Biology subjects

Hoyle, A.

Publications and source records attributed to Hoyle, A..

12 recordsLinked to original sources

bioRxiv2026

Senescent tenocytes drive an age-associated loss of cell-extracellular matrix homeostasis, which can be partially restored using dihydroresveratrol

Tendons are commonly injured, not only in athletes, but also during normal everyday activities, particularly in older age. Tendons are rich in extracellular matrix proteins, many of which are turned over extremely slowly during an individuals lifetime. As a result, tight regulation of the extracellular matrix is essential for tendons to remain resilient to the mechanical load placed on them. However, tendons are prone to age-associated functional decline, characterised by chronic inflammation, accumulation of damaged collagen, and matrix remodelling. These degenerative changes often precede injury, wherein additional inflammation and fibrotic tissue deposition make treatment difficult and reinjury highly likely. Understanding and preventing the causes of tendon functional decline is therefore vital to improving quality of life in the ageing population. Using an equine model, here we show that tendon fibroblasts, or tenocytes, isolated from aged tendons exhibit markers of senescence when cultured in vitro. Further, we describe how senescence in tenocytes drives inflammation, hypercontractility, and dysregulation of tendon matrix components. By replicatively senescing tenocytes isolated from young tendons, we observed that senescent tenocytes instigated proinflammatory signalling, had impaired capacity in wound healing assays, increased contractility, and secreted factors that induced senescence in healthy tenocytes. Further, tenocyte senescence dysregulated matrix turnover both at the gene and protein level. Finally, we demonstrate how senotherapeutic treatment of senescent tenocytes can reduce expression of senescence markers and restore proliferative capacity. Our results uncover systemic links between tendon ageing and cellular senescence, and identify mechanisms and therapeutic strategies for age-associated tendon degeneration.

cell biology↗

bioRxiv2026

Ageing impacts extracellular matrix turnover and remodelling in the kidney

At least 10% of the global population is impacted by chronic kidney disease (CKD) and ageing is a key risk factor. CKD is characterised by the build-up of extracellular matrix and a loss of functional nephrons. However, the mechanisms that maintain matrix homeostasis across the physiological lifespan remain elusive. Using {superscript 1}3C-lysine metabolic labelling, we quantified kidney matrix protein turnover in healthy mice at four timepoints (8, 22, 52, and 78 weeks). We found that basement membrane components, including collagen IV, laminin-521, nidogens and perlecan, were more long-lived over age, with collagen IV half-lives extending from weeks in young kidneys to years in aged kidneys, suggesting a reduced capacity for basement membrane renewal. The half-lives of fibrillar collagens I and III also increased over age up to forty-fold, which is consistent with minimal degradation. In contrast, collagen XV retained rapid turnover despite increased abundance, indicating a persistent role in tissue remodelling. Using peptide location fingerprinting to predict structural alterations and proteolytic processing we identified age-dependent meprin oligomerisation and altered nidogen-laminin interaction states. We predicted structural alterations within assembly domains of collagen VI and reduced accessibility of integrin-binding regions, suggesting altered microfibril organisation and cell-surface binding. Collagen XV had predicted structural changes across the NC1 domain encoding the matrikine restin, consistent with altered protease accessibility and matrikine release during ageing. These findings indicate that age-related kidney fibrosis is primarily caused by impaired matrix degradation, with protease accessibility and altered matrix interactions likely playing key roles in this remodeling process.

systems biology↗

bioRxiv2026

In silico degradomics reveals disease- and endotype-specific alterations in the joint tissue landscape

