- Email: [email protected]
Topographical and histological examination of osteophytes taken from arthrotic femoral heads
ANNALS OF ANATOMY
Topographical and histological examination of osteophytes taken from arthrotic femoral heads Horst Claassen 1 and Thomas Tschirner 2
11nstitut ftir Anatomie der Universitgt Rostock, Gertrudenstrasse 9, 18055 Rostock, Germany, 2Anatomisches Institut der Universit~t Kiel, Olshausenstrasse 40, 24098 Kiel, Germany
Summary. Until now it is not known whether osteophytes
Key words: Articular cartilage - Femoral head - Osteo-
of the femoral head develop because of pathological joint phytes - Topographical localization - Extracellular matrix alterations or arise from normal remodeling processes - Proteoglycans - Glycosaminoglyeans secondary to osteoarthrosis. Firstly, we analysed the topographical localization of osteophytes. We then compared the extracellular matrix components of macroscopically normal cartilage from the margin of osteophytes with os- I n t r o d u c t i o n teophytic cartilage from weight bearing and non-weight bearing zones by histochemical staining of low and Osteoarthritis is characterized by pathological changes of heavily sulfated glycosaminoglycans. For examination 65 the hyaline articular cartilage, the subchondral bone and femoral heads were taken during endoprosthetic hip surthe synovial tissue (Jeffrey 1975). In the zone of articular gery Osteophytes from different locations and macroscocartilage bearing the greatest load, degradation of hyaline pically normal cartilage from the margin of osteophytes cartilage and denudation of the subchondral bone can be were excised, decalcified and embedded in paraplast. A observed. The disappearance of articular chondrocytes is lateral or medial localization of osteophytes (47 cases) accompanied by a reduction in synthesis of collagens and was more common than a ventral or dorsal position (18 proteoglycans (Annefeld 1983; Fassbender 1983; Broom cases). Histochemical staining for low and heavily sul1984; Miesenb6ck et al. 1987; Seibel 1989; Tilhnann et al. fated glycosaminoglycans from normal cartilage at the 1991). rim of osteophytes was stronger in the unmineralized carRepair attempts were observed in parallel to these detilaginous zones compared to the mineralized cartilagigenerative changes. Mitotic cell divisions took place in nous zone. Weight bearing zones of osteophytic cartilage, the destroyed cartilage. These processes form so-called on the other hand, showed an even distribution of the clusters of chondrocytes; i.e. groups of cells that lie close two differently sulfated glycosaminoglycans. Surprisingly, together. These cells are not able to replace the destroyed non-weight bearing zones of osteophytic cartilage showed cartilage matrix and so degradation continues. While cara weaker staining for low and especially for heavily sultilage was destroyed in the weight bearing zones, producfated glycosaminoglycans in the superficial cartilage layer tive processes took place in the non-weight bearing than in the deep cartilage layer. Altogether, osteophytic marginal zones, which led to exostoses or osteophytes cartilage can be regarded as a reparative phenomenon for two reasons: Firstly, osteophytes arise very often at the (Pauwels 1973). Because osteophytes succumb to pressure weight bearing lateral and medial femoral head. Secondly, demands like normal articular cartilage, this process leads despite local differences in osteophytic cartilage, the same to an enlargement of the joint surface (Carstens 1982). types of glycosaminoglycans are synthesized as in normal Therefore, osteophytes diminish the load of the damaged joint surface (Williams et al. 1988). cartilage at the margin of osteophytes. Until now it is unclear whether osteophytes are formed because of pathological joint alterations or they are due to normal remodeling processes in the course of osteoarthritis. In the present study low and heavily sulCorrespondence to: H. Claassen fated glycosaminoglycans were stained histochemically in E-mail: [email protected]
II
Ann Anat (2003) 185:67-71 © Urban & FischerVerlag
http:llwww.urbanfischer.deljournalslannanat
0940-9602103118511-67$I 5.0010
