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Novel role for Selenoprotein S in regulating skeletal muscle contractile function in fast twitch mouse muscles

A. Addinsall,1 C.-H. Weng,3 N.L. McRae,1 C. Wright,2 S. Andrikopoulos,3 N. Wong3 and N. Stupka,1 1School of Medicine, Deakin University, Waurn Ponds, VIC 3216, Australia, 2School of Exercise and Nutrition Sciences, Deakin University, Waurn Ponds, VIC 3216, Australia and 3Department of Medicine - Austin Health, The University of Melbourne, Heidelberg VIC 3084, Australia.

Selenoprotein S (SEPS1) is an antioxidant protein with oxioreductase activity against H2O2. In humans, polymorphisms in the SEPS1 promoter have been linked to diseases with a heightened pro-inflammatory state. SEPS1 is also protective against endoplasmic reticulum (ER) stress by facilitating the translocation of misfolded proteins out of the ER lumen for degradation in the cytosol. SEPS1 is highly expressed in skeletal muscle, yet its physiological function in vivo is poorly characterised. In cultured C2C12 myoblasts, SEPS1 gene suppression using siRNA increased H2O2 levels and was associated with a more oxidised cellular redox state, as indicated by the oxidised to reduced glutathione (GSH:GSSG) ratio.

  Skeletal muscle contractile function and metabolism are sensitive to oxidative, ER and inflammatory stress, and our aim was to characterise the effects of reduced SEPS1 expression on whole body metabolism and skeletal muscle function. Male global SEPS1 deleted mice were generated by PGK-Cre and fed a standard chow diet. The animal studies were approved by the Animal Ethics Committees at the University of Melbourne and Deakin University, in accordance with NH&MRC guidelines. At ∼12 weeks of age, mice were placed in metabolic cages for 25 h (Metabolic Analyser, Columbus instruments) to determine VO2, VCO2 and the respiratory exchange rate ratio (RER). The physical activity of the mice was measured during this time (Animal Activity Meter: Opto-Varimex-Mini, Columbus instruments). Prior to contractile function testing, mouse body composition was assessed (ESF-005, EchoMRI), and then mice were anaesthetized via intraperitoneal injection of medetomidine (0.6 mg/kg), midazolam (5 mg/kg) and fentanyl (0.05 mg/kg) such that they were unresponsive to tactile stimuli. Fast twitch extensor digitorum longus (EDL) and slow twitch soleus muscles were surgically excised and muscle force production, endurance and recovery from fatigue were assessed in vitro (1300A Whole Mouse Test System, Aurora Scientific) (Stupka et al., 2008). Following functional testing, anaesthetised mice were humanely euthanized by cervical dislocation and the heart and hindlimb muscles were collected for molecular and histological analysis.

Here, we present our preliminary findings on SEPS1+/− mice and wild type littermates (N = 5-6). Hindlimb muscles, specifically the fast twitch tibialis anterior (TA) muscle, of SEPS1+/− had a 61% reduction in SEPS1 mRNA transcripts (P=0.04). Body weight, lean mean mass and fat mass were similar between SEPS1+/− and wild type mice. Physical activity, as assessed by ambulatory movement was 32% lower in SEPS1+/− mice (P=0.02). Although, no difference in VO2 and VCO2 was observed. SEPS1 heterozygosity had no effect on EDL or soleus muscle mass, twitch force (Pt), maximal force (Po) and specific force (sPo). To assess muscle endurance and force recovery, EDL and soleus muscles were stimulated submaximally at 60 Hz every 5 s for 4 min, and then again at 2 min, 5 min and 10 min post fatigue. During 4 min of contractile activity, EDL muscles from the SEPS1+/− mice fatigued more than those from wild type littermates (P=0.001; main effect – general linear model ANOVA). Furthermore, recovery from fatigue was ∼30% worse in EDL muscles from SEPS1+/− mice (P=0.006; main effect – general linear model ANOVA). Whereas in soleus muscles, SEPS1 heterozygosity did not compromise endurance and recovery from fatigue. In fast twitch muscles recovery of force following submaximal contractile activity is modulated by cellular redox state (Lamb & Westerblad, 2011). Thus, reduced SEPS1 expression in EDL muscles may increase cellular stress in response to contractile activity, and this could contribute to the decrease in physical activity observed in the SEPS1+/− mice. In conclusion, these preliminary data suggest a new role for SEPS1 in modulating skeletal muscle contractile function.

Stupka N, Schertzer JD, Bassel-Duby R, Olson EN & Lynch GS. (2008) American Journal of Physiology. Regulatory. Integrative and Comparative Physiology, 294: R983-R992.

Lamb GD & Westerblad H. (2011) The Journal of Physiology, 589: 2119-2127.