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A distinct and unique M2 transcriptional program expressed by tumor-associated macrophages

ABSTRACT: Description This series contains microarrays for tumor associated macrophages (TAMs) purified from 8 week old male C57/BL6N mice, 3 weeks after tumor implantation into the left hind limb. Peritoneal exudates cells (PECs) were also harvested from control C57/BL6N mice. The resulting adherent populations were maintained in culture and resting TAMs and PECs were treated for 4 hours with 100 ng/mL of LPS to promote macrophage activation. RNA isolation and cDNA microarrays: Total RNA was isolated from approximately 20x10e6 PEC and TAM cells using standard Trizol purification protocols (Invitrogen, Carlsbad, CA). Microarrays were manufactured at the NCI, Laboratory of Molecular Technology (Frederick MD). The 10,368 Mm UniGem2 set was printed on poly-lysine coated glass slides and hybridized according to NCI facility protocols. All slides were scanned using as Axon GenePix 4000 scanner (Axon Instruments, Foster city, CA) and resulting data was uploaded for analysis to the mAdb microarray database (http://madb.nci.nih.gov/). Keywords = tumor associated macrophage Keywords: other

ORGANISM(S): Mus musculus

PROVIDER: GSE2098 | GEO | 2005/01/31

SECONDARY ACCESSION(S): PRJNA91793

REPOSITORIES: GEO

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A distinct and unique M2 transcriptional program expressed by tumor-associated macrophages

Project description:Description This series contains microarrays for tumor associated macrophages (TAMs) purified from 8 week old male C57/BL6N mice, 3 weeks after tumor implantation into the left hind limb. Peritoneal exudates cells (PECs) were also harvested from control C57/BL6N mice. The resulting adherent populations were maintained in culture and resting TAMs and PECs were treated for 4 hours with 100 ng/mL of LPS to promote macrophage activation. RNA isolation and cDNA microarrays: Total RNA was isolated from approximately 20x10e6 PEC and TAM cells using standard Trizol purification protocols (Invitrogen, Carlsbad, CA). Microarrays were manufactured at the NCI, Laboratory of Molecular Technology (Frederick MD). The 10,368 Mm UniGem2 set was printed on poly-lysine coated glass slides and hybridized according to NCI facility protocols. All slides were scanned using as Axon GenePix 4000 scanner (Axon Instruments, Foster city, CA) and resulting data was uploaded for analysis to the mAdb microarray database (http://madb.nci.nih.gov/). Keywords = tumor associated macrophage

2010-06-10 | E-GEOD-2098 | biostudies-arrayexpress

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Project description:To test the difference genes expression of TAMs obtained from orthotopically implanted hepa1-6 tumors in C57BL/6 mice.Isolated TAMs were obtained from orthotopically implanted hepa1-6 tumors in C57BL/6 mice. Specimens were minced with scissors and digested by incubation for 1 h at 37°C in high-glucose DMEM (Life Technologies, CA) containing 0.1% collagenase IV (Sigma). After being washed in medium plus 10% FBS, the cell suspension was forced through a graded series of meshes to obtain single-cell suspensions and centrifuged at 400 g for 30–40 minutes at 18–20°C on layers of Lympholyte-M (Cedarlane Labs, Burlington, Ontario, Canada), after which the upper layer was removed, thus leaving the lymphocyte layer undisturbed at the interface. TAMs were sorted using Fluorescence-activated cell sorting (FACS, Becton Dickinson, Franklin Lakes, NJ) with CD45+, CD11b+, and F4/80+ (Becton Dickinson) according to the protocol listed at http://www.immgen.org/Protocols/ImmGen%20Cell%20prep%20and%20sorting%20SOP.pdf . TAMs growing in DMEM supplemented with 10% FBS were tested to determine their biological characteristics. We used microarrays to detail the global programme of gene expression and identified vehicle-TAM and treated-TAM during this process.

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