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Fig. 1. Lateral osteophyte from the femoral head of a 68-year-old male. Broad layer of fibrocartilage (FC) with degenerative changes visible as fissure (arrow). FC = fibrocartilage, B = bone. Azan. (x 10). Fig. 2. Medial osteophyte from the femoral head of a 73-year-old female. Osteophyte with a broad basis. The superficial layer of fibrocartilage (FC) which has different thickness shows degenerative changes. FC = fibrocartilage, B = bone. Azan. (x 10). Fig. 3. Histochemical staining of low and heavily sulfated glycosaminoglycans in different zones of normal articular cartilage at the rim of osteophytes. All zones of unmineralized cartilage show stronger staining for low (a) and heavily (b) sulfated glycosaminoglycans than the mineralized cartilage zone (MZ). Especially strong staining is observed in the radial fiber zone (RFZ). In the mineralized cartilage zone (MZ) only territorial matrix is positive for low (a) and heavily (b) sulfated glycosaminoglycans. T F Z = tangential fiber zone, TZ = transition zone, R F Z = radial fiber zone, MZ = mineralized zone. Alcian blue with 0.3 M (a) and 0.8 M (b) MgC12. a, b (x 40). Fig. 4. Histochemical staining of low and heavily sulfated glycosaminoglycans in osteophytic cartilage of weight bearing and nonweight bearing zones, a: Medial osteophyte stained for low sulfated glycosaminoglycans in general view. A weight bearing zone (c) can be discriminated from a non-weight bearing zone (e). b: Medial osteophyte stained for heavily sulfated glycosaminoglycans in general view. A weight bearing zone (d) can be discriminated from a non-weight bearing zone (f). c, d: Weight bearing zones of osteophytic cartilage: Low (c) and heavily (d) sulfated glycosaminoglycans were detected in all zones of fibrocartilage (FC) in even distribution, e: Non-weight bearing zone of osteophytic cartilage: In the upper third (broken line) of fibrocartilage staining for low sulfated glycosaminoglycans was weaker than in the lower two thirds, f: Non-weight bearing zone of osteophytic cartilage: In the upper half (broken line) of fibrocartilage staining for heavy sulfated glycosaminoglycans was nearly negative while it was weak in the lower half. Alcian blue with 0.3 M (a, c, e) or 0.8 M (b, d, f) MgC12. a, b (xl0); c, d (x 40); e, f (x 30). 68
osteophytic and normal cartilage at the rim of osteophytes. We tried to find out whether the extracellular matrix of osteophytes shows similarities to adult hyaline articular cartilage by comparing the former with normal cartilage.
Material and methods 65 resected femoral heads from patients undergoing endoprosthetic surgery for severe coxarthritis served as material for the present study. The age of the patients ranged from 30 to 90 years. The topographical localization of the osteophytes at the peripheral cartilage-bone border was documented immediately after surgery. Osteophytes and macroscopically normal cartilage marginal to osteophytes were dissected afterwards using an oscillating saw at different sites. Osteophytes and normal cartilage from 28 femoral heads were fixed in Bouin's solution (15 parts picric acid, 5 parts 4% formaldehyde, 1 part acetic acid). After washing for 2 days, specimens were decalcified in 10% buffered EDTA at 4 °C. EDTA-solution was changed every third day, decalcification was controlled weekly by X-rays. Following decalcification, specimens were dehydrated in several portions of alcohol and embedded in paraffin. Then, 7 gm paraffin sections were used for toluidine blue and azan staining. Low and heavily sulfated glycosaminoglycans were demonstrated histochemically according to Scott and Dorling (1963) with alcian blue containing 0.3 M or 0.8 M magnesium chloride, respectively.
Results
Localization of osteophytes. During postoperative macroscopieal investigation of femoral heads an area of nude bone with different extension was found in the region of the fovea capitis femoris. This area of uncovered bone was uneven and showed recesses. Adjacent to this pathologically changed region there was a cartilage of very soft quality which thinned to the rims of the femoral head. Protruding rims (osteophytes) of different size rose at the peripheral cartilage bone border. The superficial cover of the osteophytes was of a soft quality. Osteophytes at the cartilage-bone border were arranged in the shape of a closed ring in 23 femoral heads, while osteophytes were found in various places in 42 femoral heads. To summarize, in 65 femoral heads osteophytes were found more often on the medial and lateral side than on the ventral and dorsal side of the femoral head. A ventral or ventro-medial localization of osteophytes was found in 26 femoral heads, while a lateral or dorso-lateral one occurred in 21 femoral heads. A ventral localization of osteophytes was observed in 10 specimens, while a dorsal localization was observed in 8. Histological examination of osteophytes. The superficial cover of osteophytes does not show a uniform structure.
