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See relatedprovided article by Elsevier on - Publisher page 885 Connector Ehlers–Danlos Syndrome—Molecular Genetics Beyond the Collagens

Jouni Uitto and Franziska Ringpfeil Department of Dermatology and Cutaneous Biology, Jefferson Medical College, Philadelphia, Pennsylvania, USA

Ehlers–Danlos syndrome (EDS) is a prototypic connective they undergo proteolytic processing, fiber assembly, and tissue disorder with characteristic skin and joint involvement formation of stabilizing inter- and intra-molecular crosslinks. (Uitto et al, 2003). The cutaneous features include loose and In different forms of EDS, specific mutations have been fragile skin, poor wound healing, bruising tendency, and identified in type I, III, and V collagen polypeptides, as well the scars are characteristically atrophic, often resembling as in two enzymes that modify the collagen molecules (Uitto ‘‘cigarette paper’’. In addition, there are a number of extra- et al, 2003). These molecular defects explain the connective cutaneous manifestations, including hyperextensible joints tissue weakness and ultrastructural abnormalities in col- with propensity to dislocations, and in certain subtypes of lagen fibrils. Specifically, as demonstrated by transmission EDS, fragility of blood vessels, gastrointestinal tract and electronmicroscopy, the collagen fibrils show considerable uterus can result in catastrophic complications. The first variability in their diameter, and although individual fibrils clinical description detailing this disorder dates back to can be unusually large with irregular contour, the density of 1892 by Dr. Tschernogobow, a Russian dermatologist, and collagen fibrils is often reduced. Thus, EDS has been subsequently, Drs. Ehlers and Danlos, Danish and French considered as a disease of collagen. dermatologists, respectively, expanded on the systemic Evidence of molecular heterogeneity beyond the col- nature of this condition. The genetic nature of EDS was lagens was initially presented by observations on a patient recognized in 1949, and its clinical manifestations were with adrenal hyperplasia and a classical form of EDS, who subsequently suggested to result from defects in the was found to harbor a 30-kb contiguous gene deletion on collagen ‘‘wicker work’’. The genetic heterogeneity of EDS the short arm of chromosome 6 (Burch et al, 1997). The was established in the 1960s, and the first molecular deleted interval contained both the CYP21 gene that defects in collagen biosynthetic pathways were delineated encodes steroid 21-hydroxylase and the TNX gene, which in patients with EDS in 1972 (McKusick, 1972). encodes tenascin-X, a connective tissue protein develop- Traditionally, EDS has been subdivided into 11 distinct mentally associated with collagen fibrils. It was subse- variants (types I–XI), based on clinical observations, mode quently demonstrated by immunofluorescence of the skin of of inheritance, and/or molecular characterization. A con- this and similar patients that they were lacking tenascin-X, sensus conference held in 1997 (Villefranche), however, and their serum contained essentially undetectable levels of proposed a revised nosology, which recognizes six distinct this protein. The suggestion that tenascin-X is directly subtypes (Beighton et al, 1998). In fact, several previously involved in the pathogenesis of EDS was subsequently described variants, some of them exceedingly rare and not confirmed by demonstration of null mutations in both alleles well-defined, were excluded from the EDS category. of TNX, and the mutations in these patients were clearly The molecular basis of the major forms of EDS is now autosomal recessive (Schalkwijk et al, 2001). The clinical well-established, and the clinical manifestations are based features in these patients were similar to those of the primarily on mutations in the genes encoding collagen classical autosomal dominant type of EDS, except that the polypeptide subunits or enzymes that modify the primary tenascin-X deficient patients lack atrophic scars and wound collagen translation products. Collagen consists of a family healing is not consistently delayed, findings characteristic of of proteins, and there are as many as 27 distinct vertebrate classic EDS. The pathoetiologic role of tenascin-X has been collagens (types I–XXVII) (Myllyharju and Kivirikko, 2004). further confirmed by the development of TNX null mice, Each collagen molecule is composed of three a-chain which recapitulate many of the features of patients with subunits, which can be identical in homotrimers, or consist tenascin-X deficiency (Mao et al, 2002). of two or three different kinds of polypeptides in hetero- In this issue, Zweers et al (2004) have examined five trimers. Thus, there are over 40 different genes encoding TNX-deficient individuals for changes in dermal connective the distinct a-chains that are synthesized as precursor tissue, which consists of four major groups of extracellular polypeptides, pro-a-chains. These polypeptides are hy- matrix components, viz., collagen, the elastic fibers, non- droxylated and glycosylated in reactions catalyzed by specific collagenous glycoproteins, and glycosaminoglycan/proteo- enzymes, three of the pro-a-chains then fold into the glycan macromolecules. The principal fiber networks characteristic triple-helical conformation, and the collagen consist of collagen and elastin, which interact with non- molecules are secreted into the extracellular milieu where collagenous glycoproteins, such as elastin-associated

