Dual-Responsive Hydrogels Engineer Anisotropic Cellular Microenvironment to Modulate Stem Cell Organization and Fate

Biophysical and biochemical cues in the local cellular microenvironment, including topography, matrix mechanics, and growth factors, significantly regulate stem cell fate. However, strategies for in vitro replicating such complex organized three-dimensional (3D) cellular microenvironments and modulating cell alignment and differentiation by these cues in cell-laden hydrogels are far less developed. This study introduces light-responsive collagen peptide hydrogels mixed with magnetic nanoparticles as physical crosslinkers, leading to in situ formation of an organized network of human bone marrow mesenchymal stem cells (hMSCs) under magnetic-driven anisotropy. Moreover, by simply tailoring the nanoparticle surface with dopamine methacrylamide, chemical nanoparticle crosslinkers with dual magnetic-light-responsiveness are developed to tune matrix mechanical dynamics without significantly changing hydrogel stiffness and components. The encapsulated hMSCs exhibit enhanced spreading, alignment, and differentiation into ligamentocytes/tenocytes in anisotropic hydrogels with faster mechanical dynamics and transforming growth factor beta-3, contributing to the synergistic effects of these biophysical and biochemical cues. These simple manufacturing and conditioning strategies, which directly incorporate stimuli-responsive nanoparticle crosslinkers into cell-laden hydrogels, show great potential in developing advanced 3D organized in vitro models to modulate stem cell organization and fate.