CYLD regulates keratinocyte differentiation and skin cancer progression in humans

CYLD is a gene mutated in familial cylindromatosis and related diseases, leading to the development of skin appendages tumors. Although the deubiquitinase CYLD is a skin tumor suppressor, its role in skin physiology is unknown. Using skin organotypic cultures as experimental model to mimic human skin, we have found that CYLD acts as a regulator of epidermal differentiation in humans through the JNK signaling pathway. We have determined the requirement of CYLD for the maintenance of epidermal polarity, keratinocyte differentiation and apoptosis. We show that CYLD overexpression increases keratinocyte differentiation while CYLD loss of function impairs epidermal differentiation. In addition, we describe the important role of CYLD in the control of human non-melanoma skin cancer progression. Our results show the reversion of the malignancy of human squamous cell carcinomas that express increased levels of CYLD, while its functional inhibition enhances the aggressiveness of these tumors which progress toward spindle cell carcinomas. We have found that the mechanisms through which CYLD regulates skin cancer progression include the control of tumor differentiation, angiogenesis and cell survival. These findings of the role of CYLD in human skin cancer prognosis make our results relevant from a therapeutic point of view, and open new avenues for exploring novel cancer therapiesThis work was funded by grants from the Ministerio de Ciencia e Innovación PI06/1233 and PI10/01480 to MLC, and SAF2010-22156 to ARS

IKKβ overexpression together with a lack of tumour suppressor genes causes ameloblastic odontomas in mice

Odontogenic tumours are a heterogeneous group of lesions that develop in the oral cavity region and are characterized by the formation of tumoural structures that differentiate as teeth. Due to the diversity of their histopathological characteristics and clinical behaviour, the classification of these tumours is still under debate. Alterations in morphogenesis pathways such as the Hedgehog, MAPK and WNT/β-catenin pathways are implicated in the formation of odontogenic lesions, but the molecular bases of many of these lesions are still unknown. In this study, we used genetically modified mice to study the role of IKKβ (a fundamental regulator of NF-κB activity and many other proteins) in oral epithelial cells and odontogenic tissues. Transgenic mice overexpressing IKKβ in oral epithelial cells show a significant increase in immune cells in both the oral epithelia and oral submucosa. They also show changes in the expression of several proteins and miRNAs that are important for cancer development. Interestingly, we found that overactivity of IKKβ in oral epithelia and odontogenic tissues, in conjunction with the loss of tumour suppressor proteins (p53, or p16 and p19), leads to the appearance of odontogenic tumours that can be classified as ameloblastic odontomas, sometimes accompanied by foci of secondary ameloblastic carcinomas. These tumours show NF-κB activation and increased β-catenin activity. These findings may help to elucidate the molecular determinants of odontogenic tumourigenesis and the role of IKKβ in the homoeostasis and tumoural transformation of oral and odontogenic epitheliaThis work was funded by project PI17/00578, from the “Instituto de Salud Carlos III” (Ministry of Science, Innovation and Universities) and co-funded by the European Regional Development Fund, and approved by the Ethics Committee of our Institution. It has been founded also by projects CB16/12/00228, PI16/00161, RD16/0011/0011, RD12/0019/0023 and SAF2017–84248-PS

TNF-α and IL-10 downregulation and marked oxidative stress in Neuromyelitis Optica

<p>Abstract</p> <p>Background</p> <p>Neuromyelitis optica is a central nervous system demyelinating and inflammatory syndrome. The objective of this study is to identify cytokines related to the cellular immune response as well as blood brain barrier integrity and oxidative stress.</p> <p>Methods</p> <p>We performed a molecular characterization of cellular immune response and oxidative stress in serum from relapsing-NMO (R-NMO) patients and established the correlations between the clinical measurements and molecular parameters using the Bayesian approach.</p> <p>Serum samples from 11 patients with R-NMO diagnosed according to Wingerchuk criteria and matched in terms of age, gender and ethnicity with the healthy controls were analyzed. The levels of TNF-<it>α</it>, IFN-<it>γ</it>, IL-10, MMP-9, TIMP-1 and oxidative stress markers: malondialdehyde, advanced oxidation protein products, peroxidation potential, superoxide dismutase, catalase, and total hydroperoxides were measured.</p> <p>Results</p> <p>We found almost undetectable levels of TNF-<it>α</it>, a decreased production of IL-10 and a significant up-regulation of every oxidative stress biomarker studied. The insufficient production of TNF-<it>α </it>and IL-10 in R-NMO patients, which are two important players of T cell mediated immunoregulation, suggest an effector – regulator imbalance. The overproduction of oxygen reactive species as a consequence of the chronic inflammatory milieu is reflected on the excess of oxidative damage mediators detected. Furthermore, Multidimensional Scaling and a Bayesian linear regression model revealed a significant linear dependence between Expanded Disability Status Scale Kurtzke and TIMP-1; pointing to a possible predictive or prognostic value of this clinical-molecular relationship.</p> <p>Conclusion</p> <p>These results suggest that there is a breakdown in immunoregulatory mechanisms and noteworthy pro-oxidant environment contributing to NMO pathogenesis.</p

