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Disseminated cancer cells frequently lodge near vasculature in secondary organs. However, our understanding of the cellular crosstalk invoked at perivascular sites is still rudimentary. Here, we identify intercellular machinery governing formation of a pro-metastatic vascular niche during breast cancer colonization in the lung. We show that specific secreted factors, induced in metastasis-associated endothelial cells (ECs), promote metastasis in mice by enhancing stem cell properties and the viability of cancer cells. Perivascular macrophages, activated via tenascin C (TNC) stimulation of Toll-like receptor 4 (TLR4), were shown to be crucial in niche activation by secreting nitric oxide (NO) and tumor necrosis factor (TNF) to induce EC-mediated production of niche components. Notably, this mechanism was independent of vascular endothelial growth factor (VEGF), a key regulator of EC behavior and angiogenesis. However, targeting both macrophage-mediated vascular niche activation and VEGF-regulated angiogenesis resulted in added potency to curb lung metastasis in mice. Together, our findings provide mechanistic insights into the formation of vascular niches in metastasis.

Metastatic prostate cancer (PCa) in bone induces bone-forming lesions that enhance PCa progression. How tumor-induced bone formation enhances PCa progression is not known. We have previously shown that PCa-induced bone originates from endothelial cells (ECs) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition by tumor-secreted bone morphogenetic protein 4 (BMP4). Here, we show that EC-to-OSB transition leads to changes in the tumor microenvironment that increases the metastatic potential of PCa cells. We found that conditioned medium (CM) from EC-OSB hybrid cells increases the migration, invasion, and survival of PC3-mm2 and C4-2B4 PCa cells. Quantitative mass spectrometry (Isobaric Tags for Relative and Absolute Quantitation) identified Tenascin C (TNC) as one of the major proteins secreted from EC-OSB hybrid cells. TNC expression in tumor-induced OSBs was confirmed by immunohistochemistry of MDA PCa-118b xenograft and human bone metastasis specimens. Mechanistically, BMP4 increases TNC expression in EC-OSB cells through the Smad1-Notch/Hey1 pathway. How TNC promotes PCa metastasis was next interrogated by in vitro and in vivo studies. In vitro studies showed that a TNC-neutralizing antibody inhibits EC-OSB-CM-mediated PCa cell migration and survival. TNC knockdown decreased, while the addition of recombinant TNC or TNC overexpression increased migration and anchorage-independent growth of PC3 or C4-2b cells. When injected orthotopically, PC3-mm2-shTNC clones decreased metastasis to bone, while C4-2b-TNC-overexpressing cells increased metastasis to lymph nodes. TNC enhances PCa cell migration through α5β1 integrin-mediated YAP/TAZ inhibition. These studies elucidate that tumor-induced stromal reprogramming generates TNC that enhances PCa metastasis and suggest that TNC may be a target for PCa therapy.

Background Markers of stromal activation at future metastatic sites may have prognostic value and may allow clinicians to identify and abolish the pre-metastatic niche to prevent metastasis. In this study, we evaluate tenascin-C as a marker of pre-metastatic niche formation in bladder cancer patient lymph nodes. Methods Tenascin-C expression in benign lymph nodes was compared between metastatic (n = 20) and non-metastatic (n = 27) patients with muscle-invasive bladder cancer. Urinary extracellular vesicle (EV) cytokine levels were measured with an antibody array to examine potential correlation with lymph node inflammation. The ability of bladder cancer EVs to activate primary bladder fibroblasts was assessed in vitro. Results Lymph node tenascin-C expression was elevated in metastatic patients vs. non-metastatic patients, and high expression was associated with worse survival. Urinary EVs contained four cytokines that were positively correlated with lymph node tenascin-C expression. Bladder cancer EVs induced tenascin-C expression in fibroblasts in an NF-κB-dependent manner. Conclusions Tenascin-C expression in regional lymph nodes may be a good predictor of bladder cancer metastasis and an appropriate imaging target. It may be possible to interrupt pre-metastatic niche formation by targeting EV-borne tumour cytokines or by targeting tenascin-C directly.

Background Mesenchymal stem cells (MSCs) secrete paracrine factors and extracellular matrix proteins that contribute to their ability to support tissue healing and regeneration. Both the transcriptome and the secretome of MSCs can be altered by treating the cells with cytokines, but neither have been thoroughly investigated following treatment with the specific cytokine transforming growth factor (TGF)-β2. Methods RNA-sequencing and western blotting were used to compare gene and protein expression between untreated and TGF-β2-treated equine bone marrow-derived MSCs (BM-MSCs). A co-culture system was utilized to compare equine tenocyte migration during co-culture with untreated and TGF-β2-treated BM-MSCs. Results TGF-β2 treatment significantly upregulated gene expression of collagens, extracellular matrix molecules, and growth factors. Protein expression of collagen type I and tenascin-C was also confirmed to be upregulated in TGF-β2-treated BM-MSCs compared to untreated BM-MSCs. Both untreated and TGF-β2-treated BM-MSCs increased tenocyte migration in vitro. Conclusions Treating equine BM-MSCs with TGF-β2 significantly increases production of paracrine factors and extracellular matrix molecules important for tendon healing and promotes the migration of tenocytes in vitro.

This study was conducted to investigate the role of Tenascin-C (TNC) in paraquat (PQ)-induced lung injury in vivo and in vitro and explore its related mechanism during this process. Six- to eight-week-old male C57BL/6 mice were injected with 30 mg/kg PQ by intraperitoneal injection and sacrificed on 2 days, 7 days, 14 days, and 28 days after PQ administration. In vivo, we detected the expression of TNC at all time points of lung tissues in mice by reverse transcription-quantitative-polymerase chain reaction, western blotting, and immunohistochemistry. Expression of TLR4, NF-κB p65, TGF-β1, and α-SMA in lung tissues have also been tested. In vitro, siRNA was used to knock down TNC expression in A549 cells and TLR4, NF-κB p65, and TGF-β1 expressions were examined after PQ exposure. TNC expression increased in both lung tissues of mice model and A549 cells after PQ administration. In vivo, TNC mostly located at the extracellular matrix of thickened alveolar septum, especially at sites of injury, together with the increasing of TLR4, NF-κB p65, TGF-β1, and α-SMA. In vitro, PQ exposure also increased the expressions of TLR4, NF-κB p65, and TGF-β1 in A549 cells, but knocking down TNC gene expression obviously down-regulated the expressions of TLR4, NF-κB p65, NF-κB Pp65, and TGF-β1. The results of this study demonstrate, for the first time, that TNC participates in the development of lung injury induced by PQ poisoning. The role of TNC in this process is closely related to TLR4 and TGF-β signaling pathways.

Neural stem/progenitor cells (NSPCs) rely on internal and external cues determining their lineage decisions during brain development. The progenitor cells of the embryonic mammalian forebrain reside in the ventricular and subventricular zones of the lateral ventricles, where they proliferate, generate neurons and glial cells, and respond to external cues like growth factors. The extracellular matrix (ECM) surrounds NSPCs and influences the cell fate by providing mechanical scaffold, trophic support, and instructive signals. The ECM molecule tenascin-C (Tnc) is expressed in the proliferative zones of the developing forebrain and involved in the proliferation and maturation of NSPCs. Here, we analyzed the regulation of the Tnc gene expression by NSPCs cultivated under the influence of different growth factors. We observed that the epidermal growth factor (EGF) and the fibroblast growth factor (FGF)-2 strongly increased the expression of Tnc, whereas the transforming growth factor (TGF)β 1 had no effect on Tnc gene expression, in contrast to previous findings in cell cultures of neural and non-neural origin. The stimulation of the Tnc gene expression induced by EGF or FGF-2 was reversible and seen in constantly treated as well as short term stimulated NSPC cultures. The activation depended on the presence of the respective receptors, which was slightly different in cortical and striatal NSPC cultures. Our results confirm the influence of extracellular stimuli regulating the expression of factors that form a niche for NSPCs during embryonic forebrain development.

