Dataset Information

Hypoxia induced sex-difference in zebrafish brain proteome profile reveals the crucial role of H3K9me3 in recovery from acute hypoxia

ABSTRACT: 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.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Danio Rerio (zebrafish) (brachydanio Rerio)

TISSUE(S): Brain

DISEASE(S): Mixed Disorder As Reaction To Stress,Myocardial Ischemia

SUBMITTER: Tapatee Das

LAB HEAD: Tapatee Das

PROVIDER: PXD027528 | Pride | 2022-02-22

REPOSITORIES: Pride

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			Action	DRS
	hypoxia_ZF1_250316-03.msf	Msf
	hypoxia_ZF1_250316.mzML	Mzml
	hypoxia_ZF1_250316.raw	Raw

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Hypoxia Induced Sex-Difference in Zebrafish Brain Proteome Profile Reveals the Crucial Role of H3K9me3 in Recovery From Acute Hypoxia.

Das Tapatee T Kamle Avijeet A Kumar Arvind A Chakravarty Sumana S

Frontiers in genetics 20220131

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 well-controlled, non-invasive, reproducible conditions using a zebrafish model. Our earlier report on sex difference in global acute hypoxia-induced neural damage and recovery in zebrafish prompted us to conduct a comprehensive study on ...[more]

PMID: 35173759

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Project description:Adult zebrafish (TÃ¼bingen 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 Â± 0.6 mg/l, whereas normal ambient oxygen levels were 6.6 Â± 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??80Â°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 Â°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.