Correlation Between Endoglin and Malignant Phenotype in Human Melanoma Cells: Analysis of hsa-mir-214 and hsa-mir-370 in Cells and Their Extracellular Vesicles.

Endoglin (CD105) is an auxiliary receptor of transforming growth factor (TGF)-ß family members that is expressed in human melanomas. It is heterogeneously expressed by primary and metastatic melanoma cells, and endoglin targeting as a therapeutic strategy for melanoma tumors is currently been explored. However, its involvement in tumor development and malignancy is not fully understood. Here, we find that endoglin expression correlates with malignancy of primary melanomas and cultured melanoma cell lines. Next, we have analyzed the effect of ectopic endoglin expression on two miRNAs (hsa-mir-214 and hsa-mir-370), both involved in melanoma tumor progression and endoglin regulation. We show that compared with control cells, overexpression of endoglin in the WM-164 melanoma cell line induces; (i) a significant increase of hsa-mir-214 levels in small extracellular vesicles (EVs) as well as an increased trend in cells; and (ii) significantly lower levels of hsa-mir-370 in the EVs fractions, whereas no significant differences were found in cells. As hsa-mir-214 and hsa-mir-370 are not just involved in melanoma tumor progression, but they can also target endoglin-expressing endothelial cells in the tumor vasculature, these results suggest a complex and differential regulatory mechanism involving the intracellular and extracellular signaling of hsa-mir-214 and hsa-mir-370 in melanoma development and progression.

Implication of ERBB2 as a Predictive Tool for Survival in Patients with Pancreatic Cancer in Histological Studies.

Pancreatic cancer will be positioned by the year 2030 as the second cause of oncological death after lung cancer. The pathophysiology of the most common variety, which involves the adenocarcinoma of the pancreas, represents one of the main challenges for current oncology to explain its tumorigenesis and create a targeted treatment. The tumor microenvironment, metastatic capacity, and lack of early diagnosis lead patients to present advanced stages at the time of diagnosis. Despite numerous efforts, little progress has been made in clinical outcomes and with respect to the improved survival of these patients. For this reason, in recent years, numerous diagnostic tests, treatments, and possible approaches in the fields of radiotherapy, chemotherapy, immunotherapy, and surgery have been developed to find a combination of methods that improves life expectancy in patients diagnosed with this disease. On the other hand, the scientific community has made numerous advances in the molecular bases of pancreatic cancer since several oncogenetic pathways have been described and the markers expressed by the tumor have proven to be useful in the prognosis of pancreatic adenocarcinoma. These molecular alterations allow the study of possible therapeutic targets that improve the prognosis of these patients, but even numerous tumor cell-individual interactions must be explained to understand the underlying pathophysiology causing the high mortality. Therefore, the purpose of our study is to examine the expression of markers such as EGFR, Cyclin D1, andCDK4 in order to find a relationship with the possible long-term prognostic factors of patients affected by pancreatic ductal adenocarcinoma. Our results show that there is a prognostic role for ErbB2, EGFR, beta catenin, cyclin D1, and CDK4. Of these, we highlight the clinical importance of ErbB2 in the survival rates of patients who overexpress this component.

Evaluation of AIF-1 (Allograft Inflammatory Factor-1) as a Biomarker of Crohn's Disease Severity.

Background: Recently, increased tissue levels of AIF-1 have been shown in experimental colitis, supporting its role in intestinal inflammation. Therefore, we studied the levels of AIF-1 in Crohn's disease (CD). Methods: This study included 33 patients with CD (14 men and 19 women) who participated in the PREDICROHN project, a prospective multicenter study of the Spanish Group of Inflammatory bowel disease (GETECCU). Results: This article demonstrates declines with respect to baseline levels of serum AIF-1 in Crohn's disease (CD) patients after 14 weeks of treatment with anti-TNFs. Furthermore, in patients with active CD (HB ≥ 5), serum AIF-1 levels were significantly higher than those in patients without activity (HB ≤ 4). The study of serum AIF-1 in the same cohort, revealed an area under the ROC curve (AUC) value of AUC = 0.66 (p = 0.014), while for the CRP (C-reactive protein), (AUC) value of 0.69 (p = 0.0066), indicating a similar ability to classify CD patients by their severity. However, the combination of data on serum levels of AIF-1 and CRP improves the predictive ability of these analyses for classifying CD patients as active (HB ≥ 5) or inactive (HB ≤ 4). When we used the odds ratio (OR) formula, we observed that patients with CRP > 5 mg/L or AIF-1 > 200 pg/mL or both conditions were 13 times more likely to show HB ≥ 5 (active CD) than were those with both markers below these thresholds. Conclusion: The development of an algorithm that includes serum levels of AIF-1 and CRP could be useful for assessing Crohn's disease severity.

