Recurrent pneumothorax in a case of tenascin-X deficient Ehlers-Danlos syndrome: Broadening the phenotypic spectrum.

The genomic region surrounding the Tenascin-XB gene (TNXB) is a complex and duplicated region, with several pseudogenes that predispose to high rates of homologous recombination. Classical-like Ehlers-Danlos syndrome (clEDS) is the result of tenascin-X deficiency due to biallelic loss of function variants in the TNXB gene. Here we present a patient with clEDS and spontaneous pneumothorax, a feature not previously reported to be associated with this condition. Two inherited pathogenic/likely pathogenic variants were identified; a previously reported deletion resulting in a TNXA/TNXB chimeric gene and a novel frameshift variant. The Tenascin-XB gene is well described in the literature to be associated with collagen metabolism, stabilization of the fibrillar-collagen matrix and is expressed abundantly in the extracellular matrix. We propose that tenascin-X deficiency is directly related to pneumothorax predisposition. This case expands the phenotypic spectrum of clEDS and highlights the challenges with molecular analysis and diagnosis.

Rare neurological manifestations in a Saudi Arabian patient with Ehlers-Danlos syndrome and a novel homozygous variant in the TNXB gene.

We report a 38-year-old Saudi male with Ehlers-Danlos Syndrome (EDS). The patient presented with rare and unusual neurological manifestations, including but not limited to ophthalmoplegia and myopathic pattern on his electromyography. In addition to hand weakness, there was skin hyperextensibility, joint hyperflexibility, and frontal baldness. Next-generation sequencing was performed on target exon sequences, using whole exome sequencing and Burrows-Wheeler Aligner for alignment/base calling. Genome Analysis Toolkit and reference genome Homo sapiens (UCSC hg19) were used for sequence processing and analysis. Variant classification was done according to standard international recommendations. A novel homozygous variant, NM_019105.6: c.8488C>T p.(Gln2830*), was detected in the TNXB gene. This variant is not reported in the literature nor dbSNP or gnomAD databases. Additionally, this variant is predicted to create a premature stop codon and produce a truncated protein or nonsense-mediated mRNA decay. Hence, it is classified as a likely pathogenic variant. The same point variant was found in a heterozygous state in the patient's father and sister. Both presented with milder symptoms associated with Ehlers-Danlos syndromes and heritable connective tissue disorders. Therefore, the patient was diagnosed as a tenascin-X (TNX) deficient type of EDS known as classical-like Ehlers-Danlos syndrome. TNX deficient patients may present with clinical and electrophysiological manifestations that are unusual in EDS like frontal baldness, ophthalmoplegia, and myotonia, which mimic myotonic dystrophy type I. Clinicians should be aware of the potential overlap of symptoms among these two diseases to ensure correct diagnosis is made.

Classical-like Ehlers-Danlos syndrome: a clinical description of 20 newly identified individuals with evidence of tissue fragility.

PURPOSE: Currently, 31 patients with classical-like EDS (clEDS) due to tenascin-X deficiency have been reported in the literature. We report on the clinical and molecular characteristics of 20 additional patients with clEDS to expand knowledge and to enable improved management of this rare genetic disorder. METHODS: Patients diagnosed with clEDS by the national EDS service in the UK (n = 21) and abroad (n = 1) were asked for consent for publication of their clinical and molecular data. RESULTS: Of 22 patients, 20 consented. All patients had typical features of clEDS: joint hypermobility, easy bruising, and skin hyperextensibility without atrophic scars. Importantly, 3/20 patients experienced gastrointestinal complications consisting of small or large bowel ruptures and one esophageal rupture. Other notable observations included two separate occurrences of spontaneous compartment syndrome, suspicion of nonaccidental injury due to significant bruising, and significant clinical variability regarding the debilitating effect of joint dislocations. CONCLUSIONS: We propose a predisposition to tissue fragility, particularly of the gastrointestinal tract in patients with clEDS. As such, clinical and molecular confirmation of this diagnosis is essential. It is recommended to follow up these patients closely to understand the natural history to develop better recommendations for management.

