I22-Small angle scattering & Diffraction
|
Diamond Proposal Number(s):
[25197]
Open Access
Abstract: Chemically crosslinked acellular bovine pericardium (ABP) has been widely used in clinical practice as bioprostheses. To ensure its consistency and durability, crosslinkers are used in excess, with stability guided by indicators including the hydrothermal denaturation temperature, the enzymatic resistance and the degree of crosslinking. Yet, understanding of the intermolecular structure in collagen fibrils which imparts the intrinsic stability of the ABPs is lacking, and the discrepancies in the stability criteria in varied conditions are poorly explained. In this study, synchrotron small-angle X-ray scattering (SAXS) in combination with thermal and colorimetric methods are employed to investigate the changes in the structure and the stability of ABPs during crosslinking using glutaraldehyde (GA) or 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) at different concentrations. Based on the findings, a mechanism is proposed to explicate the crosslinking effects on collagen structure and the relationship between the structure and each stability indicator. At low crosslinker concentrations, the telopeptidyl-helical linkages are preferred which cause rearrangements in the intermolecular structure of collagen, and efficiently contribute to its stabilities. Excess crosslinking is revealed by a revert trend in structural changes and the plateauing of the stabilities, assigning to the helical-helical linkages and monovalent bindings. The former would improve thermal stability but not collagenase resistance, whereas the latter have negligible effects. Overall, this study provides mechanistic understanding of the chemical crosslinking of ABPs which will contribute to the future development of more efficient and economically viable strategies to produce bioprostheses.
|
Aug 2022
|
|
I13-2-Diamond Manchester Imaging
|
Nicole
Sommer
,
Daniela
Hirzberger
,
Lisa
Paar
,
Leopold
Berger
,
Hanna
Cwieka
,
Uwe Y.
Schwarze
,
Valentin
Herber
,
Begum
Okutan
,
Andrew J.
Bodey
,
Regine
Willumeit-Römer
,
Berit
Zeller-Plumhoff
,
Jörg F.
Löffler
,
Annelie M.
Weinberg
Diamond Proposal Number(s):
[25485]
Abstract: Implant removal is unnecessary for biodegradable magnesium (Mg)-based implants and, therefore, the related risk for implant-induced fractures is limited. Aging, on the other hand, is associated with low bone-turnover and decreased bone mass and density, and thus increased fracture risk. Osteoporosis is accompanied by Mg deficiency, therefore, we hypothesized that Mg-based implants may support bone formation by Mg2+ ion release in an ovariectomy-induced osteoporotic rat model. Hence, we investigated osseointegration and implant degradation of a low-alloyed, degrading Mg–Zn–Ca implant (ZX00) in ovariectomy-induced osteoporotic (Osteo), old healthy (OH), and juvenile healthy (JH) groups of female Sprague Dawley rats via in vivo micro-computed tomography (µCT). For the Osteo rats, we demonstrate diminished trabecular bone already after 8 weeks upon ovariectomy and significantly enhanced implant volume loss, with correspondingly pronounced gas formation, compared to the OH and JH groups. Sclerotic rim development was observed in about half of the osteoporotic rats, suggesting a prevention from foreign-body and osteonecrosis development. Synchrotron radiation-based µCT confirmed lower bone volume fractions in the Osteo group compared to the OH and JH groups. Qualitative histological analysis additionally visualized the enhanced implant degradation in the Osteo group. To date, ZX00 provides an interesting implant material for young and older healthy patients, but it may not be of advantage in pharmacologically untreated osteoporotic conditions.
|
May 2022
|
|
I02-Macromolecular Crystallography
I03-Macromolecular Crystallography
I22-Small angle scattering & Diffraction
|
Diamond Proposal Number(s):
[11316, 8458]
Abstract: The mechanical properties of connective tissues are tailored to their specific function, and changes can lead to dysfunction and pathology. In most mammalian tissues the mechanical environment is governed by the micro- and nano-scale structure of collagen and its interaction with other tissue components, however these hierarchical properties remain poorly understood. In this study we use the human cornea as a model system to characterise and quantify the dominant deformation mechanisms of connective tissue in response to cyclic loads of physiological magnitude. Synchronised biomechanical testing, x-ray scattering and 3D digital image correlation revealed the presence of two dominant mechanisms: collagen fibril elongation due to a largely elastic, spring-like straightening of tropocollagen supramolecular twist, and a more viscous straightening of fibril crimp that gradually increased over successive loading cycles. The distinct mechanical properties of the two mechanisms suggest they have separate roles in vivo. The elastic, spring-like mechanism is fast-acting and likely responds to stresses associated with the cardiac cycle, while the more viscous crimp mechanism will respond to slower processes, such as postural stresses. It is anticipated that these findings will have broad applicability to understanding the normal and pathological functioning of other connective tissues such as skin and blood vessels that exhibit both helical structures and crimp.
