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Abstract
Low oxygen and mechanical loading may play roles in regulating the fibrocartilaginous phenotype of the human inner meniscus, but their combination in engineered tissues remains unstudied. Here, we investigated how continuous low oxygen ("hypoxia") combined with dynamic compression would affect the fibrocartilaginous "inner meniscus-like" matrix-forming phenotype of human meniscus fibrochondrocytes (MFCs) in a porous type I collagen scaffold. Freshly-seeded MFC scaffolds were cultured for 4 weeks in either 3 or 20% O2 or pre-cultured for 2 weeks in 3% O2 and then dynamically compressed for 2 weeks (10% strain, 1 Hz, 1 h/day, 5 days/week), all with or without TGF-β3 supplementation. TGF-β3 supplementation was found necessary to induce matrix formation by MFCs in the collagen scaffold regardless of oxygen tension and application of the dynamic compression loading regime. Neither hypoxia under static culture nor hypoxia combined with dynamic compression had significant effects on expression of specific protein and mRNA markers for the fibrocartilaginous matrix-forming phenotype. Mechanical properties significantly increased over the two-week loading period but were not different between static and dynamic-loaded tissues after the loading period. These findings indicate that 3% O2 applied immediately after scaffold seeding and dynamic compression to 10% strain do not affect the fibrocartilaginous matrix-forming phenotype of human MFCs in this type I collagen scaffold. It is possible that a delayed hypoxia treatment and an optimized pre-culture period and loading regime combination would have led to different outcomes.
Highlights
The menisci play important mechanical roles in the knee [1, 2]
We showed that meniscus fibrochondrocytes (MFCs) expanded with the basal medium (BM)+T1F2 had faster proliferation rates and comparable subsequent matrix-forming capacity to those expanded in BM alone and BM+FGF-2
Hypoxic compared to normoxic culture with TGF-β3 supplementation did not enhance the fibrocartilaginous matrix-forming phenotype. This contrasts with the synergistic increase in matrix formation by continuous hypoxia and TGF-β3 in the pellet model [24] and the hypoxia-mediated enhanced matrix formation in a polymeric scaffold layered with a biomimetic surface [20] by human meniscus-derived cells
Summary
The menisci play important mechanical roles in the knee [1, 2]. In humans, their weight-bearing inner region develops a fibrocartilaginous phenotype throughout life with morphological differences from the outer region (Table 1) [3,4,5,6,7,8,9]. Low strain dynamic compression under hypoxia of human tissue engineered meniscus
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