Increased IKKα Expression in the Basal Layer of the Epidermis of Transgenic Mice Enhances the Malignant Potential of Skin Tumors

Non-melanoma skin cancer is the most frequent type of cancer in humans. In this study we demonstrate that elevated IKKα expression in murine epidermis increases the malignancy potential of skin tumors. We describe the generation of transgenic mice overexpressing IKKα in the basal, proliferative layer of the epidermis and in the outer root sheath of hair follicles. The epidermis of K5-IKKα transgenic animals shows several alterations such as hyperproliferation, mislocalized expression of integrin-α6 and downregulation of the tumor suppressor maspin. Treatment of the back skin of mice with the mitogenic agent 12-O-tetradecanoylphorbol-13-acetate causes in transgenic mice the appearance of different preneoplastic changes such as epidermal atypia with loss of cell polarity and altered epidermal tissue architecture, while in wild type littermates this treatment only leads to the development of benign epidermal hyperplasia. Moreover, in skin carcinogenesis assays, transgenic mice carrying active Ha-ras (K5-IKKα-Tg.AC mice) develop invasive tumors, instead of the benign papillomas arising in wild type-Tg-AC mice also bearing an active Ha-ras. Therefore we provide evidence for a tumor promoter role of IKKα in skin cancer, similarly to what occurs in other neoplasias, including hepatocarcinomas and breast, prostate and colorectal cancer. The altered expression of cyclin D1, maspin and integrin-α6 in skin of transgenic mice provides, at least in part, the molecular bases for the increased malignant potential found in the K5-IKKα skin tumors

Exocrine pancreatic disorders in transsgenic mice expressing human keratin 8

9 pages, 6 figures, 2 tables.-- et al.Keratins K8 and K18 are the major components of the intermediate-filament cytoskeleton of simple epithelia. Increased levels of these keratins have been correlated with various tumor cell characteristics, including progression to malignancy, invasive behavior, and drug sensitivity, although a role for K8/K18 in tumorigenesis has not yet been demonstrated. To examine the function of these keratins, we generated mice expressing the human K8 (hk8) gene, which leads to a moderate keratin-content increase in their simple epithelia. These mice displayed progressive exocrine pancreas alterations, including dysplasia and loss of acinar architecture, redifferentiation of acinar to ductal cells, inflammation, fibrosis, and substitution of exocrine by adipose tissue, as well as increased cell proliferation and apoptosis. Histological changes were not observed in other simple epithelia, such as the liver. Electron microscopy showed that transgenic acinar cells have keratins organized in abundant filament bundles dispersed throughout the cytoplasm, in contrast to control acinar cells, which have scarce and apically concentrated filaments. The phenotype found was very similar to that reported for transgenic mice expressing a dominant-negative mutant TGF-β type II receptor (TGFβRII mice). We show that these TGFβRII mutant mice also have elevated K8/K18 levels. These results indicate that simple epithelial keratins play a relevant role in the regulation of exocrine pancreas homeostasis and support the idea that disruption of mechanisms that normally regulate keratin expression in vivo could be related to inflammatory and neoplastic pancreatic disorders.This work was supported in part by grant PB 94-1230 of the Spanish Dirección General de Investigación Científica y Tecnológica.Peer reviewe

Estudio de la regulacion de la expresion y de la funcion de la queratina 8 humana de epitelios simples

Centro de Informacion y Documentacion Cientifica (CINDOC). C/Joaquin Costa, 22. 28002 Madrid. SPAIN / CINDOC - Centro de Informaciòn y Documentaciòn CientìficaSIGLEESSpai

Intermediate Filaments Interact with Dormant Ezrin in Intestinal Epithelial Cells

Ezrin connects the apical F-actin scaffold to membrane proteins in the apical brush border of intestinal epithelial cells. Yet, the mechanisms that recruit ezrin to the apical domain remain obscure. Using stable CACO-2 transfectants expressing keratin 8 (K8) antisense RNA under a tetracycline-responsive element, we showed that the actin-ezrin scaffold cannot assemble in the absence of intermediate filaments (IFs). Overexpression of ezrin partially rescued this phenotype. Overexpression of K8 in mice also disrupted the assembly of the brush border, but ezrin distributed away from the apical membrane in spots along supernumerary IFs. In cytochalasin D-treated cells ezrin localized to a subapical compartment and coimmunoprecipitated with IFs. Overexpression of ezrin in undifferentiated cells showed a Triton-insoluble ezrin compartment negative for phospho-T567 (dormant) ezrin visualized as spots along IFs. Pulse-chase analysis showed that Triton-insoluble, newly synthesized ezrin transiently coimmunoprecipitates with IFs during the first 30 min of the chase. Dormant, but not active (p-T567), ezrin bound in vitro to isolated denatured keratins in Far-Western analysis and to native IFs in pull-down assays. We conclude that a transient association to IFs is an early step in the polarized assembly of apical ezrin in intestinal epithelial cells