Andrew J Quantock

Cardiff University, Cardiff, Wales, United Kingdom

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Publications (77)233.71 Total impact

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    ABSTRACT: Genome-wide association studies have identified common variants in transcription factor 4 (TCF4) as susceptibility loci for schizophrenia, Fuchs' endothelial corneal dystrophy, and primary sclerosing cholangitis. By contrast, rare TCF4 mutations cause Pitt-Hopkins syndrome, a disorder characterized by intellectual disability and developmental delay, and have also been described in patients with other neurodevelopmental disorders. TCF4 therefore sits at the nexus between common and rare disorders. TCF4 interacts with other basic helix-loop-helix proteins, forming transcriptional networks that regulate the differentiation of several distinct cell types. Here, we review the role of TCF4 in these seemingly diverse disorders and discuss recent data implicating TCF4 as an important regulator of neurodevelopment and epithelial-mesenchymal transition.
    Trends in Molecular Medicine 03/2014; · 9.57 Impact Factor
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    ABSTRACT: Cell-directed deposition of aligned collagen fibrils during corneal embryogenesis is poorly understood, despite the fact that it is the basis for the formation of a corneal stroma that must be transparent to visible light and biomechanically stable. Previous studies of the structural development of the specialized matrix in the cornea have been restricted to examinations of tissue sections by conventional light or electron microscopy. Here, we use volume scanning electron microscopy, with sequential removal of ultrathin surface tissue sections achieved either by ablation with a focused ion beam or by serial block face diamond knife microtomy, to examine the microanatomy of the cornea in three dimensions and in large tissue volumes. The results show that corneal keratocytes occupy a significantly greater tissue volume than was previously thought, and there is a clear orthogonality in cell and matrix organization, quantifiable by Fourier analysis. Three-dimensional reconstructions reveal actin-associated tubular cell protrusions, reminiscent of filopodia, but extending more than 30 µm into the extracellular space. The highly extended network of these membrane-bound structures mirrors the alignment of collagen bundles and emergent lamellae and, we propose, plays a fundamental role in dictating the orientation of collagen in the developing cornea.
    Proceedings of the National Academy of Sciences 01/2014; · 9.81 Impact Factor
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    ABSTRACT: Type VI collagen is a primary component of the extracellular matrix of many connective tissues. It can form distinct aggregates depending on tissue structure, chemical environment, and physiology. In the current study we examine the ultrastructure and mode of aggregation of type VI collagen molecules in the human trabecular meshwork.
    Molecular vision 01/2014; 20:638-48. · 1.99 Impact Factor
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    ABSTRACT: This study investigated changes in collagen fibril architecture and the sulphation status of keratan sulphate (KS) glycosaminoglycan (GAG) epitopes from central to peripheral corneal regions. Freshly excised adult bovine corneal tissue was examined as a function of radial position from the centre of the cornea outwards. Corneal thickness, tissue hydration, hydroxyproline content, and the total amount of sulphated GAG were all measured. High and low-sulphated epitopes of keratan sulphate were studied by immunohistochemistry and quantified by ELISA. Chondroitin sulphate (CS) and dermatan sulphate (DS) distributions were observed by immunohistochemistry following specific enzyme digestions. Electron microscopy and x-ray fibre diffraction were used to ascertain collagen fibril architecture. The bovine cornea was 1021 ± 5.42 μm thick at its outer periphery, defined as 9-12 mm from the corneal centre, compared to 844 ± 8.10 μm at the centre. The outer periphery of the cornea was marginally, but not significantly, more hydrated than the centre (H = 4.3 vs. H = 3.7), and was more abundant in hydroxyproline (0.12 vs. 0.06 mg/mg dry weight of cornea). DMMB assays indicated no change in the total amount of sulphated GAG across the cornea. Immunohistochemistry revealed the presence of both high- and low-sulphated epitopes of KS, as well as DS, throughout the cornea, and CS only in the peripheral cornea before the limbus. Quantification by ELISA, disclosed that although both high- and low-sulphated KS remained constant throughout stromal depth at different radial positions, high-sulphated epitopes remained constant from the corneal centre to outer-periphery, whereas low-sulphated epitopes increased significantly. Both small angle x-ray diffraction and TEM analysis revealed that collagen fibril diameter remained relatively constant until the outer periphery was reached, after which fibrils became more widely spaced (from small angle x-ray diffraction analysis) and of larger diameter as they approached the sclera. Depth-profiled synchrotron microbeam analyses showed that, at different radial positions from the corneal centre outwards, fibril diameter was greater superficially than in deeper stromal regions. The interfibrillar spacing was also higher at mid-depth in the stroma than it was in anterior and posterior stromal regions. Collagen fibrils in the bovine cornea exhibited a fairly consistent spacing and diameter from the corneal centre to the 12 mm radial position, after which a significant increase was seen. While the constancy of the overall sulphation levels of proteoglycans in the cornea may correlate with the fibrillar architecture, there was no correlation between the latter and the distribution of low-sulphated KS.
