臺灣博碩士論文加值系統

　　A群鏈球菌是引起人類疾病中相當重要的病原菌，從輕微的感染到較為嚴重的感染常造成壞死性肌膜炎及鏈球菌毒性休克症候群。臨床研究報告指出，在嚴重侵入性感染的病人中有高量的發炎細胞激素與nitric oxide (NO) 的生成。Dextromethorphan (DM) 是一廣泛使用的止咳藥物，近年來的研究指出此藥物具有抗發炎作用，研究結果顯示此藥物可以降低reactive oxygen species和發炎細胞激素TNF-�捋PIL-6的產生。因此本研究探討具抗發炎特性之止咳藥物DM對A群鏈球菌的感染是否能提供保護的效果。研究結果證實，給予小鼠DM處理後確實能增加A群鏈球菌感染小鼠的存活率。進一步研究發現，DM能維持浸潤到感染部位嗜中性球的存活率並且減低感染部位的細菌量，另一方面也發現DM可避免細菌散佈到血液中，且降低血清中發炎細胞激素與趨化激素的產生。此外，觀察感染後小鼠肝臟的病理變化與血清中AST及ALT濃度也顯示DM可以減輕A群鏈球菌感染所造成的肝損傷。活體外細胞實驗的結果顯示A群鏈球菌可以誘發巨噬細胞轉錄因子NF-�羠活化，而DM可以降低NF-�羠的活化。分析轉錄因子NF-�羠下游所調控的發炎性蛋白質，結果亦證實DM可以抑制A群鏈球菌所誘發的iNOS表現與NO的產生。綜合這些研究結果，DM透過維持浸潤細胞的存活率與降低全身性發炎反應，進而提供A群鏈球菌感染的保護效果。

Group A streptococcus (GAS) is an important human pathogen that causes a wide spectrum of disorders, ranging from mild infections to severe invasive diseases such as necrotizing fasciitis and streptococcal toxic shock syndrome. Patients suffering from severe invasive infection have higher levels of inflammatory cytokines and nitric oxide (NO) compared with non-invasive infection. Dextromethorphan (DM) is a widely-used anti-tussive drug. Recent reports show that DM has anti-inflammatory properties through reducing the levels of reactive oxygen species and inflammatory cytokines including TNF-�� and IL-6. In the present study, we investigate the potential protective effect of DM in GAS infection. Our results showed that DM treatment increased the survival rate of GAS-infected mice. DM administration reduced bacterial numbers and increased neutrophil viability in local infection site. Moreover, DM treatment prevented bacterial dissemination to the blood and reduced serum levels of proinflammatory cytokines and chemokines. In addition, examination of liver histology and serum AST and ALT levels in GAS-infected mice revealed that DM reduced liver injury. Further in vitro studies showed that GAS induced NF-�羠 activation in macrophages and DM treatment reduced this activation. DM also reduced iNOS expression and NO production in GAS-infected macrophages. Taken together, DM treatment increased the viability of infiltrated neutrophils and reduced systemic inflammatory responses to provide the protective effect in GAS infection.

中文摘要 I
英文摘要 II
致謝 III
目錄 IV
表目錄 VII
圖目錄 VIII
縮寫檢索表 X

緒論 1
特定目標與實驗設計 7
材料與方法
I. 材料
A. 實驗動物 8
B. 細菌株 8
C. 細胞株 8
D. 儀器與試劑 8
E. 耗材 9
II. 方法
A. A群鏈球菌之感染小鼠模式
1. 細菌培養 10
2. 小鼠氣囊感染模式 10
3. 小鼠存活率之觀察 11
4. 皮下氣囊感染部位菌量的計數 11
5. 血液的菌量計數 11
6. 氣囊回抽液浸潤細胞之分析 11
7. 氣囊回抽液浸潤細胞的細胞凋亡分析 11
8. 器官的病理變化 12
9. 血清中的生化成分測定 12
10. 氣囊回抽液與血清中發炎細胞激素與趨化激素之測定 12
B. A群鏈球菌感染巨噬細胞模式
1. 細胞培養與保存 13
2. A群鏈球菌感染巨噬細胞株 (RAW264.7) 13
3. 轉錄因子NF-�羠活化之分析 14
4. 硝酸鹽與亞硝酸鹽測定 (Nitrate plus nitrite assay) 15
C. 統計分析 16
結果
A. A群鏈球菌感染模式中給予DM處理小鼠存活率之變化 17
B. 皮下氣囊感染部位之細菌量計數 17
C. DM對於細菌生長之影響 17
D. 皮下氣囊感染部位之免疫細胞浸潤情形 18
E. 浸潤免疫細胞之凋亡情形 18
F. 氣囊回抽液中發炎細胞激素與趨化激素的表現量 19
G. 感染後血液的細菌量計數 19
H. 感染後肝臟病理變化 20
I. 感染後肝發炎指數 20
J. 血清中發炎細胞激素與趨化激素的表現量 21
K. A群鏈球菌誘發轉錄因子NF-�羠活化之探討 21
L. DM對於A群鏈球菌誘發轉錄因子NF-�羠活化之影響 22
M. A群鏈球菌誘發巨噬細胞產生NO之探討 22
N. DM對於A群鏈球菌誘發iNOS與NO表現之影響 23
討論
A. DM對於A群鏈球菌感染的保護角色 25
B. DM的給予方式與時間點 25
C. Ａ群鏈球菌誘發浸潤細胞之細胞凋亡 26
D. DM抑制A群鏈球菌誘發轉錄因子NF-�羠活化之機轉 27
E. DM運用於革蘭氏陽性細菌感染 27
F. DM降低全身性發炎反應之探討 28
G. A群鏈球菌感染與NO產生之關係 29
H. DM與抗生素合併治療法 29
I. 總結 30
參考文獻 31
表 39
圖 40
附錄 56
自述 62

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