Figure 8.

Inhibition of CD4⁺ T cell apoptosis by direct T cell–ASM cell contact indicates bidirectional crosstalk. Purified OVA-stimulated CD4⁺ T cells were cocultured for 48 hours with ASM cells either separated in Transwells or in direct contact. Additionally, CD4⁺ T cells were cocultured in contact with the ASM cells without prior stimulation in vitro. The cocultures were exposed to BrdU for 24 hours and the CD4⁺ T cells analyzed for BrdU incorporation and total cell DNA content. (A) BrdU versus DNA density plots shown with the corresponding DNA histograms. T cell BrdU incorporation plotted against DNA content defines the following regions: 1, live quiescent cells (absence of BrdU incorporation and in G₀/G₁ phase of the cell cycle); 2, cells undergoing proliferation (incorporation of BrdU and in S or G₂/M phases of the cell cycle); 3, live postmitotic cells (incorporation of BrdU and again in G₀/G₁ phase of the cell cycle); 4, postactivation apoptotic cells (incorporation of BrdU and subdiploid DNA content); and 5, apoptotic quiescent cells (no BrdU incorporation and subdiploid DNA content). “A” in the DNA histograms corresponds to the subdiploid or sub-G₀/G₁ region and represents both quiescent and postactivation apoptotic T cells. M, cells in mitosis. (B) The corresponding side (SSC-H) versus forward (FSC-H) scatter dot plots are multicolor gated in the BrdU/DNA density plot regions defined in A. (C) Quantitative analysis of the cytometric regions defined in A and B. Data are from 4 independent experiments. *P < 0.05. Apop, apoptotic cells; incorp, incorporation.