) is an autosomal dominant disorder characterized by muscle dystrophy and dysfunction. Sarcopenia and DM share common features such as reduction in muscle fiber size, centrally located and/or clumped nuclei, defective satellite cells. Moreover, in both sarcopenia and DM, ribonucleoproteins accumulate in nuclei of muscle fibers and satellite cells thereby hampering transcription/maturation processes and causing a reduction in the regenerative capacity of the muscle tissue. In this work, two strategies have been explored to counteract these two pathologies namely, adapted physical exercise (considered a non-pharmacological approach) and nanotechnology (considered a tool for a pharmacological approach); five types of nanoparticles (NPs), i.e. chitosan NPs, Mn-based phospholipidic NPs, liposomes, cyanoacrylate-based NPs and mesoporous silica NPs (MSN-NPs) were investigated as carriers for the delivery of molecules able to reduce nuclear RNA aggregates. Observations at light- and transmission electron microscopy have shown that, in old mice (28 months), adapted physical exercise is able to partially restore the conditions of the skeletal muscle typical of adulthood (12 months). All NPs, tested in vitro on human and murine cell lines, proved to be biocompatible. To pave the way for valid administration protocols, the cellular mechanisms of NP internalization, their intracellular fate and their relationships with cell organelles have been investigated by fluorescence (conventional and confocal) microscopy and transmission electron microscopy Chitosan NPs proved to be efficient carriers for the tested molecule, without inducing cell alterations; however, they showed low internalization rate, maybe because of a tendency to aggregate. They persisted for very long time inside the cell and were able to enter the nucleus, thus constituting a potential risk. Mn-based NPs acted as good drug carrier systems and constitute a valid alternative to gadolinium as contrast agent for nuclear magnetic resonance; they efficiently entered the cells, probably by fusion with the plasma membrane, and remained for about 24 hours in the cytoplasm before degradation in the lysosomes. Liposomes demonstrated an excellent ability to enter the cell, probably by fusion with the plasma membrane (similar to Mn-based NPs), then they were quickly degraded; however, the cells showed some structural alterations, probably due to an excess of uptake. Cyanoacrylate-based NPs and MSN-NPs were internalized by endocytosis and no damage of cell structures was found. Cyanoacrylate-NPs underwent massive degradation within 24 hours, whereas MSN-NPs remained unaltered in the cells, although confined within vacuoles. The results obtained in this work provide a solid experimental basis for future studies on the use of physical exercise and nanotechnology for the development of new therapeutic approaches for sarcopenia and DM.