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The intervertebral disc (IVD) has a complex and multiscale extracellular matrix structure which provides unique mechanical properties to withstand physiological loading. Low back pain ...has been linked to degeneration of the disc but reparative treatments are not currently available. Characterising the disc’s 3D microstructure and its response in a physiologically relevant loading environment is required to improve understanding of degeneration and to develop new reparative treatments. In this study, techniques for imaging the native IVD, measuring internal deformation and mapping volumetric strain were applied to an in situ compressed ex vivo rat lumbar spine segment. Synchrotron X-ray micro-tomography (synchrotron CT) was used to resolve IVD structures at microscale resolution. These image data enabled 3D quantification of collagen bundle orientation and measurement of local displacement in the annulus fibrosus between sequential scans using digital volume correlation (DVC). The volumetric strain mapped from synchrotron CT provided a detailed insight into the micromechanics of native IVD tissue. The DVC findings showed that there was no slipping at lamella boundaries, and local strain patterns were of a similar distribution to the previously reported elastic network with some heterogeneous areas and maximum strain direction aligned with bundle orientation, suggesting bundle stretching and sliding. This method has the potential to bridge the gap between measures of macro-mechanical properties and the local 3D micro-mechanical environment experienced by cells. This is the first evaluation of strain at the micro scale level in the intact IVD and provides a quantitative framework for future IVD degeneration mechanics studies and testing of tissue engineered IVD replacements.
Synchrotron in-line phase contrast X-ray tomography provided the first visualisation of native intact intervertebral disc microstructural deformation in 3D. For two annulus fibrosus volumes of interest, collagen bundle orientation was quantified and local displacement mapped as strain. Direct evidence of microstructural influence on strain patterns could be seen such as no slipping at lamellae boundaries and maximum strain direction aligned with collagen bundle orientation. Although disc elastic structures were not directly observed, the strain patterns had a similar distribution to the previously reported elastic network. This study presents technical advances and is a basis for future X-ray microscopy, structural quantification and digital volume correlation strain analysis of soft tissue.
Summary
Background
Human skin has the crucial roles of maintaining homeostasis and protecting against the external environment. Skin offers protection against mechanical trauma due to the reversible ...deformation of its structure; these biomechanical properties are amenable to dynamic testing using noninvasive devices.
Objectives
To characterize the biomechanical properties of young, black African/African‐Caribbean and white Northern European skin from different anatomical sites, and to relate underlying skin architecture to biomechanical function.
Methods
Using cutometry and ballistometry, the biomechanical properties of buttock and dorsal forearm skin were determined in black African/African‐Caribbean (n = 18) and white Northern European (n = 20) individuals aged 18–30 years. Skin biopsies were obtained from a subset of the volunteers (black African/African‐Caribbean, n = 5; white Northern European, n = 6) and processed for histological and immunohistochemical detection of the major elastic fibre components and fibrillar collagens.
Results
We have determined that healthy skin from young African and white Northern European individuals has similar biomechanical properties (F3): the skin is resilient (capable of returning to its original position following deformation, R1), exhibits minimal fatigue (R4) and is highly elastic (R2, R5 and R7). At the histological level, skin with these biomechanical properties is imbued with strong interdigitation of the rete ridges at the dermoepidermal junction (DEJ) and candelabra‐like arrays of elastic fibres throughout the papillary dermis. Dramatic disruption to this highly organized arrangement of elastic fibres, effacement of the rete ridges and alterations to the alignment of the fibrillar collagens is apparent in the white Northern European forearm and coincides with a marked decline in biomechanical function.
Conclusions
Maintenance of skin architecture – both epidermal morphology and elastic fibre arrangement – is essential for optimal skin biomechanical properties. Disruption to underlying skin architecture, as observed in the young white Northern European forearm, compromises biomechanical function.
What's already known about this topic?
Human skin offers protection against the external environment via the reversible deformation of its structure: the stratum corneum and components of the dermal extracellular matrix imbue skin with these important biomechanical properties.
Skin biomechanical properties are amenable to dynamic testing in vivo using the noninvasive methods of cutometry and ballistometry.
What does this study add?
This study combines in vivo testing of black African and white Northern European skin with the histological assessment of skin biopsies in order to interrogate the relationship between underlying skin architecture and biomechanical function.