The role of tenascin-X in the uterosacral ligaments of postmenopausal women with pelvic organ prolapse: an immunohistochemical study.

INTRODUCTION AND HYPOTHESIS: Abnormalities of connective tissue structure or its repair mechanism may predispose women to pelvic organ prolapse (POP). We hypothesized that the expression of tenascin-X in the uterosacral ligament of postmenopausal women with symptomatic POP is increased compared with postmenopausal women without POP. Furthermore, we identified clinical risk factors associated with POP in our study population. METHODS: We conducted a retrospective case-control study in which 33 postmenopausal women with symptomatic POP ≥ pelvic organ prolapse quantification system (POP-Q) stage II were matched with 33 postmenopausal women without POP. Studied tissue specimens were taken from hysterectomy specimens, and tenascin-X expression was investigated by immunohistochemistry. The immunohistochemical profile of the uterosacral connective tissue of cases and controls was compared. RESULTS: Tenascin-X was expressed in 94% of POP cases and in 91% of controls. Our study failed to show any statistically significant differences in tenascin-X expression between women with and without POP (p = 0.64). However, tenascin-X was significantly more expressed in cases with severe prolapse (POP-Q stage IV) compared with moderate prolapse stages (POP-Q stage II and III) (p = 0.001). Advanced patient age as well as early menopausal age remained independent risk factors associated with POP in multiple logistic regression analysis (p = 0.001). CONCLUSION: No difference could be demonstrated between tenascin-X expression in patients with or without POP. Tenascin-X does not seem to play a major role in the pathogenesis of POP in postmenopausal women.

TNX deficiency results in bone loss due to an increase in multinucleated osteoclasts.

Tenascin-X (TNX), a glycoprotein of the extracellular matrix (ECM), is expressed in various tissues and plays an important role in ECM architecture. The TNXB gene encoding TNX is known as the gene responsible for classic-like Ehlers-Danlos syndrome (clEDS). To date, the role of TNX in dermal, muscular and obstetric features has been reported, but its role in bone homeostasis remains to be clarified. In this study, we found significant bone loss and upregulation of osteoclast marker gene expression in TNX-deficient mice. Further, TNX deficiency in the bone marrow promoted multinucleation of osteoclasts and resulted in increased bone resorption activity. These results indicate that multinucleated osteoclasts are the cause of bone loss in a TNX-deficient environment. Our findings provide new insight into the mechanism of osteoclast differentiation mediated by TNX and the pathology of clEDS.

Wound healing-related properties detected in an experimental model with a collagen gel contraction assay are affected in the absence of tenascin-X.

Patients with tenascin-X (TNX)-deficient type Ehlers-Danlos syndrome (EDS) do not exhibit delayed wound healing, unlike classic type EDS patients, who exhibit mutations in collagen genes. Similarly, in TNX-knockout (KO) mice, wound closure of the skin is normal even though these mice exhibit a reduced breaking strength. Therefore, we speculated that the wound healing process may be affected in the absence of TNX. In this study, to investigate the effects of TNX absence on wound healing-related properties, we performed collagen gel contraction assays with wild-type (WT) and TNX-KO mouse embryonic fibroblasts (MEFs). Collagen gels with embedded TNX-KO MEFs showed significantly greater contraction than those containing WT MEFs. Subsequently, we assessed collagen gel contraction-related properties, such as the activities of matrix metalloproteinase (MMP)-2 and MMP-9 and the protein and mRNA expression levels of transforming growth factor ß1 (TGF-ß1) in the collagen gels. The activities of MMP-2 and MMP-9 and the expression level of TGF-ß1 were elevated in the absence of TNX. Furthermore, filopodia-like protrusion formation, cell proliferation, migration, and collagen expression in MEFs were promoted in the absence of TNX. These results indicate that these wound healing-related properties are affected in a TNX-deficient extracellular environment.

Measurement of Serum Tenascin-X in Joint Hypermobility Syndrome Patients.

