Oncorhynchus mykiss muscle cells: Small vs. Large fish
ABSTRACT: Transcriptional profiling of rainbow trout muscle cells comparing muscle cells from small fish with muscle cells from large fish at two time periods. Two-condition experiment, small vs. large-fish muscle cells. Sept. and Dec. spawning fish. Biological replicates: 4 small replicates, 4 large replicates for each time period.
Project description:Transcriptional profiling of rainbow trout liver cells comparing liver cells from small fish with liver cells from large fish at two time periods. Two-condition experiment, small vs. large-fish liver cells. Sept. and Dec. spawning fish. Biological replicates: 4 small replicates, 4 large replicates for each time period.
Project description:Transcriptional profiling of rainbow trout liver and muscle cells comparing small fish with large fish within a population of neomale offspring. Small vs. large-fish liver and muscle cells from neomale offspring. Biological replicates: 4 small replicates, 4 large replicates.
Project description:This SuperSeries is composed of the SubSeries listed below. Growth in fishes is a complex trait, controlled by both genetic and environmental factors, that impacts many components of fitness. Gene expression studies may lead to the identification of candidate genes for growth and microarrays offer the opportunity to examine the expression of thousands of genes simultaneously. Gene expression differences in the liver and white muscle were examined in normally growing, 15 month-old, large and small size-selected rainbow trout (Oncorhynchus mykiss) derived from two different seasonal spawning groups (Sept. and Dec.). Examination of the gene expression differences in both liver and white muscle tissue allowed us to assess the seasonal influences upon gene expression patterns that occur in this species, and facilitated the identification of genes that may possess similar expression patterns regardless of seasonal effects. The analysis of global gene expression in large and small fish reared under standard conditions provides an understanding of typical growth patterns that may be observed in this species. The identification of candidate genes by this study may provide insight into the mechanisms of growth in fishes and may help to identify candidate genes for growth.
Project description:Although studies have established that exogenous growth hormone (GH) treatment stimulates growth in fish, its effects on target tissue gene expression are not well characterized. We assessed the effects Posilac® (Monsanto Co., St. Louis, MO), a recombinant bovine somatotropin, on tissue transcript levels. Transcript abundance was measure in liver and muscle using the GRASP 16 K cDNA microarray. A selection of the genes identified as altered with the microarray, and also transcripts for insulin-like growth factors, growth hormone receptors (GHR) and myostatins were measured by realtime PCR in the liver, muscle, brain, kidney, intestine, stomach, gill and heart. In general, transcripts identified as differentially regulated in the muscle on the microarray showed similar direction of expression in the other non-hepatic tissues. Rainbow trout were selected from two high growth rate and two low growth rate families. A total of 113 and 67 transcripts were identified by microarray as differentially expressed with GH treatment across growth rate for muscle and liver respectively. The largest proportion of the transcripts represented novel transcripts, followed by immune and metabolism related genes. The immune related genes were primarily modulated in the liver and indicate activation of a non-specific immune response. The metabolic genes include lipid metabolism, oxidative phosphorylation and one carbon metabolism pathway transcripts. Most notable among the growth axis genes measured by realtime PCR were increases in GHR1 and-2 transcript in liver and muscle. Our results indicate that short-term GH treatment activates the immune system, shifts the metabolic sectors and modulates growth regulating genes. Keywords: Growth Hormone Injection Muscle and Liver Gene Expression Rainbow trout (hatched March 2005) selected for extreme growth rate were obtained from NCCCWA brood stock. Families were selected based on body weight at 7 months of age and thermal growth coefficient for the final month of growth. The two high growth families used in the study were in the top 2% in terms of growth rate, and the low growth families were in the lowest 10% for growth rate. Fish acclimated to the new tanks for two weeks prior to initiation of the treatments. Fish from each family were randomly selected to receive one of three treatments: 1) Posilac® injection (120 mg/kg BW, n = 4 per family); 2) vehicle injection (n = 4 per family); or 3) untouched controls (n = 2 per family). We had determined there was no effect to growth or the GH/IGF-I axis in the vehicle treated fish, and therefore, all of the microarray hybridizations were made between the GH and vehicle injected groups. This study included a total of 16 two-channel arrays designed for the direct comparison of GH treatment levels. That is, for each of the four groups, 1) High Growth Rate Liver; 2) Low Growth Rate Liver; 3) High Growth Rate Muscle; and 4) Low Growth Rate Muscle, four slide were hybridizes using individual RNA samples from individual fish. RNA isolation from each tissue/organ sample was handled separately (without pooling) with the purpose of using biological replications. We hybridized two slides with the GH cDNA labeled with Alexa 555 and vehicle cDNA labeled with Alexa 647; and two slides, using unique RNA samples, for the with GH cDNA labeled with Alexa 647 and tissue from the vehicle injected group labeled with Alexa 647 within each tissue and growth rate. Sixteen slides were used in the current study representing 32 individual tissue samples, meaning a total of four biological replicates for each treatment group.
