Hidetoshi Takahashi, Akemi Ishida-Yamamoto, Hajime Iizuka

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Presentation transcript:

Ultraviolet B Irradiation Induces Apoptosis of Keratinocytes by Direct Activation of Fas Antigen Hidetoshi Takahashi, Akemi Ishida-Yamamoto, Hajime Iizuka Journal of Investigative Dermatology Symposium Proceedings Volume 6, Issue 1, Pages 64-68 (November 2001) DOI: 10.1046/j.0022-202x.2001.00020.x Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Effects of UVB irradiation on cultured keratinocytes from p53-knockout and MRL/lpr mice. The cultured keratinocytes (1 × 106) derived from wild-type, homozygous, and heterozygote p53 deficient mice and MRL/lpr mice were irradiated by UVB (600 mJ per m2). The keratinocytes were then cultured for 24 h and viable cells were counted. p53(+/+), p53(–/–), p53(–/+), MRL/lpr indicate wild-type, homozygous, heterozygous p53 deficient mice, and MRL/lpr mice, respectively. Data are expressed as a percentage of viable cells + SD (n = 4). *p < 0.05 compared with wild-type (p53(+/+)) mice. **p < 0.01 compared with wild-type (p53(+/+)) mice. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Effects of UVB irradiation on p53 knockout and MRL/lpr mice epidermis. The epidermis of wild-type, homozygous, heterozygous p53 deficient mice, and MRL/lpr mice, were irradiated by UVB (1250 J per m2). After 24 h, mice were sacrificed, the skin was resected and hematoxylin and eosin staining was performed. p53(+/+), p53(–/–), p53(–/+), and MRL/lpr indicate wild-type, homozygous, heterozygous p53 deficient mice, and MRL/lpr mice, respectively. Arrows indicated apoptotic keratinocytes. Scale bar: 50 µm. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Effects of UVB irradiation on caspase 3 and 8 activities. SVHK cells (1 × 106) were irradiated by UVB (600 J per m2). Then SVHK cells of caspase activities of indicated time were measured. Open and closed circles indicate the activities of caspase 8 and caspase 3, respectively. Data are the means + SD (all n = 4 dishes). *p < 0.05 compared with control SVHK cells. **p < 0.01 compared with control SVHK cells. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Induction of apoptosis following UVB irradiation. SVHK cells were irradiated by 600 J per m2 UVB. SVHK cells were then cultured for the indicated time and viable cells were counted. Data are the means + SD (all n = 4 dishes). *p < 0.01 compared with control SVHK cells. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 Effects of caspase 3 and 8 inhibitors on UVB-induced apoptosis. SVHK cells (1 × 106) were pretreated with caspase 3 or 8 inhibitor (10 µM) for 1 h. Following the preincubation, cells were irradiated with UVB (600 J per m2). Then SVHK cells were cultured for 24 h and viable cells were counted. Data are the means + SD (all n = 4 dishes). *p < 0.01 compared with control SVHK cells. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 6 Fas antigen expression pattern following UVB irradiation or treatment of agonistic anti-Fas antibody. SVHK cells were preincubated in the presence or absence of antagonistic anti-Fas antibody, ZB4 (10 µg per ml), for 1 h. Cells were then either exposed to UVB (600 J per m2) or stimulated with agonistic anti-Fas antibody, CH11 (1 µg per ml). Two hours later, the expression of Fas antigen was visualized. (A) untreated cells; (B) CH11 treated cells; (C) UVB irradiated cells; (D) CH11 stimulation of ZB4-pretreated cells; (E) UVB irradiation of ZB4-pretreated cells. Scale bar: 5 µm. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 7 Effects of membrane bounded or soluble Fas ligand on UVB-irradiated SVHK cells. SVHK cells were irradiated by 600 J per m2 UVB, and cultured for the indicated time. (A) Cell lysates of UVB-irradiated SVHK cells were extracted and western blot analysis was performed. (B) The concentration of soluble Fas ligand was measured. Data are the means + SD (all n = 4 dishes). *p < 0.01 compared with control SVHK cells. Journal of Investigative Dermatology Symposium Proceedings 2001 6, 64-68DOI: (10.1046/j.0022-202x.2001.00020.x) Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions