Light Microscopic and UI t rast ructu ral Observations of the Calcifying Zone of the Mandibular Condyle in the Rat

AKE LARSSON Department of oral histopathology, Faculty of Odontology, Universi ty of Lund, Malmo, S w e d e n

ABSTRACT The hypertrophic stage of development of the rat mandibular condyle was investigated in 16 and 26-day-old rats by electron microscopy. Inter- est was focused on the zone of mineralization and erosion. It was observed that capillaries invaded the lower level of the hypertrophic zone, without any previous chondroclastic resorption of calcified partitions. The partitions surrounding the hypertrophic chondrocytes were not mineralized around their entire circumfer- ence at the level of capillary penetration. The capillaries were accompanied by perivascular cells but these showed no similarities to chondroclasts. Multi- nucleated chondro- or osteoclasts were however present at a lower level of the subchondral area. It is suggested that there are no inherent differences with re- spect to the pattern of mineralization and erosion between the epiphyseal growth plate and the developing mandibular condyle.

While the light microscopic features of the developing mandibular condyle are well defined (Durkin et al., '73 for a compre- hensive review), the cellular activities at the ultrastructural level have not appar- ently been subjected to a critical analysis. Thus, the concepts of a pericellular min- eralization of the whole circumference of the chondrocytic lacunae, of a chondro- clastic rather than of a capillary primary erosion of the cartilage and of a lack of primary spongiosa in the developing con- dyle still lack electron microscopic confir- mation. These concepts have also formed the basis for the concept that there are in- herent differences between the activities of the cartilage cells of the condyle and those of the epiphyseal growth plate.

In this study, the zone of mineralization and erosion of the developing condyle in the rat have been investigated by electron microscopy. The present observations indi- cate that serious objections must be raised against the validity of some of the con- cepts maintained by Durkin et al. ('73). Thus, the walls of chondrocytic lacunae did not show a complete mineralization at the level of initial erosion and a capillary rather than a chondroclastic activity was evident in the zone of primary erosion.

ANAT. REC., 185: 171-186.

MATERIALS AND METHODS

Three animals from each of two litters of sixteen and 26-day-old Sprague-Dawley rats were included in this study. The rats were anesthetized with sodium pentobar- bital (Nembutalo) and perfused through their left cardiac ventricle with ( a ) 0.1 % Prokain-HC1 (Forssmann et al., '67) in 0.05 M cacodylate buffer (pH 7.2) contain- ing 0.2 M sucrose for a short time, imme- diately followed by (b ) 2% glutaraldehyde (Poly-Sciences Inc., Warrington, Pennsyl- vania) in the same buffer containing 2% Dextran (mwt 40.000, Pharmacia Fine Chemicals, Uppsala, Sweden) for several minutes. The mandibular condyles were exposed by first removing the overlying soft tissues, The condyles were then rapidly extirpated and fixed for another 3-4 hours at 4°C in 4% glutaraldehyde in 0.1 M ca- codylate buffer (pH 7.2). Fixation was ended by several rinsings in the cacodylate- sucrose buffer at 4°C. Thin slices of the condyles were cut free-hand in the medio- lateral plane with a sharp razor blade under a stereomicroscope. The slices were postfixed in 1% OsOI in the cacodylate- sucrose buffer (pH 7.2) for one hour at

Received Oct. 17, '75. Accepted Dec. 4, '75.

171

172 AKE LARSSON

4”C, rapidly dehydrated in ethanol and embedded in Epon.

Thick sections ( 1 p ) were stained with toluidine blue for light microscopic orien- tation. Ultrathin sections were cut in an LKB Ultrotome, using glass knives, floated on 20% ethanol and rapidly transferred to naked copper grids. Uncontrasted or uranyl-lead contrasted sections were exam- ined in a Philips EM 300 electron micro- scope.

OBSERVATIONS

The condyles of the 26-day-old rats were approximately 2.5 times broader in the mediolateral plane than those of the 16- day-old rats. The cartilage cells of the older age group were also larger and gen- erally more numerous and the separate zones of the condyle were broader than in the younger condyles. However, the gen- eral histologic architecture of the condyles was similar in the two age groups. Part of the developing condyle of a 26-day-old rat is illustrated in figure 1. A brief de- scription of some of the features seen in the 1 thick sections will be given, since this will facilitate the recognition of those areas that will be examined in detail at the ultras tructur a1 level.

Light microscopic observations The area indicated by a rectangle in

figure 1 was interpreted as being within the region of initial mineralization. A transition was seen here from a slightly metachromatic stain reaction in the walls (= partitions, trabeculae or septa) sur- rounding the hypertrophic chondrocytes (“level I”, fig. 2 ) to a deep blue staining corresponding to deposited mineral in the more deeply situated trabeculae (“level 2”). Patches of mineral were also present in the partitions in the lower parts of level 1. In the intermediate area between level 1 and 2, there were several empty lacunae, where remnants of hypertrophic degenerat- ing chondrocytes were no longer discern- ible. Thin-walled capillaries were clearly seen in this area (fig. 2 ) . Some of these capillaries apparently invaded the empty lacunae via openings between neighbour- ing partitions (fig. 2) . Roundish or more elongated cells were seen accompanying the capillaries into the area. These cells

always appeared at a level slightly below the penetrating ends of the capillaries

In the lower part of level 1, figure 2, the chondrocytic lacunae usually contained remnants of degenerating chondrocytes or seemed empty. However, other structures were occasionally found within the la- cunae, as being illustrated in figures 3a,b. Sometimes these structures gave the im- pression of sprouts penetrating the parti- tion closest to the terminal end of a capil- lary. These structures were also highly prominent due to a high uptake of tolui- dine blue and they sometimes contained a roundish element, similar to a nucleus (fig. 3b). Serial sectioning showed that these structures did not merely represent staining artefacts (figs. 3a,b).

The lacunae in the subchondral area, below level 2 in figure 2, were occupied by cellular elements, which is readily seen in figure 4. These cells covered the sur- faces of the calcified trabeculae, and a gradual loss of the transverse walls was also a prominent finding at this level. The cells were rather small and of uniform size. Larger and multinucleated cells appeared further down in the subchondral area (fig. 4 ) . These cells were also intimately associated with the surfaces of the calci- fied septa. The distance between the pene- trating ends of the capillaries at level 1, figure 2, and the level where the multinu- cleated cells were identified, was in the order of 200-300 pn in the central por- tions of the condyle (fig. 4).

Electron microscopic observations The ultrastructural findings to be de-

scribed below will mainly concern features seen at the level of mineralization of the condyle, corresponding to the rectangle in figure 1. At the level corresponding to “1” in figure 2, the hypertrophic chondrocytes exhibited an advanced but somewhat vary- ing degree of degeneration. In some la- cunae, fairly well preserved nuclear struc- tures were seen (fig. 5), whereas in others only a few membrane remnants were visi- ble. Still other lacunae contained no identi- able cellular elements at all.

The walls of the hypertrophic chondro- cytes (fig. 5 ) were composed of a network of fine fibrils, with interspersed smaller or

(fig. 2 ) .

CALCIFICATION AND RESORPTION OF THE MANDIBULAR CONDYLE 173

larger electron-dense dots and patches. Higher magnification of the smaller dots revealed the presence of globular mem- brane-limited vesicles with a homogeneous, moderately electron dense matrix (fig. 6). Clusters of crystallites were seen in the immediate vicinity of these vesicles and needle-like crystallites were also present within the vesicles themselves (fig. 7). The latter crystallites were occasionally found to permeate the limiting membrane of the vesicles (fig. 7). The surrounding matrix consisted of a meshwork of thin fibrils and many small electron-dense granules (fig. 6). Similar granules were seen in large num- bers in the matrix at the periphery of the chondrocytic lacunae (fig. 8).

At higher magnification, the larger patches of electron dense material illus- trated in figure 5 were found to represent islands of crystallites, seemingly fusing with adjacent crystallite islands (fig. 8). The vesicles described above were less nu- merous here, with only a few appearing at the periphery of the crystal clusters.

