Identification and functional modelling of plausibly causative cis-regulatory variants in a highly-selected cohort with X-linked intellectual disability.

Identifying causative variants in cis-regulatory elements (CRE) in neurodevelopmental disorders has proven challenging. We have used in vivo functional analyses to categorize rigorously filtered CRE variants in a clinical cohort that is plausibly enriched for causative CRE mutations: 48 unrelated males with a family history consistent with X-linked intellectual disability (XLID) in whom no detectable cause could be identified in the coding regions of the X chromosome (chrX). Targeted sequencing of all chrX CRE identified six rare variants in five affected individuals that altered conserved bases in CRE targeting known XLID genes and segregated appropriately in families. Two of these variants, FMR1CRE and TENM1CRE, showed consistent site- and stage-specific differences of enhancer function in the developing zebrafish brain using dual-color fluorescent reporter assay. Mouse models were created for both variants. In male mice Fmr1CRE induced alterations in neurodevelopmental Fmr1 expression, olfactory behavior and neurophysiological indicators of FMRP function. The absence of another likely causative variant on whole genome sequencing further supported FMR1CRE as the likely basis of the XLID in this family. Tenm1CRE mice showed no phenotypic anomalies. Following the release of gnomAD 2.1, reanalysis showed that TENM1CRE exceeded the maximum plausible population frequency of a XLID causative allele. Assigning causative status to any ultra-rare CRE variant remains problematic and requires disease-relevant in vivo functional data from multiple sources. The sequential and bespoke nature of such analyses renders them time-consuming and challenging to scale for routine clinical use.

Tenascin-C Deficiency Is Associated With Reduced Bacterial Outgrowth During Klebsiella pneumoniae-Evoked Pneumosepsis in Mice.

Tenascin C (TNC) is an extracellular matrix glycoprotein that recently emerged as an immunomodulator. TNC-deficient (TNC-/-) mice were reported to have a reduced inflammatory response upon systemic administration of lipopolysaccharide, the toxic component of gram-negative bacteria. Here, we investigated the role of TNC during gram-negative pneumonia derived sepsis. TNC+/+ and TNC-/- mice were infected with Klebsiella pneumoniae via the airways and sacrificed 24 and 42 h thereafter for further analysis. Pulmonary TNC protein levels were elevated 42 h after infection in TNC+/+ mice and remained undetectable in TNC-/- mice. TNC-/- mice showed modestly lower bacterial loads in lungs and blood, and a somewhat reduced local-but not systemic-inflammatory response. Moreover, TNC-/- and TNC+/+ mice did not differ with regard to neutrophil recruitment, lung pathology or plasma markers of distal organ injury. These results suggest that while TNC shapes the immune response during lipopolysaccharide-induced inflammation, this role may be superseded during pneumosepsis caused by a common gram-negative pathogen.

Mechanical allodynia in mice with tenascin-X deficiency associated with Ehlers-Danlos syndrome.

Tenascin-X (TNX) is a member of the extracellular matrix glycoprotein tenascin family, and TNX deficiency leads to Ehlers-Danlos syndrome, a heritable human disorder characterized mostly by skin hyperextensibility, joint hypermobility, and easy bruising. TNX-deficient patients complain of chronic joint pain, myalgia, paresthesia, and axonal polyneuropathy. However, the molecular mechanisms by which TNX deficiency complicates pain are unknown. Here, we examined the nociceptive behavioral responses of TNX-deficient mice. Compared with wild-type mice, TNX-deficient mice exhibited mechanical allodynia but not thermal hyperalgesia. TNX deficiency also increased pain sensitivity to chemical stimuli and aggravated early inflammatory pain elicited by formalin. TNX-deficient mice were significantly hypersensitive to transcutaneous sine wave stimuli at frequencies of 250 Hz (Aδ fiber responses) and 2000 Hz (Aß fiber responses), but not to stimuli at frequency of 5 Hz (C fiber responses). In addition, the phosphorylation levels of extracellular signal-related kinase, an active neuronal marker, and the activity of NADPH-diaphorase, a neuronal nitric oxide activation marker, were enhanced in the spinal dorsal horns of TNX-deficient mice. These results suggest that TNX deficiency contributes to the development of mechanical allodynia and hypersensitivity to chemical stimuli, and it induces hypersensitization of myelinated A fibers and activation of the spinal dorsal horn.

