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Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of mouse decrement in Norrin/Frizzled4/Lrp signaling in retinal endothelial cells

ABSTRACT: To characterize the long-term effect on the transcriptome of a decrement in Norrin/Fz4/Lrp signaling, microarray hybridization was performed with RNA from acutely dissociated and anti-PECAM immunoaffinity-purified adult WT, Fz4-/-, Lrp5-/-, and Norrin- retinal vascular cells. Experiment Overall Design: Retinal endothelial cells from 5-10 adult mice were pooled for each sample, the cell immuno-purification procedure follows that described by Matsubara et al. (2000) and Su et al. (2003). 3 replicates of each mutant and 6 replicates of control wild type animals were analyzed.

ORGANISM(S): Mus musculus

SUBMITTER: Xin Ye

PROVIDER: E-GEOD-16703 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Long-term effect on the transcriptome of a decrement in Norrin/Frizzled4/Lrp signaling in retinal endothelial cells

Project description:To characterize the long-term effect on the transcriptome of a decrement in Norrin/Fz4/Lrp signaling, microarray hybridization was performed with RNA from acutely dissociated and anti-PECAM immunoaffinity-purified adult WT, Fz4-/-, Lrp5-/-, and Norrin- retinal vascular cells.

2009-10-01 | GSE16703 | GEO

Long-term effect on the transcriptome of loss of Frizzled4 signaling in cerebellar endothelial cells

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Transcription profiling of mouse Frizzled4 signaling in cultured retinal endothelial cells

Project description:Transcriptional profiles of Fz4-/- retinal endothelial cells were compared to that of wild type endothelial cells under various culture conditions. The goal was to identify the transcriptional response to Frizzled 4 signaling in cultured retinal endothelial cells. To analyze the Norrin response of WT and Fz4-/- retinal endothelial cells in culture, we co-cultured these cells either with HEK293 cell line that stably expresses Norrin or with control 293 cells. Experiment Overall Design: To isolate the requisite retinal endothelial cell lines, we crossed Fz4-/- and control animals into the Immorto-mouse line in which a gamma interferon responsive H2Kb promoter directs the expression of a temperature sensitive SV40 large T-antigen (Jat et al., 1991). When cultured at 33°C in the presence of gamma interferon, this conditional oncogene system extends the proliferative capacity of a variety of cell types, which can be subsequently analyzed under phenotypically reverting conditions (37°C without gamma interferon). The cell immuno-purification procedure follows that described by Matsubara et al. (2000) and Su et al. (2003). The purified cells were then cultured and passaged at 33°C on dishes coated with 1% gelatin. For RNA preparation, cells were cultured at 37°C in the absence of interferon gamma. For the Matrigel experiments, endothelial cells previously starved for 24 hrs in DMEM with 1% FBS were trypsinized and plated on the Matrigel layer. The cells were incubated in DMEM with 1% FBS for 6 hours at 37°C before harvesting for RNA extraction. For the 293 cell co-culture experiments, endothelial cells were co-cultured with a human embryonic kidney (HEK) 293 cell line that stably expresses Norrin, or with control 293 cells.

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