Fibrocartilage was often found beneath the superficial layer of connective tissue containing collagen fibers. Moreover, the surface of some osteophytes contained only fibrocartilage. In another form of osteophytes, a superficial layer of connective tissue containing collagen fibers converted to fibrocartilage after a certain distance (Fig. 1, 2). Often fibrocartilage showed degenerative changes in form of fibrillations and fissures (Fig. 1). If fissures occurred, the subchondral bone was sometimes completely uncovered. The characteristic zones of articular cartilage, the tangential fiber zone, the transitional zone and the radial fiber zone were detected only in osteophytes with a thick layer of fibrocartilage. The cells of fibrocartilage kept their round shape from the deep to the superficial cartilaginous zone and did not flatten in the direction of the superficial cartilage layer. Moreover, chondrocytes were not localized in groups, but were positioned singularly. The collagen fibers were arranged vertically from the deep to the superficial layer of fibrocartilage. This observation was made in demasked collagen fibers (Fig. 2).
Histochemical staining of low and heavily sulfated glycosaminoglycans in osteophytes. As far as ,,normal" cartilage in the periphery of osteophytes is concerned, more low and heavily sulfated glycosaminoglycans were detected in the different zones of unmineralized cartilage than in the mineralized cartilage (Fig. 3 a, b). Low and heavily sulfated glycosaminoglycans were distributed evenly in the tangential fiber zone and transitional zone (Fig. 3 a, b). However, the extracellular matrix of the radial fiber zone was stained stronger for the two glycosaminoglycans of different sulfation in comparison to the superficial cartilage layers. In osteophytic cartilage, it is of interest that the distribution of low and heavily sulfated glycosaminoglycans differed from corresponding distribution in normal cartilage at the margin of osteophytes (Fig. 4 a ,b). The typical zones of articular cartilage were not found in osteophytic cartilage which mainly contained fibrocartilage. Weight bearing and non-weight bearing regions can be discriminated in osteophytic cartilage (Fig. 4 a, b). In the weight bearing zone the extracellular matrix of osteophytic cartilage contained low and heavily sulfated glycosaminoglycans in even distribution (Fig. 4c, d). In the nonweight bearing zone, the histochemical staining of low sulfated glycosaminoglycans was weaker in the upper third near to the joint surface than in the lower two thirds (Fig. 4 e). The histochemical detection of heavily sulfated glycosaminoglycans was weaker all in all in the nonweight bearing zone compared to the low sulfated glycosaminoglycans. In the upper half near the joint surface only a few heavily sulfated glycosaminoglycans were detected. In the lower half of osteophytic cartilage, the histochemical reaction with alcian blue was somewhat stronger than in the upper half (Fig. 4 f).
69
collagen specific antibodies and cDNA probes. The authors identified a fibrocartilaginous chondrocyte phenotype that expresses type II- and III-, but not typ Icollagen. In conclusion, osteophytic cartilage contains extracellular matrix components of normal articular cartilage like type II-collagen and low and heavily sulfated glycosaminoglycans. On the basis of these results it seems likely that osteophytic cartilage shows signs of a repair phenomen.
