Identification of novel DC subsets in the RA joint which induce T cell function, and activation of which are potentiated by the hypoxic environment of the joint
Canavan M, O’Rourke M, Orr C, Fletcher J, Veale DJ, Fearon U
Centre for Arthritis and Rheumatic Diseases, Dublin Academic Medical Centre and Conway Institute of Biomolecular and Biomedical Research, Dublin 4, Ireland
Dendritic cells (DC) are antigen presenting cells which link both innate and adaptive immunity.
In this study we characterise novel DC population within the inflamed RA synovium, examine their effect on T-cell function, and demonstrate that the hypoxic microenvironment of the joint can potentiate these mechanisms.
RA synovial tissue was obtained from site of inflammation at arthroscopy. For characterisation of synovial tissue DC, biopsies were digested, and in parallel with synovial fluid/peripheral blood mononuclear cells (SFMC/PBMC), gated on CD45+ cells. DC were then defined as HLADR+, Lineage-, and cell surface expression of CD11c, CD1, CD141, CD123, CD80, CD83 and CD40 was assess by Flow-cytometry. To analyse the effect of the inflamed microenvironment on DC activation and function, cells were cultured with RA synovial explant conditioned media (ECM) and synovial fluid (SF) under normoxic or hypoxic conditions, and activation/function assessed by flow cytometry. Finally monocyte (Mo) derived-DC were treated with ECM and RA SF and co-cultured with CD4+ T-cells for 5 days, supernatants harvested and T-cells restimulated with PMA (50ng/ml) and Ionomycin (500ng/ml) and intracellular cytokines expression quantified.
In this study we demonstrated a significant increased gradient in DC maturation markers CD40 and CD80 as DC migrate from blood, to fluid and finally to the synovial tissue (p<0.05;p<0.05;p<0.05).
We have identified differential DC sub-population localized to the RA inflamed joint which are more activated, induce T-cell function and effects of which are potentiated by the hypoxic nature of the joint.