AuPS Logo Programme
Previous Next PDF

A reduction in Selenoprotein S (SEPS1) amplifies the inflammatory profile of fast twitch skeletal muscle in the mdx dystrophic mouse

G. Keefe,1 A. Addinsall,2 S. Andrikopoulos,3 N. Stupka2 and C. Wright,1 1Centre for Physical Activity and Nutrition (C-PAN), School of Exercise and Nutrition Sciences, Deakin University, Waurn Ponds, VIC 3216, Australia, 2School of Medicine, 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 hypothesized to protect against inflammatory stress. Polymorphisms in the human SEPS1 gene are associated with elevated TNFα, IL-1β and IL-6 gene expression (Curran et al., 2005), and are associated with inflammatory related diseases such as type II diabetes (Walder et al., 2002; Gao et al., 2004) and obesity (Olsson et al., 2011). In several cell culture lines, the siRNA knockdown of SEPS1 upregulates inflammatory cytokines (Kim et al., 2007; Zeng et al., 2008; Fradejas et al., 2011). SEPS1 is highly expressed in human skeletal muscle, however the role of SEPS1 in mediating skeletal muscle inflammation is unknown.

Here we investigated the effects of a reduction in SEPS1 on the inflammatory profile of the mdx dystrophic mouse, a murine model of Duchenne Muscular Dystrophy. Male C57BL6 mice with a global heterozygous deletion of SEPS1 were generated using the Cre-LoxP system, and crossbred with female mdx mice to produce F1 male mdx mice with a heterozygous deletion of SEPS1 (mdx-SEPS1−/+) and mdx male controls. Body composition was measured between six and 12 weeks, after which the EDL and soleus muscles underwent in situ analysis of force, fatigability and force recovery. Briefly, mice were anaesthetized via IP 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 EDL and slow twitch soleus muscles were surgically excised, and muscle force production at increasing stimulation frequency (force frequency curve), endurance and recovery from fatigue were assessed (Aurora Scientific; n = 11-13 mice per group). Anaesthetized mice were then humanely euthanized by cervical dislocation. Following muscle function testing, the EDL and soleus were collected for mRNA and protein expression. All procedures were carried out with full approval from the Deakin University Animal Ethics Committee (AEC#: G29/2014).

Western blotting revealed that global heterozygous deletion of SEPS1 in the mdx mouse caused a robust reduction (51 %) in SEPS1 protein in the fast twitch muscle tibialis anterior (P=0.034), however there were no differences in EDL or soleus muscle mass; therefore a reduction in SEPS1 is probably not affecting skeletal muscle growth. SEPS1 knockdown mice had a 2.4 fold increase in monocyte chemoattractant protein 1 (MCP-1) mRNA (P=0.044), a two-fold increase in macrophage marker F4/80 mRNA (P=0.047) and a trend for elevated transforming growth factor beta 1 (TGF-β1) mRNA in the fast-twitch EDL muscle (P=0.056). Unlike the EDL muscles, mRNA levels of pro-inflammatory cytokines and F4/80 were not altered in the soleus muscles of mdx-SEPS1−/+ mice. This suggests that reduced SEPS1 expression may elevate inflammation in fast-twitch, but not slow twitch muscles of mdx mice. The effects of these changes in gene expression on inflammatory cell infiltration, degeneration and regeneration need to be assessed using histological and immunohistochemical techniques. It should be noted that in EDL and soleus muscles of 12 week old mdx mice, the genetic reduction of SEPS1 had no effects on muscle force, fatigability or recovery in vitro. Further morphometric analyses of skeletal muscle in mdx-SEPS1-/+ mice are required. Therefore, these preliminary data suggest that genetic reduction of SEPS1 in the mdx mouse appears to exacerbate the inflammatory profile of EDL muscles without affecting muscle function.

Curran JE, Jowett JB, Elliott KS, Gao Y, Gluschenko K, Wang J, Abel Azim DM, Cai G, Mahaney MC, Comuzzie AG, Dyer TD, Walder KR, Zimmet P, MacCluer JW, Collier GR, Kissebah AH, Blangero J. (2005). Nature Genet 37, 1234-41.

Fradejas N, Del Carmen Serrano−Pérez ZM, Tranque P, Calvo S. (2011). Glia 59(6), 959-72.

Gao Y, Feng HC, Walder K, Bolton K, Sunderland T, Bishara N, Quick M, Kantham L, Collier GR. (2004). FEBS Lett 563, 185-90.

Kim K-H, Gao Y, Walder K, Collier GR, Skelton J, Kissebah AH. (2007). Biochem Biophys Res Commun 354, 127-32.

Olsson M, Olsson B, Jacobson P, Thelle DS, Björkegren J, Walley A, Froguel P, Carlsson LM, Sjöholm K (2011). Metabolism 60, 114-20.

Walder K, Kantham L, McMillan JS, Trevaskis J, Kerr L, De Silva A, Sunderland T, Godde N, Gao Y, Bishara N, Windmill K, Tenne-Brown J, Augert G, Zimmet PZ, Collier GR. (2002). Diabetes 51, 1859-66.

Zeng J, Du S, Zhou J, Huang K. (2008). Arch Biochem Bioph 478, 1-6.