Biology subjects

Sherratt, M. J.

Publications and source records attributed to Sherratt, M. J..

10 recordsLinked to original sources

bioRxiv2025

Comparative circadian transcriptome analysis reveals dampened and phase-advanced rhythms in sun-exposed human skin

BackgroundDaily molecular rhythms modulate skin physiology. However, the effects of chronic sun exposure on these rhythms remain unstudied. ObjectivesThis study aimed to identify and compare rhythmic genes and pathways in photoprotected and chronically photoexposed human skin in vivo. MethodsTwenty healthy White women, aged 51-63, with moderate-severe photoageing of the dorsal forearm were recruited. Skin biopsies (3mm) were taken from photoprotected (upper buttock) and photoexposed (dorsal forearm) skin of each individual at noon, 6PM, midnight, and 6AM, across a 24-hour cycle. Skin biopsies were analysed by RNA sequencing. Cosinor analysis identified cycling genes along with their amplitudes and peak expression phases. Nested models were used to identify genes that were differentially rhythmic between the photoprotected and photoexposed sites. Phase set and gene set analyses identified pathways enriched among rhythmic transcripts or altered between the two sites. ResultsIn the photoprotected buttock skin, 1546 genes (12%) met the criteria for cycling. In photoexposed forearm skin, the number was reduced to 959 (8%). As a group, transcripts that cycled in both sites had overall higher amplitude in photoprotected skin (p < 2.2e-16). Peak expression times for these transcripts showed a pronounced bimodal distribution and were clustered in the early morning and mid-afternoon. Distributions of peak times were significantly different between photoexposed and photoprotected skin (p < 0.00025), with peak times advanced in photoexposed skin. We identified 480 genes with significantly different rhythmic properties between the skin sites. Genes involved in DNA repair, MYC targets, E2F and G2M checkpoint pathways were enriched among those that showed higher amplitude oscillations in photoprotected skin. Genes involved in epithelial mesenchymal transition and apical junction pathways showed higher amplitude oscillations in photoexposed skin. ConclusionsTemporal rhythms have a marked influence on skin molecular physiology and are altered in photoaged skin. Temporally advanced cycling patterns and a reduced number of rhythmic genes in photoexposed as compared to photoprotected skin suggest that chronic UV exposure may disrupt and/or reprogram circadian output rhythms to further alter skin physiology.

cell biology↗

bioRxiv2025

Multimodal X-ray imaging reveals hierarchical fibre mechanics

Fibrous materials--ranging from connective tissues to engineered composites--are vital to many biological and man-made systems, optimised to withstand complex in-operando or in-vivo loading. The spines intervertebral discs (IVD) load-bearing capacity depends on a hierarchical extracellular matrix, where plywood-like lamellae of collagen fibres in the annulus fibrosus contain nanometre-scale fibrils built from staggered triple-helical monomers. How intact IVDs couple fibril-scale mechanics to fibre-scale organisation under load remains unresolved. Here we introduce TomoSAXS, a full-field 3D small-angle X-ray scattering tomography that maps fibril-to-fibre mechanics across an intact tissue. We show that intrafibrillar molecular pre-strain (D-period stagger) is lamellar textured and tightly correlated with microscale fibre strain. Pre-strain is inversely related to fibril strain and its variability, consistent with load-sharing through molecular unwinding. Radial strain bridges and high-curvature zones at the annulus fibrosus-nucleus pulposus interfaces emerge as critical regulators of local mechanics. These findings reveal concerted fibril-fibre interactions that sustain mechanical equilibrium in the IVD, preserving elasticity and shape. More broadly, TomoSAXS establishes a platform to visualise nano- to micro-scale matrix mechanics across biological and synthetic fibrous materials, with applications in ageing and disease, therapeutic evaluation, and the design of bio-based and bioinspired materials.

bioengineering↗

bioRxiv2023

Novel in-silico predicted matrikines are differential mediators of in vitro and in vivo cellular metabolism

