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Microarray gene expression analysis in atrophying rainbow trout muscle: An unique non-mammalian muscle degradation model

ABSTRACT: Muscle atrophy is a physiological response to diverse physiological and pathological conditions that trigger muscle deterioration through distinct cellular stimuli. Despite different physiological signals, the resulting biochemical changes in atrophying muscle share many common cascades. Muscle deterioration as a physiological response to the energetic demands of fish spawning represents a unique model for studying the mechanisms of muscle degradation in non-mammalian animals. A salmonid microarray, containing 16,006 cDNAs, was used to study the transcriptome response to atrophying fast-switch muscles from gravid rainbow trout in comparison to sterile fish. A total of 202 unique transcripts were reproducibly identified as 82 up- and 120 down-regulated genes in atrophying muscles. Transcripts having gene ontology biological process identifiers were grouped according to their functions. Muscle deterioration was associated with elevated expression of genes involved in the catheptic and collagenase proteolytic pathways, the aerobic ATP production, buffering, utilization and growth arrest. Whereas, atrophying muscle showed down-regulation of genes encoding serpin (a serine proteinase inhibitor), enzymes of anaerobic respiration, muscle proteins as well as genes required for RNA and protein biosynthesis/processing. Therefore, gene transcription of atrophying trout muscle changed in a manner similar to mammalian muscle atrophy. These changes result in an arrest of normal cell growth, net protein loss, and a decreased dependence on glycolytic cellular respiration characteristic of fast-switch muscle to aerobic respiratory mechanisms. Other changes/mechanisms unique to the fish model were discussed including genes associated with muscle atrophy reported for the first time. Keywords: muscle atrophy

ORGANISM(S): Salmo salar Oncorhynchus mykiss

PROVIDER: GSE4787 | GEO | 2006/10/04

SECONDARY ACCESSION(S): PRJNA95723

REPOSITORIES: GEO

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Project description:We have constructed a rainbow trout high-density oligonucleotide microarray by using all the available tentative consensus (TC) sequences from the Rainbow Trout Gene Index database (The Computational Biology and Functional Genomics Lab., Dana Farber Cancer Institute and Harvard School of Public Health). The Rainbow Trout Gene Index integrates research data from all available international rainbow trout genomic research projects. The newly designed microarray incorporates 37,394 unique transcript-specific oligonucleotide probes, 60-mer long each. The microarray was printed according to our design by Agilent Technologies using the 4 X 44-design format and contains 1417 Agilent control spots. The performance of the new microarray platform was evaluated by analyzing gene expression associated with the rainbow trout vitellogenesis-induced muscle atrophy. These chips can be ordered from Agilent using design number 016320. This microarray is anticipated to open new avenues of research that will aid in the development of novel strategies to enhance growth efficiency and quality in salmonid species. Keywords: Development of an oligo-array for rainbow trout The performance of the new microarray platform was evaluated by analyzing transcriptome response associated with the rainbow trout vitellogenesis-induced muscle atrophy. Severe muscle deterioration accompanies the physiological responses of the energetic demands of the rainbow trout spawning/vitellogenesis. Atrophying muscle of fertile fish had 11% less extractable muscle (g/bw) and 11% less protein content compared to non-atrophying muscle of sterile fish (p<0.01). The rainbow trout was used to profile changes in gene expression of atrophying muscles. Gene expression levels were determined by comparing the amount of mRNA transcript present in the experimental sample (fertile fish) to the control (sterile fish). RNAs isolated from each experimental fish were run on separate microarrays in independent experiments, with no pooling. A total of 8 fish were used in the microarray experiments (4 replicates x 2 groups). Fluorophors (Cy3 and Cy5) were randomly assigned to RNA from each of the atrophying and nonatrophying muscles to limit the dye effect.

Project description:Training at sustainable swimming speeds can produce changes in fish skeletal muscle that are important for aquaculture due to their growth-potentiating effects. Such changes may be even more relevant when fish are fed diets containing an increasing proportion of carbohydrates as an energy source. We evaluated the effects of moderate-intensity sustained swimming on the transcriptomic response of red and white muscle in rainbow trout fed a carbohydrate-rich diet using microarray and qPCR. Analysis of the red and white muscle transcriptome revealed significant changes in the expression of a large number of genes (395 and 597, respectively), with a total of 218 differentially expressed genes (DEGs) common for both muscles. A large number of the genes involved in glucose use and energy generation, contraction, development, synthesis and catabolism of proteins were up-regulated in red and white muscle. Additionally, DEGs in both muscles were involved in processes of defense response and apoptosis. Skeletal muscle contraction activates a transcriptional program required for the successful adaptation of both muscles to the changing demands imposed by swimming conditions. Future studies should further clarify the mechanisms involved in the adaptation of both tissues to exercise and assess possible benefits of such conditions for cultured fish. Total RNA from pooled red and white skeletal muscle samples of individual rainbow trout from each group (resting fish, n=8; swimming fish, n=8) was labeled with Cy3-dUTP and Cy5-dUTP (GE Healthcare, Barcelona, Spain). We used a dye swap experimental design and each cDNA from a pooled RNA sample was hybridized to two microarrays. For the first slide, cDNAs from resting and swimming fish were labeled with Cy5 and Cy3, respectively, and for the second array dye assignment was reversed. Therefore, samples from individual fish within each group were pooled and expression values shown represent the means of 6 M-CM-^W 2 = 12 technical replicates. A total of four slides were used in this study

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Microarray gene expression analysis in atrophying rainbow trout muscle: An unique non-mammalian muscle degradation model

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