The purposes of these experiments were to study the biosynthetic and postbiosynthetic relationships between proteoglycans in noncalcified growth cartilage and calcified cartilage in metaphysis from the costochondral junctions of immature rabbits. Based on in vivo experiments in which 35 S-sodium sulfate was injected into rabbits, it is shown that proteoglycans from the hypertrophic region becomes part of the calcified cartilage matrix which is to be incorporated into the metaphysis. The proteoglycan aggregates in the growth apparatus undergo partial disaggregation and degradation. There is approximately a 25% decrease in aggregation from regions of the rib distal to the metaphyseal-growth plate junction (69%) to the region proximal to it (50%). In contrast, in their final state in calcified cartilage, the proteoglycans are more completely disaggregated and the proteoglycans subunits are smaller, as adjudged from gel chromatography. Control experiments indicate that although some artifactual disaggregation is produced by the extraction process, it is not of the same magnitude as that seen in the actual isolation experiments nor are the subunits reduced in size.

A B S T R A C TTibiae and humeri were removed from suckling rats at intervals of time after intraperitoneal injection of C14-L-phenylalanine, C14-L-leucine, S35-sulfate, or Ca ~ C12. Autoradiograms of sections of the bones were prepared. Ca 45 was removed from sections treated with dilute ace tic acid; neither the concentration of S ~5 nor that of C 14 was thereby markedly decreased. The S .5 was removed from the demineralized sections on incubation in a solution of testicular hyaluronidase; the C 14 was not. These results are interpreted as indicating that most of the S 35 was present in the bones as chondroitin sulfate and that most of the C a4 in the bones was present as protein. In the epiphyses, the C ~4 was initially concentrated in the proliferaing and hypertrophic chondrocytes, as was the S aS. Secretion of S 35-and CX4-1abeled materials into the matrix followed. Thereafter, however, although the S3~-labeled material (chondroitin sulfate) persisted in the matrix, albeit at a diminished concentration, and was incorporated into metaphyseal bone, the C14-1abeled material (protein) was almost completely removed from the matrix. When rats were given repeated doses of 1 7-13-estradiol benzoate so as to inhibit resorption of their metaphyses, repeated doses of S35-sulfate were discerned as strata of S 35 in their metaphyses. This was not the case if the rats received repeated doses of C~*-L-phenylalanine or C14-L-leucine. On the basis of the results in these experiments it is suggested that although a portion of the chondroitin sulfate produced by the chondrocytes of the epiphyseal plate is retained and becomes part of the cores of metaphyseal spicules of bone, the protein of the proteinpolysaccharide is somehow removed before calcification of the cartilage ensues.Following the isolation of chondroitin sulfate-S ~5 (1, 2) from the cartilage of rats given S35-1abeled inorganic sulfate, autoradiographic studies showed

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On the basis of an examination of autoradiograms of knee-joints fixed so as to remove chondroitin sulfate or inorganic sulfate, or to minimize the loss of both, it is suggested that the cartilage is permeable to inorganic sulfate in vivo and in vitro. In vivo and in vitro, almost as rapidly as it enters the cartilage, inorganic sulfate is utilized by the cells in the synthesis of chondroitin sulfate. The net result is a continuing low concentration of inorganic sulfate in the cartilage.