系統識別號 U0026-0812200910434973
論文名稱(中文) Fas傳遞的訊息誘發A549細胞表現IL-8及GRO-alpha機制之探討
論文名稱(英文) Fas signaling mediates the expression of IL-8 and GRO-alpha in A549
校院名稱 成功大學
系所名稱(中) 微生物及免疫學研究所
系所名稱(英) Department of Microbiology & Immunology
學年度 91
學期 2
出版年 92
研究生(中文) 張雅南
研究生(英文) Ya-Nien Chang
學號 s4690102
學位類別 碩士
語文別 中文
論文頁數 88頁
口試委員 指導教授-楊倍昌
中文關鍵字 介白素8 
英文關鍵字 Fas  IL-8  GRO-alpha 
中文摘要 Fas-L ( CD95-Ligand ) 是屬於腫瘤壞死因子家族的一員。Fas-L 作用在帶有 Fas 的細胞上而導致細胞進行細胞凋亡。Fas/FasL 的系統在免疫反應中扮演一個重要的角色。Fas-L 不僅表現在活化的 T 細胞以及自然殺手細胞上,也表現在許多不同的腫瘤細胞上。若使不表現 Fas-L 的腫瘤細胞表現 Fas-L 後有時會誘導大量的嗜中性球浸潤,引發強烈的發炎反應,使腫瘤細胞受到排斥與破壞。Fas-L 誘導嗜中性球浸潤的機制,目前還不清楚。我們推測 Fas-L 誘發嗜中性球浸潤的機制,可能與刺激內皮細胞或是上皮細胞表現趨化因子有關。我們使用 in vitro 細胞培養系統研究 Fas-L 如何影響內皮細胞或上皮細胞趨化因子的表現。人類微血管內皮細胞 HMEC-1 以及人類呼吸道上皮細胞 A549 上都能表現 Fas。以抗體活化 Fas 後,能引起 HMEC-1 細胞凋亡,但不能誘發 HMEC-1 表現 IL-8 及GRO-a。但是,A549 細胞上的 Fas 受到刺激後,不僅引起細胞凋亡,同時也能表現 IL-8 及 GRO-a。雖然性荷爾蒙可能具有調控免疫反應與基因表現的功能,但是在本研究中,性荷爾蒙對於 A549 細胞表現 IL-8 及 GRO-a 的程度並無影響。另外,當 A549 細胞在CH-11 刺激時合併處理 caspase 抑制劑:Z-VAD-fmk、NF-kB 抑制劑:SN-50、JNK 抑制劑:SP600125、MAPK/ERK 抑制劑:PD98059、p38 MAPK 抑制劑:SB202190、PKA 抑制劑:KT5720以及HSP70抑制劑後,由 CH-11 所誘發的 IL-8 及 GRO-a 表現會受到抑制。綜合以上的研究結果證明,Fas 訊號能誘發 IL-8 及 GRO-a 表現。是否它是導至免疫細胞浸潤的主因則有待進一步的研究來驗證。
英文摘要 Fas-L ( CD95-Ligand ) is a member of the TNF family. Engagement of Fas-L with Fas induces apoptosis in Fas-bearing cells. Fas-L is not only expressed on activated CTL and NK cells but also on tumors of diverse cellular organs. Recent reports suggested that forced expression of Fas-L in tumors unexpectedly induced migration of neutrophils into the tumors and the FasL-expressing tumors were rejected due to the inflammatory reaction. The mechanism of the neutrophilic response to Fas-L expression is unknown. We speculate that Fas-L can induce the chemokines expression on endothelial or epithelial cells which in turn leading to cell infiltration. Fas were detected on the surface of human microvascular endothelial cell line-1 (HMEC-1). When treated with anti-Fas antibody, CH-11, HMEC-1 cells became apoptotic, but did not express IL-8 and GRO-a at transcription and translation level. Besides, A549 cells, a human airway epithelial cell line expressing Fas on the surface, were apoptotic and expressed IL-8 and GRO-a upon CH-11 treatment. In addition, sex hormones had no effects on CH-11-induced IL-8 and GRO-a productions in A549 cells. Application of caspase inhibitor:Z-VAD-fmk, NF-kB inhibitor:SN-50, JNK inhibitor:SP600125, MAPK/ERK inhibitor:PD98059, p38 MAPK inhibitor:SB202190, PKA inhibitor:KT5720 or HSP70 inhibitor could decrease the CH-11 induced productions of IL-8 and GRO-a in A549. These data suggest that Fas signal mediates induce IL-8 and GRO-a expression in A549 cells.
論文目次 中文摘要 1
英文摘要 2
目錄 3
圖目錄 4
前言 6
實驗設計 9
儀器與試劑 11
材料與方法 14
結果 29
討論 41
References 46
Figures (結果) 53


Figure 1. Fas expression on HMEC-1 cells. 53
Figure 2. Fas expression on A549 cells. 54
Figure 3. CH-11 induce Fas-expressing HMEC-1 cells apoptosis. 55
Figure 4. CH-11 induce Fas-expressing A549 cells apoptosis. 56
Figure 5. CH-11 induce HaCaT cells apoptosis. 57
Figure 6. Anti-Fas antibody, CH-11 did not induce IL-8, GRO-a and MCP-1 protein production in HMEC-1 cells. 58
Figure 7. Anti-Fas antibody, CH-11 did not induce IL-8, GRO-a and MCP-1 mRNA expression in HMEC-1 cells. 59
Figure 8. Anti-Fas antibody, CH-11 did not induce IL-8, GRO-a protein production in HaCaT cells. 60
Figure9. Anti-Fas antibody, CH-11 induced IL-8, GRO-a but not MCP-1 protein production in A549 cells. 61
Figure 10.Anti-Fas antibody, CH-11 induced IL-8, GRO-a but not MCP-1 protein production in A549 cells. 62
Figure 11.Anti-Fas antibody, CH-11 induced IL-8, GRO-a but not MCP-1 mRNA expression in A549 cells. 63
Figure 12.CH-11 did not induce HUVEC cells apoptosis. 64
Figure 13.Anti-Fas antibody, CH-11 did not induce IL-8, GRO-a and MCP-1 protein production in HUVEC cells. 65
Figure 14.TNF-a and IL-1b induced IL-8, but not GRO-a and MCP-1 protein production in HMEC-1 cells. 66
Figure 15.TNF-a and IL-1b induced IL-8, but not GRO-a and MCP-1 mRNA expression in HMEC-1 cells. 67
Figure 16.TNF-a and IL-1b induce IL-8, GRO-a and MCP-1 protein production in HUVEC cells. 68
Figure 17.CH-11 and TNF-a did not induce ICAM expression on HMEC-1 cells. 69
Figure 18.CH-11 and TNF-a did not induce VCAM expression on HMEC-1 cells. 70
Figure 19.Estrogen receptor-a and Clucocorticoid receptor-a mRNA expression in A549 cells. 71
Figure 20.Sex hormones had no effect on CH-11-induced apoptosis in A549. 72
Figure 21.Sex hormones had no effect on CH-11-induced IL-8 and GRO-a protein production in A549. 73
Figure 22.Z-VAD-fmk inhibited CH-11-induced apoptosis in A549. 74
Figure 23.Z-VAD-fmk suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 75
Figure 24.PD98059 enhanced CH-11-induced apoptosis in A549. 76
Figure 25.PD98059 suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 77
Figure 26.SN-50 had no effect on CH-11-induced apoptosis in A549. 78
Figure 27.SN-50 suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 79
Figure 28.SP600125 had no effect on CH-11-induced apoptosis in A549. 80
Figure 29.SP600125 suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 81
Figure 30.SB202190 enhanced CH-11-induced apoptosis in A549. 82
Figure 31.SB202190 suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 83
Figure 32.KT5720 enhanced CH-11-induced apoptosis in A549. 84
Figure 33.KT5720 suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 85
Figure 34.HSP70 inhibitor enhanced CH-11-induced apoptosis in A549. 86
Figure 35.HSP70 inhibitor suppressed CH-11-induced IL-8 and GRO-a protein production in A549 cells. 87
Figure 36.Z-VAD-fmk, SP600125, SN-50 inhibited CH-11-induced IL-8 and GRO-a mRNA expression in A549 cells. 88
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