JAGGED1/NOTCH3 activation promotes aortic hypermuscularization and stenosis in elastin deficiency
Jui M. Dave, … , Kathleen A. Martin, Daniel M. Greif
Jui M. Dave, … , Kathleen A. Martin, Daniel M. Greif
Published January 6, 2022
Citation Information: J Clin Invest. 2022;132(5):e142338. https://doi.org/10.1172/JCI142338.
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Research Article Vascular biology

JAGGED1/NOTCH3 activation promotes aortic hypermuscularization and stenosis in elastin deficiency

Abstract

Obstructive arterial diseases, including supravalvular aortic stenosis (SVAS), atherosclerosis, and restenosis, share 2 important features: an abnormal or disrupted elastic lamellae structure and excessive smooth muscle cells (SMCs). However, the relationship between these pathological features is poorly delineated. SVAS is caused by heterozygous loss-of-function, hypomorphic, or deletion mutations in the elastin gene (ELN), and SVAS patients and elastin-mutant mice display increased arterial wall cellularity and luminal obstructions. Pharmacological treatments for SVAS are lacking, as the underlying pathobiology is inadequately defined. Herein, using human aortic vascular cells, mouse models, and aortic samples and SMCs derived from induced pluripotent stem cells of ELN-deficient patients, we demonstrated that elastin insufficiency induced epigenetic changes, upregulating the NOTCH pathway in SMCs. Specifically, reduced elastin increased levels of γ-secretase, activated NOTCH3 intracellular domain, and downstream genes. Notch3 deletion or pharmacological inhibition of γ-secretase attenuated aortic hypermuscularization and stenosis in Eln–/– mutants. Eln–/– mice expressed higher levels of NOTCH ligand JAGGED1 (JAG1) in aortic SMCs and endothelial cells (ECs). Finally, Jag1 deletion in SMCs, but not ECs, mitigated the hypermuscular and stenotic phenotype in the aorta of Eln–/– mice. Our findings reveal that NOTCH3 pathway upregulation induced pathological aortic SMC accumulation during elastin insufficiency and provide potential therapeutic targets for SVAS.

Authors

Jui M. Dave, Raja Chakraborty, Aglaia Ntokou, Junichi Saito, Fatima Z. Saddouk, Zhonghui Feng, Ashish Misra, George Tellides, Robert K. Riemer, Zsolt Urban, Caroline Kinnear, James Ellis, Seema Mital, Robert Mecham, Kathleen A. Martin, Daniel M. Greif

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Figure 3

Elastin deficiency reduces DNA methylation (5mC) epigenetic mark and downregulates DNMT1 levels.

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Elastin deficiency reduces DNA methylation (5mC) epigenetic mark and dow...
(A) Transverse sections of ascending aorta from WT or Eln–/– pups at P0.5 stained for 5-methylcytosine (5mC), SMA, and propidium iodide (PI, nuclei). Lu, lumen. Scale bar: 50 μm. (B) Histogram representing the percentage of 5mC+ SMCs in A (n = 3 mice). **P < 0.01 vs. WT by Student’s t test. (C–E) haSMCs were pretreated with Scr or siELN and then lysates were analyzed. In C, histogram depicts transcript levels of DNMT1, DNMT3A, and DNMT3B from qRT-PCR relative to 18S rRNA and normalized to Scr (n = 6). **P < 0.01 vs. Scr by Student’s t test. (D) Western blots for DNMT1 and GAPDH and densitometry of protein bands in E relative to GAPDH and normalized to Scr (n = 4). ***P < 0.001 vs. Scr by Student’s t test. (F and G) Aortic lysates from WT or Eln–/– mice at P0.5 analyzed by Western blotting for DNMT1 and GAPDH (F) with densitometry of protein bands relative to GAPDH and normalized to WT (G) (n = 3 mice). **P < 0.01 vs. WT by Student’s t test. (H) Genomic DNA was isolated from haSMCs pretreated with Scr or siELN and then subjected to methylated DNA chromatin immunoprecipitation (5mC ChIP). Histogram represents 5mC levels at the promoter regions of PSEN1, PSEN2, or THS2B (constitutive 5mC positive control) as assessed by qPCR and normalized to Scr (n = 4). **P < 0.01, ***P < 0.001 vs. Scr by Student’s t test. All data are averages ± SD.