[Ultrastructural studies of initial calcification of mouse fetal parietal bone in vitro]

The parietal bone fragment (1.5 mm square) extracted from murine fetal calvaria (periosteum was removed) in the latter stage of gestation was cultured in alpha-MEM containing 10% fetal bovine serum over 10-day period, and the initial calcification site was subsequently examined utilizing both electron microscopy and histochemical methods. At the initial calcification site, the so-called "matrix vesicle" (width 100 nm), derived from osteoblasts which induce matrix formation, and the larger vesicular structures (width 100-600 nm) which originated from degenerated or dead cells were observed. Additionally dead osteoblast-like cells phagocytosing bone crystals participated in bone nodul formation. In the border region between the degenerated or disrupted cells and the functioning cells, multilamellar structures (mainly composed of phospholipids) were frequently observed. Concurrently it was concluded that there is a possibility of partial deposition of calcium salts on the surface of the above mentioned cells under some circumstances. Through selective decalcification, it was determined that bone trabecule was formed by fusion of respective bone noduli and degraded collagen fibrils which filled the intervening spaces. From the above results, it was evident that both the matrix vesicles derived from functioning osteoblast, and the vesicular structures derived from the degenerated or dead cells concurrently phagocytosing spicules and collagen fibrils were involved in the initial calcification process of the membranous bone in vitro.