論文名稱:

單純皰疹病毒第ㄧ型對於人類口腔細胞中第ㄧ型干擾素、介白素-1

 

β和介白素-6的釋放與MyD88蛋白質依賴路徑的影響

 

Effects of herpes simplex virus type 1 on the release of

 

type I interferon, interleukin-1β and interleukin-6 and

 

the MyD88-dependent pathway in human oral cells

研究生:

梁維哲  Wei-Zhe Liang

指導教授:

洪善鈴  Shan-Ling Hung

 

洪善鈴  Shan-Ling Hung

學位類別(Degree):

碩士

學校名稱(School):

國立陽明大學

系所名稱(Department):

口腔生物研究所

學號(Student Number):

39517005

學年度(Academic Year):

96

語文別(Language):

中文

 

中文

出版年(Thesis Year):

97

關鍵字:

第一型干擾素  type I IFN

 

牙齦纖維母細胞  gingival fibroblast

 

口腔鱗狀細胞上皮癌細胞株  oral carcinoma 3

 

單純皰疹病毒第一型  herpes simplex virus type 1

 

MyD88依賴路徑  MyD88 dependent pathway

 

phosphatidylinositol 3 kinase/Akt及nuclear factor-kappa B活化路徑  phosphatidylinositol 3 kinase /Akt and nuclear factor-kappa B signaling pathway

全文說明(Fulltext description):

(本論文 20100716 對校內公開)

  論文頁數(Page):

107

摘要:

第ㄧ型的干擾素(type I interferon, IFN)包括了干擾素α(IFN-α)和干擾素β(IFN-β),兩者在人體先天免疫系統中,具有抗病毒的作用。而介白素-1β (interleukin-1β, IL-1β)和介白素-6interleukin-6, IL-6)乃屬於發炎反應的細胞激素(cytokine),當宿主受到外來病原感染時,IL-1β和IL-6常常會在感染引起的部位被偵測到。先前的研究指出,單純皰疹病毒第ㄧ型(herpes simplex virus type 1, HSV-1)感染牙齦纖維母細胞(gingival fibroblast, GF)和口腔上皮細胞(oral epithelial cell),會誘發發炎性蛋白的產生,但對於HSV-1 造成type I IFNIL-1β和IL-6的分泌及調控影響,仍不清楚。本研究主要目的乃探討HSV-1對於人類口腔細胞中IFN-α、IFN-β、IL-1β和IL-6的釋放以及MyD88依賴路徑的影響,利用酵素連結免疫吸附定量法(enzyme-linked immunosorbent assay, ELISA),探討HSV-1感染GF和口腔鱗狀細胞上皮癌細胞株(oral carcinoma 3, OC3),分泌IFN-α、IFN-β、IL-1β和IL-6的情況,並利用西方墨點法(Western blotting analysis),探討HSV-1感染對MyD88蛋白質的影響。進一步利用phosphatidylinositol 3 kinase (PI3K)抑制劑(LY294002Wortmannin)、Akt抑制劑(Triciribine)以及nuclear factor-kappa B (NF-κB)抑制劑(pyrollidine dithiocarbamate, PDTC)和(N-acetyl-L-cysteine, NAC),探討MyD88依賴路徑中的PI3K/AktNF-κB轉錄因子對於HSV-1造成GF細胞或OC3細胞株分泌較多的細胞激素是否有關聯。本論文結果顯示:(1) HSV-1感染GF細胞和OC3細胞株,兩者細胞分泌IFN-α的情形並無明顯的變化,IFN-β和IL-6的分泌卻有上升的趨勢,而IL-1β的分泌情形在OC3細胞株中有上升的趨勢,但在GF細胞中卻無明顯的變化;(2) HSV-1感染GF細胞和OC3細胞株,會增強MyD88蛋白質的表現量;(3) PI3KAktNF-κB抑制劑能抑制HSV-1感染口腔細胞所造成IFN-β、IL-1β或IL-6上升之現象。因此,由本論文得知,HSV-1感染能誘發GF細胞IFN-β和IL-6的分泌上升,且誘發OC3細胞株IFN-β、IL-1β和IL-6的分泌上升,而兩者細胞受HSV-1感染後皆能增加MyD88蛋白質的表現。此外這些細胞激素分泌量的上升和MyD88依賴路徑中PI3K/AktNF-κB活化路徑可能有關。

 

Type I interferon (IFN) includes IFN-α and IFN-β which have antiviral effects in human immune system. Interleukin-1β (IL-1β) and IL-6 belong to inflammatory cytokines. When host is infected by pathogens, IL-1β and IL-6 can be detected at infected site. Previous studies showed that infection of human gingival fibroblasts (GF) and oral epithelial cells with herpes simplex virus type 1 (HSV-1) induced secretion of inflammatory proteins. However, little is known about the effects of HSV-1 on secretion of type I IFN, IL-1β and IL-6 and possible mechanisms involved in the regulation. The main purpose of this study was to determine the effects of HSV-1 infection on the release of IFN-α, IFN-β, IL-1β and IL-6 and MyD88 protein-dependent pathway in human oral cells. Whether infection of GF and oral carcinoma 3 (OC3) cells with HSV-1 affected secretion of IFN-α, IFN-β, IL-1β and IL-6 was determined using the enzyme-linked immunosorbent assay (ELISA). The effects of HSV-1 infection on the expression of MyD88 protein were examined using Western blotting analysis. Whether phosphatidylinositol 3 kinase (PI3K)/Akt and nuclear factor-kappa B (NF-κB), factors in the MyD88-dependent pathway, were involved in the stimulating effects of cytokine secretion in GF and OC3 cells were determined using PI3K inhibitors (LY294002 and Wortmannin), Akt inhibitor (Triciribine) and NF-κB inhibitors (pyrollidine dithiocarbamatePDTCand N-acetyl-L-cysteineNAC). The results were as followed. (1) In HSV-1infected GF and OC3 cells, secretion of IFN-α was not changed, but secretion of IFN-β and IL-6 were increased. However, increased secretion of IL-1β after HSV-1 infection was observed in OC3 cells, but not in GF. (2) Expression of MyD88 protein was induced by HSV-1 in GF and OC3 cells. (3) PI3K, Akt and NF-κB inhibitors inhibited the increased release of IFN-β, IL-1β or IL-6 by HSV-1 in human oral cells. In conclusion, the results suggested that HSV-1 induced the secretion of IFN-β and IL-6 in GF and secretion of IFN-β, IL-1β and IL-6 in OC3 cells. Expression of MyD88 protein was induced by HSV-1 in both GF and OC3 cells. Besides, the increased secretion of these cytokines may be through PI3K/Akt and NF-κB signaling pathway in the MyD88-dependent pathway.

論文目次
Table of Content
:

目錄

 

頁次

 

目錄……………………………………………………………………I

 

表次目錄………………………………………………………………II

 

圖次目錄………………………………………………………………III

 

中文摘要………………………………………………………………VI

 

英文摘要……………………………………………………………...VIII

 

導論……………………………………………………………............1

 

材料與方法……………………………………………………………15

 

結果……………………………………………………………………28

 

討論……………………………………………………………………37

 

表列……………………………………………………………………45

 

圖列……………………………………………………………………46

 

參考文獻………………………………………………………………73

 

附錄……………………………………………………………………92