title: Improved flow cytometric assessment reveals distinct microvesicle (cell-derived microparticle) signatures in joint diseases.
abstract: BACKGROUND
  Microvesicles (MVs), earlier referred to as microparticles, represent a major type of extracellular vesicles currently considered as novel biomarkers in various clinical settings such as autoimmune disorders. However, the analysis of MVs in body fluids has not been fully standardized yet, and there are numerous pitfalls that hinder the correct assessment of these structures.
  
  
  METHODS
  In this study, we analyzed synovial fluid (SF) samples of patients with osteoarthritis (OA), rheumatoid arthritis (RA) and juvenile idiopathic arthritis (JIA). To assess factors that may confound MV detection in joint diseases, we used electron microscopy (EM), Nanoparticle Tracking Analysis (NTA) and mass spectrometry (MS). For flow cytometry, a method commonly used for phenotyping and enumeration of MVs, we combined recent advances in the field, and used a novel approach of differential detergent lysis for the exclusion of MV-mimicking non-vesicular signals.
  
  
  RESULTS
  EM and NTA showed that substantial amounts of particles other than MVs were present in SF samples. Beyond known MV-associated proteins, MS analysis also revealed abundant plasma- and immune complex-related proteins in MV preparations. Applying improved flow cytometric analysis, we demonstrate for the first time that CD3(+) and CD8(+) T-cell derived SF MVs are highly elevated in patients with RA compared to OA patients (p=0.027 and p=0.009, respectively, after Bonferroni corrections). In JIA, we identified reduced numbers of B cell-derived MVs (p=0.009, after Bonferroni correction).
  
  
  CONCLUSIONS
  Our results suggest that improved flow cytometric assessment of MVs facilitates the detection of previously unrecognized disease-associated vesicular signatures.
authors.affiliation: Department of Genetics, Cell- and Immunobiology, Semmelweis University, Budapest, Hungary.
authors.forename: Bence
authors.initials: B
authors.lastname: György
authors.forename: Tamás G
authors.initials: TG
authors.lastname: Szabó
authors.forename: Lilla
authors.initials: L
authors.lastname: Turiák
authors.forename: Matthew
authors.initials: M
authors.lastname: Wright
authors.forename: Petra
authors.initials: P
authors.lastname: Herczeg
authors.forename: Zsigmond
authors.initials: Z
authors.lastname: Lédeczi
authors.forename: Agnes
authors.initials: A
authors.lastname: Kittel
authors.forename: Anna
authors.initials: A
authors.lastname: Polgár
authors.forename: Kálmán
authors.initials: K
authors.lastname: Tóth
authors.forename: Beáta
authors.initials: B
authors.lastname: Dérfalvi
authors.forename: Gergő
authors.initials: G
authors.lastname: Zelenák
authors.forename: István
authors.initials: I
authors.lastname: Böröcz
authors.forename: Bob
authors.initials: B
authors.lastname: Carr
authors.forename: György
authors.initials: G
authors.lastname: Nagy
authors.forename: Károly
authors.initials: K
authors.lastname: Vékey
authors.forename: Steffen
authors.initials: S
authors.lastname: Gay
authors.forename: András
authors.initials: A
authors.lastname: Falus
authors.forename: Edit I
authors.initials: EI
authors.lastname: Buzás
mesh_terms: D000293:Adolescent; D000368:Aged; D001171:Arthritis, Juvenile / Q000097:blood / Q000175:diagnosis*; D015415:Biomarkers / Q000378:metabolism; D055252:Cell-Derived Microparticles; D002648:Child; D002675:Child, Preschool; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D016130:Immunophenotyping / Q000379:methods; D007223:Infant; D007592:Joint Diseases / Q000097:blood*; D008297:Male; D008854:Microscopy, Electron / Q000379:methods; D008875:Middle Aged; D053758:Nanoparticles / Q000737:chemistry; D010003:Osteoarthritis / Q000097:blood / Q000175:diagnosis*; D010641:Phenotype; D013582:Synovial Fluid / Q000378:metabolism
chemical_list: D015415:Biomarkers