Biology subjects

Hopkinson, M.

Publications and source records attributed to Hopkinson, M..

4 recordsLinked to original sources

bioRxiv2026

Regulation and function of trans-physeal growth plate bridges: evidence for a mechanical base isolation role to minimise epiphyseal shear stress

The cartilaginous growth plate (GP) is responsible for all bone elongation during post-natal growth yet must simultaneously contribute to mechanical epiphyseal stability for articulation. How the GP balances these dual functions across the bone-cartilage-bone epiphyseal interface during complex load-growth transitions is not defined. Herein, we examine regulation and mechanobiology of GP bridges - mineralised trans-physeal GP structures - to explore whether they serve these dual GP functions. We have determined the effects of age and sex, short- and long-term joint loading and several established and new osteotropic pharmacological agents on mouse tibial GP bridge number and areal density using micro-computed tomography. We also explored temporal formation and progression of GP bridges by serial in vivo scanning and we imaged epiphyseal load-transfer in young and mature mice in situ via synchrotron X-ray computed tomography (sCT) of intact joints under physiologically oriented load. Our utilisation of digital volume correlation revealed regional 3D load-induced strain inhomogeneities in the GP that are synchronised to bridge location and this was substantiated using finite element modelling. Furthermore, direct micro- and sCT examinations showed that bridges consistently contain a singular epiphyseal/metaphyseal discontinuity which appear to serve a novel mechanical base isolation role to minimise shear stress across the GP. Our data indicate that bridges are regulatable, dynamic structures that synchronise GP strains and exhibit sensitivity to local joint mechanics. We highlight that trans-physeal bridges may contribute to longitudinal bone growth cessation whilst simultaneously stabilising the epiphysis by absorbing compression and shear strains.

bioengineering↗

bioRxiv2024

Sexually dimorphic effects of prenatal alcohol exposure on the murine skeleton

BackgroundPrenatal alcohol exposure (PAE) can result in lifelong disabilities known as foetal alcohol spectrum disorder (FASD) and is associated with childhood growth deficiencies and increased bone fracture risk. However, the effects of PAE on the adult skeleton remain unclear and any potential sexual dimorphism is undetermined. Therefore, we utilised a murine model to examine sex differences with PAE on in vitro bone formation, and in the juvenile and adult skeleton. MethodsPregnant C57BL/6J female mice received 5% ethanol in their drinking water during gestation. Primary calvarial osteoblasts were isolated from neonatal offspring and mineralised bone nodule formation and gene expression assessed. Skeletal phenotyping of 4- and 12-week-old male and female offspring was conducted by micro-computed tomography ({micro}CT), 3-point bending, growth plate analyses, and histology. ResultsOsteoblasts from male and female PAE mice displayed reduced bone formation, compared to control ([≤]30%). Bglap and Ahsg were upregulated with PAE in both sexes compared to control, whereas Vegfa, Bmp6, Tgfbr1 and Flt1 were downregulated in PAE male osteoblasts only. In 12-week-old mice, {micro}CT analysis revealed a sex and exposure interaction across several trabecular bone parameters. PAE was detrimental to the trabecular compartment in male mice compared to control, yet PAE females were unaffected. Both male and female mice had significant reductions in cortical parameters with PAE. Whilst male mice were negatively affected along the tibia length, females were only distally affected. Posterior cortical porosity was increased in PAE females only. Mechanical testing revealed PAE males had significantly reduced bone stiffness compared to controls; maximum load and yield was reduced in both sexes. PAE had no effect on total body weight or tibial bone length in either sex. However, total growth plate width in male PAE mice compared to control was reduced, whilst female PAE mice were unaffected. 4-week-old mice did not display the altered skeletal phenotype with PAE observed in 12-week-old animals. ConclusionsEvidence herein suggests for the first time that PAE exerts divergent sex effects on the skeleton, possibly influenced by underlying sex specific transcriptional mechanisms of osteoblasts. Establishing these sex differences will support future policies and clinical management of FASD. Plain English summaryPrenatal alcohol exposure (PAE) can lead to a set of lifelong cognitive, behavioural, and physical disabilities known as foetal alcohol spectrum disorder (FASD). FASD is a significant burden on healthcare, justice and education systems, which is set to worsen with rising alcohol consumption rates. FASD children have an increased risk of long bone fracture and adolescents are smaller in stature. However, sex differences and the long-term effects of PAE on the skeleton have not been investigated and was the aim of this study. Using a mouse model of PAE, we examined the function and gene expression of bone-forming cells (osteoblasts). We then analysed the skeletons of male and female mice at 12-weeks-old (adult) and 4-weeks-old (juvenile). PAE reduced osteoblast bone formation in both sexes, compared to control. Differential gene expression was predominantly observed in PAE males and largely involved genes related to blood vessel formation. High resolution x-ray imaging (micro-CT) revealed PAE had a detrimental effect on the inner trabecular bone component in 12-week-old male mice only. Analysis of the outer cortical bone revealed that whilst both male and female PAE mice were negatively affected, anatomical variations were observed. Mechanical testing also revealed differences in bone strength in PAE mice, compared to control. Interestingly, 4-week-old mice did not possess these sex differences observed in our PAE model at 12 weeks of age. Our data suggest PAE has detrimental and yet sex-dependent effects on the skeleton. Establishing these sex differences will support future policies and clinical management of FASD. HighlightsO_LIPrimary calvarial osteoblasts isolated from male and female PAE mice displayed reduced mineralised bone nodule formation and differential gene expression compared to control. C_LIO_LIPAE had a detrimental effect on trabecular bone parameters in 12-week-old male mice only. C_LIO_LIPAE leads to spatial variation in cortical bone parameters and geometry, with male mice negatively affected along the tibia length and female mice only affected at the distal end. C_LIO_LIMechanical testing revealed PAE male mice had significantly reduced bone stiffness compared to controls; PAE in both sexes reduced maximum load and yield. C_LIO_LI4-week-old mice did not display the altered skeletal phenotype with PAE observed in 12-week-old animals. C_LI

