Systemic sclerosis and lupus: points in an interferon-mediated continuum

Abstract

Objective: To investigate peripheral blood (PB) cell transcript profiles of systemic sclerosis (SSc) and its subtypes in direct comparison with systemic lupus erythematosus (SLE).

Methods: We investigated PB cell samples from 74 SSc patients, 21 healthy controls, and 17 SLE patients using Illumina Human Ref-8 BeadChips and quantitative polymerase chain reaction confirmation. None of the study participants were receiving immunosuppressive agents other than low-dose steroids and hydroxychloroquine. In addition to conventional statistical and modular analysis, a composite score for the interferon (IFN)-inducible genes was calculated. Within the group of patients with SSc, the correlation of the IFN score with the serologic and clinical subtypes was investigated, as were single-nucleotide polymorphisms in a selected number of IFN pathway genes.

Results: Many of the most prominently overexpressed genes in SSc and SLE were IFN-inducible genes. Forty-three of 47 overexpressed IFN-inducible genes in SSc (91%) were similarly altered in SLE. The IFN score was highest in the SLE patients, followed by the SSc patients, and then the controls. The difference in IFN score among all 3 groups was statistically significant (P < 0.001 for all 3 comparisons). SSc and SLE PB cell samples showed striking parallels to our previously reported SSc skin transcripts in regard to the IFN-inducible gene expression pattern. In SSc, the presence of antitopoisomerase and anti-U1 RNP antibodies and lymphopenia correlated with the higher IFN scores (P = 0.005, P = 0.001, and P = 0.004, respectively); a missense mutation in IFNAR2 was significantly associated with the IFN score.

Conclusion: SLE and SSc fit within the same spectrum of IFN-mediated diseases. A subset of SSc patients shows a "lupus-like" high IFN-inducible gene expression pattern that correlates with the presence of antitopoisomerase and anti-U1 RNP antibodies.

Figure 1

Differentially expressed pathways in systemic lupus erythematosus (SLE) patients and systemic sclerosis (SSc) patients compared with controls (further information is available online at http://www.uth.tmc.edu/scleroderma). IRF = interferon regulatory factor; PKR = RNA-dependent protein kinase.

Figure 2

Mapping modular blood transcriptional activity. The heatmap represents levels of activity for a set of predetermined transcriptional modules. Average gene expression levels were obtained for each module. For each patient, 28 vectors were thus obtained, and the data set was subjected to an unsupervised analysis: modules for which at least 1 sample showed a deviation from the median of >1.5-fold were selected and arranged based on their activity pattern across patients. Conversely, patients were arranged using the same hierarchical clustering algorithm based on their activity patterns across modules. Red indicates a relative increase in transcriptional activity; blue indicates a relative decrease. The distribution of the different study groups is indicated by a color code. Functional interpretations available for some of these modules are also provided. See Figure 1 for definitions.

Figure 3

A and B, Interferon (IFN) score based on 43 IFN-inducible genes in the 3 study groups (A) and in the serologic subtypes of SSc (B). The IFN score was also dichotomized according to a threshold value corresponding to the 95th percentile in controls (horizontal line). C and D, Average score of 3 IFN-inducible genes based on quantitative polymerase chain reaction in the 3 study groups (C) and in the serologic subtypes of SSc (D). Data are shown as box plots. Each box represents the 25th to 75th percentiles. The depth of the box is the interquartile range (IQR). The line inside the box represents the median. The upper adjacent value is the largest observation that is less than or equal to the 75th percentile plus 1.5 times the IQR. The lower adjacent value is the smallest observation that is greater than or equal to the 25th percentile minus 1.5 times the IQR. ACA = anticentromere antibody; ARNA = anti–RNA polymerase III; ATA = antitopoisomerase; RNP = anti–U1 RNP (see Figure 1 for other definitions).

Figure 4

Unsupervised hierarchical clustering of 43 interferon-inducible transcripts. Samples are labeled according to study groups (SSc patients = green; SLE patients = red; controls = blue). See Figure 1 for definitions.