Discovery and analysis of inflammatory disease-related genes using cDNA microarrays

Abstract

cDNA microarray technology is used to profile complex diseases and discover novel disease-related genes. In inflammatory disease such as rheumatoid arthritis, expression patterns of diverse cell types contribute to the pathology. We have monitored gene expression in this disease state with a microarray of selected human genes of probable significance in inflammation as well as with genes expressed in peripheral human blood cells. Messenger RNA from cultured macrophages, chondrocyte cell lines, primary chondrocytes, and synoviocytes provided expression profiles for the selected cytokines, chemokines, DNA binding proteins, and matrix-degrading metalloproteinases. Comparisons between tissue samples of rheumatoid arthritis and inflammatory bowel disease verified the involvement of many genes and revealed novel participation of the cytokine interleukin 3, chemokine Gro alpha and the metalloproteinase matrix metallo-elastase in both diseases. From the peripheral blood library, tissue inhibitor of metalloproteinase 1, ferritin light chain, and manganese superoxide dismutase genes were identified as expressed differentially in rheumatoid arthritis compared with inflammatory bowel disease. These results successfully demonstrate the use of the cDNA microarray system as a general approach for dissecting human diseases.

Figures

Figure 2

Time course for LPS/PMA-induced MM6 cells. Array elements are described in Fig. 1. (A) Pseudocolor representations of fluorescent scans correspond to gene expression levels at each time point. The array is made up of 8 Arabidopsis control targets and 86 human cDNA targets, the majority of which are genes with known or suspected involvement in inflammation. The color bars provide a comparative calibration scale between arrays and are derived from the Arabidopsis mRNA samples that are introduced in equal amounts during probe preparation. Fluorescent probes were made by labeling mRNA from untreated MM6 cells or LPS and PMA treated cells. mRNA was isolated at indicated times after induction. (B I–III) The two-color samples were cohybridized, and microarray scans provided the data for the levels of select transcripts at different time points relative to abundance at time zero. The analysis was performed using normalized data collected from 8-bit images.

Figure 3

Time course for IL-1β and TNF-induced SW1353 cells using the inflammation array (Fig. 1). (A) Pseudocolor representation of fluorescent scans correspond to gene expression levels at each time point. (B I–IV) Relative levels of selected genes at different time points compared with time zero.

Figure 4

Expression profiles for early passage primary synoviocytes and chondrocytes isolated from RA tissue, cultured in the presence of 10% fetal calf serum and activated with PMA and IL-1β, or TNF and IL-1β, or TGF-β for 18 hr. The color bars provide a comparative calibration scale between arrays and are derived from the Arabidopsis mRNA samples that are introduced in equal amounts during probe preparation

Figure 5

Expression profiles of RA tissue (A) and IBD tissue (B). mRNA from RA tissue samples obtained from the same individual was isolated directly after excision (RA 21.5A) or maintained in culture without serum for 2 hr (RA 21.5B) or for 6 hr (RA 21.5C). Profiles from tissue samples of two other individuals (data not shown) were remarkably similar to the ones shown here. IBD-A and IBD-CI are from mRNA samples prepared directly after surgery from two separate individuals. For the IBD-CII probe, the tissue sample was cultured in medium without serum for 2 hr before mRNA preparation.

Figure 1

Ninety-six-element microarray design. The target element name and the corresponding gene are shown in the layout. Some genes have more than one target element to guarantee specificity of signal. For TNF the targets represent decreasing lengths of 1, 0.8, 0.6, 0.4, and 0.2 kb from left to right.