Cell Death and Disease (Nov 2021)

Genetic inhibition of RIPK3 ameliorates functional outcome in controlled cortical impact independent of necroptosis

  • Limin Wu,
  • Joon Yong Chung,
  • Tian Cao,
  • Gina Jin,
  • William J. Edmiston,
  • Suzanne Hickman,
  • Emily S. Levy,
  • Jordyn A. Whalen,
  • Eliza Sophie LaRovere Abrams,
  • Alexei Degterev,
  • Eng H. Lo,
  • Lorenzo Tozzi,
  • David L. Kaplan,
  • Joseph El Khoury,
  • Michael J. Whalen

DOI
https://doi.org/10.1038/s41419-021-04333-z
Journal volume & issue
Vol. 12, no. 11
pp. 1 – 16

Abstract

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Abstract Traumatic brain injury (TBI) is a leading cause of death and disability with no specific effective therapy, in part because disease driving mechanisms remain to be elucidated. Receptor interacting protein kinases (RIPKs) are serine/threonine kinases that assemble multi-molecular complexes that induce apoptosis, necroptosis, inflammasome and nuclear factor kappa B activation. Prior studies using pharmacological inhibitors implicated necroptosis in the pathogenesis of TBI and stroke, but these studies cannot be used to conclusively demonstrate a role for necroptosis because of the possibility of off target effects. Using a model of cerebral contusion and RIPK3 and mixed lineage kinase like knockout (MLKL −/−) mice, we found evidence for activation of RIPK3 and MLKL and assembly of a RIPK1-RIPK3-MLKL necrosome complex in pericontusional brain tissue. Phosphorylated forms of RIPK3 and MLKL were detected in endothelium, CD11b + immune cells, and neurons, and RIPK3 was upregulated and activated in three-dimensional human endothelial cell cultures subjected to CCI. RIPK3 −/− and MLKL −/− mice had reduced blood-brain barrier damage at 24 h (p < 0.05), but no differences in neuronal death (6 h, p = ns in CA1, CA3 and DG), brain edema (24 h, p = ns), or lesion size (4 weeks, p = ns) after CCI. RIPK3 −/−, but not MLKL −/− mice, were protected against postinjury motor and cognitive deficits at 1–4 weeks (RIPK3 −/− vs WT: p < 0.05 for group in wire grip, Morris water maze hidden platform trials, p < 0.05 for novel object recognition test, p < 0.01 for rotarod test). RIPK3 −/− mice had reduced infiltrating leukocytes (p < 0.05 vs WT in CD11b + cells, microglia and macrophages), HMGB1 release and interleukin-1 beta activation at 24–48 h (p < 0.01) after CCI. Our data indicate that RIPK3 contributes to functional outcome after cerebral contusion by mechanisms involving inflammation but independent of necroptosis.