Unknown,Transcriptomics,Genomics,Proteomics

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Zebrafish embryos: exposure to 3,4-dichloraniline, 2,4-dichlorophenol, pentachlorophenol, and cadmium chloride

ABSTRACT: Large-scale toxicogenomic screening approaches offer great promise for generating a bias-free, system-wide view of toxicological effects and modes-of-action of chemicals and ecotoxicants. Early applications of microarray technology have identified relatively small groups of responding genes with which to define new targets for analysis by conventional means. We have trialled a more intensive approach to the design and interpretation of array experiments incorporating a balanced interwoven ANOVA design with higher levels of biological replication, a more thorough analysis of errors and false discovery rates, and an analysis of response patterns using gene networks. Zebrafish embryos were exposed to ecotoxicants representing different classes - 2,4-dichlorophenol (DCP), 3,4-dichloroaniline (DCA), pentachlorophenol (PCP), and cadmium chloride - at low concentrations producing a developmental disturbance to 10% of embryos, and half of this dose. Extracted RNA was then analysed on microarrays. Analysis revealed responses of 3000–5000 genes, which is 10–1000 times greater than previously reported, with significance at lower levels of fold-change. Some gene responses were common to multiple toxicants and others that were restricted to just one or two toxicants. The gene expression profiles for the different toxicants were distinctive, and analysis using network-based packages provides a high level of detail of affected processes, some of which were novel. This approach provides a more highly refined view of toxic effects, from which meaningful patterns of response can be discerned and related to functional deficits, and from which more reliable indicators of toxicological effect can be predicted. Interwoven loop design consisting of 12 separate conditions - two exposure concentrations plus control for each of the four toxicants - using six replicates for each conditions, each array was hybridised with independent replicates from each of two conditions resulting in 72 samples on 36 arrays

ORGANISM(S): Danio rerio

SUBMITTER: Ashley Sawle

PROVIDER: E-GEOD-21680 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:With the aim of shedding light on the protection conferred by the DNA vaccines based in the G glycoprotein of viral haemorrhagic septicaemia virus (VHSV) in turbot (Scophthalmus maximus) we have used a specific microarray highly enriched in antiviral sequences to carry out the transcriptomic study associated to VHSV DNA vaccination/infection. The differential gene expression pattern in response to empty plasmid (pMCV1.4) and DNA vaccine (pMCV1.4-G860) intramuscular administration with regard to non-stimulated turbot was analyzed in head kidney at 8, 24 and 72 hours post-vaccination. Moreover, the effect of VHSV infection one month after immunization was also analyzed in vaccinated and non-vaccinated fish at the same time points. A total number of 204 juvenile turbot were divided into 3 groups, two of them containing 72 fish and the last one 60 fish. Turbot were anaesthetized by immersion in 50 mg/ml buffered tricaine methanesulfonate (MS-222; Sigma) and then, fish from the first two groups were intramuscularly (i.m.) injected with 50 µl of PBS containing 2 µg of pMCV1.4 or pMCV1.4-G860. Turbot from the last batch were i.m. inoculated with 50 µl of PBS. At 8, 24 and 72 h after injection, 12 fish were removed from the first two tanks and, at 8 h after PBS inoculation, other 12 fish were taken from the last tank. These turbot were sacrificed by anaesthetic overdose and the head kidney was removed. Equal amounts of tissue from three fish belonging to the same tank and sampling point were pooled, obtaining 4 biological replicates for each treatment and time point (3 turbot/replicate). The remaining fish (36 in the plasmid-injected groups and 48 in the PBS-inoculated tank) were maintained during one month and then, 12 fish from the PBS injected group were separated to another tank. This new group of fish was intraperitoneally (i.p.) injected with 50 µl of MEM + penicillin and streptomycin + 2% FBS (PBS - MEM group), whereas the other turbot were i.p. infected with a dose of VHSV860 of 5 x 105 TCID50/fish (pMCV1.4 - VHSV and pMCV1.4-G860 - VHSV groups). At 8, 24 and 72 hours after infection, 12 fish were removed from the VHSV-infected tanks, and at 8 h after MEM injection the 12 fish were taken from the non-infected tank. The fish were sacrificed by anaesthetic overdose and the head kidney was removed. Equal amounts of tissue from three fish belonging to the same tank and sampling point were pooled, obtaining 4 biological replicates for treatment and time point (3 turbot/replicate)

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Zebrafish embryos: exposure to 3,4-dichloraniline, 2,4-dichlorophenol, pentachlorophenol, and cadmium chloride

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