Theranostics 2020; 10(10):4589-4605. doi:10.7150/thno.34065

Research Paper

Carbon black nanoparticles induce cell necrosis through lysosomal membrane permeabilization and cause subsequent inflammatory response

Xia Yuan, Wen Nie, Zhiyao He, Jingyun Yang, Bin Shao, Xuelei Ma, Xiangxian Zhang, Zhenfei Bi, Lu Sun, Xiao Liang, Yan Tie, Yu Liu, Fei Mo, Dan Xie, Yuquan Wei, Xiawei Wei

Laboratory of Aging Research and Cancer Drug Target, State Key Laboratory of Biotherapy and Cancer Center, National Clinical Research Center for Geriatrics, West China Hospital, Sichuan University, No. 17, Block 3, Southern Renmin Road, Chengdu, Sichuan 610041, PR China.

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Citation:
Yuan X, Nie W, He Z, Yang J, Shao B, Ma X, Zhang X, Bi Z, Sun L, Liang X, Tie Y, Liu Y, Mo F, Xie D, Wei Y, Wei X. Carbon black nanoparticles induce cell necrosis through lysosomal membrane permeabilization and cause subsequent inflammatory response. Theranostics 2020; 10(10):4589-4605. doi:10.7150/thno.34065. Available from http://www.thno.org/v10p4589.htm

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Abstract

Rationale: The adverse health effects of nano-particulate pollutants have attracted much attention in recent years. Carbon nanomaterials are recognized as risk factors for prolonged inflammatory responses and diffuse alveolar injury. Previous research indicated a central role of alveolar macrophages in the pathogenesis of particle-related lung disease, but the underlying mechanism remains largely unknown.

Methods: C57BL/6 mice were intratracheally instilled with carbon black nanoparticles (CBNPs). Cell necrosis and the infiltrated neutrophils in the lungs were detected by flow cytometry. Release of mitochondria was observed with Mito Tracker and mitochondrial DNA (mtDNA) was quantified by qPCR via Taqman probes. TLR9-p38 MAPK signaling pathway was detected by Western blotting. The production of lipid chemoattractant leukotriene B4 (LTB4) in the supernatant and bronchoalveolar lavage fluid (BALF) was quantitated using an enzyme immunoassay (EIA).

Results: In the present study, we found that a single instillation of CBNPs induced neutrophil influx in C57BL/6 mice as early as 4 h post-exposure following the rapid appearance of cell damage indicators in BALF at 30 min. Macrophages exposed to CBNPs showed necrotic features and were characterized by lysosome rupture, cathepsin B release, reactive oxygen species generation, and reduced intracellular ATP level. Necrosis was partly inhibited by a specific lysosomal cathepsin B inhibitor CA074 Me. Further analyses suggested that the resulting leakage of mtDNA from the necrotic cells activated neutrophils and triggered severe inflammation in vivo. Pulmonary neutrophilic inflammation induced by mtDNA was reduced in TLR9-/- mice. Additionally, mtDNA induced LTB4 production from macrophages, which may contribute to neutrophil recruitment.

Conclusion: We demonstrated here that CBNPs induce acute cell necrosis through lysosomal rupture and that mtDNA released from necrotic cells functions as a key event mediating pulmonary neutrophilic inflammation. This study described a novel aspect of the pathogenesis of particle-induced inflammatory response and provided a possible therapeutic target for the regulation of inflammation.

Keywords: Carbon black nanoparticles, Macrophages, Necrosis, Mitochondrial DNA, Inflammation