Objectives To mine the serum proteome of patients with systemic sclerosis-associated pulmonary arterial hypertension (SSc-PAH) and to detect biomarkers that may assist in earlier and more effective diagnosis and treatment.
Methods Patients with limited cutaneous SSc, no extensive interstitial lung disease and no PAH-specific therapy were included. They were classified as cases if they had PAH confirmed by right heart catheterisation (RHC) and serum collected on the same day as RHC; and as controls if they had no clinical evidence of PAH.
Results Patients were mostly middle-aged females with anticentromere-associated SSc. Among 1129 proteins assessed by a high-throughput proteomic assay (SOMAscan), only 2 were differentially expressed and correlated significantly with pulmonary vascular resistance (PVR) in SSc-PAH patients (n=15): chemerin (ρ=0.62, p=0.01) and SET (ρ=0.62, p=0.01). To validate these results, serum levels of chemerin were measured by ELISA in an independent cohort. Chemerin levels were confirmed to be significantly higher (p=0.01) and correlate with PVR (ρ=0.42, p=0.04) in SSc-PAH patients (n=24). Chemerin mRNA expression was detected in fibroblasts, pulmonary artery smooth muscle cells (PA-SMCs)/pericytes and mesothelial cells in SSc-PAH lungs by single-cell RNA-sequencing. Confocal immunofluorescence revealed increased expression of a chemerin receptor, CMKLR1, on SSc-PAH PA-SMCs. SSc-PAH serum seemed to induce higher PA-SMC proliferation than serum from SSc patients without PAH. This difference appeared neutralised when adding the CMKLR1 inhibitor α-NETA.
Conclusion Chemerin seems an interesting surrogate biomarker for PVR in SSc-PAH. Increased chemerin serum levels and CMKLR1 expression by PA-SMCs may contribute to SSc-PAH pathogenesis by inducing PA-SMC proliferation.
- systemic sclerosis
- autoimmune diseases
Data availability statement
Data are available on reasonable request.
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Handling editor Josef S Smolen
SS and LR contributed equally.
Contributors All individuals listed as authors met the ICMJE guidelines for determining authorship. SS and RL conceived, designed and supervised the overall study. LR, JCM and RL screened and included patients in the discovery cohort, and contributed samples for the serum experiments. SS, DM and J-LC screened and included patients in the validation cohort, and contributed samples for the serum experiments. SS, LR and JCM performed experiments on serum samples. AMB contributed additional clinical data. SS and LR created and managed the database. CT and GM performed statistical analyses and created Figure 1. SS created figures 2 and 3. EV and RL provided lung samples for the single-cell RNA sequencing. EV performed sequencing experiments, analysed the generated dataset and created Figure 4. LT, DM, MH and CG provided lung samples for the confocal and PASMCs for the proliferation experiments. EH and DL contributed serum samples for the proliferation experiments. LT performed the confocal stainings and PASMC proliferation experiments, and created figure 5 and supplemental figure S1. CT and GM performed statistical analyses. SS created figure 6 and supplemental figure S2. SS wrote the first draft of the manuscript and created the Tables. LT, CG and RL made major revisions to the manuscript. MH provided his PAH expertise. EH and DL provided their SSc expertise. All authors read and approved the submitted version. SS acts as the guarantor of this study.
Funding This work was supported by NIH National Institute of Arthritis Musculoskeletal and Skin Disease grants to RL: Scleroderma Core Centers (5P30AR061271), Scleroderma Center of Research Translation (1P50AR060780) and 2R01AR051089.
Competing interests SS reports travel grants from Shire, Sanofi-Genzyme, SOBI and Novartis; consulting fees from Novartis; outside the submitted work. J-LC reports grants from United Therapeutic, Bioprojet and Topadur, outside the submitted work. DM reports grants and personal fees from Actelion, grants and personal fees from Bayer, personal fees from GSK, personal fees from Pfizer, personal fees from MSD, personal fees from Chiesi, outside the submitted work. EH reports consulting fees from Actelion, Boehringer Ingelheim, GSK, Roche-Chugai, Sanofi-Genzyme; speaking fees from Actelion, GSK, Roche-Chugai; and research funding from CSL Behring, GSK, Roche-Chugai, outside the submitted work. DL reports personal fees from Actelion, grants and personal fees from Takeda, grants and personal fees from CSL Behring, outside the submitted work. MH reports personal fees from Actelion, grants and personal fees from Bayer, grants and personal fees from GSK, personal fees from Merck, personal fees from United Therapeutics, personal fees from Acceleron, outside the submitted work. RL reports research grants from Formation, Elpidera and Kiniksa, outside the submitted work. RL has served as a consultant for Bristol Myers Squibb, Boehringer-Mannheim, Merck, Magenta and Genentech/Roche, outside the submitted work. Over the last three years, CG, reports grants from Acceleron, ShouTi, and Janssen and grants and personal fees from Merck, outside the submitted work.
Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Provenance and peer review Not commissioned; externally peer reviewed.
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