The exogenous application of small peptides can beneficially affect clinical skin appearance (wrinkles) and architecture (collagen and elastic fibre deposition and epidermal thickness). However, the discovery of new bioactive peptides has not been underpinned by any guiding hypothesis. As endogenous extracellular matrix (ECM)-derived peptides produced during tissue remodelling can act as molecular signals influencing cell metabolism, we hypothesised that protease cleavage site prediction could identify putative novel matrikines with beneficial activities. Here, we present an in silico to in vivo discovery pipeline, which enables the prediction and characterisation of peptide matrikines which differentially influence cellular metabolism in vitro. We use this pipeline to further characterise a combination of two novel ECM peptide mimics (GPKG and LSVD) which act in vitro to enhance the transcription of ECM organisation and cell proliferation genes and in vivo to promote epithelial and dermal remodelling. This pipeline approach can both identify new matrikines and provide insights into the mechanisms underpinning tissue homeostasis and repair.

cell biology↗

bioRxiv2022

Fibrillin microfibril structure identifies long-range effects of inherited pathogenic mutations affecting a key regulatory TGFβ-binding site

Genetic mutations in fibrillin microfibrils cause a range of serious inherited diseases such as Marfan syndrome (MFS) and Weill-Marchesani syndrome (WMS). These diseases typically show major dysregulation of tissue development and growth, particularly in skeletal long bones, but links between the mutations and the diseases are unknown. In this study we reveal the detailed cryo-EM structure of native fibrillin microfibrils from mammalian tissue. The major bead region showed pseudo 8-fold symmetry and a buried protease resistant N-terminal core. Based on this structure, we show a WMS deletion mutant induces a rearrangement with long-range effects blocking interaction with latent TGF{beta}-binding protein (LTBP)-1 at a remote site. Separate deletion of this binding site resulted in the assembly of shorter fibrillin microfibrils with structural alterations. These results establish that in complex extracellular protein assemblies, such as in fibrillin, mutations may have long-range structural consequences to disrupt growth factor signalling and cause disease.

biochemistry↗

bioRxiv2022

Peptide location fingerprinting identifies species- and tissue-conserved structural remodelling of proteins as a consequence of ageing and disease

Extracellular matrix (ECM) in the intervertebral disc (IVD), lung and artery are thought to undergo the age-dependant accumulation of damage by chronic exposure to mechanisms such as reactive oxygen species, proteases and glycation. It is unknown whether this damage accumulation is species-dependant (via differing lifespans and hence cumulative exposures) or whether it can influence the progression of age-related diseases such as atherosclerosis. Peptide location fingerprinting (PLF) is a new proteomic analysis method, capable of the non-targeted identification of structure-associated changes within proteins. Here we applied PLF to publicly available ageing human IVD (outer annulus fibrosus), ageing mouse lung and human arterial atherosclerosis datasets and identified novel target proteins alongside common age-associated differences within protein structures which were conserved between tissue regions, organs, sexes and species and in age-related disease. We identify peptide yield differences across protein structures which coincide with biological regions, potentially reflecting the functional consequences of ageing or atherosclerosis for macromolecular assemblies (collagen VI and fibrin), enzyme/inhibitor activity (cathepsin B and alpha-2 macroglobulin), activation states (complement C3 and thrombin) and interaction states (laminins, perlecan, fibronectin, filamin-A, collagen XIV and apolipoprotein-B). Furthermore, we show that alpha-2 macroglobulin, prothrombin, collagen XIV and apolipoprotein-B all exhibit possible shared structural consequences in IVD ageing and arterial atherosclerosis, providing novel links between an age-related disease and intrinsic ageing. Crucially, we also demonstrate that fibronectin, laminin beta chains and filamin-A all exhibit conserved age-associated structural differences between mouse lung and human IVD, providing evidence that ECM, and their associating proteins, may be subjected to potentially similar mechanisms or consequences of ageing across species, irrespective of differences in lifespan and tissue function.

molecular biology↗

bioRxiv2021

Matrix-Bound Growth Factors are Released upon Cartilage Compression by an Aggrecan-Dependent Sodium Flux that is Lost in Osteoarthritis

