Cyclic freeze-thaw grinding to decellularize meniscus for fabricating porous, elastic scaffolds

Decellularized meniscus extracellular matrix (dmECM)-based biological scaffolds in the forms of sponge, hydrogel, nanofiber, and composite have gained increasing interest in meniscus tissue engineering and regeneration. A common shortcoming of those scaffolds is insufficient mechanical strength and poor elasticity. Herein, we report a practicable protocol for milder meniscus decellularization to prepare elastic, porous dmECM scaffolds. Porcine meniscus was pulverized by cyclic freeze-thaw grinding and then treated with DNase to obtain fine dmECM particles. Individual dmECM particles were condensed to bulk preparation by centrifuge, followed by lyophilization to form blocks, and finally crosslinked by dehydrothermal treatment to obtain porous dmECM scaffolds. Our results show that the freeze-thaw grinding method was effective in removing cellular DNA with good retention of meniscus-derived bioactive components. The dmECM scaffold had porous structure with interconnected mesopores and good mechanical properties. Primary articular chondrocytes proliferated robustly and maintained chondrogenic characteristics and produce abundant collagen on dmECM scaffolds. Evaluation of biocompatibility in a rat model shows that the dmECM scaffold elicited minor foreign body reactions, indicating effective antigen removal from dmECM. This study provides an alternative for preparing dmECM and fabricating porous scaffolds for meniscus repair and regeneration.