(651e) Biomolecular Templated Assembly of Droplet-Derived Hydrogel Microtissues

Paracrine and autocrine cell signaling are critical factors guiding tissue development and maintenance, and dysregulation of these cues contributes to the pathogenesis of diseased states such as cancer. Patterning multiple cell types is thus a critical step for engineering functional tissue [1], but current “top-down” approaches such as dielectrophoresis and photopatterning are challenging to scale-up for the construction of mesoscale tissues. On the other hand, “bottom-up” methods wherein small tissue building blocks are assembled into larger structures have potential for constructing multicellular tissues in a facile, scalable fashion. Synthetic microtissues comprised of cell-laden hydrogels in this size range represent appropriate fundamental building blocks of such bottom-up methods [2].

To specify the placement of many different microtissues relative to one another, we have developed a “bottom-up” approach for fabricating multicellular tissue constructs that utilizes specific biomolecular interactions to template the assembly of 3D cell-laden hydrogel microtissues. A flow focusing-generated emulsion of photopolymerizable prepolymer is used to produce 100 µm monodisperse microtissues at rates of 30 Hz (10⁵/hr). Multiple cell types, including suspension and adherently cultured cells, can be encapsulated into the microtissues with high viability (~97%). We then use a surface encoding scheme to self-assemble microtissues “bottom-up” from a template that is defined using “top-down” techniques. The microtissues are derivatized with interactive functional groups using a biotin-streptavidin linkage to the polymer network, and are then assembled by specific binding onto patterned arrays. Using orthogonal interactions, we have achieved multiplexed patterning of multiple microtissue types with high binding efficiency and >90% patterning specificity. We have also demonstrated the ability to organize multicomponent constructs composed of epithelial and mesenchymal microtissues while preserving each cell type in a 3D microenvironment. The combination of high throughput microtissue generation with scalable surface-templated assembly offers the potential to dissect mechanisms of cell-cell interaction in three dimensions in healthy and diseased states as well as provide a framework for templated assembly of larger structures for implantation.

[1]        S. March, E. E. Hui, G. H. Underhill, S. Khetani, and S. N. Bhatia, "Microenvironmental regulation of the sinusoidal endothelial cell phenotype in vitro," Hepatology, vol. 50, pp. 920-8, 2009.

[2]        A. A. Chen, G. H. Underhill, and S. N. Bhatia, "Multiplexed, high-throughput analysis of 3D microtissue suspensions," Integr Biol (Camb), vol. 2, pp. 517-27, 2010.