    Matrix Biology. 01/2014;
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    ABSTRACT: The chemical composition of tissues can influence their form and function. As a prime example, the lattice-like arrangement of collagen fibrils required for corneal transparency is controlled, in part, by sulfated proteoglycans, which, via core proteins, bind to the collagen at specific locations along the fibril axis. However, to date, no studies have been able to directly identify and characterize sulfur (S) in the cornea as a function of tissue location. In this study, X-ray absorption near-edge structure spectroscopy and micro-beam X-ray fluorescence (μ-XRF) chemical contrast imaging were employed to probe the nature of the mature (bovine) cornea as a function of position from the anterior sub-epithelial region into the deep stroma. Data indicate an inhomogeneity in the composition of S species in the first ≈50 μm of stromal depth. In μ-XRF chemical contrast imaging, S did not co-localize with phosphorous (P) in the deep stroma where sulfates are prominent. Rather, P is present only as isolated micrometric spots, presumably identifiable as keratocytes. This study lends novel insights into the elemental physiology of mature cornea, especially in relation to its S distribution; future studies could be applied to human tissues. Moreover, it defines an analytical protocol for the interrogation of S species in biological tissues with micrometric resolution.
    Analytical and Bioanalytical Chemistry 06/2013; · 3.66 Impact Factor
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    ABSTRACT: To examine the effect of riboflavin/UVA corneal crosslinking on stromal ultrastructure and hydrodynamic behaviour. One hundred and seventeen enucleated ungulate eyes (112 pig and 5 sheep) and 3 pairs of rabbit eyes, with corneal epithelium removed, were divided into four treatment groups: Group 1 (28 pig, 2 sheep and 3 rabbits) were untreated; Group 2 (24 pig) were exposed to UVA light (3.04 mW/cm(2)) for 30 minutes and Group 3 (29 pig) and Group 4 (31 pig, 3 sheep and 3 rabbits) had riboflavin eye drops applied to the corneal surface every 5 minutes for 35 minutes. Five minutes after the initial riboflavin instillation, the corneas in Group 4 experienced a 30 minute exposure to UVA light (3.04 mW/cm(2)). X-ray scattering was used to obtain measurements of collagen interfibrillar spacing, spatial order, fibril diameter, D-periodicity and intermolecular spacing throughout the whole tissue thickness and as a function of tissue depth in the treated and untreated corneas. The effect of each treatment on the hydrodynamic behaviour of the cornea (its ability to swell in saline solution) and its resistance to enzymatic digestion were assessed using in vitro laboratory techniques. Corneal thickness decreased significantly following riboflavin application (p<0.01) and also to a lesser extent after UVA exposure (p<0.05). With the exception of the spatial order factor, which was higher in Group 4 than Group 1 (p<0.01), all other measured collagen parameters were unaltered by cross-linking, even within the most anterior 300 microns of the cornea. The cross-linking treatment had no effect on the hydrodynamic behaviour of the cornea but did cause a significant increase in its resistance to enzymatic digestion. It seems likely that cross-links formed during riboflavin/UVA therapy occur predominantly at the collagen fibril surface and in the protein network surrounding the collagen.
