Increased substrate stiffness, modeling fibrotic myocardium, drives cardiac myocyte hypertrophy through enhanced FAK phosphorylation and PIP2 signaling at the Z-disc, leading to increased actin assembly.
Hypothesis-generating for stiffness-driven myocyte hypertrophy; leaves open in vivo relevance or therapeutic targeting of FAK/PIP2.
Much diseased human myocardial tissue is fibrotic and stiff, which increases the work that the ventricular myocytes must perform to maintain cardiac output. The hypothesis tested is that the increased load due to greater stiffness of the substrata drives sarcomere assembly of cells, thus strengthening them. Neonatal rat ventricular myocytes (NRVM) were cultured on polyacrylamide or polydimethylsiloxane substrates with stiffness of 10 kPa, 100 kPa, or 400 kPa, or glass with stiffness of 61.9 GPa. Cell size increased with stiffness. Two signaling pathways were explored, phosphorylation of focal adhesion kinase (p-FAK) and lipids by phosphatidylinositol 4,5-bisphosphate (PIP2). Subcellular distributions of both were determined in the sarcomeric fraction by antibody localization, and total amounts were measured by Western or dot blotting, respectively. More p-FAK and PIP2 distributed to the sarcomeres of NRVM grown on stiffer substrates. Actin assembly involves the actin capping protein Z (CapZ). Both actin and CapZ dynamic exchange were significantly increased on stiffer substrates when assessed by fluorescence recovery after photobleaching (FRAP) of green fluorescent protein tags. Blunting of actin FRAP by FAK inhibition implicates linkage from mechano-signalling pathways to cell growth. Thus, increased stiffness of cardiac disease can be modeled with polymeric materials to understand how the microenvironment regulates cardiac hypertrophy.
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Li et al. (2016) studied this question.