A Modular Bioinstructive Platform for Additive-Free, Topography-Driven Stem Cell Differentiation and Patterning

Recreating 3D bone formation in vitro without biochemical inducers remains a longstanding challenge in preclinical testing. We present a scalable, bioinstructive platform based on polylactic acid microparticles with controlled dimpled surface features that direct mesenchymal stem cell differentiation through endogenous topography-mediated mechanotransduction, establishing a mechanistically validated, additive-free platform. These 3D topographical cues drive cytoskeletal reorganisation and induce osteogenesis via canonical Hedgehog signalling. RNA-Seq revealed early significant upregulation of cytoskeletal components and osteochondral transcription factors, including runt-related transcription factor 2 (RUNX2) and SRY-box transcription factor 9 (SOX9), followed by activation of the insulin growth factor-II pathway and osteogenic commitment. To demonstrate translational potential, two-photon polymerisation lithography was employed to engineer precisely-patterned 3D topographies, inducing graded GLI1 expression without added soluble cues. This establishes a modular, versatile platform for stem cell engineering, offering a topography-driven, non-genetic analogue to mechanogenetics with broad utility for regenerative medicine and human-relevant development of bone models. O_FIG O_LINKSMALLFIG WIDTH=162 HEIGHT=200 SRC="FIGDIR/small/664383v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@1b06d7dorg.highwire.dtl.DTLVardef@1e464e1org.highwire.dtl.DTLVardef@19b1299org.highwire.dtl.DTLVardef@1e21f89_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG