Cornea Large Proteoglycan Complexes and Disturbed Collagen Architecture in the Corneal Extracellular Matrix of Mucopolysaccharidosis Type VII (Sly Syndrome) Robert D. Young,1 Petra Liskova,2 Christian Pinali,1 Barbara P. Palka,1 Michalis Palos,3 Katerina Jirsova,2 Enkela Hrdlickova,3 Marketa Tesarova,4 Milan Elleder,5 Jiri Zeman,4 Keith M. Meek,1 Carlo Knupp,1 and Andrew J. Quantock1 PURPOSE. Deficiencies in enzymes involved in proteoglycan CONCLUSIONS. Cellular changes in the MPS VII cornea resemble (PG) turnover underlie a number of rare mucopolysacchari- those in other MPS. However, the wide range of collagen fibril doses (MPS), investigations of which can considerably aid un- diameters throughout the stroma and the extensive matrix derstanding of the roles of PGs in corneal matrix biology. Here, presence of supranormal-sized PG structures appear to be the authors analyze novel pathologic changes in MPS VII (Sly unique features of this disorder. The findings suggest that the syndrome) to determine the nature of PG-collagen associations accumulation of stromal chondroitin-, dermatan-, and heparan- ␤ in stromal ultrastructure. sulfate glycosaminoglycans in the absence of -glucuronidase- mediated degradation can modulate collagen fibrillogenesis. (In- METHODS. Transmission electron microscopy and electron to- vest Ophthalmol Vis Sci. 2011;52:6720–6728) DOI:10.1167/ mography were used to investigate PG-collagen architectures iovs.11-7377 and interactions in a cornea obtained at keratoplasty from a 22-year-old man with MPS VII, which was caused by a com- he mucopolysaccharidoses (MPS) are a group of lysosomal pound heterozygous mutation in the GUSB gene. Tstorage diseases caused by inherited defects in specific ESULTS enzymes involved in the degradation of glycosaminoglycans R . Transmission electron microscopy showed atypical 1–3 morphology of the epithelial basement membrane and Bow- (GAGs). The result is a widespread accumulation of derma- man’s layer in MPS VII. Keratocytes were packed with cyto- tan sulfate (DS), chondroitin sulfate (CS), heparan sulfate (HS), or keratan sulfate (KS) in the majority of organs and tissues and plasmic vacuoles containing abnormal glycosaminoglycan a wide range of clinical manifestations. Ocular involvement (GAG) material, and collagen fibrils were thinner than in nor- attributed to GAG deposition in MPS includes progressive cor- mal cornea and varied considerably throughout anterior neal clouding, glaucoma, retinal degeneration, and optic disc (14–32 nm), mid (13–42 nm), and posterior (17–39 nm) re- swelling followed by its atrophy.4 In a mouse model of MPS gions of the MPS VII stroma. PGs viewed in three dimensions VII, progressive photoreceptor death has also been observed.5 were striking in appearance in that they were significantly The cornea is affected in all the MPS subtypes classified to date, larger than PGs in normal cornea and formed highly extended with a significant overlap of corneal phenotype between, and linkages with multiple collagen fibrils. also within, the MPS types and a variety of treatment op- tions.6,7 MPS VII (Sly syndrome; Mendelian Inheritance in Man (MIM) #253220) is a rare, autosomal recessive, inherited con- From the 1Structural Biophysics Group, School of Optometry and Vision Sciences, Cardiff University, Cardiff, Wales, United Kingdom; dition with an estimated incidence of 1 in 250,000 births, the 2Ocular Tissue Bank and Laboratory of the Biology and Pathology which is one of the lowest of all MPS. Accumulation of unde- of the Eye, Institute of Inherited Metabolic Disorders, the Departments graded GAGs typically leads to short stature, bone dysplasia, of 3Ophthalmology and 4Pediatrics and Adolescent Medicine, and the hepatosplenomegaly, hydrops fetalis, and mental retarda- 5Institute of Inherited Metabolic Disorders, First Faculty of Medicine, tion.8,9 The course of the disease is considered clinically atten- Charles University in Prague and General University Hospital in Prague, uated if there is no hydrops fetalis or severe mental retardation Prague, Czech Republic. and if the condition does not lead to death within a year.8 Most Supported by Biotechnology and Biological Sciences Research children with MPS VII live into teenage or young adult years. Council UK Project Grant BB/F022077/1 (CK, AJQ), Engineering and Various approaches toward therapy have been investigated Physical Sciences Research Council UK Research Grant EP/F034970/1 in mutant MPS VII mice, including bone marrow transplanta- (AJQ, KMM, RDY), and Medical Research Council UK Programme 10 11 Grant G0600755 (KMM, AJQ), the Research Project of the Ministry of tion, enzyme replacement therapy, transplacental delivery 12 Education, Youth and Sports of the Czech Republic Grant of the deficient enzyme tagged to Fc into the fetal circulation, MSM0021620806 (PL, KJ, ME), and the Research Project of the Ministry gene therapy,13,14 encapsulation,15 and stem cell therapy.16 of Health of the Czech Republic Grant