EB病毒潛伏感染對星狀細胞產生細胞激素之影響

Regulatory Effects of EBV Latent Infection on the Cytokine Production of Astrocytes

吳宜芳(I-Fang Wu)

指導教授 ： 陳美如

國立臺灣大學/醫學院/微生物學研究所/碩士(2014年)

https://doi.org/10.6342/NTU.2014.01497

摘要

EB病毒 (Epstein–Barr virus) 是一種人類皰疹病毒，主要感染B細胞和上皮細胞。目前已知大多數的感染性單核球增多症 (Infection mononucleosis, IM) 是由EBV所引起。因EBV感染IM的部分病人會產生中樞神經系統 (Central nervous system, CNS) 發炎併發症，例如腦炎和脊髓炎，在這些CNS發炎患者的腦脊髓液中，能測到EBV基因或anti-EBV的抗體。且根據文獻，當EBV存在中樞神經系統時，可能會加重由其他病原體所造成的CNS發炎或病症。而CNS中數量最多的星狀細胞，具有維持CNS生理機能的多種功能外，亦能透過釋放細胞激素 (cytokine) 等方式調控CNS的免疫反應。已知EBV能在體外感染人類星狀細胞株，因此本研究使用本實驗室建立的兩種EB病毒陽性星狀細胞株探討EB病毒感染對星狀細胞功能的影響。先以促發炎因子IL-1β或TNF-α刺激這兩種EBV陽性星狀細胞株，再以Q- PCR測定細胞中細胞激素IL-6 與IL-8 mRNA表現量的變化。在EB病毒陽性星狀細胞株U-87E以IL-1β刺激後增加了IL-6表現量，但IL-8的表現量沒有明顯改變。為了避免實驗結果是因為混合株間特性的差異，使用其他EBV 陽性細胞株U251-E、U87-E2Pl1與U87-E2Pl2來驗證IL-1β處理後的這些細胞也有EB病毒感染增強IL-6 RNA產量的影響。因為EB病毒陽性細胞株會表現潛伏期蛋白LMP1和EBNA1，以慢病毒感染的方式讓星狀細胞株U-87MG大量表現LMP1，表現LMP1的星狀細胞株會促進IL-6與IL-8的表現，再給予IL-1β刺激後，可以觀察到LMP1的表現與IL-1β刺激兩者有加乘的效果。結果顯示LMP1在EB病毒陽性星狀細胞產生細胞激素的調控上可能扮演重要的角色，這個研究對於EB病毒如何影響CNS感染而引起的的發炎有更多的了解。

關鍵字

EB病毒 ； 星狀細胞 ； 中樞神經系統 ； 發炎反應 ； 細胞激素 ； IL-1β ； IL-6

並列摘要

Epstein–Barr virus (EBV) is a human herpesvirus, and the major viral tropism is B cells and epithelial cells. Most of infectious mononucleosis (IM) is caused by EBV infection. Central nervous system (CNS) complications, such as encephalitis and myelitis occur in some IM patients. EBV genomes and anti-EBV antibodies were found in the cerebrospinal fluid (CSF) from some IM patients with CNS inflammation. According to previous studies, EBV existing in CNS most likely accentuates the illness of CNS disease or CNS inflammation. Astrocytes are the most abundant cells and have multiple physiological functions in the CNS, also can modulate immune response by secreting cytokines. EBV was known to infect human astrocyte cell lines in vitro. In this study, two EBV-positive astrocyte cell lines were established in our laboratory to characterize effects of EBV infection to astrocyte function. These two EBV-positive astrocyte cell lines were stimulated with IL-1β or TNF-α and detected for IL-6 or IL-8 mRNA expression by quantitative real-time PCR. Expression of IL-6 was enhanced with IL-1β stimulation in EBV-positive astrocyte cell lines U87-E, but no obvious change was observed in the expression of IL-8 mRNA. To avoid clonal variation, additional EBV infected pooled clones, U251-E, U87-E2Pl1 and U87-E2Pl2 were used for IL-1β treatment to demonstrate the enhancing effect of EBV infection on IL-6 mRNA production. Because EBV-positive astrocyte cell lines expressed latent protein LMP1 and EBNA1, LMP1 were overexpressed in parental astrocyte cell lines by lentiviral infection. The expression of IL-6 and IL-8 mRNA were further stimulated in LMP1 over-expressing astrocyte cell lines than control cells. Data here suggest that LMP1 may regulate the expression of IL-6 and IL-8 mRNA in astrocytes. In addition, LMP1-overexpressing parental astrocytes have synergistic effect with IL-1β in the expression of IL-6 mRNA.Data here suggest that LMP1 may play a major role in regulating cytokine production of EBV positive astrocytes. This study thus shed light on how EBV infection may affect the inflammation process of CNS infection.

並列關鍵字

EBV ； Astrocyte ； CNS ； Inflammation ； Cytokine ； IL-1β ； IL-6

參考文獻

許嘉琳，(2012)EB病毒對於星狀細胞株之感染及影響，國立台灣大學醫學院微生物所微生物與免疫學組碩士論文。

An, Y., Chen, Q. & Quan, N. (2011). Interleukin-1 exerts distinct actions on different cell types of the brain in vitro. Journal of inflammation research 2011, 11-20.

Ascherio, A. (2013). Environmental factors in multiple sclerosis. Expert review of neurotherapeutics 13, 3-9.

Ascherio, A. & Munger, K. L. (2007). Environmental risk factors for multiple sclerosis. Part I: the role of infection. Annals of neurology 61, 288-299.

Babcock, G. J., Hochberg, D. & Thorley-Lawson, A. D. (2000). The expression pattern of Epstein-Barr virus latent genes in vivo is dependent upon the differentiation stage of the infected B cell. Immunity 13, 497-506.