Background/Purpose: Dendritic cells (DC) are a heterogeneous population of professional antigen presenting cells which link both the innate and adaptive arms of the immune system. Myeloid and plasmacytoid DC represent the two major DC subsets and can be distinguished based on their morphology, expression of surface markers and gene expression profiles. In this study we compared the percentage and activation status of  myeloid versus plasmacytoid DC at the site of inflammation in RA compared to systemic circulation.

Methods: DC whole blood phenotyping was assessed using multicolour flow cytometry on the Beckman Coulter Cyan system using FlowJo software for subsequent analysis. DC were defined as HLADR⁺, Lineage^–, and further subdivided as either myeloid (CD11c⁺, CD1⁺ or CD141⁺ ) or plasmacytoid DC ( CD123⁺ ). Cell surface expression of CD80, CD83 and CD40 was used to assess the activation and maturation status of each subtype. For characterisation of synovial tissue DC, biopsies were digested using the GentleMACs mechanical and enzymatic digestion system. Viable CD45 cells were gated and subsequently assessed for DC pan markers in addition to maturation and activation markers. In parallel, synovial fluid and peripheral blood were comparatively assessed to profile DC from blood, fluid and tissue. To assess the effect of the synovial environment on DC maturation, immature DC were derived from CD14+ monocytes in the presence of GM-CSF (70ng/ml) and IL-4 (50ng/ml). Synovial tissue explants were cultured for 24hr allowing the spontaneous release of cytokines and soluble mediators into the culture medium. Monocyte derived dendritic cells (MoDC) were cultured in the presence of this explant conditioned media for 24hr after which the expression of maturation markers was analysed on CD11c⁺ CD14^–DC.

Results: RA patients have a decreased percentage of CD11c mDC circulating in peripheral blood compared to that of healthy controls. The percentage of CD123 pDC between RA and healthy controls is not significantly different however the expression of CD40 on pDC in RA patients is significantly increased compared to healthy control (p<0.001). A comparative analysis of mDC and pDC in peripheral blood, synovial fluid and synovial tissue highlighted an increase in DC maturation as DC migrate from blood, to fluid and finally tissue. CD40 and CD83 expression on mDC is increased in tissue compared to that of fluid or blood. Similarly an increase in CD40 and CD83 on pDC was found in synovial tissue compared to that of fluid or peripheral blood. Finally immature monocyte derived DC cultured in the presence of explant conditioned media have increased expression of  CD80 and CD83 compared to basal DC medium.

Conclusion: DC have a more mature and activated phenotype in the synovial tissue compared to that in synovial fluid or peripheral blood. Given that there are also lower circulating levels of DC in RA patients compared to controls our data suggest that peripheral blood DC are recruited to the joint where they undergo a programme of maturation. Ongoing studies aim to elucidate the mechanisms in which these subsets are activated and matured.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/investigating-myeloid-and-plasmacytoid-dendritic-cell-activation-within-the-synovium-and-peripheral-blood-of-rheumatoid-arthritis-patients/