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Abstract

Tenascin-X (TNX) is a large extracellular matrix protein that is involved in cell adhesion, cell motility, and fibril deposition. Over the last years, interest in TNX and its relationship to EDS grew.

Ehlers-Danlos syndrome (EDS) is a group of hereditary multisystem connective tissue disorders, and its severity can vary from mild to life-threatening forms. The Villefranche Nosology classifies six main EDS types which do not include all of the TNX related EDS subtypes I want to review in this article.

Tenascin-X and especially its relationship to EDS is largely unknown which is why I try to give a brief overview of the so far published types and their overlap to other diseases.

Introduction

Ehlers-Danlos syndrome describes a group of hereditary and multisystem connective tissue disorders with a variety of different phenotypes that can vary from very mild forms to life-threatening (1). There is little known about why even people with the same mutation have completely different symptoms and progression. Research is moving forward, but still many questions remain unanswered.

By now all of the main types of the Villefranche Nosology except one have a clear genetic cause – mutations that are described in the literature and that can be measured by genetic blood testing. The only type with a mostly unclear genetic basis is the hypermobile type. Over the last decade in very few EDS patients (less than 10 percent) one gene could be identified as the possible cause of their EDS phenotype – Tenascin-XB (TNXB).

What is Tenascin XB (TNXB) and Tenascin (TNX)?

TNXB (Tenascin-XB) is a gene on the short arm of chromosome 6 which encodes for an extracellular matrix protein called Tenascin-X (2). Tenascin-X belongs to a huge family of proteins (3, 4) including Tenascin-C, which is expressed in growing tumors, as well as Tenascin-W and Tenascin-R that is primarily expressed in the nervous tissue.

All Tenascins have some similarities concerning their structures (5):

N-terminal domain
Epidermal growth factor-like domains (EGF-like domains)
Fibronectin type III- like domains (FN-like domains)
C-terminal fibrinogen-like domain

figure-1-001Figure 1

Tenascins interact with cells and change their adhesion which then leads to increased cell motility (3).

Of all Tenascins, Tenascin-X is the one we know only very little about (4).

The function of Tenascin-X in the human organism is mainly unclear but studies showed that it is expressed in muscles and loose connective tissue and probably important to preserve the structure of the dermis and other connective tissue throughout the body by regulating the space between fibrils through binding to collagen, fibril-associated collagens, decorin and other matrix proteins (3, 4, 6).

Beyond its architectural function, Tenascin-X controls cell to cell adhesion (7) and was found to activate transforming growth factor beta (TGFb) (8) which plays a major role in some other connective tissue disorders such as Loeys-Dietz Syndrome.

The underlying mechanism of TNX related EDS subtypes might not be the interference with collagen processing, as recently thought, but more likely due to dermal fibroblast fibril deposition regulation into the matrix (4).

Which Tenascin X related EDS subtypes are there?

There is a lot of confusion about the TNX related EDS subtypes, but I tried to get some recent information classified according to four TNX related subtypes.

The types of TNX related EDS at the moment are:

TNX-deficient EDS

EDS patients with mutations on TNXB that lead to the complete deficiency of Tenascin-X do mostly look like the classical EDS types with features of hypermobility, skin hyperextensibility, bruising but without the atrophic scarring (1). This suggests that even if the TNX-deficient type of EDS might have similarities to the classical type, it might be a completely new type.

There are indications that this type of EDS has to be present in a homozygous and dominant manner (5). Skin biopsies of TNX-deficient patients revealed disrupted structures and reduced length of the elastic fibers.

Mao et al. (9) created TNXB knock out mice that showed the same skin findings as patients with EDS. Their skin biopsies histologically looked normal, but collagen was reduced, although the shape and size of single fibers were normal, the overall density was reduced. Interestingly in vitro studies showed that collagen synthesis was not affected in the knock out mice but deposition was.

It also seems like the Tenascin-X deficient type of EDS has many neuromuscular findings, more than the classical or hypermobile type of EDS and suggests that this type is a very debilitating form of EDS (10).

Pénisson-Besnier et al. (11) described one case of TNX deficiency with mainly myopathic features. This patient harbored a 30 kb deletion and a missense mutation on TNXB and suffered from progressive muscle weakness which biopsy findings confirmed.

Voermans et al. (12) reported two cases that had reduced quantitative muscle function with normal findings on EMG and muscle biopsy.