Towards an updated view on the clinical management of pancreatic adenocarcinoma: Current and future perspectives.

Pancreatic cancer has a dire prognosis and will represent the second leading cause of cancer death in the next 10 years. The multifactorial approach represents one of the main issues in controlling the extension of this neoplasm. In recent years, the characteristics of the tumor microenvironment, metastasis mechanisms and the relationship between immune system and neoplastic cells have been described, which has made it possible to understand the pathophysiology of pancreatic adenocarcinoma. Currently, there is a failure to provide an effective preventive method or early detection, so patients present with an advanced stage at the time of diagnosis. Despite numerous efforts, little progress has been made in clinical outcome and in improving survival in long term. Therefore, in the recent years, diverse diagnostic tests, treatments and possible approaches have been developed in the fields of radiotherapy, chemotherapy and surgery to find a combination of them that improves life expectancy in patients diagnosed with pancreatic cancer. At the moment, numerous clinical trials are being conducted to evaluate preventive diagnostic procedures such as serological markers or perfecting available imaging tests. On the other hand, implementation of immunotherapy is being studied in a neoplasm that has lagged in the application of this procedure since present possible treatments do not substantially improve quality of life. Therefore, the purpose of our study is to summarize the main progresses that have been made in the diagnosis, treatment and screening of this disease, explaining the limitations that have been observed and analyzing future prospects in the management of this illness.

Generation of a Soluble Form of Human Endoglin Fused to Green Fluorescent Protein.

Endoglin (Eng, CD105) is a type I membrane glycoprotein that functions in endothelial cells as an auxiliary receptor for transforming growth factor ß (TGF-ß)/bone morphogenetic protein (BMP) family members and as an integrin ligand, modulating the vascular pathophysiology. Besides the membrane-bound endoglin, there is a soluble form of endoglin (sEng) that can be generated by the action of the matrix metalloproteinase (MMP)-14 or -12 on the juxtamembrane region of its ectodomain. High levels of sEng have been reported in patients with preeclampsia, hypercholesterolemia, atherosclerosis and cancer. In addition, sEng is a marker of cardiovascular damage in patients with hypertension and diabetes, plays a pathogenic role in preeclampsia, and inhibits angiogenesis and tumor proliferation, migration, and invasion in cancer. However, the mechanisms of action of sEng have not yet been elucidated, and new tools and experimental approaches are necessary to advance in this field. To this end, we aimed to obtain a fluorescent form of sEng as a new tool for biological imaging. Thus, we cloned the extracellular domain of endoglin in the pEGFP-N1 plasmid to generate a fusion protein with green fluorescent protein (GFP), giving rise to pEGFP-N1/Eng.EC. The recombinant fusion protein was characterized by transient and stable transfections in CHO-K1 cells using fluorescence microscopy, SDS-PAGE, immunodetection, and ELISA techniques. Upon transfection with pEGFP-N1/Eng.EC, fluorescence was readily detected in cells, indicating that the GFP contained in the recombinant protein was properly folded into the cytosol. Furthermore, as evidenced by Western blot analysis, the secreted fusion protein yielded the expected molecular mass and displayed a specific fluorescent signal. The fusion protein was also able to bind to BMP9 and BMP10 in vitro. Therefore, the construct described here could be used as a tool for functional in vitro studies of the extracellular domain of endoglin.

Relationship between IGF-1 and body weight in inflammatory bowel diseases: Cellular and molecular mechanisms involved.

