Orbital pseudotumor can be a localized form of granulomatosis with polyangiitis as revealed by gene expression profiling

Abstract

Biopsies and ANCA testing for limited forms of granulomatosis with polyangiitis (GPA) are frequently non-diagnostic. We characterized gene expression in GPA and other causes of orbital inflammation. We tested the hypothesis that a sub-set of patients with non-specific orbital inflammation (NSOI, also known as pseudotumor) mimics a limited form of GPA. Formalin-fixed, paraffin-embedded orbital biopsies were obtained from controls (n=20) and patients with GPA (n=6), NSOI (n=25), sarcoidosis (n=7), or thyroid eye disease (TED) (n=20) and were divided into discovery and validation sets. Transcripts in the tissues were quantified using Affymetrix U133 Plus 2.0 microarrays. Distinct gene expression profiles for controls and subjects with GPA, TED, or sarcoidosis were evident by principal coordinate analyses. Compared with healthy controls, 285 probe sets had elevated signals in subjects with GPA and 1472 were decreased (>1.5-fold difference, false discovery rate adjusted p<0.05). The immunoglobulin family of genes had the most dramatic increase in expression. Although gene expression in GPA could be readily distinguished from gene expression in TED, sarcoidosis, or controls, a comparison of gene expression in GPA versus NSOI found no statistically significant differences. Thus, forms of orbital inflammation can be distinguished based on gene expression. NSOI/pseudotumor is heterogeneous but often may be an unrecognized, localized form of GPA.

Keywords: Gene expression profiling; Granulomatosis with polyangiitis; Microarray analysis; Molecular pathology; Orbit pathology; Orbital pseudotumor.

Figure 1. PCA reveals clustering of expression profiles of samples within each experimental group except NSOI

In both data sets, the NSOI samples were much more heterogeneous and not clustered. The distance between points on the plots is indicative of the difference between gene expression profiles. Rotatable views of the 3-dimensional plots are in online supplemental files (Set1_PCA and Set2_PCA). C: Control; G: GPA; N: NSOI; S: Sarcoidosis; T: TED.

Figure 2. Orbital adipose tissues from some NSOI patients have cytokine expression profile similar to GPA patients

The GPA profile differs from the sarcoidosis, TED, and control profiles. A heat map of relative transcript levels was generated by GSEA from data set 2.

Figure 3. Venn diagrams comparing gene expression in orbital adipose from GPA patients with the other experimental groups

Expression levels in tissues from subjects with GPA were compared to levels in uninflamed control, sarcoidosis, NSOI, and TED orbital adipose samples. Probe sets were considered to indicate a significant difference between groups when their signals had at least a 1.5-fold change with a FDR adjusted p-value less than 0.05 in both sets. Shown are the numbers of unique genes identified by NIH DAVID from probe set lists for each category.