At a somewhat lower level of the calci- fying zone, capillaries were observed in close proximity to the chondrocytic parti- tions (fig. 9). There were signs of discon- tinuity in the endothelial lining of these capillaries. At the same level, the earlier observed clusters of crystallites in the chondrocytic walls had grown to broad bands of mineralized trabeculae. However, a lack of mineralization in parts of the walls was evident (figs. 9, 10, 12) which was also confirmed by the fact that no crystallites were seen here in uncontrasted sections.

With respect to the surface morphology, there was a clear difference between the chondrocytic and the capillary side of the mineralized trabeculae (fig. 10). On the chondrocytic side the surface was covered by a cartilaginous matrix rich in fibrils and granules. A gradually increased density was seen when going from this matrix to the area occupied by the fusing clusters of crystallites. On the capillary side, on the other hand, no cartilage matrix was found covering the surfaces. A sharp dermarca- tion line was present between the calcified trabeculae and the surrounding medium. This line of demarcation exhibited a promi- nent electron density (figs. 9, 10, see also

figs. 13-16). Rather thick fibrils were fre- quently observed at these surfaces (fig. 10). Similar fibrils were also seen on the capil- lary side opposite non-mineralized sections of the partitions (figs. lo , 11). At higher magnification, many of these fibrils were found to be cross-striated (fig. 11).

In association with lacunae, where an extensive break-down of the chondrocytes was evident and where penetrating capil- laries had not yet entered, the calcified trabeculae frequently exhibited a sharp and electron dense demarcation line. There was also an absence of cartilaginous matrix at the surfaces facing the lacunae (fig. 12).

The capillaries invading the zone of partly calcified walls (cf., figs. 2 , 3 and 4 ) were accompanied by a number of cells. Mononuclear blood cells were identified within the vascular lumina (figs. 2 , 9). Nucleated cells were also seen in the extra-vascular spaces but none of these cells was convincingly multinucleated. The cells sometimes approached the level of, but often appeared about one chondrocytic lacuna behind, the capillary sprouts (figs. 2, 3a,b, 9, 13). Here they clearly tended to encroach upon the surfaces of the cal- cified trabeculae. The peculier, cell-like ele- ments observed light microscopically in lacunae in front of the penetrating capil- laries (cf., figs. 3a,b), will be the subject of a later, separate study.

The area of contact between the cells accompanying the capillaries, and the tra- becular surfaces, exhibited some interest- ing morphologic features (figs. 13-16). In some places (figs. 13, 14), broad cell pro- jections rather than the main body of the cells were in intimate contact with the electron dense demarcation line of the cal- cified trabeculae. At other sites, there was a wider space between the cellular projec- tions and the trabeculae (fig. 15) and fine fibrils were observed (figs. 9, IS) , often exhibiting a cross-striation similar to that illustrated in figure 11.

DISCUSSION

The findings presented in this paper support previous suggestions (Durkin et al., '73) that mineralization of the condylar cartilage commences in the walls surround- ing the chondrocytic lacunae. Furthermore, the subcellular pattern of initial formation

174 AKE LARSSON

of crystallites shows great similarities to that seen in the epiphyseal growth plate. The vesicles observed in the cartilaginous matrix of the condyle exhibit the same fea- tures as those, having been proposed to mediate the initial crystal formation in the growth plate (Anderson, '69; Bonucci, '70). The matrix surrounding the chondrocytic lacunae in the hypertrophic zone of the condyle also shows all the other features associated with the corresponding matrix of the growth plate, i.e., thin fibrils and small granules, the latter looking exactly like the proteoglycan granules described by Thyberg et al. ('73).

