Dysregulated CD25 and cytokine expression by γδ T cells of systemic sclerosis patients stimulated with cardiolipin and zoledronate

Objectives: γδ T cells, a non-conventional innate lymphocyte subset containing cells that can be activated by lipids and phosphoantigens, are abnormally regulated in systemic sclerosis (SSc). To further evaluate the significance of this dysregulation, we compared how exposure to an autoantigenic lipid, cardiolipin (CL), during co-stimulation with an amino-bisphosphonate (zoledronate, zol), affects the activation and cytokine production of SSc and healthy control (HC) γδ T cells. Methods: Expression of CD25 on Vγ9⁺, Vδ1⁺, and total CD3⁺ T cells in cultured peripheral blood mononuclear cells (PBMCs), their binding of CD1d tetramers, and the effect of monoclonal antibody (mAb) blockade of CD1d were monitored by flow cytometry after 4 days of in vitro culture. Intracellular production of IFNγ and IL-4 was assessed after overnight culture. Results: Percentages of CD25⁺ among CD3⁺ and Vδ1⁺ T cells were elevated significantly in short-term cultured SSc PBMC compared to HC. In SSc but not HC, CL and zol, respectively, suppressed %CD25⁺ Vγ9⁺ and Vδ1⁺ T cells but, when combined, CL + zol significantly activated both subsets in HC and partially reversed inhibition by the individual reagents in SSc. Importantly, Vδ1⁺ T cells in both SSc and HC were highly reactive with lipid presenting CD1d tetramers, and a CD1d-blocking mAb decreased CL-induced enhancement of %SSc CD25⁺ Vδ1⁺ T cells in the presence of zol. %IFNγ⁺ cells among Vγ9⁺ T cells of SSc was lower than HC cultured in medium, CL, zol, or CL + zol, whereas %IFNγ⁺ Vδ1⁺ T cells was lower only in the presence of CL or CL + zol. %IL-4⁺ T cells were similar in SSc and HC in all conditions, with the exception of being increased in SSc Vγ9⁺ T cells in the presence of CL. Conclusion: Abnormal functional responses of γδ T cell subsets to stimulation by CL and phosphoantigens in SSc may contribute to fibrosis and immunosuppression, characteristics of this disease.