Tissues dynamically remodel extracellular matrix to maintain homeostasis, alterations in which are an early pathogenic hallmark of disease. Protein degradation, essential for tissue remodelling, is often dismissed as indiscriminate damage, despite evidence of its specificity. A major determinant of protein tissue levels and activity, matrix proteolysis also creates circulating degradation products that are emerging biomarkers, with specific collagen fragments capable of tracking disease severity. Understanding intentional matrix destruction therefore is key to understanding tissue biology. Unbiased, holistic analysis, extending our knowledge beyond ubiquitously expressed collagens, will uncover tissue- and disease-specific remodelling. However, degradomics technical demands, requiring labelling and enrichment for neo-epitopes generated by cleavage events, restricts its inclusion in omics research. Here, we develop an in-silico pipeline (DegrAID) that identifies semi-tryptic peptides in unlabelled/unenriched proteomic datasets, mapping neo-epitopes within matrix domain organization and 3D structure, correlating these with known/predicted protease sites, and applies this to rare patient cohorts. Validation with matched degradomic data showed good conservation across degraded proteins and cleavage sites. Interrogation of multiple, independent cohorts including cartilage, synovial tissue and synovial fluid from osteoarthritis (OA) or rheumatoid arthritis (RA) patients identified distinct degradomes between disease and tissue compartments. Further investigating RA heterogeneity revealed myeloid and lymphoid endotypes that display different treatment responses, have substantially different degradation patterns. Proteoglycans were more degraded in myeloid-RA, while collagens more so in lymphoid-RA, with notable exceptions, and endotype-specific fingerprints were conserved between synovial tissue and fluid. Thus, this tool provides new insights into tissue remodelling by unlocking degradomes from any proteomic dataset. One Sentence SummaryDisease- and endotype- specific degradomes generated from clinical proteomics datasets, reveal distinct tissue remodeling patterns.

bioinformatics↗

bioRxiv2025

Thrombospondin Encapsulated Dense Particles accelerate wound healing and downregulate fibrotic phenotypes

Extracellular particles (EP) released by cells are heterogenous, with extracellular vesicles (EV) believed to be the major component. To investigate different EV subpopulations, we conducted a panel screen of 15 fluorescent EV markers in 4 adherent cell lines. Nanoflow cytometry identifies TSPAN14 as the best EV marker in 3 of 4 cell lines. Surprisingly, less than 50% of EPs are labelled by the top 2 EV markers in half the cell lines. The EV marker negative EPs are detergent resistant, non-lipophilic, dense and have Thrombospondin shells by dSTORM; and were thus named Thrombospondin Encapsulated Dense particles (ThrEDs). ThrEDs are ubiquitous, contain extracellular matrix regulators and are released following cell stress. ThrEDs accelerate scratch-wound closure and downregulate the fibrotic phenotype in Dupuytrens disease patient myofibroblasts. The TSP-1 C-terminus is exposed on the surface of ThrEDs, opening engineering applications.

cell biology↗

bioRxiv2024

Golgins support extracellular matrix secretion by collectively maintaining the Golgi structure-function relationship

The secretion of extracellular matrix (ECM) proteins is vital to the maintenance of tissue health. One major control point of this process is the Golgi apparatus, whose dysfunction causes numerous connective tissue disorders. Golgi function is tightly linked to its structure, which is maintained by the cytoskeleton and Golgi organising proteins. We sought to investigate the role of two of these organising proteins, the golgins GMAP210 and Golgin-160, in ECM secretion. We found that loss of either protein had distinct impacts on Golgi organisation. GMAP210 loss caused cisternal fragmentation and dilation, alongside the accumulation of tubulovesicular structures. Meanwhile, Golgin-160 knockout lead to Golgi fragmentation and vesicle build-up. Nonetheless, loss of each protein had a similar impact on ECM secretion and glycosaminoglycan synthesis. We therefore propose that golgins are collectively required to create the correct physical-chemical space to support efficient ECM protein secretion and modification. This is the first time that Golgin-160 has been shown to be required for ECM secretion. SummaryIn this study, Thompson et al demonstrate that two cis-Golgi golgins, GMAP210 and Golgin-160, have distinct, non-redundant roles in maintaining Golgi organisation and that both are required to support the efficient secretion, assembly, and modification of extracellular matrix proteins.

cell biology↗

bioRxiv2024

Circadian and permanent pools of extracellular matrix co-exist in tendon tissue, but have distinct rates of turnover and differential responses to ageing