Tenascin-C deficiency impairs alveolarization and microvascular maturation during postnatal lung development.

After the airways have been formed by branching morphogenesis the gas exchange area of the developing lung is enlarged by the formation of new alveolar septa (alveolarization). The septa themselves mature by a reduction of their double-layered capillary networks to single-layered ones (microvascular maturation). Alveolarization in mice is subdivided into a first phase (postnatal days 4-21, classical alveolarization), where new septa are lifted off from immature preexisting septa, and a second phase (day 14 to adulthood, continued alveolarization), where new septa are formed from mature septa. Tenascin-C (TNC) is a multidomain extracellular matrix protein contributing to organogenesis and tumorigenesis. It is highly expressed during classical alveolarization, but afterward its expression is markedly reduced. To study the effect of TNC deficiency on postnatal lung development, the formation and maturation of the alveolar septa were followed stereologically. Furthermore, the number of proliferating (Ki-67-positive) and TUNEL-positive cells was estimated. In TNC-deficient mice for both phases of alveolarization a delay and catch-up were observed. Cell proliferation was increased at days 4 and 6; at day 7, thick septa with an accumulation of capillaries and cells were observed; and the number of TUNEL-positive cells (dying cells or DNA repair) was increased at day 10. Whereas at days 15 and 21 premature microvascular maturation was detected, the microvasculature was less mature at day 60 compared with wild type. No differences were observed in adulthood. We conclude that TNC contributes to the formation of new septa, to microvascular maturation, and to cell proliferation and migration during postnatal lung development.NEW & NOTEWORTHY Previously, we showed that the extracellular matrix protein tenascin-C takes part in prenatal lung development by controlling branching morphogenesis. Now we report that tenascin-C is also important during postnatal lung development, because tenascin-C deficiency delays the formation and maturation of the alveolar septa during not only classical but also continued alveolarization. Adult lungs are indistinguishable from wild type because of a catch-up formation of new septa.

Tenascin-C expression controls the maturation of articular cartilage in mice.

OBJECTIVE: Expression of the de-adhesive extracellular matrix protein tenascin-C (TNC) is associated with the early postnatal development of articular cartilage which is both load-dependent and associated with chondrocyte differentiation. We assessed morphological changes in the articular cartilage of TNC deficient mice at postnatal ages of 1, 4 and 8 weeks compared to age-matched wildtype mice. RESULTS: Cartilage integrity was assessed based on hematoxylin and eosin stained-sections from the tibial bone using a modified Mankin score. Chondrocyte density and cartilage thickness were assessed morphometrically. TNC expression was localized based on immunostaining. At 8 weeks of age, the formed tangential/transitional zone of the articular cartilage was 27% thicker and the density of chondrocytes in the articular cartilage was 55% lower in wildtype than the TNC-deficient mice. TNC protein expression was associated with chondrocytes. No relevant changes were found in mice at 1 and 4 weeks of age. The findings indicate a role of tenascin-C in the post-natal maturation of the extracellular matrix in articular cartilage. This might be a compensatory mechanism to strengthen resilience against mechanical stress.

Tenascin-C accelerates adverse ventricular remodelling after myocardial infarction by modulating macrophage polarization.