However, the present observations also raise several doubts about the reliability of previous light microscopic results concern- ing the general pattern of mineralization and resorption of the condylar cartilage. Based on the finding that the degenerating chondrocytes eventually become completely entrapped within mineralized partitions, Durkin et al. ('73) claimed that chondro- clasts rather than capillaries were the pri- mary agents in the erosion process. As seen in the present results, this suggestion must however be seriously questioned. It was observed that at the level of initial erosion of chondrocytic partitions, the walls were not completely mineralized and capillaries rather than multinucleated cells were in- vading the empty lacunae. Nucleated cells were accompanying the capillaries but there was little or no evidence to support the idea that any of these cells were ever clearing the path for the penetrating capillaries.

Furthermore, the cells were lacking those cytologic features normally associ- ated with osteoclasts as being defined by Lucht ('73). The cells showed many simi- larities to the perivascular and monocytic cells described by Schenk et al. ('67, '68) in the erosion zone of the growth plate. These authors suggested, but were unable to show, a direct involvement by these cells in the resorption of the mineralized and unmineralized cartilage. There was no in- dication in the present study that any of the cells accompanying the capillaries originated from hypertrophic chondrocytes. Thus, the results do not confirm the sug- gestion made by Silberman and Frommer

('72), that hypertrophic cells fuse to form multinucle ar chondroclas ts.

The peculiar cell-like structures seen in front of the capillaries (cf., figs. 3a,b) may be an exception to the rule that cells never penetrate ahead of the invading capillaries in the condyle. The appearance and posi- tion of these structures are intriguing and certainly deserve further study.

The data obtained in the present study, with endothelial cells being in a very close connection with the partitions, are thus highly suggestive of a direct involvement by the invading capillaries in the erosion of the condylar cartilage. Pertinent to the recognition of such a primary role by the capillaries is also the finding of unmin- eralized gaps in the chondrocytic walls in the erosion zone. This is a situation simi- lar to the one in the epiphyseal growth plate, where the uncalcified transverse chondrocytic partitions in the erosion zone obviously constitute the avenue of choice for the penetrating capillaries (Schenk et al., '68). However, Durkin et al. ('73) have proposed that the hypertrophic chon- drocytes of the condyle are completely sur- rounded by mineralized matrix at the stage of initial erosion. These authors concluded that the only way by which such trabecu- lae could be resorbed was by means of actively functioning chondroclasts. Multi- nucleated chondroclasts were also illus- trated at this level by Durkin et al. ('73). These authors also demonstrated the pene- tration of capillaries into the area. How- ever, the erosion mediated by the capillar- ies was proposed to be passively adaptive, with depth of capillary invasion being pri- marily determined by the level of previously eroded chondrocytic partitions.

It should be pointed out that resorption of the mineralized longitudinal septa of the growth plate has been claimed to require the presence of active chondro- or osteo- clasts (Schenk et al., '67). Such multinu- cleated cells were also identified in the present study but they were at a level far beyond the zone of capillary invasion. There is no reason to argue against the assumption that resorption of mineralized septa require the presence of specialized cells. But the present results do not give support to the assumption that there are

CALCIFICATION AND RESORPTION OF THE MANDIBULAR CONDYLE 175

major differences between the condyle and the growth plate with respect to the pattern of pericellular wall mineralization and ini- ial erosion.

Based on studies of animals labeled with 3H-proline, Durkin et al. (’73) concluded that the spicules of calcified cartilage in the zone of erosion are not used as the scaffolding for active bone formation. This assumption was based on the finding that the labeled areas appeared well below the calcified cartilage spicules in the erosion zone. This would imply that no synthesis of bone matrix takes place in the unlabeled zone. However, primary erosion of the cal- cified trabeculae is mediated by chondro- clasts in this zone according to Durkin et al. (’73), and the findings further support the idea that there is no primary spongiosa in the developing condyle (Irving and Dur- kin, ’ 6 5 ) .

