The mechanism by which hypertrophy of skeletal muscle occurs in response to anabolic androgens is incompletely understood. This study used a cell culture model of human skeletal muscle-derived cells, with samples obtained from consenting subjects and local ethics committee approval. The objective was to determine changes in myogenicity upon exposure to exogenous androgen +/− IGF-1 and to see whether an additive effect would be observed. IGF-1 has been shown to accelerate differentiation in human primary cultures and thus acted as a positive control.
Upon growth to confluence, differentiation of muscle-derived cells was induced using 2% fetal calf serum +/− androgen (testosterone 0500 nM or DHT 030 nM) and/or IGF-1 050 ng/ml. Specificity of IGF-1 mediated effects were assessed using a specific IGF-1 receptor antagonist (010 ug/ml). All cultures were performed in triplicate.
After 3 (early) or 7 (late) days differentiation, immunostaining was performed using primary antibodies directed towards the muscle-specific proteins desmin, and α-sarcomeric actin. The DNA-binding probe DAPI was used for nuclear staining. Using fluorescence microscopy, the relative numbers of myogenic (desmin +) vs non-myogenic (desmin −) cells were counted and % desmin+ve myogenicity calculated. Fusion index was obtained by calculating percentages of nuclei incorporated into myotubes.
Increased myogenicity occurred in cells exposed to IGF-1 (P<0.02) in early differentiation. After 7 days differentiation supraphysiological doses of testosterone (+) and IGF-1(+++) both increased fusion index with no observable synergistic effect (P<0.05).
Although androgens may increase muscle hypertrophy by increasing myogenicity of pluripotent muscle-derived cells, this does not appear to occur in the earliest stage of differentiation; mechanisms for increased later fusion need to be further explored. Moreover, this system has potential for investigating the signalling mechanisms associated with the effects of anabolic steroids on skeletal muscle.
06 - 07 Nov 2006
Society for Endocrinology