Inflammatory bowel diseases (IBD), represented by ulcerative colitis (UC) and Crohn's disease (CD), are characterized by chronic inflammation of the gastrointestinal tract, what leads to diarrhea, malnutrition, and weight loss. Depression of the growth hormone-insulin-like growth factor-1 axis (GH-IGF-1 axis) could be responsible of these symptoms. We demonstrate that long-term treatment (54 weeks) of adult CD patients with adalimumab (ADA) results in a decrease in serum IGF-1 without changes in serum IGF-1 binding protein (IGF1BP4). These results prompted us to conduct a preclinical study to test the efficiency of IGF-1 in the medication for experimental colitis. IGF-1 treatment of rats with DSS-induced colitis has a beneficial effect on the following circulating biochemical parameters: glucose, albumin, and total protein levels. In this experimental group we also observed healthy maintenance of colon size, body weight, and lean mass in comparison with the DSS-only group. Histological analysis revealed restoration of the mucosal barrier with the IGF-1 treatment, which was characterized by healthy quantities of mucin production, structural maintenance of adherers junctions (AJs), recuperation of E-cadherin and ß-catenin levels and decrease in infiltrating immune cells and in metalloproteinase-2 levels. The experimentally induced colitis caused activation of apoptosis markers, including cleaved caspase 3, caspase 8, and PARP and decreases cell-cycle checkpoint activators including phosphorylated Rb, cyclin E, and E2F1. The IGF-1 treatment inhibited cyclin E depletion and partially protects PARP levels. The beneficial effects of IGF-1 in experimental colitis could be explained by a re-sensitization of the IGF-1/IRS-1/AKT cascade to exogenous IGF-1. Given these results, we postulate that IGF-1 treatment of IBD patients could prove to be successful in reducing disease pathology.

Homozygous GDF2 nonsense mutations result in a loss of circulating BMP9 and BMP10 and are associated with either PAH or an "HHT-like" syndrome in children.

BACKGROUND: Disrupted endothelial BMP9/10 signaling may contribute to the pathophysiology of both hereditary hemorrhagic telangiectasia (HHT) and pulmonary arterial hypertension (PAH), yet loss of circulating BMP9 has not been confirmed in individuals with ultra-rare homozygous GDF2 (BMP9 gene) nonsense mutations. We studied two pediatric patients homozygous for GDF2 (BMP9 gene) nonsense mutations: one with PAH (c.[76C>T];[76C>T] or p.[Gln26Ter];[Gln26Ter] and a new individual with pulmonary arteriovenous malformations (PAVMs; c.[835G>T];[835G>T] or p.[Glu279Ter];[Glu279Ter]); both with facial telangiectases. METHODS: Plasma samples were assayed for BMP9 and BMP10 by ELISA. In parallel, serum BMP activity was assayed using an endothelial BRE-luciferase reporter cell line (HMEC1-BRE). Proteins were expressed for assessment of secretion and processing. RESULTS: Plasma levels of both BMP9 and BMP10 were undetectable in the two homozygous index cases and this corresponded to low serum-derived endothelial BMP activity in the patients. Measured BMP9 and BMP10 levels were reduced in the asymptomatic heterozygous p.[Glu279Ter] parents, but serum activity was normal. Although expression studies suggested alternate translation can be initiated at Met57 in the p.[Gln26Ter] mutant, this does not result in secretion of functional BMP9. CONCLUSION: Collectively, these data show that homozygous GDF2 mutations, leading to a loss of circulating BMP9 and BMP10, can cause either pediatric PAH and/or "HHT-like" telangiectases and PAVMs. Although patients reported to date have manifestations that overlap with those of HHT, none meet the Curaçao criteria for HHT and seem distinct from HHT in terms of the location and appearance of telangiectases, and a tendency for tiny, diffuse PAVMs.