Articular cartilage is a dense extracellular matrix-rich tissue that degrades following chronic mechanical stress, resulting in osteoarthritis (OA). The tissue has low intrinsic repair especially in aged and osteoarthritic joints. Here we describe three pro-regenerative factors; fibroblast growth factor 2 (FGF2), connective tissue growth factor, bound to transforming growth factor-beta (CTGF-TGF{beta}), and hepatoma-derived growth factor (HDGF), that are rapidly released from the pericellular matrix (PCM) of articular cartilage upon mechanical injury. All three growth factors bound heparan sulfate, and were displaced by exogenous NaCl. We hypothesised that sodium, sequestered within the aggrecan-rich matrix, was freed by injurious compression, thereby enhancing the bioavailability of pericellular growth factors. Indeed, growth factor release was abrogated when cartilage aggrecan was depleted by IL-1 treatment, and in severely damaged human osteoarthritic cartilage. A flux in free matrix sodium upon mechanical compression of cartilage was visualised by 23Na magnetic resonance imaging (MRI) just below the articular surface. This corresponded to a region of reduced tissue stiffness, measured by scanning acoustic microscopy and second harmonic generation microscopy, and where Smad2/3 was phosphorylated upon cyclic compression. Our results describe a novel intrinsic repair mechanism, controlled by matrix stiffness and mediated by the free sodium concentration, in which heparan sulfate-bound growth factors are released from cartilage upon injurious load. They identify aggrecan as a depot for sequestered sodium, explaining why osteoarthritic tissue loses its ability to repair. Treatments that restore matrix sodium to allow appropriate release of growth factors upon load are predicted to enable intrinsic cartilage repair in osteoarthritis. Significance StatementOsteoarthritis is the most prevalent musculoskeletal disease, affecting 250 million people worldwide 1. We identify a novel intrinsic repair response in cartilage, mediated by aggrecan-dependent sodium flux, and dependent upon matrix stiffness, which results in the release of a cocktail of pro-regenerative growth factors after injury. Loss of aggrecan in late-stage osteoarthritis prevents growth factor release and likely contributes to disease progression. Treatments that restore matrix sodium in osteoarthritis may recover the intrinsic repair response to improve disease outcome.

cell biology↗

bioRxiv2020

Peptide location fingerprinting reveals modification-associated biomarkers of ageing in human tissue proteomes

Although dysfunctional protein homeostasis (proteostasis) is a key factor in many age-related diseases, the untargeted identification of structural modifications in proteins remains challenging. Peptide location fingerprinting is a proteomic analysis technique capable of identifying structural modification-associated differences in mass spectrometry (MS) datasets of complex biological samples. A new webtool (Manchester Peptide Location Fingerprinter), applied to photoaged and intrinsically aged skin proteomes, can relatively quantify peptides (spectral counting) and map statistically significant differences to regions within protein structures. New photoageing biomarkers were identified in multiple proteins including matrix components (collagens and proteoglycans), oxidation and protease modulators (peroxiredoxins and SERPINs) and cytoskeletal proteins (keratins). Crucially, for many extracellular biomarkers, structural modification-associated differences were not correlated with relative abundance (by ion intensity). By applying peptide location fingerprinting to published MS datasets, (identifying biomarkers including collagen V and versican in ageing tendon) we demonstrate the potential of the MPLF webtool to discover novel biomarkers.

molecular biology↗

bioRxiv2020

Loss of PKCα increases arterial medial calcification in a uremic mouse model of chronic kidney disease