Disruption to skin architecture, particularly loss of oxytalan fibres and effacement of rete ridges, correlates with perturbation of biomechanical function.
What is the translational message?
Quantitative measurement of biomechanical properties combined with histological assessment of underlying extracellular matrix architecture are useful tools to provide us with an understanding of skin's normal functions.
These techniques may be useful to examine the clinical progression of skin conditions that affect skin elasticity and provide objective measurement of the efficacy of treatment options designed to improve skin health.
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Linked Comment: Dobrev. Br J Dermatol 2017; 177:622–623
Plain language summary available online
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.
The analyses presented here are applicable to many collagenous soft tissues which suffer from regional damage. We aimed to investigate regional intervertebral disc (IVD) structural and functional differences by characterising collagen fibre architecture and linking specific fibre- and tissue-level deformation behaviours. Synchrotron CT provided the first demonstration of tracking discrete fibres in 3D within an intact IVD. Detailed analysis of regions was performed using over 200k points, spaced every 8 μm along 10k individual fibres. Such comprehensive structural characterisation is significant in informing future computational models. Morphological indicators of tissue compliance (change in fibre curvature and orientation) and fibre strain measurements revealed localised and regional differences in tissue behaviour.
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Summary
Many biological tissues have a complex hierarchical structure allowing them to function under demanding physiological loading conditions. Structural changes caused by ageing or disease can ...lead to loss of mechanical function. Therefore, it is necessary to characterise tissue structure to understand normal tissue function and the progression of disease. Ideally intact native tissues should be imaged in 3D and under physiological loading conditions. The current published in situ imaging methodologies demonstrate a compromise between imaging limitations and maintaining the samples native mechanical function. This review gives an overview of in situ imaging techniques used to visualise microstructural deformation of soft tissue, including three case studies of different tissues (tendon, intervertebral disc and artery). Some of the imaging techniques restricted analysis to observational mechanics or discrete strain measurement from invasive markers. Full‐field local surface strain measurement has been achieved using digital image correlation. Volumetric strain fields have successfully been quantified from in situ X‐ray microtomography (micro‐CT) studies of bone using digital volume correlation but not in soft tissue due to low X‐ray transmission contrast. With the latest developments in micro‐CT showing in‐line phase contrast capability to resolve native soft tissue microstructure, there is potential for future soft tissue mechanics research where 3D local strain can be quantified. These methods will provide information on the local 3D micromechanical environment experienced by cells in healthy, aged and diseased tissues. It is hoped that future applications of in situ imaging techniques will impact positively on the design and testing of potential tissue replacements or regenerative therapies.
Lay Description
The soft tissues in our bodies, such as tendons, intervertebral discs and arteries, have evolved to have complicated structures which deform and bear load during normal function. Small changes in these structures can occur with age and disease which then leads to loss of function. Therefore, it is important to image tissue microstructure in 3D and under functional conditions. This paper gives an overview of imaging techniques used to record the deformation of soft tissue microstructures. Commonly there are compromises between obtaining the best imaging result and retaining the samples native structure and function. For example, invasive markers and dissecting samples damages the tissues natural structure, and staining or clearing (making the tissue more transparent) can distort tissue structure. Structural deformation has been quantified from 2D imaging techniques (digital image correlation) to create surface strain maps which help identify local tissue mechanics. When extended to 3D (digital volume correlation), deformation measurement has been limited to bone samples using X‐ray micro‐CT. Recently it has been possible to image the 3D structure of soft tissue using X‐ray micro‐CT meaning that there is potential for internal soft tissue mechanics to be mapped in 3D. Future application of micro‐CT and digital volume correlation will be important for soft tissue mechanics studies particularly to understand normal function, progression of disease and in the design of tissue replacements.
Instructing others to move is fundamental for many populations, whether animal or cellular. In many instances, these commands are transmitted by contact, such that an instruction is relayed directly ...(e.g. by touch) from signaller to receiver: for cells, this can occur via receptor–ligand mediated interactions at their membranes, potentially at a distance if a cell extends long filopodia. Given that commands ranging from attractive to repelling can be transmitted over variable distances and between cells of the same (homotypic) or different (heterotypic) type, these mechanisms can clearly have a significant impact on the organisation of a tissue. In this paper, we extend a system of nonlocal partial differential equations (integrodifferential equations) to provide a general modelling framework to explore these processes, performing linear stability and numerical analyses to reveal its capacity to trigger the self-organisation of tissues. We demonstrate the potential of the framework via two illustrative applications: the contact-mediated dispersal of neural crest populations and the self-organisation of pigmentation patterns in zebrafish.