Joint hypermobility syndrome (JHS) (also termed hypermobility type Ehlers-Danlos syndrome, hEDS) is a heritable connective tissue disorder that is characterized by generalized joint hypermobility, chronic pain, fatigue, and minor skin changes. Initially, it was reported that there is a small subset of patients with JHS/hEDS who have haploinsufficiency of tenascin-X (TNX). However, the relationship between TNXB and JHS/hEDS has not been reported at all afterwards. EDS was reclassified into thirteen types in 2017, and the causative gene of JHS/hEDS remained to be identified. Therefore, in this study in order to determine whether JHS/hEDS can be diagnosed by the concentrations of serum form of TNX (sTNX), we measured the concentrations of sTNX in 17 JHS/hEDS patients. The sTNX concentrations in half of the JHS/hEDS patients were significantly lower than those in healthy individuals. No mutations, insertions or deletions were detected in the TNX exon sequence of the JHS/hEDS patients except for one in patient. That patient has a heterozygous mutation. A correlation between sTNX concentration and mutation of the TNXB genomic sequence was not found in the JHS/hEDS patients. These results indicate that the decrease in sTNX concentration could be used as a risk factor for JHS/hEDS.

Loss of tenascin X gene function impairs injury-induced stromal angiogenesis in mouse corneas.

To determine the contribution by tenascin X (Tnx) gene expression to corneal stromal angiogenesis, the effects were determined of its loss on this response in TNX knockout (KO) mice. In parallel, the effects of such a loss were evaluated on vascular endothelial growth factor (VEGF) and transforming growth factor ß1 (TGFß1) gene and protein expression in fibroblasts and macrophages in cell culture. Histological, immunohistochemical and quantitative RT-PCR changes determined if Tnx gene ablation on angiogenic gene expression, inflammatory cell infiltration and neovascularization induced by central corneal stromal cauterization. The role was determined of Tnx function in controlling VEGF-A or TGFß1 gene expression by comparing their expression levels in ocular fibroblasts and macrophages obtained from wild-type (WT) and body-wide Tnx KO mice. Tnx was up-regulated in cauterized cornea. In Tnx KO, macrophage invasion was attenuated, VEGF-A and its cognate receptor mRNA expression along with neovascularization were lessened in Tnx KOs relative to the changes occurring in their WT counterpart. Loss of Tnx instead up-regulated in vivo mRNA expression of anti-angiogenic VEGF-B but not VEGF-A. On the other hand, TGFß1 mRNA expression declined in Tnx KO cultured ocular fibroblasts. Loss of Tnx gene expression caused VEGF-A expression to decline in macrophages. Tnx gene expression contributes to promoting TGFß1 mRNA expression in ocular fibroblasts and VEGF-A in macrophages, macrophage invasion, up-regulation of VEGF-A expression and neovascularization in an injured corneal stroma. On the other hand, it suppresses anti-angiogenic VEGF-B mRNA expression in vivo.

Recognizing the tenascin-X deficient type of Ehlers-Danlos syndrome: a cross-sectional study in 17 patients.

The tenascin-X (TNX) deficient type Ehlers-Danlos syndrome (EDS) is similar to the classical type of EDS. Because of the limited awareness among geneticists and the challenge of the molecular analysis of the TNXB gene, the TNX-deficient type EDS is probably to be under diagnosed. We therefore performed an observational, cross-sectional study. History and physical examination were performed. Results of serum TNX measurements were collected and mutation analysis was performed by a combination of next-generation sequencing (NGS), Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA). Included were 17 patients of 11 families with autosomal recessive inheritance and childhood onset. All patients had hyperextensible skin without atrophic scarring. Hypermobility of the joints was observed in 16 of 17 patients. Deformities of the hands and feet were observed frequently. TNX serum level was tested and absent in 11 patients (seven families). Genetic testing was performed in all families; 12 different mutations were detected, most of which are suspected to lead to non-sense mRNA mediated decay. In short, patients with the TNX-deficient type EDS typically have generalized joint hypermobility, skin hyperextensibility and easy bruising. In contrast to the classical type, the inheritance pattern is autosomal recessive and atrophic scarring is absent. Molecular analysis of TNXB in a diagnostic setting is challenging.

