EB病毒感染B細胞之基因表現圖譜

Gene expression profile of EBV-infected B cells

10.6342/NTU.2011.02969

林冠廷

EB病毒 ； B細胞活化 ； 細胞標記分子 ； 細胞激酶干擾素誘導基因 ； EBV ； B cell activation ； CD markers ； cellular kinases ； interferon-stimulated genes

臺灣大學微生物學研究所學位論文

2011年

碩士

蔡錦華

繁體中文

B細胞受到EB病毒感染後會被活化，並且被轉形成具有持續增生能力之淋巴母細胞。然而其他細胞分裂原或B細胞抗原的刺激，只能促進B細胞的短暫活化。經由本實驗室之前所完成的cDNA 微陣列分析，發現多種細胞表面標記分子、細胞激酶以及干擾素誘導基因的表現量在EB病毒感染CD19+ B細胞三天後以及在LCL都受到顯著影響。本研究為了解B細胞受到EB病毒感染後與受到其它刺激活化後，細胞基因表現有何不同。利用EB病毒、CD40+IL-4、polyinosinic: polycytidylic acid (poly I:C)和lipopolysaccharide (LPS)刺激B細胞活化三天後，收取細胞進行流式細胞儀的分析，發現B細胞活化標記分子CD80的表現在所有的刺激後表現量都有顯著上升，而其他標記分子的表現在不同刺激下則有明顯差異。為了解細胞激酶和干擾素誘導基因在B細胞受到EB病毒感染和不同的活化刺激後的表現量，利用EB病毒、CD40+IL-4、poly I:C、LPS和 Staphylococcus protein A (SpA) 刺激B細胞活化三天後，經由即時定量聚合酶連鎖反應分析，發現多種細胞激酶以及干擾素誘導基因的表現都只有在EB病毒感染後會有顯著上升的情形。為了進一步了解EB病毒調控干擾素誘導基因的機制，在Akata EBV (-) 細胞株中短期表現EB病毒潛伏期膜蛋白LMP1後，以即時定量聚合酶連鎖反應分析，發現可以激發大量干擾素誘導基因的表現。表現LMP1刪除突變株亦顯示LMP1的CTAR2區段對於調控這些干擾素誘導基因是必須的，而利用NF-

Human B cells infected by Epstein-Barr virus (EBV) are activated and transformed to continuously proliferative lymphoblastoid cell line (LCL). The other B cell mitogens or antigens, however, only activate B cell activation for a limited time. Through the previous cDNA microarray analysis of our lab, the expression of various CD markers, cellular kinases and interferon-stimulated genes (ISGs) are altered significantly in human CD19+ B cells 3 days post EBV infection and in LCLs. To investigate the differences of CD markers expression profile among EBV infected B cells and the other mitogen activated B cells, the expression of the CD markers in EBV, CD40+IL-4, poly I:C and LPS infected/activated B cells are analyzed by flow cytometry. The B cell activation marker CD80 is up-regulated in all activated cells and the other CD markers are expressed in distinct level in different treatments. In addition, the mRNA expression of the cellular kinases and ISGs in EBV, CD40+IL-4, poly I:C, LPS and SpA infected/activated B cells are determined by RT-Q-PCR. Expression of a good proportion of cellular kinases and ISGs are modulated only in EBV infected B cells. In order to further dissect the regulation of ISGs by EBV, we demonstrate that transient expression of EBV latent membrane protein 1 (LMP1) in Akata EBV (-) cells significantly induced ISGs expression. Furthermore, the LMP1 C-terminal activating region 2 (CTAR2) domain is essential for ISGs induction and NF-

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