Perspectives on the role of nanotechnology in bone tissue engineering

Abstract

Objective: This review surveys new developments in bone tissue engineering, specifically focusing on the promising role of nanotechnology and describes future avenues of research.

Methods: The review first reinforces the need to fabricate scaffolds with multi-dimensional hierarchies for improved mechanical integrity. Next, new advances to promote bioactivity by manipulating the nanolevel internal surfaces of scaffolds are examined followed by an evaluation of techniques using scaffolds as a vehicle for local drug delivery to promote bone regeneration/integration and methods of seeding cells into the scaffold.

Results: Through a review of the state of the field, critical questions are posed to guide future research toward producing materials and therapies to bring state-of-the-art technology to clinical settings.

Significance: The development of scaffolds for bone regeneration requires a material able to promote rapid bone formation while possessing sufficient strength to prevent fracture under physiological loads. Success in simultaneously achieving mechanical integrity and sufficient bioactivity with a single material has been limited. However, the use of new tools to manipulate and characterize matter down to the nano-scale may enable a new generation of bone scaffolds that will surpass the performance of autologous bone implants.

Figure 1

Bone regeneration requires three essential elements: osteoconductive matrix (scaffold), osteoconductive signals, osteogenic cells that can respond to these signals and an adequate blood supply [2]. The first step, fabrication of strong and porous scaffolds remains the Achilles' heel of the whole process. Natural composites or hybrid structures, such as bone and teeth, display properties that are invariably far superior to their individual constituent phases. The understanding of the mechanisms to achieve these remarkable properties has become far clearer in recent years, and consequently the notion of biomimicry has received much interest in the materials communities; however, resulting advances in new bone materials have been few if any, primarily due to the fact that such materials are difficult to fabricate. Fabrication alone, however, will not be enough to create an optimum scaffold. In this respect, nanotechnology provides new and useful tools to engineer the scaffold's internal surfaces and to create devices for drug delivery with carefully controlled spatial and temporal release patterns. Synthetic scaffolds can also serve as a vehicle for the delivery of cells to build new tissue. Different techniques have been proposed to successfully seed scaffolds with cells. They can be roughly divided into two main groups: attaching the cells to the internal scaffold surface, or distributing them in the scaffold porosity using a gel-like vehicle [14]. Injectable gels containing cells could also be used directly in non-load bearing applications. Seeding with skeletal stem cells has attracted much attention, but it is critical to develop the adequate chemical and physical extracellular milieu to promote differentiation towards the osteoblastic lineage. For example, it has been observed that the presence of calcium within the matrix favors osteogenic differentiation of the appropriate progenitor cell population [86].

Figure 2

Natural composites or hybrid structures, such as bone and teeth, display properties that are invariably far superior to their individual constituent phases. The origin of these remarkable properties derives from the deformation and fracture of its hierarchical structure, spanning molecular to macroscopic length-scales. The macro-scale arrangements of bones are either compact/cortical (dense material found at the surface of all bones) or spongy/cancellous (foam-like material whose struts are some 100-μm thick). Compact bone is composed of osteons surrounding and protecting blood vessels. Osteons have a lamellar structure, with each individual lamella composed of fibers arranged in geometrical patterns. These fibers are the result of several collagen fibrils, each linked by an organic phase to form fibril arrays. These mineralized collagen fibrils are the basic building blocks of bone, which are composed of collagen protein molecules (tropocollagen) formed from three chains of amino acids and nanocrystals of hydroxyapatite. Adapted from Reference [132], copyright © 1993, with permission from Macmillan Publishers Ltd.

Figure 3

Solid freeform fabrication techniques that are precise and reproducible, such as direct ink-jet printing, robotic-assisted deposition or robocasting (top of the figure), and hot-melt printing—which usually involve “building” structures layer-by-layer following a computer design, or image sources such as MRI—can be used to fabricate custom-designed scaffolds with complex architectures. Freeze casting, a technique that uses the microstructure of ice to template the architecture of ceramic scaffolds, can be used to produce porous lamellar materials that replicate the structure of the inorganic component of nacre at multiple-length scales [42, 133]. These materials can be much stronger than others with similar porosity described in the literature.

Figure 4

Urea-mediated solution mineralization of hydroxyapatite (HA) onto pHEMA hydrogel scaffolds. Thermo-decomposition of urea produces a gradual increase in pH, resulting in the hydrolysis of surface 2-hydroxyethyl esters and the precipitation of HA from the aqueous solution. The in situ generated surface carboxylates strongly interact with calcium ions and facilitate the heterogeneous nucleation and 2D growth of a high-affinity calcium-phosphate (CP) layer on the pHEMA surface. Prolonged mineralization allows for the growth of a thicker CP layer that covers the entire hydrogel surface. The SEM micrograph on the right shows the 2D circular outward growth of a calcium apatite layer from multiple nucleation sites on the acidic surface of pHEMA. The calcium phosphate layer did not delaminate even after Vickers indentation with a load of 5 N (bottom micrograph). Further functionalization of the hydrogel with either carboxylate or hydroxy ligands can be used to manipulate organic/inorganic interactions [55, 134]. Reprinted with permission from Reference [55], copyright © 2005, American Chemical Society.

Figure 5. Biomineralization by diffusion

The interdiffusion process can be used to nucleate and grow inorganic CPs on the hydrogel matrix. In this process, the hydrogel is placed between two ion solution reservoirs (Ca²⁺ and HPO₄²⁻-PO₄³⁻). (a) When a current flows, the ions are forced to diffuse accordingly through the hydrogel. (b) In situ SEM shows the homogeneous distribution of the small (< 1 μm) mineralized spherical vesicles (white dots) formed inside the polymer through phase separation. The pH in the hydrogel changes according to the movement of the OH⁻ solution front. (c-d) The ions meet at a certain position in the hydrogel, where minerals precipitate due to the pH change and the associated decrease in solubility. The precipitation also creates a local ion-concentration deficiency. The Ca²⁺ and PO₄³⁻ will keep precipitating in the gel along the OH⁻ path, creating local concentration gradients that promote ion transport to the mineral-forming location. The mineral concentration in the gel can be controlled by the movement of the OH⁻ front. Reprinted with permission from Reference [59], copyright © 2009, American Chemical Society.

Figure 6

A robocasted scaffold like the glass one shown in the scanning electron micrograph in the left could be coated with polymer layers containing drug delivery spheres to manipulate the spatio-temporal release of chemicals. The inside of the sphere will be made from porous hydrogel, or gelatin (sponge) into which the drug (BMP-2, VEGF, etc.) will be infiltrated. The outside will be made from degradable PLGA, or polycaprolactone, to protect the hydrogel and prevent burst release of the drug. The micro-capillaries will be introduced with a laser. The optimum amount and diameter of the micro-capillaries could be determined using diffusion calculations to project the release of the drug for the desired time, also assuming the thinning of the PLGA capsule. The coating composition can also be tailored to manipulate drug release and control its biodegradation, while sensors embedded in the coating monitor the biochemical environment.