In the present study, the surfaces of the partitions at the level of mineralization of the condylar cartilage showed some inter- esting features which raise further doubts about the validity of the “lack-of-primary- spongiosa”-concept held by Durkin et al. (’73). The electron-dense demarcation lines observed at the surfaces of the miner- alized trabeculae are equivalent to the “lamina limitans,” as defined by Scherft (’72). The presence of this lamina reflects a temporary arrest in mineralization rather than a progressive resorption (Scherft, ’72). Thus the presence of fibrils at the surface of the laminae limitantes cannot readily be accounted for by an active re- sorption exposing cartilage fibrils. The nature of the fibrils is unknown but the aperiodic fibrils may be related to deposi- tion of fibrin, suggested to take place in the growth plate (Schenk et al., ’68). The fact that some fibrils exhibited a periodic banding raises a suspicion that there may even be an osteoblastic activity at this level. The true nature of the activity of the perivascular cells is, however, unknown and will be the subject of further studies.

It is also of interest to note that a lamina limitans was evident not only at the capil- lary side but also at the chondrocytic side of the mineralized septa. In addition, the development of a lamina was often seen at the lacunar surfaces before the pene-

trating capillary sprouts had entered the lacuna. This would seem to imply that the endothelial cells of the capillaries are not directly involved in the removal of the cartilaginous matrix from the lacunar sur- faces. Schenk et al. (’67) have suggested that resorption of the corresponding ma- trix in the growth plate may be accom- plished by hydrolytic enzymes released upon breakdown of the chondrocytes. Based on the present results this may also be the case in the developing condyle.

In his definition of secondary or embry- onic cartilage, Durkin (’62) stated that such cartilages present with a pericellular type of calcification, circumscribing the hy- pertrophic chondrocytes. It was also stated that erosion of such calcified cartilage takes place by chondroclastic resorption and that primary spongiosa is practically nonexistent. It should be understood that the developing condyle of the mandible fulfills the criteria of such a cartilage, while another pattern of cellular activity is seen in the primary cartilages, e.g. the epiphyseal growth plate of long bones. A prominent feature of the growth plate is the columnar arrangement of the hy- pertrophying chondrocytes, with miner- alization of the vertical septa. The chon- drocytes in the condyle, on the other hand, are more haphazardly organized, with no clear-cut vertical mineralization pattern. This difference in cell organization may re- flect inherent differences in cellular activ- ity, related to different growth patterns of the two cartilages. While the epiphyseal growth plate exhibits the features of an active growth centre, the activity of the cells in the condylar cartilage would merely be a compensatory response providing enough growth for the condyle to remain in contact with the articular fossa, while the mandible is being carried down as the result of forces exerted by the surrounding soft tissues (Moss, ’60). The results ob- tained in the present study are not in con- a c t with such a concept. However, as evi- denced by the present observations, the ultrastructural pattern of mineralization and erosion of the condylar cartilage are fundamentally similar to those of the growth plate. These patterns can no longer form the basis for arguing that there are

176 AKE LARSSON

inherent differences between the growth plate and the developing condyle. The pos- sibility of other cellular factors should therefore be considered in order to explain the different growth patterns of the two cartilages.

ACKNOWLEDGMENTS

I am most grateful to Miss Catrine Jo- hansson and Mrs. Birgitta Sluga for valu- able technical assistance. I am also grate- ful to the Institute of Pathology, Malmo General Hospital, (Head: Prof. F. Linell) for the use of electron microscopic facili- ties for this study. The investigation was supported by grants from The Swedish Medical Research Council (Project No. 12X-2298 ) .

LITERATURE CITED Anderson, H. C. 1969 Vesicles associated with

calcification in the matrix of epyphyseal carti- lage. J. Cell Biol., 41 : 59-72.

Bonucci, E. 1970 Fine structure and histochem- istry of “calcifying globules” in epiphyseal cartilage. Z. Zellforsch. Mikrosk. Anat., 103:

Durkin, J. F. 1972 Secondary cartilage: A m i s - nomer? Am. J. Orthod., 62: 15-41.

Durkin, J. F., J. D. Heeley and J. T. Irving 1973 The cartilage of the mandibular condyle. In: Oral Sciences Reviews. Vol. 2. A. H. Melcher and G. A. Zarb, eds., pp. 29-99.

192-217.

Forssmann, W. G., G. Siegrist, L. Orci, L. Giradier, R. Pictet and C. Rouiller 1967 Fixation par perfusion pour la microscopie electronique. J. Microscopie, 6: 279-304.

