Cellular and Molecular Neurobiology Research Group, Department of Anatomy, Faculty of Medicine, Masaryk University, CZ-62500 Brno, Czech Republic.
Cells. 2020 Dec 11;9(12):2661. doi: 10.3390/cells9122661.
Following nerve injury, disintegrated axonal mitochondria distal to the injury site release mitochondrial formylated peptides and DNA that can induce activation and inflammatory profiling of Schwann cells via formyl peptide receptor 2 (Fpr2) and toll-like receptor 9 (TLR9), respectively. We studied RT4 schwannoma cells to investigate the regulation of Fpr2 and TLR9 after stimulation with fMLF as a prototypical formylated peptide. RT4 cells were treated with fMLF at various concentrations and times with and without pretreatment with inhibitors (chloroquine for activated TLR9, PBP10 for Fpr2). Western blots of Fpr2, TLR9, p-p38, p-NFκB, and IL-6 were compared in relation to inflammatory profiling of RT4 cells and chemokine receptors (CCR2, CXCR4) as potential co-receptors of Fpr2. fMLF stimulation upregulated Fpr2 in RT4 cells at low concentrations (10 nM and 100 nM) but higher concentrations were required (10 µM and 50 µM) when the cells were pretreated with an activated TLR9 inhibitor. Moreover, the higher concentrations of fMLF could modulate TLR9 and inflammatory markers. Upregulation of Fpr2 triggered by 10 nM and 100 nM fMLF coincided with higher levels of chemokine receptors (CCR2, CXCR4) and PKCβ. Treating RT4 cells with fMLF, as an in vitro model of Schwann cells, uncovered Schwann cells' complex responses to molecular patterns of release from injured axonal mitochondria.
在神经损伤后，损伤部位远端崩解的轴突线粒体释放出的线粒体甲酰肽和 DNA，可分别通过甲酰肽受体 2（Fpr2）和 Toll 样受体 9（TLR9）诱导雪旺细胞的激活和炎症表型。我们使用 RT4 雪旺细胞瘤系来研究 Fpr2 和 TLR9 在受到典型甲酰肽 fMLF 刺激后的调控。RT4 细胞用不同浓度和时间的 fMLF 处理，同时用和不用 TLR9 激活抑制剂（氯喹）和 Fpr2 抑制剂（PBP10）预处理。Fpr2、TLR9、p-p38、p-NFκB 和 IL-6 的 Western blot 与 RT4 细胞的炎症表型以及 Fpr2 的潜在共受体趋化因子受体（CCR2、CXCR4）进行了比较。fMLF 刺激在低浓度（10 nM 和 100 nM）时上调 RT4 细胞中的 Fpr2，但当细胞用激活的 TLR9 抑制剂预处理时，需要更高的浓度（10 μM 和 50 μM）。此外，更高浓度的 fMLF 可以调节 TLR9 和炎症标志物。10 nM 和 100 nM fMLF 引发的 Fpr2 上调与更高水平的趋化因子受体（CCR2、CXCR4）和 PKCβ 同时发生。用 fMLF 处理 RT4 细胞，作为雪旺细胞的体外模型，揭示了雪旺细胞对损伤轴突线粒体释放的分子模式的复杂反应。
