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RIPK3 deficiency or catalytically inactive RIPK1 provides greater benefit than MLKL deficiency in mouse models of inflammation and tissue injury.

ABSTRACT: Necroptosis is a caspase-independent form of cell death that is triggered by activation of the receptor interacting serine/threonine kinase 3 (RIPK3) and phosphorylation of its pseudokinase substrate mixed lineage kinase-like (MLKL), which then translocates to membranes and promotes cell lysis. Activation of RIPK3 is regulated by the kinase RIPK1. Here we analyze the contribution of RIPK1, RIPK3, or MLKL to several mouse disease models. Loss of RIPK3 had no effect on lipopolysaccharide-induced sepsis, dextran sodium sulfate-induced colitis, cerulein-induced pancreatitis, hypoxia-induced cerebral edema, or the major cerebral artery occlusion stroke model. However, kidney ischemia-reperfusion injury, myocardial infarction, and systemic inflammation associated with A20 deficiency or high-dose tumor necrosis factor (TNF) were ameliorated by RIPK3 deficiency. Catalytically inactive RIPK1 was also beneficial in the kidney ischemia-reperfusion injury model, the high-dose TNF model, and in A20(-/-) mice. Interestingly, MLKL deficiency offered less protection in the kidney ischemia-reperfusion injury model and no benefit in A20(-/-) mice, consistent with necroptosis-independent functions for RIPK1 and RIPK3. Combined loss of RIPK3 (or MLKL) and caspase-8 largely prevented the cytokine storm, hypothermia, and morbidity induced by TNF, suggesting that the triggering event in this model is a combination of apoptosis and necroptosis. Tissue-specific RIPK3 deletion identified intestinal epithelial cells as the major target organ. Together these data emphasize that MLKL deficiency rather than RIPK1 inactivation or RIPK3 deficiency must be examined to implicate a role for necroptosis in disease.

SUBMITTER: Newton K

PROVIDER: S-EPMC5072432 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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RIPK3 deficiency or catalytically inactive RIPK1 provides greater benefit than MLKL deficiency in mouse models of inflammation and tissue injury.

Newton K K Dugger D L DL Maltzman A A Greve J M JM Hedehus M M Martin-McNulty B B Carano R A D RA Cao T C TC van Bruggen N N Bernstein L L Lee W P WP Wu X X DeVoss J J Zhang J J Jeet S S Peng I I McKenzie B S BS Roose-Girma M M Caplazi P P Diehl L L Webster J D JD Vucic D D

Cell death and differentiation 20160513 9

Necroptosis is a caspase-independent form of cell death that is triggered by activation of the receptor interacting serine/threonine kinase 3 (RIPK3) and phosphorylation of its pseudokinase substrate mixed lineage kinase-like (MLKL), which then translocates to membranes and promotes cell lysis. Activation of RIPK3 is regulated by the kinase RIPK1. Here we analyze the contribution of RIPK1, RIPK3, or MLKL to several mouse disease models. Loss of RIPK3 had no effect on lipopolysaccharide-induced s ...[more]

PMID: 27177019

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Project description:Necroptosis is a pro-inflammatory cell death, which happens in the context of caspase-8 inhibition, allowing activation of the receptor interacting protein kinase 1-receptor interacting protein kinase 3-mixed lineage kinase domain-like (RIPK1-RIPK3-MLKL) axis. Recently, necroptosis has emerged as a key component of resistance against pathogens including infected macrophage by Leishmania infantum, the ethiologic agent of Visceral leishmaniasis (VL). VL is the most severe form of Leishmaniasis, characterized by systemic inflammation and neutropenia. However, the role of neutrophil cell death in VL has not been characterized. Here, we showed that VL patients exhibited increased lactate dehydrogenase levels in the serum, a hallmark of cell death and tissue damage. We investigated the effect of necroptosis in neutrophil infection in vitro. Human neutrophils pretreated with zVAD-fmk (pan-caspase inhibitor) and zIETD-fmk (caspase-8 inhibitor) increased reactive oxygen species (ROS) level in response to Leishmania infection, which is associated with necroptotic cell death. MLKL, an important effector molecule downstream of necroptosis pathway, was also required for Leishmania killing. Moreover, in absence of caspases-8, murine neutrophils displayed loss of membrane integrity, higher levels of ROS, and decreased L. infantum viability. Pharmacological inhibition of RIPK1 or RIPK3 increased parasite survival when caspase-8 was blocked. Electron microscopy assays revealed morphological features associated with necroptotic death in L. infantum infected-neutrophils pretreated with caspase inhibitor, whereas infected cells pretreated with RIPK1 and RIPK3 inhibitors did not show ultra-structural alterations in membrane integrity and presented viable Leishmania within parasitophorous vacuoles. Taken together, these findings suggest that inhibition of caspase-8 contributes to elimination of L. infantum in neutrophils by triggering necroptosis. Thus, targeting necroptosis may represent a new strategy to control Leishmania replication.

Dataset Information

RIPK3 deficiency or catalytically inactive RIPK1 provides greater benefit than MLKL deficiency in mouse models of inflammation and tissue injury.

Publications

RIPK3 deficiency or catalytically inactive RIPK1 provides greater benefit than MLKL deficiency in mouse models of inflammation and tissue injury.

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