Excavatolide B inhibits nonsmall cell lung cancer proliferation by altering peroxisome proliferator activated receptor gamma expression and PTEN/AKT/NF-Kß expression.

Marine organisms are proven to be rich source of secondary metabolites that can be used to treat various diseases. Excavatolide B (Exc.B), the most abundant metabolite was found in the marine coral Briareum excavatum exhibits cytotoxic effects against lung cancer cell. Treatment of the A549 cells with Exc.B significantly reduced its cell viability and induced cell cycle arrest at subG1 phase in a dose- and time-dependent manner, respectively. Apoptosis induction by Exc.B was further confirmed by decreased pro-caspase 3 expressions and increased proteolytic cleavage of poly (ADP-ribose) polymerase (PARP) expression. Furthermore, Exc.B increased reactive oxygen species (ROS) and reactive nitrogen species (RNS) and also decreased the antioxidant enzymes such as, Catalase, GPx, SOD, GST, and GSH. The proteomic analysis data revealed that total thirty six proteins were altered by Exc.B. STRING database showed that most of the altered proteins have no interaction between each other. Based on these data, KSR1, RuVBL2, PPAR-γ, and Tenascin X proteins were chosen to validate the 2DE data by Western blotting. Additional experiments demonstrated that Exc.B induced PTEN expression and inhibited pAKT and NF-kB expression. These results provide a novel insight into mechanisms underlying the inhibition of A549 cells growth by excavatolide B. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 290-301, 2017.

Ehlers-Danlos Syndrome Caused by Biallelic TNXB Variants in Patients with Congenital Adrenal Hyperplasia.

Some variants that cause autosomal-recessive congenital adrenal hyperplasia (CAH) also cause hypermobility type Ehlers-Danlos syndrome (EDS) due to the monoallelic presence of a chimera disrupting two flanking genes: CYP21A2, encoding 21-hydroxylase, necessary for cortisol and aldosterone biosynthesis, and TNXB, encoding tenascin-X, an extracellular matrix protein. Two types of CAH tenascin-X (CAH-X) chimeras have been described with a total deletion of CYP21A2 and characteristic TNXB variants. CAH-X CH-1 has a TNXB exon 35 120-bp deletion resulting in haploinsufficiency, and CAH-X CH-2 has a TNXB exon 40 c.12174C>G (p.Cys4058Trp) variant resulting in a dominant-negative effect. We present here three patients with biallelic CAH-X and identify a novel dominant-negative chimera termed CAH-X CH-3. Compared with monoallelic CAH-X, biallelic CAH-X results in a more severe phenotype with skin features characteristic of classical EDS. We present evidence for disrupted tenascin-X function and computational data linking the type of TNXB variant to disease severity.

Congenital adrenal hyperplasia with a CYP21A2 deletion overlapping the tenascin-X gene: an atypical presentation.

OBJECTIVES: Congenital Adrenal Hyperplasia (CAH) is a group of genetic diseases characterized by impaired cortisol biosynthesis. 95% of CAH cases result from mutation in the CYP21A2 gene encoding 21-hydroxilase. TNX-B gene partially overlaps CYP21A2 and encodes a matrix protein called Tenascin-X (TNX). Complete tenascin deficiency causes Enlers-Danlos syndrome (EDS). A mono allelic variant called CAH-X CH-1 was recently described, resulting from a CYP21A2 complete deletion that extends into the TNXB. This haploinsufficiency of TNX may be associated with a mild hypermobility form of EDS, as well as other connective tissue comorbidities such as hernia, cardiac defects and chronic arthralgia. CASE PRESENTATION: We report four patients heterozygous for a CAH-X CH-1 allele that do not present clinical manifestations of the EDS. CONCLUSIONS: All CAH patients, carriers of these TNXA/TNXB chimeras, should be evaluated for clinical manifestations related to connective tissue hypermobility, cardiac abnormalities and other EDS features, allowing for better clinical surveillance management.

Tenascin-X as a causal gene for classical-like Ehlers-Danlos syndrome.

