Insulin activation of p-AKT in IHMGECs is dependent on IGF-1R. (A) Overnight incubation of IHMGECs with IGF-1R blocking antibody, and not IgG1 isotype control or negative control (i.e., nothing added) diminished IGF-1R expression. Treatment with IGF-1 or insulin for 15 minutes did not change IGF-1R expression. (B) Following overnight incubation with IGF-1R blocking antibody, insulin and IGF-1 both showed decreased activation of p-AKT after 15 minutes of stimulation. (C) Treatment with IGF-1R blocking antibody for 7 days diminished IGF-1R expression, but not IR expression. Immortalized HMGECs were cultured in DMEM/F12/10% FBS for 5 to 7 days before protein expressions were evaluated. Insulin, 200 nM; IGF-1, 10 nM; IGF-1R blocking antibody, 10 nM; IgG1 isotype, 10 nM.