Discussion The present paper has two goals: (1) Evaluation of the localization of osteophytes of the femoral head, (2) low and heavily sulfated glycosaminoglycans of the extracellular matrix of osteophytes were analyzed histochemically. Osteophytes are bone-cartilage-like protrusions at articular joints (Pottenger et al. 1990). Beside narrowing of the joint space, sclerosis and subchondral cysts, the occurrence of osteophytes is a criterion in establishing the diagnosis of osteoarthrosis. Observation of the localization of osteophytes in 65 femoral heads showed that osteophytes can grow at any site (Jeffery 1975). In 47 femoral heads more pronounced osteophytic growth was observed in medial and lateral localizations than in ventral and dorsal areas. In 18 femoral heads, osteophyte growth was more pronounced on the ventral and dorsal side than on the medial and lateral side. These results are in accordance with Pauwels (1973). Pauwels described unloaded zones medial and lateral to the femoral head where de novo synthesis of bony tissue is possible. Histochemical staining of low and heavily sulfated glycosaminoglycans differed in normal cartilage at the rim of osteophytes compared to osteophytic cartilage. In normal cartilage, staining for low and heavily sulfated glycosaminoglycans was stronger in unmineralized cartilaginous zones than in the mineralized cartilaginous zone. In the loaded osteophytic cartilage low and heavily sulfated glycosaminoglycans were evenly distributed, while in unloaded osteophytic cartilage staining of low sulfated glycosaminoglycans and especially heavily sulfated glycosaminoglycans was weaker in the superficial cartilage than in the deep cartilage. Christensen (1985) detected only chondroitin sulfate in osteoarthritic cartilage but not keratan sulfate. Other work groups (Sweet et al. 1977; Malemud et al. 1982, 1984; Williams et al. 1988) propose that the de novo synthesized extracellular matrix of osteophytic cartilage is immature and compared it with fetal cartilage. The present results show that osteophytic cartilage contains heavily sulfated glycosaminoglycans as for example keratan sulfate. Loaded osteophytic cartilage stains stronger especially for heavily sulfated glycosaminoglycans in comparison to unloaded osteophytic cartilage. Physiologically aged articular cartilage differs from the arthritic in the keratan sulfate content. In aged cartilage the concentration of keratan sulfate is increased (Thonar et al. 1986). In arthritic cartilage keratan sulfate is decreased in comparison to unchanged cartilage of age-matched persons (Bayliss 1986, 1992). All in all, osteophytic cartilage synthesizes heavily sulfated glycosaminoglycans like keratan sulfate only under weight bearing conditions. This observation is in accordance with Huber et al. (2000) who inferred that immobilization leads to a reduction in proteoglycan synthesis and loss of cartilage. Aigner et al. (1995) showed discrete stages of cartilage differentiation in osteophytes using type I-, II- and III-
Acknowledgement. We would like to thank Ms. Antje Hanpt for her skilful assistance during the experiments.
References Aigner T, Dietz U, St0ss H, Mark K von der (1995) Differential expression of collagen types I, II, III and X in human osteophytes. Lab Invest 73:236-243 Annefeld M (1983) Gelenkknorpel und Arthrose. Huber, Bern, Stuttgart, Wien, pp 2%40 Bayliss MT (1986) Proteoglycan structure in normal and osteoarthrotic human cartilage. In: Kuettner KE, Schleyerbach R, Hascall VC (Eds) Articular cartilage biochemistry. Raven Press, New York, p 295 Bayliss MT (1992) Metabolism of animal and human osteoarthritic cartilage. In: Kuettner KE, Schleyerbach R, Peyron JG, Hascall VC (Eds) Articular cartilage and osteoarthritis. Raven Press, New York, p 487 Broom ND (1984) The altered biomechanical state of human fe'{noral head osteoarthritic articular cartilage. Arthritis Rheum 27:1028 Carstens C (1982) Untersuchungen zur mechanischen Beanspruchung der Osteophyten des arthrotischen Femurkopfes. Z Orthop 120:698-701 Christensen SB (1985) Osteoarthrosis. Changes of bone, cartilage and synovial membrane in relation to bone scintigraphy. Acta Orthop Scand Suppl 214:1M3 Fassbender HG (1983) Die Arthrose - nicht nur ein degenerativer Prozeg. Gelenkknorpel