Activation of the NOD-like receptor (NLR) family, pyrin domain-containing protein 3 (NLRP3) inflammasome has recently been implicated in driving the pathogenesis of diabetic complications. While diabetes is known to increase NETosis propensity, little is known whether neutrophil NLRP3 mediates NETosis in diabetes, and if organelles that govern metabolic homeostasis contribute to aberrant activation of NLRP3. In this study, we sought to examine if neutrophil NLRP3 is activated in diabetes, and if key metabolic organelles are involved in the activation of NLRP3, resulting in NETosis upregulation. We found that pre-treatment of blood neutrophils isolated from mice and humans with diabetes using MCC-950, a potent and specific NLRP3 inhibitor, normalized NET formation to levels similar to those of neutrophils isolated from healthy mice and humans. Similar to blood neutrophils, HL-60-derived neutrophils primed in high glucose also showed reduced NETosis when pre-treated with MCC-950. Notably, compared to their normoglycemic counterparts, neutrophils isolated from mice and humans with diabetes exhibited increased speck formation of the inflammasome adaptor protein, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) even at baseline, indicative of heightened NLRP3 activation in diabetes. In mouse neutrophils, confocal microscopy revealed that oxidized DNA release into the cytoplasm is a prerequisite for ASC speck formation, which co-localized with mitochondria. In addition, MCC-950 yielded no further reduction in NET formation when HL-60-derived neutrophils were pre-treated with tauroursodeoxycholic acid, an alleviator of endoplasmic reticulum (ER) stress. In summary, our findings suggest that mitochondria and ER stress may synergize in the activation of neutrophil NLRP3 that exacerbates NETosis in diabetes. Therapeutics that aim at intercepting the pathway may alleviate NET-mediated chronic inflammation in diabetes.


L.Wang: None. R.Dalan: None. S.Wong: None.


Nanyang Assistant Professorship; Singapore Ministry of Education Academic Research Fund Tier 1 (RG25/20); Nanyang President's Graduate Scholarship

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