X-Linked Spondyloepiphyseal Dysplasia Tarda: Molecular Cause of a Heritable Platyspondyly

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1 X-Linked Spondyloepiphyseal Dysplasia Tarda: Molecular Cause of a Heritable Platyspondyly SPINE Volume 28, Number 22, pp E478 E , Lippincott Williams & Wilkins, Inc. Jörg Fiedler, PhD,* Anne-Marie Frances, MD, Martine Le Merrer, MD, Markus Richter, MD,* and Rolf E. Brenner, MD* Study Design. Report of a family affected with X-linked spondyloepiphyseal dysplasia tarda with special respect to radiologic alterations of the spine from puberty to the forth decade and to molecular analysis of the underlying genetic defect. Objectives. To report the typical radiologic presentation of patients with X-linked spondyloepiphyseal dysplasia tarda and the diagnostic tool of mutation screening for that disease in order to avoid confusion with similar occurrences. Summary of Background Data. Spondyloepiphyseal dysplasia tarda is a genetically heterogeneous disorder that frequently manifests itself with back pain starting around puberty. The X-linked recessive form (X-linked spondyloepiphyseal dysplasia tarda) affects males and is clinically characterized by an arm span markedly exceeding total height, a barrel chest deformity, and early development of degenerative joint disease. The disorder is caused by mutations in the SEDL gene located on Xp22.12-p Methods. Radiologic alterations of the cervical, thoracal, and lumbar spine were assessed in the affected family members and one suspected female carrier in correlation to age. All 6 exon codings for the SEDL gene were analyzed by primer cycle sequencing. Results. In 3 male patients from a French family, we identified a 5 base pair deletion in SEDL, exon 5 at position (delaagac). Carrier status for the mutation could be confirmed in one female member of the family, which is inconspicuous in terms of spine and joint diseases. Radiologic abnormalities of the patients comprised generalized platyspondyly, a hump-shaped deformity of cervical, thoracal, and lumbar vertebral bodies as well as signs of retrospondylophytes, osteochondrosis, and spondylarthrosis. Conclusions. X-linked spondyloepiphyseal dysplasia tarda should be kept in mind as a differential diagnosis in men with early onset of back pain and radiologic abnormalities of the vertebral bodies comprising platyspondyly and a central hump. [Key words: spondyloepiphyseal dysplasia, SEDL, sedlin, platyspondyly] Spine 2003;28:E478 E482 From the *University of Ulm, Department of Orthopedics, Ulm, Division for Biochemistry of Joint and Connective Tissue Diseases, Hôpital Font-Pré CHIC Toulon Service de Génétique et Cytogénétique, Assistance Hôpitaux Publique de Paris, Département de Génétique, Unité de Génétique Clinique. Acknowledgment date: April 9, Acceptance date: June 6, The manuscript submitted does not contain information about medical device(s)/drug(s). No funds were received to support this work. No benefits in any form have been or will be received from a commercial party related directly or indirectly to the subject of this manuscript. Address correspondence to Rolf E. Brenner, MD, University of Ulm, Department of Orthopedics, Division for Biochemistry of Joint and Connective Tissue Diseases, RKU, Oberer Eselsberg 45, Ulm 89081, Germany; rolf.brenner@medizin.uni-ulm.de Back pain starting in the second to third decade is widespread, and its differential diagnosis may be difficult, especially when early degenerative alterations are associated. Besides rather common diseases like Morbus Scheuermann, several skeletal dysplasias, some of which include marked platyspondyly, may be underlying such complaints. One example is the X-linked spondyloepiphyseal dysplasia tarda (SEDL), which is a mild variant with usually low normal stature, barrel chest deformity, and minor epiphyseal but characteristic vertebral body dysplasia comprising platyspondyly and a central hump. 1,2 The first signs of SEDL occur around puberty with back pain, and later degenerative joint disease is common among the adult patients and often makes hip joint replacement necessary in the fourth or fifth decade. In 1988, the locus responsible for SEDL was mapped to Xp22. 3 Recently, several mutations in a gene located within that region have been described in patients with SEDL. 2,4 9 This gene, termed SEDL according to the disease, is expressed in cartilage and various other tissues. 