Spatial mechanopriming of multicellular interfaces drives cytoskeletal-to-nuclear remodeling to boost therapeutic efficacy of stem cells

Engineering multicellular constructs through biophysical cues, independent of soluble biochemical factors, is a promising strategy to boost the therapeutic potential of stem cells for regenerative medicine. However, the mechanisms by which spatially organized cell-cell and cell-matrix interfaces govern multicellular signaling and subsequent behavioral outcomes remain poorly understood. Here, we elucidate these mechanistic events by introducing 2D nanoplates (∼350 nm) compartmentalized within the intercellular space of mesenchymal stem cell (MSC) spheroids to form de novo cell-matrix spatial interfaces. These interfaces redistribute internal stress across the spheroid, amplifying integrin-cytoskeleton-nucleus force transmission and restoring YAP-dependent mechanotransduction to achieve spatial mechanopriming. Cells within these mechanoprimed spheroids exhibit profound nuclear deformation, epigenetic remodeling, and spatially patterned metabolic activation. Finite element modeling and spatial transcriptomic analyses further reveal a spheroid-wide reorganization of cytoskeletal and transcriptional networks. Mechanistically, elevated cytoskeletal tension across the reorganized interfaces is associated with a putative, tension-associated pseudo-hypoxic signature involving HIF-1α-related signaling, establishing a mechanometabolic niche that robustly enhances angiogenic secretion. Consequently, mechanoprimed spheroids demonstrate superior engraftment, accelerated vascular regeneration, and improved perfusion recovery in a murine ischemia model. Collectively, this work establishes spatial mechanopriming as an effective strategy to remodel cytoskeletal-to-nuclear mechanics and enhance the therapeutic potential of 3D multicellular systems.