Multiwalled-Carbon Nanotube Properties and Macrophage Proinflammatory Responses

多壁碳纳米管(MWNTs)的生产和使用在世界范围内迅速增加，尽管 它们可能对人类健康造成的不利影响。特别令人关切的是，有报道称，多国非关税壁垒 可导致实验室动物的肺纤维化，可能导致间皮瘤，类似于石棉。文学作品 关于MWNTs的哪些物理特性导致吞噬细胞释放促炎物质存在冲突 肺纤维化前的细胞因子。也有人认为，吞噬酶体膜 MWNTs引起的损伤是导致吞噬细胞释放细胞因子的关键事件，但MWNTs 财产造成损害的原因也不清楚。这里提出的研究的广泛目标是更好地 了解MWNT的长度和分散程度是否是导致细胞因子释放的因素 并探索一种相对新的方法，激光扫描共聚焦拉曼显微镜 评估多壁碳纳米管是否在诱导细胞因子释放的过程中破坏吞噬小体的膜。那里 为实现该提案的目标有两个具体目标： 具体目标1：量化两种吞噬细胞摄取的多壁碳纳米管的数量 MWNT的吸收与MWNT长度、分散剂、团聚、毒性和释放的相关性 促炎细胞因子。将研究两种不同的MWNT类型，长的和短的。每种类型，两个 分散体将用高度分散的分散剂牛血清白蛋白或Pluronic(R)F-108制备 而且凝聚在一起。实验设计将使用一种新的方法来测量 在培养的吞噬细胞内积聚的多壁碳纳米管，然后测量两种细胞因子的释放， 肿瘤坏死因子-β和白介素1-β。这一目标的结果将第一次将体内多壁碳纳米管的实际数量联系起来 吞噬细胞与MWNT长度、分散剂类型、聚集、毒性和释放 促炎细胞因子。 特定目标2：确定多壁碳纳米管的亚细胞分布和潜在的吞噬溶酶体 激光扫描共聚焦拉曼显微镜(LSCRM)的损伤。LSCRM，一种相对较新的方法 细胞成像，可以根据MWNTs独特的拉曼信号直接定位细胞内的MWNTs，并用于 从共聚焦切片重建显示MWNTs亚细胞分布的3D模型。LSCRM可以 同时，基于细胞膜脂质的C-H键拉曼散射对细胞膜进行成像。有了这个 多种能力的结合可能确定细胞摄取的MWNTs是否会损害 吞噬酶体膜，并重新分布在整个细胞质。这一目标的结果应该有助于解决 MWNT的长度和分散状态是否是破坏吞噬酶体膜的因素。

The production and use of multi-walled carbon nanotubes (MWNTs) are rapidly increasing world-wide, despite the possible adverse effects they may have on human health. Of particular concern are reports that MWNTs can cause pulmonary fibrosis in lab animals that may lead to mesothelioma, similar to asbestos. The literature is conflicted regarding what physical features of MWNTs cause phagocytic cells to release proinflammatory cytokines that precede pulmonary fibrosis. It has also been suggested that phagolysosome membrane damage caused by MWNTs is a key event leading to cytokine release by phagocytic cells, but what MWNT properties cause damage is also not clear. The broad objectives of the research proposed here are to better understand whether MWNT length and extent of dispersion are factors leading to cytokine release by phagocytic cells and to explore a relatively new approach, laser scanning confocal Raman microscopy, to assess whether MWNTs damage the phagolysosomal membrane en route to eliciting cytokine release. There are two specific aims to achieve the objectives of the proposal: Specific aim 1: To quantify the amount of MWNTs taken up by two types of phagocytic cells and to correlate the uptake with MWNT length, dispersant, agglomeration, toxicity, and the release of proinflammatory cytokines. Two different MWNT types will be studied, long and short. For each type, two dispersions will be prepared with the dispersants bovine serum albumin or Pluronic(R) F-108, highly dispersed and agglomerated. The experimental design will use a new method to measure the actual amount of the MWNTs that accumulate inside cultured phagocytic cells, followed by measuring the release of two cytokines, TNF-¿ and IL-1¿. The results of this aim will, for the first time, correlate the actual amount of MWNTs inside phagocytic cells with MWNT length, type of dispersant, agglomeration, toxicity, and the release of proinflammatory cytokines. Specific aim 2: To determine the subcellular distribution of MWNTs and potential phagolysosome damage by laser scanning confocal Raman microscopy (LSCRM). LSCRM, a relatively new approach to cell imaging, can directly locate MWNTs inside cells by their unique Raman signature and be used to reconstruct 3D models from confocal sections showing the subcellular distribution of MWNTs. LSCRM can also image membranes in cells based on the C-H bond Raman scattering of membrane lipids. With this combination of capabilities it may be possible to determine whether MWNTs taken up by cells damage the phagolysosome membrane and redistribute throughout the cytosol. Results from this aim should help address whether MWNT length and state of dispersion are factors in damaging the phagolysosome membrane.