In patients with mainly neuromuscular findings, lack of atrophic scarring and no affected family members this diagnosis should be thought of (10).

Fortunately, one study suggests that there might be a little benefit people with TNX deficiency have compared to other people: TNX deficiency might protect against cardiovascular diseases like high blood pressure, prevents against abnormal pressure wave propagation and accommodates atherosclerotic plaques (13). There does not seem to be a higher risk for an aneurysm or other cardiovascular complications that are reported mainly in vascular EDS (14).

TNX haploinsufficient EDS

Haploinsufficiency means that patients had around 50 percent less Tenascin-X in blood compared to healthy people. This EDS type is at the moment considered to resemble the hypermobile type of EDS, but patients did not show the typical skin features or vascular fragility.

Studies suggest that patients with heterozygous mutations in the TNXB gene will not show a complete deficiency of TNX but more a haploinsufficiency (5). Interestingly not all patients with heterozygous mutations had signs of EDS which led to the conclusion that EDS caused by haploinsufficiency needs some outside influences to uncover the whole phenotype.

Zweers et al. (5) also described that there was a comparable high number of TNX haploinsufficient people found in the Joint Hypermobility Syndrome (JHS) group which again suggests that EDS-HT and JHS are the same conditions.

Congenital adrenal hyperplasia (CAH)-EDS

The connection between CAH and TNXB EDS was the first to be described and led to the discovery of the role of TNX in EDS.

CAH is a congenital disease of the adrenal glands (congenital adrenal hyperplasia) and caused by mutations in the CYP21B gene that encodes for the steroid enzyme 21-hydroxylase. This enzyme is very important for the synthesis of cortisol and mineralocorticoids. Deficiency of this enzyme leads to a deficiency in above mentioned and has to be substituted for the rest of the life.

CYP21B is the neighbor gene of TNXB and patients with gene defects that include the junction between both genes showed the phenotypes of CAH and EDS – called CAH-X. That means that these group of people have characteristics of endocrine imbalances and an apparent EDS phenotype including joint hypermobility and skin laxity with normal healing (15).

The first described patient was a young man showing signs of salt wasting and hyperextensible skin, hypermobile joints and bruising, but no atrophic scarring. Due to this patient, Tenascin-XB was discovered and studied as a potential candidate gene in EDS.

It was believed that those larger deletions that encompass CYP21B as well as TNXB occur in 1 of 10 CAH patients (5).

Recent studies suggest a prevalence of 9 % of CAH-X in all CAH patients (15).

In 2014 two studies suggested an involvement of the transforming growth factor beta in the pathogenesis of CAH-X. This was proven individually by Alcaraz et al. (8) who have found that a TNX domain is crucial for regulating the bioavailability of TGFbeta and Morisette et al. (16) who showed in their study that TGFbeta biomarkers like pSmad (bone morphogenic protein) were increased in fibroblast cultures together with TGFbeta3 and MMP-13, suggesting a link between CAH-X and the TGFbeta pathway.

In a small case report consisting of a three generation family Chen et al. (17) showed the variety of CAH-X phenotypes. Additionally to the joint hypermobility the family suffered from midline defects, cardiac valvular abnormalities, single kidney, bicornuate uterus and a bifid uvula. Suggesting that CAH-X patient should have routine screening for heart abnormalities.

Merke et al. (18) described the phenotype of their CAH-X patients with TNX haploinsufficiency as hypermobile, with piezogenic papules, soft tissue rheumatism, hernia, spondylosis und functional bowel disorder, osteoporosis, cardiac abnormalities, and prolapses.

figure-2-001Figure 2

TNXB mutations which do not lead to TNX deficiency

Tenascin-X deficiency was only described in around 20 individuals by now, whereas CAH-X seems to be much more common. But the most common form of TNXB abnormalities seems to be mutations on TNXB that do NOT lead to TNX haploinsufficiency or deficiency. Those group of patients have a variety of missense mutations everywhere on TNXB without knowing if those are the cause of EDS or not. There are only two publications that tried to find out if missense mutations on TNXB can also lead to EDS.

In one publication Zweers et al. (19) described 3 cases of missense mutations that did not lead to TNX deficiency or haploinsufficiency. Two of those three patients were examined and showed EDS phenotypes.

The two clinically examined patients were tested for abnormalities in their skin biopsy and one showed an increase in elastic fiber length whereas the other one showed no difference at all. Due to mutation prediction software the one that did not show any elastic fiber difference was later proven to be not disease causing.

