Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Gene expression in the pancreas of healthy control, auto-antibody positive, and type 1 diabetic subjects

ABSTRACT: These experiments were performed to identify differentially expressed genes in the pancreas of healthy humans, auto-antibody positive and type 1 diabetic patients. All samples were obtained from the network of pancreatic organ donors with diabetes (nPOD). ID numbers are specified. Patient information can be obtained at http://www.jdrfnpod.org/ Pancreas tissues were homogenized in Trizol reagent (Invitrogen). Total RNA was extracted from the aqueous phase using the RNeasy mini kit (Qiagen). RNA quality was assessed using the Agilent 2100 Bioanalyzer and the RNA 6000 Nano Reagent Kit (Agilent). One color microarrays were performed at the Stanford Human Immune Monitoring Center using the SurePrint G3 Human GE 8x60K microarray kit (Agilent Technologies). All samples were processed and run at the same time.

ORGANISM(S): Homo sapiens

SUBMITTER: Linda Yip

PROVIDER: E-GEOD-72492 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:A microarray study performed in the pancreatic lymph nodes of Deaf1 knock-out and BALB/c control mice to identify genes that are regulated by the transcriptional regulator Deaf1. These experiments constitute a portion of the study described below: Abstract: Type 1diabetes (T1D) can result from a breakdown in peripheral tolerance which is controlled by peripheral tissue antigen (PTA) expression in lymph nodes. Here, we identified a transcriptional regulator, deformed epidermal autoregulatory factor 1 (Deaf1), which regulates the expression of various PTAs in the pancreatic lymph node (PLN). We found, by microarray, that Deaf1 controls the expression of ~600 genes in the PLN. In the non-obese diabetic (NOD) mouse model of T1D, we identified a wild-type form of Deaf1 (DF1) and a truncated alternatively spliced variant (DF1-VAR1) that hetero-dimerizes with and decreases the transcriptional activity of DF1. The expression of DF1 correlates with the expression of various pancreatic PTAs such as insulin, and during the onset of destructive insulitis in NOD mice, DF1 expression is downregulated, while the DF1-VAR1 expression is upregulated in the PLN. A reduction in DF1-controlled PTA expression in the PLN, leading to decreased peripheral tolerance, could underlie the pathogenesis of NOD disease. Deaf1-KO mice (4 wk old) and age-matched BALB/c control mice were sacrificed, and the PLN were removed and immediately homogenized in Trizol Reagent. RNA was extracted in Trizol and then purified using the RNeasy kit (Qiagen). RNA quality was assessed using the Agilent RNA 6000 Nano Reagents, RNA Nano chips, and the Agilent 2100 bioanalyzer (Agilent Technologies), according to manufacturerâs instructions. Control and Deaf1-KO mouse RNA was amplified, labeled with Cy3 and Cy5, respectively, and combined with spike A and spike B mix, respectively, using the Agilent low RNA input fluorescence linear amplification kit (Agilent Technologies). The amplified cRNA was purified using the RNeasy kit (Qiagen), and specific activity was determined with the NanoDrop 1000 spectrophotometer (Thermo Scientific). Samples were prepared with the gene expression hybridization kit (Agilent Technologies) and two color microarrays were performed using the whole mouse genome (4x44K) Oligo microarray kit, according to manufacturerâs instructions. Microarray chips were washed and scanned using the DNA microarray scanner (Agilent Technologies). Data was processed with Feature Extraction Software (Agilent Technologies), and analyzed using GeneSpring GX 7.3 Software (Agilent Technologies). This submission shows the data obtained from two individual Deaf1 knockout mice measured against a pool of 4 BALB/c control mice.

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Gene expression in the pancreas of healthy control, auto-antibody positive, and type 1 diabetic subjects

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