P6291Post-myocardial infarction activation of P2X7 dependent inflammasome is crucial to develop an appropriate scar integrity

Abstract

Abstract Introduction Cardiovascular diseases are the main cause of death worldwide. Acute ischemia results in cell death of cardiomyocytes accompanied with the release of so called damage associated molecular patterns (DAMPs) such as nucleotides. High concentration of ATP in the extracellular fluid leads to the opening of the ionotropic purinergic receptor P2X7. The following transmembranous ion flux triggers the assembly of the NLRP-3 inflammasome and proteolytic caspase-1 activation which cleaves pro-IL-1β and provokes the release of fully active pro-inflammatory IL-1β. Myocardial scar formation can be divided into an early remodelling IL-1β-dependent and a late scar forming TGFβ phase. Both phases negatively regulate each other. The current stuy aimed to investigate the role of this master regulator of NLRP3 inflammasome assembly P2Y7 in myocardial remodeling during prolonged ischemic conditions. Methods 10 weeks old P2X7 knock-out and C57Bl6/J mice received a full ligation of the left anterior descending (LAD) artery. Mice three and seven days post-infarction underwent echocardiography. Myocardial scar formation was assessed by histological staining and flow cytometry. Furthermore caspase-1 activity was measured using FLICA in histology. Gene expression was assessed using TaqMan realtime PCR. Results Macrophages in the myocardial infarct area showed high P2X7 expression by Co-staining with fluorescent antibodies against F4/80, CD68 and P2X7. Intriguingyl P2X7 deficient animals showed a significantly worse survival rate in a Kaplan-Meier survival analysis compared to wt littermated with LAD ligation (Mortality after 21 days P2X7+/+ 50%; P2X7−/− 100%, p<0.05). Cause of death assessed by autopsy was myocardial rupture in P2X7−/− mice. Accordingly histological analysis revealed a less compact infarct area in P2X7 knock-out animals with abundant coagulation necrosis. In agreement with that we observed a thickened infarcted anterior wall by echocardiography in P2X7 animals. Furthermore whereas the infarcted area of P2X7 competent mice showed high signals for active caspase-1 in histology, we were not able to detect any signal of caspase-1 activity in P2X7 deficient mice. In coherence with this observation we detected a premature increased TGFβ gene transcript upregulation in infarct tissue of P2X7 deficient animals. Conclusion The knockout of the NLRP3 inflammasome activating P2X7 receptor impairs the outcome after myocardial infarction by reduced monocyte infiltration and deranged scar formation. Disruption of the fine-tuned IL-1β/TGFβ sequence with an early block of IL-1β signaling and premature TGFβ activation could explain the missing clean-up of necrotic debris and impaired scar formation. Taken together these findings highlight the importance of an early inflammasome phase during myocardial scar formation.