Elastic fibres Kielty, Cay M; Sherratt, Michael J; Shuttleworth, C Adrian
Journal of cell science,
2002-Jul-15, Letnik:
115, Številka:
Pt 14
Journal Article
Recenzirano
Odprti dostop
Elastic fibres are essential extracellular matrix macromolecules comprising an elastin core surrounded by a mantle of fibrillin-rich microfibrils. They endow connective tissues such as blood vessels, ...lungs and skin with the critical properties of elasticity and resilience. The biology of elastic fibres is complex because they have multiple components, a tightly regulated developmental deposition, a multi-step hierarchical assembly and unique biomechanical functions. However, their molecular complexity is at last being unravelled by progress in identifying interactions between component molecules, ultrastructural analyses and studies of informative mouse models.
Age-related aortic stiffening is associated with cardiovascular diseases such as heart failure. The mechanical functions of the main structural components of the aorta, such as collagen and elastin, ...are determined in part by their organisation at the micrometer length scale. With age and disease both components undergo aberrant remodelling, hence, there is a need for accurate characterisation of the biomechanical properties at this length scale. In this study we used a frequency-modulated atomic force microscopy (FM-AFM) technique on a model of ageing in female sheep aorta (young: ~18 months, old: >8 years) to measure the micromechanical properties of the medial layer of the ascending aorta. The novelty of our FM-AFM method, operated at 30kHz, is that it is non-contact and can be performed on a conventional AFM using the ׳cantilever tune’ mode, with a spatial (areal) resolution of around 1.6μm2. We found significant changes in the elastic and viscoelastic properties within the medial lamellar unit (elastic lamellae and adjacent inter-lamellar space) with age. In particular, there was an increase in elastic modulus (Young; geometric mean (geometric SD)=42.9 (2.26)kPa, Old=113.9 (2.57)kPa, P<0.0001), G′ and G″ (storage and loss modulus respectively) (Young; G′=14.3 (2.26)kPa, Old G′=38.0 (2.57)kPa, P<0.0001; Young; G″=14.5 (2.56)kPa, Old G″=32.8 (2.52)kPa, P<0.0001). The trends observed in the elastic properties with FM-AFM matched those we have previously found using scanning acoustic microscopy (SAM). The utility of the FM-AFM method is that it does not require custom AFM hardware and can be used to simultaneously determine the elastic and viscoelastic behaviour of a biological sample.
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We propose a new model for the alignment of fibrillin molecules within fibrillin microfibrils. Automated electron tomography was used to generate three-dimensional microfibril reconstructions to ...18.6-Å resolution, which revealed many new organizational details of untensioned microfibrils, including heart-shaped beads from which two arms emerge, and interbead diameter variation. Antibody epitope mapping of untensioned microfibrils revealed the juxtaposition of epitopes at the COOH terminus and near the prolinerich region, and of two internal epitopes that would be 42-nm apart in unfolded molecules, which infers intramolecular folding. Colloidal gold binds microfibrils in the absence of antibody. Comparison of colloidal gold and antibody binding sites in untensioned microfibrils and those extended in vitro, and immunofluorescence studies of fibrillin deposition in cell layers, indicate conformation changes and intramolecular folding. Mass mapping shows that, in solution, microfibrils with periodicities of <70 and >140 nm are stable, but periodicities of ∼100 nm are rare. Microfibrils comprise two in-register filaments with a longitudinal symmetry axis, with eight fibrillin molecules in cross section. We present a model of fibrillin alignment that fits all the data and indicates that microfibril extensibility follows conformation-dependent maturation from an initial head-to-tail alignment to a stable approximately one-third staggered arrangement.
