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The recovery of adult rabbit articular cartilage after nine weekly intra-articular injections of hydrocortisone acetate was investigated using biochemical, autoradiographic, and tracer methods. After the cessation of steroid injections, the rates of synthesis of proteoglycans and collagen increased by up to 900 per cent. The increase occurred because of accelerated metabolism of existing cells and an increase (caused by mitosis) in the number of functioning chondrocytes. The proteoglycan concentration only returned to normal values after six months.

The repair of articular cartilage removed from the femora of adult rabbits at the knee was studied. When multiple perforations were made through the subchondral bone a cartilaginous material, staining heavily with safranin O, first filled the holes. This material resembled hyaline cartilage, and by twelve months there was complete resurfacing of the joint, although the material lost its hyaline appearance after eight months and at one year it resembled dense collagenous tissue.

We carried out histochemical and electron-microscopic studies to examine the relationship between mineral deposition and changes in the organization of proteoglycans during mineralization in the cartilaginous growth plate of the rat. To preserve the distribution and organization of proteoglycans in the extracellular matrix and to stain proteoglycans, acridine orange was included in the solutions that were used for fixation and demineralization. In undecalcified sections, mineral crystals that initially appeared at the level of the penultimate hypertrophic chondrocyte grew into dense, spherical mineral clusters, roughly one micrometer in diameter, in the longitudinal septa of the lowermost hypertrophic zone. Following the removal of mineral with EDTA in the presence of acridine orange, proteoglycan structures with a characteristic rosette-like architecture were revealed within the matrix of the longitudinal septa of the lower hypertrophic zone. The location of these rosette-like proteoglycan structures was identical to that of the dense, spherical mineral clusters in the undemineralized controls, and they were remarkably similar in morphology and size. In the electron micrographs stained with uranyl acetate and lead citrate, the greatly increased density of the rosette-like structures and large diameter of the proteoglycan structures located in mineral clusters contrasted sharply with the slender strands of more faintly stained proteoglycan that was diffusely distributed throughout most of the extracellular matrix. X-ray microprobe analysis for sulphur confirmed the existence of proteoglycans in the rosette-like structures and demonstrated that the concentration of proteoglycans was selectively increased in these regions. The cores of the metaphyseal calcified cartilage also exhibited rosette-like proteoglycan structures, which before demineralization were totally obscured by the diffuse, dense homogeneous deposition of mineral. Without stabilization of the proteoglycans in the extracellular matrix with acridine orange, the rosette-like proteoglycan structures could not be demonstrated. These results clearly indicate that there is a selective increase in the concentration of proteoglycans in exactly the same regions where mineral clusters are formed in the hypertrophic zone of the cartilaginous growth plate.

Unlabelled: Using a recently developed technique for measuring proteoglycan by x-ray microprobe analysis, we examined six-week-old rat growth plates. In the proliferating zone, the proteoglycan concentration was found to be relatively low in the transverse septa, but somewhat higher in the longitudinal septa between the proliferating chondrocytes. In the lower hypertropic zone, in the region of the degenerating hypertrophic chondrocytes, the immediate pericellular area had a very high concentration of proteoglycan, as was obtained. These results indicate that either proteoglycan concentrations fall to very low levels in fully mineralized cartilage, or that the mineralization of cartilage interferes in some way with the measurement of microprobe signals from proteoglycans. We speculate that high levels of proteoglycan are necessary for mineralization in the epiphysis of the rat., Clinical Relevance: The spot-to-spot analysis of proteoglycan by microprobe demonstrates the usefulness of this tool in analyzing minute volumes of cartilage for proteoglycan. It further demonstrates that mineralization of growth-plate cartilage is associated with changing patterns of concentration of proteoglycan. These normal patterns may constitute a basis for comparison in similarly evaluating concentrations of proteoglycan in diseases of growth disturbance.

Intra-articular fractures were created in the distal end of the femur in adult rabbits. Fractures were then reduced in three different fashions: incomplete reduction, adequate reduction without compression of the fregments, and ideal reduction, which consisted of reduction with strong compression of the fracture fragments. Animals were killed at intervals ranging from seven weeks to one year, and the cartilage fractures were studied by light, transmission, and scanning electron microscopy, using proteoglycan stains. Cartilage fractures that were inadequately reduced or were adequately reduced without compression healed by fibrocartilage only. Fractures that were reduced with compression across the fragments healed with a tissue which, by light and electron microscopy, appeared to be hyaline cartilage, which showed the collagen and proteoglycan orientation and relationships of normal hyaline cartilage. The healing of these defects appears to be the result of a proliferation of cells emanating from the region of the tidemark, and perhaps of migration of chondrocytes from the surface into the lesion. We postulate that compression of the cartilage surfaces either creates a physical environment that allows certain chondrocytes to heal the defect with hyaline cartilage, or, by coapting the surfaces, prevents ingrowth of granulation tissue from the subchondral bone that might interfere with repair by hyaline cartilage. This study suggests a hitherto unappreciated capacity for hyaline cartilage to repair structural defects.

The destruction of rabbit articular cartilage after intra-articular injections of hydrocortisone acetate was investigated using histological, biochemical, and tracer methods. Fissures and cysts increased in number as increasing amounts of hydrocortisone were given. A linear decrease of hexosamine to less than 50 per cent after twelve injections was accompanied by insigificant changes in deoxyribonucleic acid and hydroxyproline content. The synthesis of proteoglycans and proteins was reduced to one-third, while the production of collagen dropped to less than one-fifth. The changes in thymidine incorporation were not significant. Based on these data, a model indicating the sequence of events which leads to joint destruction after intra-articular injections of glucocorticoid is proposed.

Knee joints of mature rabbits were exposed to room air for periods of time ranging from thirty minutes to one hour in an attempt to mimic the human situation in the operating room. The animals were killed after the joint had been closed and activity in a cage had been allowed for twenty-four hours. When the animal was killed, cartilage was removed and was incubated with radioactive proline for four hours before light microscopy autoradiographs were made. Other samples were prepared for study by electron microscopy. The results in the animals that were killed immediately after the cartilage was exposed to room air and in those that were killed twenty-four hours after closure of the joint were identical. Both the ultrastructural and the autoradiographic findings indicated that the entire thickness of the articular cartilage was necrotic after sixty minutes of drying. Forty-five minutes of drying produced complete necrosis of the cartilage in half of the animals. In the other half, some cells survived, although many areas of the cartilage had complete necrosis, top to bottom. Thirty minutes of drying produced patchy necrosis that extended only to the middle zone of the cartilage. In joints that were exposed to room air for one hour, necrosis of the chondrocytes was completely prevented by irrigating the joint every five minutes with Ringer lactate solution.