Arterial medial calcification is an independent risk factor for mortality in chronic kidney disease. We previously reported that knock-down of PKC expression increases high phosphate-induced mineral deposition by vascular smooth muscle cells in vitro. This new study tests the hypothesis that PKC regulates uremia-induced medial calcification in vivo. Female wild-type and PKC-/- mice underwent a two-stage subtotal nephrectomy and were fed a high phosphate diet for 8 weeks. X-ray micro computed tomography demonstrated that uremia-induced medial calcification was increased in the abdominal aorta and aortic arch of PKC-/- mice compared to wild-types. Blood urea nitrogen was also increased in PKC-/- mice compared to wild-types; there was no correlation between blood urea nitrogen and calcification in PKC-/- mice. Phosphorylated SMAD2 immunostaining was detected in calcified aortic arches from uremic PKC-/- mice; the osteogenic marker Runx2 was also detected in these areas. No phosphorylated SMAD2 staining were detected in calcified arches from uremic wild-types. PKC knock-down increased TGF-{beta}1-induced SMAD2 phosphorylation in vascular smooth muscle cells in vitro, whereas the PKC activator prostratin decreased SMAD2 phosphorylation. In conclusion, loss of PKC increases uremia-induced medial calcification. The PKC/TGF-{beta} signaling axis could therefore represent a new therapeutic target for arterial medial calcification in chronic kidney disease.

cell biology↗

bioRxiv2019

Estrogen mediates acute elastic fibre homeostasis in skin

Remodelling of the dermal extracellular matrix makes a major contribution to skin fragility in the elderly. The peri-menopausal period in females is also associated with an age-like phenotype which can be reversed by hormone replacement therapy. This suggests a direct link between circulating hormone levels and tissue ageing. Despite work investigating the role of estrogen as a regulator of collagen fibril abundance and structure, the influence of estrogen on the elastic fibre system remains poorly defined. Here we used an ovariectomised (Ovx) mouse surgical menopause model to show that just 7 weeks of acute hormone deficiency significantly decreased skin tensile strength and elasticity. Systemic replacement of 17{beta}-estradiol to physiological levels protected against these changes to the skin mechanical properties. Moreover, acute hormone deficiency differentially influenced dermal structural networks, significantly decreasing dermal elastic fibre abundance without discernible effect on collagen fibril organisation or abundance. We suggest that this specific elastic fibre proteolysis may be driven by extracellular protease activity, or be a consequence of significant adipocyte hypertrophy. 17{beta}-estradiol supplementation in Ovx mice in vivo protected the elastic fibre system. Treatment of human dermal fibroblasts with 17{beta}-estradiol in vitro induced the selective upregulation of tropoelastin, fibrillin-1 and associated elastic fibre-associated proteins (including EMILINs and fibulins). In summary, these data show that the elastic fibre system is significantly perturbed by estrogen deprivation. Thus, pharmacological intervention may slow the acute effects of menopause and potentially the chronic effects of ageing in skin.

biophysics↗

bioRxiv2019

Distinct matrix composition and mechanics in aged and estrogen-deficient mouse skin

Hormone deficiency has been widely linked to accelerated tissue ageing, and increased incidence of chronic degenerative disease. Furthermore, age-associated hormonal dysregulation is thought to be a major contributing factor to the increased fragility of aged skin. The ageing process is driven by an aggregation of damage to cells and extracellular matrix, which can directly influence the mechanical properties of the tissue. Here we report on the correlation between mechanical properties and composition of skin from ovariectomised and aged mice, to assess the extent to which estrogen deprivation drives dermal ageing. We found that age and estrogen abrogation affected skin mechanical properties in contrasting ways: ageing led to increased tensile strength and stiffness while estrogen deprivation had the opposite effect. Mass spectrometry proteomics showed that the quantity of extractable fibrillar collagen-I decreased with ageing, but no change was observed in ovariectomised mice. This observation, in combination with measurements of tensile strength, was interpreted to reflect changes to the extent of extracellular matrix crosslinking, supported by a significant increase in the staining of advanced glycation endpoints in aged skin. Loss of mechanical strength in the skin following ovariectomy was consistent with a loss of elastic fibres. Other changes in extracellular matrix composition broadly correlated between aged and ovariectomised mice, confirming the important role of estrogen-related pathways in ageing. This study offers new insight into the relationship between tissue composition and mechanics, and suggests that the deleterious effects of intrinsic skin ageing are compounded by factors beyond hormonal dysregulation.

biophysics↗