Restoring the Chondron: Pericellular Matrix Reconstitution Enhances Mechano-Inflammatory Resilience and Modulates Chondrocyte-Neuron Crosstalk

Abstract Chondrocytes in native articular cartilage are enclosed within a collagen VI (COL VI)-rich pericellular matrix (PCM), forming functional units known as chondrons. However, enzymatic isolation disrupts the PCM, and the functional consequences of its loss and restoration remain poorly understood. In this study, primary human chondrocytes were cultured in alginate beads to promote PCM reconstitution and subsequently recovered as reconstituted chondrons. Chondrocytes and reconstituted chondrons were encapsulated in gelatin methacryloyl hydrogels and compared under inflammatory and mechanical stimulation. Alginate preconditioning generated chondron-like units with a distinct COL VI-positive PCM that was retained after transfer to three-dimensional culture. Under inflammatory conditions, reconstituted chondrons exhibited reduced inflammatory, catabolic, neuroinflammatory, and angiogenic responses compared with isolated chondrocytes at both gene and protein levels. Under interleukin-1{beta} stimulation, mechanical loading further increased inflammatory gene expression in chondrocytes in a donor-dependent manner, whereas responses remained comparatively limited in reconstituted chondrons. Conditioned medium from reconstituted chondrons was also associated with lower capsaicin- and potassium chloride-evoked calcium responses in human induced pluripotent stem cell-derived sensory neurons than corresponding chondrocyte-conditioned medium. These findings demonstrate that reconstitution of a COL VI-rich PCM restores a chondron-like pericellular microenvironment that attenuates inflammatory activation and buffers load-associated inflammatory amplification, while exploratory sensory neuron experiments suggest downstream modulation of neuronal responsiveness. PCM restoration may therefore provide a promising strategy for cartilage tissue engineering and regenerative applications.