    PLoS ONE 01/2013; 8(1):e52860. · 3.53 Impact Factor
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    ABSTRACT: PURPOSE:: To determine the short-term fate of the host endothelium and Descemet membrane after non-Descemet stripping automated endothelial keratoplasty (nDSAEK). METHODS:: Eight unilateral DSAEK (n = 4) or nDSAEK (n = 4) surgeries were performed in the right eyes of 8 rabbits. Corneal transparency and thickness were followed-up by slit-lamp microscopy, and 2 weeks postoperatively, corneas were evaluated by immunohistochemistry and transmission electron microscopy. RESULTS:: Corneas remained clear after both DSAEK and nDSAEK. One week after DSAEK, the stroma-to-stroma surgical interface was identifiable as a zone of fibrotic tissue a few microns thick, whereas in the nDSAEK group, the recipient corneal endothelium and Descemet membrane were clearly visible at the graft-host interface. The retained endothelial cells were positive for Na/K-ATPase but assumed a markedly different morphology from healthy endothelial cells, with cell processes extending into the graft stroma or engulfing strands of irregularly dissected grafted stromal tissue where they occasionally appeared to compartmentalize the transplanted matrix and became detached from the underlying Descemet membrane. CONCLUSIONS:: Host endothelial cells 2 weeks after nDSAEK express markers of pump function, but appear to be morphologically altered, occasionally detaching from the adjacent Descemet membrane, extending into the graft stroma or engulfing strands of the grafted stroma at the interface. The short-term persistence and subsequent phenotypical alternation of residual endothelial cells, aligned to structural changes to Descemet membrane, might influence graft adherence after nDSAEK.
    Cornea 09/2012; · 1.75 Impact Factor
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    ABSTRACT: The chemical composition and sulfur (S) speciation of developing chick corneas at embryonic days 12, 14, and 16 were investigated using synchrotron scanning x-ray fluorescence microscopy and x-ray absorption near-edge structure spectroscopy. The aim was to develop techniques for the analysis of bulk tissue and identify critical physicochemical variations that correlate with changes in corneal structure-function relationships. Derived data were subjected to principal component analysis and linear discriminant analysis, which highlighted differences in the elemental and S species composition at different stages of embryonic growth. Notably, distinct elemental compositions of chlorine, potassium, calcium, phosphorus, and S altered with development during the transition of the immature opaque cornea to a mature transparent tissue. S structure spectroscopy revealed developmentally regulated alterations in thiols, organic monosulfides, ester sulfate, and inorganic sulfate species. The transient molecular structures and compositional changes reported here provide a deeper understanding of the underlying basis of corneal development during the acquisition of transparency. The experimental and analytical approach is new, to our knowledge, and has wide potential applicability in the life sciences.
    Biophysical Journal 07/2012; 103(2):357-64. · 3.67 Impact Factor
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    ABSTRACT: The aim of this study was to develop a method for refining the optical and mechanical properties of human amniotic membrane (AM) to provide ophthalmic transparent implants for use during severe donor cornea shortages. AM was allowed to gradually dehydrate at 4-8 °C with and without chemical cross-linking. To improve the transparency of AM, a simple dehydration process using a refrigerator at 4-8 °C overnight was examined. For further improvements, dehydrated AM was then cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxy-succimide before rehydration. Light transmittance and tensile strength of individual specimens were evaluated. Light transmittance of AM improved from 50.9-77.7% at 550 nm by this simple low temperature dehydration process. Its high light transmittance was partially maintained at 70.1%, even after rehydration with normal saline. Interestingly, chemically cross-linked AM showed a significantly greater light transmittance of 81.5% under wet conditions. In addition, tensile strength was significantly increased after cross-linking from 2.32 N/mm(2) (native tissue) to 11.78 N/mm(2) (cross-linked specimens). Light transmittance and tensile strength were successfully improved by these approaches, including low temperature dehydration with and without chemical cross-linking. The use of refined AM could be feasible for use in current and future ophthalmic treatments. Copyright © 2012 John Wiley & Sons, Ltd.
    Journal of Tissue Engineering and Regenerative Medicine 04/2012; · 4.43 Impact Factor
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    ABSTRACT: This article investigates in vitro light transmission through the human cornea in the ultraviolet (UV) portion of the electromagnetic spectrum as a function of position across the cornea from center to periphery. Spectrophotometry was used to measure UV transmission in the wavelength range 310-400 nm, from the central cornea to its periphery. UV transmission decreases away from the center, and this is attributed to scattering and absorbance. Corneal endothelial cells, which line the back of the cornea and are more numerous in the periphery, therefore receive a lower dose of UV than do those in the central cornea. This is consistent with the recent observation that endothelial cells in the corneal periphery exhibit less nuclear oxidative DNA damage than those in the central cornea.
