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Abstract Number: 0060

Spectral Cytometry Shows Increased CD56hi NK Cells and New HLA-DR+CD56+ Phenotypes in RA

Estelle Khairallah1, Haani Qudsi2, Jihad Ben Gabr1, Andras Perl3 and Christian Geier1, 1SUNY Upstate Medical University, Syracuse, NY, 2Norton College of Medicine, Syracuse, NY, 3SUNY, Syracuse, NY

Meeting: ACR Convergence 2024

Keywords: Dendritic cells, immunology, Inflammation, Natural Killer Cells, rheumatoid arthritis

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Session Information

Date: Saturday, November 16, 2024

Title: RA – Etiology & Pathogenesis Poster

Session Type: Poster Session A

Session Time: 10:30AM-12:30PM

Background/Purpose: Previous studies of RA patients have shown changes in NK cell populations (as defined by CD56 expression). The nature of these cellular changes remains controversial. We aim to comprehensively characterize CD56 expression in RA immune cells, including conventional NK cells and other CD56 expressing cells, such as myeloid cells.

Methods: Using flow cytometry, we studied peripheral blood mononuclear cells (PBMC) from RA patients satisfying the 2010 ACR classification criteria and matched healthy donors (n = 16 each). To comprehensively characterize potential CD56 changes in RA, we excluded only lymphocytes and monocytes (CD3−CD19−CD14−; Lineage), retaining all other CD56+ cells (including but not limited to conventional NK cells; CD56+Lin−). We measured mean fluorescence intensities (MFI) including CD56, CD16, CD11c, CD1c, CD45RA, CD163, CCR2, CCR3, CCR7, and HLA-DR. We used SPADE (Spanning-tree Progression Analysis of Density-normalized Events) on a patient with severe polyarticular RA to screen for marker combinations defining other potential CD56 expressing populations. Mann-Whitney U testing with a threshold of p < 0.05 was performed to assess for significant differences.

Results: The CD56hi NK subset was increased in RA (0.35% of live lymphocytes in RA, compared to 0.14% in health; p = 0.016) but the phenotype of CD56hi cells did not differ between RA and health (p > 0.05). SPADE clustering of CD56+Lin− showed that CD56hi NK cells were composed of five subpopulations defined by differential MFIs of CD45RA, CD16, CD163, CCR3. A separate cluster showed a CD56+HLA-DR+CD11c+CD1c+ phenotype. Total NK cells (CD56+CD3−) as a percentage of live lymphocytes were not different in RA patients (p > 0.05).

Conclusion: The total NK cell population is shifted towards CD56hi in RA which may contribute to RA pathogenesis through increased cytokine production. Other CD56 changes in RA patients include new phenotypes such as the CD56+HLA-DR+CD11c+CD1c+ population, which phenotypically resembles dendritic cells. These observations warrant further mechanistic study of a potential connection between NK cells and antigen-presenting cells in the pathogenesis of RA.

Supporting image 1

Spectral Flow Cytometry of CD56hi NK cells in health and RA. Gated on CD3−CD19−CD14− (Lineage) live lymphocytes. Representative plots of a healthy donor (A) and an RA patient with severe polyarticular RA (B). (C) Quantification of CD56hi NK cells in health and RA as percent of live lymphocytes (n=16 each).

Supporting image 2

SPADE analysis of a patient with severe polyarticular RA. (A) Circled is the CD56hi NK cell population (red) and the CD56+HLA-DR+CD11c+CD1c+ population (blue). The remaining clusters largely represent classical NK cells. Circle size represents the number of events in each cluster. (B) Overlays of MFIs for molecules defining CD56hi (red) and CD56+HLA-DR+CD11c+CD1c+ (blue) populations.


Disclosures: E. Khairallah: None; H. Qudsi: None; J. Ben Gabr: None; A. Perl: None; C. Geier: None.

To cite this abstract in AMA style:

Khairallah E, Qudsi H, Ben Gabr J, Perl A, Geier C. Spectral Cytometry Shows Increased CD56hi NK Cells and New HLA-DR+CD56+ Phenotypes in RA [abstract]. Arthritis Rheumatol. 2024; 76 (suppl 9). https://acrabstracts.org/abstract/spectral-cytometry-shows-increased-cd56hi-nk-cells-and-new-hla-drcd56-phenotypes-in-ra/. Accessed .
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