Titin Strucure And Extensibility In Healthy And Failing Hearts

The giant sarcomeric protein, titin is the primary determinant of myocardial passive stiffness. In failing human hearts however, it has been proposed that reduced myofilament passive tension is due to an altered titin isoform expression profile1, 2. In this study we set out to directly quantify the tensile strength of titin molecules isolated from healthy and diseased myocardium. Heart failure was induced in adult male ferrets by ascending aortic coarctation. All procedures accorded with The United Kingdom Animals (Scientific Procedures) Act, 1986. Protein extracts from sham and failing hearts were separated by 2% SDS PAGE. The mean ratio of the major titin isoforms N2BA:N2B increased from 0.3 in control to 0.5 in failing hearts (n=6, p<0.01). Titin molecules were isolated from left ventricle, aligned and stretched by a receding liquid meniscus (which applied a tensile force of ∼60pN)3 and visualised by atomic force microscopy. Combed titin molecules exhibited a straightened and beaded appearance. The mean molecular diameter of titin decreased in failing hearts compared to control (0.26±0.001nm vs 0.33±0.001nm, p<0.001, n=104-130 molecules, 3 animals per group). This difference was more pronounced in the shorter molecules (<3.5 μm). The mean distance between beads was increased in failing hearts (49.3±1.5nm vs 126.8±4.5nm, p<0.001, n=370-429, 3 animals per group). The decreased titin molecular diameter combined with an increased inter-bead distance suggests that titin from failing hearts is less resistant to tensile forces when compared to control, and may help to explain the decreased titin-based passive tension observed in diseased hearts. Reference List 1. Makarenko, I. et al. Circ. Res.95, 708-716 (2004). 2. Neagoe, C. et al. Circulation106, 1333-1341 (2002). 3. Tskhovrebova, L. & Trinick,J. J. Mol. Biol.265, 100-106 (1997).