cell biology↗

bioRxiv2021

Ablation of the FACIT collagen XII disturbs musculoskeletal ECM organization and causes patella dislocation and myopathy

Collagen XII, belonging to the fibril-associated collagens with interrupted triple helix (FACIT) family, assembles from three identical -chains encoded by the COL12A1 gene. The trimeric molecule consists of three N-terminal noncollagenous NC3 domains joined by disulfide bonds followed by a short interrupted collagen triple helix at the C-terminus. Collagen XII is expressed widely in the musculoskeletal system and mutations in the COL12A1 gene cause an Ehlers-Danlos/myopathy overlap syndrome, which is associated with skeletal abnormalities and muscle weakness. Our study defines the role of collagen XII in patella development using the Col12a1-/- mouse model. Deficiency in Col12a1 expression causes malformed facies patellaris femoris grooves at an early stage, which leads to patella subluxation and growth retardation. Due to the patella subluxation, more muscle fibers with centralized nuclei occur in the quadriceps than in the gastrocnemius muscles indicating a local effect. To further understand the role of collagen XII in the skeletal tissues single cell RNAseq (scRNA-seq) was performed. Comparison of the gene expression in the tenocyte cell sub-population of wild type and Col12a1-/- mice showed that several matrix genes are altered. Finally, we reinvestigated collagen XII deficient patients and observed a patella instability.

genetics↗

bioRxiv2020

Bimodal Whole-Mount Imaging Of Tendon Using Confocal Microscopy And X-Ray Micro-Computed Tomography

BACKGROUND3-dimensional imaging modalities for optically dense connective tissues such as tendons are limited and typically have a single imaging methodological endpoint. Here, we have developed a bimodal procedure that utilises fluorescence-based confocal microscopy and x-ray micro-computed tomography for the imaging of adult tendons to visualise and analyse extracellular sub-structure and cellular composition in small and large animal species. RESULTSUsing fluorescent immunolabelling and optical clearing, we visualised the expression of the basement membrane protein laminin-4 in 3D throughout whole rat Achilles tendons and equine superficial digital flexor tendon 5 mm segments. This revealed a complex network of laminin-4 within the tendon core that predominantly localises to the interfascicular matrix compartment. Furthermore, we implemented a chemical drying process capable of creating contrast densities enabling visualisation and quantification of both fascicular and interfascicular matrix volume and thickness by x-ray micro-computed tomography. We also demonstrated that both modalities can be combined using reverse clarification of fluorescently labelled tissues prior to chemical drying to enable bimodal imaging of a single sample. CONCLUSIONSWhole-mount imaging of tendon allowed us to identify the presence of an extensive network of laminin-4 within tendon, the complexity of which cannot be appreciated using traditional 2D imaging techniques. Creating contrast for x-ray micro-computed tomography imaging of tendon using chemical drying is not only simple and rapid, but also markedly improves on previously published methods. Combining these methods provides the ability to gain spatio-temporal information and quantify tendon substructures to elucidate the relationship between morphology and function.

cell biology↗