The literature has shown the beneficial effects of microcurrent (MC) therapy on tissue repair. We investigated if the application of MC at 10 μA/90 s could modulate the expression of remodeling genes transforming growth factor beta (Tgfb), connective tissue growth factor (Ctgf), insulin-like growth factor 1 (Igf1), tenascin C (Tnc), Fibronectin (Fn1), Scleraxis (Scx), Fibromodulin (Fmod) and tenomodulin in NIH/3T3 fibroblasts in a wound healing assay. The cell migration was analyzed between days 0 and 4 in both fibroblasts (F) and fibroblasts + MC (F+MC) groups. On the 4th day, cell viability and gene expression were also analyzed after daily MC application. Higher expression of Ctgf and lower expression of Tnc and Fmod, respectively, were observed in the F+MC group in relation to F group (p

Vernal keratoconjunctivitis (VKC) is a severe and recurrent allergic conjunctivitis, the mechanism of which is not well understood. In this study, we investigated the role of oncostatin M (OSM) in the pathogenesis of VKC, with a focus on tissue remodeling. Clinical and experimental. The OSM concentrations in tear fluid samples obtained from VKC patients and healthy controls were measured using ELISA, and the expression of OSM mRNA and protein in giant papillae resected from VKC patients was investigated using RT-PCR and immunohistochemistry, respectively. In cultured human conjunctival epithelial cells (HconEpiCs), expression of OSM receptor β (OSMRβ) was detected using immunocytochemical and FACS analyses. Finally, we investigated whether recombinant OSM activated STAT1 and STAT3 to induce the expression of various genes related to tissue remodeling in HconEpiCs, by using Western blot analysis, microarray analysis, and RT-PCR. The OSM concentration was higher in the tear fluid of VKC patients than in that of the healthy controls, and strong expression of OSM mRNA was found in the giant papillae. We also detected T cells expressing OSM in the giant papillae. In addition, HconEpiCs showed surface expression of OSMRβ. Recombinant human OSM strongly activated both STAT1 and STAT3 in HconEpiCs and induced various tissue remodeling-related genes, including MMP-1, MMP-3, IL-24, IL-20, serpinB3, S100A7, tenascin C, and SOCS3. Our results suggest that OSM is one of the key molecules involved in remodeling of giant papillae in VKC.

Necroptosis, a form of programmed cell death, is characterized by the loss of membrane integrity and release of intracellular contents, the execution of which depends on the membrane-disrupting activity of the Mixed Lineage Kinase Domain-Like protein (MLKL) upon its phosphorylation. Here we found myofibers committed MLKL-dependent necroptosis after muscle injury. Either pharmacological inhibition of the necroptosis upstream kinase Receptor Interacting Protein Kinases 1 (RIPK1) or genetic ablation of MLKL expression in myofibers led to significant muscle regeneration defects. By releasing factors into the muscle stem cell (MuSC) microenvironment, necroptotic myofibers facilitated muscle regeneration. Tenascin-C (TNC), released by necroptotic myofibers, was found to be critical for MuSC proliferation. The temporary expression of TNC in myofibers is tightly controlled by necroptosis; the extracellular release of TNC depends on necroptotic membrane rupture. TNC directly activated EGF receptor (EGFR) signaling pathway in MuSCs through its N-terminus assembly domain together with the EGF-like domain. These findings indicate that necroptosis plays a key role in promoting MuSC proliferation to facilitate muscle regeneration.

Squamous cell carcinoma (SCC) of the head and neck region is the sixth most frequent malignancy with high mortality rate. Due to its poor prognosis it is considered a growing public health problem worldwide inspite of existing treatment modalities. Thus, early diagnosis of new diseases and recurrences is emerging on one hand, but on the other hand troublesome in the lack of reliable tumor markers in this field. The rapid development of proteomics has opened new perspectives in tumor marker discovery. Liquid chromatography/mass spectrometry (LC/MS) as the gold standard in proteomics enables the semi-quantitative analysis of proteins within various tissues. Abundance differences between tumor and normal tissue also can be interpreted as tumor specific changes. The aim of this study was to identify potential tumor markers of laryngeal/hypopharyngeal SCC by revealing abundance changes between cancerous and the surrounding phenotypically healthy tissue. After separating the phenotypically cancerous and healthy parts of formalin-fixed paraffin-embedded tissues, each sample underwent protein recovery process and tryptic digestion for label-free semi-quantitative LC/MS analysis. Eight proteins showed significantly higher abundance in tumor including tenascin, transmembrane emp24 domain-containing protein 2, cytoplasmic dynein light chain 1, coactosin-like protein, small proline-rich protein 2D, nucleolin, U5 small nuclear RNP 200-kDa helicase and fatty aldehyde dehydrogenase. Desmoglein-1 and keratin type I cytoskeletal 9 were down-regulated in tumor. Using Ingenuity Pathway Analysis we mapped the signaling pathways these proteins play role in regarding other tumors. Based on these findings these proteins may serve as promising biomarkers in the fight against laryngeal/hypopharyngeal SCCs.

Fibro-inflammatory processes in the extracellular matrix are closely associated with progressive structural remodeling in atrial fibrillation (AF). Serum concentrations of tenascin-C (TNC), an extracellular matrix glycoprotein, and of high-sensitivity C-reactive protein (CRP) might serve as a marker of remodeling and progressive inflammation of the aorta and in myocardial diseases. This study aimed to clarify relationships between TNC and CRP in patients with AF. This study included 38 patients with AF and five controls without left ventricular dysfunction who underwent catheter ablation. Blood was collected immediately before ablation from the left atrium (LA), right atrium (RA), and femoral artery (FA), and left and right atrial pressure was measured. Levels of TNC in the LA (TNC-LA), RA (TNC-RA), and FA (TNC-FA) and high-sensitivity C-reactive protein (CRP) were measured. Atrial size was also determined by echocardiography. Levels of TNC corrected by atrial size were maximal in the LA, followed by the RA (3.69 ± 0.32 and 2.87 ± 0.38 ng/mL/cm, respectively). Mean transverse diameter corrected by body surface area was larger and mean atrial pressure was greater in the LA than the RA. A relationship was found between CRP from the femoral vein and TNC-LA and TNC-RA, but not TNC-FA. None of TNC-LA, TNC-RA, or TNC-FA correlated with ANP or BNP in the femoral vein. Intracardiac (atrial) TNC expression plays an important role in the development of remodeling processes in the atrium with AF. Tenascin-C from the LA and RA (but not TNC, ANP, and BNP from FA) might serve as novel markers of these processes.

Gastric cancer is one of the most common malignancies worldwide and vasculogenic mimicry (VM) is considered to be the leading cause for the failure of anti-angiogenesis therapy in advanced gastric cancer patients. In the present study, we investigate the role of tenascin-c (TNC) in the formation of VM in gastric cancer and found that TNC was upregulated in gastric cancer tissue than in the corresponding adjacent tissues and correlated with VM and poor prognosis of gastric cancer. Furthermore, knockdown of TNC significantly inhibited VM formation and proliferation of gastric cancer cells in vitro and in vivo, with a reduction in cell migration and invasion. Mechanistically, TNC knockdown suppressed the phosphorylation of ERK and subsequently inhibited the process of EMT, both of which play an important role in VM formation. Our results indicated that TNC plays an important role in VM formation in gastric cancer. Combining inhibition of TNC and ERK may be a potential therapeutic approach to inhibit gastric cancer growth and metastasis and decrease antiangiogenic therapeutic resistance.

Background Lymphovascular space invasion is an independent prognostic factor in early-stage cervical cancer. However, there is a lack of non-invasive methods to detect lymphovascular space invasion. Some researchers found that Tenascin-C and Cyclooxygenase-2 was correlated with lymphovascular space invasion. Radiomics has been studied as an emerging tool for distinguishing tumor pathology stage, evaluating treatment response, and predicting prognosis. This study aimed to establish a machine learning model that combines radiomics based on PET imaging with tenascin-C (TNC) and cyclooxygenase-2 (COX-2) for predicting lymphovascular space invasion (LVSI) in patients with early-stage cervical cancer. Methods One hundred and twelve patients with early-stage cervical squamous cell carcinoma who underwent PET/CT examination were retrospectively analyzed. Four hundred one radiomics features based on PET/CT images were extracted and integrated into radiomics score (Rad-score). Immunohistochemical analysis was performed to evaluate TNC and COX-2 expression. Mann-Whitney U test was used to distinguish differences in the Rad-score, TNC, and COX-2 between LVSI and non-LVSI groups. The correlations of characteristics were tested by Spearman analysis. Machine learning models including radiomics model, protein model and combined model were established by logistic regression algorithm and evaluated by ROC curve. Pairwise comparisons of ROC curves were tested by DeLong test. Results The Rad-score of patients with LVSI was significantly higher than those without. A significant correlation was shown between LVSI and Rad-score (r = 0.631, p r = 0.244, p = 0.024) and COX-2 (r = 0.227, p = 0.036). The radiomics model had the best predictive performance among all models in training and external dataset (AUCs: 0.914, 0.806, respectively, p Conclusion The machine learning model of the combination of PET radiomics with COX-2 and TNC provides a new tool for detecting LVSI in patients with early-stage cervical cancer. In the future, multicentric studies on larger sample of patients will be used to test the model. Trial registration This is a retrospective study and there is no experimental intervention on human participants. The Ethics Committee has confirmed that retrospectively registered is not required.