Abstract
Bacterial DNA gyrase introduces negative supercoils into chromosomal DNA and relaxes positive supercoils introduced by replication and transiently by transcription. Removal of these positive ...supercoils is essential for replication fork progression and for the overall unlinking of the two duplex DNA strands, as well as for ongoing transcription. To address how gyrase copes with these topological challenges, we used high-speed single-molecule fluorescence imaging in live Escherichia coli cells. We demonstrate that at least 300 gyrase molecules are stably bound to the chromosome at any time, with ∼12 enzymes enriched near each replication fork. Trapping of reaction intermediates with ciprofloxacin revealed complexes undergoing catalysis. Dwell times of ∼2 s were observed for the dispersed gyrase molecules, which we propose maintain steady-state levels of negative supercoiling of the chromosome. In contrast, the dwell time of replisome-proximal molecules was ∼8 s, consistent with these catalyzing processive positive supercoil relaxation in front of the progressing replisome.
Although the composition and structure of cartilaginous tissues is complex, collagen II fibrils and aggrecan are the most abundant assemblies in both articular cartilage (AC) and the nucleus pulposus ...(NP) of the intervertebral disc (IVD). Whilst structural heterogeneity of intact aggrecan ( containing three globular domains) is well characterised, the extent of aggrecan fragmentation in healthy tissues is poorly defined. Using young, yet skeletally mature (18-30 months), bovine AC and NP tissues, it was shown that, whilst the ultrastructure of intact aggrecan was tissue-dependent, most molecules (AC: 95 %; NP: 99.5 %) were fragmented (lacking one or more globular domains). Fragments were significantly smaller and more structurally heterogeneous in the NP compared with the AC (molecular area; AC: 8543 nm2; NP: 4625 nm2; p < 0.0001). In contrast, fibrillar collagen appeared structurally intact and tissue-invariant. Molecular fragmentation is considered indicative of a pathology; however, these young, skeletally mature tissues were histologically and mechanically (reduced modulus: AC: ≈ 500 kPa; NP: ≈ 80 kPa) comparable to healthy tissues and devoid of notable gelatinase activity (compared with rat dermis). As aggrecan fragmentation was prevalent in neonatal bovine AC (99.5 % fragmented, molecular area: 5137 nm2) as compared with mature AC (95.0 % fragmented, molecular area: 8667 nm2), it was hypothesised that targeted proteolysis might be an adaptive process that modified aggrecan packing (as simulated computationally) and, hence, tissue charge density, mechanical properties and porosity. These observations provided a baseline against which pathological and/or age-related fragmentation of aggrecan could be assessed and suggested that new strategies might be required to engineer constructs that mimic the mechanical properties of native cartilaginous tissues.
Cranial cruciate ligament disease (CCLD) is the most common cause of pelvic limb lameness in dogs but its precise aetiopathogenesis is uncertain. Fibrillin microfibrils (FM) are complex ...macro-molecular assemblies found in many tissues including ligaments, where they are thought to play an important mechanical role. We hypothesised that FM ultrastructural variation correlates with the differing predisposition of canine breeds to CCLD.
Non-diseased cranial and caudal cruciate ligaments (CCLs and CaCLs) were obtained from Greyhound (GH) and Staffordshire Bull Terrier (SBT) cadavers. Fibrillin microfibrils were extracted from the ligaments by bacterial collagenase digestion, purified by size-exclusion chromatography and subsequently visualized by atomic force microscopy (AFM). With AFM, FMs have a characteristic beads-on-a-string appearance. For each FM, periodicity (bead-bead distance) and length (number of beads/FM) was measured.
