Matrix Vesicles Mediate Mineralization of Human Thyroid Cartilage

Abstract.

Mineralization and ossification of human thyroid cartilage first starts after the end of adolescence when the previously cartilaginous human skeleton has become ossified and the epiphyseal discs are in the process of closing. However, the mechanisms involved in mineralization and ossification of human thyroid cartilage are not well understood. Ultrastructural analysis of human thyroid cartilage revealed that mineralization started close to cartilage canals in a matrix containing gigantic collagen fibers (asbestoid fibers). Matrix vesicles were detected in mineralized areas and were often associated with needle-like crystals. For the first time we were able to isolate matrix vesicles from human thyroid cartilage by mild enzymatic digestions and ultracentrifugation. These particles were oval and varied in size; some were heavily calcified. They were enriched in alkaline phosphatase, calcium, and inorganic phosphate, suggesting that the particles contain Ca²⁺-P_i complexes. Immunoblot analysis of these vesicles revealed the presence of annexins II, V, and VI, membrane-associated, channel-forming proteins, which allow influx of Ca²⁺ into the vesicles and intralumenal crystal growth. In addition, the vesicles were associated with types II and X collagen, suggesting that this association not only anchors the vesicles to the extracellular matrix, but, as shown previously, also stimulates Ca²⁺ influx into these particles. In conclusion, matrix vesicles isolated from human thyroid cartilage contain all the components, enabling them to initiate and mediate the mineralization process in human thyroid cartilage.