Lesions of Articular, Sternal and Growth Plate Cartilage in Rats

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1 Vet. Pathol. 22: (1 985) Lesions of Articular, Sternal and Growth Plate Cartilage in Rats K. YAMASAKI and S. INUI Chemical Biotesting Center, Chemicals Inspection & Testing Institute, Hita; and Research Institute for Animal Science in Biochemistry and Toxicology, Sagamihara, Japan Abstract. Degenerative osteoarthritis was observed in the femur and sternum in specific pathogen free Fisher 344 rats at and months of age. Histological changes consisted of degeneration of the matrix and erosion in the articular cartilage of the femur. In the sternal cartilage, degeneration and necrosis of the matrix, formation of cysts, and cleft-like fractures were observed. In addition, degeneration of the matrix, appearance of eosinophilic streaking, and necrosis and fissure between the growth plate and epiphyseal trabeculae were seen in the femur. The changes in the sternum were essentially the same as the femur. Degenerative joint disease (osteoarthritis) is by far the most common form of articular disease, and usually occurs in large or small synovial joints. This condition is encountered in man,4. lo domestic animals,l x. 11- fowls,s. l4 and laboratory animals, - ~ l3 although old rats sometimes have localized, chondromucoid cystic degeneration of the articular cartilage which does not progress to degenerative osteoarthritk8.. In addition, no changes in the growth plate are observed in laboratory animals. The pathological findings of degenerative osteoarthritis and changes in the growth plate occurring in the femur and sternum of rats are described. Materials and Methods One hundred twenty specific pathogen free Fisher 344 rats were studied: males and females, months of age; and males and females at both and months of age. The rats were placed in wire-mesh cages with three rats per cage and kept in a barrier system animal room conditioned at a temperature of 24 & 1 C with a relative humidity of 55 k 5%. Food and water were supplied ad libitum. Diet consisted of M for rats (Oriental Yeast Co., Ltd., Itabashi, Tokyo, Japan). Rats were killed by bleeding from the posterior vena cava by opening the abdominal cavity under ether anesthesia. After necropsy, the femur and sternum from each rat were fixed in 10% formalin solution. After decalcification in 10% formic acid-formalin solution, bone tissue blocks were embedded in paraffin, cut into sections, stained with hematoxylin and eosin (HE), Mallory-azan, periodic acid-schiff (PAS), toluidine blue, phosphotungstic acid hematoxylin, and Gomori s silver impregnation. We graded the change in the growth plate of the femur and 46 sternum. A change graded mild indicated the disappearance and dissolution of the eosinophilic homogeneity of the matrix in a small area. Moderate was when the mild change spread to a large area or when cartilage necrosis appeared in a small area. Severe indicated that cartilage necrosis was observed in a large area. Results No macroscopic abnormalities were observed, however, microscopically there were changes seen in the articular, sternal, and growth plate cartilage. Articular cartilage: Abnormalities were observed in the femur of -month-old rats (table I). Eosinophilic homogeneity of the matrix was lost with a decrease in PAS- and toluidine blue-positive substance and appearance of collagen fibers. Focal areas of the tangential layer were destroyed and resulted in a roughened surface (figs. 1, 2). Degeneration and necrosis of chondrocytes were noted in the radial zone and led to the appearance of small clusters. The above lesions were limited to the articular cartilage and did not extend to the subchondral bone. Degeneration did not change with time, however, erosion increased markedly with time. Furthermore, no articular lesions were seen in seven-month-old rats (table I). Sternal cartilage: Acidophilic staining tendencies in swollen interstitial substance were seen beginning at seven months of age. Cleft-like fractures were observed parallel to the rib near the incisura costales (fig. 3). Some enlarged chondrocytes degenerated and disappeared. Cartilage sometimes was focally dissolved, and

2 Cartilagenous Lesions in Rats Table I. Prevalence of lesions in the articular cartilage of the femur Sex Male Female Age Number (months) examined Lesions Degeneration a (100) (100) (8) (100) Erosion 1 1 () 2 () 10 (6) Number affected (%). resulting cysts contained fibrous or amorphous materials in their spaces (fig. 4). Dissolutive changes with formation of cysts were observed in the center of the sternal cartilage of rats at months of age (fig. 5). In 4 the costal cartilage, unmasking and fibrillation of collagen in the matrix, appearance of eosinophilic streaking, and fissures were noted. Growth plate: Enchondral ossification was active in the femur of seven-month-old rats with no distinction of sex (figs. 6, ). Primary changes were the unmasking and fibrillation of collagen in the matrix, appearance of eosinophilic streaking and necrosis in the growth plate, and fissure between the growth plate and epiphyseal trabeculae. The disappearance and dissolution of the eosinophilic homogeneity of the matrix with decreased PAS- and toluidine blue-positive substances were characteristic-the collagen fibers were unmasked. Changes were extended from the epiphyseal hyalin cartilage zone to the diaphyseal side of the growth plate, Fig. 1: Normal articular cartilage in distal femur of seven-month-old male rat. HE. Fig. 2: Articular cartilage in distal femur of -month-old male rat; erosion in tangential zone. HE. Fig. 3: Sternal cartilage between third and fourth sternebra of -month-old male rat. Cleft-like fracture near incisura costales with unmasking and fibrillation of collagen in matrix; degenerative chondrocytes (arrows). HE. Fig. 4: Sternal cartilage between fourth and fifth sternebra of -month-old male rat. Cyst contained eosinophilic substance in space. HE.

