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DO THE CHONDROCYTES OF OSTEOARTHRITIC CARTILAGE UNDERGO CLASSICAL APOPTOSIS?



Abstract

Programmed cell death (PCD) contributes to the pathogenesis of many diseases including osteoarthritis. The principal method is apoptosis that has a well-defined and very characteristic morphology and biochemistry.

The aim of the present study was examine whether the mechanism of cell death in OA chondrocytes was classical apoptosis.

Rat thymocytes were used as controls since these cells are known to undergo classical apoptosis. Human OA cartilage was obtained from femoral head of patients (50 – 80 years) who were undergoing joint replacement surgery. Pieces of OA samples were processed into paraffin and sections incubated with the following antibodies: M3O, an antibody that recognizes the cleavage of cytokeratin 18 by caspases; annexin V, which recognizes phosphatidylserine “flip-flop” that occurs early in the apoptotic process; bcl-2, a protein whose presence protects apoptosis and c-myc, a transcription factor thought to be associated with apoptosis. To induce apoptosis, some samples were incubated with etoposide and staurosporine.

In sections of thymus we noticed the presence of numerous apoptotic bodies. The number increased when the tissue was treated with etoposide and staurosporine. Some thymocytes were immunopositive for M3O and annexin V, and the number of positive cells increased when treated for 2h with etoposide. Chondrocytes of the articular cartilage showed chromatin condensation and many vacuoles but no fragmentation into apoptotic bodies, even when treated with etoposide or staurosporine. The OA chondrocytes were immunonegative for M3O and annexin-V, even after incubation with etoposide and staurosporine. With respect to c-myc and bcl-2, both non-weight bearing and weight-bearing areas in OA sample showed more positive cells then the thymus. More chondrocytes stained for c-myc in the superficial zone of the articular cartilage in the non-weight bearing, while in the weight-bearing areas it was more in the intermediate zone. On the other hand, there were no differences in the distribution of the cells stained for bcl-2 in the articular cartilage. It is known that some events like the phosphatidylserine flip, caspase activation and apoptotic bodies fragmentation occur quickly during apoptosis, so may be difficult to detect.

The results suggest that some features of classical apoptosis, such as phosphatidylserine flip,caspase activation and apoptotic bodies formation did not take place in OA cartilage. It is known that the molecular machinery for apoptosis is not always present in tissues that are undergoing programmed cell death, which seems to be case for OA chondrocytes.

Abstracts prepared by Dr P E Watkins, Hodgkin Building, Guys Campus, King’s College London.