10 We present a family in which we identified a mutation in the SEDL gene that causes typical deformities of the vertebral bodies in the affected members. Retrospective evaluation of radiographs allowed us to follow the spinal manifestations over more than a decade. The long time span until the correct diagnosis was set reflects the overlap of spinal alterations in SEDL with other diseases like Scheuermann disease, Stickler Syndrome, Morquio disease, or brachyolmia. According to the degenerative process in older age, diagnosis remains difficult. In this situation, mutation screening for SEDL proved to be an important new diagnostic tool. Patients and Methods Family Report. The family was presented for genetic counseling in view of a planned pregnancy. Several family members had a history of early back pain and radiologic signs of dysplastic vertebral bodies that had lead to the diagnosis of Morquio disease, dating from Additional enzymatic studies to confirm the diagnosis failed. Therefore, clinical data were reconsidered, and another radiologic examination of an affected male was done. The family s pedigree indicated an X-linked recessive trait of inheritance (Figure 1) with two women (I-2 and II-6) suspected to be carriers of the mutant allele. Clinical and anthropometric data as well as blood samples were obtained from all patients and the presumptive two female carriers with informed consent. Mutation Analysis. For mutational screening, all 6 exon of the SEDL gene were analyzed by primer cycle sequencing. E478

2 X-Linked Spondyloepiphyseal Dysplasia Tarda Fiedler et al E479 Results Figure 1. Pedigree of the described family with X-linked spondyloepiphyseal dysplasia tarda. Genomic DNA was isolated from blood samples with QIAmp DNA Blood Mini Kit (QIAGEN, Hilden, Germany) following the manufacturer s instructions. Amplification, primers, and polymerase chain reaction (PCR) conditions are described elsewhere. 11 Sequencing was performed by using the Thermo Sequenase Primer Cycle Sequencing Kit. Samples were run on a fluorescent automated DNA sequencer, ALFexpress, and the ALFwin Sequence Analyzer Software V2.1 was used for data processing (all Amersham Biosciences Europe GmbH, Freiburg, Germany). Clinical Phenotype The pedigree of the family and the X-linked inheritance is presented in Figure 1. The affected male patients had barrel chest deformity and a height at the lower extreme of normal range. Patient II-1 had an arm span of 1.78 m, markedly exceeding his height of 1.58 m. The female carrier (I-2) showed no such disproportion, with a nearly equal height of 1.60 m and arm span of 1.63 m. From the end of the first decade, the complaints of the male patients led to repeated radiologic evaluation of the spine and other joints. They developed early onset degenerative joint disease, and Patient II-1 underwent replacements of both hips and the left shoulder until the age of 39 years. As shown in Figure 2, radiographs of the lumbar spine revealed platyspondyly and dysplasia of the vertebral bodies with a more or less pronounced hump in the central or dorsal region. Similar alterations could be observed in the thoracic (Figure 3) and cervical spine (Figure 4). The female II-2 did not show these abnormalities. Interestingly, in the patients, the degree of kyphosis was less severe than expected from the severity of vertebral body deformation. From Patient II-3, a radiograph of the cervical spine was already performed at the age of 11, 12, and 33 years (Figure 4) showing the typical hump-shaped deformities Figure 2. Lateral radiographs of the lumbar spine of the affected male family members (B, II-3, age of 8 years; C, II-1, age of 14 years; D, II-4, age of 26 years) at different ages and the nonaffected female (A, II-2, age of 18 years).

3 E480 Spine Volume 28 Number Figure 3. Lateral radiographs of the thoracic spine. A, II-2, age of 18 years; B, Patient II-3, age of 7 years; C, II-1, age of 14 years. of vertebral bodies with endplate deformities similar to the lumbar region. In addition, clear signs of retrospondylophytes, osteochondrosis, and spondyloarthrosis were detectable in the radiographs. Overall, developments of spinal alterations included early onset and rapid progression of degenerative spine disease. Mutation Analysis Direct sequencing of PCR-amplified SEDL exon 1 to 6 with their flanking regions revealed hemizygosity for a 5 bp deletion in the index Patient II-4. This mutation affects the nucleotides at position 267 to 271 of exon 5 and causes a frameshift after amino acid 90, followed by 8 missense amino acids and a termination codon. Thus, 50 amino acids from the C-terminus were deleted. Segregation of the mutant allele could be confirmed by sequence analysis in all affected family members and the female carrier I-2 (Figure 5). No changes in other regions of the SEDL gene could be detected. Discussion The French family presented confirms previous reports that the phenotype of SEDL is clinically not very impressive at an early age and that there is an overlap of spinal alterations with other diseases like Morquio Syndrome, brachyolmia, Stickler Syndrome, Morbus Scheuermann, or early degenerative disease of weight bearing joints and the spine in general. 