Figure 5.

Accelerated uptake of R1 aptamer on HUVECs treated with mixed Raji/BM CMs. (A). Raji or HUVECs were treated with Raji CM, BM CM or their mixture thereof prior to R1 incubation. R1 uptake was assessed with flow cytometry, and representative figures are shown. R1 uptake of HUVECs was enhanced with co-incubation of mixed Raji/BM CMs. Such phenomenon cannot be found in Raji cells. (B). Impact of CM mixture ratio. (C). Enhanced R1 uptake on HUVECs activated with BM CM and CMs from selected cells. Most of lymphoma CMs, but not myeloid leukemia or solid tumor cell lines, can accelerate R1 uptake. (D) Internalization of R1 on CM activated HUVECs. HUVECs were stimulated by mixed Raji CM and BM CM, followed by R1 incubation for different periods. Cells were then co-stained with anti-EEA1 antibody to visualize early endosomes. (E) Identification of internalization pathways. Activated HUVECs were treated with different endocytosis inhibitors, and incubated with 1 nM Cy5-R1 afterwards. (F) Competitive inhibition of cellular uptake by flow cytometry. Binding of Cy5-R1 was measured with or without co-incubation of 50-fold non-labeled R1.