PATHOGENESIS OF LYME NEUROBORRELIOSIS IN THE RHESUS MONKEY: STUDIES IN VITRO

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 众所周知，伯氏疏螺旋体能诱导多种细胞产生炎症介质。我们推测，在这种炎性环境中，神经细胞可能发生凋亡，导致神经疏螺旋体病。在伯氏杆菌与原代小胶质细胞共培养的实验中，我们观察到IL-6和IL-8、CCL2(MCP-1)、CCL3(MIP-1)、CCL4(MIP-1)和CCL5(RANTES)的强烈释放，但我们没有检测到诱导小胶质细胞凋亡。相比之下，与伯氏杆菌共培养的SH-SY5Y(SY)神经母细胞瘤细胞表达的炎症介质数量可以忽略不计，但仍对凋亡具有抵抗力。当SY细胞与小胶质细胞和伯氏杆菌共培养时，神经元持续发生凋亡。对这些细胞培养物进行细胞凋亡染色和细胞类型特异性标记的共聚焦显微镜成像证实，主要是SY细胞正在死亡。微阵列分析进一步表明，伯氏杆菌存在强烈的小胶质细胞介导的炎症反应，包括促炎细胞因子、Toll样受体2(TLR-2)和核因子的基因转录上调。有趣的是，在炎症信号方面表现出最深刻变化的途径是触发髓样细胞上表达的受体-1(TREM1)。与单独培养的SY细胞相比，在小胶质细胞和伯氏假单胞菌存在下培养的神经细胞中，必要的P53途径基因的转录也发生了显著的变化，这表明细胞从存活转变为准备凋亡。这些发现表明，伯氏杆菌对SY细胞没有直接毒性；相反，这些细胞通过旁观者效应在小胶质细胞产生的炎症环境中痛苦并死亡。这种神经元损伤可能最终会导致神经疏松症的神经认知症状。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Borrelia burgdorferi is known to induce the production of inflammatory mediators in a variety of cells. We hypothesized that in this inflammatory milieu neuronal apoptosis may occur, leading to neuroborreliosis. In experiments where B. burgdorferi was co-cultured in vitro with primary microglia, we observed robust release of IL-6 and IL-8, CCL2 (MCP-1), CCL3 (MIP-1¿), CCL4 (MIP-1¿) and CCL5 (RANTES), but we detected no induction of microglial apoptosis. In contrast, SH-SY5Y (SY) neuroblastoma cells co-cultured with B. burgdorferi expressed negligible amounts of inflammatory mediators but also remained resistant to apoptosis. When SY cells were co-cultured with microglia and B. burgdorferi, neuronal apoptosis consistently occurred. Confocal microscopy imaging of these cell cultures stained for apoptosis and with cell type-specific markers confirmed that it was predominantly the SY cells that were dying. Microarray analysis further demonstrated an intense microglia mediated inflammatory response to B. burgdorferi including up-regulation in gene transcripts for proinflammatory cytokines, Toll-like receptor 2 (TLR-2) and NF¿¿. Interestingly, the pathway that exhibited the most profound changes in regard to inflammatory signaling was triggering receptor expressed on myeloid cells-1 (TREM1). Significant transcript alterations in essential p53 pathway genes also occurred in neuronal cells cultured in the presence of microglia and B. burgdorferi, which indicated a shift from cell survival to preparation for apoptosis when compared to SY cells cultured in the presence of B. burgdorferi alone. These findings indicate that B. burgdorferi is not directly toxic to SY cells; rather, these cells become distressed and die in the inflammatory surroundings generated by microglia through a bystander effect. Such a neuronal impairment may eventually contribute to the neurocognitive symptoms seen in neuroborreliosis.