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01.04 Enrichment of activated cd141+ dendritic cells in inflammatory arthritis
  1. Mary Canavan1,
  2. Trudy McGarry1,
  3. Kate Killick2,
  4. Barry Moran3,
  5. Deborah Cluxton3,
  6. Carl Orr4,
  7. Siobhan Wade1,
  8. Hannah Convery4,
  9. Sarah Wade1,
  10. Ronan Mullan5,
  11. Jean Fletcher3,
  12. Douglas Veale4,
  13. Ursula Fearon1
  1. 1Department of Molecular Rheumatology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Ireland
  2. 2Systems Biology Ireland, Conway Institute, University College Dublin, Ireland
  3. 3School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Ireland
  4. 4Centre for Arthritis and Rheumatic Diseases, Dublin Academic Medical Centre, University College Dublin, Ireland
  5. 5Department of Rheumatology, Adelaide and Meath Hospital, Dublin, Ireland

Abstract

Background Dendritic cell (DC) are a heterogeneous group of antigen presenting cells that can be subdivided into CD1c and CD141+DC. CD141+DC were first discovered in 2010 and little is known about their function in health or disease. No previous studies on CD141+DC in inflammatory arthritis (IA) have been performed therefore our aim was to examine if these newly described DC play a pathogenic role in IA.

Methods CD141+DC were purified from SFMC/PBMC, stimulated and stained with a panel of DC specific activation markers. CD141+DC were cocultured with T cells for 6d basally or in the presence of TREM-1 after which intracellular cytokine production was assessed by flow cytometry. Supernatants from DC-T cell cocultures were used to treat synovial fibroblasts and the expression of adhesion molecules, cytokines and MMPs was measured. CD141+DC were also identified in synovial-tissue by immunohistology/flow cytometry for CD141/CLEC9A/TREM-1. Finally using sorted populations of CD141+DC from SFMC and PBMC, RNA sequencing was performed and differentially expressed genes and interaction network analysis were identified using the DeSeq2 R package, Ingenuity® Pathway Analysis (IPA) and InnateDB and Cytoscape.

Results SF-CD141+DC are significantly enriched compared to WB and express higher levels of activation markers CD80/CD86 and CD40. SF-CD141+DC induced both CD8+ and CD4+T cell proliferation, granzyme B production from CD8+ and TNFα, IFNγ and GMCSF from CD4+T cells. Supernatants from CD141+DC activated T cells subsequently induced synovial fibroblast expression of ICAM-1, IL-6, IL-8, MMP1 and MMP3. SF-CD141+DC expressed significantly higher levels of the the hypoxia marker TREM-1, activation of which further induces expression of CD80, CD86 and CD40. Coculture of these TREM-1 activated CD141+DC with CD3+ T cells increased IFNγ and IL-17a production. Finally RNASeq analysis revealed that there are 2089 differentially expressed genes between SF-CD141+DC and PBMC CD141+DC, with induction of a number of key network pathways identified in SF CD141+ DC, including energy metabolism, chemokine and cytokine signalling.

Conclusion CD141+ DC are enriched in the IA joint in an active state. RNASeq analysis revealed they are distinct from blood CD141+DC and our in vitro data would support the hypothesis that these CD141+DC contribute to synovial inflammation and joint destruction.

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