Oral Presentation Australasian Cytometry Society 41st Annual Conference

Utilising R to assist in interrogation of B cell subsets by mass cytometry in multiple sclerosis and clinically isolated syndrome patients (24186)

Felix Marsh-Wakefield 1 , Thomas Ashhurst 2 , Anna Zinger 1 , Helen McGuire 3 4 , Simon Hawke 1 , Georges Grau 1 , Prue Hart 5 , Scott Byrne 6
  1. Vascular Immunology Unit, Department of Pathology, University of Sydney, Sydenham, NSW, Australia
  2. Viral Immunopathology Laboratory, Department of Pathology, University of Sydney, Camperdown, NSW, Australia
  3. Discipline of Pathology, Charles Perkins Centre, University of Sydney, Camperdown, NSW, Australia
  4. Ramaciotti Facility for Human Systems Biology, University of Sydney, Camperdown, NSW, Australia
  5. Telethon Kids Institute, University of Western Australia, Crawley, WA, Australia
  6. Applied Medical Sciences, Westmead Institute for Medical Research, Westmead, NSW, Australia

Mass cytometry allows interrogation of more than 40 markers in a single panel, allowing very powerful analysis of numerous immune cell subsets. This leads to huge amounts of data, but analysis can be laborious due to the high number of parameters. To assist in data analysis and interpretation, the R script CAPX has been utilised, which includes clustering (FlowSOM) and visualisation (tSNE) packages in a single script. Together, these algorithms allow fast differentiation and visualisation of complex data for easier interpretation. In our context, B cells from multiple sclerosis (MS) and clinically isolated syndrome (CIS: a “pre-MS” disease) patients were analysed. CIS and MS are autoimmune diseases of the central nervous system, with MS currently treated with disease-modifying therapeutics that greatly alter circulating immune cells. This includes the use of monoclonal antibodies against CD20, which leads to the depletion of most (but not all) B cells, which has had beneficial effects for MS patients. In contrast, the use of anti-BAFF/APRIL (important B cell development cytokines) has had detrimental effects to MS patients. For these reasons, we believe there is a subset of B cells that are protective against disease that are not depleted following anti-CD20. As such, mass cytometry allowed us to interrogate a wide range of B cell subsets, and investigate changes occurring between disease states and healthy controls. Our results found an alteration in various subsets of circulating B cells between diseased and healthy controls, including changes in CD20– B cells. The changes in specific B cell subsets provides further insight into the role of B cells during CIS and MS. By utilising CAPX, we were able to digest a large amount of data in a more efficient workflow than previously achieved, in turn assisting in our investigation of B cells within MS and CIS patients.