ABSTRACT
MRLlpr mice develop spontaneous systemic autoimmunity with many hallmarks of the human disease systemic lupus erythematosus. Although a variety of genes have been implicated in this model, disease pathogenesis is still poorly understood. In an effort to identify novel genes and pathways, we performed genome-wide mRNA expression analysis in the spleens and kidneys of MRLlpr mice throughout the disease course. Samples were collected from cohorts of C57BL/6, MRL+/+ and MRLlpr mice, and profiled by flow cytometry and gene expression microarrays. Serum autoantibodies and renal pathology were studied in parallel. We identified 236 genes in MRLlpr spleen that showed significant threefold or greater changes in expression between 6 and 20 weeks. Of interest, a number of interferon-responsive genes were expressed early, and remained dysregulated throughout the disease course. Many chemokines, cell surface proteins, transcription factors and cytokines, including IFN-gamma, also showed altered expression as disease progressed. Analysis of kidneys indicated the presence of severe inflammation that coincided with evidence for changes in kidney function and elevated expression of IFN-inducible genes, complement components and antigen presentation genes. These data provide a unique genomic view of the progression to fatal autoimmunity in MRLlpr mice, and provide new candidate genes and pathways to explore.
Subject(s)
Autoimmunity , Genome , Lupus Erythematosus, Systemic/genetics , Lupus Erythematosus, Systemic/immunology , Animals , Chemokines/genetics , Chemokines/physiology , Disease Progression , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Gene Expression Profiling , Interferon-gamma/genetics , Interferon-gamma/physiology , Kidney/immunology , Kidney/pathology , Lupus Erythematosus, Systemic/pathology , Membrane Proteins/genetics , Membrane Proteins/physiology , Mice , Mice, Inbred C57BL , Mice, Inbred MRL lpr , Oligonucleotide Array Sequence Analysis , Spleen/immunology , Spleen/pathology , Transcription Factors/genetics , Transcription Factors/physiologyABSTRACT
Monitoring of gene and protein expression in peripheral blood cells has significant potential for improving the diagnosis and therapy of many human diseases. As genomic-scale microarray and proteomic technologies are applied to peripheral blood, it is important to consider the variables that may affect interpretation of data. Here we report experiments performed to identify genes that are particularly sensitive to ex vivo handling prior to RNA extraction for gene expression microarrays or quantitative real-time RT-PCR assays. We examined Affymetrix gene expression in samples from eight normal individuals where blood was processed for RNA either immediately after blood draw or the next day following overnight incubation. These studies identified hundreds of genes that are sensitive to ex vivo handling of blood, and suggest that this is an important variable to consider when designing and interpreting human PBMC experiments.