3 48 Yamasaki and Inui Fig. 5: Sternal cartilage between third and fourth sternebra of -month-old male rat. Cyst in center of sternal cartilage. HE. Fig. 6: Growth plate in distal femur of seven-month-old male rat. Necrotic center with fissures between epiphyseal trabeculae and growth plate. Unmasking and fibrillation of collagen in matrix with disturbance of the arrangement of chondrocytes and appearance of eosinophilic streaks (arrows). HE. Fig. : Growth plate in distal femur of -month-old female rat. Closure of growth plate and unmasking and fibrillation of collagen in matrix with eosinophilic streaking (arrow). HE. Fig. 8: Growth plate in distal femur of seven-month-old male rat. Abnormal proliferation of epiphyseal hyalin cartilage zone. HE. and were severe under the fissures between the growth plate and epiphyseal trabeculae. As a result of unmasking and fibrillation of collagen in the matrix, a few chondrocytes were present in groups. Eosinophilic streaking, which was positive with phosphotungstic acid hematoxylin and stained red in Mallory-azan, appeared in severely changed parts of the matrix and around the necrosis of the cartilage. Advanced necrosis in the growth plate was observed in masses or bands with fissures and exudation of fibrin (fig. 6). Necrosis sometimes extended from the epiphyseal hyalin cartilage zone of the growth plate to the metaphyseal trabeculae. The above degenerative and necrotic changes in the growth plate were observed beginning at seven months of age and increased in severity with age (table 11). There was an abnormal proliferation of the epiphyseal hyalin cartilage cells which extended to the epiphysis (fig. 8). The cartilage mass was observed some distance from the epiphysis in some cases. In the metaphysis, many trabeculae contained necrotic cartilage (fig. 9). Cleft-like fractures frequently were present between the growth plate and cortex with activation of periosteal osteogenesis. These changes were observed in both antenor and posterior parts. Closure of the growth plate began at months of age, and ended by months of age. Changes in the sternum consisted of unmasking and fibrillation of collagen in the matrix, appearance of eosinophilic streaking and necrosis of the growth

4 Cartilagenous Lesions in Rats 49 Table 11. Seventy of degenerative and necrotic changes in the growth plate of the femur and sternum Sex Age (months) Number examined Femur Sternum " 11 (3) 2 () (63) 8 (54) 2() 11 (3) (8) 10 (6) 5 (33) (8) 5 (33) 18 (60) 6 (40) 2() 20 (6) 4 (26) 12(40) 9 (60) ll(4) 10 (33) 12(80) 8 (54) 2() ~~ Male Female (63) 2 () - = not observed; + = mild; ++ = moderate; +++ = severe. Number affected (%). plate. Necrosis of the growth plate was associated with many fissures and exudation of fibrin and had a proliferation of fibrocytes, osteoclasts and osteoblasts, and formation of osteoids (fig. 10). These sternal changes increased in severity with age in both sexes (table 11). Discussion The pathological condition of large joints affected with primary osteoarthritis, which is encountered in man,4.10 domestic animals, fowls,'. l 4 and laboratory l 3 is characterized by alterations in the cartilage and subchondral bone. In laboratory l3 animals, the lesions are common in guinea pigs,'.'. " and some strains of mice,-'. l 3 but older rats have localized, chondromucoid cystic degeneration of the articular cartilage which does not progress to degenerative o~teoarthritis.~.~. l 3 In spite of the above reports, the lesions seen in this study were generally identical to those in degenerative osteoarthritis of the synovial joints of other animals. Pathogenesis is unknown, although disturbance of the articular cartilage with age, weight-bearing on the articuli, and genetics have been suggested. It is suggested that the regressive changes of the growth plate in the femur and sternum are caused by mechanical forces which intensify with age. In addition, mechanical disruption of the epiphyseal blood supply at the growth plate-epiphysis junction is considered an essential factor in the pathogenesis. The fact that the regressive changes of the growth plate were distinct beneath the fissures between the growth plate and epiphyseal trabeculae is said to divert or prevent the passage of blood from the epiphyseal vasculature to the growth plate. Furthermore, the growth plate-metaphysis junction is stronger than the growth plate-epiphysis junction in rats, so mechanical force may be placed on the latter parts. Fig. 9: Growth plate in distal femur of seven-month-old male rat. Epiphyseal side (E); diaphyseal side (D). Necrosis from growth plate to metaphysis surrounds mature trabeculae. HE. Fig. 10: Necrotic growth plate in third sternebra of seven-month-old male rat. Reaction of fibrocytes, osteoclasts, and osteoblasts with formation of the osteoid. HE.

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