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Zebrafish fins: normoxic Controls vs. hypoxic fins

ABSTRACT: Transcriptional profiling of zebrafish fins comparing control fins and hypoxic fins (cobalt chloride (CoCl2, 1 mM)

ORGANISM(S): Danio rerio

PROVIDER: GSE58568 | GEO | 2017/02/09

SECONDARY ACCESSION(S): PRJNA252909

REPOSITORIES: GEO

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Project description:Using the orthotopic breast cancer xenograft model of basal-like breast cancer MDA-MB-231 line, we have found that the expression of miRNAs encoded by MIR17HG was significantly decreased in cells isolated from spontaneous lung metastases compared to cells from primary tumors grown in orthotopic sites. We investigated the role of a MIR17HG family member, miR-18a, in primary tumor growth and pulmonary metastasis from the orthotopic site. We demonstrated that enforced expression of exogenous miR-18a, significantly limited continuous growth of primary tumors in mammary gland fat pads and reduced spontaneous lung metastasis. Further investigation on the mechanism of miR-18a action led to the finding that the expression of HIF1A, a key regulator of tumor metastasis, was regulated by miR-18a. Enforced miR-18a expression significantly decreased HIF1A expression at both mRNA and protein levels, resulting in altered transcriptional response and decreased survival of cells in response to Cobalt(II) chloride (CoCl2), a hypoxia-mimicking agent. Conversely, miR-18a knockdown significantly increased HIF1A expression levels and enhanced cell survival in response to CoCl2. Analysis of expression data of human breast tumor tissues showed that miR-18a expression is inversely correlated with HIF1A expression in basal-like breast tumors, supporting a role of miR-18a in restricting HIF1A expression in this subtype of breast cancer. In addition, we demonstrated that hypoxia inhibits miR-18a expression, likely through MYC inactivation. Furthermore, gene expression and functional analysis revealed that miR-18a also plays a role in regulating cell adhesion, migration and invasion. Taken together, this study provides evidence for a novel role of miR-18a to inhibit breast cancer metastasis. Our results suggest that miR-18a downregulation might provide tumor cells survival/growth advantage under hypoxic pressure in basal-like breast cancer. A lung metastatic subline, designated as MB231RN-LM, was derived from MDA-MB-231 breast cancer cells through in vivo selection. The MB231RN-LM cells were stably transfected with has-miR-18a or control vector and treated with 200uM Cobalt(II) chloride (CoCl2, a hypoxia-mimicking agent) for 4 hr. A total of 8 samples were subjected to microarray analysis, with two biological repeats for each experiment condition.

Project description:Cobalt is a transition group metal present in trace amounts in the human diet, but in larger doses it can be acutely toxic or cause adverse health effects in chronic, long term exposures. Its use in many industrial processes and alloys worldwide presents opportunities for occupational exposures, as well as exposures to military personnel. While the toxic effects of cobalt have been widely studied, the exact mechanisms of toxicity remain unclear. In order to further elucidate these mechanisms and identify potential biomarkers of exposure or effect, we exposed two rat liver-derived cell lines, H4-II-E-C3 and MH1C1, to two concentrations of cobalt chloride. We examined changes in gene expression using DNA microarrays in both cell lines and examined changes in cytoplasmic protein abundance in MH1C1 cells using mass spectrometry. We chose to closely examine differentially expressed genes and proteins changing in abundance in both cells lines in order to remove cell line specific effects. We identified enriched pathways, networks, and biological functions using commercial bioinformatic tools and manual annotation. Many of the genes, proteins, and pathways modulated by exposure to cobalt appear to be due to an induction of a hypoxic-like response and oxidative stress. Genes that may be differentially expressed due to a hypoxic-like response are involved in Hif-1Î± signaling, glycolysis, gluconeogenesis, and other energy metabolism related processes. Gene expression changes linked to oxidative stress are also known to be involved in the NRF2-mediated response, protein degradation, and glutathione production. Using microarray and mass spectrometry analysis, we were able to identify modulated genes and proteins, further elucidate the mechanisms of toxicity of cobalt, and identify biomarkers of exposure and effect in vitro, providing targets for focused in vitro studies. H4-II-E-C3 and MH1C1 cells were treated with two doses of CoCl2 for 24 h in serum free DMEM. For each condition, we ran four biological replicates, with each replicate having its own untreated control. Therefore, this study contains 24 samples.

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Zebrafish fins: normoxic Controls vs. hypoxic fins

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