Lysosomal-mediated degradation of apoptotic thymocytes within thymic nursecells

A thymic epithelial cell line (tsTNC-1) that maintains the ability to selec tively bind and internalize immature alpha beta TCR(lo)CD4(+)CD8(+) thymocy tes in vitro was used in long-term coincubation experiments to determine th e ultimate fate of thymocytes that remained within intracytoplasmic vacuole s of thymic nurse cells (TNCs). In an earlier report, a subset of the popul ation released from the TNC interaction was shown to mature to the alpha be ta TGR(hi)CD69(hi) stage of development, while thymocytes that bided within the TNC cytoplasm died through the process of apoptosis. Here, we show the presence of both apoptotic and nonapoptotic thymocytes within the cytoplas m of freshly isolated TNCs as well as in tsTNC-1 cells in culture. A micros copic analysis revealed total degradation of the cytoplasmic apoptotic thym ocyte population that remained in tsTNC-1 cells after an 8- to 10-h incubat ion period. A quantitative analysis showed an increase of cytoplasmic thymo cyte degradation over time to almost 80% after 9 h of incubation. However, in the presence of bafilomycin A1, which is used to inhibit acidification o f lysosomal vesicles, degradation of apoptotic thymocytes never reached 10% . These data suggest that lysosomes within TNCs play a role in the degradat ion of apoptotic thymocytes. We examined tsTNC-1 cells before the addition of thymocytes to cultures and found lysosomes to be clustered around the nu cleus in the cytoplasm of TNCs. Shortly after the internalization event, ap optotic thymocytes move to the area of the cytoplasm containing lysosomes, Using the confocal microscope, we obtained evidence that shows the degradat ion event to be facilitated through the fusion of lysosomes with the specia lized vacuoles within TNCs containing apoptotic cells. (C) 1999 Academic Pr ess.