BackgroundThe extracellular matrix has been critically associated with the tumorigenesis and progression of Ewing sarcoma (ES). However, the regulatory and prognostic roles of tenascin-C (TNC) in ES remain unclear.MethodsTNC expression was examined in specimens by immunohistochemistry, and the association of TNC expression with ES patient survival was also analysed. TNC-knockout cell lines were constructed using CRISPR/Cas9 methods. In vitro experiments and in vivo bioluminescent imaging using BALB/c nude mice were conducted to evaluate the effect of TNC on ES tumour progression. RNA sequencing was performed, and the underlying mechanism of TNC was further explored.ResultsTNC was overexpressed in ES tissue and cell lines, and TNC overexpression was associated with poor survival in ES patients. TNC enhanced cell proliferation, migration and angiogenesis in vitro and promoted ES metastasis in vivo. The oncoprotein EWS-FLI1 profoundly increased TNC expression by directly binding to the TNC promoter region. Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) upregulation induced by Yes-associated protein (YAP) activation was responsible for TNC-regulated ES tumour progression. Activated integrin α5β1 signalling might be correlated with YAP dephosphorylation and nuclear translocation.ConclusionsTNC may promote ES tumour progression by targeting MALAT1 through integrin α5β1-mediated YAP activation.

Pancreatic cancer tissue contains abundant stromal components, including extracellular matrix proteins such as tenascin C (TNC), which exists as large (TNC-L) and non-large splice variants. Here, we examined human pancreatic cancer specimens for the expression of total TNC (TNC-ALL) and TNC-L in the stroma and annexin A2 (ANXA2), a cell surface receptor for TNC, and evaluated their significance as prognostic markers for pancreatic cancer. Expression of ANXA2, TNC-ALL, and TNC-L was examined in 106 pancreatic cancer tissues from patients who underwent curative resection and who had not received prior therapy or surgery. Protein expression was measured by immunohistochemistry and scored on a semi-quantitative scale. The relationships between protein expression, clinicopathological factors, and prognosis were evaluated by Cox proportional hazards analysis. TNC-ALL and TNC-L were detected mainly in the stroma, whereas ANXA2 was predominantly expressed in cancer cell membranes. TNC-ALL was also expressed in non-tumor pancreatic tissue. High levels of stromal TNC-L and membranous ANXA2, but not stromal TNC-ALL, were independently associated with cancer progression and poor prognosis. Moreover, high co-expression of stromal TNC-L and membranous ANXA2 was a superior indicator of poor prognosis compared with detection of TNC-ALL, TNC-L, or ANXA2 alone. Our data suggest that co-expression of stromal TNC-L and membranous ANXA2 is a poor prognostic marker compared with detection of TNC-L or ANXA2 alone for pancreatic cancer patients. Additionally, targeting of crosstalk between stromal TNC and cancer cell ANXA2 could be a promising therapeutic strategy to overcome refractory pancreatic cancer.

A central and unique aspect of high-grade serous ovarian carcinoma (HGSC) is the extensive transcoelomic spreading of tumor cell via the peritoneal fluid or malignant ascites. We and others identified tumor-associated macrophages (TAM) in the ascites as promoters of metastasis-associated processes like extracellular matrix (ECM) remodeling, tumor cell migration, adhesion, and invasion. The precise mechanisms and mediators involved in these functions of TAM are, however, largely unknown. We observed that HGSC migration is promoted by soluble mediators from ascites-derived TAM, which can be emulated by conditioned medium from monocyte-derived macrophages (MDM) differentiated in ascites to TAM-like asc-MDM. A similar effect was observed with IL-10-induced alternatively activated m2c-MDM but not with LPS/IFNγ-induced inflammatory m1-MDM. These observations provided the basis for deconvolution of the complex TAM secretome by performing comparative secretome analysis of matched triplets of different MDM phenotypes with different pro-migratory properties (asc-MDM, m2c-MDM, m1-MDM). Mass spectrometric analysis identified an overlapping set of nine proteins secreted by both asc-MDM and m2c-MDM, but not by m1-MDM. Of these, three proteins, i.e., transforming growth factor beta-induced (TGFBI) protein, tenascin C (TNC), and fibronectin (FN1), have been associated with migration-related functions. Intriguingly, increased ascites concentrations of TGFBI, TNC, and fibronectin were associated with short progression-free survival. Furthermore, transcriptome and secretome analyses point to TAM as major producers of these proteins, further supporting an essential role for TAM in promoting HGSC progression. Consistent with this hypothesis, we were able to demonstrate that the migration-inducing potential of asc-MDM and m2c-MDM secretomes is inhibited, at least partially, by neutralizing antibodies against TGFBI and TNC or siRNA-mediated silencing of TGFBI expression. In conclusion, the present study provides the first experimental evidence that TAM-derived TGFBI and TNC in ascites promote HGSC progression.

Background In order to optimize the tenogenic differentiation of mesenchymal stem cells (MSCs), researchers should consider various factors. However, this requires testing numerous experimental settings, which is costly and time-consuming. We aimed to assess the differential effects of transforming growth factor beta-3 (TGF-β3) on the tenogenesis of tonsil-derived MSCs (T-MSCs) and bone marrow-derived MSCs (BM-MSCs) using response surface methodology (RSM). Methods Bone marrow and tonsillar tissue were collected from four patients; mononuclear cells were separated and treated with 5 or 10 ng/mL of TGF-β3. A full factorial experimental design with a categorical factor of 0 was employed to study the effect of tension based on T-MSCs. Eighty-four trials were fitted with RSM and then used to obtain mathematical prediction models. Results Exposure of T-MSCs and BM-MSCs to TGF-β3 increased the expression of scleraxis (SCX), tenomodulin (TNMD), decorin, collagen I, and tenascin C. Expression of most of these factors reached a maximum after 2–3 days of treatment. The model predicted that the values of the tenocyte lineage-related factors assessed would be significantly increased at 2.5 days of culture with 2.7 ng/mL of TGF-β3 for T-MSCs and at 2.3 days of culture regardless of TGF-β3 concentration for BM-MSCs. Conclusions This study demonstrated that the RSM prediction of the culture time necessary for the tenogenic differentiation of T-MSCs and BM-MSCs under TGF-β3 stimulation was similar to the experimentally determined time of peak expression of tenocyte-related mRNAs, suggesting the potential of using the RSM approach for optimization of the culture protocol for tenogenesis of MSCs.

ObjectiveExpression 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.ResultsCartilage 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.

Objective: Maternal plasma is a source of circulating placental nucleic acids. In this study, we validated previous observations on abnormal levels of circulating messenger RNA (mRNA) for the tenascin-X gene in pregnancies with ventricular septal defects in the second trimester of pregnancy. Methods: This was a bicentric retrospective study conducted from March 2016 to July 2017. Real-time polymerase chain reaction was used to identify abnormally expressed genes, comparing ten women carrying a euploid fetus with ventricular septal defects to 30 controls at 19–24 weeks of gestation. The univariable analysis was used to determine whether the mean mRNA for the tenascin-X gene values would differ from the expected values for the controls. Results: mRNA for tenascin-X gene values was higher in ventricular septal defects, 4.38 ± 3.01 versus 1.00 ± 0.80. The result was still significant even after adjustment for gestational age. Conclusions: These data confirm previous studies on the specific association of mRNA species and type of congenital heart defect and confirm that ventricular septal defects are associated with abnormal mRNA for the tenascin-X gene. The positive predictive value of this molecular marker in the general population should be assessed through prospective studies.

Obesity is related to a dynamic extracellular matrix (ECM) remodeling, which involves the synthesis and degradation of different proteins, such as tenascin C (TNC) in the adipose tissue (AT). Given the functional relationship between leptin and inducible nitric oxide synthase (iNOS), our aim was to analyze the impact of the absence of the iNOS gene in AT inflammation and ECM remodeling in ob/ob mice. The expression of genes involved in inflammation and ECM remodeling was evaluated in 10-week-old male double knockout (DBKO) mice simultaneously lacking the ob and iNOS genes as well as in ob/ob mice classified into three groups [control, leptin-treated (1 mg kg−1 day−1) and pair-fed]. Leptin deficiency increased inflammation and fibrosis in AT. As expected, leptin treatment improved the obesity phenotype. iNOS deficiency in ob/ob mice improved insulin sensitivity, AT inflammation, and ECM remodeling, as evidenced by lower AT macrophage infiltration and collagen deposition, a downregulation of proinflammatory and profibrogenic genes Tnf, Emr1, Hif1a, Col6a1, Col6a3, and Tnc, as well as lower circulating TNC levels. Interestingly, leptin upregulated TNC expression and release in 3T3-L1 adipocytes, and iNOS knockdown in 3T3-L1 fat cells produced a significant decrease in basal and leptin-induced Tnc expression. Ablation of iNOS in leptin-deficient mice improved AT inflammation and ECM remodeling-related genes, attenuating fibrosis, and metabolic dysfunction. The activation of iNOS by leptin is necessary for the synthesis and secretion of TNC in adipocytes, suggesting an important role of this alarmin in the development of AT inflammation and fibrosis.

