ABSTRACT
DNA microarray technology has developed rapidly in recent years and has become an essential tool, providing novel approaches to biomedical research. In this paper, we describe a self-designed ImmunoChip cDNA array for immunological research. With a comprehensive selection of genes of interest, we can focus on key signalling pathways and molecular mechanisms at relatively low cost compared to commercial platforms which are usually targeted at global screening of gene expression. To validate the efficiency of the ImmunoChip, we studied T helper cell polarization to functionally distinct subsets (Th1 and Th2). We also developed a tool for quality control of cDNA microarrays that assesses the technical quality of an ImmunoChip. The information produced with the quality control tool is shown to be valuable for extracting correct information from cDNA microarrays. Gene expression measurements with ImmunoChip are in agreement with the results obtained using oligonucleotide microarrays and with published quantitative RT-PCR data. The ImmunoChip provides reliable measurements and gives new insights into various aspects of human immune responses.
Subject(s)
DNA, Complementary , Immunity, Cellular , Oligonucleotide Array Sequence Analysis/methods , Gene Expression Profiling , Gene Expression Regulation/immunology , Humans , Interleukin-2/physiology , Interleukin-4/physiology , Quality Control , Th1 Cells/immunology , Th1 Cells/metabolism , Th2 Cells/immunology , Th2 Cells/metabolism , Transforming Growth Factor beta/physiologyABSTRACT
An association between cytomegalovirus (CMV) infection and alloresponse has been suggested. CMV increases inflammation and adhesion molecule expression in graft, and induces cytokines and growth factors, linked with transplant vasculopathy and chronic rejection. We have investigated the gene expression of various inflammatory factors in the CMV-associated immune response and compared this with the immune response of acute rejection in liver transplants by using DNA microarray technology. Gene expression was studied at mRNA level in biopsies from liver transplant patients experiencing CMV infection or acute rejection. RNA extracted from liver grafts after reperfusion was used as control material. Among the strongly upregulated genes in the specimens obtained from liver transplants during CMV infection were IFN-gamma, caspases 1 and 3, granzymes A and B, TGF-beta receptors II and III, IL-10 receptor alpha, VCAM-1, TNF receptor, IL-4, TNF-alpha, IL-10, IL-2 receptor beta, IL-1beta, PDGF-receptor beta, vascular adhesion protein-1, TGF-beta2, and ICAM-1. In biopsies with acute liver allograft rejection, the most significantly upregulated genes were MHC class II, IFN-gamma, caspases 1 and 3, IL-2R beta and gamma, granzymes A and B, VLA-4, L-selectin, E-selectin, VCAM-1, and IL-1beta. Upregulated genes common for CMV and alloresponse were granzyme A and B, E-selection, IFN-gamma, VCAM-1, VLA-4, TNF, caspases 1, 3, and 8, and PDGF. Microarray analysis defined different entities in the immune responses of CMV infection and acute rejection. The differences and similarities of the gene expression profiles related to those in CMV infection and rejection may help to understand the intragraft immunologic events.
