In situ localization of apoptotic changes in the interface membrane of aseptically loosened orthopaedic implants.

Twenty specimens of bone-implant interface membrane from THR/TKR were used for in situ localization of apoptotic changes. A panel of antibodies was used to label leukocyte antigens (CD68 and CD3) cytokines (IL-1alpha and IL-1beta) and apoptosis inhibiting and promoting proteins (bcl-2 and bax) by means of immunohistochemical techniques. A DNA fragment test on the tissue sections was also carried out to confirm actual cell death using the enzyme terminal deoxy nucleotidyl transferase (TdT) to incorporate biotinylated nucleotide with the 3'-OH DNA ends. Leukocyte antigen staining showed that there were large numbers of CD68 positive macrophages as well as multinucleate giant cells (MNGC) but that CD3 positive lymphocytes were also present in the interface membrane. The leukocyte surface antigen staining pattern corresponded to previous findings [1]. Immunostaining with bcl-2 and bax antibodies revealed that both of these proteins were expressed in the cytoplasm of the cells in the interface membrane but they showed different cellular patterns. Bcl-2 was localized in a small number of lymphocyte-like cells while bax was expressed by large numbers of cells, mainly macrophages. The number of cells which expressed bcl-2 was significantly lower than that of bax (P<0.01). DNA fragment localization occurred mostly in a layer of cells (1- 3 cells deep) next to the implant surface. Again the level of DNA fragment-containing cells was significantly lower than that of bax positive cells (P<0.01). The results, for the first time, indicate that there is an apoptotic activity occurring in cells in the interface membrane, but not all the cells which express apoptosis-promoting protein (i.e. bax) will enter into the phase of cell death. Copyright 1999 Kluwer Academic Publishers