Abstract 13089: Reperfusion Injury is Reduced by Antagonism of NLRP3's Caspase-1

VX-765, inhibitor of caspases-1, 3 and 4, at reperfusion reduces infarction in isolated rat hearts by 50% after ischemia/reperfusion (I/R). It was assumed cardioprotection by this pharmaceutical-grade drug was the result of inhibition of caspase-1 (cas-1) activation by the NLRP3 inflammasome which would trigger lethal pyroptosis. However, there has been no direct proof of that hypothesis. To test which, if any, caspase was responsible for protection, we measured infarct size after 50-min global ischemia/2-h reperfusion in isolated wild type (WT) mouse hearts and hearts from mice in which cas-1 had been genetically deleted (offspring of breeders from Jackson Labs genotyped to confirm mutation). Infarct size was measured objectively by Image J analysis of tetrazolium-stained heart slices using the same threshold settings for all hearts. No gender difference in infarct size was found in WT hearts (see figure) so genders were mixed, especially in the in the knockout (KO) groups. Because Krebs perfusate lacks blood-borne esterases required to convert pro-drug VX-765 to its active metabolite, VRT-043198 (VRT), we employed VRT in our model. Infarct size in hearts reperfused with buffer containing 180 nM VRT was halved to 43.7±4.8% of ventricular mass (p<0.001 vs either cohort of WT hearts). Infarct size in cas-1 KO mice averaged 42.2±5.1% (pNS vs VRT). To eliminate possibility of a protected phenotype unrelated to cas-1, cas-1 KO mice were treated with 180 nM VRT. There was no further change in infarct size (44.0±4.3%) confirming cas-1 is VRT’s only target. NLRC4 also activates cas-1, but NLRC4 KO mice exposed to I/R were not protected (89.2±1.2% infarction), whereas inhibition of NLRP3 with MCC950 resulted in modest protection (65.5±5.3% infarction). These results strongly support NLRP3 as the source of the lethal cas-1. VX-765 at reperfusion is a promising adjunct intervention to decrease infarct size in patients treated for ST-segment elevation myocardial infarction.