    Biophysical Journal 03/2012; 102(6):1258-64. · 3.67 Impact Factor
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    ABSTRACT: Biospectroscopy tools are increasingly being recognized as novel approaches toward interrogating complex biological structures in a nondestructive fashion. This study was conducted to apply these tools to interrogate alterations in the molecular signatures of developing chick corneas during the onset and development of transparency. Embryonic chick corneas (n = 46) were obtained at 2-day intervals from embryonic day (E)10 to E18 of incubation and investigated with attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy and Raman microspectroscopy. Resultant spectra were analyzed for variance by using principal component analysis and linear discriminant analysis (PCA-LDA). Mean spectra after ATR-FTIR spectroscopy or Raman microspectroscopy derived from corneas at each developmental stage showed some overlap; however, in PCA-LDA scores plots, a clear segregation of spectra was evident, and two-category discrimination indicated that significant molecular alterations occur during tissue morphogenesis. Notable by both techniques was the increasing intensity of DNA signal (1080 cm⁻¹) from E10 onward. Major segregating biomarkers identified by ATR-FTIR spectroscopy between E10 and E18 were in the DNA/RNA (1126 cm⁻¹), glycogen (1045 cm⁻¹), protein (1470 cm⁻¹), and amide II (1512 cm⁻¹ and 1524 cm⁻¹) spectral regions. Raman spectroscopy also identified major distinguishing vibrational modes that included proteins, amino acids (tyrosine, proline phenylalanine, and valine), and secondary structures of proteins (amide I and amide II). The developing chick cornea undergoes significant changes in its biomolecular composition in the E10 to E18 developmental period, with the major changes occurring in the spectral regions associated with DNA/RNA, proteins, glycogen, and secondary protein structures.
    Investigative ophthalmology & visual science 01/2012; 53(3):1162-8. · 3.43 Impact Factor
  • Biophysical Journal 01/2012; 103(2):357. · 3.67 Impact Factor
  • Investigative ophthalmology & visual science 01/2012; 53(3):1162. · 3.43 Impact Factor
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    ABSTRACT: Our study examined the effect of a selective Rho kinase inhibitor, Y-27632, on corneal wound healing and potential stromal scarring after superficial keratectomy. Rabbit keratocytes were induced into myofibroblasts by transforming growth factor β1 (TGFβ1) either with or without Y-27632. Then α-smooth muscle actin (α-SMA) was examined by immunohistochemistry and western blotting, and the contractility of the seeded collagen gels was measured. Y-27632 eye drops (or vehicle only) were administered to eyes after a superficial keratectomy, and the tissue was examined by immunohistochemistry for α-SMA, collagen types I, II, and III, and keratan sulfate. Electron microscopy was conducted with and without histochemical contrasting of sulfated proteoglycans. Spindle-like cells in culture constituted 99.5±1.1% with TGFβ1 stimulation, but 3.5±1.0% after TGFβ1 and Y-27632 treatment (p<0.01, n=6). α-SMA was seen in 4% of TGFβ1-treated cells, but in only 0.3% of cells with Y-27632 added (p<0.01, n=6), which was confirmed by western blotting. Y-27632 also inhibited the TGFβ1-induced contraction of seeded collagen gels. After superficial keratectomies, collagen type I and keratan sulfate were unchanged by Y-27632 application. Collagen type II was not detected in Y-27632 or vehicle-only corneas. With Y-27632 treatment, α-SMA expression increased and the collagen type III signal became in the weaker subepithelial area. Interestingly, bundles of aligned and uniformly spaced collagen fibrils were more prevalent in keratocytes in Y-27632-treated corneas, which is reminiscent of fibripositor-like structures that have been proposed as a mechanism of matrix deposition in embryonic connective tissues. Y-27632 inhibits keratocyte-to-myofibroblast transition, and its topical application after a superficial lamellar keratectomy elicits an altered wound healing response, with evidence of an embryonic-type deposition of collagen fibrils.