|
Jan 2022
|
|
I13-2-Diamond Manchester Imaging
|
Diamond Proposal Number(s):
[19322]
Open Access
Abstract: Many soft tissues, such as the intervertebral disc (IVD), have a hierarchical fibrous composite structure which suffers from regional damage. We hypothesise that these tissue regions have distinct, inherent fibre structure and structural response upon loading. Here we used synchrotron computed tomography (sCT) to resolve collagen fibre bundles (∼5μm width) in 3D throughout an intact native rat lumbar IVD under increasing compressive load. Using intact samples meant that tissue boundaries (such as endplate-disc or nucleus-annulus) and residual strain were preserved; this is vital for characterising both the inherent structure and structural changes upon loading in tissue regions functioning in a near-native environment. Nano-scale displacement measurements along >10,000 individual fibres were tracked, and fibre orientation, curvature and strain changes were compared between the posterior-lateral region and the anterior region. These methods can be widely applied to other soft tissues, to identify fibre structures which cause tissue regions to be more susceptible to injury and degeneration. Our results demonstrate for the first time that highly-localised changes in fibre orientation, curvature and strain indicate differences in regional strain transfer and mechanical function (e.g. tissue compliance). This included decreased fibre reorientation at higher loads, specific tissue morphology which reduced capacity for flexibility and high strain at the disc-endplate boundary.
|
Oct 2021
|
|
|
|
Open Access
Abstract: Biomechanical changes to the collagen fibrillar architecture in articular cartilage are believed to play a crucial role in enabling normal joint function. However, experimentally there is little quantitative knowledge about the structural response of the Type II collagen fibrils in cartilage to cyclic loading in situ, and the mechanisms that drive the ability of cartilage to withstand long-term repetitive loading. Here we utilize synchrotron small-angle X-ray scattering (SAXS) combined with in-situ cyclic loading of bovine articular cartilage explants to measure the fibrillar response in deep zone articular cartilage, in terms of orientation, fibrillar strain and inter-fibrillar variability in healthy cartilage and cartilage degraded by exposure to the pro-inflammatory cytokine IL-1β. We demonstrate that under repeated cyclic loading the fibrils reversibly change the width of the fibrillar orientation distribution whilst maintaining a largely consistent average direction of orientation. Specifically, the effect on the fibrillar network is a 3-dimensional conical orientation broadening around the normal to the joint surface, inferred by 3D reconstruction of X-ray scattering peak intensity distributions from the 2D pattern. Further, at the intrafibrillar level, this effect is coupled with reversible reduction in fibrillar pre-strain under compression, alongside increase in the variability of fibrillar pre-strain. In IL-1β degraded cartilage, the collagen rearrangement under cyclic loading is disrupted and associated with reduced tissue stiffness. These finding have implications as to how changes in local collagen nanomechanics might drive disease progression or vice versa in conditions such as osteoarthritis and provides a pathway to a mechanistic understanding of such diseases.
|
Sep 2021
|
|
I13-2-Diamond Manchester Imaging
|
Diamond Proposal Number(s):
[22575, 20132]
Abstract: Digital volume correlation (DVC) in combination with high-resolution micro-computed tomography (microCT) imaging and in situ mechanical testing is gaining popularity for quantifying 3D full-field strains in bone and biomaterials. However, traditional in situ time-lapsed (i.e., interrupted) mechanical testing cannot fully capture the dynamic strain mechanisms in viscoelastic biological materials. The aim of this study was to investigate the time-resolved deformation of bone structures and analogues via continuous in situ synchrotron-radiation microCT (SR-microCT) compression and DVC to gain a better insight into their structure-function relationships. Fast SR-microCT imaging enabled the deformation behaviour to be captured with high temporal and spatial resolution. Time-resolved DVC highlighted the relationship between local strains and damage initiation and progression in the different biostructures undergoing plastic deformation, bending and/or buckling of their main microstructural elements. The results showed that SR-microCT continuous mechanical testing complemented and enhanced the information obtained from time-lapsed testing, which may underestimate the 3D strain magnitudes as a result of the stress relaxation occurring in between steps before image acquisition in porous biomaterials. Altogether, the findings of this study highlight the importance of time-resolved in situ experiments to fully characterise the time-dependent mechanical behaviour of biological tissues and biomaterials and to further explore their micromechanics under physiologically relevant conditions.
