RABBIT SLOW AND FAST SKELETAL MUSCLE-DERIVED SATELLITE MYOBLAST PHENOTYPES DO NOT INVOLVE CONSTITUTIVE DIFFERENCES IN THE COMPONENTS OF THEINSULIN-LIKE GROWTH-FACTOR SYSTEM

The insulin-like growth factor (IGF) system is actively involved in th e control of proliferation and differentiation of several myogenic cel l lines, and phenotypic differences between myoblasts are associated w ith modifications of the equilibrium of the components of the IGF syst em. To determine whether this observation is a physiologic feature tha t also concerns the phenotypes of ex vivo adult satellite myoblasts in primary cell culture, we investigated the IGF system in rabbit slow-t witch muscle-derived satellite myoblasts (SSM), which differ phenotypi cally from fast-twitch muscle-derived satellite myoblasts (FSM) by the ir proliferation and differentiation kinetics in vitro. The expression of IGF-I and IGF-II were similar in SSM and FSM as well as their conc entrations measured in cell-conditioned media. Ligand blotting of cond itioned media samples indicated the presence of five IGF binding prote in (IGFBP) species of Mr 37-40, 32, 30-31, 28, and 24 kDa. The 30-31 k Da doublet was visible in SSM-conditioned medium only and associated w ith the presence of a 22-kDa protein, which may represent a proteolyti c fragment. In contrast, the 32-kDa band was observed in FSM-condition ed medium only. The other IGFBP moieties were present in both SSM- and FSM-conditioned media. Cross-linking experiments revealed the presenc e of the M6P/IGF-II receptor on both SSM and FSM membranes. We also ob served an IGF-I receptor form bearing unusual high affinity for IGF-II : the binding of [I-125]IGF-I on this receptor was preferentially disp laced by IGF-I but that of [I-125]IGF-II was mostly inhibited by IGF-I I, suggesting that the two tracers did not bind on the same epitopes. [I-125]IGF-II binding to this receptor was greater on SSM than on FSM membranes. Autophosphorylation of WGA-purified receptors revealed an s imilar to 400-kDa band after SDS-PAGE under nonreducing conditions, wh ich corresponded to the alpha 2 beta 2 form of the IGF-I receptor, and two beta subunit moieties of Mr 101 and 105 kDa under reducing condit ions in both SSM and FSM extracts. Phosphorylation of the 105-kDa moie ty was more intensively increased than that of the 101-kDa protein aft er growth factor stimulation. Basal phosphorylation state of the two b eta subunits was similarly stimulated by IGF-I and IGF-II and less by insulin. Since both insulin and IGF-I receptors were expressed in FSM and SSM, one of the two beta subunits may actually correspond to that of the insulin receptor. We conclude that the IGF system is not consid erably affected by the phenotypes of SSM and FSM. The differences obse rved, which mostly concern IGFBP species, more likely appear as regula tory adaptations than as phenotypic changes targeting the components o f the IGF system. (C) 1996 Wiley-Liss, Inc.