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bioRxiv · 10.1101/2022.11.24.517638

Inhibition of MLKL impairs abdominal aortic aneurysm development by attenuating smooth muscle cell necroptosis

Nemade, H. N.·Mehrkens, D.·Lottermoser, H. S.·Yilmaz, Z. E.·Schelemei, P.·Picard, F. R.·Geissen, S.·Schwab, G. F.·Hoyer, F. F.·Guthoff, H.·Hof, A.·Nettersheim, F. S.·Sachinidis, A.·Winkels, H.·Baldus, S.·Pasparakis, M.·Adam, M.·Mollenhauer, M.
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Abstract

BackgroundReceptor-interacting serine/threonine-protein kinase 1 and 3 (RIPK1 and RIPK3) dependent cell death has been identified as a crucial mediator of abdominal aortic aneurysm (AAA) development. RIPK3 mediates phosphorylation of Mixed lineage kinase domain like pseudokinase (MLKL) thereby inducing its oligomerization and translocation to the cell membrane. Given the dual role of RIPKs being involved in necroptosis as well as in apoptosis induction, the specific role of MLKL-induced necroptotic cell death in AAA remains unclear. MethodsWe monitored elastase-perfusion (PPE) induced progression of AAA in C57BL/6N (WT), RIPK1 kinase-inactive (Ripk1D138N/D138N), MLKL knockout (Mlkl-/-) and MLKL phospho-deficient (MlklAA) mice by ultrasound measurements, histological analyses and bulk mRNA sequencing to assess structural and molecular aortic changes. Bone marrow transplantations in WT and MlklAA mice were utilized to dissect the role of MLKL in smooth muscle cells (SMCs) and myeloid cells in AAA development. MLKL expressing human SMCs were generated to investigate necroptosis-induced proinflammatory cytokine secretion and subsequent polymorphonuclear neutrophil (PMN) migration and activation in vitro. ResultsUltrasound analysis showed that ~70% of the WT animals developed PPE induced-AAA with significant aortic structural alterations and enhanced myeloid cell infiltration. In contrast, Ripk1D138N/D138N, MlklAA, and Mlkl-/- mice were protected from AAA. This protection was associated with reduced adverse extracellular matrix (ECM) remodeling and leukocyte infiltration. MLKL deficiency was associated with a significant downregulation of genes involved in fibrinolysis, anti-inflammatory response, immune response and complement activation in aortic tissue in AAA. Bone marrow transplantation studies showed the lack of MLKL in SMCs to be the main driver of AAA protection. Proinflammatory cytokine secretion was elevated in necroptosis induced SMCs and resulted in a significant accumulation and activation of PMN. ConclusionsOverall, these findings indicate that MLKL-induced necroptotic SMC death and subsequent proinflammatory leukocyte activation play a causative role in AAA development and suggest that pharmacological inhibition of MLKL may represent a promising treatment strategy for AAA disease.

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Nemade, H. N., Mehrkens, D., Lottermoser, H. S., Yilmaz, Z. E., Schelemei, P., Picard, F. R., Geissen, S., Schwab, G. F., Hoyer, F. F., Guthoff, H., Hof, A., Nettersheim, F. S., Sachinidis, A., Winkels, H., Baldus, S., Pasparakis, M., Adam, M., Mollenhauer, M.. 2022-11-24. Inhibition of MLKL impairs abdominal aortic aneurysm development by attenuating smooth muscle cell necroptosis. https://doi.org/10.1101/2022.11.24.517638

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