Loss and shedding of surface markers from the leukemic myeloid monocytic line THP-1 induced to undergo apoptosis.

Previous studies have established that a relationship exists between apoptosis and cell surface (ecto-) peptidase activity. Thus dose-dependent increases were found both in ectopeptidase activities and in the proportion of cells undergoing apoptosis in HeLa cell monolayers after exposure to UV and other perturbants causing arrest of DNA synthesis (indirectly or directly as a result of DNA damage). The nature of the correlation made no distinction as to whether an increase in peptidase activity was causal of, or consequential to apoptosis, nor whether the increase was a general response by all cells. As a wider approach to understanding the possible role played by ectopeptidases in apoptosis, we report the effect on expression of a known ectopeptidase, aminopeptidase N (CD13), by a myelomonocytic cell line induced to undergo apoptosis. Using THP-1 cultures exposed to low concentrations of ethanol, we used FACS technology to sort for early apoptotic cells that have an increased ability to sequester the vital dye Hoechst 33342 while excluding nonvital dyes. Apoptosis was verified by light, fluorescence, and transmission electron microscopy, and the presence of DNA fragmentation. These early apoptotic cells showed a significant loss in CD13 labeling. Another surface marker, CD33, behaved similarly, whereas CD14 was lost globally, and not just by the apoptotic cells. Peptidase assays confirmed that an aminopeptidase was shed into the bathing media and that this activity was inhibitable both by bestatin and by a CD13 neutralizing monoclonal antibody. In treated cells, there was no evidence for an increase in cell surface protease activity directed toward a highly aliphatic nonapeptide substrate used as a model for TGF-alpha scission from its precursor form. However, other cell surface proteases of different specificity are presumably responsible for the observed shedding of CD13.