MZOVFN2005 (JZ, MT). However, no effective treatment is yet available for affected Submitted for publication February 11, 2011; revised June 9, 2011; human patients. MPS VII is caused by deficiencies of ␤-glucu- accepted June 28, 2011. ronidase enzyme (GUSB), leading to an accumulation of DS, Disclosure: R.D. Young, None; P. Liskova, None; C. Pinali, None; HS, and CS GAGs in the cells and tissues.17 Forty-nine unique B.P. Palka, None; M. Palos, None; K. Jirsova, None; E. Hrdlickova, disease-causing mutations in the ␤-glucuronidase gene (GUSB None; M. Tesarova, None; M. Elleder, None; J. Zeman, None; K.M. Meek, None; C. Knupp, None; A.J. Quantock, None MIM #611499) have been identified in persons with MPS VII Corresponding author: Andrew J. Quantock, Structural Biophysics from various populations. Heterogeneity of pathogenic Group, School of Optometry and Vision Sciences, Cardiff University, changes, with different effects on the function of the mutant Maindy Road, Cardiff CF24 4LU, Wales, United Kingdom; proteins, manifests as extensive clinical variability among pa- [email protected]. tients with MPS VII.18 Investigative Ophthalmology & Visual Science, August 2011, Vol. 52, No. 9 6720 Copyright 2011 The Association for Research in Vision and Ophthalmology, Inc. Downloaded from iovs.arvojournals.org on 09/26/2021 IOVS, August 2011, Vol. 52, No. 9 Proteoglycans in Mucopolysaccharidosis Type VII 6721 Proteoglycans (PGs) with their GAG side chains, in partic- suggesting MPS diagnosis were present at birth. The ␤-glucuronidase ular those of the small leucine-rich PG family of molecules, are activity in isolated leukocytes was Ͻ2% of controls. Molecular genetic implicated in the regulation of fibrillar order in cornea. Recent analysis of the GUSB gene revealed compound heterozygosity for two studies on corneal three-dimensional structure using electron missense mutations, c.[155CϾT] ϩ [1120CϾT], leading to amino acid tomography have provided new insights into how PGs may changes p.Ser52Phe and p.Arg374Cys.22 Medical examination at the help to maintain the ordered architecture of the stroma essen- age of 24 years revealed short stature (155 cm), a body weight of 65 kg, tial for tissue transparency.19–21 Here, we describe a case of and head circumference 61 cm, plus retroflexion of the head with a MPS VII in which grafting was carried out to treat a reduction short neck, sternal prominence, lumbar hyperlordosis, and hepato- of corneal transparency. Clinical, biochemical, molecular ge- splenomegaly. An x-ray survey showed typical signs of dysostosis netic, histologic, and ultrastructural examinations of explanted multiplex. There were thickened aortic and mitral valves on echocar- MPS VII corneal tissue were performed, with comparisons with diography and an obstructive type of ventilation with a forced vital data obtained from a healthy human donor cornea acquired capacity of 74%. The urinary GAG excretion, measured on several from an eye bank. Data indicate that disturbance in PG turn- occasions, was increased between 8 and 25 g/mol creatinine compared over resulting from a GUSB deficiency leads to alterations in with a normal value in adults of 1.8 Ϯ 0.6 g/mol,23 with higher both fibrillar and interfibrillar components of the corneal fractions of CS and DS. The patient was not mentally retarded and was stroma. able to read, and several IQ tests performed since childhood revealed scores of 81 to 85. MATERIALS AND METHODS At 5 years of age, the patient’s best-corrected visual acuity (BCVA) was 0.4 in both eyes and mild corneal clouding was noticed. Refraction The study conformed to the tenets of the Helsinki Declaration and local was ϩ4.0/Ϫ5.0 D ϫ 7° in the right eye and ϩ3.0/Ϫ4.5 D ϫ 174° in the ethics codes. left. At 14 years of age, the patient was referred to the Department of Ophthalmology in Prague with BCVAs of 0.33 (right eye) and 0.4 (left Medical and Ocular History eye). Because of progressive corneal clouding, a penetrating kerato- The patient reported here was a male, an only child, born to healthy, plasty (8-mm diameter button) was performed in the right eye at the unrelated parents after an unremarkable pregnancy. Clinical features age of 20 years and in the left eye at the age of 22 years. Before surgery, FIGURE 1. Ocular findings in a pa- tient with mucopolysaccharidosis VII. (A, B) Diffuse fine cloudiness in corneal stroma of the right eye be- fore surgery. (C) Videotopography reveals regular astigmatism in the left eye before grafting. (D) Six months after keratoplasty in the right eye. (E) Normal postoperative appearance of fundus in the right eye. (F) Normal appearance of the macular region on OCT in the right eye. Downloaded from iovs.arvojournals.org on 09/26/2021 6722 Young et al. IOVS, August 2011, Vol. 52, No. 9 diffuse opacification involving all corneal layers was observed bilater- were fixed for 20 hours at room temperature with 2.5% glutaraldehyde ally (Fig. 1), with a BCVA of 0.33 in the right eye and 0.11 in the left.