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 × 2 = 12 technical replicates. A total of four slides were used in this study
Project description:The main findings of the current study were that exposing adult sockeye salmon Oncorhynchus nerka to a warm temperature that they regularly encounter during their river migration induced an mRNA-level heat shock response that is exacerbated with swimming. Similar immune defense-related responses were also observed. Microarray analyses revealed that 347 genes were differentially expressed between the cold (12-13° C) and warm (18-19° C) treated fish (P < 0.01), with stress response (GO:0006950; P = 0.014) and response to fungus (GO:0009620; P = 0.003) elevated with warm treatment, while expression for genes involved in oxidative phosphorylation (GO:0006119; P = 0.0019) and electron transport chain (GO:0022900; P = 0.00043) increased in cold-treated fish. By studying single genes with RT-qPCR, warm treatment fish from the Chilko population of O. nerka induced expression of heat shock protein (hsp) 90α, hsp90β and hsp30, as well as interferon-inducible protein (P < 0.05). A Nechako population of O. nerka with a narrower thermal tolerance window than the Chilko population showed even more pronounced responses to the warm treatment. In conclusion, it appears that during their once-in-the-lifetime migration these adult sockeye salmon encounter conditions that induce several cellular defense mechanisms. As river temperatures continue to increase, it remains to be seen whether or not these cellular defenses provide enough protection for all sockeye salmon populations. Two condition experiment; cold treated fish vs. warm treated fish, n=4 in both group
Project description:Treatment with insulin-like growth factor-I (IGF-I) during feed deprivation attenuates the weight loss response in both mammals and fish, therefore the reduction in plasma IGF-I concentrations during fasting likely serves as a signal that contributes to the catabolic response. To better understand the physiological mechanisms responsible for this effect rainbow trout were administered IGF-I during a 2 wk period of feed deprivation and changes in gene expression in white muscle were determined using microarray analysis. Weight loss was reduced by 15% (P<0.05) in IGF-I treated fish. A total of 440 transcripts were identified as differentially regulated (P<0.05) between saline and IGF-I treated fish. Genes related to protein degradation were down-regulated and included protease and peptidase genes and genes involved in ubiquitin-proteasome and cathepsin-mediated proteolytic pathways. IGF-I increased expression of myosin binding protein H and coronin-1C, while decreasing expression of other myofibrillar and cytoskeleton-associated genes like troponin-C and parvalbumin-2. Polyadenylate-binding protein 2, a transcription factor that positively regulates myoD and myogenin expression, was upregulated with IGF-I treatment. Additional genes that were differentially regulated are associated with lipid and carbohydrate metabolism, mitochondrial biogenesis and electron transport, gene transcription, signal transduction and regulation of apoptosis. In summary, these data suggest that IGF-I plays a central role in regulating protein degradation, especially via the ubiquitin-proteasomal pathway, and that reductions in protein degradation and the subsequent effects on other physiological systems are largely responsible for the IGF-I-induced reduction in weight loss. Eight approximately 1-year old full-sibling rainbow trout families were housed in individual tanks, according to family, at the National Center for Cool and Cold Water. Each fish was pit-tagged at 7-months of age. At the beginning of the study, fish were anesthetized with tricaine methanesulfonate (MS-222), weighed, and an osmotic pump was surgically inserted into the peritoneal cavity of each fish (n=4 fish/family/treatment, N=16 fish). The osmotic pump contained either recombinant human IGF-I, which released hormone at 25 ug/kg/day, or saline, which was the vehicle used to resuspend IGF-I. Feed was withheld 2-days prior to surgery and for the experimental 2 week period. Fish were harvested using as overdose of MS-222, weighed, and white muscle samples were removed, frozen in liquid N, and stored at -80 C until analysis. Muscle RNA was isolated from muscle of the two families that exhibited the greatest difference in weight loss between IGF-I and saline treatments; families 64 and 107. Hybridizations were performed using a dye-swap design with an IGF-I and saline treated fish within the same family, therefore a total of 16 slides were hybridized using Alexa 647 and Alexa 555 dyes.