High resolution and dynamic imaging of biopersistence and bioreactivity of extra and intracellular MWNTs exposed to microglial cells.
High resolution and dynamic imaging of biopersistence and bioreactivity of extra and intracellular MWNTs exposed to microglial cells.
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DOI：
10.1016/j.biomaterials.2015.08.019
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发表时间：
2015-11
期刊：
影响因子：
14
通讯作者：
Porter AE
中科院分区：
文献类型：
--
作者：
Goode AE;Gonzalez Carter DA;Motskin M;Pienaar IS;Chen S;Hu S;Ruenraroengsak P;Ryan MP;Shaffer MS;Dexter DT;Porter AE
多壁碳纳米管（MWNTs）越来越多地被开发为神经治疗药物输送系统到大脑和神经支架，以驱动组织再生的病变部位。具有不同程度的酸氧化的多壁碳纳米管可能具有不同的生物活性和在细胞外环境中聚集的倾向，并且可以预期在大脑中发现个体化和聚集的多壁碳纳米管。在实际应用之前，了解聚集体和单个MWNT如何与局部吞噬免疫细胞（小胶质细胞）相互作用，并最终确定它们在大脑中的生物持久性是至关重要的。通过关联一系列动态、定量和多尺度技术，包括：紫外-可见光谱、光学显微镜、聚焦离子束扫描电子显微镜和透射电子显微镜，在多个长度尺度上表征小胶质细胞对细胞外和细胞内MWNT（原始和酸氧化时）的处理。动态，活细胞成像显示的能力，小胶质细胞分裂和内化的酸氧化的MWNT，但不是原始的MWNT的微米级细胞外聚集体。通过小胶质细胞内化或强烈结合的MWNT的总量被定量为时间的函数。无论是氧化的多壁碳纳米管的显着吸收，也不完全吸收的原始多壁碳纳米管影响小胶质细胞的活力，促炎细胞因子的释放或一氧化氮的生产。然而，在暴露于原始MWNTs 24小时后，观察到活性氧物质的产生显著增加。72小时后，细胞质和囊泡（包括多层小体）中存在小聚集体和个体化氧化MWNT。观察到氧化MWNT结构的形态损伤的一些证据，包括高度无序的石墨结构，表明可能的生物降解。这项工作展示了动态，定量和多尺度技术在理解功能化纳米材料的不同细胞加工途径中的实用性。这种相关的方法具有广泛的影响，用于评估MWNT聚集体在体内其他地方的生物持久性，特别是它们与肺中巨噬细胞的相互作用。
Multi-walled carbon nanotubes (MWNTs) are increasingly being developed both as neuro-therapeutic drug delivery systems to the brain and as neural scaffolds to drive tissue regeneration across lesion sites. MWNTs with different degrees of acid oxidation may have different bioreactivities and propensities to aggregate in the extracellular environment, and both individualised and aggregated MWNTs may be expected to be found in the brain. Before practical application, it is vital to understand how both aggregates and individual MWNTs will interact with local phagocytic immune cells, the microglia, and ultimately to determine their biopersistence in the brain. The processing of extra- and intracellular MWNTs (both pristine and when acid oxidised) by microglia was characterised across multiple length scales by correlating a range of dynamic, quantitative and multi-scale techniques, including: UV-vis spectroscopy, light microscopy, focussed ion beam scanning electron microscopy and transmission electron microscopy. Dynamic, live cell imaging revealed the ability of microglia to break apart and internalise micron-sized extracellular agglomerates of acid oxidised MWNT, but not pristine MWNTs. The total amount of MWNTs internalised by, or strongly bound to, microglia was quantified as a function of time. Neither the significant uptake of oxidised MWNTs, nor the incomplete uptake of pristine MWNTs affected microglial viability, pro-inflammatory cytokine release or nitric oxide production. However, after 24 hrs exposure to pristine MWNTs, a significant increase in the production of reactive oxygen species was observed. Small aggregates and individualised oxidised MWNTs were present in the cytoplasm and vesicles, including within multilaminar bodies, after 72 hours. Some evidence of morphological damage to oxidised MWNT structure was observed including highly disordered graphitic structures, suggesting possible biodegradation. This work demonstrates the utility of dynamic, quantitative and multi-scale techniques in understanding the different cellular processing routes of functionalised nanomaterials. This correlative approach has wide implications for assessing the biopersistence of MWNT aggregates elsewhere in the body, in particular their interaction with macrophages in the lung.