Oncorhynchus mykiss liver and muscle cells: Small vs. Large neomale offspring
ABSTRACT: 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: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 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: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:Exposure to environmental contaminants like nonylphenol can disrupt smolt development and may be a contributing factor in salmon population declines. We used GRASP 16K cDNA microarrays to identify genes that are differentially expressed in the liver, gill, hypothalamus, pituitary, and olfactory rosettes of Atlantic salmon smolts treated with nonylphenol compared to control smolts. Nonylphenol treatment was confirmed using physiological assays: nonylphenol-treatment significantly decreased gill Na+,K+-ATPase activity and plasma cortisol and T3 levels. Microarray analyses were used to compare expression in nonylphenol-injected fish with expression in vehicle-injected fish: eight arrays each for liver, gill, olfactory rosettes, hypothalamus, and pituitary tissues. Total RNA was isolated from the tissues of eight nonylphenol-injected fish (six males and two females) and eight vehicle-injected fish (two males and six females) and reverse transcribed separately (not pooled); each slide represents a biological replicate. For each tissue, the eight arrays were balanced for dye: nonylphenol-injected fish were labeled with Alexa Fluor 555 and vehicle-injected fish were labeled with Alexa Fluor 647 on four slides, nonylphenol-injected fish were labeled with Alexa Fluor 647 and vehicle-injected fish were labeled with Alexa Fluor 555 on four slides. Liver, gill, hypothalamus, pituitary, and olfactory rosette tissues were analyzed separately.
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:The angio-suppressive effect of 20(R)-ginsenoside Rg3 (Rg3-R) has been previously demonstrated, and microRNAs (miRNAs) are a vital group of small non-coding RNAs that function as post-transcriptional modulator of gene expression. Thus, using human umbilical vein endothelial cells (HUVEC) as model, we compared the microRNA (miRNA) expression profile of vascular endothelial growth factor (VEGF)-induced cells with the profile of the cell co-treated with VEGF and Rg3-R. Among the screened 553 human miRNAs, 6 up-regulated (miR-520h, miR-487b, miR-197, miR-524*, miR-342 and miR-219) and 3 down-regulated (miR-23a, miR-489 and miR-377) miRNAs were detected in Rg3-R treated vascular endothelial growth factor (VEGF)-induced HUVECs compared to VEGF alone. Real time RT-PCR was subsequently performed to verify the miRNA microarray result. Two condition experiment: VEGF-induced HUVEC and VEGF-induced HUVEC treated with Rg3-R. Three independent microarray experiments, with triplicate per microarray.
Project description:To identify genes involved in the developmental process of Atlantic salmon smoltification, gene expression was compared between smolt and parr in tissues involved in osmoregulation (gill), metabolism (liver), imprinting (olfactory rosettes) and neuroendocrine control (hypothalamus and pituitary). Tissue samples were harvested from laboratory-reared parr and smolts on the same date. Smolts were distinguished from parr by size and appearance; developmental status was confirmed by physiological assays. Eight biological replicates (16 fish) balanced for sex and for dye were used in the liver, gill, olfactory rosette, and hypothalamus comparisons. Four male parr were compared to four male smolts and four female parr were compared to four female smolts; smolts were labeled with Alexa Fluor 555 on four arrays and with Alexa Fluor 647 on four arrays. Six biological replicates (12 fish) were used for the pituitary comparison (two female and four male).
Project description:The long-term viability of Pacific salmon stocks and the fisheries they support are threatened if large numbers die prematurely en-route to spawning grounds. Physiological profiles that were correlated with the fate of wild sockeye salmon during river migration were discovered using functional genomics studies on biopsied tissues. Three independent biotelemetry studies tracked the biopsied fish after tagging in the marine environment over 200 km from the Fraser River, in the lower river 69 km from the river mouth and at the spawning grounds. Salmon carrying the poor performance (unhealthy) profile in the ocean exhibited a 4-times lower probability of arriving to spawning grounds than those with a healthy genomic signature, although generally migrated into the river and to the spawning grounds faster. A related unhealthy signature observed in the river was associated with a 30% reduction in survival to spawning grounds in one of the three stocks tested. At spawning grounds, the same poor performance signature was associated with twice the pre-spawning mortality compared with healthy fish. Functional analysis revealed that the unhealthy signature, which intensified during migration to spawning grounds, was consistent with an intracellular pathogenic infection, likely a virus. These results are the first to suggest a pathogen present in salmon in the marine environment could be a major source of mortality during migration and spawning in the river. This series are gill expression profiles from the study of fish at the Weaver creek spawning grounds, and were observed for pre-spawning mortality or successful spawning. Weaver creek sockeye salmon (a late-run stock) spawn in an artificial spawning channel which has a controlled entrance and no exit, situated 100 km from the ocan. Females were dip-netted out of the entrance of the spawning channel, and placed into a sampling trough with flowing ambient water for the gill biopsy procedure and tagged with Petersen discs and returned to the spawning channel. Moribund fish were recovered daily and their gonads examined to assess whether they had spawned or not. Gene expression was assayed on the GRASP salmonid 32K cDNA microarray.