und Arthrose. Huber, Bern, Stuttgart und Wien, pp 7-28 Huber M, Trattnig S, Lintner F (2000) Anatomy, biochemistry, and physiology of articular cartilage. Invest Radiol 35: 573580 Jeffery AK (1975) Osteophytes and the osteoarthritic femoral head. J Bone Joint Surg 57B: 314324 Malemud CJ, Goldberg VM, Moskowitz RW (1982) Biosynthesis of proteoglycan in vitro by cartilage from human osteochondrophytic spurs. Biochem J 206:329-341 Malemud CJ, Moskowitz RW, Goldberg VM (1984) Biosynthesis of sulfated proteoglycan in vitro by cells derived from human osteoehondrophytic spurs of the femoral head. Connective Tissue Res 12:319-335 Miesenb6ck G, Siebels W, Bliimel G, Giinther R (1987) Mechanochemie des Gelenkknorpels und Arthrose - Ein pathophysiologischer Zusammenhang. Z Rheumatol 46:311-316 Pauwels F (1973) Atlas zur Biomechanik der gesunden und kranken Htifte. Springer, Berlin Pottenger LA, Phillips F, Draganich L (1990) The effect of marginal osteophytes on reduction of varus-valgus instability in osteoarthritic knees. Arthritis Rheum 33:853-858
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Scott JE, Dorling J (1965) Differential staining of acid glycosaminoglycans (mucopolysaccharides) by alcian blue in salt solutions. Histochemie 5:221-233 Seibel MJ (1989) Komponenten der extrazellul~iren Gewebematrix als potentieller ,,Marker" des Bindegewebs-, Knorpel- und Knochenmetabolismus bei Erkrankungen des Bewegungsapparates. Z Rheumatol 48:6-18 Sweet MBE, qlaonar EJ, Immelman AR, Solomon L (1977) Biomechanical changes in progressive osteoarthritis. Ann Rheum Dis 36:387-398
Tillmann B, Schtinke M (1991) Struktur und Funktion extrazellularer Matrix. Anat Anz Suppl 168:23-36 Williams JM, Katz RJ, Childs D, Lenz ME, Thonar EJ (1988) Keratan sulfate content in the superficial and deep layers of osteophytic and nonfibrillated human articular cartilage in osteoarthritis. Calcif Tissue Int 42:162-166 Accepted June 5, 2002
71
Topographical and histological examination of osteophytes taken from arthrotic femoral heads Horst Claassen 1 and Thomas Tschirner 2
11nstitut ftir Anatomie der Universitgt Rostock, Gertrudenstrasse 9, 18055 Rostock, Germany, 2Anatomisches Institut der Universit~t Kiel, Olshausenstrasse 40, 24098 Kiel, Germany
Summary. Until now it is not known whether osteophytes
Key words: Articular cartilage - Femoral head - Osteo-
of the femoral head develop because of pathological joint phytes - Topographical localization - Extracellular matrix alterations or arise from normal remodeling processes - Proteoglycans - Glycosaminoglyeans secondary to osteoarthrosis. Firstly, we analysed the topographical localization of osteophytes. We then compared the extracellular matrix components of macroscopically normal cartilage from the margin of osteophytes with os- I n t r o d u c t i o n teophytic cartilage from weight bearing and non-weight bearing zones by histochemical staining of low and Osteoarthritis is characterized by pathological changes of heavily sulfated glycosaminoglycans. For examination 65 the hyaline articular cartilage, the subchondral bone and femoral heads were taken during endoprosthetic hip surthe synovial tissue (Jeffrey 1975). In the zone of articular gery Osteophytes from different locations and macroscocartilage bearing the greatest load, degradation of hyaline pically normal cartilage from the margin of osteophytes cartilage and denudation of the subchondral bone can be were excised, decalcified and embedded in paraplast. A observed. The disappearance of articular chondrocytes is lateral or medial localization of osteophytes (47 cases) accompanied by a reduction in synthesis of collagens and was more common than a ventral or dorsal position (18 proteoglycans (Annefeld 1983; Fassbender 1983; Broom cases). Histochemical staining for low and heavily sul1984; Miesenb6ck et al. 1987; Seibel 1989; Tilhnann et al. fated glycosaminoglycans from normal cartilage at the 1991). rim of osteophytes was stronger in the unmineralized carRepair attempts were observed in parallel to these detilaginous zones compared to the mineralized cartilagigenerative changes. Mitotic cell divisions took place in nous zone. Weight bearing zones of osteophytic cartilage, the destroyed cartilage. These processes form so-called on the other