Transmission microscopy of freeze dried, unstained epiphyseal cartilage of the guinea pig

Abstract

The question of the initial mineralization in the epiphyseal plate has been investigated to date in specimens prepared by conventional electron microscopical techniques. As conventional techniques can produce artifacts, either a loss of mineral substance or a secondary nucleation, the mineralization process was investigated using freeze dried, vacuum embedded growth cartilage which was neither contrasted not stained and which had a very short contact with water. The prevailing theory that the first mineralization begins within extracellular matrix vesicles and that the mineralization outside these vesicles is a secondary process was confirmed. Mineralized matrix vesicles were found in the fully mineralized long septa down to the opening zone. In several cases a mineralization could be observed in those transverse septa in which organic substance was present between the cells. The typical radial arrangement of the apatitic needles and platelets in the matrix vesicles could be explained by the formation of a mineralization in an ionotropic gel, the orientation of the matrix macromolecules to be produced by a vectorial influx of calcium ions and phosphate groups coming from different directions. Thin strands of mineral substance with low contrast, which follow the direction of the longitudinal septum, were assumed to be the mineralized collagen fibrils. In several needles dot-like formations were seen and the distance between the middle of neighbouring dots was found to lie mainly in the range 30-56 A, while the lateral separation distance between the middle of closely packed parallel chains and needles was found to lie mainly in the range 30-42 A. Parallel periodic structures which could be visualized in apatitic chains and needles 20-40 A in diameter were assumed to be the 8.2 A-(100)-lattice planes of apatite, being an indication that these formations already possess criteria of the apatite lattice.