Expression data from primary murine osteoblasts stimulated with C5a

The anaphylatoxin C5a is a potent mediator of innate immunity and promotes inflammation via its receptor C5aR1 upon complement system activation danger-associated molecular patterns. Both C5a and C5aR1 are thought to be contributing factors in inflammatory and infectious conditions of the bone. Bone fracture healing, for example, was significantly improved when applying a C5aR1-antagonist in a rodent model of severe systemic inflammation and osteoblasts were found to be target cells for C5a in this setting. Interestingly, osteoblasts up-regulate C5aR1 during osteogenic differentiation and after bone injury. Further, C5a induces inflammatory cytokines, such as IL-6, and the osteoclastogenic mediator RANKL in osteoblasts. However, the molecular mechanisms underlying C5a-C5aR1 signaling axis in osteoblasts remain unclear, and further targets of C5a are still elusive. Using microarray analysis, we analyzed intracellular events following C5aR1 activation in osteoblasts and defined up- or down-regulated genes and their belonging biological pathways. Primary osteoblasts were derived from three different male WT mice (C57BL6, aged 8-12 weeks). Those cells were osteogenically differentiated for 14 days and then each cell pool derived from a specific mouse was either left untreated (n = 3) or treated with recomninant murine C5a for 4h.

过敏毒素C5a（anaphylatoxin C5a）是固有免疫（innate immunity）的强效介质，在补体系统（complement system）激活及损伤相关分子模式（danger-associated molecular patterns）触发下，可通过其受体C5aR1（C5aR1）介导炎症反应。现有研究认为，C5a与C5aR1均参与骨骼的炎症与感染性疾病进程。例如，在重度全身性炎症（systemic inflammation）啮齿动物模型（rodent model）中，应用C5aR1拮抗剂可显著改善骨折愈合（bone fracture healing），且该实验体系下成骨细胞（osteoblasts）被证实为C5a的靶细胞。值得注意的是，成骨细胞在成骨分化（osteogenic differentiation）阶段及骨损伤后会上调C5aR1的表达。此外，C5a可在成骨细胞中诱导IL-6等炎症细胞因子及破骨细胞生成介质RANKL（osteoclastogenic mediator RANKL）的产生。然而，成骨细胞中C5a-C5aR1信号轴发挥功能的分子机制仍未明确，C5a的其他潜在靶点也尚未阐明。本研究通过基因芯片分析（microarray analysis），解析了成骨细胞中C5aR1激活后的胞内信号事件，明确了差异表达基因及其所属的生物学通路。实验所用原代成骨细胞（primary osteoblasts）取自3只8~12周龄的雄性野生型（WT）C57BL/6小鼠（C57BL6）。将这些细胞进行14天成骨分化后，每只特异性小鼠来源的细胞池要么不予处理（n=3），要么用重组小鼠C5a（recombinant murine C5a）处理4小时。