MicroRNA and Gene expression profiles in colorectal cancer stromal tissue
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE35602 | GEO | 2012/03/14
SECONDARY ACCESSION(S): PRJNA152525
REPOSITORIES: GEO
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Project description:Using Laser Microdissection System (Leica Microsystems), RNA samples specific for stroma or epithelium were separately collected from thirteen colorectal cancer tissues and four normal tissues. Both microRNA microarray and gene expression microarray were performed in these samples. Tissue samples from thirteen colorectal cancer tissues and four normal tissues were microdissected using Laser Microdissection System (Leica Microsystems), and RNA samples specific for stroma or epithelium were separately collected.MicroRNA and Gene expression profiles obtained from cancer stromal tissues were compared with that of normal colon stromal tissues. we used the commercially available Human Whole Genome Oligo DNA Microarray Kit (Agilent Technologies) for gene expresson analysis and microRNA array for microRNA analysis.
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Project description:Tissues from another series of 132 patients with colorectal cancer were collected by laser micro-dissection with the Leica Laser Microdissection System (Leica Microsystems).
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Project description:Tissues from another series of 74 patients with colorectal cancer were collected by laser micro-dissection with the Leica Laser Microdissection System (Leica Microsystems).
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Project description:Tissues from another series of 132 patients with colorectal cancer were collected by laser micro-dissection with the Leica Laser Microdissection System (Leica Microsystems). Gene expression levels obtained from 132 microdissected colorectal tumors were compared with microdissected normal colon epitheliums. we used the commercially available Human Whole Genome Oligo DNA Microarray Kit (Agilent Technologies). Labeled cRNAs were fragmented and hybridized to an oligonucleotide microarray (Whole Human Genome 4×44K Agilent G4112F). Fluorescence intensities were determined with an Agilent DNA Microarray Scanner and were analyzed using G2567AA Feature Extraction Software Version A.7.5.1 (Agilent Technologies).
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Project description:Tissues from another series of 74 patients with colorectal cancer were collected by laser micro-dissection with the Leica Laser Microdissection System (Leica Microsystems). Gene expression levels obtained from 74 microdissected colorectal tumors were compared with 5 microdissected normal colon epitheliums. We used the commercially available Human Whole Genome Oligo DNA Microarray Kit (Agilent Technologies). Labeled cRNAs were fragmented and hybridized to an oligonucleotide microarray (Whole Human Genome 4×44K Agilent G4112F). Fluorescence intensities were determined with an Agilent DNA Microarray Scanner and were analyzed using G2567AA Feature Extraction Software Version A.7.5.1 (Agilent Technologies).
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Project description:Compared to what is known in model species, reproductive biology in citrus is still poorly understood. Although in recent years several efforts have been made to study pollen-pistil interaction and self-incompatibility, little information is available about the molecular mechanisms regulating these processes. We performed microarray analysis for the identification of candidate genes involved in pollen-pistil interaction and self-incompatibility in clementine (Citrus clementina Hort. ex Tan.). The analysis was performed comparing the transcriptome of laser-microdissected stylar canal cells isolated from two clementine genotypes differing for self-incompatibility response (‘Comune’, self-incompatible; and ‘Monreal’, a self compatible mutation of ‘Comune’). Styles with stigmas, collected 24 hours after self pollination, were immediately snap-frozen in OCT embedding medium (Sakura Finetek, Zoeterwoude, Netherlands) in Peel-A-Way plastic embedding molds (Polysciences, Polysciences, Warrington, PA, USA). Transversal sections 10 µm thick at the upper part of the style were cut with a Leica CM1900 cryostat (Leica Microsystems, Germany) at -20°C. A Leica AS Laser Microdissection system (Leica Microsystems) was used for the isolation of stylar canals from transversal sections. Canals from the stigma were discarded to avoid contamination with pollen or pollen tubes. Three biological replicates were prepared for each genotype. Each biological replicate consisted of bulks of about 200 microdissected areas (composed of an average of 50 cells) coming from two different molds
2012-02-15 | E-GEOD-33014 | biostudies-arrayexpress
Project description:We explored the hypothesis that maternal cocaine exposure could alter the fetal epigenetic machinery sufficiently to cause lasting neurochemical and functional changes in the offspring. These data were used as preliminary results for paper:; Maternal cocaine administration in mice alters DNA methylation and gene expression in hippocampal neurons of neonatal and prepubertal offspring. Svetlana I. Novikova, Fang He, Jie Bai, Nicholas J. Cutrufello, Ashiwel S. Undieh and Michael S. Lidow Experiment Overall Design: Pregnant CD1 mice were administered either saline or 20 mg/kg cocaine twice daily on gestational days 8-19. Male pups from each of ten litters of the cocaine and control groups were analyzed at 3 (P3) day postnatum. Pyramidal layer tissue from the right hemisphere of one of these brains (On every section, the pyramidal layer, encompassing the C1, C2 and C3 hippocampal regions, was carefully cut out)was used for analyzing gene expression. Laser microdissection was performed on a Leica ASLMD laser capture microdissection system (Leica Microsystems, Wetzlar, Germany).
