Regulation of signaling pathways downstream of IGF-I/insulin by androgen in skeletal muscle of glucocorticoid-treated rats

Abstract

Background: The mechanisms by which androgens ameliorate glucocorticoid-induced muscle wasting are still under investigation. In the present study, we tested the hypothesis that androgen's effects in reversing muscle wasting are related to activating the signaling pathways downstream of insulin-like growth factor-1 (IGF-I)/insulin.

Methods: Forty female Sprague-Dawley rats were randomly divided into four groups: control group, dexamethasone (DEX) group, testosterone (TES) group, and TES + DEX group. Each group was injected with saline or DEX (0.1 mg/100 g/d) for 10 days and sesame oil or TES (0.5 mg/100 g/d) for 13 days. Several downstream targets of IGF-I/insulin in skeletal muscle including protein kinase B (Akt), p70 ribosomal protein S6 kinase (p70S6K), and glycogen synthase kinase-3beta (GSK-3beta) that are associated with protein synthesis were examined. Two proteolysis-related ubiquitin E3-ligases, muscle atrophy F-box, and muscle RING finger-1 that are also regulated by IGF-I/insulin were also assessed.

Results: TES attenuated gastrocnemius muscle atrophy induced by DEX. TES prevented the DEX-induced decrease of IGF-I expression in gastrocnemius muscle, but not in serum. TES ameliorated DEX-induced dephosphorylation of Akt and p70S6K and promoted the phosphorylation of GSK-3beta in gastrocnemius muscle. The total amount of Akt, p70S6K, or GSK-3beta proteins was not changed among these groups. TES did not show any effects on the DEX-induced upregulation of muscle atrophy F-box, and muscle RING finger-1 mRNA in gastrocnemius muscle.

Conclusion: This findings suggest that the effects of TES in reversing DEX-induced muscle atrophy are related to signaling pathways downstream of IGF-I/insulin that are associated with protein synthesis.

Fig. 1

Effects of DEX and TES treatment on serum TES concentration. Serum testosterone was analyzed by Radioimmuno-assay. * P<0.05 vs. CON group, ##P<0.01 vs. DEX group. N=10.

Fig. 2

Effects of dexamethasone (DEX) and testosterone (TES) treatment on body weights. **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. N=10 per group.

Fig. 3

Effects of DEX and TES treatment on gastrocnemius muscle weight and muscle fiber cross-sectional area (CSA). *P<0.05 and **P<0.01 vs. CON group, #P<0.05 and ##P<0.01 vs. DEX group. N=10 per group.

Fig. 4

Effects of DEX and TES treatment on IGF-I mRNA expression in skeletal muscle (N=6 per group) and serum IGF-I concentration (N=10 per group). IGF-I mRNA expression in gastrocnemius muscle was analyzed by real-time PCR. Serum IGF-I was analyzed by ELISA. **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; A.U., arbitrary units.

Fig. 5

Effects of DEX and TES treatment on phosphorylation of Akt in skeletal muscle. Top: Akt protein expression in gastrocnemius muscle was analyzed by Western blot. Bottom: Akt protein was quantified by optical densitometry. **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. N=5 per group.

Fig. 6

Effects of DEX and TES treatment on phosphorylation of GSK-3β in skeletal muscle. Top: GSK-3β protein expression in gastrocnemius muscle was analyzed by Western blot. Bottom: GSK-3β protein was quantified by optical densitometry. **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. N=5 per group.

Fig. 7

Effects of DEX and TES treatment on phosphorylation of p70S6K in skeletal muscle. Top: p70S6K protein expression in gastrocnemius muscle was analyzed by Western blot. Bottom: p70S6K protein was quantified by optical densitometry. **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. N=5 per group.

Fig. 8

Effects of DEX and TES treatment on MAFbx and MuRF1 mRNA expression in skeletal muscle. mRNA expression in gastrocnemius muscle was analyzed by real-time PCR. *P<0.05 and **P<0.01 vs. CON group, ##P<0.01 vs. DEX group. N=6 per group. GAPDH, glyceraldehyde-3-phosphate dehydrogenase; A.U., arbitrary units.