AIMS: Tenascin-C (TN-C) is an extracellular matrix protein undetected in the normal adult heart, but expressed in several heart diseases associated with inflammation. We previously reported that serum TN-C levels of myocardial infarction (MI) patients were elevated during the acute stage, and that patients with high peak TN-C levels were at high risk of left ventricular (LV) remodelling and poor outcome, suggesting that TN-C could play a significant role in the progression of ventricular remodelling. However, the detailed molecular mechanisms associated with this process remain unknown. We aimed to elucidate the role and underlying mechanisms associated with TN-C in adverse remodelling after MI. METHODS AND RESULTS: MI was induced by permanent ligation of the coronary artery of TN-C knockout (TN-C-KO) and wild type (WT) mice. In WT mice, TN-C was expressed at the borders between intact and necrotic areas, with a peak at 3 days post-MI and observed in the immediate vicinity of infiltrating macrophages. TN-C-KO mice were protected from ventricular adverse remodelling as evidenced by a higher LV ejection fraction as compared with WT mice (19.0 ± 6.3% vs. 10.6 ± 4.4%; P < 0.001) at 3 months post-MI. During the acute phase, flow-cytometric analyses showed a decrease in F4/80+CD206lowCD45+ M1 macrophages and an increase in F4/80+CD206highCD45+ M2 macrophages in the TN-C-KO heart. To clarify the role of TN-C on macrophage polarization, we examined the direct effect of TN-C on bone marrow-derived macrophages in culture, observing that TN-C promoted macrophage shifting into an M1 phenotype via Toll-like receptor 4 (TLR4). Under M2-skewing conditions, TN-C suppressed the expression of interferon regulatory factor 4, a key transcription factor that controls M2-macrophage polarization, via TLR4, thereby inhibiting M2 polarization. CONCLUSION: These results suggested that TN-C accelerates LV remodelling after MI, at least in part, by modulating M1/M2-macrophage polarization.

Deficiency of Tenascin-C Alleviates Neuronal Apoptosis and Neuroinflammation After Experimental Subarachnoid Hemorrhage in Mice.

Tenascin-C (TNC), a matricellular protein, is upregulated in brain parenchyma after experimental subarachnoid hemorrhage (SAH). Recent studies emphasize that early brain injury (EBI) should be overcome to improve post-SAH outcomes. The aim of this study was to investigate effects of TNC knockout (TNKO) on neuronal apoptosis and neuroinflammation, both of which are important constituents of EBI after SAH. C57BL/6 wild-type (WT) mice or TNKO mice underwent sham or filament perforation SAH modeling. Twenty-five WT mice and 25 TNKO mice were randomly divided into sham+WT (n = 10), sham+TNKO (n = 8), SAH+WT (n = 15), and SAH+TNKO (n = 17) groups. Beam balance test, neurological score, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining, immunostaining of Toll-like receptor 4 (TLR4), and Western blotting were performed to evaluate neurobehavioral impairments, neuronal apoptosis, and neuroinflammation at 24 h post-SAH. Deficiency of TNC significantly alleviated post-SAH neurobehavioral impairments and neuronal apoptosis. The protective effects of TNKO on neurons were associated with the inhibition of a caspase-dependent apoptotic pathway, which was at least partly mediated by TLR4/nuclear factor-κB/interleukin-1ß and interleukin-6 signaling cascades. This study first provided the direct evidence that TNC causes post-SAH neuronal apoptosis and neuroinflammation, potentially leading to the development of a new molecular targeted therapy against EBI.

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.

Effects of Tenascin-C Knockout on Cerebral Vasospasm After Experimental Subarachnoid Hemorrhage in Mice.

A matricellular protein tenascin-C (TNC) has been suggested to play a role in the pathogenesis of cerebral vasospasm after subarachnoid hemorrhage (SAH), but the direct evidence remains lacking. In this study, we examined effects of TNC knockout (TNKO) on cerebral vasospasm after experimental SAH in mice. C57BL/6 wild-type (WT) or TNKO mice were subjected to SAH by endovascular puncture. Ten WT and ten TNKO mice were randomized to WT sham (n = 4), TNKO sham (n = 4), WT SAH (n = 6), and TNKO SAH (n = 6) groups. In addition to neurobehavioral impairments and severity of SAH, cerebral vasospasm was assessed by morphometric measurements of the left internal carotid artery (ICA). Infiltration of inflammatory cells in the subarachnoid periarterial space was also assessed, and expressions of TNC and mitogen-activated protein kinases (MAPKs) in the ICA were immunohistochemically evaluated at 24 h post-surgery. TNC was induced in the smooth muscle cell layers and the adventitia in the spastic ICAs as well as the periarterial inflammatory cells in WT SAH mice. Compared with WT SAH mice, TNKO SAH mice showed better neurological scores and less severe cerebral vasospasm, as well as fewer inflammatory cell infiltration in the periarterial space. Post-SAH activation of MAPKs in the smooth muscle cell layers of the ICAs was also prevented in TNKO SAH mice. The findings in the present study suggest that TNC causes the development of cerebral vasospasm via pro-inflammatory effects and activation of MAPKs.

