Gene expression profiling in equine muscle tissues using mouse cDNA microarrays

Progress could be achieved by using microarrays to understand metabolic adaptations and disorders in equine muscle in response to exercise.

To test the feasibility of using mouse cDNA microarrays to analyse gene expression profile in normal equine muscles.

Muscular biopsies of dorsal gluteus medius and longissimus lumborum were done in 4 healthy Standardbreds. Total RNA was extracted from the muscle samples. The concentration and quality of RNA were measured before and after amplification. Gene expression profiles were measured using mouse cDNA microarrays including 15,264 unique genes representing about 11,000 documented genes. Three hybridisation tests were performed to check interspecificity, reproducibility and to compare gene expression in these muscles. For each test, a dye-swap hybridisation with Cy3 and Cy5 fluoromarkers were done and the gene list filtered according the signal level.

According to the specificity test, the mouse cDNA microarrays were correctly hybridised by equine muscle cDNA. All positive control genes (GAPDH, HPRT and beta-Actin) and no negative control gene (yeast, plant) hybridised. The reproducibility test demonstrated a good linearity between the duplicate hybridisations: 99.99% of the significant expressed genes have an expression ratio between 1.4 and 1/1.4 = 0.71. These limits can be considered as the thresholds to qualify as up-regulated (ratio >1.4) or downregulated (ratio <0.71). In the muscle comparison test between gluteus medius vs. longissimus lumborum, 63 genes were found up-regulated and 8 genes down-regulated. The range of gene expression ratios in the gluteus medius was 0.61-8.31 x the longissimus lumborum. This list of modulated genes was classified by functions using a gene ontology data basis.

Mouse microarrays could be used to hybridise equine RNA extracted from muscle tissues. For many genes there are large sequence identities that allowed interspecific cDNA hybridisation. The sensitivity of the method allowed quantification of up- and down-regulated genes after applying appropriate filters.

Expression profiling could be used to explore the muscle metabolism changes related to exercise, training, pathology and illegal medication in horses.