Figure 4.

Mitogenic activities of EGF and recombinant HB-EGF proteins in BE, B108, and BE2 cells. (A) Schematic structures of HB1 and HB2. HB1 is a GST fusion protein containing the EGF-like domain of human HB-EGF. HB2 is essentially similar to HB1 except for three basic amino acids within the EGF-like domain that are changed to the indicated amino acids. Amino acids are shown as one-letter symbols, and numbers in the figure indicate the number of amino acids from the N-terminus of human HB-EGF. (B) Flow cytometric analysis of cell surface EGFR and EGFR-EpoR chimeric receptor with anti-EGFR antibody. Ba/F3, BE, BE2, and B108 cells treated with anti-EGFR antibody recognizing the extracellular domain of EGFR were stained with FITC-conjugated secondary antibody. (C–E) The mitogenic activities of EGF, HB1, and HB2 toward BE cells (C), B108 cells (D), and BE2 cells (E) were determined by measuring DNA synthesis. BE cells, B108 cells, and BE2 cells were cultured with EGF, HB1, or HB2 for 24 h, followed by incubation with [³H]thymidine (37 kBq/ml) at 37°C for 4 h. Cells were harvested, and the amounts of [³H]thymidine incorporated into DNA were measured. Data are shown as means ± SD obtained from three independent experiments.