Multi-directional cellular alignment in 3D guided by electrohydrodynamically-printed microlattices

Abstract

Recapitulating aligned cellular architectures of native tissues in vitro is important to engineer artificial tissue analogs with desired biological functions. Here a novel strategy is presented to direct three-dimensional (3D) cellular alignment by embedding cell/collagen hydrogel into the predefined electrohydrodynamically-printed microlattices. The cell/collagen hydrogel, originally filled within the printed microlattices uniformly, was found to gradually develop into densely-populated and highly-aligned bands along the longitudinal direction of the printed microlattices. The cellular alignment was highly dependent on the height, spacing and orientation of the microlattices. The presented method was applicable to multiple cell types including primary cardiomyocytes and the gaps formed between the aligned bands and the lateral walls of the microlattice facilitated the subsequent seeding and rapid alignment of other cell types which enables to engineer anisotropic multicellular tissue constructs. The engineered cardiac patches expressed mature cardiomyocyte-specific phenotypes and exhibited synchronous contractive activities. Multilayer cellular alignment with varied orientation in 3D collagen hydrogel was successfully achieved by using electrohydrodynamically-printed microlattices with layer-specific orientations. This exploration offers a promising way to engineer complex 3D tissue constructs with predefined cellular alignments. Statement of significanceFabrication of biomimetic highly-aligned complex cellular architectures has a great significance to recapitulate the unique mechanical and physiological functions of the engineered tissues (e.g., heart tissue, neuron, muscle). Here, we introduced a novel strategy to direct 3D cellular alignment by embedding cell/collagen hydrogel into the predefined electrohydrodynamically-printed microlattices without any external stimuli. The microscopical study of the dynamic alignment process of cells and collagen fibers contributed to exploring the mechanism of autonomous formation of highly-aligned cellular bands. Multilayer cellular alignment with varied orientation in 3D collagen hydrogel was successfully achieved by using the microlattices with layer-specific orientations, which showed a promising way to engineer complex 3D tissue constructs with predefined cellular alignments.