Abstract
Ca2+ is a vital molecule involved in many cellular processes that occur within skeletal muscle fibres, including influencing the cells oxidative state. The primary contributor of reactive oxygen species within myofibres is NADPH oxidase enzymes. Cross-talk between reactive oxygen species and Ca2+ is carefully maintained within myofibres to prevent the pathophysiology seen in disease, such as Duchenne Muscular Dystrophy. The hypothesis for this thesis was that there are reciprocal changes in Ca2+-handling protein expression with Nox enzymes that differ with fibre-types, particularly in the mouse model of Duchenne Muscular Dystrophy.
The skeletal muscle plasma membrane invaginates to form t-tubules which comes into proximity with the internal store of Ca2+, the sarcoplasmic reticulum, forming the major signalling region called the triad. Within the triad are the proteins responsible for excitation-contraction coupling, Ca2+-handling and redox homeostasis. Fast and slow-twitch skeletal muscle fibre types differ in their Ca2+ handling capabilities due to differences in expression of these proteins and can also differ in reactive oxygen species handling. Nox isoforms, NADPH oxidase-2 and NADPH oxidase-4, have been shown to display a reciprocal amplification with Ca2+ and this reciprocal interaction has been linked to disease pathology. In Duchenne Muscular Dystrophy, myofibres have increased oxidative stress and fast-twitch fibres preferentially degenerate in the progression of the disease. The project aimed to examine the total expression of these Ca2+ handling proteins and reciprocal changes in Nox enzymes in fast, extensor digitorum longus and slow-twitch soleus muscle from healthy wild-type and dystrophic mice via western blot. Western blot revealed that no Ca2+-handling proteins differed in expression between wild-type soleus and dystrophic soleus, but calsequestrin-1, both isoforms of stromal interaction molecular-1, and the redox protein NADPH oxidase-2 were increased in the dystrophic extensor digitorum longus compared to wild-type extensor digitorum longus. Interestingly there was a significant difference in expression of calsequestrin-1 and dihydropyridine receptor in the dystrophic extensor digitorum longus compared to the dystrophic soleus.