Arthritis & Rheumatism, Volume 62,
November 2010 Abstract Supplement

Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Atlanta, Georgia November 6-11, 2010.


Lupus Immune Complex Gene Expression Arrays Reveal Divergent Pro- and Anti-Inflammatory Pathways Induced Depending on the Presence of Plasmacytoid Dendritic Cells and C1q.

Santer,  Deanna M., Elkon,  Keith B.

Background:

Patients with Systemic Lupus Erythematosus (SLE) have an 'IFN-signature' that correlates with disease severity. RNA-containing immune complexes (ICs) stimulate IFN-alpha (IFN-a) production by plasmacytoid dendritic cells (pDCs) through activation of TLR7. We recently demonstrated that normal human serum contains potent inhibitors of IFN-a including IgG and the classical complement protein C1q. We showed that C1q inhibition of IFN-a required monocytes to indirectly inhibit pDCs, but thus far we only knew that the inhibition was soluble factor independent. The purpose of this study was 2-fold: 1) To determine what genes are induced by C1q-containing ICs that could explain the mechanism by which C1q regulates IFN-a induction, and 2) To identify what other pathways, in addition to type I IFNs, are induced by SLE ICs.

Methods:

SLE ICs were formed with diluted SLE serum (1:2000) or purified IgG and necrotic cell extract. ICs were added to normal donor peripheral blood mononuclear cells (PBMCs) or purified monocytes (>97% pure) in the absence (IC) or presence of C1q (C1q-IC) for 6 hours (microarray) or 40 hours (ELISA and flow cytometry). Controls included unstimulated and C1q alone treated cells. IFN-a and other pro-inflammatory cytokines were quantified by ELISA in culture supernatants and the expression of the activation markers CD86 and CD40 were determined by flow cytometry. Gene expression was quantified using Illumina Ref 8 beadchips and the results were confirmed by qRT-PCR with a total of 4 donors. ICs and necrotic extracts contained <0.06EU/ml endotoxin contamination.

Results:

We found that SLE ICs altered the expression of 105 genes greater than 2-fold for PBMCs and only 7 genes for purified monocytes. Surprisingly, none of the ICs tested, whether formed with purified IgG or diluted serum, induced a typical pro-inflammatory response (e.g. TNF-a and IL-6) by PBMCs or monocytes. Rather, 40 of the top 50 genes induced in PBMCs were known type I IFN stimulated genes (ISGs). These ISGs were not induced in our monocyte microarrays. Upon addition of C1q to ICs, we observed a striking downmodulation of the expression of the 40 ISGs. In agreement with our findings that C1q is anti-inflammatory, we found that C1q-ICs specifically induced inhibitors of the NFkB, MAPK and Syk signaling pathways >1.5 fold compared to ICs alone. SLE ICs also upregulated the activation markers CD86 and CD40 on CD14+ monocytes within PBMC cultures, but upon depletion of pDCs or when using purified monocytes, ICs were significantly less stimulatory (P<0.01) despite the fact that C1q strongly promoted binding to, and endocytosis of ICs, by monocytes.

Conclusions:

(1) In the absence of pDCs, SLE ICs are remarkably inefficient at stimulating PBMCs or isolated monocytes. (2) The presence of C1q in SLE ICs significantly downregulates the expression of 40 ISGs in both PBMCs and monocytes. (3) The presence of C1q in SLE ICs also alters the expression of genes that may impact inflammation through modulation of the NFkB, MAPK and Syk signaling pathways.

To cite this abstract, please use the following information:
Santer, Deanna M., Elkon, Keith B.; Lupus Immune Complex Gene Expression Arrays Reveal Divergent Pro- and Anti-Inflammatory Pathways Induced Depending on the Presence of Plasmacytoid Dendritic Cells and C1q. [abstract]. Arthritis Rheum 2010;62 Suppl 10 :499
DOI: 10.1002/art.28268

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