Nanotechnology can be conveniently defined in terms of linear scale; between 1 and 100 nanometres. Unfortunately, the definition, although correct in terms of scale does not describe what this science represents. This is perhaps why there are so many varied opinions of nanotechnology. This article aims to offer a clearer insight into this topic and its relationship to orthopaedics.
Nanotechnology hit the headlines a few years ago and public perception has shifted with time. Potential advantages were the focus of the media in the late 90s, but there was a change after the millennium. In 2003, The Times ran an article, “Prince (Charles) launches crusade against ‘grey goo’ science.”1 The subsequent interviews suggest this was not against nanotechnology per se, but simply about increasing awareness and research into the potential dangers. Since this time, it seems that society has become more acceptant of the concept; this has coincided with the adoption of many products into the marketplace that feature nanotechnology.
In simple terms, small-scale science can occur from the bottom up or the top down. In the first, small elements can be brought together bit by bit to create more complex structures. An example would be the interaction of molecules designed to self-assemble for a specific purpose. In top down approaches, devices can be used to manufacture scaled down or miniature devices and structures. An example of this would be the move of solid-state electronics from the micro- to the nano- scale, allowing smaller and more complex electronic components for items such as telephones and computers.
Biomolecules can be measured in nanometres. It is thus no surprise that nanotechnology has prioritised healthcare applications for potential exploitation. This has given rise to a new branch of science – nanomedicine. This encompasses the breadth of the topic from pharmacological agents and surface modification through to regulation and toxicology. There are many branches of medicine that could benefit in terms of diagnostics and treatment, e.g. oncology. This article cannot be exhaustive but can give a selection of recent examples of how nanotechnology can be applied to orthopaedic surgery.
Nanoscale surface modification
The characteristic shape of a surface at the nano and micro scale has a significant effect on the biological systems with which it interfaces. The ability to characterise and modify topography on a nanoarchitectural level has been a strategy to optimise surfaces for interaction with cells. The underlying premise has been that certain textures and chemical properties of surfaces might be more conducive to control and dictate cell interactions and thereby promote integration of artificial materials with biological tissues.
An in vitro study by Kay et al2 with nanostructured ceramic composites showed improved chondrocyte adhesion to such surfaces. The study also confirmed enhanced osteoblast adhesion as was first identified by Webster et al3. Nanotexturing was subsequently found to upregulate expression of bone sialoprotein and osteopontin in osteoblasts.4
In 2004, Webster and Ejiofor5 investigated the potential for nanotopography of metals to influence osteoblast adhesion and found similar results to those of nanostructured versus conventional ceramics. Popat et al6 carried out a study with nanoporous alumina membranes and found significantly increased matrix production as opposed to amorphous alumina. The authors also confirmed that more protein was adsorbed onto the nanoporous membranes.
The potential molecular mechanisms that underpin the way in which surface topography at the nanoscale influences the behaviour of bone cells are beginning to be unravelled. Studies by Dalby et al7 have identified how semi-ordered nanotopography alone can direct osteogenesis and emphasises how length scales of the same magnitude as the protein complexes that anchor cells to their surroundings can be engineered to control cell fate.
Over subsequent years, different topographies, methods and materials have been compared for effects on osteoblast adhesion and function. One interesting approach has been the conjugation of substances such as BMP-2 to such surfaces. On nanofibrous chitosan membranes, this led to a significantly increased proliferation of osteoblasts, alkaline phosphatase activity and calcium deposition compared with a BMP-2-adsorbed membrane.8 Another group investigated the effect of different extents of strontium substitution for calcium on hydroxyapatite thin films and found enhanced osteoblast proliferation and function combined with decreased osteoclast formation.9
In terms of in vivo work, Mendes, Moineddin and Davies10 assessed the osteoconduction of titanium and titanium alloy implants with and without a coating of a discrete crystalline deposition of calcium phosphate nanoparticles. These implants were placed bilaterally into the femora of Wistar rats and harvested after nine days. The authors found a significant increase in bone-implant contact in the presence of the calcium phosphate nanocrystals.