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Project description:The mammalian lens consists of an aged core of quiescent cells enveloped by a layers of mature fully elongated and younger, continuously elongating and transcriptionaly active cells. In the present study, we compared global gene expression profiles in elongating and mature, non-elongating fiber cells. The microarray-based comparison of gene expression profiles was performed using Agilent 22K Mouse Oligo microarrays with two rounds of RNA amplification from samples obtained by laser capture microdissection (LCM) of paraffin-embedded lens sections. Lenses were removed and immediately fixed using 4% paraformaldehide or methanol-based UMFIX reagent (Sakura Finetek USA, Inc.). To map the exact location of both elongating and maturing fibers on the lens slices, lenses were vibratome-sliced (Vibratome 1000, St. Louis, MO) as described previously and the GFP expression pattern was captured by confocal microscopy. For RNA extraction lens tissue was sliced into 5 micron-thick paraffin sections and microdissected using LCM (Leica Microsystems, Bannockburn, IL). The mid-saggital slices were used both for syncytium border measurements and for LCM. Control measurement (data not shown) confirmed that similar rates of shrinkage in paraformaldehyde- and UMFIX-fixed samples did not affect precision of LCM dissection. Fiber cell samples were dissected out of 5 micron-thick slices. In one experiment we typically processed about 40 slices, which was sufficient to collect the minimum of 200 zone-specific cells pooled from P5 littermate lenses. This sample size provided a reliable representation of RNA species in experimental procedure originally designed and tested for just 1-10 cells. Localization of the syncytium border characterized by the abrupt change of GFP labeling pattern was captured by confocal microscopy as described previously. In brief, GFP fluorescence was visualized using LSM510 instrument (Carl Zeiss, NY) equipped with an argon/krypton laser at 488 nm excitation and a 515-565 nm band pass emission filter. Physical parameters of the zones containing young and maturing fibers were measured in fixed lens slices using the morhphometric software provided by Zeiss. Lenses were fixed in 4% paraformaldehyde/PBS and sectioned with a vibratome. Cells from elongating and maturing fiber regions were dissected out using the Leica DMLA laser capture microscope, (Leica Microsystems, Bannockburn, IL). The cut-out pieces containing captured cells were put directly into tubes containing the lysis buffer supplied in the Absolutely RNA® nanoprep kit (Stratagene, La Jolla, CA). Total RNA from the microdissected tissue sections was extracted and purified using the Absolutely RNA nanoprep kit according to the manufacturer's protocol. Caps briefly placed onto the section without laser activation were used as negative controls. Samples from several age-matched lenses were pooled together to obtain differentiation-specific samples for microarray analysis. Target RNA amplification and labeling with Cy-3 or Cy-5 dyes from CyDye Post Labelling Reactive Dye Pack (Amersham, USA) was carried out in two rounds using the Amino Allyl MessageAmp™ aRNA Kit (Ambion, USA) as specified by the manufacturer. Quality and size distribution of the targets were determined by the Agilent 2100 Bioanalyzer (Agilent Technologies, USA). The Amino Allyl MessageAmp aRNA Kit is configured to incorporate the modified nucleotide, 5-(3-aminoallyl)-UTP (aaUTP) into the aRNA during in vitro transcription. Once purified and fragmented, the dye labeled aRNA was used for microarray hybridization. Labeled amplified RNA was hybridized to the 22K Mouse Oligo microarrays (Agilent Technologies)) according to the manufacturer's instructions. After hybridization microarrays were washed and scanned using GenePix 4000A (Axon Instruments,Inc.). Keywords = Lens Keywords = differentiation Keywords = microarray Keywords = gene expression profiling Keywords = laser capture microdissection Keywords = RNA amplification Keywords: repeat sample
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Project description:Background: The molecular determinants of carcinogenesis, tumor progression and patient prognosis can be deduced from simultaneous comparison of thousands of genes by microarray analysis. However, the presence of stroma cells in surgically excised carcinoma tissues might obscure the tumor cell-specific gene expression profiles of these samples. To circumvent this complication, laser microdissection can be performed to separate tumor epithelium from the surrounding stroma and healthy tissue. In this report, we compared RNAs isolated from macrodissected, of which only surrounding healthy tissue had been removed, and microdissected rectal carcinoma samples by microarray analysis in order to determine the most reliable approach to detect the expression of tumor cell-derived genes by microarray analysis. Results: As microdissection yielded low tissue and RNA quantities, extra rounds of mRNA amplification were necessary to obtain sufficient RNA for microarray experiments. These second rounds of amplification influenced the gene expression profiles. Moreover, the presence of stroma cells in macrodissected samples had a minor contribution to the tumor cell gene expression profiles, which can be explained by the observation that more RNA is extracted from tumor epithelial cells than from stroma. Conclusion: These data demonstrate that the more convenient procedure of macrodissection can be adequately used and yields reliable data regarding the identification of tumor cell-specific gene expression profiles. Keywords: macrodissection, microdissection, RNA amplification, stroma, rectal carcinoma
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Project description:E12.5 mouse lens epithelium and fiber cells were collected using Leica LMD 6000 Laser microdissection system. Total RNA was isolated from epithelium and fiber cells using Qiagen RNeasy kit
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