Cardiac hypertrophy is an adaptive growth mechanism that is mediated by several signaling pathways including those coupled through extracellular matrix (ECM) proteins to sites of cell adhesion. Integrins are an important class of receptors for the ECM proteins that can mediate both force transmission and signal transduction. Consequently, integrins have been proposed to be an important mechanosensor in myocytes and to play a central role as mechanotransducers during normal development and in response to mechanical forces associated with physiological and pathophysiological states. However, the precise mechanisms that define how integrins are involved in sensing and transducing physical forces into cellular signals are unclear. Our long-term goal is to understand the mechanisms involved in integrin-based mechanical signaling in the myocardium. Our central hypothesis is that initiation of mechanically induced cell signaling in cardiac myocytes by integrins (alpha3beta1 and alpha5beta1) involves either bond formation, bond stressing and/or bond dissociation between integrins and ECM proteins, fibronectin (FN), collagen (CN) and laminin (LN). We hypothesize that the integrin-mediated cell signaling events in cardiac myocytes will involve alterations in cytosolic calcium concentration ([Ca2+]j), cell stiffness/elasticity or a cellular mechanical response. In this exploratory/developmental study, we will establish a method to study the mechanotransduction processes in mouse cardiac myocytes. A unique aspect of this study is the use of Atomic Force Microscopy (AFM) as a tool to probe integrin function and cellular mechanics.
The specific aims of this proposal are: 1) to determine if integrin-ECM bond formation, bond stressing or forced bond dissociation on the myocyte membrane result in changes of [Ca2+]i; and 2) to determine if the [Ca2+]j changes result in detectable alterations of cortical myocyte stiffness/elasticity or mechanical responses. The binding forces between ECM and their integrin receptors will be measured in the myocytes using the AFM. The myocytes responses to integrin-ECM interactions will be investigated by measuring cytosolic Ca2+, cell stiffness and cellular mechanical responses to a mechanical stimulus. These studies will provide some of the first measurements of many critical quantitative parameters of integrin-ECM bonding in single cardiac myocytes. This will provide novel insight into transduction mechanisms of myocytes.
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