    Molecular vision 01/2012; 18:1727-39. · 1.99 Impact Factor
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    ABSTRACT: We have discovered that a combination of fibroblast growth factor 2 and transforming growth factor β1 induce profound morphologic changes in immature articular cartilage. The purpose of this study was to test the hypothesis that these changes represent accelerated postnatal maturation. Histochemical and biochemical assays were used to confirm the nature of the morphologic changes that accompany growth factor stimulation of immature bovine articular cartilage explants in serum-free culture medium. Growth factor-induced apoptosis, cellular proliferation, and changes in the collagen network were also quantitatively analyzed. Growth factor stimulation resulted in rapid resorption from the basal aspect of immature cartilage explants that was simultaneously opposed by cellular proliferation from the apical aspect driven from a pool of chondroprogenitor cells we have previously described. Maturation-dependent changes in tissue stiffness, collagen crosslinking, and collagen fibril architecture as well as differentiation of the extracellular matrix into distinct pericellular, territorial, and interterritorial domains were all present in growth factor-stimulated cartilage samples and absent in control samples. Our data demonstrate that it is possible to significantly enhance the maturation of cartilage tissue using specific growth factor stimulation. This may have applications in transplantation therapy or in the treatment of diseased cartilage, through phenotype modulation of osteoarthritic chondrocytes in order to stimulate growth and maturation of cartilage repair tissue.
    Arthritis & Rheumatology 07/2011; 63(11):3417-27. · 7.48 Impact Factor
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    ABSTRACT: Deficiencies in enzymes involved in proteoglycan (PG) turnover underlie a number of rare mucopolysaccharidoses (MPS), investigations of which can considerably aid understanding of the roles of PGs in corneal matrix biology. Here, the authors analyze novel pathologic changes in MPS VII (Sly syndrome) to determine the nature of PG-collagen associations in stromal ultrastructure. Transmission electron microscopy and electron tomography were used to investigate PG-collagen architectures and interactions in a cornea obtained at keratoplasty from a 22-year-old man with MPS VII, which was caused by a compound heterozygous mutation in the GUSB gene. Transmission electron microscopy showed atypical morphology of the epithelial basement membrane and Bowman's layer in MPS VII. Keratocytes were packed with cytoplasmic vacuoles containing abnormal glycosaminoglycan (GAG) material, and collagen fibrils were thinner than in normal cornea and varied considerably throughout anterior (14-32 nm), mid (13-42 nm), and posterior (17-39 nm) regions of the MPS VII stroma. PGs viewed in three dimensions were striking in appearance in that they were significantly larger than PGs in normal cornea and formed highly extended linkages with multiple collagen fibrils. Cellular changes in the MPS VII cornea resemble those in other MPS. However, the wide range of collagen fibril diameters throughout the stroma and the extensive matrix presence of supranormal-sized PG structures appear to be unique features of this disorder. The findings suggest that the accumulation of stromal chondroitin-, dermatan-, and heparan-sulfate glycosaminoglycans in the absence of β-glucuronidase-mediated degradation can modulate collagen fibrillogenesis.
    Investigative ophthalmology & visual science 07/2011; 52(9):6720-8. · 3.43 Impact Factor
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    ABSTRACT: In this protocol, we describe a 3D imaging technique known as 'volume electron microscopy' or 'focused ion beam scanning electron microscopy (FIB/SEM)' applied to biological tissues. A scanning electron microscope equipped with a focused gallium ion beam, used to sequentially mill away the sample surface, and a backscattered electron (BSE) detector, used to image the milled surfaces, generates a large series of images that can be combined into a 3D rendered image of stained and embedded biological tissue. Structural information over volumes of tens of thousands of cubic micrometers is possible, revealing complex microanatomy with subcellular resolution. Methods are presented for tissue processing, for the enhancement of contrast with osmium tetroxide/potassium ferricyanide, for BSE imaging, for the preparation and platinum deposition over a selected site in the embedded tissue block, and for sequential data collection with ion beam milling; all this takes approximately 90 h. The imaging conditions, procedures for alternate milling and data acquisition and techniques for processing and partitioning the 3D data set are also described; these processes take approxiamtely 30 h. The protocol is illustrated by application to developing chick cornea, in which cells organize collagen fibril bundles into complex, multilamellar structures essential for transparency in the mature connective tissue matrix. The techniques described could have wide application in a range of fields, including pathology, developmental biology, microstructural anatomy and regenerative medicine.
