Simvastatin represses protein synthesis in the muscle-derived C2C12 cell line with a concomitant reduction in eukaryotic initiation factor 2B expression

Statins are a widely prescribed class of cholesterol lowering drugs whose use is frequently associated with muscle-related ailments. A number of mechanisms have been implicated in statin-induced myotoxicity including alterations in both protein synthesis and protein degradation. The objective of the present study was to explore the mechanism(s) contributing to the statin-induced reduction in protein synthesis in the muscle-derived C2C12 cell line. Cells were treated with 10 μM simvastatin or vehicle alone for 24 h in 1% serum. Cells exposed to simvastatin exhibited reduced rates of protein synthesis, as evidenced by [35S]methionine and [35S]cysteine incorporation into protein. The reduction in protein synthesis occurred with a concomitant decrease in expression and activity of eukaryotic initiation factor 2B (eIF2B), a regulated and rate-controlling guanine nucleotide exchange factor known to affect global rates of protein synthesis. The reductions in protein synthesis and eIF2B expression were prevented by coincubation with mevalonate. Simvastatin treatment also resulted in a proteasome-sensitive reduction in the protein expression of all the subunits of the eIF2B heteropentameric complex. Finally, increased phosphorylation of the catalytic ϵ-subunit at Ser535 was observed, an event consistent with an observed reduction in eIF2B activity. These results suggest that repression of eIF2B expression and activity may contribute, at least in part, to the statin-induced reduction in protein synthesis

Ectopic expression of eIF2Bε in rat skeletal muscle rescues the sepsis-induced reduction in guanine nucleotide exchange activity and protein synthesis

Eukaryotic initiation factor 2B (eIF2B) is a guanine nucleotide exchange factor (GEF) whose activity is both tightly regulated and rate-controlling with regard to global rates of protein synthesis. Skeletal muscle eIF2B activity and expression of its catalytic ε-subunit (eIF2Bε) have been implicated as potential contributors to the altered rates of protein synthesis in a number of physiological conditions and experimental models. The objective of this study was to directly examine the effects of exogenously expressed eIF2Bε in vivo on GEF activity and protein synthetic rates in rat skeletal muscle. A plasmid encoding FLAG-eIF2Bε was transfected into the tibialis anterior (TA) of one leg, while the contralateral TA received a control plasmid. Ectopic expression of eIF2Bε resulted in increased GEF activity in TA homogenates of healthy rats, demonstrating that the expressed protein was catalytically active. In an effort to restore a deficit in eIF2B activity, we utilized an established model of chronic sepsis in which skeletal muscle eIF2B activity is known to be impaired. Ectopic expression of eIF2Bε in the TA rescued the sepsis-induced deficit in GEF activity and muscle protein synthesis. The results demonstrate that modulation of eIF2Bε expression may be sufficient to correct deficits in skeletal muscle protein synthesis associated with sepsis and other muscle-wasting conditions

Minimally Invasive Sampling of Transdermal Body Fluid for the Purpose of Measuring Insulin-Like Growth Factor-I During Exercise Training Military Metabolic Monitoring

ABSTRACT Insulin-like growth factor-I (IGF-I) is a ubiquitous hormone that is secreted in both an endocrine and an autocrine/paracrine manner. IGF-I has conventionally been measured in serum; however, transdermal body fluid (TDF) remains as an unexplored biocompartment in which IGF-I also resides and may be more biologically relevant because of its proximity to tissues and cells. The purpose of this study was to compare IGF-I in serum versus IGF-I in TDF before and after 8 weeks of physical training. Twenty-eight healthy men (28 Ϯ 5 years old, 176 Ϯ 8 cm tall, weighing 83 Ϯ 11 kg) had TDF obtained by a novel, minimally invasive method that included the application of continuous vacuum pressure on forearm skin perforated with tiny micropores created by a focused beam from a laser system and also had blood obtained by venipuncture. An enzyme-linked immunosorbent assay measured total IGF-I concentrations. A repeated-measures analysis of variance (biocompartment ϫ time) and Pearson Product Moment Correlation coefficients (P Յ 0.05) were used for statistical analyses. Data are presented as mean Ϯ SE. Total TDF IGF-I was significantly lower than serum IGF-I both before (TDF, 91 Ϯ 6 ng/mL; serum, 375 Ϯ 17 ng/mL) and after (TDF, 83 Ϯ 5 ng/mL; serum, 363 Ϯ 19 ng/mL) the exercise training. Serum and TDF IGF-I values were not significantly different pre-to post-training. Serum and TDF IGF-I levels were significantly correlated pre-training (r ϭ 0.41), but not post-training (r ϭ 0.34). The percent change between serum and TDF was not correlated (r ϭ 0.09). This study has demonstrated that total IGF-I can be sampled and measured in TDF via a minimally invasive manner and is appreciably (ϳ76%) less than total IGF-I measured in serum. Additionally, the IGF-I measurements in these two biocompartments were not closely associated, possibly indicating an uncoupled, rather than a linked, regulation of IGF-I among the body's biocompartments