Loss of CDKN2B Promotes p53-Dependent Smooth Muscle Cell Apoptosis and Aneurysm Formation

Abstract

Objective Genome wide association studies have implicated allelic variation at 9p21.3 in multiple forms of vascular disease, including atherosclerotic coronary heart disease and abdominal aortic aneurysm. As for other genes at 9p21.3, human eQTL studies have associated expression of the tumor suppressor gene CDKN2B with the risk haplotype, but its potential role in vascular pathobiology remains unclear. Methods and Results Here we employed vascular injury models and found that Cdkn2b knockout mice displayed the expected increase in proliferation after injury, but developed reduced neointimal lesions and larger aortic aneurysms. In situ and in vitro studies suggested that these effects were due to increased smooth muscle cell apoptosis. Adoptive bone marrow transplant studies confirmed that the observed effects of Cdkn2b were mediated through intrinsic vascular cells and were not dependent on bone marrow-derived inflammatory cells. Mechanistic studies suggested that the observed increase in apoptosis was due to a reduction in MDM2 and an increase in p53 signaling, possibly due in part to compensation by other genes at the 9p21.3 locus. Dual inhibition of both Cdkn2b and p53 led to a reversal of the vascular phenotype in each model. Conclusions These results suggest that reduced CDKN2B expression and increased SMC apoptosis may be one mechanism underlying the 9p21.3 association with aneurysmal disease.

Cyclin-dependent kinase inhibitor 2B regulates efferocytosis and atherosclerosis

“…CDKN2B is a well-described cell cycle inhibitor that is frequently lost during malignant transformation, but had not previously been implicated in vascular disease. We recently reported that CDKN2B does play a role in vascular SMC physiology and that Cdkn2b knockout mice develop advanced aneurysms related to accelerated SMC apoptosis and medial thinning (16). Because the mechanism(s) by which 9p21 promote coronary disease remain unclear and the subject of debate (17)(18)(19)(20)(21)(22), we hypothesized that CDKN2B's effects on programmed cell death contribute to the development of CAD by stimulating the accumulation of necrotic debris within the evolving plaque.…”

Genetics of Coronary Artery Disease

“…A third kinase inhibitor, ARF, is an alternate reading frame of the CDKN2a gene, which inhibits the cell cycle by either the retinoblastoma pathway or the p53 pathway arresting the cell cycle in G1 and G2 phases, and also inhibits apoptosis. 58 ARF does this by degrading MDM2, a protein common to both pathways. All of these cell cycle kinase inhibitors are found in many tissues, including coronary vasculature, adipose tissue, and circulating leukocytes.…”

FOXE3 mutations predispose to thoracic aortic aneurysms and dissections

“…We then sought to determine whether p53 inhibition could prevent aortic apoptosis in vivo by injecting PFT (2.2 mg/kg) i.p. into WT and Foxe3 -/-mice 24 hours before TAC and every 48 hours after TAC (21). PFT significantly reduced the incidence of aortic rupture and intramural hematomas in Foxe3 -/-mice that underwent TAC (50% to 17%, P < 0.05).…”

“…PFT, a pharmacological p53 inhibitor, was purchased from (EMD Millipore) and diluted in PBS. An in vitro p53 inhibition study was performed by adding 10 μM PFT to the cell culture media as previously described (21). Rates of programmed cell death were assessed with 3 independent in vitro assays.…”

“…injection at a dosage of 2.2 mg/kg, then dissolved in PBS 1 hour before TAC and then every 48 hours. Animals were euthanized 2 weeks after the surgery, and the ascending aortic tissues were harvested for either RNA, total protein, histomorphometric analysis, or TUNEL assay, as described previously (21).…”