The third mutation where no skin biopsy was obtained is probably disease causing too because this patient carried a mutation that changed one Arginine residue of TNX to a Tryptophan residue, both of which are biochemically very different.

Based on the above-mentioned missense mutations a Canadian group picked one of them and performed homology modeling, denaturation, single molecular, atomic force microscopy and molecular dynamic techniques based on already known Tenascin X domain structures that were available. Those simulations revealed that the mutation is altered the flexibility of a loop which then negatively affected binding between TNX and other molecules (20).

Diagnostics

Tenascin-X protein Essay (blood sample) (10)
TNXB gene analysis (blood sample or mouth swab)

Overlap with Type VI Myopathies

Since Tenascin X plays a huge role in the expression of type VI collagen, it is not surprising that especially the TNX-deficient type shows some huge overlaps with type VI collagen myopathies like Ulrich congenital muscular dystrophy (UCMD) and Bethlem myopathy (BM) (1). Those Myopathies show moderate to severe muscular dystrophy and joint hypermobility, especially of the distal joints. The main characteristics of those myopathies are the typical joint contractures which are usually not present in EDS.

Kirschner et al. (21) discovered through electron microscopy of skin biopsies from UCMD many abnormalities of collagen fiber morphology which led them to conclude that there is a morphological overlap between UCMD and EDS.

Another study showed that TNX deficiency led to in vitro decrease of type IV collagen suggestive of an overlap between TNX-deficient EDS and type VI collagen myopathies (22).

Those in vitro studies and results are supported by clinical findings in TNX-deficient patients accomplished by Voermans et al. (23, 24) who proves a clear relationship between neuromuscular involvement including muscle weakness and contractures and TNX deficiency.

 

figure-3-001Figure 3

Other conditions with possible TNX involvement

One study in Japan identified a single nucleotide polymorphism that seems to be the cause for some Japanese Lupus cases (25).

Another study showed the involvement of Tenascin C and XB in Neurofibromatosis type 1 (26).

Gene expression analysis showed that TNXB was highly expressed in malignant mesothelioma which makes TNX a possible new diagnostic marker to differentiate malignant mesothelioma against other serosal cavity tumors (27).

Very recently an overlapping Osteogenesis imperfecta and EDS phenotype due to COL1A1 mutation and a biallelic missense mutation in TNXB was identified via whole exome sequencing by a German group (28).

Conclusion and Discussion

A lot of research has been happening since the first discovery of Tenascin X in a CAH patient but still, most is unknown about the mechanism of TNX in the human body and how TNXB mutations lead to EDS.

Only around 20 patients with TNX deficiency or Haploinsufficiency were published so far, some more related to CAH-X and only 2 TNXB missense mutations were proven to be causative for EDS-HT.

And despite the clearly severe disease progression and its overlap with myopathies, there is no commercially available TNX test in the U.S., and only a few EDS patients are screened for TNXB mutations.

I believe that there are a lot of EDS-HT patients that are positive for especially TNXB missense mutation and only basic screening for those mutations can lead us to any progress concerning the disease mechanism and also the progression.

It is not only important for doctors to know about their patient’s risks due to TNX deficiency but more important is the patient’s personal peace of mind. Many patients are severely suffering from the pressure of having a disease and not knowing what caused it. For the patients understanding the cause makes it easier to cope with their limitations.

Most of the time the diagnosis of hypermobile EDS is based on clinical findings without genetic evidence which makes it likely for other physicians or insurances to doubt those findings and the patient can end up without a support system – socially and financially.

Bristow et al. (4) stated in 2005 that the discovery of other proteins that are involved in collagen formation, deposition or organization could provide potentially causative genes in EDS-HT and lead to 30 – 50 percent resolution of the at the moment genetic unclear EDS-HT cases.

11 years later and we are still at the same point and have not identified more potential genes. This fact shows that there is a lot more research needed to give those patient group the answers they desperately need.

