NLRP3 inflammasome-induced pyroptosis is increased in patients with cardiogenic shock

Alexander Kogel (Leipzig)1, L. Baumann (Leipzig)1, C. Mäder (Leipzig)1, P. Büttner (Leipzig)2, H. Thiele (Leipzig)2, J. M. Kneuer (Leipzig)1, J.-N. Boeckel (Leipzig)1, U. Laufs (Leipzig)1, S. Gaul (Leipzig)1

1Universitätsklinikum Leipzig Klinik und Poliklinik für Kardiologie Leipzig, Deutschland; 2Herzzentrum Leipzig - Universität Leipzig Klinik für Innere Medizin/Kardiologie Leipzig, Deutschland


Introduction: Cardiogenic shock (CS) is a highly heterogeneous syndrome associated with high mortality. Tailored therapies and the identification of subgroups are needed to improve these outcomes. Since revascularisation may lead to reperfusion injury and stress response, inflammation, and associated inflammatory cell death have a key role in reperfusion tissue damage. Therefore, we aimed to study whether pyroptosis, a pro-inflammatory cell death, is increased in patients with CS and whether the revascularisation affects the NLRP3 inflammasome activation and pyroptosis.
Materials and Methods: We collected serum and plasma (n=44) as well as peripheral blood mononuclear cells (PBMC, n=7) of patients with CS complicating acute myocardial infarction on admission and post-revascularization (24 h, 48 h, 72 h) and of healthy controls (n=75 serum and plasma, n=12 additional PBMC). Gene expression was measured in isolated PBMC by qPCR to determine if the necessary priming for inflammasome activation is present in CS patients. Interleukin concentrations in serum were measured by ELISA and Caspase 1 activity as well as circulating ASC specks were assessed by flow cytometry to assess inflammasome activity and the resulting cytokine release.
Gene expression of NLRP3, Caspase1, ASC, and IL1B (Interleukin-1ß), as well as IL18 (Interleukin-18), was increased in patients with CS. Caspase-1 activity in PBMC of patients with CS was transiently increased 24h after revascularisation compared to healthy controls (1.6-fold, p<0.01). We further found increased time-dependent pyroptotic cell death in these patients with the highest level 48 h after revascularization (control: 11.4% vs. CS before revascularization: 11.4% vs. CS 48h post-revascularization 17.2%, Fig 1A), and significantly increased IL-1ß levels after revascularization. IL-18 and IL-6 showed no significant regulation before and after revascularization. Pro-inflammatory ASC specks were increased in serum of CS patients vs healthy controls and increased after revascularization (CS at admission: 11.1±4.6 specks/µl vs. CS after 24h: 19.0±3.9, p<0.02, Fig 1B). There was no correlation of pyroptosis rate or circulating ASC specks to conventional blood parameters. 
Conclusion: Our data proved that pyroptosis and circulating ASC specks are increased in patients with CS and that revascularization mediates NLRP3 inflammasome activation and pyroptotic cell death. This adds a new and independent parameter of inflammatory activity in these patients and may be used to tailor anti-inflammatory therapeutic strategies to susceptible patients.
Fig 1. 
(A) Percentage of FLICA-Caspase-1 positive and 7-AAD positive cells of all measured PBMCs measured by flow cytometry as a surrogate for pyroptosis (B) Absolute values of ASC specks per microliter serum measured by flow cytometry. Data are presented as mean ± standard deviation *p < 0.05 
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