Irving, J. T., and J. F. Durkin 1965 A com- parison of the changes i n the mandibular con- dyle with those in the upper tibial epiphysis during the onset and healing of scurvy. Arch. Oral Biol., 10: 179-185.

Lucht, U. 1972 Osteoclasts and their relation- ship to bone as studied by electron microscopy. Z. Zellforsch., 135: 211-228.

Moss, M. L. 1960 Embryology, growth and mal- formations of the temporomandibular joint. In: Disorders of the Temporomandibular Joint. L. Schwartz, ed. W. B. Saunders Co., Philadel- phia and London, pp. 89-103.

Schenk, R. K., D. Spiro and J. Wiener 1967 Cartilage resorption in the tibial epiphyseal plate of growing rats. J. Cell Biol., 34: 275-291.

Schenk, R. K., J. Wiener and D. Spiro 1968 Fine structural aspects of vascular invasion of the tibial epiphyseal plate of growing rats. Acta anat., 69: 1-17.

Scherft, J. P. 1972 The lamina limitans of the organic matrix of calcified cartilage and bone. J. Ultrastruct. Res., 38: 318-331.

Silbermann, M., and J. Frommer 1972 The na- ture of endochondral ossification in the man- dibular condyle of the mouse. Anat. Res., 172:

1973 Electron microscopic demonstration of proteo- glycans in guinea pig epiphyseal cartilage. J. Ultrastruct. Res., 45: 407-427.

659-667. Thyberg, J., S. Lohmander and U. Friberg

PLATES

Figures 1 4 are light microscopic pictures; figures 5-16 are electron micrographs.

PLATE 1

EXPLANATION O F FIGURES

1 Coronal section of mandibular condyle of 26-day-old rat, showing the immature stage of development, where the following indistinctly separated zones are identified: resting or articular zone (AZ) , transi- tional or proliferating zone (PZ), hypertrophic zone (HZ) , zone of erosion (EZ) and zone of subchondral bone formation (BZ). Rec- tangle indicates the border region between HZ and EZ, where min- eralization of chondrocytic partitions is first recognized. x 115.

From a level of the condyle corresponding to rectangle in figure 1. “Level 1” represents the lower part of HZ, with degenerating hyper- trophic chondrocytes (HC) and metachromatically stained pericellu- lar trabeculae. At “level 2,” the trabeculae are mineralized and nucleated cells (NC) cover their surfaces. Patches of mineral are also seen in trabeculae at the lower part of level 1 (arrows). At the border between the two levels, several lacunae look empty (EL) and capillaries are invading the area (C). x 600.

From two different sectioning depths of the same condyle, at the level of capillary penetration, cf. figure 2. Capillaries (C) accom- panied by nucleated cells (NC) have entered the empty chondro- cytic lacunae. Occasionally, cell-like structures (double arrows) ap- pear in the lacunae in front of the capillary sprouts. The structures contain deeply stained and sometimes also nucleus-like (arrow) elements. x 500.

From central section of condyle, showing the hypertrophic (HZ) and erosion (EZ) zone as well as deeper subchondral area. Immedi- ately below EZ, the lacunae are occupied by nucleated cells ( a - rows), accompanying the capillaries (C). Loss of transverse trabecu- lae is evident at this level. Large, multinucleated cells are observed further down in the subchondral area (double arrow). X 270.

2

3a,b

4

178

CALCIFICATION A N D RESORPTION OF THE MANDIBULAR CONDYLE Ake Larsson

PLATE 1

179

PLATE 2

EXPLANATION OF FIGURES

5 From lower level of the hypertrophic zone, cf . figure 2. Degenerating hypertrophic chondrocytes are seen in the lacunae (L) where rem- nants of nuclei may still be observed ( N ) . The surrounding trabecu- lae are composed of a fine fibrillar network, with interspersed smaller (arrow) and larger (double arrow) deposits of electron-dense mate- rial. x 12,300.

