Fig. 3.

Differences in the ultrastructure of VSMC calcification and the mineralised bone nodules formed by osteoblasts. Cultures of (A) early osteoblasts and (B) non-calcifying VSMCs showed no evidence of any extracellular mineralisation. (C) In bone-forming osteoblasts there was widespread collagen [COL] deposition and clear mineralised matrix which formed part of a bone nodule [BN]. (D) Cultures of calcifying VSMCs contained amorphous regions of calcification [Cal] and extracellular fibril-like structures that appeared to be cellular debris [CD] but no deposited collagen fibres. (E) Higher power images showed that mineralised matrix [MM] was formed in and around the collagen fibres [COL] in bone-forming osteoblasts. (F) The extracellular calcification [Cal] present in VSMCs was less widespread, morphologically distinct from the osteoblast bone nodules and was sometimes associated with filamentous cellular debris [CD]. (G) In osteoblasts, matrix vesicles [MV] were seen associated with collagen fibres [COL]. (H) In calcifying VSMCs, extracellular vesicles [EV] were observed associated with filamentous cellular debris [CD]. Images shown are representative of the calcification and matrix deposition observed in three different cell preparations. Scale bars: A-B = 5  μm, C-D = 2 μm, E-F = 1  μm, G-H = 200 nm.