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| Status |
Public on Jun 21, 2006 |
| Title |
Genome-wide expression profile in rat model of renal isografts from brain dead donors |
| Organism |
Rattus norvegicus |
| Experiment type |
Expression profiling by array
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| Summary |
The clinical results obtained with organ allografts from marginal or extended donors are less satisfactory over both the short-term and long-term than the outcome of grafting from living donors. Early postoperative graft function depends on several pretransplant factors, with the major donor influences being brain death (BD) and ischemia/reperfusion(I/R) injury. The effects of BD and I/R injury not only reduce the number of functioning nephrons, but also trigger a host immune response to the grafted kidney. It has been hypothesized that allografts from marginal sources may not be biologically inert at the time of surgery, but may already be programmed to initiate or amplify a host response. To exclude the effects of allogenicity, we established a rat renal isograft model using dead donors and compared the results with those obtained after grafting from living donors. In this study, we performed an analysis of the changes of gene expression in rat kidney isografts using samples obtained at 0(pre-transplant) and 1 hour time points since transplantation procedure. The results enhanced our insight into the pathways and cascades that are activated or down-regulated by BD and/or I/R injury. Better knowledge of the key components of BD or I/R injury will provide clues to the factors triggering progression that leads to a decline of organ viability, as well as ideas on how to overcome such graft failures. Such data may also allow us to identify novel biomarkers as predictors of adverse outcomes.
Keywords: disease state analysis
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| Overall design |
Inbred male Lewis rats were used for these experiments as recipients and donors. The left kidney was transplanted orthotopically by end-to-end anastomosis. The contralateral right kidney was removed at the time of transplantation. The time of operative ischemia for transplantation was 30 min, which did not vary between animals. Donor animals were divided into two groups. The experimental group received kidneys from BD rats, while kidneys from living donors were used in the control group. Brain death was produced by gradually increasing the intracranial pressure that led to brain stem herniation. The rats were mechanically ventilated for a period of 6 hours. We compared the gene expression profiles of renal isografts from BD donors and grafts from living donors using a high-density oligonucleotide microarray that contained approximately 20,500 genes.
For DNA microarray experiments, 200 ng aliquots of total RNA were labeled using the Agilent Low RNA Input Fluorescent Linear Amplification Kit (Agilent Technologies Product) according to the manufacturer's instructions. RNA purified from each kidney graft was used for microarray analysis (Cy3-labeled), with pooled RNA derived from normal kidneys as a template control (Cy5-labeled). After checking the labeling efficiency, 1ug aliquots of Cy5-labeled normal control RNA and Cy3-labeled RNA from individual grafts (control 0 hour, 1 hour, BD 0 hour, BD 1 hour, n=3/group) were mixed, and then hybridized to Agilent Rat Oligo Microarrays (Product No. G4130A). After washing, the microarray slides were analyzed with an Agilent Microarray scanner and software (scanner model G2565BA). Data analysis was performed using Agilent Feature Extraction software (Ver. A.7.1.1), and Excel 2003 (Microsoft). The data were imported into GeneSpring 7.0 (Silicon Genetics, Redwood City, CA), with per spot, per chip, and intensity dependent (lowess) normalization being applied for each array. The ratio of the normalized channels (Cy3/Cy5) was used to assess the level of expression.
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| Citation(s) |
15808645 |
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| Submission date |
Jun 19, 2006 |
| Last update date |
Dec 06, 2012 |
| Contact name |
Mamoru Kusaka |
| E-mail(s) |
[email protected]
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| Phone |
81-562-93-2181
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| Fax |
81-562-93-7863
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| Organization name |
Fujita-Health University
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| Department |
Department of Urology
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| Street address |
1-98 Kutsukake
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| City |
Toyoake |
| State/province |
Aichi |
| ZIP/Postal code |
470-1192 |
| Country |
Japan |
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| Platforms (1) |
| GPL890 |
Agilent-011868 Rat Oligo Microarray G4130A (Feature Number version) |
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| Samples (12)
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| GSM115033 |
Kidney sample from brain dead donor (6hrs after induction of brain death) BD1 |
| GSM115034 |
Normal kidney sample NK1 |
| GSM115035 |
1 hr transplanted kidney sample from brain dead donor B21 |
| GSM115036 |
1 hr transplanted kidney sample from normal donor C22 |
| GSM115037 |
1 hr transplanted kidney sample from brain dead donor B23 |
| GSM115038 |
1 hr transplanted kidney sample from brain dead donor B27 |
| GSM115039 |
Kidney sample from brain dead donor(6hrs after induction of brain death) BD2 |
| GSM115040 |
Kidney sample from brain dead donor(6hrs after induction of brain death) BD3 |
| GSM115041 |
1 hr transplanted kidney sample from normal donor C24 |
| GSM115042 |
1 hr transplanted kidney sample from normal donor C25 |
| GSM115043 |
Normal kidney sample NK2 |
| GSM115044 |
Normal kidney sample NK3 |
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| Relations |
| BioProject |
PRJNA95363 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE5104_RAW.tar |
2.8 Mb |
(http)(custom) |
TAR |
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