Daclizumab Blocks Expansion Of ThIL17 Cells in Human Peripheral Blood Mononuclear Cells (PBMC)

Abstract

Purpose:: Uveitis is a T-cell mediated intraocular inflammatory disease of presumed autoimmune etiology. Although administering IFNα//ß or inhibiting IL-2 receptor α subunit by Daclizumab is a partially effective therapy, pathogenic T-cell subtypes that cause uveitis or mechanism of IL-2 or IFNα//ß actions remain unknown. Here, we have used PBMC/CD4⁺ T cells from normal human donors to examine potential role of IL-2 in expansion of Th_IL17 cells, a newly described pathogenic T-helper subtype implicated in multiple sclerosis, inflammatory bowel disease and arthritis.

Methods:: Human PBMC was isolated from buffy coats by density gradient centrifugation. Human CD4⁺ T cells were enriched (98% purity) by RosettSep Cocktail (StemCell Tec.). PBMC and CD4⁺ T cells were cultured in medium containing various combinations of IL-2, IL-7, IL-15, IL-23, TGF-ß1/IL-6, anti-CD3/-CD28 Abs or Daclizumab for 4 days. Cytokine analysis was performed by ELISA or intracellular staining and surface phenotype by FACS analysis.

Results:: We showed: (i) Exposure of normal human PBMC/CD4⁺ T cells to exogenous IL-2 induces significant expansion of Th_IL17 cells; (ii) Analysis of NK cells, monocytes, polymorphonuclear cells, and B-cells, reveal that T cells are the sole source of IL-17 in human PBMC; (iii) Among T cell subsets examined, including, CD8⁺, CD25, CD45RO⁺ or CD45RA⁺, CD62L, CD69 CD4⁺ and γ/Δ T-cells, IL-2-induced increase in IL-17 expression is primarily in CD4⁺ CD45RO⁺ CD25⁺population; (iv) TGF-ß1 and IL-6 or IL-23 alone is not as efficient as IL-2 in expanding Th_IL17 cells in CD4⁺ T cells; (v) The IL-2-induced expansion is blocked by Daclizumab.

Keywords: immunomodulation/immunoregulation • cytokines/chemokines • signal transduction