Mechanically graded granular scaffolds for osteochondral tissue engineering

This study developed photoannealed polyethylene glycol granular scaffolds with a continuous stiffness gradient to model the mechanical transition of the osteochondral unit without introducing the sharp interfaces typical of biphasic scaffold designs. Using microfluidically generated PEG microgels of three diameters, the authors created macroporous scaffolds whose stiffness increased continuously across an 8 mm region from approximately 2–5 kPa to 50–60 kPa. Human mesenchymal stromal cells cultured in these constructs showed position-dependent responses to the local mechanical environment. Softer regions promoted rounded cell morphology, increased glycosaminoglycan-rich matrix deposition, and elevated chondrogenic-associated responses, while stiffer regions promoted cell elongation, greater cytoskeletal tension, increased alkaline phosphatase activity, mineral deposition, and osteogenic marker expression. Smaller and medium microgels amplified these regional effects relative to large microgels, indicating that scaffold architecture and available interfacial surface area influenced how cells interpreted the gradient. Inhibition of actomyosin contractility with blebbistatin reduced cytoskeletal organization and collapsed gradient-dependent transcriptional patterning, showing that tension-dependent mechanotransduction was required for MSC interpretation of the graded scaffold. Overall, the work establishes a single-material, macroporous gradient scaffold platform for studying mechanically regulated osteochondral tissue formation.