ALTERATIONS IN CYTOSKELETAL AND NUCLEAR MATRIX-ASSOCIATED PROTEINS DURING APOPTOSIS

Evidence exists that apoptosis or programmed cell death plays an impor tant role in tumor growth. Morphologically, apoptosis is characterized by membrane blebbing and nuclear fragmentation in individual cells. I n this study me investigated changes in the nuclear organization in sp ontaneously occurring apoptotic cells in several cancer cell lines usi ng several antibodies to nuclear matrix constituents. It appeared that nuclear matrix components remained detectable in cells undergoing spo ntaneous apoptosis. The same results were found when apoptosis was ind uced by cycloheximide in the non-small cell lung cancer cell line MR65 . Using this induction method, the percentage of apoptotic cells in MR 65 cells increased, allowing a more detailed and extensive examination of nuclear matrix alterations together with cytoskeletal changes. To study the expression of cytokeratins, type A- and B lamins, a nuclear matrix-associated 13 kDa U(1)RNP particle and the Ki67-antigen, immuno cytochemistry in combination with confocal scanning laser microscopy w as used. Apoptotic cells were identified based on nuclear morphology a nd the in situ nick translation assay. Whereas immunoreactivity agains t lamins and Ki67-Ag was rapidly lost during apoptosis, expression of the 13 kDa protein and, in early apoptotic stages, also cytokeratin ex pression, was observed to remain present. Dead cells lacked reactivity with all the antibodies tested. The persistence of nuclear matrix com ponents is therefore a useful marker for the detection of apoptosis.