Investigation into Mechanisms NLRP3 Activation by Nickel Associated Multi-Walled

描述（由申请人提供）：某些工程纳米材料（ENM）已被证明在动物模型中引起显著的肺部病理变化，引起了人们对随着使用和接触的增加将出现人类健康影响的担忧。然而，尚未建立基于ENM的物理和表面性质的机械预测模型来帮助保护人类健康。我们实验室的其他工作已经显示了体外NLRP 3炎性体（NLRP 3）活化和体内炎症之间的关联。在初步数据中，我们已经表明，镍相关的多壁碳纳米管（Ni-MWCNT）在体外使用原代肺泡巨噬细胞（AM）或THP-1细胞激活NLRP 3的能力与肺部炎症和病理学之间可能存在相关性。因此，我们认为NLRP 3的体外激活似乎是肺部炎症的可靠预测因子。区分生物活性ENM和良性ENM的基础很可能与ENM被吞噬的能力和/或破坏溶酶体导致组织蛋白酶B释放的能力有关。我们假设ENM的体内炎症潜力与溶酶体破坏活性和NLRP 3活化密切相关。根据初步数据，我们提出，多壁碳纳米管的炎症潜力将取决于相关的镍的量。因此，我们的目标是：1。确定AM中NLRP 3炎性体活性与暴露于限定且充分表征的Ni-MWCNT后小鼠的肺病理学之间的关系。2.使用THP-1细胞确定Ni-MWCNT表面性质（镍量和表面电荷）对引起细胞毒性的重要性和NLRP 3炎性体的活化程度。最后，我们将确定MWCNT-Ni的内化，随后是溶酶体膜破坏和组织蛋白酶B释放是NLRP 3炎性体激活的起始事件。因此，从这项研究中获得的信息将是重要的，在确定安全ENM的特点和建立的作用机制，并可能导致在体外筛选平台，使用THP-1细胞。 公共卫生相关性：这些研究将确定纳米材料破坏溶酶体膜并激活NRLP 3炎性体以引起体内炎症和病理的潜力。此外，在提案中进行的测定可以作为ENM的高通量筛选的工具。!

DESCRIPTION (provided by applicant): Certain engineered nanomaterials (ENM) have been shown to cause significant lung pathological changes in animal models raising concern that human health effects will emerge with increasing use and exposure. However, a mechanistic predictive model based on physical and surface properties of ENM has not been established to aid in protecting human health. Other work in our laboratory has shown an association between in vitro NLRP3 inflammasome (NLRP3) activation and in vivo inflammation. In preliminary data, we have shown that there may be a correlation between the ability of nickel associated MWCNT (Ni-MWCNT) to activate NLRP3 in vitro using primary alveolar macrophages (AM) or THP-1 cells with lung inflammation and pathology. Therefore, we propose that in vitro activation of NLRP3 appears to be a reliable predictor of lung inflammation. The basis for the distinction between bioactive and benign ENM is most likely associated with the ability of ENM to be phagocytosed and/or ability to disrupt lysosomes causing cathepsin B release. We hypothesize that the in vivo inflammatory potential of ENM correlates well with lysosomal disrupting activity and NLRP3 activation. Based on preliminary data we propose that the inflammatory potential of MWCNT will be dependent on the amount of associated nickel. Therefore, our aims are: 1. Determine the relationship between NLRP3 inflammasome activity in AM and lung pathology in mice following exposure to defined and well- characterized Ni-MWCNT. 2. Determine the importance of Ni-MWCNT surface properties (amount of Nickel and surface charge) to cause cytotoxicity and degree of activation of the NLRP3 inflammasome using THP-1 cells. Finally, we will establish that internalization of MWCNT-Ni followed by lysosomal membrane disruption and Cathepsin B release is the initiating event in activation of the NLRP3 inflammasome. Thus, information from this study will be important in determining characteristics of safe ENM and establish mechanisms of action and may lead to an in vitro screening platform using THP-1 cells. PUBLIC HEALTH RELEVANCE: These studies will establish the potential of nanomaterials to disrupt lysosomal membranes and activate the NRLP3 inflammasome to cause in vivo inflammation and pathology. Furthermore that the assays performed in the proposal may serve as tools for high throughput screening of ENM. !