Abstract
Biomimetic bone tissue engineering strategies partially recapitulate development. We recently showed functional restoration of femoral defects using scaffold-free human mesenchymal stem cell (hMSC) condensates featuring localized morphogen presentation with delayed in vivo mechanical loading. Possible effects of construct geometry on healing outcome remain unclear. Here, we hypothesized that localized presentation of transforming growth factor (TGF)-β1 and bone morphogenetic protein (BMP)-2 to engineered hMSC tubes mimicking femoral diaphyses induces endochondral ossification, and that TGF-β1 + BMP-2-presenting hMSC tubes enhance defect healing with delayed in vivo loading vs. loosely packed hMSC sheets. Localized morphogen presentation stimulated chondrogenic priming/endochondral differentiation in vitro. Subcutaneously, hMSC tubes formed cartilage templates that underwent bony remodeling. Orthotopically, hMSC tubes stimulated more robust endochondral defect healing vs. hMSC sheets. Tissue resembling normal growth plate was observed with negligible ectopic bone. This study demonstrates interactions between hMSC condensation geometry, morphogen bioavailability, and mechanical cues to recapitulate development for biomimetic bone tissue engineering.
Highlights
Biomimetic bone tissue engineering strategies partially recapitulate development
Transcript analysis of key differentiation markers, normalized to controls without growth factors, revealed only minimally elevated mRNA expression of the chondrogenic genes sex-determining region Y-box 9 (SOX9), aggrecan (ACAN), and collagen type 2A1 (COL2A1), and the early osteogenic gene alkaline phosphatase (ALP) in bone morphogenetic protein (BMP)-2-loaded human mesenchymal stem cell (hMSC) tubes compared to transforming growth factor (TGF)-β1-presenting constructs (Fig. 2b)
These findings suggest that while hMSC tubes across all groups were phenotypically undifferentiated at day 8 of culture, in situ presentation of transforming growth factor-β1 (TGF-β1), bone morphogenetic protein-2 (BMP-2), or TGFβ1 + BMP-2 imparted robust chondrogenic lineage priming in a morphogen-dependent manner
Summary
Biomimetic bone tissue engineering strategies partially recapitulate development. We recently showed functional restoration of femoral defects using scaffold-free human mesenchymal stem cell (hMSC) condensates featuring localized morphogen presentation with delayed in vivo mechanical loading. Scaffold-free cartilage templates derived from self-assembled human MSC (hMSC) condensations have been shown to progress through in vivo endochondral ossification, contingent on in vitro morphogen priming[22,23,24,25,26,27,28] Limitations of this approach include (i) the necessity to pre-culture hMSC condensations for ≥3 weeks in an induction medium supplied with morphogens (e.g., transforming growth factor (TGF)-β1) to stimulate cartilage template formation, and (ii) and the inability to generate tubular tissue constructs. The incorporation of TGF-β1-presenting gelatin microspheres for in situ chondrogenic priming[32] and BMP-2-presenting mineral-coated hydroxyapatite microparticles to induce bony remodeling of the cartilaginous template[33] circumvents the need for lengthy predifferentiation and enables early in vivo implantation Using this approach, we recently demonstrated functional femoral bone defect healing via endochondral ossification with microparticlecontaining hMSC condensate sheets for a localized presentation of TGF-β134 or TGF-β1 + BMP-235 contingent on in vivo mechanical loading. The effects of implant geometry—mimicking the geometry of long bone diaphyses with a channel for nutrient transport and endogenous cell invasion—on the healing outcome have not been assessed far
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