Hypoxia and TGF-β3 Synergistically Mediate Inner Meniscus-Like Matrix Formation by Fibrochondrocytes

Abstract

Objective:

To investigate hypoxia and TGF-β3 effects on inner meniscus-like extracellular matrix (ECM) formation and related gene expression by meniscus fibrochondrocytes (MFCs).

Design:

Aggregates of human MFCs were cultured for 3 weeks under hypoxia (3% O₂) or normoxia (atmospheric O₂) with or without TGF-β3 supplementation (10 ng/mL). Inner meniscus-like ECM formation was assessed by biochemistry, histology, and immunofluorescence. mRNA expression of ECM-related genes, TGF-β isoforms 1–3, and hypoxia-inducible factor-1 (HIF-1) targets were assessed by quantitative real-time polymerase chain reaction.

Results:

Hypoxia and TGF-β3 supplementation synergistically induced inner meniscus-like ECM formation at the protein level with similar effects on ECM-related gene expression. Hypoxia alone did not induce an inner meniscus-like ECM-forming response nor upregulate mRNA of TGF-β isoforms. Expression of HIF-1α and HIF-1 target genes suggested that HIF-1 was a possible contributor to the observed synergistic interactions of hypoxia and TGF-β3 supplementation.

Conclusion:

Hypoxia and TGF-β3 supplementation synergistically induced inner meniscus ECM formation by adult human MFCs. Hypoxia alone is insufficient to induce an inner meniscus ECM-forming response in this culture model.

Impact Statement

The interactions of hypoxia and TGF-β3 in aggregates of human meniscus fibrochondrocytes are synergistic in nature, suggesting combinatorial strategies using these factors are promising for tissue engineering the inner meniscus regions. Hypoxia alone in the absence of TGF-β supplementation may be insufficient to initiate an inner meniscus-like extracellular matrix-forming response in this model.