Abstract
The creation of engineered 3D microtissues has attracted prodigious interest because of the fact that this microtissue structure is able to mimic in vivo environments. Such microtissues can be applied extensively in the fields of regenerative medicine and tissue engineering, as well as in drug and toxicity screening. Here, we develop a novel method of fabricating a large number of dense honeycomb concave microwells via surface tension-mediated self-construction. More specifically, in order to control the curvature and shape of the concavity in a precise and reproducible manner, a custom-made jig system was designed and fabricated. By applying a pre-set force using the jig system, the shape of the honeycomb concave well was precisely and uniformly controlled, despite the fact that wells were densely packed. The thin wall between the honeycomb wells enables the minimization of cell loss during the cell-seeding process. To evaluate the performance of the honeycomb microwell array, rat hepatocytes were seeded, and spheroids were successfully formed with uniform shape and size. Liver-specific functions such as albumin secretion and cytochrome P450 were subsequently analyzed. The proposed method of fabricating honeycomb concave wells is cost-effective, simple, and reproducible. The honeycomb well array can produce multiple spheroids with minimal cell loss, and can lead to significant contributions in tissue engineering and organ regeneration.