A phenotypic re-evaluation of human γδ T cells in health and disease: Is Vδ2⁽⁺⁾ T cell depletion a risk predictor for BRONJ?

Objectives: Bisphosphonate Related Osteonecrosis of the Jaw (BRONJ) is a chronic jawbone necrosis occurring in ≈ 0.1-7% of patients receiving bisphosphonate medication for osteoporosis and certain cancers. Bisphosphonates potently activate human γδ T cells and specifically the Vδ2⁽⁺⁾ subtype. The pathogenesis of BRONJ is still uncertain but to date an immune-mediated pathology has largely been ignored. The objectives of the study were to firstly re-evaluate the most appropriate Vδ2⁽⁺⁾ phenotypic markers to describe Vδ2⁽⁺⁾ subsets, secondly to determine the functional potential of these subsets, and finally to assess whether Vδ2⁽⁺⁾ T cells are implicated in BRONJ pathogenesis.
Methods: Peripheral blood mononuclear cells were isolated by density gradient separation from 63 healthy individuals. Flow cytometry was used to determine extracellular marker expression on Vδ2⁽⁺⁾ T cells with further characterisation using intracellular cytokine staining and proliferation assays. A gene microarray was subsequently performed to further characterise sorted Vδ2⁽⁺⁾ subsets. Finally, the healthy cohort was compared to 8 BRONJ and 10 long-term bisphosphonate patients without BRONJ to determine peripheral blood Vδ2⁽⁺⁾ levels and phenotype.
Results: Re-evaluation of conventional Vδ2⁽⁺⁾ T cell surface markers in healthy individuals showed that CD45RA is an unreliable marker for accurate identification of functionally-relevant subsets. An alternative marker set (CD28, CD27, and CD16) unambiguously identifies four Vδ2⁽⁺⁾ T cell subsets which can be used to subdivide human individuals into six stable “Vδ2-profiles”. Analysis of these profiles demonstrates a surprisingly wide spectrum of Vδ2⁽⁺⁾ phenotypes. Patients on long-term bisphosphonates including BRONJ patients had significantly depleted peripheral blood Vδ2⁽⁺⁾ T cells with no apparent characteristic disease signature phenotype (p < 0.01).
Conclusions: Depletion of peripheral blood Vδ2⁽⁺⁾ T cells is characteristic of BRONJ and patients on long-term bisphosphonates and may represent a useful risk predictor for BRONJ.