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Introduction Rheumatoid arthritis (RA) is an autoimmune disease with unknown etiopathogenesis where systemic autoimmunity precedes clinical disease onset. Adaptive immunity is initiated in lymphoid tissue where lymph node stromal cells (LNSC) play a crucial role in shaping the immune response and maintaining peripheral tolerance. Objectives We developed an experimental model for studying the functional capacities of human LNSC during the earliest phases of RA and compared their cellular and molecular characteristics to LNSC from healthy volunteers. Methods ACPA +RA patients (n=24), ACPA +RA risk individuals (n=23) and seronegative healthy controls (n=14;HC) underwent ultrasound-guided inguinal lymph node biopsy. Human LNSCs were isolated and expanded in vitro for cellular (flow cytometry), molecular (methylome, transcriptome and microRNA) and functional analyses. Results Key LN chemokines CCL19, CCL21 and CXCL13 were induced in LNSCs upon stimulation with TNFα and lymphotoxin α1β2, but to a lesser extent in LNSCs from RA patients. RNA sequencing was performed on LNSC of HC (n=5), ACPA +RA risk individuals (n=6) and ACPA +RA patients (n=4). Of interest, LNSC from ACPA +RA risk individuals and ACPA +RA patients revealed a common significantly differential expressed gene signature compared with HC LNSC. Pathway analysis of this common signature showed, among others, significant enrichment of pathways affecting actin cytoskeleton, focal adhesion and cell junction. Accordingly, in a gel contraction assay LNSC from ACPA +RA risk individuals and RA patients showed impaired collagen contraction compared to healthy LNSC. In RA LNSC a significant enrichment was observed for genes involved in TGFb signalling while in RA-risk LNSC cell cycle genes were differentially expressed compared with HC. DNA methylation analyses revealed common differentially methylated CpG sites (DMS) in LNSC from ACPA +RA patients (n=5) and ACPA +RA risk individuals (n=3) compared with HC (n=4). These DMS were significantly hypomethylated and associated with antigen processing and presentation (HLA-DRB1). Conclusions This data point towards alterations in the cytoskeleton and antigen-processing and presentation in LNSC from ACPA+RA risk individuals and RA patients. Further studies are required to investigate the influence of this LNSC abnormality on immune responses. Disclosure of interest C. Ospelt: None declared, E. Karouzakis: None declared, J. Hahnlein: None declared, H. Semmelink: None declared, R. Gay: None declared, P.-P. Tak Employee of: Currently: Senior Vice President R and D Pipeline, Global Development Leader and Chief Immunology Officer, GSK, D. Gerlag Employee of: Currently: Head Clinical Unit Cambridge at GSK, S. Gay: None declared, L. Van Baarsen: None declared |
