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A3.11 Immunosuppressive Effects of Glucocorticoids and Regulatory T Cells on CD28null T Cells in Vitro
  1. Jayesh Pandya,
  2. Mohammad Shahadat Hossain,
  3. Ingrid E Lundberg,
  4. Vivianne Malmström
  1. Rheumatology Unit, Department of Medicine, Karolinska University Hospital, Solna, Karolinska Institutet, Stockholm, Sweden

Abstract

Background and Objectives CD28null T cells are terminally differentiated T cells lacking CD28 co-receptor. These cells display properties of proinflammatory killer cells and are suggested to be resistant to apoptosis in vivo. Frequencies of CD28null T cells are increased in various chronic, inflammatory diseases. CD28null T cells dominate both in the affected muscle and peripheral blood of patients with idiopathic inflammatory myopathies (myositis), suggesting a role these cells in disease mechanism and muscle pathology. Recently, it was found in our lab that after conventional glucocorticoid treatment, the relative number of regulatory T cells (Tregs) was unchanged or decreased, while the CD28null T cell proportion was mainly increased in muscle tissue of myositis patients. This lead to our working hypothesis that CD28null T cells are resistant to immunosuppression mediated by glucocorticoids in the setting of myositis. Such resistance could also be against Tregs mediated immunosuppression due to distinct phenotype of CD28null T cells. The aim of this study was to evaluate the immunosuppressive effects of glucocorticoids and Tregs on CD28null T cells in an in vitro system.

Method Peripheral blood mononuclear cells (PBMCs) were obtained from 3 healthy individuals using Ficoll separation. CD3+CD4+ CD25++(high) cells were sorted as Tregs by flow cytometry. For glucocorticoid or Tregs mediated T cell suppression assays, PBMCs were stimulated with plate bound α-CD3 antibody in presence of 4uM glucocorticoid (methyl prednisolone sodium succinate) or optimal proportion of Tregs. Up-regulation of the early activation marker CD69 was measured by flow cytometry. Suppression was estimated based on % reduction in CD69 mean fluorescent intensity compared to stimulated control cells.

Results CD4+CD28null T cells (median % suppression: 40.1%) displayed lower sensitivity towards glucocorticoid-mediated suppression compared to CD28+ counterparts (median: 54.7%), seen in all individuals tested. Similarly, CD4+CD28null T cells (median: 17.5%) were less sensitive towards Tregs mediated suppression compared to CD28+ counterparts (median: 34.4%) in all individuals. No clear trend could be observed in CD8 compartment so far.

Conclusions Although, more individuals need to be tested, the above in vitro data support our in vivo findings that CD28null T cells are relatively resistant to glucocorticoid and Tregs mediated immunosuppression. Lower sensitivity of CD28null T cells towards glucocorticoid and Tregs mediated suppression support their treatment resistance nature in myositis and a role in chronic inflammation and autoimmunity.

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