26 CHAPTER 2 Materials and methods Patients We included 13 RA patients meeting the 2010 American College of Rheumatology/ European League Against Rheumatism Classification Criteria for RA who had active disease (disease activity score evaluated in 28 joints >3.2) [26]. All but one patient were autoantibody positive (anticyclic citrullinated peptide test >25 kAU/l and/or IgM rheumatoid factor >12.5 kU/l). All patients were typed for HLA class II alleles (Supplemental Table 1). Two patients were treated with a biological at the time of arthroscopy (infliximab and rituximab; last infusion 1 month before sampling). We did not observe a significant difference in the number of TCRβ clones, the number of highly expanded TCRβ clones (HECs), or the impact of these HECs on the total TCR repertoire between the different types of treatment (data not shown). More details on patient characteristics are shown in Table 1. From 10 patients, ST biopsy specimens were taken from either one (n = 1) or two inflamed joints (n = 9), all of which were paired with PB. In seven of these patients, we also collected SF from the same joint on which biopsy was performed prior to the arthroscopy. Three additional patients were included for paired SF and PB analysis. The study was approved by the independent Medical Ethics Committee of the Academic Medical Center/University of Amsterdam and performed according to the Declaration of Helsinki. All patients gave written informed consent. Sampling of synovial biopsy specimens, SF, and PB To obtain ST biopsy specimens, a minimally invasive arthroscopy was performed from a clinically inflamed knee or ankle, as described previously [27]. ST biopsy specimens from 11 inflamed knee joints were taken from two locations: the infrapatellar (IP) and the suprapatellar (SP) regions. If biopsy was performed on multiple joints, this was done within the same day. SF was obtained by arthrocentesis. In case of a combined collection of ST and SF, the SF was collected prior to the arthroscopy to avoid contamination of SF by hemorrhagic fluid. In all patients, PB was drawn at the time of the arthroscopy and/or arthrocentesis.
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