Resources:

  1. Van Damme T, Syx D, Coucke P, Symoens S, De Paepe A, Malfait F. Genetics of the Ehlers–Danlos syndrome: more than collagen disorders. Expert Opinion on Orphan Drugs. 2015 Apr 3;3(4):379-92.
    OMIM entry 600985 Tenascin XB; TNXB
  2. Chiquet-Ehrismann R, Tucker RP. Tenascins and the importance of adhesion modulation. Cold Spring Harbor perspectives in biology. 2011 May 1;3(5):a004960.
  3. Bristow J, Carey W, Egging D, Schalkwijk J. Tenascin‐X, collagen, elastin, and the Ehlers–Danlos syndrome. InAmerican Journal of Medical Genetics Part C: Seminars in Medical Genetics 2005 Nov 15 (Vol. 139, No. 1, pp. 24-30). Wiley Subscription Services, Inc., A Wiley Company.
  4. Zweers MC, Hakim AJ, Grahame R, Schalkwijk J. Joint hypermobility syndromes: The pathophysiologic role of tenascin‐X gene defects. Arthritis & Rheumatism. 2004 Sep 1;50(9):2742-9.
  5. Byers PH, Murray ML. Ehlers–Danlos syndrome: A showcase of conditions that lead to understanding matrix biology. Matrix Biology. 2014 Jan 31;33:10-5.
  6. Valcourt U, Alcaraz LB, Exposito JY, Lethias C, Bartholin L. Tenascin-X: beyond the architectural function. Cell adhesion & migration. 2015 Jan 2;9(1-2):154-65.
  7. Alcaraz LB, Exposito JY, Chuvin N, Pommier RM, Cluzel C, Martel S, Sentis S, Bartholin L, Lethias C, Valcourt U. Tenascin-X promotes epithelial-to-mesenchymal transition by activating latent TGF-β. The Journal of cell biology. 2014 May 12;205(3):409-28.
  8. Mao JR, Taylor G, Dean WB, Wagner DR, Afzal V, Lotz JC, Rubin EM, Bristow J. Tenascin-X deficiency mimics Ehlers-Danlos syndrome in mice through alteration of collagen deposition. Nature genetics. 2002 Apr 1;30(4):421-5.
    Schalkwijk J, Zweers MC, Steijlen PM, Dean WB, Taylor G, van Vlijmen IM, van Haren B, Miller WL, Bristow J. A recessive form of the Ehlers–Danlos syndrome caused by tenascin-X deficiency. New England Journal of Medicine. 2001 Oct 18;345(16):1167-75.
  9. Pénisson-Besnier I, Allamand V, Beurrier P, Martin L, Schalkwijk J, van Vlijmen-Willems I, Gartioux C, Malfait F, Syx D, Macchi L, Marcorelles P. Compound heterozygous mutations of the TNXB gene cause primary myopathy. Neuromuscular Disorders. 2013 Aug 31;23(8):664-9.
  10. Voermans NC, Altenburg TM, Hamel BC, de Haan A, Van Engelen BG. Reduced quantitative muscle function in tenascin-X deficient Ehlers-Danlos patients. Neuromuscular Disorders. 2007 Aug 31;17(8):597-602.
  11. Petersen JW, Douglas JY. Tenascin-X, collagen, and Ehlers–Danlos syndrome: Tenascin-X gene defects can protect against adverse cardiovascular events. Medical hypotheses. 2013 Sep 30;81(3):443-7.
  12. Peeters AC, Kucharekova M, Timmermans J, Van Den Berkmortel FW, Boers GH, Novakova IR, Egging D, Heijer MD, Schalkwijk J. A clinical and cardiovascular survey of Ehlers-Danlos syndrome patients with complete deficiency of tenascin-X.
  13. Morissette R, Chen W, Perritt AF, Dreiling JL, Arai AE, Sachdev V, Hannoush H, Mallappa A, Xu Z, McDonnell NB, Quezado M. Broadening the spectrum of Ehlers-Danlos syndrome in patients with congenital adrenal hyperplasia. The Journal of Clinical Endocrinology & Metabolism. 2015 Jun 15;100(8): E1143-52.
  14. Morissette R, Merke DP, McDonnell NB. Transforming growth factor-β (TGF-β) pathway abnormalities in tenascin-X deficiency associated with CAH-X syndrome. European journal of medical genetics. 2014 Feb 28;57(2):95-102.
    Chen W, Kim MS, Shanbhag S, Arai A, VanRyzin C, McDonnell NB, Merke DP. The phenotypic spectrum of contiguous deletion of CYP21A2 and tenascin XB: quadricuspid aortic valve and other midline defects. American Journal of Medical Genetics Part A. 2009 Dec 1;149(12):2803-8.
