Project description:Brain transcriptome at 0h, 1h, 3h, 6h, 12h, 24h, 48h after hypoxia stress Large yellow croakers (body weight at 90-100 g) were purchased from the mariculture farm in Ningde, Fuzhou, China. The fish were maintained at 25 °C in aerated water tanks (dissolved oxygen concentration: 7.8±0.5 mg per liter) with a flow through seawater supply. After 7 days of acclimation, these fish were used for the following experiments. Hypoxic time-course experiments were conducted at 25 °C using published method 30, by bubbling nitrogen gas into an aquarium. The desired pO2 was controlled by using dissolved oxygen meter (, Canada). At the onset of the time course, the oxygen content of the tank was lowered from an aerated pO2 of 100% (7.8 mg per liter) down to 20% (1.6±0.2 mg per liter) over a 30-min period. At the 1-, 3-, 6-, 12-, 24-, and 48-h time points, fish were sampled and sequenced.
Project description:Understanding the molecular basis of sex differences in neural response to acute hypoxic insult has profound implications for the effective prevention and treatment of ischemic stroke. Global hypoxic-ischemic induced neural damage has been studied recently under the well-controlled, non-invasive, reproducible conditions using zebrafish model. Our earlier report on sex difference in global acute hypoxia induced neural damage and recovery in zebrafish prompted us for comprehensive study on the mechanisms underlying the recovery. An omics approach for studying quantitative changes in brain proteome upon hypoxia insult following recovery was undertaken using iTRAQ-based LC-MS/MS approach. The results shed light on altered expression of many regulatory proteins in zebrafish brain upon acute hypoxia following recovery. The sex difference in differentially expressed proteins along with the proteins expressed in uniform direction in both the sexes was studied. Core expression analysis by Ingenuity Pathway analysis (IPA) showed a distinct sex difference in the disease function heatmap. Most of the upstream regulators obtained through IPA were validated at the transcriptional level. Translational upregulation of H3K9me3 in male led us to elucidate the mechanism of recovery by confirming transcriptional targets through ChIP-qPCR. The upregulation of H3K9me3 level in male at 4 hr post-hypoxia appears to affect the early neurogenic markers nestin, klf4 and sox2, which might explain the late recovery in male, compared to female. Acute hypoxia-induced sex-specific comparison of brain proteome led us to reveal many differentially expressed proteins, which can be further studied for the development of novel targets for better therapeutic strategy.
Project description:Hypoxia is an important environmental stressor in aquatic ecosystems, with increasingly impacts on global biodiversity. Yellow catfish is an economically important farmed fish in China, which has increased dramatically. We investigated the response of hybrid yellow catfish to hypoxia under experimental conditions and focused on the analysis of the differential expression patterns of specific genes associated with hypoxia response by RNA-seq and qPCR analysis. A total of 1556 genes were captured significantly differentially expressed, and were categorized into immune response and energy metabolism. Functional enrichment analysis revealed the NLR signaling pathway play pivotal roles in hypoxia tolerance and resistance. Our study provides important insights into the physiological acclimation, immune response and defense activity of hybrid yellow catfish under hypoxia challenge.
Project description:Adult zebrafish (Tübingen strain, sex not specified) at approximately 1 year of age were analysed. For experiments conducted under low oxygen conditions, nitrogen gas was bubbled through water to deplete oxygen before exposure of individual fish to the medium. Oxygen concentrations were measured using a dissolved oxygen meter (DO 6+, EUTECH instruments, Singapore). The dissolved oxygen level for hypoxia treatment was measured to be 1.20 ± 0.6 mg/l, whereas normal ambient oxygen levels were 6.6 ± 0.45 mg/l. Zebrafish were exposed to the hypoxic medium for 3 hours. Briefly, after each hypoxia trial, the animals were euthanized by hypothermic shock and then decapitated to remove the brain. Total RNA was extracted from samples mentioned above using the QIAGEN RNeasy mini kit (QIAGEN, GmbH, Hilden, Germany) and stored at â??80°C before further analysis. RNA concentration was determined with a NanoVueâ?¢ UVâ??vis spectrophotometer (GE Healthcare Life Sciences, Fairfield, USA). RNA integrity and quality were then estimated using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA) and the RNA integrity number (RIN) index was calculated for each sample. Only RNAs with a RIN number >7.0 were processed further. Microarray analysis of gene expression was performed using the Zebrafish Gene 1.0 ST Array (Affymetrix Inc. Santa Clara, CA). Briefly, 300ng of total RNA derived from a single adult brain was converted to amplified sense strand cDNA using the Ambion WT Expression Kit (Life Technologies, Carlsbad, CA). The resulting sense cDNA was fragmented and Biotin end-labelled using the Affymetrix Genechip WT Terminal Labeling Kit prior to hybridisation to the array at 45 °C for 16 hours. Two treatments including hypoxia and normoxia were studied. Each treatment had three biological replicates (i.e. three fish exposed to hypoxia and three fish exposed to normoxia). Six samples were analysed. Microarray analysis of gene expression was performed using the Zebrafish Gene 1.0 ST Array. The signal intensity of the chip was scanned using a GeneChipR Scanner 3000TG and analysed using Expression Console software (www. Affymetrix.com). CEL files were imported and intensities adjusted by RMA background correction and quantile normalization.
Project description:We investigated the transition from juvenile yellow to the adult sexually maturing, migrating silver eel (Anguilla anguilla) by examining the hypothesis that: The brain is the central organ for the co-ordination of environmental cues (day length, photoperiod, temperature and environmental salinity) with the anatomical and physiological adaptations which accompany pre-migrational morphogenesis and the osmoregulatory plasticity seen in post-migrational, salinity-adapted fish. We have characertised the mRNA expression profiles for the brains of fresh water, yellow and silver eel using a highly representative brain cDNA microarray. The array comprises 5760 cDNA clones from A.anguilla ranging from 0.5 -10 kb and an estimated redundancy of > 5 %.
Project description:Since normal brain function depends upon continuous oxygen delivery and short periods of hypoxia can precondition against subsequent ischemia, this study examined the effects of brief hypoxia on the whole genome transcriptional response in adult mouse brain. Genomic expression profiling was perfromed for individual brain regions of the adult mice following the entire time course of hypoxia preconditioning. Adult C57BL/6 male mice were exposed to systemic preconditioning hypoxia (8% O2 ) for 3 hr and allowed to recover in normoxia for 24 hr. The mouse brains were removed and dissected into individual brain regions at multiple time points during the 3hr hypoxia and subsequent 24hr reoxygenation periods. Total RNA was purified from the human whole blood or individual mouse brain regions. Genomic scale gene expression was then measured with Affymetrix Mouse Expression 430 2.0 arrays.
Project description:Since normal brain function depends upon continuous oxygen delivery and short periods of hypoxia can precondition against subsequent ischemia, this study examined the effects of brief hypoxia on the whole genome transcriptional response in adult mouse brain. Genomic expression profiling was perfromed for individual brain regions of the adult mice following the entire time course of hypoxia preconditioning.