A developmentally inspired combined mechanical and biochemical signaling approach on zonal lineage commitment of mesenchymal stem cells in articular cartilage regeneration

Abstract

Articular cartilage is organized into multiple zones including superficial, middle and calcified zones with distinct cellular and extracellular components to impart lubrication, compressive strength, and rigidity for load transmission to bone, respectively. During native cartilage tissue development, changes in biochemical, mechanical, and cellular factors direct the formation of stratified structure of articular cartilage. The objective of this work was to investigate the effect of combined gradients in cell density, matrix stiffness, and zone-specific growth factors on the zonal organization of articular cartilage. Human mesenchymal stem cells (hMSCs) were encapsulated in acrylate-functionalized lactide-chain-extended polyethylene glycol (SPELA) gels simulating cell density and stiffness of the superficial, middle and calcified zones. The cell-encapsulated gels were cultivated in a medium supplemented with growth factors specific to each zone and the expression of zone-specific markers was measured with incubation time. Encapsulation of 60 × 10⁶ cells per mL hMSCs in a soft gel (80 kPa modulus) and cultivation with a combination of TGF-β1 (3 ng mL⁻¹) and BMP-7 (100 ng mL⁻¹) led to the expression of markers for the superficial zone. Conversely, encapsulation of 15 × 10⁶ cells per mL hMSCs in a stiff gel (320 MPa modulus) and cultivation with a combination of TGF-β1 (30 ng mL⁻¹) and hydroxyapatite (3%) led to the expression of markers for the calcified zone. Further, encapsulation of 20 × 10⁶ cells per mL hMSCs in a gel with 2.1 MPa modulus and cultivation with a combination of TGF-β1 (30 ng mL⁻¹) and IGF-1 (100 ng mL⁻¹) led to up-regulation of the middle zone markers. Results demonstrate that a developmental approach with gradients in cell density, matrix stiffness, and zone-specific growth factors can potentially regenerate zonal structure of the articular cartilage.