Investigation into Mechanisms NLRP3 Activation by Nickel Associated Multi-Walled

描述(申请人提供)：某些工程纳米材料(ENM)已被证明能在动物模型中引起显着的肺部病变，这引起了人们的担忧，即随着使用和暴露的增加，人类健康将受到影响。然而，基于ENM的物理和表面性质的机理预测模型还没有建立起来，以帮助保护人类健康。我们实验室的其他工作表明，体外NLRP3炎症小体(NLRP3)激活和体内炎症之间存在联系。在初步的数据中，我们已经证明了镍相关的MWCNT(Ni-MWCNT)体外利用原代肺泡巨噬细胞(AM)或THP-1细胞激活NLRP3的能力可能与肺部炎症和病理有关。因此，我们认为NLRP3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的体内炎症潜能与溶酶体破坏活性和NLRP3激活有很好的相关性。基于初步数据，我们认为MWCNT的炎症潜能将取决于相关的镍量。因此，我们的目标是：1.确定明确的和具有良好特征的Ni-MWCNT暴露后小鼠AM中NLRP3炎症小体活性与肺病理的关系。2.利用THP-1细胞，确定Ni-MWCNT表面性质(镍量和表面电荷)对NLRP3炎症小体的细胞毒性和激活程度的重要性。最后，我们将确定MWCNT-Ni的内化、溶酶体膜的破坏和组织蛋白酶B的释放是NLRP3炎症体激活的起始事件。因此，这项研究的信息对于确定安全的ENM的特征和建立作用机制将是重要的，并可能导致使用THP-1细胞的体外筛选平台。

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.