Modular Biofabrication of Functional Skeletal Muscle Incorporated with Endothelialized Microvascular Seed‐Derived Lumenized Microvessels

ABSTRACT The clinical translation of engineered skeletal muscle (eSM) for volumetric muscle regeneration is hindered by the challenge of establishing a functional vascular network capable of sustaining its high metabolic demand and ensuring graft survival. Here, we present a bottom‐up biofabrication strategy to generate a pre‐vascularized in vitro eSM model through the modular assembly of independently matured muscle and vascular compartments. C2C12 myoblasts were encapsulated within core–shell fibers using rotary wet‐spinning (RoWS), yielding anisotropically aligned, multinucleated, and contractile myofibers expressing myosin heavy chain and sarcomeric α‐actinin. In parallel, gelatin methacryloyl (GelMA)‐based microvascular seeds (µVS), pre‐endothelialized with human umbilical vein endothelial cells, were engineered to guide rapid and structurally stable vascular formation while preventing uncontrolled capillary self‐organization. Fully endothelialized µVS were incorporated into a pro‐angiogenic bioink and processed via RoWS to generate tubular vascular fibers with physiological diameters (100–200 µm) and continuous CD31‐positive lumens. After independent maturation, muscle and vascular constructs were bioassembled into a hierarchically organized tissue and co‐cultured. By decoupling myogenic and angiogenic differentiation, this strategy overcomes medium incompatibility typical of conventional co‐cultures, preserving compartment‐specific architecture and function and establishing a versatile platform for muscle‐vascular modeling and translational muscle repair.