Heavy carbon isotopes in the tendons of people who grew up in the age of nuclear bomb testing have shown that the extracellular matrix (ECM), assembled during development, stays with us for life. However, recent work suggests that type-I collagen in ECM-rich mouse tendon exists in two pools: a permanent matrix, and a more soluble, circadian-regulated matrix. Despite this, the underlying regulation of such distinct pools is not understood. Here, we demonstrate using stable isotope labelling coupled with mass spectrometry proteomics that circadian and permanent matrix pools have significantly different half-lives. Furthermore, the properties of the matrix pools are altered during development and ageing. Tail tendon tissue was harvested from mice fed on a heavy-lysine diet; protein was then extracted for analysis using a sequential two-step protocol. The first, soluble fraction ( F1) was found to contain intracellular proteins, and a range of core and associated extracellular matrix proteins, including a pool of type-I collagen shown to be circadian-regulated. The remaining fraction ( F2) contained primarily collagens, including type-I collagen which did not show rhythmicity. In adult mice, matrix proteins extracted in the F1 pool had significantly shorter half-lives than F2, including type-I collagen which had half-lives of 4 {+/-} 2 days in F1, compared to 700 {+/-} 100 days in F2. Circadian-regulated matrix proteins were found to have significantly faster turnover than non-circadian in adult mice, but this distinction was lost in older animals. This work identifies protein turnover as the underlying mechanism for the circadian/permanent model of tendon matrix, and suggests a loss of circadian regulation as a characteristic of ECM ageing.

molecular biology↗

bioRxiv2024

Collagen fibril formation at the plasma membrane occurs independently from collagen secretion

Collagen fibrils are the primary supporting scaffold of vertebrate tissues but how they are assembled is unclear. Here, using CRISPR-tagging of type I collagen and SILAC labelling, we elucidate the cellular mechanism for the spatiotemporal assembly of collagen fibrils, in cultured fibroblasts. Our findings reveal multifaceted trafficking of collagen, including constitutive secretion, intracellular pooling, and plasma membrane-directed fibrillogenesis. Notably, we differentiate the processes of collagen secretion and fibril assembly and identify the crucial involvement of endocytosis in regulating fibril formation. By employing Col1a1 knockout fibroblasts we demonstrate the incorporation of exogenous collagen into nucleation sites at the plasma membrane through these recycling mechanisms. Our study sheds light on the assembly process and its regulation in health and disease. Mass spectrometry data are available via ProteomeXchange with identifier PXD036794.

cell biology↗

bioRxiv2023

Optimising a self-assembling peptide hydrogel as a Matrigel alternative for 3-dimensional mammary epithelial cell culture

Three-dimensional (3D) organoid models have been instrumental in understanding molecular mechanisms responsible for many cellular processes and diseases. However, established organic biomaterial scaffolds used for 3D hydrogel cultures, such as Matrigel, are biochemically complex and display significant batch variability, limiting reproducibility in experiments. Recently, there has been significant progress in the development of synthetic hydrogels for in vitro cell culture that are reproducible, mechanically tuneable, and biocompatible. Self-assembling peptide hydrogels (SAPHs) are synthetic biomaterials that can be engineered to be compatible with 3D cell culture. Here we investigate the ability of PeptiGel(R) SAPHs to model the mammary epithelial cell (MEC) microenvironment in vitro. The positively charged PeptiGel(R)Alpha4 supported MEC viability, but did not promote formation of polarised acini. Modifying the stiffness of PeptiGel(R) Alpha4 stimulated changes in MEC viability and changes in protein expression associated with altered MEC function, but did not fully recapitulate the morphologies of MECs grown in Matrigel. To supply the appropriate biochemical signals for MEC organoids, we supplemented PeptiGels(R) with laminin. Laminin was found to require negatively charged PeptiGel(R) Alpha7 for functionality, but was then able to provide appropriate signals for correct MEC polarisation and expression of characteristic proteins. Thus, optimisation of SAPH composition and mechanics allows tuning to support tissue-specific organoids.

cell biology↗

bioRxiv2022

Molecular landscape and functional characterization of centrosome amplification in ovarian cancer

