Quantitative and Qualitative Tracking of Expanded CD4+ T Cell Clones in Rheumatoid Arthritis Patients

Kazuyoshi Ishigaki¹, Hirofumi Shoda², Yuta Kochi³, Tetsurou Yasui⁴, Yuho Kadono⁴, Sakae Tanaka⁴, Keishi Fujio² and Kazuhiko Yamamoto¹, ¹Department of Allergy and Rheumatology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan, ²Department of Allergy and Rheumatology, The University of Tokyo, Tokyo, Japan, ³Center for Integrative Medical Sciences, RIKEN, Yokohama, Japan, ⁴Department of Orthopaedic Surgery, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan

Meeting: 2014 ACR/ARHP Annual Meeting

Keywords: T cells and rheumatoid arthritis (RA)

Session Information

Title: Rheumatoid Arthritis - Human Etiology and Pathogenesis

Session Type: Abstract Submissions (ACR)

Background/Purpose

The purpose of this study is to elucidate the characteristics of expanded CD4⁺ T cell clones (ECs) in peripheral blood and synovium of rheumatoid arthritis (RA) patients by T cell receptor (TCR) repertoire and single cell transcriptome analysis.

Methods

We obtained peripheral blood from 5 RA patients and 5 control subjects. We performed TCR repertoire analysis by combination of single cell and next-generation sequencer (NGS) in CD4⁺ T cells. We also performed single cell RNA-Seq and real time PCR analysis of ECs and non-expanded clones (NECs). Similarly, CD4⁺ T cells from synovium were analyzed in 1 patient.

Results

We detected significantly higher frequency of ECs in peripheral blood of 4 RA patients compared with controls both in naïve and memory subset of CD4⁺ T cells. Within memory CD4⁺ T cells of peripheral blood, non-Th1/Th17/Tfh subset (negative for CXCR3, CCR6 and CXCR5) contained majority of expanded clones in PBMC of 3 RA patients and synovium-infiltrating clones in 1 RA patient. Gene expression analysis of the mostly expanded CD4⁺ T cell clones in synovium of 1 RA patient and peripheral blood of 2 RA patients revealed up-regulation of GZMB, TBX21 and CD5 and down-regulation of CD28 compared with NECs. The tracking of gene expression profile of one unique EC in peripheral blood showed down-regulation of CXCR4 and CCR5 compared with that of the identical clone in synovium of the same patient.

Conclusion

ECs were more frequently observed in non-Th1/Th17/Tfh subset in peripheral blood and had gene expression profiles consistent with senescent CD4⁺ T cells in both of peripheral blood and synovium. Given that there are no reliable markers of clonal expansion, single cell transcriptome analysis is the only method to investigate gene expression profiles of expanded clones. Combined with the robustness of NGS TCR repertoire analysis, our approach should be a rational strategy to characterize ECs and might be able to identify pathogenic clones in RA.

Disclosure:

K. Ishigaki,
None;

H. Shoda,
None;

Y. Kochi,
None;

T. Yasui,
None;

Y. Kadono,
None;

S. Tanaka,
None;

K. Fujio,
None;

K. Yamamoto,
None.

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