Professor • Hudson Moore Professor • Biomedical, Mechanics of Materials, Materials Paul M. Rady Department of Mechanical Engineering, University of Colorado, Boulder, United States
Introduction Osteochondral implants are promising treatment strategies for treating articular cartilage defects that lead to osteoarthritis. However, scaffolds used for cartilage regeneration are often too soft to support in vivo loads, leading to degradation of the surrounding tissue. Additive manufacturing techniques like bio-ink printing have emerged to reinforce soft scaffolds, however, these techniques have limited design freedom and resolution. In contrast, digital light processing (DLP) is a layer-by-layer printing approach where custom-built systems permit the use of any resin viscosity, a major benefit over bioprinting, over-projection stereolithography, and programmability of each print layer (i.e., thickness by controlling parameters including light intensity and exposure time. DLP thus establishes an ideal platform to fabricate architected micro-truss structures with sub-millimeter control over depth-dependent stiffness gradients while maintaining an open pore structure to enable cell migration. We present a 3D DLP printed composite scaffold that uses a unit-cell micro-truss with graded stiffness to reinforce a biomimetic hydrogel. The system is designed to control mechanical properties to both support physiological loads and guide cell mechanobiological responses. We hypothesize that incorporating a stiff structure into a soft gel scaffold will prevent the degradation of adjacent native cartilage tissue as compared to a soft gel alone after implantation in within an osteochondral defect within a swine knee.
Materials & Methods We designed and fabricated unit-cell based micro-truss structures to possess spatially varying geometry and controlled stiffness gradients. Using a custom digital light processing system and diacrylate-based resin with a dark light absorber and thermal post cure, we designed and fabricated three micro-truss designs with feature sizes < 200 μm: 1) uniform structure with 1 MPa structural modulus designed to match equilibrium modulus of healthy articular cartilage, 2) E = 1 MPa gradient structure designed to vary strain with depth, and 3) osteochondral bilayer with distinct cartilage (E = 1 MPa) and bone (E = 7-10 MPa) layers. Finite element models (FEM) predicted structural stiffness of the printed scaffold and strains within the infilling hydrogel. Empty and hydrogel-infilled composite trusses were stained with iodine for contrast and subjected to uniaxial compression during imaging using X-ray microscopy (XRM) to experimentally evaluate for the structural stiffness of the cartilage and bone layers. Next, the 3D printed structure was infilled with soft biomimetic hydrogels which were formulated with extracellular matrix analogs and tethered growth factors as biochemical cues to promote guide chondrogenesis in the cartilage layer and osteogenesis in the bone layer of the scaffold. Composite scaffolds were implanted into osteochondral defects in the knee of 12-week-old Yorkshire pigs for six weeks, followed by histological assessment and micromechanical (indentation) testing to evaluate properties of surrounding cartilage.
Results, Conclusions, Discussion This study demonstrated a 3D printed osteochondral scaffold that captured the ability to reproduce tissue-specific strains under a physiological compressive load. FEM was used to iteratively design a an octet truss, unit-cell micro-truss structure with high (80%) porosity that bore high (10%) strains while matching cartilage stiffness (E = 1 MPa). We demonstrated the ability to control stiffness, and strain, gradients in cartilage that could be further refined to emulate the functions of zonal cartilage. Our DLP system demonstrated that ability to accurately print large (millimeters) structures with small ( < 100’s of micrometers) feature sizes. By spatially grading the design of the printed structures, we generated controlled gradients in stiffness that produced strain distributions matching that of articular cartilage under uniaxial compression. We also generated a bilayer osteochondral design that combined a cartilage-mimetic micro-truss structure with a ~6× stiffer bone layer. FEM predicted cartilage layer structural stiffness at ~1 MPa and bone layer at ~9 MPa. Displacement tracking while compressing the hydrogel infilled structures during XRM imaging enabled calculation of structural stiffnesses of ~1 MPa and ~6.4 MPa, respectively. We further demonstrated improvements in cartilage properties adjacent to bilayer structures that were infilled with biomimetic hydrogels after six weeks of implantation in osteochondral defects. Histological assessment demonstrated other positive outcomes including robust bone formation in the bone, and not the cartilage, layer. Future studies will address the need to limit implant subsidence, better encourage hyaline articular cartilage and not fibrocartilage within the cartilage layer of the bilayer scaffolds, and develop a fully degradable printed material that is removed as new, load-bearing osteochondral tissue is formed.