Monocytes of patients with ankylosing spondylitis (AS) over-express toll-like receptor (TLR) 4. Tenascin-C (TNC) is an endogenous TLR4 ligand. Thus, we studied the serum and synovial fluid levels of TNC in AS. TNC was measured in serum of 36 AS patients (ASAS 2010 criteria) and 39 healthy controls by ELISA. Twenty-two patients were followed up after 3 months of standard treatment. Five paired serum-synovial fluid samples were also analyzed. Disease activity was assessed by BASDAI, ASDAS, swollen joint count, ESR, and CRP. All values are in median (IQR). Median age was 30 (20–35) years, and disease duration was 5.5 (1.3–10) years. Thirty-one were male. Twenty-five (69.5%) had peripheral arthritis. Median BASDAI was 5.3 (3.3–6.7). HLA B27 was positive in 34 (94.5%) cases. Median serum tenascin C levels were higher in AS [578.5 ng/ml] as compared to healthy controls [32.88 ng/ml, p < 0.0001]. Serum tenascin C levels correlated with ASDAS ESR [r = 0.367, p = 0.028] and ESR [r = 0.39, p = 0.035]. In patients with early disease (duration ≤ 5 years), serum levels had better correlation with ESR [r = 0.59, p = 0.009] and CRP [r = 0.479, p = 0.044]. On ROC analysis for active (PhGA ≥ 6) vs. inactive (PhGA ≤ 4) disease, tenascin-C (AUC = 0.60) performed as well as CRP (AUC = 0.65) and ESR (AUC = 0.73). Synovial fluid levels [11.61 (5.99–176.9) ng/ml] were lower than in serum [627.4 (488.5–779.1) ng/ml, p = 0.008]. Tenascin C fell levels with treatment [n = 11, 630.8 ng/ml to 376.4 ng/ml p = 0.0006] in treatment responders but not in non-responders [n = 11, 562.3 to 445.6, p = 0.33]. Serum TNC levels are raised in AS and may serve as marker of inflammation in early disease.

Background: Tumour stroma has important roles in the development of colorectal cancer (CRC) metastasis. We aimed to clarify the roles of microRNAs (miRNAs) and their target genes in CRC stroma in the development of liver metastasis. Methods: Tumour stroma was isolated from formalin-fixed, paraffin-embedded tissues of primary CRCs with or without liver metastasis by laser capture microdissection, and miRNA expression was analysed using TaqMan miRNA arrays. Results: Hierarchical clustering classified 16 CRCs into two groups according to the existence of synchronous liver metastasis. Combinatory target prediction identified tenascin C as a predicted target of miR-198, one of the top 10 miRNAs downregulated in tumour stroma of CRCs with synchronous liver metastasis. Immunohistochemical analysis of tenascin C in 139 primary CRCs revealed that a high staining intensity was correlated with synchronous liver metastasis (P

Background:Oral tongue squamous cell carcinoma (OTSCC) metastasises early, especially to regional lymph nodes. There is an ongoing debate on which early stage (T1-T2N0) patients should be treated with elective neck dissection. We need prognosticators for early stage tongue cancer. Methods: Mice immunisation with human mesenchymal stromal cells resulted in production of antibodies against tenascin-C (TNC) and fibronectin (FN), which were used to stain 178 (98 early stage), oral tongue squamous cell carcinoma samples. TenascinC and FN expression in the stroma (negative, moderate or abundant) and tumour cells (negative or positive) were assessed. Similar staining was obtained using corresponding commercial antibodies. Results: Expression of TNC and FN in the stroma, but not in the tumour cells, proved to be excellent prognosticators both in all stages and in early stage cases. Among early stages, when stromal TNC was negative, the 5-year survival rate was 88%. Correspondingly, when FN was negative, no cancer deaths were observed. Five-year survival rates for abundant expression of TNC and FN were 43% and 25%, respectively. Conclusions: Stromal TNC and, especially, FN expressions differentiate patients into low-and high-risk groups. Surgery alone of early stage primary tumours might be adequate when stromal FN is negative. Aggressive treatments should be considered when both TNC and FN are abundant.

To study ROCK2 activation in carcinogenesis, mice expressing 4-hydroxytamoxifen (4HT)-activated ROCK2 (K14.ROCKer) were crossed with mice expressing epidermal-activated rasHa (HK1.ras1205). At 8 weeks, 4HT-treated K14.ROCKer/HK1.ras1205 cohorts exhibited papillomas similar to HK1.ras1205 controls; however, K14.ROCKer/HK1.ras1205 histotypes comprised a mixed papilloma/well-differentiated squamous cell carcinoma (wdSCC), exhibiting p53 loss, increased proliferation and novel NF-κB expression. By 12 weeks, K14.ROCKer/HK1.ras1205 wdSCCs exhibited increased NF-κB and novel tenascin C, indicative of elevated rigidity; yet despite continued ROCK2 activities/p-Mypt1 inactivation, progression to SCC required loss of compensatory p21 expression. K14.ROCKer/HK1.ras1205 papillomatogenesis also required a wound promotion stimulus, confirmed by breeding K14.ROCKer into promotion-insensitive HK1.ras1276 mice, suggesting a permissive K14.ROCKer/HK1.ras1205 papilloma context (wound-promoted/NF-κB+/p53-/p21+) preceded K14.ROCKer-mediated (p-Mypt1/tenascin C/rigidity) malignant conversion. Malignancy depended on ROCKer/p-Mypt1 expression, as cessation of 4HT treatment induced disorganized tissue architecture and p21-associated differentiation in wdSCCs; yet tenascin C retention in connective tissue extracellular matrix suggests the rigidity laid down for conversion persists. Novel papilloma outgrowths appeared expressing intense, basal layer p21 that confined endogenous ROCK2/p-Mypt1/NF-κB to supra-basal layers, and was paralleled by restored basal layer p53. In later SCCs, 4HT cessation became irrelevant as endogenous ROCK2 expression increased, driving progression via p21 loss, elevated NF-κB expression and tenascin C-associated rigidity, with p-Mypt1 inactivation/actinomyosin-mediated contractility to facilitate invasion. However, p21-associated inhibition of early-stage malignant progression and the intense expression in papilloma outgrowths, identifies a novel, significant antagonism between p21 and rasHa/ROCK2/NF-κB signalling in skin carcinogenesis. Collectively, these data show that ROCK2 activation induces malignancy in rasHa-initiated/promoted papillomas in the context of p53 loss and novel NF-κB expression, whereas increased tissue rigidity and cell motility/contractility help mediate tumour progression.

Diffuse intrinsic pontine glioma (DIPG), an infiltrative, high grade glioma (HGG) affecting young children, has the highest mortality rate of all pediatric cancers. Despite treatment, average survival is less than twelve months, and five-year survival under 5%. We previously detected increased expression of Tenascin-C (TNC) protein in DIPG cerebrospinal fluid and tumor tissue relative to normal specimens. TNC is an extracellular matrix (ECM) glycoprotein that mediates cell-matrix interactions, guides migrating neurons during normal brain development and is thought to maintain the periventricular stem cell niche in the developing brain. Tumor TNC expression is reported in adult glioma and other cancers. However, the pattern and effects of TNC expression in DIPG has not been previously explored. Here, we characterize TNC expression in patient derived pediatric supratentorial HGG (n = 3) and DIPG (n = 6) cell lines, as well as pediatric glioma tumor (n = 50) and normal brain tissue specimens (n = 3). We found tumor specific TNC gene and protein overexpression that directly correlated with higher tumor grade (WHO III and IV, p = 0.05), H3K27 M mutation (p = 0.012), shorter progression free survival (p = 0.034), and poorer overall survival (0.041) in association with these factors. TNC knockdown via lentiviral shRNA transfection of HGG (n = 1) and DIPG (n = 3) cell lines resulted in decreased cell proliferation, migration, and invasion in vitro (p

