In situ self-assembled organoid for osteochondral tissue regeneration with dual functional units

The regeneration of hierarchical osteochondral units is challenging due to difficulties in inducing spatial,directional and controllable differentiation of mesenchymal stem cells(MSCs)into cartilage and bone compartments.Emerging organoid technology offers new opportunities for osteochondral regeneration.In this study,we developed gelatin-based microcryogels customized using hyaluronic acid(HA)and hydroxyapatite(HYP),respectively for inducing cartilage and bone regeneration(denoted as CH-Microcryogels and OS-Microcryogels)through in vivo self-assembly into osteochondral organoids.The customized microcryogels showed good cytocompatibility and induced chondrogenic and osteogenic differentiation of MSCs,while also demonstrating the ability to self-assemble into osteochondral organoids with no delamination in the biphasic cartilage-bone structure.Analysis by mRNA-seq showed that CH-Microcryogels promoted chondrogenic differentiation and inhibited inflammation,while OS-Microcryogels facilitated osteogenic differentiation and suppressed the immune response,by regulating specific signaling pathways.Finally,the in vivo engraftment of pre-differentiated customized microcryogels into canine osteochondral defects resulted in the spontaneous assembly of an osteochondral unit,inducing simultaneous regeneration of both articular cartilage and subchondral bone.In conclusion,this novel approach for generating self-assembling osteochondral organoids utilizing tailor-made microcryogels presents a highly promising avenue for advancing the field of tissue engineering.

A low dose cell therapy system for treating osteoarthritis:In vivo study and in vitro mechanistic investigations

Mesenchymal stem cells(MSCs)can be effective in alleviating the progression of osteoarthritis(OA).However,low MSC retention and survival at the injection site frequently require high doses of cells and/or repeated injections,which are not economically viable and create additional risks of complications.In this study,we produced MSC-laden microcarriers in spinner flask culture as cell delivery vehicles.These microcarriers containing a low initial dose of MSCs administered through a single injection in a rat anterior cruciate ligament(ACL)transection model of OA achieved similar reparative effects as repeated high doses of MSCs,as evaluated through imaging and histological analyses.Mechanistic investigations were conducted using a co-culture model involving human primary chondrocytes grown in monolayer,together with MSCs grown either within 3D constructs or as a monolayer.Co-culture supernatants subjected to secretome analysis showed significant decrease of inflammatory factors in the 3D group.RNA-seq of co-cultured MSCs and chondrocytes using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis revealed processes relating to early chondrogenesis and increased extracellular matrix interactions in MSCs of the 3D group,as well as phenotypic maintenance in the co-cultured chondrocytes.The cell delivery platform we investigated may be effective in reducing the cell dose and injection frequency required for therapeutic applications.