Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptome analysis of CD11b+ myeloid cells isolated from the infarction area of Reg3beta knock-out Reg3beta-/- vs. WT mice

ABSTRACT: Adult wild-type (C57/Bl6J) and mice with constitutive knock-out of regenerating-islet derived protein 3 beta (Reg3beta-/-) at the age of 8-12 weeks were subjected to experimental myocardial infarction (LAD ligation). After 4 days, infarcted tissue was dissected, minced with scissors and enzymatically digested with liberase blendzyme (Roche, 0.15 mg/ml) to obtain a single-cell suspension. Isolated cells were washed and resuspended in PBS buffer supplemented with 0.5 % BSA and 2 mM EDTA and passed through 30 um pre-separation filters. The myeloid cells were isolated using the mouse/human CD11b Microbead Kit (Miltenyi Biotec, 130-049-061) and RNA was isolated using the TRIzol method (Invitrogen).

ORGANISM(S): Mus musculus

SUBMITTER: Thomas Boettger

PROVIDER: E-MTAB-3000 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:We examined the transformation susceptibility of different cerebellar stem/progenitors by developing several new Group3 medulloblastoma murine models using orthotopic transplantation and in utero electroporation (EP)-based in vivo gene transfer with Cre/LoxP-mediated conditional Myc gene activation and loss of Trp53 function. We used microarrays to compared the transcriptome of these novel Group3 medulloblastoma mouse models to existing mouse models of medulloblastoma subgroups and used cross-species analysis to compare these models to human medulloblastoma subgroups This study aimed to investigate the cell of origin of Group3 MB using our orthotopical MYC model followed by a novel electroporation approach. Orthotopic cell-lineage specific MYC tumors were generated by enforced Myc expression in P6 GNPs isolated from P0-1 tamoxifen treated [Atoh1-CreER;Trp53fl/-] and [Prom1-CreER;Trp53fl/-] mice followed by cortical implants in immunocompromised mice. These tumors are referred to as Atoh1ER-MYC [dka072-075], Prom1-CreER [dka077-081]. The first Group3 MB models in which tumors developed in situ were generated by electroporation of plasmids containing Myc and dominant negative Trp53 flanked by loxP sites into the fourth ventricle of E13.5 Blbp-Cre [dka081, 087, 089, 090, 091 and blm121], Gad2-IRES-Cre [blm128-130 and blm134] and Ptf1a-Cre [blm135-137] mouse embryos. The gene expression profile of these tumors were compared to our previously published Group3 MB model as well as SHH and WNT models of medulloblastoma. For SHH subgroup medulloblastoma, [dka001-005, 009, 033 and 034] and [dka050-057], spontaneous medulloblastomas from [Cdkn2c-/-; Trp53Fl/Fl; Nestin-Cre] and [Cdkn2c-/-; Ptch1+/-] (Uziel et al.,2005 Genes Dev) were used, respectively. For Group3 medulloblastomas, [dka013-16, 049 and 065-067], in which Myc was overexpressed in Cdkn2c-/-, Trp53-/- cerebellar cells and implanted into the cortices of immunocompromised nude mice prior to tumor isolation. For WNT subgroup medulloblastomas [pgr003, 016 and 066], spontaneously developed tumors from CTNNB1+/lox (ex3); BLBP-Cre; Trp53Fl/Fl (Gibson et al., 2010, Nature) were removed for RNA extraction.

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Transcriptome analysis of CD11b+ myeloid cells isolated from the infarction area of Reg3beta knock-out Reg3beta-/- vs. WT mice

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