Monitoring the gene expression profiles of chronic allograft dysfunction by cDNA microarray

Abstract

Background: Gene expression profiles of chronic allograft dycfunction is essential for understanding the distinct mechanism of allograft dysfunction and exploring new target genes for therapeutic purposes. In the study, we investigate the gene expression profile of 1081 known genes during chronic dysfunction of cardiac allograft in rats. Methods: Heterotropic heart transplantation was performed using male LEW (RT1l ) and F344 (RT1avl) rats as donor and recipient, respectively. Cyclosporin A (2.5mg/kg/ d. i.m. d.0-4) was given to recipients to overcome the acute rejection episodes. 60 % of long-term survival of allograft was achieved after the treatment. Grafts were harvested on day 100 (histology showed obliterative arteriopathy, fibrosis and tissue demage). Syngeneic LEW grafts that harvested at similar time point served as control. High purity of total RNA was isolated form grafts. Total RNA was then transcripted reversely to cDNA and labeled with Cy3 (allograft control) or Cy5 (isograft). Samples were premixed and hybridized with the DNA sequences on the Atlas Glass Microarray (Clontech). Slides were detected with Scan Array 5000 and the data were analyzed using Quantarray. Results: After normalization with 9 housekeeping genes, total 238 of 1081 genes (mean of 5 experiments) were over-expressed (fold > 2.0) and 2 genes were under-expressed (fold < 0.5) in rejecting allografts on day 100. Overexpressed genes encoding proteins involved in interleukin and interferon receptors (5 genes, CXCR2, IL-4R, IL-1R, etc.), hormone and hormone receptors (30 genes, TGF-beta superfamily receptors, somatoliberin, thymosin beta-like protein, etc), growth factors (27 genes, activin type receptor, bone morphogenetic protein, TGFbeta, heparin-binding growth factor, IGF, FGF, hGF, etc.), proteinase inhibitors (9 genes, calpastatin, plasma proteinase inhibitor, plasminogen activator inhibitor, tissue inhibitor of metalloprotinase, etc.), G-protein and G-protein-coupled receptors (36 genes, ras-related proteins and GTPase, GPR6, PAR2, GPR19, etc.) and energy metabolism (14 genes, COX5B, NSE, MCAD, etc.) Conclusions: The data provide new insights into the molecular process of chronic allograft dysfunction involving the over-expression of hormones, growth factors and proteinase inhibitors.