Knowledge of the microenvironment of articular cartilage in health and disease is the key to accomplishing fundamental disease-modifying treatments for osteoarthritis. The proteins comprising the CCN Family are matricellular proteins with a remarkable relevance within the context of cartilage metabolism. CCN2 displays a great capability for regenerating articular cartilage, and CCN3 has been shown to activate the expression of genes related to articular chondrocytes and to repress genes related to endochondral ossification in epiphyseal chondrocytes. Moreover, mice lacking CCN3 protein have been shown to display ostearthritic changes in their knee articular cartilage. In this study, we employed a monoiodoacetic acid (MIA)-induced osteoarthritic model to investigate whether osteoarthritic changes in the cartilage are reciprocally accompanied by CCN3 down-regulation and an inducible overexpression system to evaluate the effects of CCN3 on articular chondrocytes in vitro. Finally, we also investigated the effects of exogenous CCN3 in vivo during the early stages of MIA-induced osteoarthritis. We discovered that CCN3 is expressed by articular chondrocytes in normal rat knees, whereas it is rapidly down-regulated in osteoarthritic knees. In vitro, we also discovered that CCN3 increases the proteoglycan accumulation, the gene expression of type II collagen, tenascin-C and lubricin, as well as the protein production of tenascin-C and lubricin in articular chondrocytes. In vivo, it was discovered that exogenous CCN3 increased tidemark integrity and produced an increased production of lubricin protein. The potential utility of CCN3 as a future therapeutic agent and possible strategies to improve its therapeutic functions are also discussed.

Increased overall survival for patients with glioma brain tumours is associated with mutations in the metabolic regulator isocitrate dehydrogenase 1 (IDH1). Gliomas develop within a mechanically challenged microenvironment that is characterized by a dense extracellular matrix (ECM) that compromises vascular integrity to induce hypoxia and activate HIF1α. We found that glioma aggression and patient prognosis correlate with HIF1α levels and the stiffness of a tenascin C (TNC)-enriched ECM. Gain- and loss-of-function xenograft manipulations demonstrated that a mutant IDH1 restricts glioma aggression by reducing HIF1α-dependent TNC expression to decrease ECM stiffness and mechanosignalling. Recurrent IDH1-mutant patient gliomas had a stiffer TNC-enriched ECM that our studies attributed to reduced miR-203 suppression of HIF1α and TNC mediated via a tension-dependent positive feedback loop. Thus, our work suggests that elevated ECM stiffness can independently foster glioblastoma aggression and contribute to glioblastoma recurrence via bypassing the protective activity of IDH1 mutational status.

Although gut flexures characterize gut morphology, the mechanisms underlying flexure formation remain obscure. Previously, we analyzed the mouse duodenojejunal flexure (DJF) as a model for its formation and reported asymmetric morphologies between the inner and outer bending sides of the fetal mouse DJF, implying their contribution to DJF formation. We now present the extracellular matrix (ECM) as an important factor for gut morphogenesis. We investigate ECM distribution during mouse DJF formation by histological techniques. In the intercellular space of the gut wall, high Alcian-Blue positivity for proteoglycans shifted from the outer to the inner side of the gut wall during DJF formation. Immunopositivity for fibronectin, collagen I, or pan-tenascin was higher at the inner than at the outer side. Collagen IV and laminins localized to the epithelial basement membrane. Beneath the mesothelium at the pre-formation stage, collagen IV and laminin immunopositivity showed inverse results, corresponding to the different cellular characteristics at this site. At the post-formation stage, however, laminin positivity beneath the mesothelium was the reverse of that observed during the pre-formation stage. High immunopositivity for collagen IV and laminins at the inner gut wall mesenchyme of the post-formation DJF implied a different blood vessel distribution. We conclude that ECM distribution changes spatiotemporally during mouse DJF formation, indicating ECM association with the establishment of asymmetric morphologies during this process.

Cocaine-associated environmental cues sustain relapse vulnerability by reactivating long-lasting memories of cocaine reward. During periods of abstinence, responding to cocaine cues can time-dependently intensify a phenomenon referred to as ‘incubation of cocaine craving'. Here, we investigated the role of the extracellular matrix protein brevican in recent (1 day after training) and remote (3 weeks after training) expression of cocaine conditioned place preference (CPP). Wild-type and Brevican heterozygous knock-out mice, which express brevican at ~50% of wild-type levels, received three cocaine–context pairings using a relatively low dose of cocaine (5 mg/kg). In a drug-free CPP test, heterozygous mice showed enhanced preference for the cocaine-associated context at the remote time point compared with the recent time point. This progressive increase was not observed in wild-type mice and it did not generalize to contextual-fear memory. Virally mediated overexpression of brevican levels in the hippocampus, but not medial prefrontal cortex, of heterozygous mice prevented the progressive increase in cocaine CPP, but only when overexpression was induced before conditioning. Post-conditioning overexpression of brevican did not affect remote cocaine CPP, suggesting that brevican limited the increase in remote CPP by altering neuro-adaptive mechanisms during cocaine conditioning. We provide causal evidence that hippocampal brevican levels control time-dependent enhancement of cocaine CPP during abstinence, pointing to a novel substrate that regulates incubation of responding to cocaine-associated cues.

Background The obesity paradox in COPD describes protective effects of obesity on lung pathology and inflammation. However, the underlying relationships between obesity, diet and disease outcomes in COPD are not fully understood. In this study we measured the response to dietary fatty acids upon markers of inflammation and remodelling in human lung cells from people with and without COPD. Methods Pulmonary fibroblasts were challenged with ω-3 polyunsaturated fatty acids (PUFAs), ω-6 PUFAs, saturated fatty acids (SFAs) or the obesity-associated cytokine TNFα. After 48–72 h release of the pro-inflammatory cytokines interleukin (IL)-6 and CXCL8 was measured using ELISA and mRNA expression and deposition of the extracellular matrix (ECM) proteins fibronectin, type I collagen, tenascin and perlecan were measured using qPCR or ECM ELISA, respectively. Results Challenge with the ω-6 PUFA arachidonic acid (AA), but not ω-3 PUFAs or SFAs, resulted in increased IL-6 and CXCL8 release from fibroblasts, however IL-6 and CXCL8 release was reduced in COPD (n = 19) compared to non-COPD (n = 36). AA-induced cytokine release was partially mediated by downstream mediators of cyclooxygenase (COX)-2 in both COPD and non-COPD. In comparison, TNFα-induced IL-6 and CXCL8 release was similar in COPD and non-COPD, indicating a specific interaction of AA in COPD. In patients with or without COPD, regression analysis revealed no relationship between BMI and cytokine release. In addition, AA, but not SFAs or ω-3 PUFAs reduced the basal deposition of fibronectin, type I collagen, tenascin and perlecan into the ECM in COPD fibroblasts. In non-COPD fibroblasts, AA-challenge decreased basal deposition of type I collagen and perlecan, but not fibronectin and tenascin. Conclusions This study shows that AA has disease-specific effects on inflammation and ECM protein deposition. The impaired response to AA in COPD might in part explain why obesity appears to have less detrimental effects in COPD, compared to other lung diseases. Electronic supplementary material The online version of this article (10.1186/s12931-018-0919-4) contains supplementary material, which is available to authorized users.

Background Perineuronal nets (PNNs), which are localized around neurons during development, are specialized forms of neural extracellular matrix with neuroprotective and plasticity-regulating roles. Hyaluronan and proteoglycan link protein 1 (HAPLN1), tenascin-R (TNR) and aggrecan (ACAN) are key elements of PNNs. In diseases characterized by neuritogenesis defects, the expression of these proteins is known to be downregulated, suggesting that PNNs may have a role in neural differentiation. Methods In this study, the mRNA and protein levels of HAPLN1, TNR and ACAN were determined and compared at specific time points of neural differentiation. We used PC12 cells as the in vitro model because they reflect this developmental process. Results On day 7, the HAPLN1 mRNA level showed a 2.9-fold increase compared to the non-differentiated state. However, the cellular HAPLN1 protein level showed a decrease, indicating that the protein may have roles in neural differentiation, and may be secreted during the early period of differentiation. By contrast, TNR mRNA and protein levels remained unchanged, and the amount of cellular ACAN protein showed a 3.7-fold increase at day 7. These results suggest that ACAN may be secreted after day 7, possibly due to its large amount of post-translational modifications. Conclusions Our results provide preliminary data on the expression of PNN elements during neural differentiation. Further investigations will be performed on the role of these elements in neurological disease models.

