Consistent self-organized emergence of hyaline cartilage in hiPSC-derived multi-tissue organoids

ABSTRACT: Existing protocols for in vitro hyaline cartilage production utilizing human induced pluripotent stem cells (hiPSC) have a number of challenges including a complex culturing process that uses undefined culture media, phenotypic instability, and batch-to-batch variability of the cell product. Here, we describe a simple, xeno- and feeder-free protocol for the generation of human hyaline cartilage utilizing multi-tissue organoids (MTOs). We investigate gene regulatory networks during hiPSC-MTO differentiation using RNA sequencing and bioinformatic analyses as well as histological and immunohistochemical methods. Interplays between BMPs and neural FGF pathways associated with the phenotype transition of MTOs are described. Comparisons across transcriptomes reveal that expression of chondrocyte-specific genes in MTOs correlates strongly with fetal lower-limb chondrocytes. Single-cell RNA-sequencing findings confirm that the majority of cells belong to the chondrogenic lineage, and that they are similar across MTO batches suggesting uniformity of the culture process. Collectively, these findings demonstrate consistent emergence of hyaline cartilage in MTOs and the molecular pathways that govern this process, thereby establishing an accessible source of functional chondrocytes for future therapeutic evaluation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE276142 | GEO | 2025/08/01

REPOSITORIES: GEO