Tenascin-X (TNX) is an extracellular matrix glycoprotein for which a deficiency results in a recessive form of classical-like Ehlers-Danlos syndrome (clEDS), a heritable connective tissue disorder with hyperextensible skin without atrophic scarring, joint hypermobility, and easy bruising. Notably, patients with clEDS also suffer from not only chronic joint pain and chronic myalgia but also neurological abnormalities such as peripheral paresthesia and axonal polyneuropathy with high frequency. By using TNX-deficient (Tnxb -/-) mice, well-known as a model animal of clEDS, we recently showed that Tnxb -/- mice exhibit hypersensitivity to chemical stimuli and the development of mechanical allodynia due to the hypersensitization of myelinated A-fibers and activation of the spinal dorsal horn. Pain also occurs in other types of EDS. First, we review the underlying molecular mechanisms of pain in EDS, especially that in clEDS. In addition, the roles of TNX as a tumor suppressor protein in cancer progression have been reported. Recent in silico large-scale database analyses have shown that TNX is downregulated in various tumor tissues and that high expression of TNX in tumor cells has a good prognosis. We describe what is so far known about TNX as a tumor suppressor protein. Furthermore, some patients with clEDS show delayed wound healing. Tnxb -/- mice also exhibit impairment of epithelial wound healing in corneas. TNX is also involved in liver fibrosis. We address the molecular mechanism for the induction of COL1A1 by the expression of both a peptide derived from the fibrinogen-related domain of TNX and integrin α11.

Clinical and molecular delineation of classical-like Ehlers-Danlos syndrome through a comprehensive next-generation sequencing-based screening system.

Classical-like Ehlers-Danlos syndrome (clEDS) is an autosomal recessive disorder caused by complete absence of tenascin-X resulting from biallelic variation in TNXB. Thus far, 50 patients from 43 families with biallelic TNXB variants have been identified. Accurate detection of TNXB variants is challenging because of the presence of the pseudogene TNXA, which can undergo non-allelic homologous recombination. Therefore, we designed a genetic screening system that is performed using similar operations to other next-generation sequencing (NGS) panel analyses and can be applied to accurately detect TNXB variants and the recombination of TNXA-derived sequences into TNXB. Using this system, we identified biallelic TNXB variants in nine unrelated clEDS patients. TNXA-derived variations were found in >75% of the current cohort, comparable to previous reports. The current cohort generally exhibited similar clinical features to patients in previous reports, but had a higher frequency of gastrointestinal complications (e.g., perforation, diverticulitis, gastrointestinal bleeding, intestinal obstruction, rectal/anal prolapse, and gallstones). This report is the first to apply an NGS-based screening for TNXB variants and represents the third largest cohort of clEDS, highlighting the importance of increasing awareness of the risk of gastrointestinal complications.

COL1A1 expression induced by overexpression of both a 15­amino acid peptide from the fibrinogen domain of tenascin­X and integrin α11 in LX­2 cells.

Extracellular matrix tenascin­X (TNX) is the largest member of the tenascin family. Our previous study demonstrated that TNX was involved in hepatic dysfunction, including fibrosis, in mice that were administered a high­fat and high­cholesterol diet with high levels of phosphorus and calcium. The present study investigated whether overexpression of both the fibrinogen domain of TNX (TNX­FG) and integrin α11, one of the TNX cell surface receptors, induces in vitro fibrosis in LX­2 human hepatic stellate cells. Overexpression of both a 15­amino acid peptide (hTNX­FGFFFF) derived from the TNX­FG domain and integrin α11 induced the expression of type I collagen α1 chain (COL1A1). Treatment with verteporfin [YAP (Yes­associated protein) inhibitor] attenuated the elevated COL1A1 expression elicited by overexpression of both hTNX­FGFFFF and integrin α11. In addition, small interfering RNA­mediated knockdown of YAP1 resulted in a decrease in COL1A1 expression induced by overexpression of both hTNX­FGFFFF and integrin α11. These results indicated that overexpression of both hTNX­FGFFFF and integrin α11 induced COL1A1 expression via the YAP signaling pathway.

Role of Tenascin-X in regulating TGF-ß/Smad signaling pathway in pathogenesis of slow transit constipation.