Copyright r 2004 by The Society for Investigative Dermatology, Inc. xii 122 : 4 APRIL 2004 EHLERS–DANLOS SYNDROME xiii microfibrillar proteins (MFAPs) and glycosaminoglycan/ the distribution of decorin could be detected between proteoglycans. The supramolecular organization of these tenascin-X deficient and normal skins. The authors suggest extracellular matrix components, as well as their interac- that this discrepancy between reduced collagen content tions with cellular components of the connective tissue, and normal skin thickness could possibly be explained by have previously allowed identification of critical metabolic edema, a suggestion that clearly does not conform with and structural features that are prerequisites for normal the clinical observation of soft and velvety skin in these physiological function of the skin. Zweers et al (2004) now patients. report that, in addition to previously reported reduction in Collectively, the data reported by Zweers et al (2004) dermal collagen, there are major abnormalities in elastic confirm that tenascin-X deficiency can result in connective fibers and associated microfibrils in the dermis of these tissue abnormalities, which manifest as classical EDS with patients. The elastic fibers are fragmented, clumped, and autosomal recessive inheritance. These findings add to the devoid of microfibrils. Based on these observations, the molecular complexity of EDS beyond the collagens, but authors suggest that abnormality in elastic fibers contri- they also provide a diagnostic test through serum assay of butes to the clinical phenotype, and they put forward a tenascin-X, helpful in subclassification of EDS as well as proposal that tenascin-X may serve as an ‘‘organizer of providing information on the mode of inheritance with connective tissue assembly’’. implications for genetic counseling. The authors’ interpretation raises a couple of questions. First, all skin biopsies were obtained from TNX-deficient patients with the age range of 32–56 years. Since these DOI: 10.1111/j.0022-202X.2004.22435.x patients are reported to have clinical findings since early childhood, could the changes in elastic fibers just be secondary to overall instability of the connective tissue due References to collagen abnormalities? In this context, it should be noted that elastin abnormalities have been demonstrated in Beighton P, De Paepe A, Steinmann B, Tsipouras P, Wenstrup RJ: Ehlers–Danlos syndromes: Revised nosology, Villefranche, 1997. Ehlers–Danlos National the skin of EDS patients with clear cut collagen defects Foundation (USA) and Ehlers–Danlos Support Group (UK). Am J Med and presumably no tenascin-X deficiency. It should also Genet 77:31–37, 1998 be noted that although fibrillin-1, a major non-collagenous Burch GH, Gong Y, Liu W, et al: Tenascin-X deficiency is associated with Ehlers– glycoprotein associated with the elastic fiber network, was Danlos syndrome. Nature Genet 17:104–108, 1997 Mao JR, Taylor G, Dean WB, et al: Tenascin-X deficiency mimics Ehlers–Danlos deficient in the skin of the patients studied by Zweers et al syndrome in mice through alteration of collagen deposition. Nature Genet (2004), the primary synthesis of fibrillin-1 by TNX-deficient 30:421–425, 2002 fibroblasts was not altered. McKusick VA: Heritable Disorders of Connective Tissue. 4th edn., St. Louis: C.V. Another puzzling question relates to the fact that the Mosby, 1972 Myllyharju J, Kivirikko KI: Collagens, modifying enzymes and their mutations in collagen density was reported to have been reduced to humans, flies and worms. Trends Genet 20:33–43, 2004 about 40% of the control, yet the thickness of the dermis Schalkwijk J, Zweers MC, Steijlen PM, et al: A recessive form of the Ehlers– was unaltered. Considering the fact that collagen comprises Danlos syndrome caused by tenascin-X deficiency. N Engl J Med 345:1167–1175, 2001  80% of the extracellular matrix of the dermis, one would Uitto J, Ringpfeil F, Pulkkinen L: Heritable Disorders of Connective Tissue — expect significant atrophy, unless there is substitution of Ehlers–Danlos Syndrome, Pseudoxanthoma Elasticum and Cutis Laxa. collagen by some other material. The authors studied the In: Bolognia JL, Jorizzo JL, Rapini RP (eds). Dermatology. Harcourt presence of glycosaminoglycans/proteoglycans with spe- Publishers, London; p 1519–1530, 2003 Zweers MC, van Vlijmen-Willems IM, van Kuppevelt TH, et al: Deficiency of cific histopathologic and immunohistochemical stains. No tenascin-X causes abnormalities in dermal elastic fiber morphology. difference in the negatively charged polysaccharides or in J Invest Dermatol 122:885–891, 2004