hand, showed an even distribution of the clusters of chondrocytes; i.e. groups of cells that lie close two differently sulfated glycosaminoglycans. Surprisingly, together. These cells are not able to replace the destroyed non-weight bearing zones of osteophytic cartilage showed cartilage matrix and so degradation continues. While cara weaker staining for low and especially for heavily sultilage was destroyed in the weight bearing zones, producfated glycosaminoglycans in the superficial cartilage layer tive processes took place in the non-weight bearing than in the deep cartilage layer. Altogether, osteophytic marginal zones, which led to exostoses or osteophytes cartilage can be regarded as a reparative phenomenon for two reasons: Firstly, osteophytes arise very often at the (Pauwels 1973). Because osteophytes succumb to pressure weight bearing lateral and medial femoral head. Secondly, demands like normal articular cartilage, this process leads despite local differences in osteophytic cartilage, the same to an enlargement of the joint surface (Carstens 1982). types of glycosaminoglycans are synthesized as in normal Therefore, osteophytes diminish the load of the damaged joint surface (Williams et al. 1988). cartilage at the margin of osteophytes. Until now it is unclear whether osteophytes are formed because of pathological joint alterations or they are due to normal remodeling processes in the course of osteoarthritis. In the present study low and heavily sulCorrespondence to: H. Claassen fated glycosaminoglycans were stained histochemically in E-mail: [email protected]
II
Ann Anat (2003) 185:67-71 © Urban & FischerVerlag
http:llwww.urbanfischer.deljournalslannanat
0940-9602103118511-67$I 5.0010
"i!:~i!~!i!:~ ~ ~: ~!i~
/ii
ii i ~I! iil/i:i
i.....
Fig. 1. Lateral osteophyte from the femoral head of a 68-year-old male. Broad layer of fibrocartilage (FC) with degenerative changes visible as fissure (arrow). FC = fibrocartilage, B = bone. Azan. (x 10). Fig. 2. Medial osteophyte from the femoral head of a 73-year-old female. Osteophyte with a broad basis. The superficial layer of fibrocartilage (FC) which has different thickness shows degenerative changes. FC = fibrocartilage, B = bone. Azan. (x 10). Fig. 3. Histochemical staining of low and heavily sulfated glycosaminoglycans in different zones of normal articular cartilage at the rim of osteophytes. All zones of unmineralized cartilage show stronger staining for low (a) and heavily (b) sulfated glycosaminoglycans than the mineralized cartilage zone (MZ). Especially strong staining is observed in the radial fiber zone (RFZ). In the mineralized cartilage zone (MZ) only territorial matrix is positive for low (a) and heavily (b) sulfated glycosaminoglycans. T F Z = tangential fiber zone, TZ = transition zone, R F Z = radial fiber zone, MZ = mineralized zone. Alcian blue with 0.3 M (a) and 0.8 M (b) MgC12. a, b (x 40). Fig. 4. Histochemical staining of low and heavily sulfated glycosaminoglycans in osteophytic cartilage of weight bearing and nonweight bearing zones, a: Medial osteophyte stained for low sulfated glycosaminoglycans in general view. A weight bearing zone (c) can be discriminated from a non-weight bearing zone (e). b: Medial osteophyte stained for heavily sulfated glycosaminoglycans in general view. A weight bearing zone (d) can be discriminated from a non-weight bearing zone (f). c, d: Weight bearing zones of osteophytic cartilage: Low (c) and heavily (d) sulfated glycosaminoglycans were detected in all zones of fibrocartilage (FC) in even distribution, e: Non-weight bearing zone of osteophytic cartilage: In the upper third (broken line) of fibrocartilage staining for low sulfated glycosaminoglycans was weaker than in the lower two thirds, f: Non-weight bearing zone of osteophytic cartilage: In the upper half (broken line) of fibrocartilage staining for heavy sulfated glycosaminoglycans was nearly negative while it was weak in the lower half. Alcian blue with 0.3 M (a, c, e) or 0.8 M (b, d, f) MgC12. a, b (xl0); c, d (x 40); e, f (x 30). 68
osteophytic and normal cartilage at the rim of osteophytes. We tried to find out whether the extracellular matrix of osteophytes shows similarities to adult hyaline articular cartilage by comparing the former with normal cartilage.
