Award details

The mechanics of the collagen fibrillar network in ageing cartilage

Reference	BB/R003610/1
Principal Investigator / Supervisor	Professor Himadri Shikhar Gupta
Co-Investigators / Co-Supervisors	Professor Martin Knight
Institution	Queen Mary University of London
Department	School of Engineering & Materials Scienc
Funding type	Research
Value (£)	369,875
Status	Current
Type	Research Grant
Start date	01/10/2017
End date	24/07/2023
Duration	70 months

Abstract

Type II collagen fibrils in cartilage tissue play a critical but experimentally less understood role in joint biomechanics. Using time-resolved synchrotron X-ray nanomechanics as a novel probe of cartilage fibrillar mechanics, our preliminary data has uncovered hitherto unknown aspects of fibrillar deformation under loading, including transient loss of pre-strain, intrafibrillar disordering indicative of water movement, alteration of collagen pre-strain levels under proteoglycan digestion and in-phase changes of fibril strain, orientation and disorder under cyclic loading. We hypothesize that age-related changes in cartilage, including crosslinking and hydration, will critically alter these fibrillar deformation dynamics, which will have significant effects on joint biomechanical deterioration in ageing. We will use synchrotron X-ray nanomechanics combined with proteomic characterization of ageing human tissues to test this hypothesis. Further, using crosslinking and hydration as mechanistic variables, we will test their effect under controlled conditions in bovine cartilage. We will characterise age-related alteration in collagen fibrillar dynamics under loading and possible fatigue-related changes. By linking fibrillar response to localized loading, we will clarify how focal damage to cartilage can spread to overall joint degradation. We will obtain a comprehensive understanding of the structural and mechanical role of the collagen fibrillar network in cartilage, and quantify the mechanical homeostatic changes in ageing. By integrating molecular-level information through microscale mapping, we will be able to link small scale alterations to whole joint deterioration in ageing and musculoskeletal degeneration. This will in turn enable potential clinical impact (understanding the influence of drugs on the mechanics of cartilage at the nanoscale) or in investigating the downstream effect of genetic knockout models of disease.

Summary

The connective tissues in our bodies are made up of both cells as well as a fibrous matrix around the cells. The fibrous matrix plays the major role in giving the tissue its mechanical properties needed for function. Despite having very different functions, the fibrous matrices of different soft tissues are at the molecular level made up of similar building blocks: collagen molecules, long sugar chains linked by protein (proteoglycans), and water. In particular, collagen molecules form long thin fibrils, which assemble into a network along with the gel-like material of proteoglycans and water. To achieve a range of diverse functions from the same building blocks, different soft tissues often vary the relative proportion of fibrils to the proteoglycan gel, or their orientation or interconnection to form complex composite materials at very small scales, below the thickness of a human hair. When we age, the properties of our connective tissues tend to deteriorate: e.g. skin becomes stiffer, and cartilage breaks down in osteoarthritis. These adverse changes arise from changes in either the intrinsic properties of the building blocks, or in their architecture. Because these changes occur at very small (nanometre) length scales, it is challenging to find out both the change and its effect on mechanics. To address this, our group has developed a high resolution X-ray imaging technique which works like a diffraction grating for collagen: it picks up regularities in the arrangement of the nanoscale collagen fibril networks in tissues, and when used with a very bright X-ray source like a synchrotron, can track how the fibrils stretch, reorient or otherwise respond to loads. In this project, we will apply this method to understand how the nanoscale mechanics of the collagen fibrillar network in cartilage changes in ageing. Articular cartilage serves as a frictionless bearing surface in joints, and cushions the load transfer between bones. If overloaded, the fibrous matrixbreaks down and leads to osteoarthritis, joint pain and immobility. We aim to understand how the compositional changes in collagen link to the alterations in its nanoscale mechanics - and eventually to joint breakdown. We will combine the X-ray technique with high-level characterisation of the protein composition and structure in the tissue as it ages. Such a combination is completely novel: the X-ray technique has not been applied to cartilage before, and its combination with proteomics enables a clear link between structural change and mechanical function. In cartilage, the collagen fibrillar network resists the swelling pressure of the proteoglycan gel. We first aim to understand how this load-balance changes in ageing, and by varying the chemical structure and relative proportion of different components in cartilage, to understand the mechanisms linking changes at the molecular level to disruption of mechanical equilibrium. Secondly, we will study real-time deformation of collagen fibrils as they are subjected to the types of load observed in real life and how ageing affects these dynamics. This is especially relevant because ageing leads to fibrillated and disrupted cartilage, but the mechanism by which collagen fibrils fail to resist loading is not understood. We will then focus on two types of relevant biomechanics: repeated loading or local traumatic impact. First, we will investigate whether the fibrillar response to repetitive loading is altered in ageing. Then, we will map, with micron-resolution, how collagen fibrils around the site of a local injury deform, testing the hypothesis that compositional change in ageing enables the damage to spread across the joint. To achieve these aims, we have brought together complementary expertise in X-ray nanomechanics (Gupta), cartilage mechanics (Knight), proteomics of ageing tissues (Swift) and synchrotron technology (Terrill), all of whom are internationally leading in their fields.

