Basic Research
Copyright ©2005 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Mar 7, 2005; 11(9): 1303-1316
Published online Mar 7, 2005. doi: 10.3748/wjg.v11.i9.1303
Identification of differentially expressed genes after partial rat liver ischemia/reperfusion by suppression subtractive hybridization
Christine Fallsehr, Christina Zapletal, Michael Kremer, Resit Demir, Magnus von Knebel Doeberitz, Ernst Klar
Christine Fallsehr, Magnus von Knebel Doeberitz, Institute of Molecular Pathology, University of Heidelberg, Germany
Christina Zapletal, Department of Surgery, University of Frankfurt am Main, Germany
Michael Kremer, Department of Surgery, University of Heidelberg, Germany
Resit Demir, Department of Surgery, University of Erlangen, Germany
Ernst Klar, Department of Surgery, University of Rostock, Germany
Author contributions: All authors contributed equally to the work.
Correspondence to: Professor Magnus von Knebel Doeberitz, Institute of Molecular Pathology, University of Heidelberg, Im Neuenheimer Feld 220, 69120 Heidelberg, Germany. knebel@med.uni-heidelberg.de
Telephone: +49-6221-562876 Fax: +49-6221-565981
Received: August 13, 2004
Revised: August 14, 2004
Accepted: October 5, 2004
Published online: March 7, 2005
Abstract

AIM: To identify potential diagnostic target genes in early reperfusion periods following warm liver ischemia before irreversible liver damage occurs.

METHODS: We used two strategies (SSH suppression subtractive hybridization and hybridization of cDNA arrays) to determine early changes in gene expression profiles in a rat model of partial WI/R, comparing postischemic and adjacent nonischemic liver lobes. Differential gene expression was verified (WI/R; 1 h/2 h) and analyzed in more detail after warm ischemia (1 h) in a reperfusion time kinetics (0, 1, 2 and 6 h) and compared to untreated livers by Northern blot hybridizations. Protein expression was examined on Western blots and by immunohistochemistry for four differentially expressed target genes (Hsp70, Hsp27, Gadd45a and IL-1rI).

RESULTS: Thirty-two individual WI/R target genes showing altered RNA levels after confirmation by Northern blot analyzes were identified. Among them, six functionally uncharacteristic expressed sequences and 26 known genes (12 induced in postischemic liver lobes, 14 with higher transcriptional expression in adjacent nonischemic liver lobes). Functional categories of the verified marker genes indicate on the one hand cellular stress and tissue damage but otherwise activation of protective cellular reactions (AP-1 transcription factors, apoptosis related genes, heat shock genes). In order to assign the transcriptional status to the biological relevant protein level we demonstrated that Hsp70, Hsp27, Gadd45a and IL-1rI were clearly up-regulated comparing postischemic and untreated rat livers, suggesting their involvement in the WI/R context.

CONCLUSION: This study unveils a WI/R response gene set that will help to explore molecular pathways involved in the tissue damage after WI/R. In addition, these genes especially Hsp70 and Gadd45a might represent promising new candidates indicating WI/R liver damage.

Keywords: Organ injury, Liver ischemia, AP-1 transcription factor, Heat shock protein, Gadd45A, Suppression subtractive hybridization