RIPK1 or RIPK3 deletion prevents progressive neuronal cell death and improves memory function after traumatic brain injury

Antonia Clarissa Wehn,Igor Khalin,Nikolaus Plesnila,Carsten Culmsee,Peter Vandenabeele,Marco Duering,Farida Hellal,Nicole Angela Terpolilli

doi:10.1186/s40478-021-01236-0

Abstract

Traumatic brain injury (TBI) causes acute and subacute tissue damage, but is also associated with chronic inflammation and progressive loss of brain tissue months and years after the initial event. The trigger and the subsequent molecular mechanisms causing chronic brain injury after TBI are not well understood. The aim of the current study was therefore to investigate the hypothesis that necroptosis, a form a programmed cell death mediated by the interaction of Receptor Interacting Protein Kinases (RIPK) 1 and 3, is involved in this process. Neuron-specific RIPK1- or RIPK3-deficient mice and their wild-type littermates were subjected to experimental TBI by controlled cortical impact. Posttraumatic brain damage and functional outcome were assessed longitudinally by repetitive magnetic resonance imaging (MRI) and behavioral tests (beam walk, Barnes maze, and tail suspension), respectively, for up to three months after injury. Thereafter, brains were investigated by immunohistochemistry for the necroptotic marker phosphorylated mixed lineage kinase like protein(pMLKL) and activation of astrocytes and microglia. WT mice showed progressive chronic brain damage in cortex and hippocampus and increased levels of pMLKL after TBI. Chronic brain damage occurred almost exclusively in areas with iron deposits and was significantly reduced in RIPK1- or RIPK3-deficient mice by up to 80%. Neuroprotection was accompanied by a reduction of astrocyte and microglia activation and improved memory function. The data of the current study suggest that progressive chronic brain damage and cognitive decline after TBI depend on the expression of RIPK1/3 in neurons. Hence, inhibition of necroptosis signaling may represent a novel therapeutic target for the prevention of chronic post-traumatic brain damage.

Highlights

With an estimated case load of 69 million per year [1], traumatic brain injury (TBI) represents a leading causeWehn et al acta neuropathol commun (2021) 9:138 memory deficits, and loss of fine motor skills, have difficulties returning to their previous occupation, and require lifelong support [4,5,6]
Three months after TBI large amounts of Phosphorylated MLKL (pMLKL) were found in the rim of the traumatic cavity, the presumed site of progressive chronic post-trauma brain damage (Fig. 2a, upper panel). pMLKL was found by high resolution confocal imaging in the cytoplasm of selected neurons as small dots, suggesting that pMLKL is part of a protein complex such as the necrosome (Fig. 2a, lower panel, white arrowheads). pMLKL staining was almost absent in neuronal RIPK3 deficient mice suggesting that necroptotic signaling in neurons did essentially not occur in these animals (Fig. 2b)
Chronic posttraumatic brain damage is reduced in RIPK1 or RIPK3 deficient mice After demonstrating neuronal necroptotic signaling three months after TBI in wild type mice and showing that RIPK1 or 3 deficiency prevented this process, we evaluated lesion volume in cortex and hippocampus of wild type and Receptor Interacting Protein Kinases (RIPK) deficient mice by longitudinal MR imaging

Summary

Introduction

With an estimated case load of 69 million per year [1], traumatic brain injury (TBI) represents a leading causeWehn et al acta neuropathol commun (2021) 9:138 memory deficits, and loss of fine motor skills, have difficulties returning to their previous occupation, and require lifelong support [4,5,6]. While the pathophysiology of acute brain damage has been investigated in detail in experimental animals and in humans over the past decades, relatively little is known about the mechanisms determining long-term outcome after TBI. Chronic functional deficits in TBI patients may be caused by progressive brain atrophy in cortex and hippocampus [8, 9] and hydrocephalus formation [10,11,12,13]. Clinical and experimental studies suggest that inflammation plays an important role for the development of chronic posttraumatic brain damage [14,15,16,17,18], the cellular and molecular mechanisms downstream of this process are not fully understood. The trigger and the intracellular signaling cascades causing neuronal cell death weeks and months after TBI are still unknown

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