Impact Summary

The project will deliver in three areas: public engagement, academic engagement and industrial/application impact. A significant part of the academic impact activity is integrated into the "Academic Beneficiaries" section above. In terms of industrial/application impact, by the development of software methods for rapid, high throughput extraction and visualization of nanoscale structural parameters in collagenous fibrous biocomposites like cartilage, we will facilitate the application of high brilliance synchrotron X-ray microprobe scattering SAXS by the wider bioengineering and biomechanics community. Taking advantage of the structural and compositional commonalities across hydrated collagenous tissues like cartilage, tendon, skin and intervertebral disc, we will develop software scripts which will perform reduction, fitting and display of nanostructural parameters arising from the fibrillar collagen SAXS pattern. These parameters include but are not limited to: fibril D-period, fibril orientation, degree of orientation, fibrillar radial distribution, gap/overlap ratio and lateral intermolecular spacing. By working with the software team developing the general X-ray diffraction analysis program DAWN (www.dawnsci.org) we will embed these scripts into the pipelines for data processing present in this software. As a result, non-specialist users (like biologists, bioengineers or clinicians) would be able to drop in a series of acquired experimental synchrotron X-ray diffraction patterns acquired from a spatial map or time series of a collagenous tissue into DAWN, and with minimal user intervention, acquire a 2D map or time-plot of collagen fibrillar orientation, pre-strain or any other parameter extractable from the SAXS pattern. By combining high throughput data analysis, advanced and attractive data graphical representations and full automatization, such software scripts will significantly lower the technical barrier surrounding certain synchrotron X-ray imagingmethods in the eyes of non-specialists, and facilitate their application to a wide range of biological and bioengineering questions. To enable this, Diamond Light Source has committed a significant portion of an expert software PDRA's time to work with our team to develop and implement these routines into DAWN over the course of the Research Objectives. The second area of impact is a planned series of public engagement activities at the Royal Society Summer Exhibition, the Big Bang Science Fair and at the Centre of the Cell at the Blizzard Institute, Whitechapel (London). These activities will engage and enlighten the public on the excitement of a relatively overlooked aspect of biology - the role of the extracellular matrix - and to convey the spirit of multidisciplinary research in STEM subjects. The stand will consist of two main parts. The first will enable users to generate fibril-matrix architectures of the types found in the body (cartilage, skin etc.) using 3D printing, combined with interactive visualization of the X-ray diffraction patterns produced by the structures generated and supported by movies of acquired experimental data and tissue-microscopy. The second will emphasize the role of the fibre-matrix interactions by combining interactive modification of relative material properties of fibre and matrix, along with their orientations, with simulations of observed mechanical behaviours. These activities are chosen to cover different audiences: while the Royal Society and the Big Bang Science Fair will attract visitors across the UK, the Centre of the Cell exhibitions in East London have a special focus on widening participation and science outreach in the community. Lastly, this project will impact academic research via application of novel combinations of methods - like the synchrotron X-ray and proteomics approaches - which will impact bioengineering, biophysics, cartilage development, mechanobiology and development of X-ray methods.

Committee	Research Committee C (Genes, development and STEM approaches to biology)
Research Topics	Ageing
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

Associated awards:

BB/R004773/1 The mechanics of the collagen fibrillar network in ageing cartilage

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