Project description:Infectious diseases among fish present an important economic burden for the aquaculture and fisheries industries around the world. For example, the infectious salmon anemia virus (ISAV) is known to infect farmed Atlantic salmon (Salmo salar), and results in millions of dollars of lost revenue to salmon farmers. Although improved management and husbandry practices over the last few years have minimized the losses and the number of outbreaks, the risk of new virulent isolates emerging is still a looming threat to the viability and sustainability of this industry. An understanding of the host-pathogen interactions at the molecular level during the course of an infection thus remains of strategic importance for the development of molecular tools and efficient vaccines capable of minimizing losses in the eventual case of a new outbreak. Using a 32 k cDNA microarray platform (cGRASP), we have studied various signaling pathways and immune regulated genes, activated or repressed, in Atlantic salmon head-kidney during the course of an ISAV infection. Gene expressions were measured at 5 different time-points: 6h, 24h, 3d, 7d and 16d post infection to get an overall view of changes as they occurred in time. The earliest time points showed only a few differentially expressed genes in infected fish, relative to controls, although as time progressed, many additional genes involved in key defense pathways were up-regulated including MHC type I, beta-2 microglobulin, TRIM 25 and CC-chemokine 19. During the latest stage of the infection process, many genes related to oxygen transportation were under-expressed, which correlates well with the anemia observed prior to death in Atlantic salmon infected with virulent strains of ISAV. Atlantic salmon smolts from 2 families of Atlantic salmon were IP injected with either 0.1mL of 10e5 TCID50 mL-1 of virus or 0.1mL of sham solution (L15 culture medium) and divided equally in four 1000 L tanks: 2 duplicate tanks containing ISAV injected fish and 2 duplicate control tanks containing sham solution injected fish. Four fish per family were sampled immediately prior to injection. An additional two fish per family per tank (four fish per family total) were sampled at 6h, 24h, 3d, 7d and 16d post injection. Head-kidney was dissected from each fish and used for microarray analysis. ISAV infected Atlantic salmon were compared to non-infected Atlantic salmon for each time-point.
Project description:The ability to interrogate circulating tumor cells (CTC) and disseminated tumor cells (DTC) is restricted by the small number detected and isolated (typically <10). We wanted to determine if a commercially available technology could provide a transcriptomic profile of a single prostate cancer (PCa) cell. We clonally selected and cultured a single passage of cell cycle synchronized C4-2B PCa cells. Ten sets of single, 5-, or 10-cells were isolated using a micromanipulator under direct visualization with an inverted microscope. Additionally, we analyzed 10 individual DTC isolated from each of 2 patients with metastatic PCa. We have shown that a transcriptomic profile can be obtained from a single cell using commercially available technology. As expected, fewer amplified genes are detected from a single-cell sample than from pooled cell samples, but this method can be used to obtain a transcriptomic profile from DTC isolated from the bone marrow of patients with PCa. Custom Agilent 44K whole human genome expression oligonucleotide microarrays were used to profile clonally selected and cultured single passage of cell cycle synchronized C4-2B PCa cells isolated using a micromanipulator under direct visualization with an inverted microscope into ten sets of single, 5-, or 10-cells. Single disseminated tumor cells were isolated from bone marrow (BM) samples of two advanced prostate cancer patients. Essentially, a two-step selection process was employed, in which anti-CD45 and anti-CD61 conjugated to immunomagnetic beads were used for negative selection, and anti-HEA was used for positive selection. Cells were then fluorescently stained for BerEP4, counter stained with RPE anti-CD45, and individually selected (10 single cells each per patient) under fluorescent light using a micropipette system for further analysis. RNA was amplified using the WT-Oviation one-direct system and hybridization against a common reference pool of prostate tumor cell lines. Data from C42B cell data and data from single cells isolated from the bone marrow of patients were normalized and analyzed separately.