|
Jun 2021
|
|
I22-Small angle scattering & Diffraction
|
Diamond Proposal Number(s):
[20365]
Abstract: Understanding interactions between nanoparticles and model membranes is relevant to functional nano-composites and the fundamentals of nanotoxicity. In this study, the effect of polyamidoamine (PAMAM) dendrimers as model nanoparticles (NP) on the mesophase behaviour of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) has been investigated using high-pressure small-angle X-ray scattering (HP-SAXS). The pressure-temperature (
p−T
) diagrams for POPE mesophases in excess water were obtained in the absence and presence of G2 and G4 polyamidoamine (PAMAM) dendrimers (29 Å and 45 Å in diameter, respectively) at varying NP-lipid number ratio (ν = 0.0002-0.02) over the pressure range p = 1-3000 bar and temperature range T = 20–80°C. The
p−T
phase diagram of POPE exhibited the Lβ, Lα and HII phases. Complete analysis of the phase diagrams, including the relative area pervaded by different phases, phase transition temperatures (Tt) and pressures (pt), the lattice parameters (d-spacing), the pressure-dependence of d-spacing (Δd/Δp), and the structural ordering in the mesophase as gauged by the Scherrer coherence length (L) permitted insights into the size- and concentration-dependent interactions between the dendrimers and the model membrane system. The addition of dendrimers changed the phase transition pressure and temperature and resulted in the emergence of highly swollen lamellar phases, dubbed Lβ-den and Lα-den. G4 PAMAM dendrimers at the highest concentration ν = 0.02 suppressed the formation of the HII phase within the temperature range studied, whereas the addition of G2 PAMAM dendrimers at the same concentration promoted an extended mixed lamellar region in which Lα and Lβ phases coexisted.
|
Jan 2020
|
|
I22-Small angle scattering & Diffraction
|
Diamond Proposal Number(s):
[17869]
Abstract: The cuticle of stomatopod is an example of a natural mineralized biomaterial, consisting of chitin, amorphous calcium carbonate and protein components with a multiscale hierarchical structure, and forms a protective shell with high impact resistance. At the ultrastructural level, cuticle mechanical functionality is enabled by the nanoscale architecture, wherein chitin fibrils are in intimate association with enveloping mineral and proteins. However, the interactions between these ultrastructural building blocks, and their coupled response to applied load, remain unclear. Here, we elucidate these interactions via synchrotron microbeam wide-angle X-ray diffraction combined with in situ tensile loading, to quantify the chitin crystallite structure of native cuticle – and after demineralization and deproteinization – as well as time-resolved changes in chitin fibril strain on macroscopic loading. We demonstrate chitin crystallite stabilization by mineral, seen via a compressive pre-strain of approximately 0.10% (chitin/protein fibre pre-stress of ∼20 MPa), which is lost on demineralization. Clear reductions of stiffness at the fibrillar-level following matrix digestion are linked to the change in the protein/matrix mechanical properties. Furthermore, both demineralization and deproteinization alter the 3D-pattern of deformation of the fibrillar network, with a non-symmetrical angular fibril strain induced by the chemical modifications, associated with loss of the load-transferring interfibrillar matrix. Our results demonstrate and quantify the critical role of interactions at the nanoscale (between chitin-protein and chitin-mineral) in enabling the molecular conformation and outstanding mechanical properties of cuticle, which will inform future design of hierarchical bioinspired composites.
|
Sep 2019
|
|
I18-Microfocus Spectroscopy
I22-Small angle scattering & Diffraction
|
Diamond Proposal Number(s):
[10311]
Open Access
Abstract: Structural and associated biomechanical gradients within biological tissues are important for tissue functionality and preventing damaging interfacial stress concentrations. Articular cartilage possesses an inhomogeneous structure throughout its thickness, driving the associated variation in the biomechanical strain profile within the tissue under physiological compressive loading. However, little is known experimentally about the nanostructural mechanical role of the collagen fibrils and how this varies with depth. Utilising a high-brilliance synchrotron X-ray source, we have measured the depth-wise nanostructural parameters of the collagen network in terms of the periodic fibrillar banding (D-period) and associated parameters. We show that there is a depth dependent variation in D-period reflecting the pre-strain and concurrent with changes in the level of intrafibrillar order. Further, prolonged static compression leads to fibrillar changes mirroring those caused by removal of extrafibrillar proteoglycans (as may occur in aging or disease). We suggest that fibrillar D-period is a sensitive indicator of localised changes to the mechanical environment at the nanoscale in soft connective tissues.
|
Jul 2019
|
|
I14-Hard X-ray Nanoprobe
I18-Microfocus Spectroscopy
|
Open Access
Abstract: Biological exposures to micro- and nano-scale exogenous metal particles generated as a consequence of in-service degradation of orthopaedic prosthetics can result in severe adverse tissues reactions. However, individual reactions are highly variable and are not easily predicted, due to in part a lack of understanding of the speciation of the metal-stimuli which dictates cellular interactions and toxicity. Investigating the chemistry of implant derived metallic particles in biological tissue samples is complicated by small feature sizes, low concentrations and often a heterogeneous speciation and distribution. These challenges were addressed by developing a multi-scale two-dimensional X-ray absorption spectroscopic (XAS) mapping approach to discriminate sub-micron changes in particulate chemistry within ex-vivo tissues associated with failed CoCrMo total hip replacements (THRs). As a result, in the context of THRs, we demonstrate much greater variation in Cr chemistry within tissues compared with previous reports. Cr compounds including phosphate, hydroxide, oxide, metal and organic complexes were observed and correlated with Co and Mo distributions. This variability may help explain the lack of agreement between biological responses observed in experimental exposure models and clinical outcomes. The multi-scale 2D XAS mapping approach presents an essential tool in discriminating the chemistry in dilute biological systems where speciation heterogeneity is expected.
|
Jun 2019
|
|