Differential Expression of Circulating Plasma miRNA-370 and miRNA-10a from Patients with Hereditary Hemorrhagic Telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant, vascular disorder that presents with telangiectases and arteriovenous malformations. HHT is a genetically heterogeneous disorder, involving mutations in endoglin (ENG; HHT1) and activin receptor-like kinase 1 (ACVRL1/ALK1; HHT2) genes that account for over 85% of all HHT patients. The current diagnosis of HHT patients remains at the clinical level, but many suspected patients do not have a clear HHT diagnosis or do not show pathogenic mutations in HHT genes. This situation has prompted the search for biomarkers to help in the early diagnosis of the disease. We have analyzed the plasma levels in HHT patients of selected micro-RNAs (miRNAs), small single-stranded RNAs that regulate gene expression at the transcriptional level by interacting with specific RNA targets. A total of 16 HHT1 and 17 HHT2 plasma samples from clinically confirmed patients and 16 controls were analyzed in this study. Total RNA was purified from plasma, and three selected miRNAs (miRNA-10a, miRNA-214, and miRNA-370), related to the pathobiology of cardiovascular diseases and potentially targeting ENG or ALK1, were measured by quantitative polymerase chain reaction. Compared with controls, levels of miRNA-370, whose putative target is ENG, were significantly downregulated in HHT1, but not in HHT2, whereas the levels of miRNA-10a, whose putative target is ALK1, were significantly upregulated in HHT2, but not in HHT1. In addition, the levels of miRNA-214, potentially targeting ENG and ALK1, did not change in either HHT1 or HHT2 patients versus control samples. While further studies are warranted, these results suggest that dysregulated plasma levels of miRNA-370 or miRNA-10a could help to identify undiagnosed HHT1 or HHT2 patients, respectively.

Endoglin Protein Interactome Profiling Identifies TRIM21 and Galectin-3 as New Binding Partners.

Endoglin is a 180-kDa glycoprotein receptor primarily expressed by the vascular endothelium and involved in cardiovascular disease and cancer. Heterozygous mutations in the endoglin gene (ENG) cause hereditary hemorrhagic telangiectasia type 1, a vascular disease that presents with nasal and gastrointestinal bleeding, skin and mucosa telangiectases, and arteriovenous malformations in internal organs. A circulating form of endoglin (alias soluble endoglin, sEng), proteolytically released from the membrane-bound protein, has been observed in several inflammation-related pathological conditions and appears to contribute to endothelial dysfunction and cancer development through unknown mechanisms. Membrane-bound endoglin is an auxiliary component of the TGF-ß receptor complex and the extracellular region of endoglin has been shown to interact with types I and II TGF-ß receptors, as well as with BMP9 and BMP10 ligands, both members of the TGF-ß family. To search for novel protein interactors, we screened a microarray containing over 9000 unique human proteins using recombinant sEng as bait. We find that sEng binds with high affinity, at least, to 22 new proteins. Among these, we validated the interaction of endoglin with galectin-3, a secreted member of the lectin family with capacity to bind membrane glycoproteins, and with tripartite motif-containing protein 21 (TRIM21), an E3 ubiquitin-protein ligase. Using human endothelial cells and Chinese hamster ovary cells, we showed that endoglin co-immunoprecipitates and co-localizes with galectin-3 or TRIM21. These results open new research avenues on endoglin function and regulation.

Characterization of a family mutation in the 5' untranslated region of the endoglin gene causative of hereditary hemorrhagic telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is a vascular disease characterized by nose and gastrointestinal bleeding, telangiectases in skin and mucosa, and arteriovenous malformations in major internal organs. Most patients carry a mutation in the coding region of the endoglin (ENG) or activin A receptor type II-1 (ACVRL1) gene. Nonetheless, in around 15% of patients, sequencing analysis and duplication/deletion tests fail to pinpoint mutations in the coding regions of these genes. In these cases, it has been shown that sequencing of the 5'-untranslated region (5'UTR) of ENG may be useful to identify novel mutations in the ENG non-coding region. Here we report the genetic characterization and functional analysis of the heterozygous mutation c.-142A>T in the 5'UTR region of ENG found in a family with several members affected by HHT. This variant gives rise to a new initiation codon of the protein that involves the change in its open reading frame. Transfection studies in monkey cells using endoglin expression vectors demonstrated that c-142A>T mutation results in a clear reduction in the levels of the endoglin protein. These results support the inclusion of the 5'UTR of ENG in the standard genetic testing for HHT to increase its sensitivity.