Primary vesicoureteral reflux (PVUR) is the most common malformation of the kidney and urinary tract, and reflux nephropathy is a major cause of chronic kidney disease in children. Recently, we reported mutations in the tenascin XB gene (TNXB) as a cause of PVUR with joint hypermobility. To define the role of rare variants in tenascin genes in the etiology of PVUR, we screened a cohort of patients with familial PVUR (FPVUR) and non-familial PVUR (NFPVUR) for rare missense variants inTNXB and the tenascin C gene (TNC) after excluding mutations in ROBO2 and SOX17. The screening procedure identified 134 individuals from 112 families with PVUR; two families with mutations in ROBO2 were excluded from further analysis. Rare missense variants in TNXB were found in the remaining 110 families, of which 5/55 (9 %) families had FPVUR and 2/55 (4 %) had NFPVUR. There were no differences in high-grade reflux or renal parenchymal scarring between patients with and without TNXB variants. All patients with TNXB rare variants who were tested exhibited joint hypermobility. Overall we were able to identify causes of FPVUR in 7/57 (12 %) families (9 % in TNXB and 3 % in ROBO2). In conclusion, the identification of a rare missense variant in TNXB in combination with a positive family history of VUR and joint hypermobility may represent a non-invasive method to diagnose PVUR and warrants further evaluation in other cohorts.

α-smooth muscle actin (α-SMA) and tenascin-C are stress-induced phenotypic features of myofibroblasts. The expression levels of these two proteins closely correlate with the extracellular mechanical microenvironment. We investigated how the expression of α-SMA and tenascin-C was altered in the periodontal ligament (PDL) under orthodontic loading to indirectly reveal the intrinsic mechanical microenvironment in the PDL. In this study, we demonstrated the synergistic effects of transforming growth factor-β1 (TGF-β1) and mechanical tensile or compressive stress on myofibroblast differentiation from human periodontal ligament cells (hPDLCs). The hPDLCs under higher tensile or compressive stress significantly increased their levels of α-SMA and tenascin-C compared with those under lower tensile or compressive stress. A similar trend was observed in the tension and compression areas of the PDL under continuous light or heavy orthodontic load in rats. During the time-course analysis of expression, we observed that an increase in α-SMA levels was matched by an increase in tenascin-C levels in the PDL under orthodontic load in vivo. The time-dependent variation of α-SMA and tenascin-C expression in the PDL may indicate the time-dependent variation of intrinsic stress under constant extrinsic loading.

Marginal iodine deficiency is a common health problem in pregnant women. Epidemiological and animal studies had shown that marginally maternal iodine deficiency could cause the mild changes of maternal thyroid function, eventually lead to a negative effect on neurodevelopment. But the underlying mechanisms responsible for the neurological impairment remain unclear. The aim of this study is to explore whether marginally maternal iodine deficiency could produce subtle changes in cell migration and cognitive function of offspring, and the optimal time of giving intervention to minimize the adverse effects. In the present study, we established a marginal iodine deficiency model, and iodine supplement was performed on pre-pregnancy (PP), G13 (gestation day 13), and postnatal day 0 (P0). Our data showed that there were changes in the cytoarchitecture and the percentage of bromodeoxyuridine (BrdU)-labeled cells in the cerebral cortex in marginal iodine deficiency rats. The Reelin expression was significantly lower, but Tenascin-C was higher in the cerebral cortex of marginal iodine deficiency group on P7 than the normal group, respectively. When iodine supplement, especially before G13 could reverse the abnormal expression of the two proteins involved in cell migration, which was consistent with the results of Morris Water Maze test. The three intervention groups had shorter escape latencies than the marginal iodine deficiency rats. The earlier that iodine is supplied, the better behavior performance would reach. Our findings suggested that iodine supplement in early stage of pregnancy could improve the cell migration of cerebral cortex and neurodevelopment of offspring.

We investigated the immunohistochemical localization of tenascin-C in 8-week-old rat periodontal ligaments. Tenascin-C immunoreactivity was detected in zones along with cementum and alveolar bone, and more intensely on the resorption surface of alveolar bone than on the formation surface. On the resorbing surface, tenascin-C immunoreactivity was detected in Howship's lacunae without osteoclasts, and in the interfibrous space of the periodontal ligaments, indicating that this molecule works as an adhesion molecule between bone and fibers of periodontal ligaments. Upon experimental tooth movement by inserting elastic bands (Waldo method), the physiological resorption surface of alveolar bone under compressive force showed enhanced bone resorption and enhanced tenascin-C immunoreactivity. However, on the physiological bone formation surface under compressive force, bone resorption was seen only occasionally, and no enhanced tenascin-C immunoreactivity was noted. In an experiment involving excessive occlusal loading to rat molars, transient bone resorption occurred within interradicular septa, but no enhanced tenascin-C immunoreactivity was seen in the periodontal ligaments. These results indicate that tenascin-C works effectively on the bone resorbing surface of physiological alveolar bone remodeling sites, rather than on the non-physiological transient bone resorbing surface. Fibronectin immunoreactivity was distributed evenly in the periodontal ligaments under experimental conditions. Co-localization of tenascin-C and fibronectin immunoreactivity was observed in many regions, but mutually exclusive expression patterns were also seen in some regions, indicating that fibronectin might not be directly involved in alveolar bone remodeling, but may play a role via interaction with tenascin-C.

Fibronectin (Fn) and tenascin-C (TnC) are two extracellular matrix proteins associated with remodeling changes. Fn and TnC gene and protein expression in lung tissue, including their predominant location in bronchial and pulmonary artery structures, have not yet been fully evaluated. The aim of the present study was to assess: (1) gene expression of Fn and TnC in lung samples from chronic obstructive pulmonary disease (COPD) and non-COPD subjects; and (2) protein content and location of Fn and TnC in both groups. Consecutive subjects requiring lung resection due to lung cancer surgery were included. Lung specimens were examined for gene expression by quantitative real-time PCR (values expressed as fold change ratio). The analysis of their protein content and location was performed by western blot and immunohistochemical studies, respectively. Patients were divided into two cohorts according to COPD status. A total of 41 patients (20 with COPD and 21 without COPD) were included. An enhanced Fn gene expression was observed in the COPD group compared to the non-COPD group (4.73 ± 0.54 vs. 2.65 ± 0.57; P = 0.012), whereas no differences in gene TnC expression were observed (2.91 ± 0.44 vs. 2.60 ± 0.48; P = 0.633). No differences in lung protein content and location were found between groups. Immunohistochemical evaluation showed a predominantly vascular and bronchial location of Fn and TnC in both groups. An enhanced lung gene expression of Fn was observed in COPD subjects compared to non-COPD subjects. No differences were found in Fn protein expression or in TnC gene or protein expression among groups.

The aims of this study were to develop a new coil, gellan sulfate core platinum coil (GSCC), that delivers tenascin-C (TNC) to an aneurysm (GSCC-TNC) and to evaluate the effects on intra-aneurysmal organization. We performed in vitro adsorption tests of TNC to gellan sulfate (GS). GSCC-TNC was produced by immersing GSCC in TNC solution under the following conditions (TNC concentration 10, 50, or 100 μg/mL; TNC immersion time 15, 30, or 60 min) by monitoring intra-aneurysmal organization in a rat blind-ended aneurysm model. In addition, 20 rats randomly underwent implantation of a platinum coil or the GSCC-TNC produced under optimum conditions into an aneurysm, whose organization effects were compared in a blind fashion at 2 weeks post-surgery. GS demonstrated a high affinity to TNC in a dose-dependent fashion (affinity constant = 1.79 × 10(10) (M(-1))). GSCC immersed in 10 μg/mL of TNC solution for 30 and 60 min induced similar and better organization of aneurysmal cavity compared with that for 15 min (the ratio of the organized areas in an aneurysmal cavity-15 min, 27.2 ± 11.8 %; 30 min, 75.6 ± 11.9 %; 60 min, 82.6 ± 19.7 %, respectively) with the preservation of the aneurysmal wall structure, while higher TNC concentrations caused the destruction of the aneurysmal wall. GSCC-TNC produced under 10 μg/mL of TNC solution for 30 min showed a significantly better organization of aneurysms compared with bare platinum coils in rats. A newly developed coil, GSCC-TNC, may be effective for improving intra-aneurysmal organization after coil embolization.

