DNA damage response, Runx2 and vascular calcification

Vascular calcification ultimately robs the arterial wall of its elasticity, rendering the arteries stiff and hardened ^{1, 2}. The physiological changes resulting from such hardening lead to difficult-to-treat hypertension and congestive heart failure. If occurring in the setting of atherosclerosis, it furthers destabilization of plaques and interferes with interventions aimed at reducing coronary obstructions. Patients that are especially vulnerable to vascular calcification include those suffering from chronic kidney disease, diabetes and hypertension; disorders that often travel together as we age. Indeed, clinical studies have reported that arterial calcification is present in more than 90% of men and 67% women over 70 years of age ³. Calcification of the vascular media (media sclerosis) is widely accepted as a cell-driven process, where VSMCs (vascular smooth muscle cells) are phenotypically modulated and undergo transition into osteogenic cells, or alternatively, undergo apoptosis. Although many calcific pathways have been elucidated, clear gaps exist in our understanding of how pathways interact with pro-calcific conditions and how pathology can modify cellular responses.

In this issue of ATVB, Shanahan and her colleagues put the spotlight on the connection between nuclear DNA damage, the DDR (DNA damage response) and how these may enhance a relatively mild calcific stimuli through interference with Runx2 (Runt-related transcription factor 2) ⁴. High levels of oxidative stress and propensity for stress-induced DNA damage are linked to premature onset of age-related disorders ⁵, among them atherosclerosis, diabetic vasculopathy, and aortic valve disease, all with elements of calcification. Although it could also be the result of what we perceive as normal aging, eventually affecting us all to a certain extent.

Runx2 is an essential transcription factor in bone development and osteogenic differentiation ⁶. It acts as a scaffold to organize enhancers and repressors and its activity depends on the localization in the nuclear matrix. Despite its importance during the developmental period, later activation in the vascular wall may have deleterious effects and trigger vascular calcification ⁷; an event that can be prevented in mice by deleting the Runx2 gene ⁸. In highly mechanistic studies, the Shanahan team focused on the role of Runx2 in the DDR in VSMCs and how it might add to an already calcific milieu ⁴. Their thought-provoking results suggest that Runx2 is a direct link between DDR, osteogenesis and apoptosis in VSMCs.

Their results reveal that low to moderate DNA damage (genotoxic stress) in VSMCs that were already subjected to calcification stimuli (such as elevated levels of phosphate), results in poly(ADP-ribose)ylation (PARylation) of the Runx2 protein. It increases the accumulation of Runx2, targets downstream osteogenic genes, and promotes the reprogramming of the VSMCs (Figure 1 for schematic model). Runx2 localizes to sites of DNA damage and participates in the DNA repair by regulating phosphorylation events on histone H2AX. However, an overwhelming or persistent DNA injury that leads to more Runx2 also promotes the activation of JNK dependent apoptosis through retention of p142-H2AX and suppression of DNA repair. Thus, Runx2 drives vascular calcification through a combination of osteogenic transition and apoptosis, implicating Runx2 as an opportunistic and deleterious driver of vascular calcification as part of the DDR in VSMCs.

Figure 1. Schematic model of the role of Runx2 in the DNA damage response (DDR) in vascular smooth muscle cells (VSMCs).

The addition of genotoxic stress to VSMCs in a pro-calcific milieu triggers the DDR with PARylation and accumulation of Runx2. Low to moderate DNA damage increases the Runx2-mediated induction of osteogenic target genes, whereas persistent or excessive DNA damage results in excessive Runx2 and an enhancement of JNK-mediated apoptosis through retention of p142-H2AX and suppression of DNA repair.

ATM, ataxia-telangiectasia, mutated; PARP, poly(ADP-ribose) polymerase; PARylation, poly(ADP-ribose)ylation; ROS, reactive oxygen species.

The findings suggest a number of novel concepts, including how a cellular repair system can manipulate a transcription factor like Runx2 to specify the phenotype of VSMCs. Furthermore, the studies delineate how ATM (ataxia-telangiectasia, mutated) ⁹ and PARP [poly(ADP-ribose) polymerase] ¹⁰modify Runx2 to carry out the VSMC reprogramming and/or JNK mediated apoptosis. This provides insight into the interactions between pathways that trigger calcification and pathological milieus that are created by disease and can be mitigated by treating the disease. The findings may also shed light on other types of calcification such as calcific aortic valve disease and calcifying scars after myocardial infarction or pulmonary tuberculosis.

In revealing the link between the DDR and Runx2, the study provides clues that could assist in the development of strategies that limit the DDR in calcific milieus. Please note that it is not a given that aging must calcify our vasculature. Even a mild reduction or delay in the hardening of the vessels, or small increases in elasticity and contractility, could have profound benefits in maintaining a favorable physiology as we age. Future applications related to the DDR and Runx2 are likely to prove to be most useful in patients with the highest risk of calcification.

Thus, even though the prerequisite of Runx2 exists during bone development and supersedes concerns for the development of a less desirable “vascular skeleton” later in life, these studies should provide justification for further studies on how to regulate this transcription factor.