Abnormal Cardiac Cross-Bridge Kinetics in a Troponin T ILE79ASN Transgenic Mouse Model

Abstract

The missense mutation Ile79Asn in human cardiac troponin T (HcTnT-I79N) has been associated with phenotypic outcomes of familial hypertrophic cardiomyopathy, arrhythmias, and sudden cardiac death. Previous characterization of a transgenic mouse model bearing the HcTnT-I79N mutation, which expressed 52% HcTnT-I79N of total troponin T, demonstrated increased Ca2+ sensitivity of isometric force. However, little is known about the changes in cross-bridge kinetics, if any, that result from the mutation. This study investigates isometric force, sinusoidal stiffness and rate of tension redevelopment (kTR) of skinned papillary muscle fiber bundles at two sarcomere lengths (1.9 µm and 2.1 µm); cardiac muscle was isolated from the left ventricle of non-transgenic wild-type (WT) mice or transgenic mice bearing the HcTnT-I79N mutation. Sarcomere length (SL) was set at pCa8 using HeNe laser diffraction after the attachment points of each skinned fiber bundle were fixed with glutaraldehyde to minimize end compliance. Sinusoidal stiffness and kTR were obtained after recording steady-state isometric tension by oscillating the fiber bundle 0.2% of its relaxed length or by applying a ramped shortening followed by a quick re-stretch to original fiber bundle length, respectively. As previously reported, the HcTnT-I79N mutation resulted in increased Ca2+-sensitivity of isometric force although no difference in maximum force per cross-sectional area was observed. Length-dependent activation at the longer SL resulted in increased Ca2+-sensitivity of isometric force and sinusoidal stiffness, and decreased kTR for both WT and HcTnT-I79N. However, HcTnT-I79N exhibited faster kTR and greater sinusoidal stiffness at sub-maximum pCa (-log[Ca2+]) solutions compared to WT regardless of SL. These data indicate that HcTnT-I79N mutation, and SL, markedly influence not only the dynamics of individual regulatory units, but also the kinetics of cross-bridge cycling.