Characterization of the NLRP3 Inflammasome: Inhibitors, Activators, and Priming
Inflammation is an important response of the innate immune system that aids in recruiting immune cells to the site of injury or infection. Protein complexes termed the inflammasomes form in response to certain danger signals and mediate the inflammatory response through recruitment and activation of caspase-1, which leads to the processing of inactive pro-IL-1β and pro-IL-18 into active cytokines, IL-1β and IL-18. Regulated inflammation is important for the host immune response, but abnormal activation of the inflammasome has been linked to several auto-inflammatory diseases such as gout, metabolic syndromes, and neurodegeneration. Several different proteins have been identified as platforms for the formation of the inflammasome, and each has a distinct set of stimuli recognition. One such protein is NLRP3, which is a member of the NOD-like receptor (NLR) family. While very intensely studied, the mechanism of activation of the NLRP3 inflammasome still remains unknown. In this study, the NLRP3 inflammasome will first be characterized through the anti-inflammatory compounds, parthenolide and Bay 11-7082, that were found to directly inhibit the inflammasome. Results demonstrate here that parthenolide inhibits the activity of multiple inflammasomes by directly inhibitng the protease activity of caspase-1. In addition, further investigations reveal that the synthetic IκB kinase-β inhibitor Bay 11-7082 and structurally related vinyl sulfone compounds selectively inhibit the NLRP3 inflammasome independent of their anti-NF-κB activity. In vitro assays indicate that parthenolide and Bay 11-7082 are able to inhibit the ATPase activity of NLRP3, suggesting that the inhibitory effect of these compounds could be in part due to their effect on the ATPase activity of NLRP3. These results examine the molecular mechanisms for the anti-inflammatory activity of parthenolide and identify vinyl sulfones as a new class of potential therapeutic compounds that target the activity of the NLRP3 inflammasome. The second portion of this study characterizes the priming mechanism of NLRP3. Here, evidence demonstrates that priming is a rapid event that occurs within minutes of incubation with priming stimuli, such as LPS. Additional investigation reveals that the priming of NLRP3 is an MYD88-dependent mechanism through mitochondrial ROS signaling. Results through confocal microscopy reveal that the active NLRP3 inflammasome is visible as a single speck-like structure that localizes to MitoTracker labeled mitochondria. Collectively, these results help to elucidate the mechanism of NLRP3 priming and further the understanding of NLRP3 inflammasome activation.
Juliana, Christine A, "Characterization of the NLRP3 Inflammasome: Inhibitors, Activators, and Priming" (2012). ETD Collection for Thomas Jefferson University. AAI3503550.