Journal of Visualized Experiments
Gross contraction in skeletal muscle is primarily determined by a relatively small number of contractile proteins, however this tissue is also remarkably adaptable to environmental factors such as hypertrophy by resistance exercise and atrophy by disuse. It thereby exhibits remodeling and adaptations to stressors (heat, ischemia, heavy metals, etc.). Damage can occur to muscle by a muscle exerting force while lengthening, the so-called eccentric contraction. The contractile proteins can be damaged in such exertions and need to be repaired, degraded and/or resynthesized; these functions are not part of the contractile proteins, but of other much less abundant proteins in the cell. To determine what subset of proteins is involved in the amelioration of this type of damage, a global proteome must be established prior to exercise and then followed subsequent to the exercise to determine the differential protein expression and thereby highlight candidate proteins in the adaptations to damage and its repair. Furthermore, most studies of skeletal muscle have been conducted on the male of the species and hence may not be representative of female muscle. In this article we present a method for extracting proteins reproducibly from male and female muscles, and separating them by two-dimensional gel electrophoresis followed by high resolution digital imaging. This provides a protocol for spots (and subsequently identified proteins) that show a statistically significant (p < 0.05) two-fold increase or decrease, appear or disappear from the control state. These are then excised, digested with trypsin and separated by high-pressure liquid chromatography coupled to a mass spectrometer (LC/MS) for protein identification (LC/MS/MS). This methodology (Figure 1) can be used on many tissues with little to no modification (liver, brain, heart etc.).
2-D gel electrophoresis, Gender, HPLC/MS, Issue 58, Medicine, Mouse, Skeletal muscle
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Dimova, Kalina; Metskas, Lauren Ann; Kulp, Mohini; and Scordilis, Stylianos P., "Skeletal Muscle Gender Dimorphism from Proteomics" (2011). Biological Sciences: Faculty Publications, Smith College, Northampton, MA.