Isolation and culture of wapiti (Cervus elaphus) satellite cells

Myogenic satellite cells (SC) were isolated from the sternomandibularis mus cles of two, 227 kg, male wapiti (Cervus Elaphus) and studied in primary ce ll culture. Wapiti-derived SC were capable of attaching to culture substrat a and following the myogenic program of proliferation and differentiation t o form multinucleated myotubes. Wapiti SC attached equally well to pig skin gelatin (PSG), fibronectin (FN), Matrigel(R) and plastic (P > 00.05), but cell viability measured at 120 h varied depending on initial substratum typ e. Pig skin gelatin (0.02% wt vol(-1)) was chosen for the majority of subse quent experimentation for cost efficiency. The greatest amount of wapiti SC proliferation was observed in media containing 10% (vol/vol(-1)) horse ser um (HS), 15% HS and 15% fetal bovine serum (FBS) (P > 0.05). Wapiti SC prol iferated more when exposed to HS and FBS than to sheep serum (SS) (P < 0.05 ). No proliferation, differentiation or decrease in cell viability was obse rved in Dulbecco's Modified Eagle Medium (DMEM) + 1% HS, DMEM + 2% HS, DMEM + 3% HS or DMEM + 4% HS (P > 0.05) after 120 h in vitro. Proliferation of SC was doubled when insulin was added to both 10% HS- and 2% MS-containing media (P < 0.05). Although insulin alone in serum-containing media did not promote fusion of wapiti SC, two defined media (ITT and ITT-CF) that contai n insulin did promote fusion of wapiti SC cultures. ITT-CF induced 3% fusio n of wapiti SC into myotubes, and ITT induced 1% (P < 0.05). There was also an increase in total cell numbers in SC exposed to ITT-CF in comparison wi th ITT, ovine defined media (ODM) or ovine defined media-Modified (ODM-Mod) )(P < 0.05). Although defined media differed in their ability to induce pro liferation or differentiation (P < 0.05), the substrata on which the SC wer e plated did not influence the defined media effect on SC activity (P > 0.0 5). Satellite cells exposed to ITT and ITT-CF differed morphologically from SC exposed to ODM and ODM-Mod, which may suggest that formulation differen ces are influencing wapiti-derived SC proliferation and differentiation.