Abstract
Vascular smooth muscle cell damage is a key step in inducing vascular calcification that yields hydroxyapatite (HAP) as a major product. The effect of the shape of HAP on the damage to vascular smooth muscle cells has yet to be investigated. In this study, we compared the differences in toxicity of four various morphological nano-HAP crystals, namely, H-Rod, H-Needle, H-Sphere, and H-Plate, in rat aortic smooth muscle cells (A7R5). The sizes of these crystals were 39 nm × 115 nm, 41 nm ×189 nm, 56 nm × 56 nm, and 91 nm × 192 nm, respectively. Results showed that all HAPs decreased cell viability, disorganized cell morphology, disrupted cell membranes, increased intracellular reactive oxygen species concentration, decreased mitochondrial membrane potential, decreased lysosome integrity, increased alkaline phosphatase activity, and increased intracellular calcium concentration, resulting in cell necrosis. The cytotoxicity of the four kinds of HAP was ranked as follows: H-Plate > H-Sphere > H-Needle > H-Rod. The cytotoxicity of each crystal was positively correlated with the following factors: large specific surface area, high electrical conductivity and low surface charge. HAP accelerated calcium deposits on the A7R5 cell surface and induced the expression of osteogenic proteins, such as BMP-2, Runx2, OCN, and ALP. The crystals with high cytotoxicity caused more calcium deposits on the cell surface, higher expression levels of osteogenic protein, and stronger osteogenic transformation abilities. These findings elucidated the relationship between crystal shape and cytotoxicity and provided theoretical references for decreasing the risks of vascular calcification.