Multiphoton‐Guided Creation of Complex Organ‐Specific Microvasculature

Engineering functional human tissues in vitro is currently limited by difficulty replicating the small caliber, complex connectivity, cellularity, and 3D curvature of the native microvasculature. Multiphoton ablation has emerged as a promising technique for fabrication of microvascular structures with high resolution and full 3D control, but cellularization and perfusion of complex capillary‐scale structures has remained challenging. Here, multiphoton ablation combined with guided endothelial cell growth from pre‐formed microvessels is used to successfully create perfusable and cellularized organ‐specific microvascular structures at anatomic scale within collagen hydrogels. Fabrication and perfusion of model 3D pulmonary and renal microvascular beds is demonstrated, as is replication and perfusion of a brain microvascular unit derived from in vivo data. Successful endothelialization and blood perfusion of a kidney‐specific microvascular structure is achieved, using laser‐guided angiogenesis. Finally, proof‐of‐concept hierarchical blood vessels and complex multicellular models are created, using multistep patterning with multiphoton ablation techniques. These successes open new doors for the creation of engineered tissues and organ‐on‐a‐chip devices. Engineering functional tissues for regenerative medicine and human disease modeling is limited by difficulty re‐creating in vivo‐like microvasculature. This paper reports success combining multiphoton microscopy with directed angiogenesis to precisely generate perfusable 3D organ‐specific capillary beds within natural extracellular matrices. These methods enable full cellularization of complex capillary models and the creation of multicellular, heterogeneous, and hierarchical vasculature.