T Cell-Intrinsic Nod2 Protects Against Th17-Mediated Autoimmune Arthritis in SKG Mice

Ruth Napier¹, Ellen Lee¹, Emily Vance¹, Paige Snow², Clare Dawson³, Kimberly Samson⁴, Amy Moran⁵, Michael Davey⁶, Shimon Sakaguchi⁷ and Holly Rosenzweig¹, ¹Molecular Microbiology and Immunology, Oregon Health & Science University, Portland, OR, ²Molecular Microbiology and Immunology, VA Portland Health Care System, Portland, OR, ³Portland VA, PORTLAND, OR, ⁴Oregon Health & Science University, Portland, OR, ⁵Cell, Developmental, and Cancer Biology, Oregon Health & Science University, Portland, OR, ⁶VA Portland Health Care System, Portland, OR, ⁷Osaka University, Osaka, Japan

Meeting: 2018 ACR/ARHP Annual Meeting

Keywords: Arthritis, Mouse model, nod-like receptor (NLR) and rheumatoid arthritis (RA), T cells

Session Information

Date: Sunday, October 21, 2018

Title: Rheumatoid Arthritis – Animal Models Poster

Session Type: ACR Poster Session A

Session Time: 9:00AM-11:00AM

Background/Purpose: Nucleotide-binding oligomerization domain-containing protein 2 (Nod2) is an innate immune receptor known primarily for its role in host protection against bacterial infections. Intriguingly, Nod2 also appears to have a role outside of infection, as a single point mutation in this molecule leads to 100% incidence of arthritis in Blau Syndrome. Although the contribution of autoreactive CD4⁺ T cells to autoimmune arthritis is appreciated, the molecules that regulate their development and propagation remain unknown. Here we sought to elucidate the role of Nod2 in the generation of autoreactive CD4⁺ T cells using SKG mice, which are genetically predisposed to arthritis.

Methods: SKG and Nod2^-/-SKG mice housed under specific pathogen-free conditions were i.p. injected with zymosan (1.5 mg) at 6-8 wk age, clinically monitored for arthritis and analyzed histologically for joint pathology. CD4⁺ T cells from ankle-draining lymph nodes of na•ve or zymosan treated mice were stimulated in vitro with PMA/io and production of pro-inflammatory cytokines was quantified by ICS and flow cytometry. IL-17 concentration in synovial fluid was determined by ELISA. The role of IL-17 in disease was assessed by in vivo neutralization with anti-IL-17A monoclonal antibody. T regulatory (Treg) cell (CD4⁺CD25⁺Foxp3⁺) frequency and ability to suppress T effector (CD4⁺CD25^–) cell proliferation was determined by Treg suppression assays and flow cytometry. For adoptive T cell transfers, splenic CD4⁺ T cells (4×10⁶) purified from na•ve mice by magnetic positive selection were injected i.v. into lymphopenic Nude (nu/nu) recipients who were injected 24h later with zymosan. Experiments were performed three independent times (n=5-6 mice/genotype) and data were analyzed using non-parametric statistics.

Results: Nod2^-/- SKG mice developed markedly worse arthritis following zymosan injection that coincided with increased numbers of pro-inflammatory Th17 (CD4⁺IL-17⁺) cells in the ankle-draining lymph nodes and elevated IL-17 protein levels in joint synovial fluid. The increase in Th17 cell frequency was likely not due to compromised Treg responses as the frequency and suppressive capacity of Tregs was not affected by Nod2 deletion. Neutralization of IL-17A in vivo attenuated arthritis in Nod2^-/-SKG mice suggesting Nod2 is a negative regulator of IL-17. Importantly, CD4⁺ T cells from na•ve Nod2^-/-SKG mice had a fundamental capacity to produce more IL-17 and had enhanced ability to passively transfer arthritis to T cell-deplete nude recipients.

Conclusion: These data uncover a previously unappreciated role for T cell-intrinsic Nod2 as an endogenous negative regulator of the Th17 response and the arthritogenicity of CD4⁺ T cells on a single cell level. Understanding how Nod2 participates in immune tolerance mechanisms could illuminate protective mechanisms in arthritis that may be exploited for the future development of novel treatments.

Disclosure: R. Napier, None; E. Lee, None; E. Vance, None; P. Snow, None; C. Dawson, None; K. Samson, None; A. Moran, None; M. Davey, None; S. Sakaguchi, None; H. Rosenzweig, None.

To cite this abstract in AMA style:

Napier R, Lee E, Vance E, Snow P, Dawson C, Samson K, Moran A, Davey M, Sakaguchi S, Rosenzweig H. T Cell-Intrinsic Nod2 Protects Against Th17-Mediated Autoimmune Arthritis in SKG Mice [abstract]. Arthritis Rheumatol. 2018; 70 (suppl 9). https://acrabstracts.org/abstract/t-cell-intrinsic-nod2-protects-against-th17-mediated-autoimmune-arthritis-in-skg-mice/. Accessed .

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