Hydrolytic enzyme activity during endochondral ossification of secondary cartilage

Abstract

Reactions for hydrolytic enzymes such as aminopeptidase, p-glucuronidase, N-acetyl-p-glucosaminidase, arylsulfatase and non-specific esterases indicated various degrees of activity within the chondrocytes of the young mandibular condyle. Most of the reaction products of the above enzymes appeared as discrete granules, which might indicate a lysosomal origin. The most intense activity of these enzymes was observed within chondroblasts and premineralizing hypertrophic chondrocytes. However, chondrocytes within the mineralizing zone also appeared synthetically active. The latter cells, as well as those at the adjacent ossification front, revealed some activity by a majority of the enzymes tested.This could indicate that some metabolic functions, although reduced in degree, continue in the cells of these mineralizing zones of endochondral ossification.Matrical reactions, with the exception of arylsulfatase and aminopeptidase, were essentially negative. The positive reactions for the latter enzymes might be an indication of their involvement in the process of extracellular mineralization.While hydrolytic enzymes have been studied in the cartilage of the epiphyseal plate by Kuhlman ('65), Granda and Posner ('71) and Ehrlich et al. ('73), few investigations have been conducted on secondary cartilage such as that found in the mandibular condyle (Greenspan and Blackwood, '66; Silbermann and Frommer, '73a). The possibility that hydrolytic enzymes, such as nonspecific esterases, glycosidases, aminopeptidase and arylsulfatase, are associated with the synthesis and turnover of protein-polysaccharide complexes in cartilaginous tissue increases the importance of investigating these enzymes in normal cartilage during endochondral ossification. Such a study might provide further information in regard to the participation of these enzymes in matrix formation and mineralization (Silbermann and Frommer, '73b). Therefore, studies were undertaken to determine whether these enzymes are active in the condylar cartilage and, if so, to relate them to morphological and physiological alterations that are known to occur during chondrogenesis and mineralization (Brighton et al., '73). MATERIALS AND METHODSThirty male and female CD-1 mice, six AM. J. ANAT., 140: 369-382. days of age, were anesthetized with ether and decapitated. Blocks 3-4 mm thick were prepared. After fixation in formol-calcium (Baker, '44) for 24 hours at 4"C, tissues were transferred directly into hypertonic gum-sucrose (0.88 M ) medium and were left for 24-48 hours at 4°C. Tissues were then quenched for two minutes in 2-methylbutane which was cooled to -80°C by liquid nitrogen. The blocks were sectioned in the coronal plane at 10 in a microtome-cryostate at -20°C.The histochemicd methods utilized and components of the various incubation media are listed in table 1. As controls, sections were incubated in media from which the substrate had been omitted, OK were heated to 60°C for 20 minutes prior to incubation in the reaction mixtures. The following inhibitors of a-...

Development of cartilage and bone tissues of the anterior part of the mandible in cichlid fish: A light and TEM study

“…The mature flattened chondrocytes resemble the chon-drocytes described a t the light microscopical level in the flattened cell zone in epiphyseal cartilages of fish (Haines, 1934(Haines, , 1938(Haines, , 1942Meunier, 1979) and of warm-blooded vertebrates (e.g., Howlett, 1979). The chondrocytes in Meckel's cartilage of H. bimaculatus however, seem to lack one feature which is often held to be characteristic for such cells, namely deposits of glycogen (e.g., Godman and Porter, 1960;Silberberg et al, 1964Silberberg et al, , 1966Silberberg et al, , 1976Silva and Hart, 1967;Holtrop, 1972;Silbermann and Frommer, 1974;Silbermann and Lewinson, 1978). The presence of glycogen in chondrocytes is indicative of the state of maturation of the cells and related to the synthesis of matrix polysaccharides (Knese, 1979).…”

Bone and cartilage resorption in relation to tooth development in the anterior part of the mandible in cichlid fish: A light and TEM study

“…Several cell types have been proposed to be involved in resorption of uncalcified cartilage, such as multinucleated chondroclasts (Muhlhauser, 1983;Aceitero et al, 1988) and enclosed chondrocytes (Schenk et al, 1967;Melcher, 1972;Silbermann and Frommer, 1974;Yoshioka and Yagi, 1988). Except for the possible limited participation of (mononuclear) osteoclasts, we have no evidence of the involvement of such cells during the resorption process of Meckel's cartilage in Hemichromis bimaculatus.…”

Enzyme histochemical studies of induced heterotopic cartilage and bone formation in guinea pigs with special reference to acid phosphatase

“…Studies ofthe function and importance of acid extracellular matrix precedes the mioeralization phosphatase (AcP) have shown that this enzyme of hard tissues has been suggested (14,22). can be found in matrix-producing areas of bone LysosomaJ enzymes have been proposed as and cartilage (13,405,41)-Both enzyme his-mediators of this reduction of glycosaminotochemical (!7,18) and hiochemical studies glycans (19,29,31,33). Electron microscopic (5,38) have produced evidence of two groups of investigations have shown lysosome-like ex-AcP in normal hard tissue forming areas^ one tracellular bodies, matrix vesicles type I (25,37).…”