Management of acute and especially chronic rejection after human cardiac transplantation is still challenging. Chronic rejection, represented by allograft vasculopathy (CAV) and cardiac interstitial fibrosis (CIF) is known to cause severe long-term complications. Rejection associated tissue-remodelling entails the reoccurrence of fetal variants of Fibronectin (Fn) and Tenascin-C (Tn-C), which are virtually absent in adult human organs. In a rat model, an extensive re-expression could be demonstrated for ED-A(+) Fn with spatial association to CAV and CIF. Thus, it is of great interest to investigate the cardiac tissue expression and distribution in human samples. From 48 heart transplanted patients, 64 tissue specimens derived from right ventricular biopsies were available. Histopathological analysis was performed according to the International Society for Heart and Lung Transplantation (ISHLT) guidelines for the detection of acute rejection. By immunohistochemistry, protein expression of ED-A(+) Fn, B(+) Tn-C, alpha-smooth muscle actin, CD31 and CD45 was assessed and analysed semiquantitatively. Co-localisation studies were performed by means of immunofluorescence double labelling. Histopathological analysis of the 64 samples revealed different ISHLT grades (0R in 36 cases, 1R in 20 cases and 2R in 8 cases). There was a distinct and quantitatively relevant re-occurrence of ED-A(+) Fn and B(+) Tn-C in most samples. Semi-quantitative evaluation did not show any correlation to the acute rejection grade for all markers. Interestingly, significant correlations to the extent of inflammation could be shown for ED-A(+) Fn (r = 0.442, p = 0.000) and B(+) Tn-C (r = 0.408, p = 0.001) as well as between both proteins (r = 0.663, p = 0.000). A spatial association of ED-A(+) Fn and B(+) Tn-C to CAV and CIF could be demonstrated. A relevant re-occurrence of ED-A(+) Fn and B(+) Tn-C following human heart transplantation could be demonstrated with spatial association to signs of rejection and a significant correlation to tissue inflammation. These data might contribute to the identification of novel biomarkers reflecting the rejection process and to the development of promising strategies to image, prevent or treat cardiac rejection.

Tenascin-C (TNC) is a key molecule in tissue remodeling, and high levels are observed in many diseases, including heart failure, thrombosis, atherosclerosis, and cancer. High TNC expression by immunohistochemical analysis has been shown in invasive and metastasizing tissues from a variety of cancers, including colon, lung, brain, and breast. This study was conducted to investigate the serum level of TNC in breast cancer patients and its relationship with tumor progression and known prognostic parameters. Ninety-six breast cancer patients were enrolled into the study. Serum samples were obtained on first admission before adjuvant and metastatic treatments were given and at follow-up. Serum TNC levels were determined by the solid-phase sandwich enzyme-linked immunosorbent assay (ELISA) method. Median age of diagnosis was 48 years old (range, 29-80). Thirty-seven (39 %) patients had metastatic breast cancer. The mean TNC levels were found to be significantly higher in patients with breast cancer (344.1 ± 42.4 pg/mL) compared to those in healthy controls (137.2 ± 26.8 pg/mL) (p = 0.005). Serum TNC level in grade 3 tumors was found to be significantly higher than in grades 1-2 tumors (p = 0.04). No correlation was detected between serum TNC levels and other prognostic parameters analyzed, including presence of metastasis, lymph node involvement, and tumor size. Serum TNC level had no significantly adverse effect on survival in univariate and multivariate analyses (p = 0.65 and p = 0.85, respectively). In conclusion, although serum TNC levels are elevated, it has no predictive or prognostic roles on survival in breast cancer patients.

Janes and colleagues use organotypic 3D models, transcriptomic analyses, mathematical modelling and in vivo tumour data to show that single cells oscillate between two anticorrelated expression states defined by TGFBR3 and JUND. They show that this signalling circuit is controlled by the engagement of the extracellular matrix, and propose that dynamic changes in gene expression states might underlie breast tumour heterogeneity.

The extra-domain A1 of tenascin-C (TC-A1) is highly expressed in the extracellular matrix of tumours and on newly formed blood vessels and is thus a valuable target for radionuclide therapy. Tenarad is a fully human miniantibody or small immunoprotein (SIP, molecular weight 80 kDa) labelled with (131)I that is derived from a TC-A1-binding antibody. Previous phase I/II studies with a similar compound ((131)I-L19SIP) used for radioimmunotherapy (RIT) have shown preliminary efficacy in a variety of cancer types. In this ongoing phase I/II trial, Tenarad was administered to patients with recurrent Hodgkin's lymphoma (HL) refractory to conventional treatments.Eight patients (four men, four women; age range 19 - 41) were enrolled between April 2010 and March 2011. All patients had received a median of three previous lines of chemotherapy (range three to six) and seven had also undergone autologous stem cell transplantation (ASCT) or bone marrow transplantation. In addition, seven patients received external beam radiation. All patients had nodal disease, constitutional B symptoms and some showed extranodal disease in skeletal bone (four patients), lung (three), liver (two) and spleen (one). Baseline assessments included whole-body FDG PET with contrast-enhanced CT and diagnostic Tenarad planar and SPECT studies. Patients were considered eligible to receive a therapeutic dose of Tenarad (2.05 GBq/m(2)) if tumour uptake was more than four times higher than that of muscle.All patients were eligible and received the therapeutic dose of Tenarad. Only one patient developed grade 4 thrombocytopenia and leucocytopenia, requiring hospitalization and therapeutic intervention. All other patients had haematological toxicity of grade 3 or lower, which resolved spontaneously. At the first response assessment (4 - 6 weeks after therapy), one patient showed a complete response, one showed a partial response (PR) and five had disease stabilization (SD). Five patients were given up to three repeated Tenarad treatments. One patient showed SD which then improved to a PR, three showed clinical benefit while maintaining SD and one patient showed disease progression.Tenarad RIT is effective in chemorefractory HL and resulted in objective responses or clinical benefit in the majority of patients. Toxicity was acceptable despite the high load of prior treatments, previous ASCT and multiple Tenarad administrations. Further studies are planned to define the most effective schedule for this type of RIT in HL patients.

The determination of dermal injury age is important in forensic practice. It helps answer questions that are important to an investigation such as the timing of the injury and incident, the order of infliction (where there is more than one injury), the survival time after injury (post-infliction interval) and the relation of the injury to the incident. Despite the importance of injury age determination, there currently exists no reliable method to estimate dermal injury age. In this study, the expression of the following 14 mRNAs was studied in human dermal injuries and their usefulness in the estimation of human dermal injury age was evaluated: dual specificity phosphate 1 (DUSP1), interleukin 7 (IL7), vascular cell adhesion molecule 1, tenascin C, cluster of differentiation 14, interleukin 6, tumour necrosis factor alpha (TNFα), interleukin 1beta (IL1β), chymase 1 (CMA1), collagen type III alpha I, interleukin 2, collagen type I alpha I, collagen type I alpha II and vascular endothelial growth factor A (VEGFA). DUSP1, IL7, TNFα and VEGFA showed an initial decrease in expression during the early stages followed by an increase in expression towards the middle and late phases. IL1β and CMA1 expression was limited to specific time points. The remaining markers either showed inconsistent expression or were undetected in our samples. The expression patterns of the detected markers suggest they have potential to predict injury age, especially during the initial stages of injury healing, if used in combination with one another.

Ependymomas are relatively uncommon tumours of the central nervous system which arise from the ependymal lining of the ventricles and spinal canal. The molecular changes leading to ependymal oncogenesis are not completely understood. We examined chromosome 9q33-34 locus for gain, potential oncogenes at this locus (Notch-1 and Tenascin-C) and Notch pathway target genes (Hes-1, Hey-2 & C-myc) in ependymomas by fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC), respectively, to assess if they have any correlation with clinical characteristics. We analyzed 50 cases of ependymomas by FISH for 9q gain and by IHC for Notch-1 and its target gene proteins (Hes-1, Hey-2 and C-myc) expression. We also performed IHC for Tenascin-C to rule out any correlation with aggressiveness/grade of tumour. FISH study revealed significant chromosome 9q gain in ependymomas of adult onset (age > 18 years) and spinal cord origin. Notch-1 showed significantly more frequent immunohistochemical expression in supratentorial and anaplastic ependymomas. Tenascin-C (TN-C) expression was significant in intracranial, childhood (age ≤ 18 years) and anaplastic ependymomas. Of the three Notch pathway target gene proteins (Hes-1, Hey-2 and C-myc), Hes-1 and C-myc expression showed significant correlation with anaplastic and adult onset ependymomas, respectively. Genetic alterations are independent prognostic markers in ependymomas. A clinicopathological correlation with various molecular signatures may be helpful in the development of new therapeutic targets.