Material and methods 65 resected femoral heads from patients undergoing endoprosthetic surgery for severe coxarthritis served as material for the present study. The age of the patients ranged from 30 to 90 years. The topographical localization of the osteophytes at the peripheral cartilage-bone border was documented immediately after surgery. Osteophytes and macroscopically normal cartilage marginal to osteophytes were dissected afterwards using an oscillating saw at different sites. Osteophytes and normal cartilage from 28 femoral heads were fixed in Bouin's solution (15 parts picric acid, 5 parts 4% formaldehyde, 1 part acetic acid). After washing for 2 days, specimens were decalcified in 10% buffered EDTA at 4 °C. EDTA-solution was changed every third day, decalcification was controlled weekly by X-rays. Following decalcification, specimens were dehydrated in several portions of alcohol and embedded in paraffin. Then, 7 gm paraffin sections were used for toluidine blue and azan staining. Low and heavily sulfated glycosaminoglycans were demonstrated histochemically according to Scott and Dorling (1963) with alcian blue containing 0.3 M or 0.8 M magnesium chloride, respectively.
Results
Localization of osteophytes. During postoperative macroscopieal investigation of femoral heads an area of nude bone with different extension was found in the region of the fovea capitis femoris. This area of uncovered bone was uneven and showed recesses. Adjacent to this pathologically changed region there was a cartilage of very soft quality which thinned to the rims of the femoral head. Protruding rims (osteophytes) of different size rose at the peripheral cartilage bone border. The superficial cover of the osteophytes was of a soft quality. Osteophytes at the cartilage-bone border were arranged in the shape of a closed ring in 23 femoral heads, while osteophytes were found in various places in 42 femoral heads. To summarize, in 65 femoral heads osteophytes were found more often on the medial and lateral side than on the ventral and dorsal side of the femoral head. A ventral or ventro-medial localization of osteophytes was found in 26 femoral heads, while a lateral or dorso-lateral one occurred in 21 femoral heads. A ventral localization of osteophytes was observed in 10 specimens, while a dorsal localization was observed in 8. Histological examination of osteophytes. The superficial cover of osteophytes does not show a uniform structure.
Fibrocartilage was often found beneath the superficial layer of connective tissue containing collagen fibers. Moreover, the surface of some osteophytes contained only fibrocartilage. In another form of osteophytes, a superficial layer of connective tissue containing collagen fibers converted to fibrocartilage after a certain distance (Fig. 1, 2). Often fibrocartilage showed degenerative changes in form of fibrillations and fissures (Fig. 1). If fissures occurred, the subchondral bone was sometimes completely uncovered. The characteristic zones of articular cartilage, the tangential fiber zone, the transitional zone and the radial fiber zone were detected only in osteophytes with a thick layer of fibrocartilage. The cells of fibrocartilage kept their round shape from the deep to the superficial cartilaginous zone and did not flatten in the direction of the superficial cartilage layer. Moreover, chondrocytes were not localized in groups, but were positioned singularly. The collagen fibers were arranged vertically from the deep to the superficial layer of fibrocartilage. This observation was made in demasked collagen fibers (Fig. 2).
Histochemical staining of low and heavily sulfated glycosaminoglycans in osteophytes. As far as ,,normal" cartilage in the periphery of osteophytes is concerned, more low and heavily sulfated glycosaminoglycans were detected in the different zones of unmineralized cartilage than in the mineralized cartilage (Fig. 3 a, b). Low and heavily sulfated glycosaminoglycans were distributed evenly in the tangential fiber zone and transitional zone (Fig. 3 a, b). However, the extracellular matrix of the radial fiber zone was stained stronger for the two glycosaminoglycans of different sulfation in comparison to the superficial cartilage layers. In osteophytic cartilage, it is of interest that the distribution of low and heavily sulfated glycosaminoglycans differed from corresponding distribution in normal cartilage at the margin of osteophytes (Fig. 4 a ,b). The typical zones of articular cartilage were not found in osteophytic cartilage which mainly contained fibrocartilage. Weight bearing and non-weight bearing regions can be discriminated in osteophytic cartilage (Fig. 4 a, b). In the weight bearing zone the extracellular matrix of osteophytic cartilage contained low and heavily sulfated glycosaminoglycans in even distribution (Fig. 4c, d). In the nonweight bearing zone, the histochemical staining of low sulfated glycosaminoglycans was weaker in the upper third near to the joint surface than in the lower two thirds (Fig. 4 e). The histochemical detection of heavily sulfated glycosaminoglycans was weaker all in all in the nonweight bearing zone compared to the low sulfated glycosaminoglycans. In the upper half near the joint surface only a few heavily sulfated glycosaminoglycans were detected. In the lower half of osteophytic cartilage, the histochemical reaction with alcian blue was somewhat stronger than in the upper half (Fig. 4 f).