High grade serous ovarian carcinoma (HGSOC) is characterised by poor outcome and extreme chromosome instability (CIN). Therapies targeting centrosome amplification (CA), a key mediator of chromosome missegregation and CIN, may have significant clinical utility in HGSOC. However, the prevalence of CA in HGSOC, its relationship to genomic biomarkers of CIN and its potential impact on therapeutic response have not been defined. Using high-throughput multi-regional microscopy on 287 clinical HGSOC tumour tissues and 73 ovarian cancer cell lines, we show that CA through centriole overduplication is a highly recurrent and heterogeneous feature of HGSOC and is strongly associated with CIN and genome subclonality. Cell-based studies showed that high prevalence CA is phenocopied in ovarian cancer cell lines, and that high CA is associated with increased multi-treatment resistance; most notably to paclitaxel which is the most common treatment used in HGSOC. CA in HGSOC may therefore present a potential driver of tumour evolution and a powerful biomarker for response to standard-of-care treatment.

cancer biology↗

bioRxiv2018

Phospho-regulation of Atoh1 is required for plasticity of secretory progenitors and tissue regeneration

The intestinal epithelium is maintained by a small number of self-renewing stem cells in homeostasis. In addition committed progenitors can contribute to the functional stem cell compartment at a low level during homeostasis and substantially during regeneration following tissue damage. However the mechanism of, and requirement for, progenitor plasticity in mediating pathological response has not been demonstrated. Here we show that multisite phosphorylation of the transcription factor Atoh1 is required both for the contribution of secretory progenitors to the intestinal stem cell pool and for a robust regenerative response following damage. In lineage tracing experiments Atoh1+ cells (Atoh1(WT)CreERT2 mice) show stem cell activity by giving rise to multilineage intestinal clones both in the steady state and after tissue damage. Notably in the colonic epithelium a single generation of Atoh1+ progenitors sustains 1 in 15 stem cells. In an activating Atoh1(9S/T-A)CreERT2 line, the loss of phosphorylation sites on the Atoh1 protein promotes secretory differentiation and inhibits the contribution of these cells to self-renewal. Finally, in a chemical colitis model the Atoh1+ cells of Atoh1(9S/T-A)CreERT2 mice have reduced clonogenic capacity that impacts overall regenerative response of the epithelium. Thus progenitor plasticity plays an integral part in maintaining robust self-renewal in the intestinal epithelium and the balance between stem and progenitor fate behaviour is directly co-ordinated by Atoh1 multi-site phosphorylation.

cell biology↗

bioRxiv2018

Drosophila Toll links systemic immunity to long-term intestinal epithelial integrity

The intestine is an organ where immune and metabolic functions are co-ordinated with tissue renewal via progenitor somatic stem cells (PSSCs). How this is achieved is still unclear. We report that in Drosophila, a generalised infection increased PSSC numbers. This was mimicked by expressing a constitutive form of the immune receptor Toll in PSSCs and blocked when Toll was silenced via RNAi. Without infection, absence of bacterial recognition and downstream Toll signalling resulted in a short lifespan and an age-dependent decrease of PSSCs and gut microbiota. The latter implied a metabolic environment incompatible with the presence of bacteria. Indeed, infection or constitutive Toll signalling in PSSCs triggered 4E-BP transcription in enterocytes, while loss of signalling reduced it. 4E-BP controlled fat levels and sustained the microbiota suggesting that Toll-dependent regulation of 4E-BP was important for long-term gut function. Therefore, the Toll pathway is crucial for responses to both infection and microbiota.

immunology↗

bioRxiv2017

Copy-number signatures and mutational processes in ovarian carcinoma

Tumours with profound copy-number aberration elude molecular stratification due to their genomic complexity. By representing this complexity as a mixture of copy-number signatures, we provide molecular explanations for differing clinical outcomes. Here we present a method for copy-number signature identification, deriving eight signatures in 117 shallow whole-genome sequenced high-grade serous ovarian cancers (HGSOC), which validated on independent cohorts of 95 deep whole-genome sequenced, and 402 SNP array-profiled cases. Three copy-number signatures predicted longer overall survival, while the others predicted poorer outcome. We found evidence for the mutational processes giving rise to copy-number change for six of the eight signatures via correlations with other genomic features. Our results provide insights into the pathogenesis of HGSOC by uncovering multiple mutational processes that shape genomes following TP53 mutation. Importantly, our work shows that most HGSOC have a mixture of mutational processes suggesting that targeting a single mutator phenotype may be therapeutically suboptimal.

cancer biology↗