Stretch-induced MAP kinase activation in cardiac myocytes: Differential regulation through β₁-integrin and focal adhesion kinase

Abstract 

Mitogen-activated protein (MAP) kinases have been implicated in hemodynamic load induced heart failure. Both angiotensin II (Ang II) and mechanical stretch activate MAP kinases in cardiac myocytes. In this study, we used a neonatal rat ventricular myocyte (NRVM) model to determine the role of focal-adhesion kinase (FAK) in β₁ integrin mediated MAP kinase activation in response to mechanical stretch in presence and absence of Ang II receptor blockade (ATB). NRVM plated on deformable membranes coated with collagen IV were exposed to 20% equiaxial static-stretch. β₁ integrin signaling was blocked by adenovirus-mediated expression of a dominant-negative form of β_1D integrin (tac-β_1D). FAK signaling was disrupted by infecting NRVM with adenovirus expressing FAK-related non-kinase (FRNK). Western blot analysis was used to assess the phosphorylation of MAP kinases. In the presence and absence of ATB, mechanical stretch caused maximal phosphorylation of ERK, p38 and JNK at 5 min, which was significantly attenuated in NRVM expressing tac-β_1D. In the presence of ATB, FRNK overexpression significantly increased basal phosphorylation of ERK (40.2±8.6% P<0.05), p38 (39.5±11.7%, P<0.05), JNK (86±29.4%, P<0.05) and stretch-induced p38 (48.1±8.7%, P<0.05) and JNK (85.0±19.4%, P<0.05) phosphorylation. However, in the absence of ATB, FRNK overexpression significantly reduced basal and stretch-induced phosphorylation of only ERK. Examination of FAK activation revealed that β₁ integrin was required for stretch-induced phosphorylation of FAK at Y³⁹⁷ and Y⁹²⁵, but not Y⁸⁶¹. In summary, mechanical stretch-activated ERK1/2, p38 and JNK through FAK independent and dependent mechanisms. β₁ integrin was required for FAK independent activation of all three MAP kinases, whereas cross-talk between β₁ integrin and Ang II receptors mediated FAK dependent regulation of ERK1/2.

Keywords: Cardiac myocytes, Focal-adhesion kinase, Integrin, MAP kinase, Mechanical stretch