MLKL and CaMKII Are Involved in RIPK3-Mediated Smooth Muscle Cell Necroptosis

Area of Science:

• Cardiovascular Biology
• Cell Death Pathways
• Molecular Signaling

Background:

• Receptor interacting protein kinase 3 (RIPK3)-mediated necroptosis in smooth muscle cells (SMCs) contributes to abdominal aortic aneurysm (AAA) pathogenesis.
• The precise downstream signaling cascade of RIPK3 in SMC necroptosis remains largely unelucidated.

Purpose of the Study:

• To investigate the roles of mixed lineage kinase domain-like pseudokinase (MLKL) and calcium/calmodulin-dependent protein kinase II (CaMKII) in RIPK3-dependent SMC necroptosis.
• To determine the sequential order of MLKL and CaMKII activation in this pathway.

Main Methods:

• Utilized a murine CaCl2-induced abdominal aortic aneurysm model.
• Employed in vitro TNFα plus zVAD-FMK stimulation of mouse aortic SMCs to induce necroptosis.
• Investigated the effects of MLKL and CaMKII inhibition/silencing on cell death and signaling events using siRNA and specific inhibitors.

Main Results:

• Both MLKL and CaMKII were phosphorylated in SMCs during AAA development and in vitro necroptosis induction.
• RIPK3 deficiency attenuated MLKL and CaMKII phosphorylation.
• MLKL knockdown reduced SMC necroptosis and CaMKII phosphorylation, while CaMKII inhibition/silencing partially inhibited necroptosis without affecting MLKL.
• MLKL activation appears to precede CaMKII activation in the RIPK3-mediated necroptosis pathway.

Conclusions:

• MLKL and CaMKII are both critical effectors in RIPK3-mediated SMC necroptosis.
• MLKL functions upstream of CaMKII in the signaling pathway leading to SMC death in AAA pathogenesis.