  15. Merke DP, Chen W, Morissette R, Xu Z, Van Ryzin C, Sachdev V, Hannoush H, Shanbhag SM, Acevedo AT, Nishitani M, Arai AE. Tenascin-X haploinsufficiency associated with Ehlers-Danlos syndrome in patients with congenital adrenal hyperplasia. The Journal of Clinical Endocrinology & Metabolism. 2013 Jan 2;98(2): E379-87.
    Zweers, M. C., et al. “Elastic fiber abnormalities in hypermobility type Ehlers–Danlos syndrome patients with tenascin‐X mutations.” Clinical Genetics 67.4 (2005): 330-334.
  16. Zhuang S, Linhananta A, Li H. Phenotypic effects of Ehlers–Danlos syndrome‐associated mutation on the FnIII domain of tenascin‐X. Protein Science. 2010 Nov 1;19(11):2231-9.
  17. Kirschner J, Hausser I, Zou Y, Schreiber G, Christen HJ, Brown SC, Anton‐Lamprecht I, Muntoni F, Hanefeld F, Bönnemann CG. Ullrich congenital muscular dystrophy: connective tissue abnormalities in the skin support overlap with Ehlers–Danlos syndromes. American Journal of Medical Genetics Part A. 2005 Jan 30;132(3):296-301.
  18. Minamitani T, Ariga H, Matsumoto KI. Deficiency of tenascin-X causes a decrease in the level of expression of type VI collagen. Experimental cell research. 2004 Jul 1;297(1):49-60.
  19. Voermans NC, van Alfen N, Pillen S, Lammens M, Schalkwijk J, Zwarts MJ, van Rooij IA, Hamel BC, van Engelen BG. Neuromuscular involvement in various types of Ehlers–Danlos syndrome. Annals of neurology. 2009 Jun 1;65(6):687-97.
  20. Voermans NC, Jenniskens GJ, Hamel BC, Schalkwijk J, Guicheney P, Van Engelen BG. Ehlers–Danlos syndrome due to tenascin‐X deficiency: Muscle weakness and contractures support overlap with collagen VI myopathies. American Journal of Medical Genetics Part A. 2007 Sep 15;143(18):2215-9.
  21. Kamatani Y, Matsuda K, Ohishi T, Ohtsubo S, Yamazaki K, Iida A, Hosono N, Kubo M, Yumura W, Nitta K, Katagiri T. Identification of a significant association of a single nucleotide polymorphism in TNXB with systemic lupus erythematosus in a Japanese population. Journal of human genetics. 2008 Jan 1;53(1):64-73.
  22. Lévy P, Ripoche H, Laurendeau I, Lazar V, Ortonne N, Parfait B, Leroy K, Wechsler J, Salmon I, Wolkenstein P, Dessen P. Microarray-based identification of tenascin-C and tenascin XB, genes possibly involved in tumorigenesis associated with neurofibromatosis type 1. Clinical cancer research. 2007 Jan 15;13(2):398-407.
  23. Yuan Y, Nymoen DA, Stavnes HT, Rossnes AK, Bjørang O, Wu C, Nesland JM, Davidson B. Tenascin-X is a novel diagnostic marker of malignant mesothelioma. The American journal of surgical pathology. 2009 Nov;33(11):1673.
  24. Mackenroth L, Fischer‐Zirnsak B, Egerer J, Hecht J, Kallinich T, Stenzel W, Spors B, von Moers A, Mundlos S, Kornak U, Gerhold K. An overlapping phenotype of Osteogenesis imperfecta and Ehlers–Danlos syndrome due to a heterozygous mutation in COL1A1 and biallelic missense variants in TNXB identified by whole exome sequencing. American Journal of Medical Genetics Part A. 2016 Jan 1.

 

Information about the author:

karinaThe author, Karina Sturm, suffers from EDS and is classified as having Ehlers-Danlos syndrome – Hypermobile Type with an unpublished but considered pathological TNXB mutation. Because of this fact, she reads everything she can about Tenascin X. She hopes by finding out more about her mutation she somehow learns more about her disease’s progression. With her scientific background, she tries to inform other patients in her country, Germany. All information she gives is based on scientific publications.

For more information visit our TNXB EDS Facebook Group:

https://www.facebook.com/groups/TNXBandEDS

If you want to learn more about Karina, and the work she is doing, please visit her website:

http://www.instabile-halswirbelsaeule.de/en/