Biomaterial-based endochondral bone regeneration: a shift from traditional tissue engineering paradigms to developmentally inspired strategies

Abstract

There is an urgent, clinical need for an alternative to the use of autologous grafts for the ever increasing number of bone grafting procedures performed annually. Herein, we describe a developmentally inspired approach to bone tissue engineering, which focuses on leveraging biomaterials as platforms for recapitulating the process of endochondral ossification. To begin, we describe the traditional biomaterial-based approaches to tissue engineering that have been investigated as methods to promote in vivo bone regeneration, including the use of three-dimensional biomimetic scaffolds, the delivery of growth factors and recombinant proteins, and the in vitro engineering of mineralized bone-like tissue. Thereafter, we suggest that some of the hurdles encountered by these traditional tissue engineering approaches may be circumvented by modulating the endochondral route to bone repair and, to that end, we assess various biomaterials that can be used in combination with cells and signaling factors to engineer hypertrophic cartilaginous grafts capable of promoting endochondral bone formation. Finally, we examine the emerging trends in biomaterial-based approaches to endochondral bone regeneration, such as the engineering of anatomically shaped templates for bone and osteochondral tissue engineering, the fabrication of mechanically reinforced constructs using emerging three-dimensional bioprinting techniques, and the generation of gene-activated scaffolds, which may accelerate the field towards its ultimate goal of clinically successful bone organ regeneration.

Keywords: biochemical factors; endochondral ossification; hypertrophy; mesenchymal stem cell; oxygen tension; scaffold.

Fig. 1

Schematic illustrating combinations of biomaterials, cells, and signaling factors for the tissue engineering of hypertrophic cartilaginous grafts in vitro. Signaling factors such as soluble biochemical factors, mechanical stimuli, and oxygen tension can be harnessed to direct chondrogenesis and subsequent hypertrophy of cells such as chondrocytes and MSCs. Biomaterial scaffolds fabricated from natural polymers, synthetic polymers, and/or ceramics facilitate the upscaling of tissue-engineered grafts to clinically relevant sizes and can be leveraged to further guide cells down the endochondral pathway. The resultant engineered tissue should contain hypertrophic chondrocytes, to promote vascularization upon implantation, as well as the key extracellular matrix components of cartilage and hypertrophic cartilage, such as proteoglycans and collagens types II and X.

Fig. 2

Assessment of repair of rat cranial bone defects by osteogenically primed tissue-engineered constructs compared to cell-free scaffolds. Tissue-engineered constructs were formed by seeding collagen/calcium phosphate scaffolds with bone marrow–derived MSCs and culturing the constructs in an osteogenic medium for 4 weeks prior to implantation. (a) Micro–computed tomography (μCT) image of the cell-free collagen/calcium phosphate scaffold showing good levels of healing 4 weeks after implantation in comparison to (b) μCT the tissue-engineered construct which showed limited levels of repair. (c) Low magnification image of the tissue-engineered construct (stained with hematoxylin and eosin) showing the formation of a dense capsule around the periphery of the construct which at higher magnification (d) shows the original host bone adjacent to the defect site (white arrow) in comparison to the dense layer of tissue around the periphery of the implanted construct (red arrow) which has resulted the formation (black arrow) of a necrotic area in the center of the implanted construct. Modified with permission from Ref. .

Fig. 3

Schematic illustration of hypothesized biomaterial-based endochondral bone regeneration in vivo. (a) Tissue-engineered hypertrophic cartilaginous grafts are implanted into the bone defect site. The ability of chondrogenic cells to survive in avascular environments maintains the viability of the engineered tissue during this initial hypoxic phase. (b) VEGF released by hypertrophic chondrocytes within the engineered tissue promotes the invasion of blood vessels. The cartilaginous matrix begins to degrade and osteoclasts and osteoblast begin to remodel the engineered tissue into bone. (c) Blood vessels, osteoclasts, and osteoblasts encroach further into the defect site promoting remodeling of any remaining cartilaginous matrix into bone. (d) The vascular network and bone marrow cavity is fully restored, and the bone is healed. VEGF, vascular endothelial growth factor.

Fig. 4

Anatomically shaped phalanx constructs, consisting of an osseous component comprising an MSC-encapsulated alginate hydrogel and a chondral component comprising self-assembled chondrocytes, generated through spatial regulation of endochondral ossification. (a) Macroscopic image demonstrating a vascularized osseous component and an integrated chondral component which was not vascularized. (b) Picrosirius red staining for collagen. (c) Aldehyde fuchsin/alcian blue staining for sulfated glycosaminoglycan. Insets show collagen type II (top), collagen type X (center), and collagen type I (bottom staining). (d) H&E staining. Arrows indicate blood vessel structures. (e) Micro–computed tomography image of the whole construct. (f) Micro–computed tomography image of the center section of construct. Main image scale bars are 2 mm. Inset scale bars in (c) are 250 μm. Inset scale bars in (d) are 100 μm. Reproduced with permission from Ref. . MSC, mesenchymal stem cell.