Notch ligation by Delta1 inhibits peripheral immune responses to transplantation antigens by a CD8⁺ cell–dependent mechanism
J. Clin. Invest. Kenneth K. Wong, et al. 112:1741 doi:10.1172/JCI18020 [Go to this article.]

Figure 2
Dl1 is able to activate Notch signaling in reporter cell assays and in T cells. (a) CHO/N2-luc cells, which contain a construct that reports Notch pathway activation by expression of luciferase, were cultured with mitomycin C–treated K^b, K^b/Dl1, A^b, or A^b/Dl1. Untransfected CHO cells were used to control cell numbers in the assay and do not activate the Notch pathway in CHO/N2-luc cells. Cell lysates were assayed for luciferase activity after 24 hours. (b) Quantitative RT-PCR was used to assess the expression of Notch receptors and Notch ligands by L cells. All samples were normalized by assessment amplification of 18s RNA. Mouse embryonic RNA (day 11) was used as a positive control in the experiment, and transcript levels in other samples are expressed relative to this. (c) Positively selected CD4⁺ or CD8⁺ cells were cultured in the presence of immobilized CD3 Ab (0.12 μg/ml), CD28 Ab (2 μg/ml), and immobilized Dl1-Fc fusion protein (40 μg/ml). After 4 hours, cells were harvested and RNA extracted. Samples were analyzed for Hes1 transcripts and normalized using 18s amplification by quantitative RT-PCR. Data are expressed as fold induction compared with cells incubated with CD3 and CD28 Ab’s in the absence of Dl1-Fc fusion protein. All data are representative of at least three repeat experiments.