High constitutive glucocorticoid receptor β in human neutrophils enables them to reduce their spontaneous rate of cell death in response to corticosteroids

Neutrophils are markedly less sensitive to glucocorticoids than T cells, making it difficult to control inflammation in neutrophil-mediated diseases. Development of new antiinflammatory strategies for such diseases would be aided by an understanding of mechanisms underlying differential steroid responsiveness. Two protein isoforms of the human glucocorticoid receptor (GR) exist, GR alpha and GR beta, which arise from alternative splicing of the GR pre-mRNA primary transcripts. GR beta does not bind glucocorticoids and is an inhibitor of GR alpha activity. Relative amounts of these two GRs can therefore determine the level of glucocorticoid sensitivity. In this study, human neutrophils and peripheral blood mononuclear cells (PBMCs) were studied to determine the relative amounts of each GR isoform. The mean fluorescence intensity (MFI) using immunofluorescence analysis for GR alpha was 475 +/- 62 and 985 +/- 107 for PBMCs and neutrophils, respectively. For GR beta, the MFI was 350 +/- 60 and 1,389 +/- 143 for PBMCs and neutrophils, respectively (P < 0.05). After interleukin (IL)-8 stimulation of neutrophils, there was a statistically significant increase in intensity of GR staining to 2,497 +/- 140 (P < 0.05). No change in GR expression was observed. This inversion of the GR alpha /GR beta ratio in human neutrophils compared with PBMCs was confirmed by quantitative Western analysis. Increased GR beta mRNA expression in neutrophils at baseline, and after IL-8 exposure, was observed using RNA dot blot analysis. Increased levels of GR alpha /GR beta heterodimers were found in neutrophils as compared with PBMCs using coimmunoprecipitation/Western analysis. Transfection of mouse neutrophils, which do not contain GR beta, resulted in a significant reduction in the rate of cell death when treated with dexamethasone. We conclude that high constitutive expression of GR beta by human neutrophils may provide a mechanism by which these cells escape glucocorticoid-induced cell death. Moreover, upregulation of this GR by proinflammatory cytokines such as IL-8 further enhances their survival in the presence of glucocorticoids during inflammation.