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.


Multiple Expanded B-Cell Clones Are Already Present in Early Rheumatoid Arthritis: Results of a Comparison between Early and Longstanding Disease Using High Throughput Sequencing Technology.

Doorenspleet2,  Marieke E., Alivernini2,  Stefano, de Hair2,  Marjolein J., Klarenbeek2,  Paul L., Herenius2,  Marieke M., van de Sande2,  Marleen G., van Schaik1,  Barbera D.

Dept. of Clinical Epidemiology, Biostatistics and Bioinformatics Academic Medical Center-University of Amsterdam
Div. Clinical Immunology and Rheumatology Academic Medical Center-University of Amsterdam, Amsterdam, The Netherlands
Laboratory for Genome Analysis Academic Medical Center-University of Amsterdam

Background:

B-cells play an important role in the pathogenesis of rheumatoid arthritis (RA) but their exact role remains unclear. We previously found that expanded B-cell clones can be detected in inflamed synovial tissue (ST) of patients with longstanding RA. These clones were either absent or of very low frequency in peripheral blood (PB), suggesting local retention and/or proliferation in the synovium. Here we examined whether oligoclonal B-cell expansions can be detected in early stages of RA.

Objectives:

Compare expanded B-cells in paired samples (ST and PB) during different stages of disease using novel high throughput sequencing technology.

Methods:

We included 2 anti-CCP+ patients with active RA. Patient 1 (pt1) was DMARD naïve and had a disease duration less than 1 year. Patient 2 (pt2) had active disease despite methotrexate treatment and had a disease duration of more than 10 years (longstanding RA). mRNA was isolated from paired samples (ST biopsies from arthritic ankle/knee and PB, respectively). Linear amplification was performed with primers for all V(ariable)-families of the receptor heavy-chain. The amplified products contain the Complementarity Determining Region 3 (CDR3), which can be used as a 'fingerprint' for each clone. The samples were analyzed using a Genome Sequencer (454/Roche). The frequency of clones was determined by custom bioinformatics algorithms identifying the CDR3-region of each receptor (up to 1 million receptors at once). Clones with a frequency of >=1% were arbitrarily considered as highly expanded.

Results:

In the longstanding RA-patient 15 highly expanded B cell clones were detected in the ST. Of interest, we also detected 14 highly expanded clones in the ST of the early RA patient. In both patients the highest frequency observed was 7%, and none of the highly expanded clones had a clearly higher frequency than the other clones. The oligoclonal pattern observed was defined by distinctly different CDR3 sequences and use of different gene segments in both patients. In the PB samples of both patients only few expanded clones were found (5 and 6 highly expanded clones, respectively). However, most clones that were highly expanded in ST could also be detected in the PB of both patients, albeit at very low frequencies (less than 0,05% in both patients). In contrast, highly expanded clones from the PB were not at all found in ST. Comparing both patients, the highly expanded clones were all different.

Conclusion:

This is the first analysis of B-cell clones in ST and PB samples comparing RA patients with different disease duration, using novel HTS technology. Our data suggest that oligoclonal B-cell expansions can already be detected in early stages of RA. This suggests that multiple epitopes might already be involved in so-called early RA.

To cite this abstract, please use the following information:
Doorenspleet, Marieke E., Alivernini, Stefano, de Hair, Marjolein J., Klarenbeek, Paul L., Herenius, Marieke M., van de Sande, Marleen G., et al; Multiple Expanded B-Cell Clones Are Already Present in Early Rheumatoid Arthritis: Results of a Comparison between Early and Longstanding Disease Using High Throughput Sequencing Technology. [abstract]. Arthritis Rheum 2010;62 Suppl 10 :1604
DOI: 10.1002/art.29370

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