小鼠肺部幹細胞對於流感病毒和單純疱疹病毒第一型之感受性測試及其機制探討

Abstract

幹細胞處在發育最早期的階段，是一種原始且未經分化的細胞，其具有自我新生及多潛能分化性的特徵。在個體內，幹細胞負責各種組織和器官的更新與修復。2006年，林泰元老師自新生小鼠的肺部分離到一種帶有Oct-4標幟的肺部幹細胞，他們發現此種幹細胞表面帶有SARS病毒的受體，而且能夠支持病毒在此細胞中進行複製。2007年，香港大學在因SARS病毒感染而死亡的病人肺部發現被SARS感染的是一群帶有Oct-4肺部幹細胞，所以推測肺部幹細胞很有可能是SARS病毒主要攻擊的目標，因而造成嚴重的肺部感染。 除了上述報導外，其他關於肺部幹細胞和病毒有關的報導並不多，因此本篇論文的研究目的即是測試肺部幹細胞對於流感病毒和單純疱疹病毒的感受性，再進一步了解其感染的相關機制。 首先，藉由觀察感染後細胞病變產生的情況來做初步的判定。肺部幹細胞只有在高的流感病毒感染劑量下，才會產生明顯的細胞病變。利用免疫螢光染色的方法確認在受感染細胞的位置，確實有病毒蛋白的表現。在病毒生長時程實驗中發現，肺部幹細胞受到感染後病毒力價有一百倍的上升，且相較於MDCK細胞，克流感在此細胞中抑制病毒的效果較不顯著。在單純疱疹病毒方面，我們觀察到肺部幹細胞在受到感染的短時間內，即可產生嚴重的細胞病變，而且釋放出的病毒量比HEp2細胞高出一百倍以上。透過進一步分析後發現，疱疹病毒進入肺部幹細胞的效率比HEp2細胞高出兩倍，而病毒DNA複製的效率更高出HEp2細胞七倍以上。 由以上的實驗結果得知，流感病毒和單純疱疹病毒都能夠感染小鼠肺部幹細胞，而且肺部幹細胞能夠支持病毒在其中複製，並造成細胞的破壞。而肺部幹細胞在受到病毒感染後，對於其修復肺部能力的影響則須進一步的實驗來證明。

Stem cells are characterized of self-renewal and multipotent differentiation, have been implicated to repair the injured organ and tissue. In 2006, Ling et al. established a primary culture system to generate mouse pulmonary Oct-4+ stem/progenitor cells, which can support SARS-CoV infection. In 2007, Chen et al. further reported the colocalization of Oct-4 and SARS viral protein in the SARS-infected cells in lung autopsy of SARS-infected patients. In this study, we aimed at determining the susceptibility of mouse pulmonary stem/progenitor cells to influenza virus and herpes simplex virus type 1(HSV-1), and study the mechanisms of virus infection. First, cytopathic effects were immediately observed in influenza virus infected stem/progenitor cells at 24hr. The expression of in stem cell colony and the gradual increase of release infectious virus particles in the supernatant, indicated that the stem cells died support the replicate of influenza virus. The susceptibility of influenza virus to Oseltamivir in stem cells is not as significant as those observed in MDCK cell lines. Next, we found that stem cells are more sensitive to HSV-1 infect than HEp2 cell lines and cytopathic effects can be observed at 16hr post infection. The efficient replication of HSV-1 in stem cells is likely due to more efficient viral entry and DNA replication. The pulmonary stem/progenitor cells can be infected with influenza virus and HSV-1. The influences of viral infection in pulmonary stem/progenitor cells, such as repair of injured lung, require further analysis.