Characterization of a mutation in the zona pellucida module of Endoglin that causes Hereditary Hemorrhagic Telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is a vascular rare disease characterized by nose and gastrointestinal bleeding, skin and mucosa telangiectasias, and arteriovenous malformations in internal organs. HHT shows an autosomal dominant inheritance and a worldwide prevalence of approximately 1:5000 individuals. In >80% of patients, HHT is caused by mutations in either ENG (HHT1) or ACVRL1 (HHT2) genes, which code for the membrane proteins Endoglin and Activin A Receptor Type II-Like Kinase 1 (ALK1), respectively, both belonging to the TGF-ß/BMP signaling pathway. In this work, we describe a novel mutation in exon 9 of ENG (c.1145 G > A) found in five affected members of a family, all of them with characteristic symptoms of HHT. This mutation involves Cys382 residue of the Endoglin protein (p.Cys382 > Tyr) in the zona pellucida (ZP) module of its extracellular region. This is a critical residue involved in a conserved intrachain disulphide bond and in the correct folding of the protein. In fact, transfection studies in human cells using Endoglin expression vectors demonstrated that the p.Cys382 > Tyr mutation results in a marked reduction in the levels of the Endoglin protein. These results demonstrate the pathogenic role for this variant in HHT1 and confirm the key function of Cys382 in Endoglin expression.

Thyroid Hormone Receptors Regulate the Expression of microRNAs with Key Roles in Skin Homeostasis.

BACKGROUND: MicroRNAs (miRNAs) play a unique role in posttranscriptional regulation of gene expression and control different aspects of skin development, homeostasis, and disease. Although it is generally accepted that thyroid hormone signaling is important in skin pathophysiology, the role of their nuclear receptors (TRs) in cutaneous miRNA expression has yet to be explored. METHODS: RNAseq was used to compare the skin miRnome of wild-type mice and genetically modified mice lacking both TRα1 and TRß, the main thyroid hormone binding isoforms. Changes in miRNAs with a crucial role in skin physiopathology were confirmed by stem-loop quantitative polymerase chain reaction in both total skin and isolated keratinocytes, and the levels of their target mRNAs were evaluated by real-time polymerase chain reaction. RESULTS: The skin of TRα1/TRß knockout mice displays altered levels of >50 miRNAs. Among the downregulated species are several miRNAs, including miR-21, miR-31, miR-34, and miR-203, with crucial roles in skin homeostasis. TRα1 appears to be the main isoform responsible for their regulation. Increased levels of gene transcripts previously shown to be bona fide targets of these miRNAs are also found in the skin and keratinocytes of TR-deficient mice. This suggests that multiple miRNAs that are downregulated in the absence of TRs cooperate to regulate gene expression in the skin. CONCLUSIONS: The miRNAs reduced in TRα1/TRß knockout mice are known to play crucial roles in epidermal proliferation, hair cycling, wound healing, stem-cell function, and tumor development, all processes altered in the absence of TRs. These results suggest that their regulation could contribute to the skin defects found in these mice and to the skin disorders associated with altered thyroid status in humans.

Functional analysis of a novel ENG variant in a patient with hereditary hemorrhagic telangiectasia (HHT) identifies a new Sp1 binding-site.

Hereditary Hemorrhagic Telangiectasia (HHT) is a rare disease, with an autosomal dominant inheritance and a worldwide incidence of about 1: 5000 individuals. In >80% of patients, HHT is caused by mutations in either ENG or ACVRL1, which code for ENDOGLIN and Activin A Receptor Type II-Like Kinase 1 (ALK1), belonging to the TGF-ß/BMP signalling pathway. Typical HHT clinical features are mucocutaneous telangiectases, arteriovenous malformations, spontaneous and recurrent epistaxis, as well as gastrointestinal bleedings. An additional, but less frequent, clinical manifestation in some HHT patients is the presence of Pulmonary Arterial Hypertension (PAH). The aim of this work is to describe the functional role of a novel ENG intronic variant found in a patient affected by both HHT and PAH, in order to assess whether it has a pathogenic role. We proved that the variant lies in a novel binding-site for the transcription factor Sp1, known to be involved in the regulation of ENG and ACVRL1 transcription. We confirmed a pathogenic role for this intronic variant, as it significantly reduces ENG transcription by affecting this novel Sp1 binding-site.

