Osteochondral Interface Tissue Engineering Using Macroscopic Gradients of Bioactive Signals

Abstract

Continuous gradients exist at osteochondral interfaces, which may be engineered by applying spatially patterned gradients of biological cues. In the present study, a protein-loaded microsphere-based scaffold fabrication strategy was applied to achieve spatially and temporally controlled delivery of bioactive signals in three-dimensional (3D) tissue engineering scaffolds. Bone morphogenetic protein-2 and transforming growth factor-β1-loaded poly(d,llactic- co-glycolic acid) microspheres were utilized with a gradient scaffold fabrication technology to produce microsphere-based scaffolds containing opposing gradients of these signals. Constructs were then seeded with human bone marrow stromal cells (hBMSCs) or human umbilical cord mesenchymal stromal cells (hUCMSCs), and osteochondral tissue regeneration was assessed in gradient scaffolds and compared to multiple control groups. Following a 6-week cell culture, the gradient scaffolds produced regionalized extracellular matrix, and outperformed the blank control scaffolds in cell number, glycosaminoglycan production, collagen content, alkaline phosphatase activity, and in some instances, gene expression of major osteogenic and chondrogenic markers. These results suggest that engineered signal gradients may be beneficial for osteochondral tissue engineering.

Osteogenic Differentiation of Human Bone Marrow Stromal Cells in Hydroxyapatite-Loaded Microsphere-Based Scaffolds

“…[18][19][20][21][22][23][24] Although microspheres have been used in tissue engineering for stratified designs, [25][26][27] the use of continuously graded microsphere-based scaffolds is still limited. 28,29 The combination of continuously graded microsphere-only scaffolds with mineral incorporation, which provides a continuous gradient in macroscopic material composition, has previously been demonstrated as a method, 30 but the long-term response of cells in this environment has not been investigated. To address osteoblastic cell response, the most logical step is to evaluate the 3-D performance of homogenous microsphere-based scaffolds with mineral incorporation before using a continuously-graded design, as in the current study.…”

“…The advantages of this approach relate to the ability to tightly control a gradient in physical or chemical properties across the length of the scaffold by using each single microsphere as a ''building block.'' The current study, for the first time, used a precision particle fabrication technology 28,[30][31][32] to make poly(d,l-lactic acid-co-glycolic acid) (PLGA) microspheres loaded with HAp nanoparticles. The microspheres were used to construct 3-D scaffolds with various HAp contents on which human bone marrow stromal cells (hBMSC) were osteogenically differentiated for 6 weeks.…”

Microsphere-Based Gradient Implants for Osteochondral Regeneration: a Long-Term Study in Sheep

“…Two different batches of monodisperse microspheres were prepared using the aforementioned emulsions of PLGA-CS-NaHCO 3 and PLGA-β-TCP according to our previously reported technology [12][13][14]. Briefly, uniform polymer droplets were created by introducing regular jet instabilities in the polymer stream through acoustic excitation via an ultrasonic transducer.…”

“…Microsphere-based gradient implants for osteochondral regeneration: a long-term study in sheep Research Article Fabrication of gradient scaffolds Gradient scaffolds were prepared using our previously reported technology [12][13][14][15]. Briefly, 50 mg of lyophilized chondrogenic microspheres and 100 mg of osteogenic microspheres were dispersed in DI water, and separately loaded into two syringes.…”

Controlled Release Strategies for Bone, Cartilage, and Osteochondral Engineering—Part II: Challenges on the Evolution from Single to Multiple Bioactive Factor Delivery

“…Dormer et al 175 designed PLGA microsphere-sintered constructs incorporated with gradients of BMP-2 and TGF-b1, and despite obtaining promising data, acknowledged the difficulty in sustaining the gradient patterns for long term in vitro due to the quick diffusion of the molecules. Re'em et al 176 designed a bilayered system, spatially presenting the chondroinductive TGF-b1 in one layer and the osteoinductive BMP-4 in a second layer via affinity binding to the matrix.…”