Fig. 5.

Mechanical stress induced AT1R-mediated EGFR transactivation. (A) EGFR-GFP was internalized in HEK 293 cells stably expressing AT1R that were stimulated with 1 μM AngII, EGF (10 ng/ml), or 10% cyclic stretch. This internalization was blocked by pretreatment with the Src inhibitor PP2 (5 μM) or the EGFR inhibitor AG1478 (1 μM). Arrowheads, internalized EGFRs. (B) Knockdown of β-arrestin1/2 by siRNA blocked AngII- and mechanical stretch–induced EGFR transactivation in HEK 293 cells stably expressing AT1Rs (n = 4 independent experiments with 50 to 80 cells per experiment). (C) Knockdown of β-arrestin1/2 in HEK 293 cells stably expressing AT1Rs prevented EGFR phosphorylation in response to 1 μM AngII, osmotic stretch, or 10% cyclic stretch, but not to EGF ligand (10 ng/ml). *P < 0.001, control siRNA no stimulation (NS) compared to control siRNA treated with AngII and Str; †P < 0.001, control siRNA Ang or Str compared to β-arrestin1/2 siRNA AngII or Str. n = 3 independent experiments with 2 × 10⁶ to 3 × 10⁶ cells per experiment. (D) Mechanical stretch in WT hearts stimulated activation of ERK, which was sensitive to the EGFR inhibitor erlotinib (Erl) (n = 4 hearts). *P < 0.0001, Str no treatment (open bar) compared to Str treated with Erl (solid bar); *P < 0.0001, EC EGF treated (solid bar) compared to EC no treatment (open bar). Scale bar, 10 μm.