Orientation and deformation changes of left ventricular cardiomyocyte aggregates (“myofibers”) underlie many forms of cardiovascular disease. In vivo cardiomyocyte aggregate shortening (Eff) has mechanistic significance, but there exists no established technique to measure in vivo Eff. We present a pipeline to compute Eff by combining multi-slice Displacement Encoded with Stimulated Echoes (DENSE) MRI and in vivo cardiac Diffusion Tensor Imaging (cDTI) data. We show that Eff computed in healthy swine has decreased transmural variability compared to radial and circumferential strains. The spatial uniformity and mechanistic significance of Eff make it a compelling candidate for early detection of cardiac dysfunction.