High‐Resolution and Real‐Time In Situ Generation of Cellular Spheroids by Laser‐Assisted Bioprinting for Guided Microvascularization

ABSTRACT Replicating the native cellular density of tissues remains a critical bottleneck in bioprinting, as most techniques yield local concentrations orders of magnitude lower than physiological levels. Here, we report Micro‐Injection Laser‐Assisted Bioprinting (MI‐LAB), a laser‐assisted strategy that uniquely integrates in situ microspheroid formation with micrometer‐scale deposition precision. Unlike conventional laser‐assisted bioprinting (LAB), which relies on pre‐fabricated spheroids, MI‐LAB dynamically generates cellular aggregates during printing, achieving features as small as 30 µm while reaching local cell concentrations in the range of 100 million cells/mL, comparable to native tissues. This dual capability bridges the gap between single‐cell placement and multicellular assembly. As a proof of concept, we fabricated prevascularized constructs by sequentially depositing human umbilical vein endothelial cell aggregates within a methacrylated gelatin/collagen (GelMA/ColMA) matrix containing human skin fibroblasts. The engineered tissues exhibited guided vasculogenic sprouting and robust endothelial marker expression, validating the maturation of pre‐microvascular networks. By enabling programmable microspheroid placement at native‐like densities with microscale precision, MI‐LAB establishes a disruptive biofabrication paradigm for pre‐microvascularized tissues while offering a broadly applicable strategy for high‐density, biomimetic tissue engineering.