NLRP3 inflammasome-produced pro-inflammatory cytokines IL-1b and IL-18 are critical exacerbating factors of septic cardiomyopathy

Background Sepsis is life-threatening multiple organ failure due to systemic infection, and frequently causes septic cardiomyopathy (SCM). Recent evidence has established that cytokine storm, an aberrant increase in various cytokines leading to severe inflammation and tissue damage, plays a crucial role in SCM. However, it is unknown which cytokines are the most critical initiators and exacerbating factors of SCM. Purpose To investigate how IL-1β and IL-18 driven by NLRP3 inflammasome contribute to the pathophysiology of SCM, and to study whether blockage of these cytokines can ameliorate cardiac dysfunction in a murine model of lipopolysaccharide (LPS)-induced SCM. Methods Wild-type, Nlrp3-/-, Casp1/11-/-, and Il1b-/- mice were injected intraperitoneally with 6 mg/kg of LPS to induce SCM. To suppress IL-18, the adeno-associated virus (AAV) vectors encoding IL-18 binding protein (IL-18BP) were injected intramuscularly 14 days before LPS. Bone marrow transplantation (5 Gy x2) was performed 6 weeks prior to LPS. Results While 40% of wild-type mice died 48 h after LPS injection in our SCM model, none of Nlrp3-/- mice died. Cardiac function, as determined by echocardiography and BNP, was significantly decreased 6 h after LPS injection in WT mice but not in Nlrp3-/- or Casp1/11-/- mice. Serum CK-MB increase was also ameliorated in Nlrp3-/- or Casp1/11-/- mice. Serum levels of various pro-inflammatory cytokines including TNFα, IL-1β, IL-6, IL-18, and IFNγ were suppressed in Nlrp3-/- mice. These findings collectively indicated a critical role of NLRP3 inflammasome in LPS-induced SCM. However, unexpectedly, genetic deletion of IL-1β, the canonical effector molecule of NLRP3 inflammasome, in Il1b-/- mice failed to alleviate cardiac dysfunction and damage caused by LPS. In addition, IL-18 suppression by AAV-mediated overexpression of IL-18BP, a high-affinity IL-18 decoy receptor, only partially prevented LPS-induced SCM. On the other hand, intriguingly, dual inhibition of IL-1β and IL-18 successfully protected mice from LPS-induced SCM, demonstrating that concomitant blockage of two NLRP3 inflammasome products, IL-1β and IL-18, was necessary and sufficient to prevent SCM. Finally, while bone marrow transplantation experiments showed that cardiac dysfunction after LPS injection was suppressed in recipient Nlrp3-/- mice transplanted with wild-type bone marrow cells, cardiomyocyte-specific deletion of NLRP3 in αMHC-Cre;Nlrp3-/- mice did not prevent LPS-induced SCM. These data suggest that NLRP3 inflammasome in resident cells other than bone marrow-derived cells and cardiomyocytes is important in the pathophysiology of SCM. Conclusion The present study indicates that concomitant blockage of IL-1β and IL-18 driven by NLRP3 inflammasome is necessary and sufficient to prevent SCM. Our data provide fundamental evidence that combination therapy targeting both IL-1β and IL-18 could serve as a new approach to the treatment of SCM.