Fibrillin is the principal structural component of the 10-12 nm diameter elastic microfibrils of the extracellular matrix. We have previously shown that both fibrillin molecules and assembled ...microfibrils are susceptible to degradation by serine proteases. In this study, we have investigated the potential catabolic effects of six matrix metalloproteinases (MMP-2, MMP-3, MMP-9, MMP-12, MMP-13 and MMP-14) on fibrillin molecules and on intact fibrillin-rich microfibrils isolated from ciliary zonules. Using newly synthesized recombinant fibrillin molecules, major cleavage sites within fibrillin-1 were identified. In particular, the six different MMPs generated a major degradation product of approximately 45 kDa from the N-terminal region of the molecule, whereas treatment of truncated, unprocessed and furin-processed C-termini also generated large degradation products. Introduction of a single ectopia lentis-causing amino acid substitution (E2447K; one-letter symbols for amino acids) in a calcium-binding epidermal growth factor-like domain, predicted to disrupt calcium binding, markedly altered the pattern of C-terminal fibrillin-1 degradation. However, the fragmentation pattern of a mutant fibrillin-1 with a comparable E-->K substitution in an upstream calcium-binding epidermal growth factor-like domain was indistinguishable from wild-type molecules. Ultrastructural examination highlighted that fibrillin-rich microfibrils isolated from ciliary zonules were grossly disrupted by MMPs. This is the first demonstration that fibrillin molecules and fibrillin-rich microfibrils are degraded by MMPs and that certain amino acid substitutions change the fragmentation patterns. These studies have important implications for physiological and pathological fibrillin catabolism and for loss of connective tissue elasticity in ageing and disease.
England has experienced a heavy burden of COVID-19, with multiple waves of SARS-CoV-2 transmission since early 2020 and high infection levels following the emergence and spread of Omicron variants ...since late 2021. In response to rising Omicron cases, booster vaccinations were accelerated and offered to all adults in England. Using a model fitted to more than 2 years of epidemiological data, we project potential dynamics of SARS-CoV-2 infections, hospital admissions and deaths in England to December 2022. We consider key uncertainties including future behavioural change and waning immunity and assess the effectiveness of booster vaccinations in mitigating SARS-CoV-2 disease burden between October 2021 and December 2022. If no new variants emerge, SARS-CoV-2 transmission is expected to decline, with low levels remaining in the coming months. The extent to which projected SARS-CoV-2 transmission resurges later in 2022 depends largely on assumptions around waning immunity and to some extent, behaviour, and seasonality.
Genomic imprinting refers to the pattern of monoallelic parent-of-origin-dependent gene expression where one of the two alleles at a locus is expressed and the other silenced. Although some genes in ...mice are known to be imprinted, the true scope of imprinting and its impact on the genetic architecture of a wide range of morphometric traits is mostly unknown. We therefore searched for quantitative trait loci (QTL) exhibiting imprinting effects on mandible size and shape traits in a large F(3) population of mice originating from an intercross of the LG/J (Large) and SM/J (Small) inbred strains. We discovered a total of 51 QTL affecting mandible size and shape, 6 of which exhibited differences between reciprocal heterozygotes, the usual signature of imprinting effects. However, our analysis showed that only one of these QTL (affecting mandible size) exhibited a pattern consistent with true imprinting effects, whereas reciprocal heterozygote differences in the other five all were due to maternal genetic effects. We concluded that genomic imprinting has a negligible effect on these specific morphometric traits, and that maternal genetic effects may account for many of the previously reported instances of apparent genomic imprinting.
Non-pharmaceutical interventions (NPIs) are used to reduce transmission of SARS coronavirus 2 (SARS-CoV-2) that causes coronavirus disease 2019 (COVID-19). However, empirical evidence of the ...effectiveness of specific NPIs has been inconsistent. We assessed the effectiveness of NPIs around internal containment and closure, international travel restrictions, economic measures, and health system actions on SARS-CoV-2 transmission in 130 countries and territories.
We used panel (longitudinal) regression to estimate the effectiveness of 13 categories of NPIs in reducing SARS-CoV-2 transmission using data from January to June 2020. First, we examined the temporal association between NPIs using hierarchical cluster analyses. We then regressed the time-varying reproduction number (R
) of COVID-19 against different NPIs. We examined different model specifications to account for the temporal lag between NPIs and changes in R
, levels of NPI intensity, time-varying changes in NPI effect, and variable selection criteria. Results were interpreted taking into account both the range of model specifications and temporal clustering of NPIs.
