State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China.
University of Chinese Academy of Sciences, Beijing, 100049, China.
Small. 2016 Nov;12(43):5998-6011. doi: 10.1002/smll.201602410. Epub 2016 Sep 20.
Exocytosis of single-walled carbon nanotubes (SWCNTs) determines therapeutic efficiency and toxicity of nanoproducts but its underlying mechanism remains elusive. In this study, it is found that the exocytosis mechanism of SWCNTs is mediated mainly through the activation of P2X receptor (P2X R), an ATP-gated membrane receptor highly expressed in macrophages. Inhibition of P2X R signaling by either a specific inhibitor (oxidized ATP) or small interfering RNA targeting P2X R largely prevents the exocytosis of SWCNTs from Raw264.7 cells, resulting in significant accumulation of SWCNTs within cells. In contrast, activation of P2X R with ATP promotes exocytosis of SWCNTs. Specifically, it is elucidated that internalized SWCNTs are accumulated in lysosomes and induce transitional release of ATP into extracellular space, which further activates P2X R, leading to the influx of calcium ions, phosphorylation of protein kinase C, ERK1/2, p38, and JNK, as well as alkalization of lysosomes. SWCNTs exposure also induces microtubules reorganization that facilitates the secretion of SWCNTs-containing lysosomes. It is also found that P2X R simultaneously mediates secretion of IL-1β from Raw264.7 cells during the process of SWCNTs exocytosis. The combined data reveals that P2X R-mediated pathway is the predominant molecular mechanism for exocytosis of SWCNTs in Raw264.7 cells. Moreover, SWCNT-induced inflammation is closely coupled with the exocytosis of SWCNTs through P2X R.
SWCNTs 的胞吐作用决定了纳米产物的治疗效果和毒性，但其中的作用机制仍不清楚。在本研究中，研究人员发现，SWCNTs 的胞吐作用机制主要通过 P2X 受体 (P2X R) 的激活来介导，P2X R 是一种高度表达于巨噬细胞中的 ATP 门控膜受体。通过特异性抑制剂 (氧化型 ATP) 或靶向 P2X R 的小干扰 RNA 抑制 P2X R 信号转导，在很大程度上阻止了 Raw264.7 细胞中 SWCNTs 的胞吐作用，导致 SWCNTs 在细胞内的大量积累。相反，用 ATP 激活 P2X R 则促进了 SWCNTs 的胞吐作用。具体来说，阐明了内化的 SWCNTs 被积累在溶酶体中，并诱导 ATP 向细胞外空间的瞬时释放，进而激活 P2X R，导致钙离子内流、蛋白激酶 C、ERK1/2、p38 和 JNK 的磷酸化以及溶酶体的碱化。SWCNTs 暴露还诱导微管重排，促进含有 SWCNTs 的溶酶体的分泌。还发现 P2X R 同时介导了 SWCNTs 胞吐过程中 Raw264.7 细胞中 IL-1β 的分泌。综合数据表明，P2X R 介导的途径是 Raw264.7 细胞中 SWCNTs 胞吐的主要分子机制。此外，SWCNT 诱导的炎症与通过 P2X R 的 SWCNTs 胞吐密切相关。
