Skeletal Muscle Gender Dimorphism from Proteomics

Abstract

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 1 such as hypertrophy by resistance exercise and atrophy by disuse. It thereby exhibits remodeling and adaptations to stressors (heat, ischemia, heavy metals, etc.) 2,3 . Damage can occur to muscle by a muscle exerting force while lengthening, the so-called eccentric contraction 4 . 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 5 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 6 . 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) 5 . This methodology (Figure 1) can be used on many tissues with little to no modification (liver, brain, heart etc.).

Gender specific differences in the liver proteome of rats exposed to short term and low-concentration hexabromocyclododecane (HBCD)

“…Interestingly, the observed gender-specifically regulated pathways are also the ones affected by HBCD exposure in a gender specific way in the present (and previous) study. Also other reports have shown differences in gender-susceptibility (in rat serum proteins in health and inflammation, 25 , 41 in mouse proteins in muscle 42 or in the aging kidney 43 ). Our results are thus in line with a recent NIH directive/recommendation for gender-balance in further biomedical research studies.…”

Dietary fish oil supplement induces age-specific contractile and proteomic responses in muscles of male rats

“…Protein was twice-precipitated with acetone from the homogenates and 800 μg protein were suspended in urea-CHAPS buffer (8 M urea, 50 mM DTT, 4% CHAPS, 0.2% pH 5-8 carrier ampholytes, 0.0002% Bromophenol Blue) and loaded onto 11-cm, pH 5-8 IPG strips (Bio-Rad Labs) and focused 40,000 V-Hrs [34]. After focusing, immobilized pH gradient (IPG) strips were equilibrated in a denaturing SDS buffer (6 M urea, 2% SDS, 0.05 M Tris/HCl, 20% glycerol) and alkylated, embedded onto 11-cm 10.5-14% Tris-HCl gels (Bio-Rad Labs), and electrophoresed at 120 V [35].…”

Proteomics as a tool to gain more insight into sub-lethal toxicological effects

“…Also other reports have shown differences in gender-susceptibility (in rat serum proteins in health and inflammation , in mouse proteins in muscle (Dimova et al, 2011) or in the aging kidney (Amelina and Cristobal, 2009)). This approach is in line with a recent NIH directive/recommendation for gender-balance in further biomedical research studies .…”