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. 2023 Nov 3:14:1266391.
doi: 10.3389/fimmu.2023.1266391. eCollection 2023.

Reactome pathway analysis from whole-blood transcriptome reveals unique characteristics of systemic sclerosis patients at the preclinical stage

Chiara Bellocchi  1   2 , Xuan Wang  3 , Marka A Lyons  4 , Maurizio Marchini  1 , Maurizio Lorini  1 , Vincenzo Carbonelli  1 , Nicola Montano  1 , Shervin Assassi  4 , Lorenzo Beretta  2
Affiliations

Reactome pathway analysis from whole-blood transcriptome reveals unique characteristics of systemic sclerosis patients at the preclinical stage

Chiara Bellocchi et al. Front Immunol. .

Abstract

Objective: This study aims to characterize differential expressed pathways (DEP) in subjects with preclinical systemic sclerosis (PreSSc) characterized uniquely by Raynaud phenomenon, specific autoantibodies, and/or capillaroscopy positive for scleroderma pattern.

Methods: Whole-blood samples from 33 PreSSc with clinical prospective data (baseline and after 4 years of follow-up) and 16 matched healthy controls (HC) were analyzed for global gene expression transcriptome analysis via RNA sequencing. Functional Analysis of Individual Microarray Expression method annotated Reactome individualized pathways. ANOVA analysis identified DEP whose predictive capability were tested in logistic regression models after extensive internal validation.

Results: At 4 years, 42.4% subjects progressed (evolving PreSSc), while the others kept stable PreSSc clinical features (stable PreSSc). At baseline, out of 831 pathways, 541 DEP were significant at a false discovery rate <0.05, differentiating PreSSc versus HC with an AUROC = 0.792 ± 0.242 in regression models. Four clinical groups were identified via unsupervised clustering (HC, HC and PreSSc with HC-like features, PreSSc and HC with PreSSc-like features, and PreSSc). Biological signatures changed with disease progression while remaining unchanged in stable subjects. The magnitude of change was related to the baseline cluster, yet no DEP at baseline was predictive of progression. Disease progression was mostly related to changes in signal transduction pathways especially linked to calcium-related events and inositol 1,4,5-triphosphate metabolism.

Conclusion: PreSSc had distinguished Reactome pathway signatures compared to HC. Progression to definite SSc was characterized by a shift in biological fingertips. Calcium-related events promoting endothelial damage and vasculopathy may be relevant to disease progression.

Keywords: disease progression; gene expression; pathways; preclinical systemic sclerosis; systemic sclerosis.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Heat maps of differentially expressed Reactome pathways at baseline. Heat maps after unsupervised clustering analysis of the 541 Reactome pathways that were differentially expressed in preclinical systemic sclerosis patients (PreSSc) vs. healthy controls (HCs) (false discovery rate < 0.05 and moderate effect size) at baseline. The subjects can be grouped in four clusters (from top to bottom, yellow to red bars) that are enriched in HCs (top) or PreSSc (bottom) and with a clearly visible gradient of expression profile.
Figure 2
Figure 2
Baseline vs. follow-up differential expression pathway analysis. Heat maps representing the pathways whose expression significantly changed in the pre- and post-analysis in evolving vs. stable preclinical systemic sclerosis (PreSSc) and according to the baseline cluster (see Figure 1 ). In evolving patients (top charts, brown bar), there is a clear shift in the expression profile, especially in the PreSSc-enriched cluster(s); in stable patients, the expression profile remains unaltered.
See this image and copyright information in PMC

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