BACKGROUND: Chronic constipation is a gastrointestinal functional disease that seriously harms physical and mental health and impacts the quality of life of patients. Its incidence rate is 2%-27%. Slow transit constipation (STC) is a common type of chronic functional constipation, accounting for 10.3%-45.5% of such cases. Scholars have performed many studies on the pathogenesis of STC. These studies have indicated that the occurrence of STC may be related to multiple factors, such as dysfunction of the enteric nervous system, interstitial cells of Cajal (ICC) damage, and changes in neurotransmitters regulating intestinal peristalsis. AIM: To investigate the role of Tenascin-X (TNX) in regulating the TGF-ß/Smad signaling pathway in the pathogenesis of STC. METHODS: This study included an experimental group and a control group. The experimental group included 28 patients with severe colonic STC, and the control group included 18 patients with normal colon tissues. Immunohistochemistry (IHC) was used to detect c-Kit, a specific marker of the ICC. Western blot, immunofluorescence, and IHC were used to detect the localization and expression of TNX and TGF-ß/Smad. RESULTS: IHC showed that the number of ICC with positive c-Kit expression was significantly reduced in the colon of STC patients (22.17 ± 3.28 vs 28.69 ± 3.53, P < 0.05) and that the distribution was abnormal. Western blot results showed that c-Kit and Smad7 levels were significantly decreased in the colon of STC patients (c-kit: 0.462 ± 0.099 vs 0.783 ± 0.178, P < 0.01; Smad7: 0.626 ± 0.058 vs 0.799 ± 0.03, P < 0.01) and that TNX and Smad2/3 levels were higher in the STC group (TNX: 0.868 ± 0.028 vs 0.482 ± 0.032, P < 0.01). There was no significant difference in TGF-ß between the two groups (0.476 ± 0.028 vs 0.511 ± 0.044, P = 0.272). Pearson correlation analysis showed that the TNX protein exhibited a strong correlation with Smad2/3 and Smad7 (P < 0.05, |R| > 0.8) and TGF-ß (P < 0.05, |R| = 0.7). CONCLUSION: The extracellular matrix protein TNX may activate the TGF-ß/Smad signaling pathway by upregulating the Smad 2/3 signaling protein and thereby induce slight or complete epithelial stromal cell transformation, leading to an abnormal distribution and dysfunction of ICC in the diseased colon, which promotes the occurrence and development of STC.

The Roles of Tenascins in Cardiovascular, Inflammatory, and Heritable Connective Tissue Diseases.

Tenascins are a family of multifunctional extracellular matrix (ECM) glycoproteins with time- and tissue specific expression patterns during development, tissue homeostasis, and diseases. There are four family members (tenascin-C, -R, -X, -W) in vertebrates. Among them, tenascin-X (TNX) and tenascin-C (TNC) play important roles in human pathologies. TNX is expressed widely in loose connective tissues. TNX contributes to the stability and maintenance of the collagen network, and its absence causes classical-like Ehlers-Danlos syndrome (clEDS), a heritable connective tissue disorder. In contrast, TNC is specifically and transiently expressed upon pathological conditions such as inflammation, fibrosis, and cancer. There is growing evidence that TNC is involved in inflammatory processes with proinflammatory or anti-inflammatory activity in a context-dependent manner. In this review, we summarize the roles of these two tenascins, TNX and TNC, in cardiovascular and inflammatory diseases and in clEDS, and we discuss the functional consequences of the expression of these tenascins for tissue homeostasis.

Biallelic mutations in Tenascin-X cause classical-like Ehlers-Danlos syndrome with slowly progressive muscular weakness.

