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Spondyloarthritis
Proteomic and genomic profiling of plasma exosomes from patients with ankylosing spondylitis
  1. Fataneh Tavasolian1,
  2. Starlee Lively1,2,
  3. Chiara Pastrello1,2,3,
  4. Michael Tang1,
  5. Melissa Lim1,
  6. Addison Pacheco1,
  7. Zoya Qaiyum1,
  8. Enoch Yau1,
  9. Zeynep Baskurt4,
  10. Igor Jurisica1,2,3,5,6,
  11. Mohit Kapoor1,2,7,
  12. Robert D Inman1,8,9,10
  1. 1 Schroeder Arthritis Institute, Toronto Western Hospital, University Health Network, Toronto, Ontario, Canada
  2. 2 Osteoarthritis Research Program, Division of Orthopaedics, Schroeder Arthritis Institute, University Health Network, Toronto, Ontario, Canada
  3. 3 Krembil Research Institute, - Data Science Discovery Centre for Chronic Diseases, University Health Network, Toronto, Ontario, Canada
  4. 4 Department of Biostatistics, Princess Margaret Cancer Center, 610 University Ave, Toronto, Ontario, Canada
  5. 5 Departments of Medical Biophysics and Computer Science, and Faculty of Dentistry, University of Toronto, Toronto, Ontario, Canada
  6. 6 Institute of Neuroimmunology, Slovak Academy of Sciences, Bratislava, Slovakia
  7. 7 Department of Surgery, Division of Orthopaedic Surgery and Department of Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
  8. 8 Department of Medicine, Temerty Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada
  9. 9 Department of Immunology, Temerty Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada
  10. 10 Spondylitis Program, Division of Rheumatology, Schroeder Arthritis Institute, University Health Network, Toronto, Ontario, Canada
  1. Correspondence to Dr Robert D Inman; robert.inman{at}uhn.ca

Abstract

Introduction Recent advances in understanding the biology of ankylosing spondylitis (AS) using innovative genomic and proteomic approaches offer the opportunity to address current challenges in AS diagnosis and management. Altered expression of genes, microRNAs (miRNAs) or proteins may contribute to immune dysregulation and may play a significant role in the onset and persistence of inflammation in AS. The ability of exosomes to transport miRNAs across cells and alter the phenotype of recipient cells has implicated exosomes in perpetuating inflammation in AS. This study reports the first proteomic and miRNA profiling of plasma-derived exosomes in AS using comprehensive computational biology analysis.

Methods Plasma samples from patients with AS and healthy controls (HC) were isolated via ultracentrifugation and subjected to extracellular vesicle flow cytometry analysis to characterise exosome surface markers by a multiplex immunocapture assay. Cytokine profiling of plasma-derived exosomes and cell culture supernatants was performed. Next-generation sequencing was used to identify miRNA populations in exosomes enriched from plasma fractions. CD4+ T cells were sorted, and the frequency and proliferation of CD4+ T-cell subsets were analysed after treatment with AS-exosomes using flow cytometry.

Results The expression of exosome marker proteins CD63 and CD81 was elevated in the patients with AS compared with HC (q<0.05). Cytokine profiling in plasma-derived AS-exosomes demonstrated downregulation of interleukin (IL)-8 and IL-10 (q<0.05). AS-exosomes cocultured with HC CD4+ T cells induced significant upregulation of IFNα2 and IL-33 (q<0.05). Exosomes from patients with AS inhibited the proliferation of regulatory T cells (Treg), suggesting a mechanism for chronically activated T cells in this disease. Culture of CD4+ T cells from healthy individuals in the presence of AS-exosomes reduced the proliferation of FOXP3+ Treg cells and decreased the frequency of FOXP3+IRF4+ Treg cells. miRNA sequencing identified 24 differentially expressed miRNAs found in circulating exosomes of patients with AS compared with HC; 22 of which were upregulated and 2 were downregulated.

Conclusions Individuals with AS have different immunological and genetic profiles, as determined by evaluating the exosomes of these patients. The inhibitory effect of exosomes on Treg in AS suggests a mechanism contributing to chronically activated T cells in this disease.

  • Ankylosing Spondylitis
  • Arthritis
  • Autoimmune Diseases
  • Autoimmunity

Data availability statement

Data are available in a public, open access repository. Sequencing datasets used for analysis during the current study have been deposited in the National Center for Bioinformatics Information Gene Expression Omnibus repository under accession code GSE220130. The main manuscript and its accompanying supplemental files include all data created or analysed during this research

https://doi.org/10.1136/ard-2022-223791

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    Data availability statement

    Data are available in a public, open access repository. Sequencing datasets used for analysis during the current study have been deposited in the National Center for Bioinformatics Information Gene Expression Omnibus repository under accession code GSE220130. The main manuscript and its accompanying supplemental files include all data created or analysed during this research

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    Footnotes

    • Handling editor Josef S Smolen

    • Contributors This study was performed by FT under my supervision at the Krembil Research Institute. I approved the study design and had direct supervision of the experimental and analytical aspects of the study. FT, MT, ML, AP, ZQ and EY all participated in blood collection, sample processing and biobanking. All the exosome-related experiments were performed by FT. The Flow cytometry experiments and analyses were performed by FT, MT and AP. miRNA sequencing was performed by SL and FT under the supervision of MK at the Schroeder Arthritis Institute’s Centre for Arthritis Diagnostic and Therapeutic Innovation: https://www.uhn.ca/Arthritis/Research_Education/Our_Cross-Cutting_Platforms/Pages/Core_Facilities.aspx. The Bioinformatics work have performed by CP under the supervision of IJ. Statistical analysis performed by FT and ZB under the supervision of RDI. The manuscript was written by FT under the direct supervision of RDI. All authors participated in review of the manuscript. RDI accepts full responsibility for the work and/or the conduct of the study, had access to the data and controlled the decision to publish.

    • Funding This study was funded by Spondylarthritis Research Consortium of Canada (SPARCC) (SPARCC Research Pilot Program 2021); The Schroeder Arthritis Institute (The Schroeder Arthritis Institute Collaborative Pilot Grant Competition 2021); Krembil Foundation to the strategic research plan fund (2021 Spring Krembil Postdoctoral /Clinical Research Fellowship Award); Arthritis Society Canada Award (Postdoctoral Fellowship Salary Award); The Schroeder Arthritis Institute (The Schroeder Arthritis Institute Collaborative Pilot Grant Competition 2022).

    • Competing interests None declared.

    • Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.

    • Provenance and peer review Not commissioned; externally peer reviewed.

    • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.

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