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.

Comprehensive DNA microarray expression profiles of tumors in tenascin-C-knockout mice.

Tenascin-C (TNC), an extracellular matrix glycoprotein, plays a pivotal role in tumor growth. However, the mechanism whereby TNC affects tumor biology remains unclear. To investigate the exact role of TNC in primary tumor growth, a mouse mammary tumor cell line, GLMT1, was first developed. Subsequently, global gene expression in GLMT1-derived tumors was compared between wild-type (WT) and TNC-knockout (TNKO) mice. Tumors in WT mice were significantly larger than those in TNKO mice. DNA microarray analysis revealed 447 up and 667 downregulated in the tumors inoculated into TNKO mice as compared to tumors in WT mice. Validation by quantitative gene expression analysis showed that Tnc, Cxcl1, Cxcl2, and Cxcr2 were significantly upregulated in WT mice. We hypothesize that TNC stimulates the CXCL1/2-CXCR2 pathway involved in cancer cell proliferation.

Suppression of hepatic dysfunction in tenascin­X­deficient mice fed a high­fat diet.

Extracellular matrix glycoprotein tenascin­X (TNX) is the largest member of the tenascin family. In the present study, the contribution of TNX to liver dysfunction was investigated by administration of high­fat and high­cholesterol diet with high levels of phosphorus and calcium (HFCD) to wild­type (WT) and TNX­knockout (KO) mice. After 16 weeks of HFCD administration, the ratio of liver weight to body weight was approximately 22% higher in the HFCD­fed WT mice compared with the HFCD­fed TNX­KO mice, indicating hepatomegaly in HFCD­fed WT mice. Histological analyses with hematoxylin and eosin staining at 21 weeks revealed that hepatocyte hypertrophy in HFCD­fed TNX­KO mice was suppressed to 85% of that in HFCD­fed WT mice. By contrast, there was a 1.2­fold increase in lipid deposition in hepatocytes from HFCD­fed TNX­KO mice compared with HFCD­fed WT mice at 18 weeks, as demonstrated by Oil Red O staining. In addition, TNX­KO mice at 21 weeks and 27 weeks post­HFCD administration exhibited significant suppression of inflammatory cell infiltrate to 51 and 24% of that in WT mice, respectively. Immunofluorescence analysis for type I collagen and Elastica van Gieson staining demonstrated a clear hepatic fibrosis progression in HFCD­fed WT mice at 27 weeks, whereas hepatic fibrosis was undetected in HFCD­fed TNX­KO mice. The present findings indicated that TNX deficiency suppressed hepatic dysfunction induced by HFCD administration.

Enriched environment alters the behavioral profile of tenascin-C deficient mice.

Tenascin-C (TnC) is an extracellular matrix glycoprotein implicated in a variety of processes ranging from brain development to synaptic plasticity in the adult vertebrates. Although the role of the TnC gene in regulation of behavior has been investigated, it remained elusive how TnC deficiency interacts with the environment in shaping the behavioral phenotype. To address this, 3-week-old TnC+/+ and TnC-/- male mice were housed over an 8-week period in standard conditions (SC), or enriched environment (EE). A comprehensive battery of tests was used in behavioral phenotyping. When housed in SC, TnC-/- mice showed spontaneous nocturnal hyperactivity, as well as poor sensorimotor coordination and low swimming velocity. However, housing of TnC-/- mice in EE abolished hyperlocomotion, led to faster habituation to novel environment, strengthened the grasp of fore limbs and partially improved movement coordination, while the swimming ability remained deficient. Conversely, TnC deficiency attenuated both the beneficial effects of EE on learning/memory capacity and the anxiolytic effect of EE in reducing the level of acrophobia. This study expands the existing knowledge about the phenotype associated with TnC deficiency, and reveals that the effect of genetic background on the behavioral response could be altered by post-weaning housing in a highly stimulating environment.

ODZ1 allows glioblastoma to sustain invasiveness through a Myc-dependent transcriptional upregulation of RhoA.