2,12 Moreover, the X-linked recessive mode of inheritance may prevent identification of its heritable basis. Therefore, some cases might not be recognized during routine orthopedic care, and the incidence of patients with SEDL may be higher than expected so far. In males with early chronic back pain, the presence of a disproportion between rump and limb length and family history should be carefully assessed. If this additional information is compatible with the diagnosis of SEDL, the lumbar spine should be radiographically evaluated. The presence of platyspondyly associated with a humpshaped deformity of vertebral bodies should lead to further evaluation of epiphyseal regions of the hip joint, for example. Interestingly, the degree of kyphosis was less than could be expected from the degree of vertebral body deformation, indicating that intervertebral disc tissue could compensate partially for the dysplastic shape of the vertebrae. The radiologic follow-up over more than a decade also revealed that spinal abnormalities were not as progressive than those of weight bearing joints indicated by early joint replacement in one of the affected family members. Some skeletal dysplasias that have similar radiologic manifestations of the spine can be separated quite easily from SEDL by their involvement of marked short stature. Others like Morquio Syndrome or Stickler Syndrome can be excluded by their involvement of specific nonskeletal tissues and metabolic analysis of mucopolysaccharide excretion (Morquio Syndrome) or mutation screening (collagen types II and XI in Stickler Syndrome). Stickler syndrome (hereditary arthroophthalmopathy) is an autosomal dominant connective tissue disorder linked to mutations in Type II and Type XI collagen. 13 It is often associated with vitreoretinal degeneration, severe myopia, cleft palate, high frequency hearing loss, hip anomalies, and early degenerative joint disease. Spinal abnormalities in Stickler Syndrome include spondylolisthesis, scoliosis, Scheuermann-like kyphosis, and thoracolumbar abnormalities, including Schmorl nodes and platyspondyly. Endplate abnormalities could be detected in radiographs as irregular vertebral borders, sclerosis, and disc space variations. 14,15 Another skeletal dysplasia recognized during infancy and adolescence is brachyolmia. Three types of brachyolmia have been identified so far, either autosomal recessive or dominant forms. Besides distinctive characteristics concerning metaphyseal changes in the long bones and variations in the femoral neck, all patients present a shortening of the trunk, generalized platyspondyly, and kyphoscoliosis

4 X-Linked Spondyloepiphyseal Dysplasia Tarda Fiedler et al E481 Figure 4. Radiographs of patient II-3 at the age of 11 years (D), 12 years (B) and 33 years (A, C, E), showing the deformities of the cervical spine and the not age correlated retrospondylophytes, signs of spondyloarthrosis (C, E) and osteochondrosis (A, C, E). The incidence of Scheuermann disease that comprises kyphotic abnormalities, Schmorl nodes, or irregularities of the vertebral endplates has been estimated at 1% to 8% of the general population. 20 Overall, Morbus Scheuermann is a quite heterogeneous disease with still unknown etiology. Therefore, up to now, the diagnosis is based on clinical and radiologic features and has to be separated from other rather rare diseases that comprise similar spinal alterations. The shape of the vertebrae with irregularities of the vertebral plate is not typically characterized by the central hump, and there is no epiphyseal dysplasia of other joints predisposing to early development of osteoarthritis. The overlapping phenotypes indicate that SEDL mutation screening is an important new diagnostic tool to prove the diagnosis in patients or families suspected to carry that disease. Today, over 20 different mutations have been identified in patients with SEDL. 2,4 6,9,12 These mutations are spread over the gene s entire coding region comprising exon 3 6. Mumm et al first described the mutation identified in the present study. 7 It leads to a predicted loss of 36% of SEDLIN, the corresponding protein for which the precise function is not known. It has been suggested that the protein is involved in intracellular transport processes of molecules, which may be of special importance for cartilage because chondrocytes need a high synthetic capacity for extracellular matrix molecules. Furthermore, the association with early development of osteoarthrosis might depend in part on a quantitatively reduced biosynthesis of extracellular matrix molecules. A more detailed investigation of SEDL may lead to a novel pathogenetic model for vertebral body dysplasia and early development of osteoarthrosis. Conclusions Patients with SEDL show the typical morphologic deformations of the vertebral bodies comprising platyspondyly and a central hump. Because of the rapid progressive multiple degenerations of the spine, it is necessary to ensure close medical attendance and offer genetic counseling and lifestyle advisory. Heterozygote female carriers obviously show no signs of spine degeneration due to mutations in SEDL, but it is advisable to implicate all female family members in the examination for genetic counseling in order the prove the carrier status. Mutation screening is an important diagnostic tool to differ- Figure 5. Sequence analysis in one male patient (II-4) and two female family members (I-2, II-2) revealing hemizygosity (II-4) and heterozygosity (I-2) for the mutation (delaagac) at nucleotide position 267 to 271 of exon 5 of the SEDL. II-2 is obviously not affected with this mutation and therefore she has no carrier status. EXON 5DNA Genebank SEDL sequence for exon 5; EXON 5 cds SEDLIN coding sequence for exon 5.