    Nature Protocol 05/2011; 6(6):845-58. · 8.36 Impact Factor
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    ABSTRACT: The spatial distribution of collagen fibrils in the corneal stroma is essential for corneal transparency and is primarily regulated by extrafibrillar proteoglycans, which are multi-functional polymers that interact with hybrid type I/V collagen fibrils. In order to understand more about proteoglycan organisation and collagen associations in the cornea, three-dimensional electron microscopy reconstructions of collagen-proteoglycan interactions in the anterior, mid and posterior stroma from a Chst5 knockout mouse, which lacks a keratan sulphate sulphotransferase, were obtained. Both longitudinal and transverse section show sinuous, oversized proteoglycans with near-periodic, orthogonal off-shoots. In many cases, these proteoglycans traverse over 400nm of interfibrillar space interconnecting over 10 collagen fibrils. The reconstructions suggest that multiple chondroitin sulphate/dermatan sulphate proteoglycans have aggregated laterally and, possibly, end-to-end, with orthogonal extensions protruding from the main electron-dense stained filament. We suggest possible mechanisms as to how sulphation differences may lead to this increase in aggregation of proteoglycans in the Chst5-null mouse corneal stroma and how this relates to proteoglycan packing in healthy corneas.
    Journal of Structural Biology 03/2011; 174(3):536-41. · 3.36 Impact Factor
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    ABSTRACT: The lamellar architecture found in many natural fibrous tissues has a significant bearing on their specific functions. However, current engineered tissues have simultaneously no realistic structures and no adequate functions. This study demonstrates a two-step process for obtaining structurally mimicking laminates in natural fibrous tissues with good optical and mechanical characters from purified-clinically- safe collagen molecules. Stacked lamella structures can be created by repeating flow casting, with the controlling parallel/orthogonal directionalities of each thin single-layer (2e5 mm in thickness). The transparency of laminates is successfully improved by a unique multi-cyclic vitrification with chemical cross-linking. The directionalities of optical and mechanical functions in laminates are strongly related with the preferential collagen alignments in the laminates. The tensile strength of laminates is extremely higher than any other engineered materials as well as native cornea, which exhibit an orthogonal laminated collagen structure and a good optical transmission.
    Biomaterials 02/2011; 32:3358-3366. · 8.31 Impact Factor
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    ABSTRACT: Engineering of well-organized tissue constructs is active in the field of material science for biomedical applications. Here, we propose a method for orienting collagen in transparent high-density collagen hydrogels using a simple rolling method. Structural organization and mechanical function were adjusted by regulating the thickness of the construct and the cross-linking reagent. Directionality of collagen alignment on the microscopic scale was achieved parallel to the extensional flow. The preferential alignment of collagen significantly affected the mechanical properties of the construct, with strong tensile strength in the direction parallel to the collagen, and high elastic strain in the perpendicular direction. The tensile strength in the parallel direction was effectively increased by 67% by increasing the cross-linking reagents by 33%, without affecting transparency which remained at 70-80% to visible light. The constructs exhibit good biocompatibility for as a substrate for the expansion of corneal epithelial cells isolated from human donor cornea, indicating the potential for tissue engineering and biomedical applications, particularly for ocular treatments.
    Journal of Biomaterials Science Polymer Edition 01/2011; 22(11):1427-42. · 1.70 Impact Factor

Publication Stats

673 Citations
233.71 Total Impact Points

Institutions

  • 2003–2014
    • Cardiff University
      • School of Optometry and Vision Sciences
      Cardiff, Wales, United Kingdom
  • 2013
    • European Synchrotron Radiation Facility
      Grenoble, Rhône-Alpes, France
  • 2011
    • Queen Mary, University of London
      • School of Engineering and Materials Science
      London, ENG, United Kingdom
  • 2010–2011
    • Tohoku University
      • Graduate School of Medicine
      Sendai-shi, Miyagi-ken, Japan
  • 1999–2008
    • Kyoto Prefectural University of Medicine
      • Department of Ophthalmology
      Kyoto, Kyoto-fu, Japan
  • 1994
    • Saint Louis University
      Saint Louis, Michigan, United States