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Cytoprotective activated protein C averts Nlrp3 inflammasome induced ischemia reperfusion injury via mTORC1 inhibition

Sumra Nazir, Ihsan Gadi, Moh'd Mohanad Al-Dabet, Ahmed Elwakiel, Shrey Kohli, Sanchita Ghosh, Jayakumar Manoharan, Satish Ranjan, Fabian Bock, Ruediger C. Braun-Dullaeus, Charles T. Esmon, Tobias B. Huber, Eric Camerer, Chris Dockendorff, John H. Griffin, Berend Isermann and Khurrum Shahzad

Key points

  • Activated protein C (aPC) protects from myocardial and renal IRI by restricting mTORC1 mediated activation of the Nlrp3 inflammasome.

  • Nlrp3 inflammasome suppression by aPC is independent of its anticoagulant effect, depends on PAR1, and can be mimicked by parmodulin-2.

Abstract

Cytoprotection by activated protein C (aPC) following ischemia-reperfusion injury (IRI) is associated with apoptosis inhibition. However, IRI is hallmarked by inflammation and hence conceptually cell-death forms disjunct from immunologically silent apoptosis are more likely to be relevant. As pyroptosis, cell death resulting from inflammasome activation, is typically observed in IRI we speculated that aPC ameliorates IRI by inhibiting inflammasome activation. Here we analyzed the impact of aPC on inflammasome activity in myocardial and renal IRI. aPC treatment before or after myocardial IRI reduced infarct size and Nlrp3 inflammasome activation in mice. Kinetic in vivo analyses revealed that Nlrp3 inflammasome activation preceded myocardial injury and apoptosis, corroborating a pathogenic role of the Nlrp3 inflammasome. The constitutively active Nlrp3A350V mutant abolished aPC's protective effect, demonstrating that Nlrp3 suppression is required for aPC-mediated protection from IRI. In vitro aPC inhibited inflammasome activation in macrophages, cardiomyocytes, and cardiac fibroblasts via PAR-1 and mTORC1 signaling. Accordingly, inhibiting PAR-1 signaling, but not aPC's anticoagulant properties, abolished aPC's ability to restrict Nlrp3 inflammasome activity and tissue damage in myocardial IRI. Targeting biased PAR-1 signaling via parmodulin-2 restricted mTORC1 and Nlrp3 inflammasome activation and limited myocardial IRI as efficient as aPC. The relevance of aPC-mediated Nlrp3 inflammasome suppression following IRI was corroborated in renal IRI, where aPC's tissue protective effect was likewise dependent on Nlrp3 inflammasome suppression. These studies reveal that aPC protects from IRI by restricting mTORC1 dependent inflammasome activation and that mimicking biased aPC-PAR1 signaling using parmodulins may be a feasible therapeutic approach to combat IRI.

  • Submitted May 2, 2017.
  • Accepted September 1, 2017.