Tenascin-X, is an extracellular matrix glycoprotein expressed in skin, muscle, tendons, and blood vessels with an anti-adhesive function. Biallelic Tenascin-X mutations cause classical-like Ehlers-Danlos syndrome. We report a 46-year-old woman with slowly progressive weakness of the lower limbs and myalgia from age 28 years. In the past she had Raynaud's phenomenon, multiple sprains and joint dislocations, conjunctival haemorrhages and a colonic perforation during colonoscopy. Neurologic examination showed moderate asymmetric proximal and axial muscular weakness, distal amyotrophy of 4 limbs, moderate skin hyperextensibility, and hypermobility of distal joints of fingers. Whole body Magnetic Resonance Imaging showed symmetric fatty infiltration of thigh and leg muscles, with predominant atrophy of thighs. Next Generation Sequencing revealed two pathogenic TNXB variants, g.32024681C>G, c.7826-1G>C, and g.32016181dup, c.9998dupA, p.(Asn3333Lysfs*35). Western Blot and immunofluorescence studies confirmed a marked Tenascin-X reduction in both patient's serum and muscle. Here we further detail the clinical and genetic spectrum of a patient with classical-like Ehlers-Danlos syndrome and prominent muscle involvement.

Loss of Tenascin-X expression during tumor progression: A new pan-cancer marker.

Cancer is a systemic disease involving multiple components produced from both tumor cells themselves and surrounding stromal cells. The pro- or anti-tumoral role of the stroma is still under debate. Indeed, it has long been considered the main physical barrier to the diffusion of chemotherapy by its dense and fibrous nature and its poor vascularization. However, in murine models, the depletion of fibroblasts, the main ExtraCellular Matrix (ECM)-producing cells, led to more aggressive tumors even though they were more susceptible to anti-angiogenic and immuno-modulators. Tenascin-C (TNC) is a multifunctional matricellular glycoprotein (i.e. an ECM protein also able to induce signaling pathway) and is considered as a marker of tumor expansion and metastasis. However, the status of other tenascin (TN) family members and particularly Tenascin-X (TNX) has been far less studied during this pathological process and is still controversial. Herein, through (1) in silico analyses of the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases and (2) immunohistochemistry staining of Tissue MicroArrays (TMA), we performed a large and extensive study of TNX expression at both mRNA and protein levels (1) in the 6 cancers with the highest incidence and mortality in the world (i.e. lung, breast, colorectal, prostate, stomach and liver) and (2) in the cancers for which sparse data regarding TNX expression already exist in the literature. We thus demonstrated that, in most cancers, TNX expression is significantly downregulated during cancer progression and we also highlighted, when data were available, that high TNXB mRNA expression in cancer is correlated with a good survival prognosis.

Novel TNXB Variants in Two Italian Patients with Classical-Like Ehlers-Danlos Syndrome.

TNXB-related classical-like Ehlers-Danlos syndrome (TNXB-clEDS) is an ultrarare type of Ehlers-Danlos syndrome due to biallelic null variants in TNXB, encoding tenascin-X. Less than 30 individuals have been reported to date, mostly of Dutch origin and showing a phenotype resembling classical Ehlers-Danlos syndrome without atrophic scarring. TNXB-clEDS is likely underdiagnosed due to the complex structure of the TNXB locus, a fact that complicates diagnostic molecular testing. Here, we report two unrelated Italian women with TNXB-clEDS due to compound heterozygosity for null alleles in TNXB. Both presented soft and hyperextensible skin, generalized joint hypermobility and related musculoskeletal complications, and chronic constipation. In addition, individual 1 showed progressive finger contractures and shortened metatarsals, while individual 2 manifested recurrent subconjunctival hemorrhages and an event of spontaneous rupture of the brachial vein. Molecular testing found the two previously unreported c.8278C > T p.(Gln2760*) and the c.(2358 + 1_2359 - 1)_(2779 + 1_2780 - 1)del variants in Individual 1, and the novel c.1150dupG p.(Glu384Glyfs*57) and the recurrent c.11435_11524+30del variants in Individual 2. mRNA analysis confirmed that the c.(2358 + 1_2359 - 1)_(2779 + 1_2780 - 1)del variant causes a frameshift leading to a predicted truncated protein [p.(Thr787Glyfs*40)]. This study refines the phenotype recently delineated in association with biallelic null alleles in TNXB, and adds three novel variants to its mutational repertoire. Unusual digital anomalies seem confirmed as possibly peculiar of TNXB-clEDS, while vascular fragility could be more than a chance association also in this Ehlers-Danlos syndrome type.