Genetically Modifying Lactobacillus to Alter Gut Inflammation and Pathogens

本项目的重点是表征1型干扰素（IFN）在乳酸菌中的表达，并确定给予对照和转基因乳酸菌对葡聚糖硫酸钠诱导的肠道炎症的影响。我们成功地证明了表达干扰素的细菌而不是对照细菌，可以触发STAT1的快速磷酸化，并且这种作用需要细菌和巨噬细胞之间的接触。此外，这种作用依赖于干扰素受体，但不需要toll受体2或9。乳酸菌对干扰素基因的表达也诱导了原代小鼠树突状细胞和小鼠肠上皮细胞系中STAT1的快速磷酸化。我们还证实了转基因细菌直接分泌ifn - β。目前关于细菌对DSS诱导的肠道炎症的影响的研究已经取得了意想不到的结果。我们观察到，产生ifn - β的乳杆菌的管理实际上恶化了DSS诱导的结肠炎，我们目前的数据支持ifn - β表达干扰肠道树突状细胞成熟的模型，导致更促炎的环境。作为该项目的第二部分，我们开发了一种鼠诺如病毒的PCR检测方法，并确定在感染后至少10-12天的粪便颗粒中可以检测到诺如病毒。我们现在正在测试注射乳酸杆菌是否能改变感染的时间进程。作为本项目的新内容，我们发现乳酸菌分泌一种趋化活性，我们正在对培养上清进行纯化，试图鉴定其中的活性成分。

This project has focused on characterizing the expression of the Type 1 interferon (IFN) by Lactobacillus and determining the consequences of administering control and transgenic Lactobacillus on dextran sodium sulfate induced gut inflammation. We successfully demonstrated that bacteria expressing the interferon but not the control bacteria, triggered rapid STAT1 phosphorylation and that this effect required contact between the bacteria and the macrophages. Furthermore, this effect was dependent on the interferon-beta receptor but did not require toll receptors 2 or 9. The expression of the interferon gene by the Lactobacillus also induced rapid STAT1 phosphorylation in primary murine dendritic cells as well as a mouse gut epithelial cell line. We also demonstrated direct secretion of IFN-beta by the transgenic bacteria. Current studies on the effects of the bacteria on DSS induced gut inflammation have yielded unexpected results. We have observed that administration of IFN-beta producing Lactobacillus actually worsens DSS induced colitis and our current data supports a model where the IFN-beta expression interferes with gut dendritic cell maturation, resulting in a more pro-inflammatory environment. As a second part of this project, we have developed a PCR assay for murine norovirus and have determined that norovirus can be detected in fecal pellets for at least 10-12 days following infection. We are now testing if administration of the Lactobacillus can alter the time course of infection. As a new part of this project, we have found that the Lactobacillus appear to secrete a chemotactic activity and we are in the process of purifying culture supernatant in an attempt to identify the active component.