69
collagen specific antibodies and cDNA probes. The authors identified a fibrocartilaginous chondrocyte phenotype that expresses type II- and III-, but not typ Icollagen. In conclusion, osteophytic cartilage contains extracellular matrix components of normal articular cartilage like type II-collagen and low and heavily sulfated glycosaminoglycans. On the basis of these results it seems likely that osteophytic cartilage shows signs of a repair phenomen.
Discussion The present paper has two goals: (1) Evaluation of the localization of osteophytes of the femoral head, (2) low and heavily sulfated glycosaminoglycans of the extracellular matrix of osteophytes were analyzed histochemically. Osteophytes are bone-cartilage-like protrusions at articular joints (Pottenger et al. 1990). Beside narrowing of the joint space, sclerosis and subchondral cysts, the occurrence of osteophytes is a criterion in establishing the diagnosis of osteoarthrosis. Observation of the localization of osteophytes in 65 femoral heads showed that osteophytes can grow at any site (Jeffery 1975). In 47 femoral heads more pronounced osteophytic growth was observed in medial and lateral localizations than in ventral and dorsal areas. In 18 femoral heads, osteophyte growth was more pronounced on the ventral and dorsal side than on the medial and lateral side. These results are in accordance with Pauwels (1973). Pauwels described unloaded zones medial and lateral to the femoral head where de novo synthesis of bony tissue is possible. Histochemical staining of low and heavily sulfated glycosaminoglycans differed in normal cartilage at the rim of osteophytes compared to osteophytic cartilage. In normal cartilage, staining for low and heavily sulfated glycosaminoglycans was stronger in unmineralized cartilaginous zones than in the mineralized cartilaginous zone. In the loaded osteophytic cartilage low and heavily sulfated glycosaminoglycans were evenly distributed, while in unloaded osteophytic cartilage staining of low sulfated glycosaminoglycans and especially heavily sulfated glycosaminoglycans was weaker in the superficial cartilage than in the deep cartilage. Christensen (1985) detected only chondroitin sulfate in osteoarthritic cartilage but not keratan sulfate. Other work groups (Sweet et al. 1977; Malemud et al. 1982, 1984; Williams et al. 1988) propose that the de novo synthesized extracellular matrix of osteophytic cartilage is immature and compared it with fetal cartilage. The present results show that osteophytic cartilage contains heavily sulfated glycosaminoglycans as for example keratan sulfate. Loaded osteophytic cartilage stains stronger especially for heavily sulfated glycosaminoglycans in comparison to unloaded osteophytic cartilage. Physiologically aged articular cartilage differs from the arthritic in the keratan sulfate content. In aged cartilage the concentration of keratan sulfate is increased (Thonar et al. 1986). In arthritic cartilage keratan sulfate is decreased in comparison to unchanged cartilage of age-matched persons (Bayliss 1986, 1992). All in all, osteophytic cartilage synthesizes heavily sulfated glycosaminoglycans like keratan sulfate only under weight bearing conditions. This observation is in accordance with Huber et al. (2000) who inferred that immobilization leads to a reduction in proteoglycan synthesis and loss of cartilage. Aigner et al. (1995) showed discrete stages of cartilage differentiation in osteophytes using type I-, II- and III-
Acknowledgement. We would like to thank Ms. Antje Hanpt for her skilful assistance during the experiments.
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