gms | German Medical Science

Objectives: In vitro chondrogenesis of human mesenchymal stromal cells (hMSC) provides an effective model to recapitulate successive phases of embryonic chondrocyte maturation from progenitor cells to hypertrophic chondrocytes. Unfortunately, chondrogenesis of hMSC is still insufficiently understood, progresses along the endochondral pathway, and in vitro production of an articular chondrocyte phenotype remains a challenge. MicroRNAs (miRs) are small non-coding RNAs inhibiting protein production. A better understanding of their regulation during specific phases of chondrocyte maturation, especially during hypertrophic differentiation, offers new strategies to modulate differentiation outcome or to improve therapeutic strategies to prevent osteoarthritis (OA). Aim of the current work was to characterize miR regulation during well-defined in vitro induced chondrocyte maturation stages to extract cartilage-relevant and hypertrophy-related miR clusters.

Methods: hMSC were subjected to TGF-beta driven chondrogenesis in 3D-pellet culture for 42 days. At five defined time points miR profiles were determined by miRNA microarray analysis and results were evaluated via ANOVA and t-Test. Regulation of selected miRs was verified by qRT-PCR and compared between hypertrophic chondrocytes derived from hMSC and non-hypertrophic chondrocytes (hAC) from human articular cartilage.

Results and Conclusion: 169 miRs were significantly regulated during chondrogenic differentiation of hMSC. Hierarchical clustering revealed a distinct miR expression pattern representative for chondrocyte maturation stages and defined single miRs regulated between MSC, prechondrocytes, chondroblasts, chondrocytes, and hypertrophic chondrocytes. Regulation of miR-181 family members allowed discrimination of successive differentiation stages. Levels of several miRs, including miR-140, miR-181, and miR-210 positively correlated with successful chondrocyte formation. Seven miRs were differentially expressed between hypertrophic chondrocytes produced from hMSC compared to redifferentiated hAC. Overall, chondrocyte maturation-dependent miRs are promising candidates as novel diagnostic tools and targets of potential relevance for OA.