Endoglin and alk1 as therapeutic targets for hereditary hemorrhagic telangiectasia.

INTRODUCTION: Hereditary Haemorrhagic Telangiectasia (HHT) is as an autosomal dominant trait characterized by frequent nose bleeds, mucocutaneous telangiectases, arteriovenous malformations (AVMs) of the lung, liver and brain, and gastrointestinal bleedings due to telangiectases. HHT is originated by mutations in genes whose encoded proteins are involved in the transforming growth factor ß (TGF-ß) family signalling of vascular endothelial cells. In spite of the great advances in the diagnosis as well as in the molecular, cellular and animal models of HHT, the current treatments remain just at the palliative level. Areas covered: Pathogenic mutations in genes coding for the TGF-ß receptors endoglin (ENG) (HHT1) or the activin receptor-like kinase-1 (ACVRL1 or ALK1) (HHT2), are responsible for more than 80% of patients with HHT. Therefore, ENG and ALK1 are the main potential therapeutic targets for HHT and the focus of this review. The current status of the preclinical and clinical studies, including the anti-angiogenic strategy, have been addressed. Expert opinion: Endoglin and ALK1 are attractive therapeutic targets in HHT. Because haploinsufficiency is the pathogenic mechanism in HHT, several therapeutic approaches able to enhance protein expression and/or function of endoglin and ALK1 are keys to find novel and efficient treatments for the disease.

Thyroid hormones inhibit TGF-ß signaling and attenuate fibrotic responses.

TGF-ß, the most potent profibrogenic factor, acts by activating SMAD (mothers against decapentaplegic) transcription factors, which bind to SMAD-binding elements in target genes. Here, we show that the thyroid hormone triiodothyronine (T3), through binding to its nuclear receptors (TRs), is able to antagonize transcriptional activation by TGF-ß/SMAD. This antagonism involves reduced phosphorylation of SMADs and a direct interaction of the receptors with SMAD3 and SMAD4 that is independent of T3-mediated transcriptional activity but requires residues in the receptor DNA binding domain. T3 reduces occupancy of SMAD-binding elements in response to TGF-ß, reducing histone acetylation and inhibiting transcription. In agreement with this transcriptional cross-talk, T3 is able to antagonize fibrotic processes in vivo. Liver fibrosis induced by carbon tetrachloride is attenuated by thyroid hormone administration to mice, whereas aged TR knockout mice spontaneously accumulate collagen. Furthermore, skin fibrosis induced by bleomycin administration is also reduced by the thyroid hormones. These findings define an important function of the thyroid hormone receptors and suggest TR ligands could have beneficial effects to block the progression of fibrotic diseases.

Impaired hair growth and wound healing in mice lacking thyroid hormone receptors.

Both clinical and experimental observations show that the skin is affected by the thyroidal status. In hypothyroid patients the epidermis is thin and alopecia is common, indicating that thyroidal status might influence not only skin proliferation but also hair growth. We demonstrate here that the thyroid hormone receptors (TRs) mediate these effects of the thyroid hormones on the skin. Mice lacking TRα1 and TRß (the main thyroid hormone binding isoforms) display impaired hair cycling associated to a decrease in follicular hair cell proliferation. This was also observed in hypothyroid mice, indicating the important role of the hormone-bound receptors in hair growth. In contrast, the individual deletion of either TRα1 or TRß did not impair hair cycling, revealing an overlapping or compensatory role of the receptors in follicular cell proliferation. In support of the role of the receptors in hair growth, TRα1/TRß-deficient mice developed alopecia after serial depilation. These mice also presented a wound-healing defect, with retarded re-epithelialization and wound gaping, associated to impaired keratinocyte proliferation. These results reinforce the idea that the thyroid hormone nuclear receptors play an important role on skin homeostasis and suggest that they could be targets for the treatment of cutaneous pathologies.

microRNAs 424 and 503 are mediators of the anti-proliferative and anti-invasive action of the thyroid hormone receptor beta.

