Zhang Zhiren, Guo Ketai, Schluesener Hermann J
Institute of Brain Research, University of Tuebingen, Calwer Str. 3, D-72076 Tuebingen, Germany.
J Neuroimmunol. 2005 Apr;161(1-2):68-77. doi: 10.1016/j.jneuroim.2004.12.009.
Oligonucleotides (ODN) with hexameric motifs containing central unmethylated CpG dinucleotides are immunostimulatory. Also ODN with continuous guanosines (polyG motif) show a wide range of immunological activity. Depending on the position, the chemical property of the ODN backbone and the cell type, polyG motifs have either an enhancing or a suppressing effect on the immunostimulatory activity of the CpG-ODN. Microglial cells are central components of the innate immune system of the brain and are activated by CpG-ODN in vitro and in vivo. Here we present the analysis of the immunomodulatory effects of CpG-ODN carrying a polyG motif on the microglial cell line N9. Our data show that N9 cells express Toll-like receptor 9 (TLR9) and are activated by CpG-ODN, which leads to expression of interleukin-12p40 (IL12p40), tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS). A 3'-end polyG motif inhibits phosphothioate (PS) CpG-ODN immunostimulatory activity but enhances the immunostimulatory activity of phosphodiester (PE) CpG-ODN. Correspondingly, a 3'-end polyG motif improves the cellular uptake of PE CpG-ODN but does not change their cellular distribution pattern. Furthermore, PE CpG-ODN with a 3'-end polyG motif interact with a much higher number of cellular proteins than PE CpG-ODN. These data indicate that the 3'-end polyG motif could enhance the immunostimulatory activity of PE CpG-ODN in microglial N9 cells through increasing interaction with cellular proteins. Therefore PE CpG-ODN containing a 3'-end polyG motif resulting in increased immunostimulatory activity might be promising alternate analogues for studies in the central nervous system.
含有中心未甲基化CpG二核苷酸的六聚体基序的寡核苷酸具有免疫刺激作用。同样，具有连续鸟苷的寡核苷酸（多聚G基序）也表现出广泛的免疫活性。根据位置、寡核苷酸骨架的化学性质和细胞类型，多聚G基序对CpG-ODN的免疫刺激活性具有增强或抑制作用。小胶质细胞是大脑固有免疫系统的核心组成部分，在体外和体内均可被CpG-ODN激活。在此，我们展示了携带多聚G基序的CpG-ODN对小胶质细胞系N9的免疫调节作用分析。我们的数据表明，N9细胞表达Toll样受体9（TLR9）并被CpG-ODN激活，这导致白细胞介素-12p40（IL12p40）、肿瘤坏死因子-α（TNF-α）和诱导型一氧化氮合酶（iNOS）的表达。3'-末端多聚G基序抑制硫代磷酸酯（PS）CpG-ODN的免疫刺激活性，但增强磷酸二酯（PE）CpG-ODN的免疫刺激活性。相应地，3'-末端多聚G基序提高了PE CpG-ODN的细胞摄取，但不改变其细胞分布模式。此外，具有3'-末端多聚G基序的PE CpG-ODN与比PE CpG-ODN数量多得多的细胞蛋白相互作用。这些数据表明，3'-末端多聚G基序可通过增加与细胞蛋白的相互作用来增强PE CpG-ODN在小胶质N9细胞中的免疫刺激活性。因此，含有3'-末端多聚G基序从而导致免疫刺激活性增加的PE CpG-ODN可能是用于中枢神经系统研究的有前景的替代类似物。
