Anti-CD3-induced apoptosis in T-cells from young and old mice

Light scatter measurements using flow cytometry indicated that T cells from young and old mice undergo apoptosis following activation with immobilized anti-CD3. The percentage of cells in apoptosis after 20 h activation was significantly greater (p < .001) in cultures containing cells from older animals. The mean percentages of apoptotic T cells from young and old mice after 20 h activation were 19.3% and 33.0%, respectively. The proportion of viable cells after 20 h activation was significantly higher (p < .003) in the young (mean = 78.4%) than in the old animals (mean = 65.8%). Simultaneous measurements of light scatter and fluorescence indicated that apoptotic T cells contained both the CD4+ and the CD8+ T-cell phenotypes. The frequency of apoptotic CD8+ T cells was elevated (p < .007) in older animals, where the mean percentage was 15.1%, compared to 5.3% in the young. The most dramatic difference between young and old (p < .0008) was seen in the percentages of viable CD4+ T cells after 20 h activation. The mean viable CD4+ T-cell percentage was 33.7% in- the young and 21.4% in the old. CD4+ cells expressing high levels of CD45RB (CD45RBhi) after activation for 20 h possessed light scatter and bright fluorescence properties characteristic of viable cells, whereas CD4+/CD45RBlo density cells could be identified as apoptotic based on their decreased CD4 fluorescence and scatter characteristics. CD4+ cells from young animals were predominantly CD45RBhi, whereas CD4+ cells from the old had greater levels of CD454RBlo cells. In addition to light scatter changes, measurement of DNA content after 40 h activation revealed the presence of a sub-G1 DNA apoptotic peak and a viable cell cycle distribution. After 40 h of activation, there was an increase in the percentage of apoptotic cells in both young and old mice, with the greatest increase seen in the cells from older animals. Further evidence supporting the process of apoptosis in 40 h-activated cells was confirmed by the appearance of DNA strand breaks detected by in situ nick translation.