Abstract 16302: Notch3 Signaling Promotes M1 Macrophage Activation and Atherosclerosis: A Novel Therapeutic Target
Background: Pro-inflammatory (“M1”) macrophages may exert atherogenic effects while “M2” macrophages may be anti-inflammatory. The Notch pathway, involving four receptors (Notch1-4), regulates various biological processes. We previously used siRNA silencing to suggest in vitro that Notch3 skews macrophages towards M1 phenotype.
Methods and Results: Testing the hypothesis that Notch3 promotes M1 macrophage activation and athergenesis employed gain-of-function approaches in vitro and in vivo. Transient expression of constitutively active Notch3 in the murine macrophage cell line RAW 264.7 induced various pro-inflammatory genes, including IL-1β (DNA microarray analysis, qPCR). Enforced expression of a dominant negative form of RBP-Jκ or MAML1, transcriptional regulators for the canonical Notch pathway, abrogated Notch3-mediated macrophage activation. To explore in vivo evidence, we established macrophage-selective transgenic mice of the constitutively active Notch3 (Notch3tg). Peritoneal macrophages of Notch3tg mice showed features of M1 phenotype (e.g., increased expression of IL-1β and iNOS and NF-κB activation, decreased M2 marker arginase 1, Figure A) compared to cells from wild type littermates (n=6 each). In Notch3tg mice crossed with atherosclerosis-prone LDL receptor-deificient mice (Notch3tg:Ldlr-/-), atherosclerotic plaques were larger and contained more lipids (oil red O) and macrophages (Mac3 immunostain) than Ldlr-/- mice (n=10 each, Figiure B). Macrophages of Notch3tg:Ldlr-/- mice showed increased expression of the scavenger receptor CD36 and accelerated foam cell formation. The aorta of Notch3tg:Ldlr-/- mice showed greater alkaline phosphatase activity and more advanced calcification (von Kossa stain) than did Ldlr-/- mice (n=10 each, p<0.05).
Conclusion: Notch3 signaling promotes M1 macrophage activation and atherosclerosis, offering a new therapeutic target for cardiometabolic inflammation.
- © 2013 by American Heart Association, Inc.