Papillary thyroid carcinoma (PTC) is the most common thyroid cancer, and it early metastasizes into regional lymph nodes. We evaluated immunohistochemically the expression of the hypoxia marker hypoxia-inducible factor 1α (HIF-1α) as well as extracellular matrix protein tenascin C as a marker of stroma remodelling in a cohort of 160 PTCs. Expression of HIF-1α was seen focally accentuated in 100 of the 160 tumours (62.5 %) including 16 cases with equivocal staining (faint staining or only single-cell staining) and 84 cases with unequivocal staining. HIF-1α expression correlated with the degree of desmoplastic stromal reaction as well as with the expression of tenascin C (p0.001, Kruskal-Wallis test, respectively). Moreover, expression of HIF-1α was significantly associated with the presence of lymph node metastases (p0.001, Mann-Whitney test) as well as signs of invasion, namely, peritumoural and extrathyroidal invasion as well as angioinvasion (p = 0.024, p0.001, p = 0.017, Mann-Whitney test, respectively). Additionally, PTC with unequivocal HIF-1α nuclear staining was a larger tumour than PTC with negative (-) or equivocal HIF-1α expression (p0.001, Kruskal-Wallis test). Interestingly, the expression of HIF-1α was not significantly associated with BRAF V600E status (p0.05, Mann-Whitney test). Our data show that the expression of HIF-1α is associated with stroma remodelling processes within the tumour and, thus, may play an important role in an invasive behaviour of these tumours independent of BRAF mutation status.

The extracellular matrix (ECM) protein tenascin-C (TN-C) is upregulated within glioma tissues and cultured glioma cell lines. TN-C possesses a multi-modular structure and a variety of functional properties have been reported for its domains. We describe five novel monoclonal antibodies identifying different domains of TN-C. The epitopes for these antibodies were investigated by using recombinantly expressed fibronectin type III domains of TN-C. The biological effects of TN-C fragments on glioma cell proliferation and adhesion were analyzed. The expression pattern of TN-C in human glioma tissue sections and in glioma cell lines was studied with the novel library of monoclonal antibodies. The immunocytochemical analyses of the established human glioma cell lines U-251-MG, U-373-MG and U-87-MG revealed distinct staining patterns for each antibody. Robust expression of TN-C was found within the tumor mass of surgery specimens from glioblastoma. In many cases, the expression of this ECM molecule was clearly associated with blood vessels, particularly with microvessels. Three of the new antibodies highlighted individual TN-C-expressing single cells in glioma tissues. The effect of TN-C domains on glioma cells was examined by a BrdU-proliferation assay and an adhesion assay. Short fragments of constitutively expressed TN-C-domains did not exert significant effects on the proliferation of glioma cells, whereas the intact molecule increased cell division rates. In contrast, the long fragment TNfnALL containing all of the FNIII domains of TN-C decreased proliferation. Additionally, we found strong differences between the adhesion-influencing properties of the recombinant fragments on glioma cells.

The tenascin-XB (TNXB) gene has antiadhesive effects, functions in matrix maturation in connective tissues, and localizes to the major histocompatibility complex class III region. We hypothesized that it may influence adaptive physiological response through an effect on blood vessel function. We identified a novel g.1324 A→G polymorphism at a TaqI recognition site in a 454 bp fragment of ovine TNXB and genotyped it in 150 Nigerian sheep using PCR-RFLP. The missense mutation changes glutamic acid (GAA) to glycine (GGA). Among SNP genotypes, significant differences (P

The aim of this study was to establish the time-course of molecular events in intrascapular brown adipose tissue (iBAT) during the development of diet-induced obesity using microarrays and molecular network analysis.C57BL/6J male inbred mice were fed a high-fat diet (HFD) or normal diet (ND) and killed at multiple time-points over 24 weeks.Global transcriptional changes in iBAT were determined by time-course microarrays of pooled RNA (n=6, pools per time-point) at 2, 4, 8, 20 and 24 weeks using Illumina MouseWG-6 v2.0 Beadchips. Molecular networks were constructed using the Ingenuity knowledgebase based on differentially expressed genes at each time-point.Body weight and subcutaneous adipose were progressively increased over 24 weeks, whereas iBAT was significantly increased between 6 and 12 weeks in HFD-fed C57BL/6J mice compared with controls. Blood glucose and insulin levels were increased between 16 and 24 weeks. Time-course microarrays, revealed 155 differentially expressed genes at one or more time-points over 24 weeks in the iBAT of HFD-fed mice compared with controls. Time-course network analysis revealed a network of skeletal muscle development genes that was activated between 2 and 4 weeks, subsequently a network of immune trafficking genes was activated at 8 weeks. After 20 and 24 weeks, multiple lipid metabolism and immune response networks were activated. Several target genes identified by time-course microarrays were independently validated using RT-qPCR. Tnnc1 was upregulated early between 2 and 4 weeks, later Cd68 and Col1a1 were upregulated between 20 and 24 weeks, whereas 11β-hydroxysteroid dehydrogenase (Hsd11b1) was consistently downregulated during the development of diet-induced obesity.Molecular networks in iBAT are modulated in a time-dependent manner in response to a HFD. A broad range of gene targets exists to alter molecular changes within iBAT during the development of diet-induced obesity.

In vitro and in vivo studies on the role of tenascins have shown that the two paradigmatic glycoproteins of the tenascin family, tenascin-C (TnC) and tenascin-R (TnR) play important roles in cell proliferation and migration, fate determination, axonal pathfinding, myelination, and synaptic plasticity. As components of the extracellular matrix, both molecules show distinct, but also overlapping dual functions in inhibiting and promoting cell interactions depending on the cell type, developmental stage and molecular microenvironment. They are expressed by neurons and glia as well as, for TnC, by cells of the immune system. The functional relationship between neural and immune cells becomes relevant in acute and chronic nervous system disorders, in particular when the blood brain and blood peripheral nerve barriers are compromised. In this review, we will describe the functional parameters of the two molecules in cell interactions during development and, in the adult, in synaptic activity and plasticity, as well as regeneration after injury, with TnC being conducive for regeneration and TnR being inhibitory for functional recovery. Although not much is known about the role of tenascins in neuroinflammation, we will describe emerging knowledge on the interplay between neural and immune cells in autoimmune diseases, such as multiple sclerosis and polyneuropathies. We will attempt to point out the directions of experimental approaches that we envisage would help gaining insights into the complex interplay of TnC and TnR with the cells that express them in pathological conditions of nervous and immune systems.

Background Intrinsic and acquired resistance to drug therapies remains a challenge for malignant melanoma patients. Intratumoral heterogeneities within the tumor microenvironment contribute additional complexity to the determinants of drug efficacy and acquired resistance. Methods We use 3D biomimetic platforms to understand dynamics in extracellular matrix (ECM) biogenesis following pharmaceutical intervention against mitogen-activated protein kinases (MAPK) signaling. We further determined temporal evolution of secreted ECM components by isogenic melanoma cell clones. Results We found that the cell clones differentially secrete and assemble a myriad of ECM molecules into dense fibrillar and globular networks. We show that cells can modulate their ECM biosynthesis in response to external insults. Fibronectin (FN) is one of the key architectural components, modulating the efficacy of a broad spectrum of drug therapies. Stable cell lines engineered to secrete minimal levels of FN showed a concomitant increase in secretion of Tenascin-C and became sensitive to BRAFV600E and ERK inhibition as clonally- derived 3D tumor aggregates. These cells failed to assemble exogenous FN despite maintaining the integrin machinery to facilitate cell- ECM cross-talk. We determined that only clones that increased FN production via p38 MAPK and β1 integrin survived drug treatment. Conclusions These data suggest that tumor cells engineer drug resistance by altering their ECM biosynthesis. Therefore, drug treatment may induce ECM biosynthesis, contributing to de novo resistance. Electronic supplementary material The online version of this article (doi:10.1186/s12885-016-2211-7) contains supplementary material, which is available to authorized users.

Biomarkers have gained immense scientific interest and clinical value in the practice of medicine. With unprecedented advances in high-throughput technologies, research interest in identifying novel and customized disease biomarkers for early detection, diagnosis, or drug responses is rapidly growing. Biomarkers can be identified in different levels of molecular biomarkers, networks biomarkers and dynamical network biomarkers (DNBs). The latter is a recently developed concept which relies on the idea that a cell is a complex system whose behavior is emerged from interplay of various molecules and this network of molecules dynamically changes over time. A DNB can serve as an early-warning signal of disease progression, or as a leading network that drives the system into the disease state and thus unravels mechanisms of disease initiation and progression. It is therefore of great importance to identify DNBs efficiently and reliably. In this work, the problem of DNB identification is defined as a multi-objective optimization problem and a framework to identify DNBs out of time-course high-throughput data is proposed. Temporal gene expression data of a lung injury with carbonyl chloride inhalation exposure has been used as a case study and the functional role of the discovered biomarker in the pathogenesis of lung injury has been thoroughly analyzed.