Glucocorticoids induce basophil apoptosis

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Glucocorticoids induce basophil apoptosis Chitose Yoshimura, MDa,d, Misato Miyamasu, PhDa, Hiroyuki Nagase, MDb, Motoyasu Iikura, MD, PhDa, Masao Yamaguchi, MD, PhDa, Oichi Kawanami, MD, PhDe, Yutaka Morita, MD, PhDb, Tsutomu Iwata, MD, PhDd, Kazuhiko Yamamoto, MD, PhDa, Koichi Hirai, MD, PhDc Journal of Allergy and Clinical Immunology Volume 108, Issue 2, Pages 215-220 (August 2001) DOI: 10.1067/mai.2001.116575 Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 1 Time course of cell status in human basophils. Purified basophils (4 × 104/200 μL) were cultured in medium alone (○; n = 8), in the presence of 300 pmol/L IL-3 (▵; n = 8), or in the presence of 10–7 mol/L DEX (•; n = 7). Cell status was analyzed by double-staining through use of annexin V and PI. Live cells are indicated as annexin V-negative/PI-negative (A ), whereas early apoptotic cells are determined as annexin Vpositive/PI-negative (B ). Data are expressed as percentages of total cell numbers. **P < .01,*P < .05 versus medium alone. Journal of Allergy and Clinical Immunology 2001 108, 215-220DOI: (10.1067/mai.2001.116575) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 2 Determination of DNA fragmentation in cultured basophils. Basophils (1 × 105/200 μL) were cultured for 48 hours in medium alone or in the presence of 300 pmol/L IL-3 or 10–7 mol/L DEX. Cells were then fixed in ethanol, and DNA fragmentation was analyzed by means of flow cytometry through use of PI-staining. Data are representative of 6 independent analyses, which showed similar results. Journal of Allergy and Clinical Immunology 2001 108, 215-220DOI: (10.1067/mai.2001.116575) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 3 Morphology of basophils. Photomicrographs show freshly isolated (A ) and 72-hour–cultured (B ) basophils. Cytospin preparations stained with May-Grünwald-Giemsa (original magnification ×1000) representative of 5 different experiments. Journal of Allergy and Clinical Immunology 2001 108, 215-220DOI: (10.1067/mai.2001.116575) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 4 Effects of various steroids on basophil apoptosis. Basophils (4 × 104/200 μL) were incubated with serially diluted DEX, hydrocortisone (HC , 10–7 mol/L), testosterone (Test , 10–7 mol/L), progesterone (Prog , 10–7 mol/L), or estradiol (Est , 10–7 mol/L) for 72 hours. The steroid vehicle corresponding to the highest concentrations of ethanol was also tested (Vehicle ). Live cells are indicated as annexin Vnegative/PI-negative (A ), whereas early apoptotic cells are determined as annexin V-positive/PI-negative (B ). **P < .01, *P < .05 versus medium alone; n = 4, except for hydrocortisone (n = 5). Journal of Allergy and Clinical Immunology 2001 108, 215-220DOI: (10.1067/mai.2001.116575) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 5 Effects of incremental concentrations of IL-3 on DEX-induced apoptosis of basophils. Purified basophils were incubated with the indicated concentrations of IL-3 in the presence or absence of DEX (10–7 mol/L). Cell status was analyzed by double-staining through use of annexin V and PI. Live cells are indicated as annexin V-negative/PI-negative (A ), whereas early apoptotic cells are determined as annexin V-positive/PI-negative (B ). Data are expressed as percentages of total cell numbers. **P < .01,*P < .05 versus cells cultured without DEX (n = 5). Journal of Allergy and Clinical Immunology 2001 108, 215-220DOI: (10.1067/mai.2001.116575) Copyright © 2001 Mosby, Inc. Terms and Conditions