Endocrine Abstracts

Introduction

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.

Method

Upon growth to confluence, differentiation of muscle-derived cells was induced using 2% fetal calf serum +/− androgen (testosterone 0–500 nM or DHT 0–30 nM) and/or IGF-1 0–50 ng/ml. Specificity of IGF-1 mediated effects were assessed using a specific IGF-1 receptor antagonist (0–10 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.

Results

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).

Conclusions

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.