Insulin Signaling Regulates Cardiac Titin Isoform Composition in Development and Diabetic Cardiomyopathy

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Isoform switching of the giant elastic protein titin is a main mechanism for adjusting passive myocardial stiffness in perinatal heart development and chronic heart disease. Previous evidence suggested that thyroid hormone (T3) signaling converging onto the phosphoinositol-3-kinase (PI3K)/AKT pathway is an important determinant of the cardiac titin-isoform pattern in developing cardiomyocytes. We hypothesized that other activators of PI3K/AKT, particularly insulin, may similarly alter the titin-isoform composition, thereby modifying titin-based stiffness. Embryonic rat cardiomyocytes were cultured in medium containing 0.5% hormone-reduced serum and were treated with 175 nmol/L insulin for seven days. Analysis of titin-isoform expression by 2% SDS-PAGE showed a significant increase in the mean proportion of the stiff N2B titin isoform (3,000 kDa), from 53% in control cells to 64% in insulin-treated cells, the remainder being the more compliant N2BA isoform (>3,200 kDa). This insulin-dependent titin-isoform shift was blocked in the presence of PI3K-inhibitor, LY294002, suggesting that insulin regulates the cardiac titin-isoform pattern by activating the PI3K/AKT pathway. Whether this mechanism operates in vivo was studied by testing the effect of insulin deficiency on titin-isoform expression in streptozotozin-treated (STZ) rats as a model for diabetes mellitus (type 1). Within four months, STZ rats developed cardiac hypertrophy and mild left ventricular (LV) fibrosis, concomitant with elevated glucose levels. The mean proportion of N2B-titin was significantly decreased from 86% in control LV to 78% in LV of STZ rats. Wormlike chain modeling of titin elasticity suggested that such a change reduces titin-based passive stiffness by ∼6%. Results of mechanical measurements on skinned cardiac fiber bundles confirmed minor passive stiffness modifications in STZ myocardium. We conclude that insulin signaling regulates titin-isoform composition in cardiac development and could also contribute to altered diastolic function in diabetic cardiomyopathy.