Vertical distribution of elements in cells and matrix of epiphyseal growth plate cartilage determined by quantitative electron probe analysis.

Quantitative electron probe analysis was performed on chick epiphyseal growth cartilage prepared by two anhydrous methods, ultrathin cryosections and freeze-dried epoxy-embedded tissue. Levels of Na, Mg, P, S, Cl, K, and Ca were determined in cytoplasm, mitochondria, extracellular matrix, matrix vesicles, and mineral nodules in four zones of the cartilage--proliferative, prehypertrophic, early hypertrophic, and early calcification. The exceptionally high levels of Na and K (up to 550 and 200 mmol/kg wet wt, respectively) found in the matrix are believed to be largely bound to fixed anions. Within cells, Na was higher than K (140 versus 20-34 mmol/kg wet wt), a condition that may reflect hypoxia. Ca and P were low in cells and unmineralized matrix. Ca and P were high in mitochondrial granules of the early hypertrophic zone and diminished in amount in the calcifying zone; the converse occurred in matrix vesicles. Mg was low to undetectable except in heavily mineralized structures (i.e., mitochondrial granules, matrix vesicles, and mineral nodules). S levels were high in matrix (approximately 400 mmol/kg wet wt) and increased slightly with maturation. The amount of S present greatly exceeds Ca levels and implies that sulfate, the predominant form of sulfur in proteoglycans, may serve as an ion-exchange mechanism for the passage of Ca through the matrix to sites where Ca and phosphate are precipitated.

Avian tibial dyschondroplasia. I. Ultrastructure.

Tibial dyschondroplasia is an abnormality of the growth cartilage that occurs in chickens and other rapidly growing animals. The disease is characterized by a mass of avascular opaque cartilage, which is continuous with the growth plate of the proximal tibia and extends into the metaphysis. In this study electron micrographs revealed that chondrocytes in the hypertrophic zone of the growth plate were normal in appearance with the exception that the cells did not undergo complete hypertrophy. In the proximal region of the lesion, cells began to undergo necrotic changes suggestive of an energy depletion. These changes included dilatation and vesiculation of the endoplasmic reticulum, enlargement of the paranuclear space, mitochondrial swelling with dilatation of the intracristal spaces and the appearance of electron-dense, flocculent material in the mitochondrial matrix, chromatin margination, and dilatation of the Golgi saccules. Chondrocytes also occurred with rarefied cytoplasm and atrophic Golgi saccules. A few cartilage cells in the proximal region of smaller lesions contained crescentic caps of condensed chromatin in the nuclei, which is indicative of apoptosis. These cells also exhibited dilated endoplasmic reticulum and lamellar bodies; and sometimes, in the proximal region of the lesion, they appeared to be condensed and convoluted. This process continued in the mid and distal regions. The condensed necrotic cells appeared as amorphous osmiophilic masses with karyorrhexic and pyknotic nuclei. Matrix vesicles were observed at all levels of the lesion, but calcified only at the distal edge of the lesion, where mineralization of both matrix and cells occurred. The resulting shell of mineral may act as a diffusion barrier.

Avian tibial dyschondroplasia. III. Electron probe analysis.

Tibial dyschondroplastic (TD) lesions and their associated growth plates, obtained from chickens, were prepared by freeze-drying and embedding in an anhydrous epoxy resin. Quantitative electron probe analysis was performed on dry, unstained sections. Levels of Na, Mg, P, S, Cl, K, and Ca were determined in cytoplasm (endoplasmic reticulum), mitochondria, and extracellular matrix of the proliferative, prehypertrophic, and early hypertrophic zones of the growth plate and in the proximal, mid, and distal regions of the lesion. A zone of calcification in the growth plate was absent. The concentration of elements in all regions of the TD growth plate was the same as found in an earlier study for normal growth plate. The cytoplasm of proximal lesion chondrocytes was similar to that of early hypertrophic chondrocytes. However, in the remainder of the lesion there was a progressive increase in cellular Na, S, Cl, and Ca and a progressive loss of P. In matrix, there was less S and K than expected in all regions of growth plate and lesion, except in the proliferating zone. Also, in matrix of the distal lesion there was less Na and Cl. The levels of Na, S, Cl, and K in matrix may have been lowered by their adsorption into the condensed masses of dead cells. Mitochondria acquire only half as much Ca and P as normal and release it earlier than usual (ie, early prehypertrophic cells, rather than chondrocytes of the lower hypertrophic zone). There were no granules in mitochondria of the cells at all levels of the lesion, even though anhydrous methods were used. The first sign of the disease appears in the matrix of the growth plate, where it seems that S and K are in abnormally low amounts. Although there are sufficient levels of Ca and P present, the matrix does not calcify. The cartilage remains avascular, and the cells appear to be dying. The event that triggers the chondrocytes of the growth plate to form an abnormal uncalcified matrix is not known.