6 Higher magnification of the smaller electron-dense deposits seen in figure 5. Membrane-limited vesicles (large arrows) and clusters of crystallites (double arrow) are present in a matrix, rich in fibrils and minute electron-dense granules (small arrows). X 50,500.

High magnification micrograph of matrix vesicles, cf. figure 6. Crys- tallites are seen within the vesicles and the limiting, tri-laminated membrane is being penetrated by crystallites in both vesicles depicted. x 104,000.

8 Section of cartilaginous partitions (P) with adjacent chondrocytic lacunae (L) . The matrix of the partitions is rich in fibrils and gran- ules, the latter being particularly numerous at the borders of the lacunae. Clusters of crystallites are gradually fusing into larger aggre- gates (arrow). X 12,700.

7

180

CALCIFICATION AND RESORPTION OF THE MANDIBULAR CONDYLE Ake Larsson

PLATE 2

181

PLATE 3

EXPLANATION OF FIGURES

9 From area of capillary invasion, cf. figure 2. The ends of two capil- laries, ( C ) one of them containing two mononuclear cells, are i n close proximity to the partitions surrounding the chondrocytic lacunae (L) . Several nucleated cells (NC) appear at about the same level as the capillary sprouts. Non-mineralized gaps are seen in between the mineralized sections ( M j of the walls (arrows 1 and 2). x 1,800.

From area at arrow 1. figure 9, showing the cartilaginous wall sepa- rating a chondrocytic lacuna (L j from compartment with penetrating capillary ( C ) . E, endothelial cell of capillary. A non-mineralized gap is seen between sections of the mineralized wall ( M ) . The chon- drocytic side of this gap consists of cartilaginous matrix ( C M ) of normal appearance. O n the capillary side of the wall, many fibrils are present (F ) and the wall also exhibits an electron-dense demar- cation line (arrows). X 11,900,

From area at arrow 2, figure 9. The wall bordering the chondrocytic lacuna ( L ) consists of non-mineralized cartilaginous matrix (CM). Many of the fibrils seen o n the outer side of the wall show a cross- striation (arrows). X 50,500.

From a lacuna containing only very few remnants of a degenerated chondrocyte. The hypertrophic zone is in the upper direction of the picture. At the lower left corner, underneath the lacuna, the pene- trating end of a capillary is seen. Heavily mineralized sections of the wall ( M ) are interrupted by non-mineralized gaps. The chondrocytic surface of the mineralized trabeculae are partly covered by cartilagi- nous matrix (CM), but the capillary side as well as some areas of the chondrocytic side lack such a layer (arrows). X 3,100.

10

11

12

182

CALCIFICATION A N D RESORPTION OF THE MANDIBULAR CONDYLE Ake Larsson

PLATE 3

183

PLATE 4

EXPLANATION O F FIGURES

13 From the level of capillary invasion. The end of a penetrating capil- lary is seen (C) beneath a chondrocytic lacuna (L). Nucleated cells (NC) encroach upon the surface of the mineralized walls ( M ) , closely below the level of capillary ends. Note the electron-dense demarcation line of trabeculae on the capillary side (arrows), being less prominent or absent (double arrow) on the chondrocytic side. X 4,100.

14 From nucleated cell-trabecula contact area, depicted in figure 13. Cellular projections (CP) are in close contact with the electron-dense demarcation line of the mineralized wall (M) . The cytoplasm of the cell is rich in mitochondria, rough surfaced endoplasmic reticulum and various small vesicles. x 12,000.

Cellular projection (CP) between endothelial lining ( E ) of penetrat- ing capillary and surface of mineralized wall ( M ) . Fine fibrils are present in the space between CP and the electron-dense demarcation line of M. L, chondrocytic lacuna. X 7,300.

From area below arrow 2 , figure 9, showing chondrocytic lacuna (L) , cartilaginous matrix (CM). mineralized wall ( M ) and nucleated cell (NC). The space between the cell and the electron dense demarcation line of the wall contains numerous thin fibril. X 12,700.

15

16

184

CALCIFICATION AND RESORPTION OF THE MANDIBULAR CONDYLE Ake Larsson

PLATE 4

185