Abstract
The heart is a muscular organ with a wrapping, laminar structure embedded with neural and vascular networks, collagen fibrils, fibroblasts, and cardiac myocytes that facilitate contraction. We hypothesized that these non-muscle components may have functional benefit, serving as important structural alignment cues in inter- and intra-cellular organization of cardiac myocytes. Previous studies have demonstrated that alignment of engineered myocardium enhances calcium handling, but how this impacts actual force generation remains unclear. Quantitative assays are needed to determine the effect of alignment on contractile function and muscle physiology. To test this, micropatterned surfaces were used to build 2-dimensional myocardium from neonatal rat ventricular myocytes with distinct architectures: confluent isotropic (serving as the unaligned control), confluent anisotropic, and 20 μm spaced, parallel arrays of multicellular myocardial fibers. We combined image analysis of sarcomere orientation with muscular thin film contractile force assays in order to calculate the peak sarcomere-generated stress as a function of tissue architecture. Here we report that increasing peak systolic stress in engineered cardiac tissues corresponds with increasing sarcomere alignment. This change is larger than would be anticipated from enhanced calcium handling and increased uniaxial alignment alone. These results suggest that boundary conditions (heterogeneities) encoded in the extracellular space can regulate muscle tissue function, and that structural organization and cytoskeletal alignment are critically important for maximizing peak force generation.
Highlights
The striated muscle of the heart is organized into anisotropic laminae that wrap around the ventricular cavities [1]
The PDMS/PIPAAm coated cover slips were UV ozone treated (Model No 342, Jelight Company, Irvine, CA, USA.) and functionalized with the extracellular matrix (ECM) protein fibronectin (FN) according to one of three conditions; (i) isotropic myocytes randomly arrayed in a continuous monolayer (ISO), (ii) anisotropic myocytes aligned in a continuous monolayer (ANISO) or (iii) lines where multicellular muscle strands are arranged in parallel without lateral coupling between the strands (LINES)
We have described a functional role for extracellular heterogeneities in the organization of engineered cardiac muscle, which enhances systolic performance in the heart by potentiating the spatially ordered assembly of the contractile apparatus
Summary
The striated muscle of the heart is organized into anisotropic laminae that wrap around the ventricular cavities [1]. Perturbation of these cell populations and the extracellular matrix (ECM) is commonly associated with maladaptive remodeling, resulting in decreased cardiac output [6] and arrhythmogenesis [7,8] It remains unclear whether these structural changes are causative or correlative with contractile function and dysfunction. We have reported that geometric cues in the ECM act as boundary conditions that regulate myofibrillogenesis [13,14,15] and that the bundled, parallel alignment of myofibrils enhances myocyte contraction strength [16] These reports suggest that boundary conditions imposed on muscle cells in the heart may be an important regulator of cardiac tissue form and function
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