The thyroid hormone receptors (TRs) mediate tumor suppressive effects in hepatocarcinoma and breast cancer cells. Here we show that incubation of hepatocarcinoma SK-hep1 cells expressing TRb with the thyroid hormone T3 induces transcription of the polycistronic message coding for microRNAs 424 and 503. TRb binds to the promoter region of these miRNAs and T3 induces an exchange of corepressors and coactivators inducing histone acetylation and transcriptional stimulation. We have validated cell cycle components as targets of these miRNAs. Overexpression of miR-424 mimicked the repressive effect of T3 on cell proliferation, growth in suspension, migration and invasion. Knockdown of miR-424 or miR-503 reduced the inhibitory effect of the hormone. T3 increased miR-424 and miR-503 in breast cancer cells expressing TRb, and this induction is also involved in the anti-invasive effects of the hormone. Furthermore, miR-424 or miR-503 depletion enhanced extravasation to the lungs of hepatocarcinoma cells injected in the tail vein of mice. The levels of these miRNAs were reduced in xenograft tumors formed in hypothyroid nude mice that are more invasive. Therefore, miR-424 or miR-503 mediate anti-proliferative and anti-invasive actions of TRb both in cultured cells and in vivo.

The growing role of gene methylation on endocrine function.

DNA methylation is the best studied epigenetic factor, playing a key role in producing stable changes in gene expression, thus defining cell identity and function and adapting cells to environmental changes. DNA methylation has also been recently shown to mediate cell responses to physiological endocrine signals. Moreover, alterations of the normal DNA methylation pattern can also contribute to the development of endocrine and metabolic diseases and can explain the relationship between an individual's genetic background, the environment, and disease. It should be remarked that although DNA methylation and demethylation are active processes, epigenetic changes produced during development can impact adult processes, establishing the idea that endocrine function can be persistently affected by events occurring in early life. Given the complexity of the endocrine system, both genetic and epigenetic processes, including DNA methylation, must be involved in its proper development and functioning. In this study, we summarize the recent knowledge in the field of DNA methylation and endocrinology. Given that DNA methylation can be involved in a number of endocrine and metabolic disorders, understanding and manipulating this modification opens a new door for preventing and treating endocrine diseases.

The thyroid hormone receptors as tumor suppressors.

In addition to the well-known role of the thyroid hormone receptors (TRs) in growth, development and metabolism, there is increasing evidence that they have profound effects on cell proliferation and malignant transformation. TRs repress transcriptional induction of cyclin D1 by the ras oncogene and block transformation and tumor formation by Ras-transformed fibroblasts in nude mice. Mutant receptors that do not bind coactivators are able to display these actions, whereas receptors defective in corepressors binding are unable to antagonize the responses to the ras oncogene. Furthermore, expression of TRß1 in hepatocarcinoma and breast cancer cells abolishes anchorage-independent growth and migration, blocks responses to growth factors and represses expression of prometastatic genes, reducing tumor growth and strongly inhibiting invasiveness, extravasation and metastasis formation in euthyroid mice. By contrast, when cells are inoculated into hypothyroid host, tumor growth is retarded, but tumors are more invasive and metastatic growth is enhanced. Increased aggressiveness and tumor growth retardation was also observed with parental cells that do not express TRs, showing that changes secondary to hypothyroidism can modulate tumor progression and metastatic growth independently of the presence of TRs on the tumor cells. Finally, increased malignancy of skin tumors is found in mice lacking TRs, further demonstrating the role of these receptors as inhibitors of tumor progression and suggesting that they represent a potential therapeutic target in cancer.

Thyroid receptor: roles in cancer.

The thyroid hormone receptors, encoded by the TRalpha and TRbeta genes, are ligand-dependent transcription factors that belong to the nuclear receptor superfamily. In addition to the role of these receptors in growth, development and metabolism, there is increasing evidence that they also inhibit transformation and act as tumor suppressors. Aberrant TR action, as well as receptor silencing, are common events in human cancer, and TRs also have an important role in tumor progression in experimental animal models, suggesting that these receptors constitute a novel therapeutic target in cancer. This review highlights recent studies on mechanisms by which loss of expression and/or function of these receptors results in a selective advantage for cellular transformation, tumor development and metastatic growth.