The three-dimensional (3D) bioprinting technology provides programmable and customizable platforms to engineer cell-laden constructs mimicing human tissues for a wide range of biomedical applications. However, the encapsulated cells are often restricted in spreading and proliferation by dense biomaterial networks from gelation of bioinks. Herein, we report a novel cell-benign approach to directly bioprint porous-structured hydrogel constructs by using an aqueous two-phase emulsion bioink. The bioink, which contains two immiscible aqueous phases of cell/gelatin methacryloyl (GelMA) mixture and poly(ethylene oxide) (PEO), is photocrosslinked to fabricate predesigned cell-laden hydrogel constructs by extrusion bioprinting or digital micromirror device-based stereolithographic bioprinting. Porous structure of the 3D-bioprinted hydrogel construct is formed by subsequently removing the PEO phase from the photocrosslinked GelMA hydrogel. Three different cells (human hepatocellular carcinoma cells, human umbilical endothelial cells, and NIH/3T3 mouse embryonic fibroblasts) within the 3D-bioprinted porous cell-laden hydrogel patterns showed enhanced cell viability, spreading, and proliferation compared to the standard (i.e. non-porous) hydrogel constructs. The new 3D bioprinting strategy is believed to provide a robust and versatile platform to engineer porous-structured tissue constructs and their models for a variety of applications in tissue engineering, regenerative medicine, and personalized therapeutics.

Despite advances in the bioprinting technology, biofabrication of circumferentially multilayered tubular tissues or organs with cellular heterogeneity, such as blood vessels, trachea, intestine, colon, ureter, and urethra, remains a challenge. Herein, a promising multichannel coaxial extrusion system (MCCES) for microfluidic bioprinting of circumferentially multilayered tubular tissues in a single step, using customized bioinks constituting gelatin methacryloyl, alginate, and eight-arm poly(ethylene glycol) acrylate with a tripentaerythritol core, is presented. These perfusable cannular constructs can be continuously tuned up from monolayer to triple layers at regular intervals across the length of a bioprinted tube. Using customized bioink and MCCES, bioprinting of several tubular tissue constructs using relevant cell types with adequate biofunctionality including cell viability, proliferation, and differentiation is demonstrated. Specifically, cannular urothelial tissue constructs are bioprinted, using human urothelial cells and human bladder smooth muscle cells, as well as vascular tissue constructs, using human umbilical vein endothelial cells and human smooth muscle cells. These bioprinted cannular tissues can be actively perfused with fluids and nutrients to promote growth and proliferation of the embedded cell types. The fabrication of such tunable and perfusable circumferentially multilayered tissues represents a fundamental step toward creating human cannular tissues.