Orthogonally induced differentiation of stem cells for the programmatic patterning of vascularized organoids and bioprinted tissues.

The generation of organoids and tissues with programmable cellular complexity, architecture and function would benefit from the simultaneous differentiation of human induced pluripotent stem cells (hiPSCs) into divergent cell types. Yet differentiation protocols for the overexpression of specific transcription factors typically produce a single cell type. Here we show that patterned organoids and bioprinted tissues with controlled composition and organization can be generated by simultaneously co-differentiating hiPSCs into distinct cell types via the forced overexpression of transcription factors, independently of culture-media composition. Specifically, we used such orthogonally induced differentiation to generate endothelial cells and neurons from hiPSCs in a one-pot system containing either neural or endothelial stem-cell-specifying media, and to produce vascularized and patterned cortical organoids within days by aggregating inducible-transcription-factor and wild-type hiPSCs into randomly pooled or multicore-shell embryoid bodies. Moreover, by leveraging multimaterial bioprinting of hiPSC inks without extracellular matrix, we generated patterned neural tissues with layered regions composed of neural stem cells, endothelium and neurons. Orthogonally induced differentiation of stem cells may facilitate the fabrication of engineered tissues for biomedical applications.

Biomanufacturing of organ-specific tissues with high cellular density and embedded vascular channels.

Engineering organ-specific tissues for therapeutic applications is a grand challenge, requiring the fabrication and maintenance of densely cellular constructs composed of ~108 cells/ml. Organ building blocks (OBBs) composed of patient-specific-induced pluripotent stem cell-derived organoids offer a pathway to achieving tissues with the requisite cellular density, microarchitecture, and function. However, to date, scant attention has been devoted to their assembly into 3D tissue constructs. Here, we report a biomanufacturing method for assembling hundreds of thousands of these OBBs into living matrices with high cellular density into which perfusable vascular channels are introduced via embedded three-dimensional bioprinting. The OBB matrices exhibit the desired self-healing, viscoplastic behavior required for sacrificial writing into functional tissue (SWIFT). As an exemplar, we created a perfusable cardiac tissue that fuses and beats synchronously over a 7-day period. Our SWIFT biomanufacturing method enables the rapid assembly of perfusable patient- and organ-specific tissues at therapeutic scales.