Figure 4.
Genistein (Gen)-stimulated β-cell proliferation is mediated through the PKA and ERK1/2 signaling mechanisms. A, INS1 cells were incubated in RPMI 1640 containing 1 mm glucose with various concentrations of Gen for 15 min at 37 C. Western analysis was performed per protocol described in Materials and Methods to detect phosphorylated ERK1/2 (P-ERK1/2), which was normalized to total ERK 1/2 (B) from the same sample. C and D, INS1 cells were preincubated with PD (2 μm), H89 (H; 10 μm) or vehicle (Control) for 30 min. Cells were then treated with Gen (1 μm) or vehicle (Control) in the continued presence or absence of inhibitors at 37 C. Incubation was either terminated after 15 min to determine ERK1/2 phosphorylation by Western blotting or continued for 24 h to determine cell proliferation. E, INS1 cells were transfected with antibodies against PKA Cα plus Cβ or preimmune IgG per protocol as described in Materials and Methods. Transfected cells were treated with Gen (1 μm) or vehicle for 24 h followed by performing cell proliferation assay. Data are expressed as mean ± se of three to six experiments in triplicate each. *, P < 0.05 vs. vehicle alone or preimmune IgG-alone-treated cells.