Engineering Anisotropic Muscle Tissue using Acoustic Cell Patterning

Abstract

Tissue engineering has offered unique opportunities for disease modeling and regenerative medicine; however, the success of these strategies is dependent on faithful reproduction of native cellular organization. Here, it is reported that ultrasound standing waves can be used to organize myoblast populations in material systems for the engineering of aligned muscle tissue constructs. Patterned muscle engineered using type I collagen hydrogels exhibits significant anisotropy in tensile strength, and under mechanical constraint, produced microscale alignment on a cell and fiber level. Moreover, acoustic patterning of myoblasts in gelatin methacryloyl hydrogels significantly enhances myofibrillogenesis and promotes the formation of muscle fibers containing aligned bundles of myotubes, with a width of 120–150 µm and a spacing of 180–220 µm. The ability to remotely pattern fibers of aligned myotubes without any material cues or complex fabrication procedures represents a significant advance in the field of muscle tissue engineering. In general, these results are the first instance of engineered cell fibers formed from the differentiation of acoustically patterned cells. It is anticipated that this versatile methodology can be applied to many complex tissue morphologies, with broader relevance for spatially organized cell cultures, organoid development, and bioelectronics.

Using Acoustic Fields to Fabricate ECM-Based Biomaterials for Regenerative Medicine Applications

“…First, vascular sprouting and collagen fiber alignment along the direction of vessel growth have been observed in acoustically-patterned systems containing vascular endothelial cells [75], consistent with observations of sprouting angiogenesis from microvessel explants [161]. Likewise, enhanced contraction of acousticallypatterned constructs has been reported in several studies [74,80]. Critically, the direction of cell alignment can be directed to develop either parallel or perpendicular to the acoustic exposure axis by altering the orientation in which the engineered tissue is anchored [80].…”

“…The use of ultrasound to non-invasively pattern cells is of growing interest to tissue engineering, as spatial cues such as relative position, spacing, and density of cells serve as important determinants of cellular behavior [71][72][73]. USWF exposures have been used to pattern a variety of cell types, including fibroblasts, endothelial cells, Schwann cells, and myocytes to produce enhanced collagen gel contraction [74], vascular network formation [75][76][77], and cellular alignment [78][79][80][81], respectively.…”

Immunogold FIB‐SEM: Combining Volumetric Ultrastructure Visualization with 3D Biomolecular Analysis to Dissect Cell–Environment Interactions

“…Murine myoblasts (C2C12 line) grown on the surface of gelatin methacryloyl (GelMA) hydrogel substrates were differentiated into myotubes using a 7 day culture in low-serum myogenic media supplemented with insulin-like growth factor-1. [30] Undifferentiated myoblasts (day 0) and differentiated myotubes (day 7) were fixed and labeled with NPC-specific primary antibody and immunogold secondary antibody. Successful immunolabeling was confirmed using confocal fluorescence microscopy, which also revealed a relatively higher level of NPCs in myotubes compared to myoblasts (Figure 4a; Figure S2, Supporting Information).…”

Fabrication and Mechanical Properties of Engineered Protein‐Based Adhesives and Fibers

“…), endow them to form different secondary structures, such as α‐helices and β‐sheets . The diversity of protein structures results in different mechanical properties of their resulting biomaterials . With the technology of genetic engineering, many efforts have been made to investigate the relationships between the structure and mechanical functions of recombinant proteins .…”