5 E482 Spine Volume 28 Number entiate SEDL from other diseases with overlapping radiologic findings. Key Points Back pain starting around puberty is often the first sign in male SEDL patients. A hump-shaped deformity of vertebral bodies is a typical radiologic sign of SEDL. The diagnosis can be confirmed by mutation analysis of the SEDL-gene. Acknowledgments The authors thank the members of the family described and R. Schubert-Schmitt for expert technical assistance. References 1. Iceton JA, Horne G. Spondylo-epiphyseal dysplasia tarda. The X-linked variety in three brothers. J Bone Joint Surg Br 1986;68: Whyte MP, Gottesman GS, Eddy MC, et al. X-linked recessive spondyloepiphyseal dysplasia tarda. Clinical and radiographic evolution in a 6-generation kindred and review of the literature. Medicine (Balt) 1999;78: Szpiro-Tapia S, Sefiani A, Guilloud-Bataille M, et al. Spondyloepiphyseal dysplasia tarda: linkage with genetic markers from the distal short arm of the X chromosome. Hum Genet 1988;81: Gedeon AK, Colley A, Jamieson R, et al. Identification of the gene (SEDL) causing X-linked spondyloepiphyseal dysplasia tarda. Nat Genet 1999;22: Gedeon AK, Tiller GE, Le Merrer M, et al. The molecular basis of X-linked spondyloepiphyseal dysplasia tarda. Am J Hum Genet 2001;68: Grunebaum E, Arpaia E, MacKenzie JJ, et al. A missense mutation in the SEDL gene results in delayed onset of X linked spondyloepiphyseal dysplasia in a large pedigree. J Med Genet 2001;38: Mumm S, Christie PT, Finnegan P, et al. A five-base pair deletion in the sedlin gene causes spondyloepiphyseal dysplasia tarda in a six-generation Arkansas kindred. J Clin Endocrinol Metab 2000;85: Mumm S, Zhang X, Vacca M, et al. The sedlin gene for spondyloepiphyseal dysplasia tarda escapes X-inactivation and contains a non-canonical splice site. Gene 2001;273: Tiller GE, Hannig VL, Dozier D, et al. A recurrent RNA-splicing mutation in the SEDL gene causes X-linked spondyloepiphyseal dysplasia tarda. Am J Hum Genet 2001;68: Gecz J, Hillman MA, Gedeon AK, et al. Gene structure and expression study of the SEDL gene for spondyloepiphyseal dysplasia tarda. Genomics 2000; 69: Fiedler J, Bittner M, Puhl W, et al. Mutations in the X-linked spondyloepiphyseal dysplasia tarda (SEDL) coding sequence are not a common cause of early primary osteoarthritis in men. Clin Genet 2002;62: MacKenzie JJ, Fitzpatrick J, Babyn P, et al. X linked spondyloepiphyseal dysplasia: a clinical, radiological, and molecular study of a large kindred. J Med Genet 1996;33: Rose PS, Ahn NU, Levy HP, et al. Thoracolumbar spinal abnormalities in Stickler syndrome. Spine 2001;26: Letts M, Kabir A, Davidson D. The spinal manifestations of Stickler s syndrome. Spine 1999;24: Stickler GB, Hughes W, Houchin P. Clinical features of hereditary progressive arthro-ophthalmopathy (Stickler syndrome): a survey. Genet Med 2001; 3: Darcan S, Yalman O, Coker M, et al. Familial brachyolmia. J Pediatr Endocrinol Metab 2000;13: Hoo JJ, Oliphant M. Two sibs with brachyolmia type Hobaek: five-year follow-up through puberty. Am J Med Genet 2003;116: Ikegawa S, Nakamura K, Nakamura S, et al. Brachyolmia: a report of two cases. J Pediatr Orthop 1995;15: Karabiyik N, Oguz F, Sidal M, et al. A case of brachyolmia. Turk J Pediatr 1997;39: Wenger DR, Frick SL. Scheuermann kyphosis. Spine 1999;24: