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研究生:林明瑋
研究生(外文):Ming-Wei Lin
論文名稱:人類中性白血球中Mitogen-AvtivatedProteinKinase之訊號傳導
論文名稱(外文):Mitogen-Activated Protein Kinase Signal Transduction Pathways in Human Neutrophils
指導教授:許清玫
指導教授(外文):Ching-Mei Hsu
學位類別:碩士
校院名稱:國立中山大學
系所名稱:生物科學系研究所
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2002
畢業學年度:91
語文別:英文
論文頁數:59
中文關鍵詞:中性白血球訊號傳導
外文關鍵詞:signal transductionNF-kappa-BneutrophilfMLPPAFROSMAPK
相關次數:
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摘要

中性白血球是主要調控各種免疫發炎反應的細胞。活化MAPK及NF-kappaB調控著許多細胞內的功能,包括超氧離子 (superoxide) 的產生。而透過細菌的peptide (fMLP) 和血小板活化因子 (PAF) 來活化中性白血球的機制仍有待研究。實驗的中性白血球取自健康人的靜脈血管,並在給予fMLP或PAF 刺激前給予PKC的抑制劑GF109203X,PKC-gamma的抑制劑calphostin C,PI3K的抑制劑wortmannin,PLA2的抑制劑aristolochic acid,PLC的抑制劑U73122,SOC的抑制劑SKF96365,鈣離子螯合劑EGTA,p38 MAPK的抑制劑SB203580與MEK 1的抑制劑PD98059。以西方墨點法偵測MAPK的磷酸化,EMSA偵測NF-kappa-B的活化,及利用流式細胞儀測量超氧離子的產生。實驗結果顯示fMLP和PAF透過不同的MAPK訊號傳遞路徑來活化中性白血球。PI3K, PKC, PLA2, PLC, 和細胞外的鈣離子均影響fMLP活化ERK MAPK,而與PAF不同的是,PKC-gamma和細胞外的鈣離子並不影響fMLP活化ERK MAPK。雖然fMLP和PAF透過不同的訊號傳遞路徑來活化中性白血球,但是fMLP和PAF皆透過PI3K, PKC, PLA2, PLC,p38 MAPK和ERK MAPK來活化NF-kappa-B及超氧離子的產生。
Abstract

Neutrophils are the major cellular component of acute inflammatory response. The mechanism by which fMLP or PAF activates neutrophils is not fully elucidated. Stimulation of MAPKs and activation of NF-kappa B in neutrophils regulate various cell functions, including superoxide production. Neutrophils isolated from blood taken from healthy donors, were incubated with specific inhibitors, GF109203X (PKC inhibitor), calphostin C (PKC-gamma isoform inhibitor), wortmannin (PI3K inhibitor), U73122 (PLC inhibitor), aristolochic acid (PLA2 inhibitor), SKF96365 (SOC channel inhibitor), EGTA (extracellular calcium chelator), SB203580 (p38 MAPK inhibitor), and PD98059 (MEK inhibitor), followed by fMLP or PAF treatment. MAPK activation by fMLP or PAF is based on immunoblot analysis. NF-kappa B activation is detected by EMSA, and superoxide production is measured by flow cytometry. The data indicate that neutrophil MAPK signaling pathways mediated by fMLP and PAF are different. PAF-induced ERK MAPK phosphorylation was involved PI3K, PKC, PLA2, PLC, and extracellular calcium, wheres fMLP-induced phosphorylation doesn’t involve PKC�n���nisoform and extracellular calcium. Although neutrophil MAPK activation by fMLP and PAF involved different pathways, NF-kappa B activation and superoxide production were stimulated in similar fashions, suggesting that PKC, PLA2, PI3K, PLC, p38 and ERK MAPK are all participated in NF-kappa B activation and superoxide production induced by fMLP or PAF stimulation in human neutrophils.
Contents
Abstract in Chinese....................1
Abstract in English....................2
Review.................................3
Introduction...........................8
Method ....................10
Results ....................13
Discussion ....................16
Figure ....................24
Materials ....................44
References ....................45
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