Three-dimensional in vitro cell systems are a promising alternative to animals to study cardiac biology and disease. We have generated three-dimensional in vitro models of the human heart (“cardiac spheroids”, CSs) by co-culturing human primary or iPSC-derived cardiomyocytes, endothelial cells and fibroblasts at ratios approximating those present in vivo. The cellular organisation, extracellular matrix and microvascular network mimic human heart tissue. These spheroids have been employed to investigate the dose-limiting cardiotoxicity of the common anti-cancer drug doxorubicin. Viability/cytotoxicity assays indicate dose-dependent cytotoxic effects, which are inhibited by the nitric oxide synthase (NOS) inhibitor L-NIO, and genetic inhibition of endothelial NOS, implicating peroxynitrous acid as a key damaging agent. These data indicate that CSs mimic important features of human heart morphology, biochemistry and pharmacology in vitro, offering a promising alternative to animals and standard cell cultures with regard to mechanistic insights and prediction of toxic effects in human heart tissue.

Importance of the field Effective vascularization of thick three-dimensional engineered tissue constructs is a problem in tissue engineering. As in native organs, a tissue-engineered intra-organ vascular tree must be comprised of a network of hierarchically branched vascular segments. Despite this requirement, current tissue-engineering efforts are still focused predominantly on engineering either large-diameter macrovessels or microvascular networks. Areas covered in this review We present the emerging concept of organ printing or robotic additive biofabrication of an intra-organ branched vascular tree, based on the ability of vascular tissue spheroids to undergo self-assembly. What the reader will gain The feasibility and challenges of this robotic biofabrication approach to intra-organ vascularization for tissue engineering based on organ-printing technology using self-assembling vascular tissue spheroids including clinically relevantly vascular cell sources are analyzed. Take home message It is not possible to engineer 3D thick tissue or organ constructs without effective vascularization. An effective intra-organ vascular system cannot be built by the simple connection of large-diameter vessels and microvessels. Successful engineering of functional human organs suitable for surgical implantation will require concomitant engineering of a ‘built in’ intra-organ branched vascular system. Organ printing enables biofabrication of human organ constructs with a ‘built in’ intra-organ branched vascular tree.