Figure 3
(a) Western blot analysis confirmed Jagged-1 overexpression in EC transduced with LZRS-Jagged-1, and demonstrated activation of Notch-1, Notch-2, and Notch-4 in these cells (NIC arrows). Transmembrane expression of Notch-1 and Notch-2 receptors (TM, arrowheads) was also increased in the Jagged-1-transduced EC compared to control cells. Results are representative of at least two experiments. (b) Notch transcriptional activity was examined using the Hes-1A/B-luc reporter construct in EC transduced with Jagged-1 or stimulated with JAG1. Luciferase activity in Jagged-1-transduced EC was 3.0-fold higher than vector control cells (P<0.01). Similarly, EC treated with 40 μM JAG1 peptide had 3.9-fold higher luciferase activity than cells treated with the same concentration of a scrambled control peptide (P<0.01). Luciferase values were normalized for transfection efficiency. Results are combined data from three independent experiments performed in triplicate. (c) WST-1 proliferation assays revealed a modest, yet statistically significant, decrease in proliferation of EC transduced with Jagged-1 compared to vector control cells (1.3-fold decrease, P<0.01). Treatment of cells with JAG1 resulted in a 1.2-fold decrease in proliferation, which was also statistically significant (P<0.05) compared to cells treated with a scrambled control peptide. Results are combined data from at least two experiments performed in triplicate.
