Background/Purpose: Self-activating mutations in NLRP3 cause a spectrum of autoinflammatory diseases known as cryopyrin-associated periodic syndromes (CAPS). NLRP3 is a key component of a multiprotein complex known as the inflammasome that mediates the maturation of the proinflammatory cytokine IL-1beta, and can induce rapid cell death in a process known as pyronecrosis. Although several models for inflammasome activation have been proposed the precise molecular mechanism, as well as the role of NRLP3 mutations, remains to be elucidated. Emerging evidence suggests that mitochondria are involved in inflammasome activation. STAT3 associates with the mitochondrial inner membrane in a GRIM-19 dependent manner and has been implicated in regulating cellular respiration. Here we asked whether regulation of mitochondrial membrane potential plays a role in NLRP3 inflammasome activation.
Methods: We used whole blood cells from NOMID patients and healthy controls, THP-1 cells with STAT3, NLRP3, GRIM-19, or OSCP expression knocked down, and monocytes derived from NLRP3-deficient mice. Cells were stimulated with LPS in the presence of inhibitors of STAT3, followed by ATP. Cell supernatants were collected and incubated with IL-1beta-capturing beads. Cells were fixed and permeabilized. Then beads were added back to cells, and the mixture of cells with beads was stained with anti-IL-1beta, CD14, and CD16 antibodies and then evaluated by flow cytometry. LPS stimulated cells were also evaluated using immunofluorescence and western blot analysis.
Results: By flow analysis we provide evidence that inhibition of STAT3 function in NOMID and healthy control monocytes, as well as knockdown of STAT3 in THP-1 cells, results in a significant decrease in inflammasome activation. Using confocal microscopy to visualize pyronecrosis, we provide evidence that this process is NLRP3 dependent.  Knockdown of GRIM-19 in THP-1 cells also inhibited NLRP3 activation, suggesting a requirement for mitochondrial STAT3. Enhancement of the mitochondrial membrane potential in STAT3 knockdown cells bypassed the effect of STAT3 knockdown, and reconstituted inflammasome activation, whereas knockdown of OSCP significantly reduced inflammasome activation.
Conclusion: These data suggest a previously unrecognized role for STAT3 in regulating mitochondrial membrane potential, which can regulate NLRP3 inflammasome activation. These results point toward mitochondrial STAT3 as a novel therapeutic target for NOMID and other NLRP3-mediated inflammatory diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/stat3-mediated-regulation-of-mitochondrial-membrane-potential-is-critical-for-nlrp3-inflammasome-activation/