Long-term survival remains low for most patients with glioblastoma (GBM), which reveals the need for markers of disease outcome and novel therapeutic targets. We describe that ODZ1 (also known as TENM1), a type II transmembrane protein involved in fetal brain development, plays a crucial role in the invasion of GBM cells. Differentiation of glioblastoma stem-like cells drives the nuclear translocation of an intracellular fragment of ODZ1 through proteolytic cleavage by signal peptide peptidase-like 2a. The intracellular fragment of ODZ1 promotes cytoskeletal remodelling of GBM cells and invasion of the surrounding environment both in vitro and in vivo. Absence of ODZ1 by gene deletion or downregulation of ODZ1 by small interfering RNAs drastically reduces the invasive capacity of GBM cells. This activity is mediated by an ODZ1-triggered transcriptional pathway, through the E-box binding Myc protein, that promotes the expression and activation of Ras homolog family member A (RhoA) and subsequent activation of Rho-associated, coiled-coil containing protein kinase (ROCK). Overexpression of ODZ1 in GBM cells reduced survival of xenografted mice. Consistently, analysis of 122 GBM tumour samples revealed that the number of ODZ1-positive cells inversely correlated with overall and progression-free survival. Our findings establish a novel marker of invading GBM cells and consequently a potential marker of disease progression and a therapeutic target in GBM.

Tenascin-C deficiency protects mice from experimental autoimmune encephalomyelitis.

The extracellular matrix glycoprotein tenascin-C (TnC) has been increasingly appreciated as a molecule susceptibly reacting to abnormalities in the mammalian immune system. TnC expression is elevated in inflamed tissues outside the immune system, but also in lymphoid organs. It participates in the promotion of inflammatory responses. Here, the role of TnC in a paradigm of CNS autoimmunity was investigated. Experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, was induced in mice deficient in TnC (TnC-/- mice). Amelioration of EAE was observed in these mice in comparison to their wild-type (TnC+/+) littermates. Since T helper (Th)1 and Th17 cells play a dominant role in the pathogenesis of EAE, these cells were investigated in addition to analyzing locomotor functions and pro-inflammatory cytokine levels. Smaller numbers of interferon-gamma-producing Th1 cells and reduced ability of Th17 cells to produce interleukin-17 were observed in spleens of TnC-/- mice challenged by immunization with the myelin associated glycoprotein (MOG) when compared to TnC+/+ mice. There was no difference in Th1 and Th17 responses in non-immunized TnC-/- and TnC+/+ mice, thus excluding generalized immunosuppression in TnC-/- mice. These results show that TnC is important for the pathogenesis of CNS autoimmunity and that its deficiency interferes with Th1 and Th17 encephalitogenic potentials.

A method for quantification of serum tenascin-X by nano-LC/MS/MS.

BACKGROUND: Complete deficiency of an extracellular matrix tenascin-X (TNX) leads to a classical type of Ehlers-Danlos syndrome (EDS). TNX haploinsufficiency is a cause of hypermobility type of EDS. Human TNX is also present in a serum form (sTNX) with a molecular size of 140kDa. In this study, we established a method for quantification of sTNX using nano-liquid chromatography tandem mass spectrometry (LC/MS/MS) with selected/multiple reaction monitoring. METHODS: Twelve abundant protein-depleted sera were reduced, alkylated, and digested with Lys-C and trypsin. Subsequently, the digests were fractionated by strong cation exchange chromatography. Optimal and validated transitions of precursor and product ions of the peptides from sTNX were developed on a triple quadrupole mass spectrometer. RESULTS: Serum concentrations of sTNX of healthy individuals were quantified as an average of 144ng/ml. However, sTNX was not detected by this method in serum from a patient with a classical type of EDS in whom sTNX was not found by Western blot analysis. The limit of quantification (LOQ) of sTNX by nano-LC/MS/MS method was 2.8pg whereas the detection sensitivity of sTNX by Western blot analysis was 19pg. The nano-LC/MS/MS method is more sensitive than Western blot analysis. CONCLUSIONS: The quantification method will be useful for diagnosis and risk stratification of EDS caused by TNX deficiency and haploinsufficiency.

Deficiency of tenascin-C and attenuation of blood-brain barrier disruption following experimental subarachnoid hemorrhage in mice.

