Tissue-nonspecific Alkaline Phosphatase (TNAP)-induced Vascular Calcification and Atherosclerosis - Project Summary
Vascular calcification is associated with coronary artery disease and poor prognosis. This prompted us to
examine whether calcification can be a pathogenic factor in atherosclerosis. We developed a mouse model, in
which overexpression of tissue-nonspecific alkaline phosphatase (TNAP) in endothelial cells prompted the
development of complex calcified arterial lesions that originated as macro-calcific nodules in the intima. We
then tested the idea that pathologic micro-calcification can precede atherosclerosis by challenging endothelial
TNAP mice with high cholesterol diet. We documented that animals with pre-existing intimal calcification
developed accelerated coronary artery disease, whereas equally hyperlipidemic mice without TNAP
overexpression displayed a typical course of murine atherosclerosis, in which coronary arteries were
unaffected. Pharmacological inhibition of TNAP under these conditions by a specific chemical inhibitor SBI-425
reduced coronary atherosclerosis, normalized heart fraction, and improved survival. Analyzing human
myocardial samples, we detected TNAP activity in the endothelium of small coronary arteries and arterioles.
Our significant findings regarding TNAP, calcification and atherogenesis merit further investigation of the role
of this enzyme in vascular disease. Our overall hypothesis is that under conditions that promote pathologic
vascular calcification (e.g. chronic kidney disease, diabetes, and advanced age), TNAP activity might induce
micro-calcification in the intima that can accelerate lipid retention and inflammation and that inhibition of TNAP
activity could be a valuable therapeutic goal for the reduction of atherosclerosis in patients predisposed to
vascular calcification. Here we propose to determine the prevalence of intimal micro-calcification in a large
sample of individuals over the age of 65; using 250 human cadavers specimens (Aim 1.1). Going back to the
animal study and using computational flow dynamics (CFD) simulations, we will compute how micro-scale
endothelial roughness (experimentally determined by submicron-resolution optical scanning) affects shear
stress distribution. These experiments should extend our understanding of the relationship between vascular
calcification and atherogenesis associated with low shear fields – a fundamental pathophysiological process
(Aim 1.2). A potential interaction between adenosine (another product of TNAP) and alpha adrenergic signaling
in the regulation of vascular tone will also be examined to explain increased sensitivity of TNAP-positive
arteries to sympathomimetic stimulation and impaired relaxation (Aim 2.3). In Aim 2, we will interrogate TNAP
axis in a chronic kidney disease model both genetically, by targeted deletion of TNAP in endothelial cells (Aim
2.1), and pharmacologically, by SBI-425 (Aim 2.2). Upon completion of this work we will establish that TNAP-
induced vascular calcification can mechanistically precede atherosclerosis. We will also validate TNAP
inhibition as a viable pharmacological approach that could potentially be used to treat many human conditions
associated with both increased calcification and atherosclerosis.
