Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative disorder associated in 20% of the familial cases with mutations in the gene coding for the antioxidant enzyme superoxide dismutase (SOD1). ALS is a multi-systemic and multifactorial disease and recent evidences suggest that skeletal muscle is a primary target in ALS. Indeed muscle specific expression of SOD1 mutant gene in MLC/SOD1G93A transgenic mice reveals muscle atrophy, mitochondrial alterations and neuromuscular junction (NMJ) instability. A critical role in this molecular mechanism is played by Protein Kinase C theta (PKC¿), which is chronically activated in MLC/SOD1G93A mice. PKC¿ pharmacological inhibition restores mitochondrial functionality and neuromuscular damaged phenotype, sustaining the "dying back" hypothesis. Nevertheless whether PKC¿ inhibition can counteract muscle functional decline associated with the disease progression remains unclear, so it might be important to analyze the effects of PKC¿ pharmacological inhibition and/or genetic ablation in SOD1G93A mice.
Metabolic changes associated with muscle espression of SOD1G93A in the MLC/SOD1G93A mice occur independently of motor neuron degeneration, so we'll investigate glucose and lipid metabolism and muscle fiber composition in the mice model of PKC¿ inhibition. It is known that ALS is also related to hypothalamus atrophy, so we'll evaluate markers of hypothalamus atrophy in all mice models above mentioned and metabolites levels involved in melanocortin system. Moreover the superchiasmatic nucleus, that is the central circadian clock, is located in hypothalamus anterior region, so it could be interesting to study circadianicity to evaluate whether the circadian alterations occur independently of motor neuron degeneration and to verify whether NMJ stabilization, due to PKC¿ inhibition, could have effects on hypothalamus defects.
This research project proposes to deeply investigate some unclear questions about molecular mechanisms of ALS pathogenesis and find out triggers of the pathology.
The use of the mouse model SOD1G93A with PKC¿ pharmacological inhibition and the genetic ablation could let us to study and develop our research according to an investigational tool and a therapeutic tool.
In particular, genetic ablation of PKC¿ in PKC¿-/- SOD1G93A could help us to investigate PKC¿ molecular mechanism of activity during the development in the NMJ dismantlement. Whereas pharmacological treatment at different ages, could help us to elucidate the effects of PKC¿ inhibition in SOD1G93A mice both in a presymptomatic phase and at the onset, to evaluate whether the inhibition is able to improve or not disease progression. This could let us to understand whether the pharmacological inhibition in ALS mouse model is able to restore the neuromuscular damaged phenotype, like in the MLC/SOD1G93A mice, and to evaluate the effects of the treatmeant at two different ages, before symptoms or at the onset of the disease.
ALS patients and ALS mouse models are characterized by hypermetabolism condition, so thanks to these mice model with PKC¿ inhibition, we could evaluate whether the inhibition of its activity is also able to restore metabolic alterations. PKC¿ is also involved in muscle fiber metabolism and it determines a slow-twitch phenotype, so we could evaluate the effects of PKC¿ inhibition on the expression of markers of slow muscle fiber program and also on the myofibers composition.
The final goal will be to verify whether in these mice PKC¿-/- SOD1G93A and PKC¿ treated, the NMJ stabilization, due to PKC¿ inhibition, could have effects on hypothalamus and restore defects in melanocortin system neurons, at the levels of metabolites acting on hypothalamus and also in circadianicity of these mice.
Moreover whether this pharmacological treatment in SOD1G93A mice will have positive effects on mitochondrial morphology and functionality, on metabolic profile and on neuromuscular altered phenotype, we could hypothesize a translation of the "dying back" theory into a "saving back" process proposing a therapeutic tool for ALS patients.