Mechanisms of focal calcification in rabbit aortas: role of calcifying vesicles

We previously demonstrated that calcification in rabbit thoracic aortas is initiated in the lower zone of the extensively thickened plaques (LZP) adjacent to the media. Whether osteogenesis or a local increase in calcifying vesicles underlies the focal calcification remains to be established.

To determine that focal calcification is related to osteogenesis, an obligatory osteogenic biomarker, alkaline phosphatase (ALP) activity, was evaluated in the unfixed thoracic aorta sections of rabbits fed a supplemental cholesterol diet and of humans with advanced atherosclerosis. To determine whether blood shortage to the smooth muscle cells (SMCs) imposed by intima thickening may increase calcifying vesicles, the effect of serum deprivation on the biogenesis of calcifying vesicles in cultured SMCs was investigated.

(1) In contrast to positive rabbit kidney cross sections and consistent with the activity in various isolated subcellular fractions of aortas, ALP activity was absent in the media, adventitia, lesions, and LZP of rabbit aortas or in the fibrointima of human aortas. (2) Histologic assessments of the lesions indicate the absence of bone cells or osteoid. (3) Depletion of the serum from culture media caused a twofold increase in the levels of ALP-deficient and adenosine triphosphatase-rich calcifying vesicles, which were released from the cells by treatment with 0.05% trypsin-0.02% ethylenediaminetetraacetic acid for 15 minutes.

(1) Focal calcification in rabbit aortas and diffuse mineralization in human fibrointima are not associated with osteogenesis, and (2) blockade of the blood supply to SMCs may trigger the cells to produce more calcifying vesicles, thereby leading to site-specific calcification in rabbit aortas.