Conditionally immortalised equine skeletal muscle cell lines for analysis.

BACKGROUND

Thoroughbred racehorse performance is largely influenced by a major quantitative trait locus at the () gene which determines aptitude for certain race distances due to a promoter region insertion mutation influencing functional phenotypes in skeletal muscle. To develop an system for functional experiments we established three novel equine skeletal muscle cell lines reflecting the variation in phenotype associated with genotype (CC/II, CT/IN and TT/NN for SNP g.66493737C > T/SINE insertion 227 bp polymorphism). Primary equine skeletal muscle myoblasts, isolated from Thoroughbred horse , were conditionally immortalised and evaluated to determine whether cell phenotype and metabolic function were comparable to functional characteristics previously reported for skeletal muscle isolated from Thoroughbred horses with each genotype.

RESULTS

Primary myoblasts conditionally immortalised with the temperature sensitive SV40TtsA58 lentivirus vector successfully proliferated and could revert to their primary cell phenotype and differentiate into multinucleated myotubes. Skeletal muscle fibre type, gene expression, mitochondrial abundance, and mitochondrial function of the three genotype cell lines, were consistent with equivalent characterisation of skeletal muscle samples with these genotypes. Furthermore, addition of coenzyme Q (CoQ) to the cell lines improved mitochondrial function, an observation consistent with skeletal muscle samples with these genotypes following supplementation with CoQ in the diet.

CONCLUSIONS

The observation that the phenotypic characteristics and metabolic function of the cells lines are equivalent to skeletal muscle indicates that this system will enable efficient and cost-effective analyses of equine skeletal muscle for a range of different applications including understanding metabolic function, testing of nutritional supplements, drug test development and gene doping test development. In the multi-billion-euro international Thoroughbred horse industry research advances in the biological function of skeletal muscle are likely to have considerable impact. Furthermore, this novel genotype-specific system may be adapted and applied to human biomedicine to improve understanding of the effects of myostatin in human physiology and medicine.