Fibrillin microfibril length was found to be similar for GH and SBT, with non-significant inter-breed and inter-ligament differences. Fibrillin microfibril periodicity varied when comparing GH and SBT for CCL (GH 60.2 ± 1.4 nm; SBT 56.2 ± 0.8 nm) and CaCL (GH 55.5 ± 1.6 nm; SBT 61.2 ± 1.2 nm). A significant difference was found in the periodicity distribution when comparing CCL for both breeds (P < 0.00001), further, intra-breed differences in CCL vs CaCL were statistically significant within both breeds (P < 0.00001). The breed at low risk of CCLD exhibited a periodicity profile which may be suggestive of a repair and remodelling within the CCL.
•Fibrillin microfibrils are important indicators of canine cruciate disease (CCD)•Greyhounds (low risk of CCD) exhibit a unique fibrillin microfibril profile•Intra-breed ultrastructural differences between ligaments may explain injury risk
Summary
Background
Geographical ancestry plays a key role in determining the susceptibility of human skin to external insults and dermatological disease. Despite this, studies of skin from ...individuals of diverse geographical ancestry focus primarily on epidermal pigmentation. Few reports characterize the gross morphology and composition of the dermis and dermal‐epidermal junction (DEJ).
Objectives
To characterize epidermal morphology and dermal composition in skin from individuals of diverse geographical ancestry.
Methods
Immunohistochemical techniques were used to assess epidermal morphology and protein composition of the DEJ and dermal extracellular matrix in photoprotected skin from young African, Eurasian and Far East Asian individuals (n = 7 per group; age 18–30 years).
Results
The epidermis of African skin was thicker, with deeper rete ridges and a more convoluted DEJ than Eurasian and Far East Asian skin. Compared with Eurasians, protein composition of the DEJ was collagen VII poor in African and Far East Asian skin (P < 0·001 and P < 0·01, respectively); the dermis of African skin was enriched in fibrillar collagens (P < 0·05), but was relatively elastin poor (P < 0·05). African dermis was abundant in fibrillin‐rich microfibrils and fibulin‐5 (P < 0·001 and P < 0·001, respectively) compared with Eurasian and Far East Asian skin.
Conclusions
We demonstrate that fundamental differences exist in skin structure and composition in individuals of diverse geographical ancestry. Disparate environmental pressures encountered by ancestral human populations living at different latitudes may have driven adaptations in skin structure and composition. Further research into the functional significance and clinical consequences of these differences is warranted.
What's already known about this topic?
While the role of epidermal pigmentation in characterizing human skin of differing geographical origins is well characterized, potential differences in dermal composition and epidermal structure are less well defined.
What does this study add?
This unique study compares dermal extracellular matrix composition and epidermal morphology of skin from three groups with diverse geographical ancestry.
This study identifies previously unappreciated differences in the abundance and architecture of dermal fibrillar collagens, collagen VII anchoring fibrils and the elastic fibre network.
It highlights potential functional and clinical differences in skin of diverse geographical origins, which may have arisen as a consequence of adaptations to ancestral environments.
An important question in evolutionary biology is how often, and to what extent, do similar ecologies elicit distantly related taxa to evolve towards the same phenotype? In some scenarios, the ...repeated evolution of particular phenotypes may be expected, for instance when species are exposed to common selective forces that result from strong functional demands. In bivalved scallops (Pectinidae), some species exhibit a distinct swimming behaviour (gliding), which requires specific biomechanical attributes to generate lift and reduce drag during locomotive events. Further, a phylogenetic analysis revealed that gliding behaviour has independently evolved at least four times, which raises the question as to whether these independent lineages have also converged on a similar phenotype. Here, we test the hypothesis that gliding scallops display shell shape convergence using a combination of geometric morphometrics and phylogenetic comparative methods that evaluate patterns of multivariate trait evolution. Our findings reveal that the gliding species display less morphological disparity and significant evolutionary convergence in morphospace, relative to expectations under a neutral model of Brownian motion for evolutionary phenotypic change. Intriguingly, the phylomorphospace patterns indicate that gliding lineages follow similar evolutionary trajectories to not one, but two regions of morphological space, and subsequent analyses identified significant differences in their biomechanical parameters, suggesting that these two groups of scallops accomplish gliding in different ways. Thus, whereas there is a clear gliding morphotype that has evolved convergently across the phylogeny, functionally distinct morphological subforms are apparent, suggesting that there may be two optima for the gliding phenotype in the Pectinidae.
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