RIPK1 expression associates with inflammation in early atherosclerosis in humans and can be therapeutically silenced to reduce NF-κB activation and atherogenesis in mice

Karunakaran, D.; Nguyen, M.A.; Geoffrion, M.; Vreeken, D.; Lister, Z.; Cheng, H.S.; ... ; Rayner, K.J. (2021)

Article / Letter to editor

Background: Chronic activation of the innate immune system drives inflammation and contributes directly to atherosclerosis. Previously, we showed that macrophages in the atherogenic plaque undergo RIPK3-MLKL-dependent programmed necroptosis in response to sterile ligands such as oxidized LDL and damage-associated patterns (DAMPs) and necroptosis is active in advanced atherosclerotic plaques. Upstream of the RIPK3-MLKL necroptotic machinery lies RIPK1, which acts as a master switch that controls whether the cell undergoes NFκB-dependent inflammation, caspase-dependent apoptosis or necroptosis in response to extracellular stimuli. We therefore set out to investigate the role of RIPK1 in the development of atherosclerosis, which is largely driven by NFκB-dependent inflammation at early stages. We hypothesize that, unlike RIPK3 and MLKL, RIPK1 primarily drives NFκB-dependent inflammation in early atherogenic lesions and knocking down RIPK1 will reduce inflammatory cell activation and protect against the progression of atherosclerosis. Methods: We examined expression of RIPK1 protein and mRNA in both human and mouse atherosclerotic lesions, and using loss-of-function approaches in vitro in macrophages and endothelial cells to measure inflammatory responses. We administered weekly injections of RIPK1 anti-sense oligonucleotides (ASO) to Apoe^-/- mice fed a cholesterol-rich (Western) diet for 8 weeks. Results: We find RIPK1 expression is abundant in early-stage atherosclerotic lesions in both humans and mice. Treatment with RIPK1 ASOs led to a reduction in aortic sinus and en face lesion areas (47.2% or 58.8% decrease relative to control, p<0.01) and plasma inflammatory cytokines (IL-1α, IL-17A, p<0.05) compared to controls. RIPK1 knockdown in macrophages decreased inflammatory genes (NFκB, TNFα, IL-1α) and in vivo LPS- and atherogenic diet-induced NF-κB activation. In endothelial cells, knockdown of RIPK1 prevented NF-κB translocation to the nucleus in response to TNFα, where accordingly there was a reduction in gene expression of IL1B, E-selectin and monocyte attachment. Conclusions: We have identified RIPK1 as a central driver of inflammation in atherosclerosis by its ability to activate the NF-κB pathway and promote inflammatory cytokine release. Given the high levels of RIPK1 expression in human atherosclerotic lesions, our study suggests RIPK1 as a future therapeutic target to reduce residual inflammation in patients at high risk of coronary artery disease.

• All authors: Karunakaran, D.; Nguyen, M.A.; Geoffrion, M.; Vreeken, D.; Lister, Z.; Cheng, H.S.; Otte, N.; Essebier, P.; Wyatt, H.; Kandiah, J.W.; Jung, R.; Alenghat, F.J.; Mompeon, A.; Lee, R.; Pan, C.; Gordon, E.; Rasheed, A.; Lusis, A.J.; Liu, P.; Matic, L.P.; Hedin, U.; Fish, J.E.; Guo, L.; Kolodgie, F.; Virmani, R.; Gils, J.M. van; Rayner, K.J.
• Date: 2021-11-11
• Journal: Circulation
• Volume: 143
• Issue: 2
• Pages: 163 - 177
• DOI: doi:10.1161/circulationaha.118.038379
• Link: https://www.ahajournals.org/doi/10.1161/CIRCULATIONAHA.118.038379