High constitutive glucocorticoid receptor beta 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, ma king it difficult to control inflammation in neutrophil-mediated diseases. Development of new antiinflammatory strategies for such diseases would be a ided by an understanding of mechanisms underlying differential steroid resp onsiveness. Two protein isoforms of the human glucocorticoid receptor (GR) exist, GR alpha and GR beta, which arise from alternative splicing of the G R pre-mRNA primary transcripts. GR beta does not bind glucocorticoids and i s 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 stud ied 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, respect ively. 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<beta> staining to 2,497 +/- 140 (P < 0.05). No change in GR<alpha> e xpression was observed. This inversion of the GR alpha /GR beta ratio in hu man neutrophils compared with PBMCs was confirmed by quantitative Western a nalysis. Increased GR beta mRNA expression in neutrophils at baseline, and after IL-8 exposure, was observed using RNA dot blot analysis. Increased le vels of GR alpha /GR beta heterodimers were found in neutrophils as compare d 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 neutrophi ls may provide a mechanism by which these cells escape glucocorticoid-induc ed cell death. Moreover, upregulation of this GR by proinflammatory cytokin es such as IL-8 further enhances their survival in the presence of glucocor ticoids during inflammation.