Abstract
Interleukin-1β (IL-1β) is a proinflammatory cytokine and an important component of innate immunity. However, overproduction of IL-1β is implicated in a number of autoinflammatory diseases. The discovery of the nucleotide binding oligomerisation domain (NOD)-like receptor containing a pyrin domain 3 (NLRP3) inflammasome provided crucial insight into how IL-1β processing is regulated, but despite decades of research the exact mechanisms by which the inflammasome can be activated are not fully understood. One hypothesis is that increases in reactive oxygen species (ROS), as occurs during oxidative stress, leads to activation of the inflammasome but the available data are contradictory. The association between inflammation and oxidative stress is well-known and it is possible that the NLRP3 inflammasome could function as an important mechanistic link between ROS, oxidative stress and inflammatory disease.The aim of this study was to investigate the function of ROS in NLRP3 activation in primary human monocytes – an innate immune cell that extravasates into inflamed tissues and secretes IL-1β, thus playing a significant role in innate immunity, inflammation and disease.
ROS production was measured in primary human monocytes that were exposed to monosodium urate (MSU) and calcium pyrophosphate dihydrate (CPPD) crystals together with toll-like receptor 2 (TLR2) ligand Pam3Csk4. Both type of crystal induced NLRP3- dependent IL-1β secretion, but only MSU induced significant increases in ROS, suggesting that the production of ROS is not necessary for inflammasome activation. These findings were subsequently confirmed using small molecule antioxidants and antioxidant enzymes which inhibited ROS production but had no effect on MSU-induced IL-1β secretion.
The effects of more subtle changes in redox homeostasis on the activation of NLRP3 were then assessed. Uric acid is one of the major antioxidants in the blood and when present at high concentrations is the greatest risk factor for the development of gout – an inflammatory disease driven by MSU-induced IL-β secretion. Uric acid had no effect on activation of the inflammasome nor did it influence gene expression of inflammasome components in human monocytes. Although differences were observed in antioxidant capacity in sera and monocytes from individuals with gout, chronic kidney disease and rheumatoid arthritis, there were no correlations between intra- or extracellular redox state and IL-1b secretion from monocytes, suggesting that inflammasome activation is redox-independent.
The results of this work demonstrate that activation of NLRP3 in primary human monocytes is not affected by ROS and thus the inflammasome is unlikely to be redox-regulated in these
cells. This is an important finding for understanding the complex relationship between inflammation and oxidative stress, and indicates that this relationship is not always causal in nature in human monocytic cells.
| Date of Award | Feb 2019 |
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| Original language | English |
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| Supervisor | Lisa Mullen (Supervisor), Kevin A. Davies (Supervisor) & Pietro Ghezzi (Supervisor) |