OBJECT Tenascin-C (TNC), a matricellular protein, is induced in the brain following subarachnoid hemorrhage (SAH). The authors investigated if TNC causes brain edema and blood-brain barrier (BBB) disruption following experimental SAH. METHODS C57BL/6 wild-type (WT) or TNC knockout (TNKO) mice were subjected to SAH by endovascular puncture. Ninety-seven mice were randomly allocated to WT sham-operated (n = 16), TNKO sham-operated (n = 16), WT SAH (n = 34), and TNKO SAH (n = 31) groups. Mice were examined by means of neuroscore and brain water content 24-48 hours post-SAH; and Evans blue dye extravasation and Western blotting of TNC, matrix metalloproteinase (MMP)-9, and zona occludens (ZO)-1 at 24 hours post-SAH. As a separate study, 16 mice were randomized to WT sham-operated, TNKO sham-operated, WT SAH, and TNKO SAH groups (n = 4 in each group), and activation of mitogen-activated protein kinases (MAPKs) was immunohistochemically evaluated at 24 hours post-SAH. Moreover, 40 TNKO mice randomly received an intracerebroventricular injection of TNC or phosphate-buffered saline, and effects of exogenous TNC on brain edema and BBB disruption following SAH were studied. RESULTS Deficiency of endogenous TNC prevented neurological impairments, brain edema formation, and BBB disruption following SAH; it was also associated with the inhibition of both MMP-9 induction and ZO-1 degradation. Endogenous TNC deficiency also inhibited post-SAH MAPK activation in brain capillary endothelial cells. Exogenous TNC treatment abolished the neuroprotective effects shown in TNKO mice with SAH. CONCLUSIONS Tenascin-C may be an important mediator in the development of brain edema and BBB disruption following SAH, mechanisms for which may involve MAPK-mediated MMP-9 induction and ZO-1 degradation. TNC could be a molecular target against which to develop new therapies for SAH-induced brain injuries.

Recurrent gastrointestinal perforation in a patient with Ehlers-Danlos syndrome due to tenascin-X deficiency.

Ehlers-Danlos syndrome (EDS) is a clinically and genetically heterogeneous disorder. Using a customized targeted exome-sequencing system we identified nonsense mutations in TNXB in a patient who had recurrent gastrointestinal perforation due to tissue fragility. This case highlights the utility of targeted exome sequencing for the diagnosis of congenital diseases showing genetic heterogeneity, and the importance of attention to gastrointestinal perforation in patients with tenascin-X deficient type EDS.

Tenascin-C aggravates autoimmune myocarditis via dendritic cell activation and Th17 cell differentiation.

BACKGROUND: Tenascin-C (TN-C), an extracellular matrix glycoprotein, appears at several important steps of cardiac development in the embryo, but is sparse in the normal adult heart. TN-C re-expresses under pathological conditions including myocarditis, and is closely associated with tissue injury and inflammation in both experimental and clinical settings. However, the pathophysiological role of TN-C in the development of myocarditis is not clear. We examined how TN-C affects the initiation of experimental autoimmune myocarditis, immunologically. METHODS AND RESULTS: A model of experimental autoimmune myocarditis was established in BALB/c mice by immunization with murine α-myosin heavy chains. We found that TN-C knockout mice were protected from severe myocarditis compared to wild-type mice. TN-C induced synthesis of proinflammatory cytokines, including interleukin (IL)-6, in dendritic cells via activation of a Toll-like receptor 4, which led to T-helper (Th)17 cell differentiation and exacerbated the myocardial inflammation. In the transfer experiment, dendritic cells loaded with cardiac myosin peptide acquired the functional capacity to induce myocarditis when stimulated with TN-C; however, TN-C-stimulated dendritic cells generated from Toll-like receptor 4 knockout mice did not induce myocarditis in recipients. CONCLUSIONS: Our results demonstrated that TN-C aggravates autoimmune myocarditis by driving the dendritic cell activation and Th17 differentiation via Toll-like receptor 4. The blockade of Toll-like receptor 4-mediated signaling to inhibit the proinflammatory effects of TN-C could be a promising therapeutic strategy against autoimmune myocarditis.