Kick-starting mechanoadaptation in aged bones

Animals can modify the shape and mass of their individual limb bones to accommodate both habitual and new, imposed mechanical forces. This is perhaps best exemplified by the increases in bone mass that are seen in the dominant, serving arm of tennis players versus their non-serving, ball-throwing arm - we term the process by which these changes are achieved mechano-adaptation. If we understood the way such mechano-adaptive increases in bone mass were coordinated, we would be able to mimic them and, thus, alleviate any decreases in bone mass that places specific regions of bones at risk of fracture. This risk of fracture is very obvious in ageing bones, which fail to exhibit the capacity for such structural modification in response to imposed forces. There are massive, often hidden, consequences of such failure to adapt, which are increasing dramatically with enhanced longevity and lifestyle choices - the costs to health and society are huge. There is one main reason why we don't understand how bone mechano-adaptation is coordinated. The reason is that until now we have not been able to match microscopic level bone bending in response to imposed forces with the precise location of the ensuing structural increases in bone formation. We have recently made three advances that now make this possible. We have developed: i) a non-surgical model for imposing controllable forces to the tibia in living mice, ii) a means of measuring the extent of bending over the entire surface of small bones, and iii) recently acquired a machine that allows a precise microscopic 3D analysis of where bone is actively being formed. This creates a unique and timely opportunity. It was previously thought that mechano-adaptive processes acted to minimise the 'greatest' bending. We have generated pilot data that have allowed us to predict that, in contrast, bones adapt in order to minimise the 'steepest' local bending gradients on their surface. To test this we will bend some bones in mature mice that we know will adapt by increasing bone formation. To test whether steep bending gradients 'drive' formation we will: i) measure the bending pattern to generate a 'contour map' across the tibial bone surface, while it is being exposed to mechanical forces; ii) produce a 3D picture of the entire tibia describing exactly where bone formation takes place; iii) explore the overall changes in tibial structure by high-power 3D X-ray imaging, and iv) create virtual, digital computer images of the changes induced by these mechanical forces. These studies will determine which specific mechanical force is responsible for promoting bone formation and whether this occurs where bending gradients are indeed steepest. This will be useful if it can be used to modify bones which fail to fulfil their structural load-bearing role, as in ageing. Our pilot data from aged mouse bone are therefore crucial as they show that bending gradients appear less 'steep'; the general magnitude of bending engendered by force increases and the steepest gradients are consequently less severe. Using the methods described above as well as measures of bone quality and cellular responses to mechanical stimulation, we will explore if tibiae in aged mice exhibit a reduced cellular sensitivity to bending forces and/or a reduced mechanical stimulus capable of promoting adaptation. Our pilot studies have also shown that aged mouse bones do not exhibit mechano-adaptation and, intriguingly, that this can be restored to aged bones by prior imposition of a prolonged period without habitual use. We propose to explore the factors that change with this restoration, in the hope that we can identify how to kick-start mechano-adaptation in aged bones and thus alleviate the risk of fracture. Defining the drivers of adaptive increases in new bone formation in a precise 3D manner will also allow future studies to pinpoint the cellular events that are necessary to coordinate mechano-adaptation.

Adaptation to loading (mechano-adaptation) maintains bone mass and strength. Relationships between applied loads, the strain patterns generated, and ensuing structural changes remain largely unknown; precise 3D registration and measurement have made it impossible. Three advances now create a unique opportunity to identify the basis of this relationship. We have developed: i) a non-surgical in vivo mouse tibial loading model, ii) a means of 'mapping' strains across entire surfaces of such loaded bones by digital image correlation (DIC), and iii) acquired new technology for 'slice and view' fluorescence imaging and 3D reconstruction of entire bones. We will combine these approaches to address three aims. Our pilot DIC studies have mapped strains on normal tibiae and tibiae which have undergone adaptation to in vivo loading (2 weeks), to establish that this had minimised high strain gradients. Our first aim is to examine whether bone formation occurs in regions of high strain gradients. Skeletal fracture vulnerability in ageing is due to a mechano-adaptive failure. Whether aged bones lose responsiveness due to an insufficient mechanical triggering stimulus or inadequate cellular load-sensing or bone synthesis is unclear. Our pilot studies show that mechano-adaptation fails in aged bones (despite induction of generally higher strains), and our second aim will determine whether ageing bone exhibits a reduced mechanical stimulus and/or in cellular sensitivity to strain. Our final pilot data found that mechano-adaptive responses are restored to aged bones by prolonged neurectomy-induced disuse and our third aim is to establish whether the osteogenic activity in aged bone can be 'kick-started' by introducing regions of high strain gradient. This proposal will identify how we can restore adaptive responses to ageing bone and define strain: bone formation relationships allowing the pathways that translate and communicate the effects of loading to be determined.

Demand for an understanding of why aged bones become more vulnerable to fracture is increasing rapidly in lay, academic, animal welfare and healthcare communities. The healthcare implications of bone frailty are massive, with treatment costs of over 13 billion Euros/year in the EU related to bone frailty. It is clear that this impact will only rise in the aging population. Understanding the critical processes involved in maintaining appropriate bone mass and architecture will allow for more realistic and practical therapeutic approaches for controlling bone mass to be developed. The proposed work will impact significantly on our understanding of bone mechano-adaptation by establishing the importance of contributions made by specific mechanical components of the load-bearing bone environment. By defining the mechanical 'drivers' of bone adaptation in the mature skeleton and the potential for these to be restored in aged bone, suitable regimes for exploiting these processes can be identified. The following groups will benefit: 1. Industry The work proposed here is applicable to both the implant industry (prosthetic and biomaterial implants) and pharmaceutical companies. Mechanical integrity is critical for bone implants and can be promoted through mechano-adaptation. Our research will identify how bone is spatially regulated during mechano-adaptation and identify possible deficits in the mechanical environment or cellular function in aged bone. Bone-modifying drugs that target the bone's inherent adaptation pathways will have improved spatial regulation in comparison to drugs that promote uncontrolled diffuse bone formation. 2. Academics: Academics working in a number of fields across biology, aging, and bioengineering will benefit, as outlined in the 'academic beneficiaries' section of this proposal. 3. Society: The public will benefit from the development of novel and safe 'exercise' regimes aimed at optimizing impact on the skeleton. To date physical therapy to address bone loss in the elderly has had limited success. Our work will provide the understanding that is necessary for modifying the guidelines regarding how the aged should aim to conserve their bones and limit their chances of fracture. 4. Policy makers and government agencies: The NHS and the Animal Health Trust and DEFRA will benefit from insight to potential benefits to skeletal health revealed by our studies. The RVC's active involvement with public engagement in the field of animal welfare will also benefit from this research which will feed into reports, commissions and recommendations made by these and similar societies. Timescale for benefits to be realized: General conclusions about how bones adapt and details regarding how these mechanisms decline with ageing, are likely to have a relatively rapid impact on health policy. As there is relatively little advice at present, any concrete advice that can be offered with reasonable confidence is likely to be implemented in a wide range of circumstances.