Aprotinin, a Kunitz-type protease inhibitor, stimulates skeletal muscle differentiation

Aprotinin, a Kunitz-type inhibitor of serine proteases, stimulates myotube formation by mouse G8-1 and C2C12 skeletal muscle myoblasts. This stimulation of morphological differentiation is accompanied by accumulation of myogenin transcripts and production of muscle-specific proteins. In contrast, active TGF beta prevents differentiation of G8-1 and C2C12 myoblasts. When active TGF beta and aprotinin are both added to myoblast cultures, differentiation is inhibited, suggesting the active growth factor acts downstream of the protease inhibitor. TGF beta is found in serum as a latent, dimeric propolypeptide that is cleaved by limited proteolysis to release the biologically active carboxy-terminal dimer. To address the possibility that aprotinin may effect myogenesis by inhibiting proteolytic activation of latent TGF beta, levels of the endogenous growth factor were measured in differentiating myoblast cultures. Latent TGF beta is rapidly depleted from control cultures within 24 hours of plating, but remains relatively stable in aprotinin-treated cultures. Consistent with this, aprotinin-treated cultures have reduced levels of active TGF beta. These data indicate that Kunitz-domain containing protease inhibitors may help orchestrate the onset of myogenesis, possibly by regulating the activity of TGF beta-like molecules.