Evidence HDAC9 genetic variant associated with ischemic stroke increases risk via promoting carotid atherosclerosis

Abstract

Background and purpose: A novel association between a single nucleotide polymorphism on chromosome 7p21.1 and large-vessel ischemic stroke was recently identified. The most likely underlying gene is histone deacetylase 9 (HDAC9). The mechanism by which HDAC9 increases stroke risk is not clear; both vascular and neuronal mechanisms have been proposed.

Methods: We determined whether the lead single nucleotide polymorphisms were associated with asymptomatic carotid plaque (N=25 179) and carotid intima-media thickness (N=31 210) detected by carotid ultrasound in a meta-analysis of population-based and community cohorts. Immunohistochemistry was used to determine whether HDAC9 was expressed in healthy human cerebral and systemic arteries. In the Tampere Vascular Study, we determined whether HDAC9 mRNA expression was altered in carotid (N=29), abdominal aortic (N=15), and femoral (N=24) atherosclerotic plaques compared with control (left internal thoracic, N=28) arteries.

Results: Both single nucleotide polymorphisms (rs11984041 and rs2107595) were associated with common carotid intima-media thickness (rs2107595; P=0.0018) and with presence of carotid plaque (rs2107595; P=0.0022). In both cerebral and systemic arteries, HDAC9 labeling was seen in nuclei and cytoplasm of vascular smooth muscle cells, and in endothelial cells. HDAC9 expression was upregulated in carotid plaques compared with left internal thoracic controls (P=0.00000103). It was also upregulated in aortic and femoral plaques compared with controls, with mRNA expression increased in carotid compared with femoral plaques (P=0.0038).

Conclusions: Our results are consistent with the 7p21.1 association acting via promoting atherosclerosis, and consistent with alterations in HDAC9 expression mediating this increased risk. Further studies in experimental models are required to confirm this link.

Figure 1

Immunohistochemical labelling of HDAC9 in human middle cerebral artery Post mortem sample of human middle cerebral artery labelled for HDAC9 (panel A) or with no primary antibody (B). A: cellular HDAC9 immunoreactivity (brown) is evident in endothelial cells (examples marked with arrows) and in myoctes within the medial layer (examples marked with arrowheads). B: labelling was absent from neighbouring sections treated identically but with no primary antibody. Scale bars 20 μm

Figure 2

Immunohistochemical labelling of HDAC9 in human aorta. Cellular HDAC9 immunoreactivity (brown) in adult human aorta (A) was absent from a neighbouring negative control section (B). C, higher magnification shows HDAC9 in VSMC (arrowheads) and endothelial cells (arrows). In VSMC a similar pattern of labelling is seen with smooth muscle myosin (SMM; D). Haematoxylin counterstain (blue) labels nuclear chromatin. Scale bars 100 μm (A,B), 20 μm (C,D).