Orthopaedic tissue engineering has concentrated on bone and cartilage. Initially, this considered bone-substitute materials; early in the last decade there were several papers exploring variations on nanocrystalline hydroxyapatite formation, with biological supplements ranging from TGF-β, to chitosan, to BMP-2. More recently there has been interest in the potential for direct effects of these materials on osteoblasts. In 2006, Balasundaram, Sato and Webster compared the effect of nanometre-scale hydroxyapatite crystals on osteoblast adhesion as well as functionalised conventional hydroxyapatite.11 Additional work in 2009 tested nano- and micro-scale hydroxyapatite for its effect on human osteoblast-like MG63 cells. These studies demonstrated that cell proliferation was greater, and apoptosis reduced, with exposure to nano-scale hydroxyapatite as compared with the micro-scale particles.12
Further experiments on hydroxyapatite/calcium phosphate variants have introduced the artificial bone constructs into animal models of bone defects, which have then been tested for bone healing.13 This work has been extended to titanium-blast implants, with and without nanostructured calcium coating, placed into rabbit tibiae and then compared. Removal torque tests and histomorphometric analyses, demonstrated greater bone-implant contact and torque removal force being required with the calcium-coated implants, suggestive of enhanced osseointegration.14
If cartilage is now considered, in many areas the nanotechnological research into this can be seen to mirror that of bone. In recent years, there have been attempts to develop artificial cartilage. In 2008, Pan, Xiong and Gao15 tested nanohydroxyapatite-reinforced polyvinyl alcohol gel and found an optimum composite with mechanical properties similar to natural articular cartilage. In the same year, Nesti et al16 seeded multipotent, adult human mesenchymal stem cells onto an electrospun, biodegradable nanofibrous scaffold with a hyaluronic acid hydrogel centre. With appropriate growth factors to guide differentiation, a cartilaginous construct developed which had a macro- and micro-architecture resembling that of a natural intervertebral disc. In terms of potential pharmacological applications of nanotechnology in cartilage, in 2002 Horisawa et al17 developed and tested betamethasone sodium phosphate-loaded nanospheres for their effect on a chronic synovitis model in rabbits. They found a prolonged pharmacological effect with intra-articular injection of the nanosphere-loaded betamethasone as opposed to simple aqueous betamethasone solution.
A potential role for nanoscale particles in drug delivery has been a focus of research activity, with the ability to combine targeting, imaging and therapeutics leading to significant optimism that nanotechnology will lead to a new generation of treatment options. Initial work in 2003 demonstrated a possible use of polymer nanospheres coated with radioisotope. These constructs were injected into the circulation of mice, and demonstrated significantly higher bone uptake but decreased liver and spleen uptake compared with 99mTc-tin colloids.18 This suggested a potential for improved drug delivery to bone via nanotechnological means.
Other work has made use of nano-derived delivery systems where the target itself is inherently specific, such as in a mouse model of Ewing’s sarcoma with the EWS-Fli 1 fusion gene. Here, siRNA targeting the oncogene was loaded into biodegradable nanocapsules to improve intracellular penetration. The result was a dramatic inhibition of tumour growth that was not seen with free (i.e. non-encapsulated) siRNA.19 One paper has explored the potential therapeutic role for nanoparticles themselves, without added/conjugated substances. Yudoh et al20 have made use of the free radical scavenger property of water-soluble fullerene (C60) in a rat model of adjuvant-induced arthritis. In this model, intra-articular injection of C60 brought about a reduction in osteoclast number and extent of bone resorption. Finally, some work has been done into drug delivery via implants. Xin et al21 have produced strontium-releasing nanotube arrays on the surface of titanium implants which are capable of releasing strontium at a slow rate over protracted periods. The implants are yet to be tested in vivo.
This overview highlights how nanotechnology, in recent years, has become part of orthopaedic research. The research areas of implanted interfaces, tissue engineering and therapeutics have been an orthopaedic focus over many years. Nanotechnology has enhanced capability in these various areas. It has become an advanced toolkit of approaches that take traditional research priorities to a new level. A broad range of potential clinical applications in the short to medium term will feature aspects of nanotechnology.
1. Pierce A. Prince launches crusade against ‘grey goo’ science. The Times 2003 April 28.
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