Background/Purpose:

Immature dendritic cells (iDC) have been shown to act as OC precursors and are important cell players in the pathogenesis of RA. We aimed to investigate the relationship between iDC derived osteoclastogenesis and specific immunity in RA. 

Methods:

CD14⁺ monocytes from PB of ACPA⁺ RA patients and healthy individuals were first cultured in the presence of GM-CSF and IL-4 to generate iDC or in the presence of M-CSF to generate MΦ and then further differentiated to OC in the presence of RANKL and M-CSF. In parallel, cells were grown on osteoassay surfaces and bone resorption area was quantified by computer assisted image analysis. Mass spectrometry analysis was performed on different stages of differentiation of iDC and MΦ derived OC.  ACPA positive and negative polyclonal IgGs were isolated from synovial fluid (SF, n=26) and peripheral blood (PB, n=38) samples of RA patients. Cytokines were measured by cytometric bead arrays in cultures supernatants. Immunohistochemistry (IHC) was used to stain the OCs with murinized monoclonal ACPAs derived from single SF derived B cells. The effect of IL-8 inhibition was tested in the OC cultures.

Results:

Principal component analysis confirmed distinct proteomic profiles during OC maturation from iDC and MΦ, respectively. Vimentin significantly increased during iDC-OC maturation, with citrullinated vimentin peptides detectable in matured OCs. Polyclonal ACPAs enhanced osteoclastogenesis and bone resorption from iDC (fold increase of 1.6±0.2 for OC number and 2.0±0.3 for bone resorption area). Similar effect was observed when the iDC were derived from ACPA+ RA patients (fold increase of 2.3±0.9 for OC number and 2.6±0.1 for bone resorption area). PB derived ACPAs were equally effective with SF ACPAs. Increased osteoclastogenesis was associated with significantly higher levels of IL-8 levels in cultures supernatants (fold increase of 2.4±0.5). Immunohistochemistry demonstrated presence of cit peptides in both iDC precursors and iDC-derived mature OCs suggesting that citrullination is important for iDC-OC differentiation and maturation (Figure 6B). PAD2 and PAD4 showed faint staining in CD14 monocytes with increased staining intensity in both iDC precursors and more mature OCs. The importance of citrullination and PAD enzymes for iDC transdifferentiation was confirmed by a dose-dependent inhibition of OC differentiation using PADi in the presence of RANKL and M-CSF. IL-8 was the main cytokine detected in the culture supernatants of iDC-derived cultures.

Conclusion:

iDC can effeiciently transdifferentiate in OC and are targeted by RA specific antibodies.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/immature-dendritic-are-potent-oc-precursors-in-ra-and-are-targeted-by-ra-specific-antibodies/