Nakanishi Kana, Tsukimoto Mitsutoshi, Tanuma Sei-Ichi, Takeda Ken, Kojima Shuji
Department of Radiation Biosciences, Generation, Research Institute for Science and Technology, Tokyo University of Science (TUS), 2641 Yamazaki, Noda-shi, Chiba 278-8510, Japan.
Biochemistry, Faculty of Pharmaceutical Sciences, Generation, Research Institute for Science and Technology, Tokyo University of Science (TUS), 2641 Yamazaki, Noda-shi, Chiba 278-8510, Japan.
Toxicol In Vitro. 2016 Sep;35:202-11. doi: 10.1016/j.tiv.2016.06.003. Epub 2016 Jun 14.
We examined the mechanism of SNP-mediated stimulation of IL-1β and IL-18 production via P2R-mediated pathways in mouse bone marrow dendritic cells (mBMDCs). Examination of uptake of SNPs with diameters of 30, 70, and 300nm (SNP30, SNP70, and SNP300, respectively) by lipopolysaccharide-matured mBMDCs revealed that significant uptake of SNP30 occurred within as short a time as 1h. Production of IL-1β and IL-18 by cells exposed to SNPs increased dose-dependently, and was highest in cells exposed to SNP30. The SNP30-induced cytokine production was significantly inhibited by ATPase (apyrase) and by P2X7 receptor antagonist (A438079). ATP release was also highest in SNP30-exposed cells. Treatment of mBMDCs with exogenous ATP induced release of high levels of IL-1β and IL-18, and this release was also significantly inhibited by apyrase and A438079. The order of effectiveness of the three SNPs for inducing intracellular reactive oxygen species (ROS) production accorded well with those of cytokine production and ATP release. ROS production was inhibited by diphenyleneiodonium chloride (DPI). SNPs, especially SNP30, activate purinergic signaling in matured mBMDCs by inducing ATP release via P2X7 receptor. ATP induces ROS production via NADPH oxidase, and ROS activate inflammasomes, leading to caspase-1-dependent processing of pro-cytokines and release of IL-1β and IL-18.
我们研究了单核苷酸多态性（SNP）通过P2受体介导的途径刺激小鼠骨髓树突状细胞（mBMDCs）产生白细胞介素-1β（IL-1β）和白细胞介素-18（IL-18）的机制。用脂多糖成熟的mBMDCs检测直径为30、70和300nm的SNP（分别为SNP30、SNP70和SNP300）的摄取情况，结果显示，SNP30在短短1小时内就有显著摄取。暴露于SNP的细胞产生IL-1β和IL-18的量呈剂量依赖性增加，且在暴露于SNP30的细胞中最高。ATP酶（腺苷三磷酸双磷酸酶）和P2X7受体拮抗剂（A438079）可显著抑制SNP30诱导的细胞因子产生。暴露于SNP30的细胞中ATP释放量也最高。用外源性ATP处理mBMDCs可诱导高水平的IL-1β和IL-18释放，且这种释放也被腺苷三磷酸双磷酸酶和A438079显著抑制。三种SNP诱导细胞内活性氧（ROS）产生的有效性顺序与细胞因子产生和ATP释放的顺序非常一致。二苯基碘鎓氯化物（DPI）可抑制ROS产生。SNP，尤其是SNP30，通过P2X7受体诱导ATP释放，从而激活成熟mBMDCs中的嘌呤能信号通路。ATP通过烟酰胺腺嘌呤二核苷酸磷酸氧化酶诱导ROS产生，而ROS激活炎性小体，导致前细胞因子的半胱天冬酶-1依赖性加工以及IL-1β和IL-18的释放。