There was strong evidence for an association between two NPIs (school closure, internal movement restrictions) and reduced R
. Another three NPIs (workplace closure, income support, and debt/contract relief) had strong evidence of effectiveness when ignoring their level of intensity, while two NPIs (public events cancellation, restriction on gatherings) had strong evidence of their effectiveness only when evaluating their implementation at maximum capacity (e.g. restrictions on 1000+ people gathering were not effective, restrictions on < 10 people gathering were). Evidence about the effectiveness of the remaining NPIs (stay-at-home requirements, public information campaigns, public transport closure, international travel controls, testing, contact tracing) was inconsistent and inconclusive. We found temporal clustering between many of the NPIs. Effect sizes varied depending on whether or not we included data after peak NPI intensity.
Understanding the impact that specific NPIs have had on SARS-CoV-2 transmission is complicated by temporal clustering, time-dependent variation in effects, and differences in NPI intensity. However, the effectiveness of school closure and internal movement restrictions appears robust across different model specifications, with some evidence that other NPIs may also be effective under particular conditions. This provides empirical evidence for the potential effectiveness of many, although not all, actions policy-makers are taking to respond to the COVID-19 pandemic.
Scanning acoustic microscopy is potentially a powerful tool for characterizing the elastic properties of soft biological tissues and cells. In this paper, we present a method, multi-layer phase ...analysis (MLPA), which can be used to extract local speed of sound values, for both thin tissue sections mounted on glass slides and cultured cells grown on cell culture plastic, with a resolution close to 1 μm. The method exploits the phase information that is preserved in the interference between the acoustic wave reflected from the substrate surface and internal reflections from the acoustic lens. In practice, a stack of acoustic images are captured beginning with the acoustic focal point 4 μm above the substrate surface and moving down in 0.1-μm increments. Scanning parameters, such as acoustic wave frequency and gate position, were adjusted to obtain optimal phase and lateral resolution. The data were processed offline to extract the phase information with the contribution of any inclination in the substrate removed before the calculation of sound speed. Here, we apply this approach to both skin sections and fibroblast cells, and compare our data with the V(f) (voltage versus frequency) method that has previously been used for characterization of soft tissues and cells. Compared with the V(f) method, the MPLA method not only reduces signal noise but can be implemented without making a priori assumptions with regards to tissue or cell parameters.
Aortic wall remodelling is a key feature of both ageing and genetic connective tissue diseases, which are associated with vasculopathies such as Marfan syndrome (MFS). Although the aorta is a 3D ...structure, little attention has been paid to volumetric assessment, primarily due to the limitations of conventional imaging techniques. Phase-contrast microCT is an emerging imaging technique, which is able to resolve the 3D micro-scale structure of large samples without the need for staining or sectioning.
Here, we have used synchrotron-based phase-contrast microCT to image aortae of wild type (WT) and MFS
mice aged 3, 6 and 9 months old (n=5). We have also developed a new computational approach to automatically measure key histological parameters.
This analysis revealed that WT mice undergo age-dependent aortic remodelling characterised by increases in ascending aorta diameter, tunica media thickness and cross-sectional area. The MFS aortic wall was subject to comparable remodelling, but the magnitudes of the changes were significantly exacerbated, particularly in 9 month-old MFS mice with ascending aorta wall dilations. Moreover, this morphological remodelling in MFS aorta included internal elastic lamina surface breaks that extended throughout the MFS ascending aorta and were already evident in animals who had not yet developed aneurysms.
Our 3D microCT study of the sub-micron wall structure of whole, intact aorta reveals that histological remodelling of the tunica media in MFS could be viewed as an accelerated ageing process, and that phase-contrast microCT combined with computational image analysis allows the visualisation and quantification of 3D morphological remodelling in large volumes of unstained vascular tissues.
Fibulin-5 plays an important role in elastic fibre formation in vivo. We have investigated the molecular interactions between fibulin-5 and components of fibrillin-rich microfibrils which form a ...template for elastin. Fibulin-5 interacted in a dose-dependent manner with a fibrillin-1 N-terminal sequence and with tropoelastin, but not with MAGP-1 (microfibril-associated glycoprotein-1) or decorin. Fibulin-5 did not inhibit interactions between fibrillin-1 N- and C-terminal fragments, or fibrillin-1 interactions with tropoelastin. Fibulin-5 may provide a link between tropoelastin and microfibrils in the pericellular space during elastic fibre assembly.
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