NUCKS overexpression in breast cancer

<p>Abstract</p> <p>Background</p> <p>NUCKS (Nuclear, Casein Kinase and Cyclin-dependent Kinase Substrate) is a nuclear, DNA-binding and highly phosphorylated protein. A number of reports show that <it>NUCKS </it>is highly expressed on the level of mRNA in several human cancers, including breast cancer. In this work, NUCKS expression on both RNA and protein levels was studied in breast tissue biopsies consisted of invasive carcinomas, intraductal proliferative lesions, benign epithelial proliferations and fibroadenomas, as well as in primary cultures derived from the above biopsies. Specifically, in order to evaluate the level of NUCKS protein in correlation with the histopathological features of breast disease, immunohistochemistry was employed on paraffin sections of breast biopsies of the above types. In addition, NUCKS expression was studied by means of Reverse Transcription PCR (RT-PCR), real-time PCR (qRT-PCR) and Western immunoblot analyses in the primary cell cultures developed from the same biopsies.</p> <p>Results</p> <p>The immunohistochemical Results showed intense NUCKS staining mostly in grade I and II breast carcinomas compared to normal tissues. Furthermore, NUCKS was moderate expressed in benign epithelial proliferations, such as adenosis and sclerosing adenosis, and highly expressed in intraductal lesions, specifically in ductal carcinomas <it>in situ </it>(DCIS). It is worth noting that all the fibroadenoma tissues examined were negative for NUCKS staining. RT-PCR and qRT-PCR showed an increase of <it>NUCKS </it>expression in cells derived from primary cultures of proliferative lesions and cancerous tissues compared to the ones derived from normal breast tissues and fibroadenomas. This increase was also confirmed by Western immunoblot analysis. Although NUCKS is a cell cycle related protein, its expression does not correlate with Ki67 expression, neither in tissue sections nor in primary cell cultures.</p> <p>Conclusion</p> <p>The results show overexpression of the NUCKS protein in a number of non malignant breast lesions and cancerous tissues. In particular, the NUCKS overexpression in ADH and DCIS indicates a significant role of this protein in neoplastic progression.</p

Altered expression pattern of integrin alphavbeta3 correlates with actin cytoskeleton in primary cultures of human breast cancer

Background: Integrins are transmembrane adhesion receptors that provide the physical link between the actin cytoskeleton and the extracellular matrix. It has been well established that integrins play a major role in various cancer stages, such as tumor growth, progression, invasion and metastasis. In breast cancer, integrin alphavbeta3 has been associated with high malignant potential in cancer cells, signaling the onset of widespread metastasis. Many preclinical breast cancer studies are based on established cell lines, which may not represent the cell behavior and phenotype of the primary tumor of origin, due to undergone genotypic and phenotypic changes. In the present study, short-term primary breast cancer cell cultures were developed. Integrin alphavbeta3 localization was studied in correlation with F-actin cytoskeleton by means of immunofluorescence and immunogold ultrastructural localization. Integrin fluorescence intensities were semi-quantitatively assessed by means of computerized image analysis, while integrin and actin expression was evaluated by Western immunoblotting. Results: In the primary breast cancer epithelial cells integrin alphavbeta3 immunofluorescence was observed in the marginal cytoplasmic area, whereas in the primary normal breast epithelial cells it was observed in the main cell body, i.e. in the ventrally located perinuclear area. In the former, F-actin cytoskeleton appeared well-formed, consisting of numerous and thicker stress fibers, compared to normal epithelial cells. Furthermore, electron microscopy showed increased integrin alphavbeta3 immunogold localization in epithelial breast cancer cells over the area of stress fibers at the basal cell surface. These findings were verified with Western immunoblotting by the higher expression of integrin beta3 subunit and actin in primary breast cancer cells, revealing their reciprocal relation, in response to the higher motility requirements, determined by the malignant potential of the breast cancer cells. Conclusion: A model system of primary breast cancer cell cultures was developed, in an effort to maintain the closest resembling environment to the tumor of origin. Using the above system model as an experimental tool the study of breast tumor cell behavior is possible concerning the adhesion capacity and the migrating potential of these cells, as defined by the integrin alphavbeta3 distribution in correlation with F-actin cytoskeleton

Priorities for Mediterranean marine turtle conservation and management in the face of climate change

As climate-related impacts threaten marine biodiversity globally, it is important to adjust conservation efforts to mitigate the effects of climate change. Translating scientific knowledge into practical management, however, is often complicated due to resource, economic and policy constraints, generating a knowledge-action gap. To develop potential solutions for marine turtle conservation, we explored the perceptions of key actors across 18 countries in the Mediterranean. These actors evaluated their perceived relative importance of 19 adaptation and mitigation measures that could safeguard marine turtles from climate change. Of importance, despite differences in expertise, experience and focal country, the perceptions of researchers and management practitioners largely converged with respect to prioritizing adaptation and mitigation measures. Climate change was considered to have the greatest impacts on offspring sex ratios and suitable nesting sites. The most viable adaptation/mitigation measures were considered to be reducing other pressures that act in parallel to climate change. Ecological effectiveness represented a key determinant for implementing proposed measures, followed by practical applicability, financial cost, and societal cost. This convergence in opinions across actors likely reflects long-standing initiatives in the Mediterranean region towards supporting knowledge exchange in marine turtle conservation. Our results provide important guidance on how to prioritize measures that incorporate climate change in decision-making processes related to the current and future management and protection of marine turtles at the ocean-basin scale, and could be used to guide decisions in other regions globally. Importantly, this study demonstrates a successful example of how interactive processes can be used to fill the knowledge-action gap between research and management.This work was conducted under FutureMares EU project that received funding from the European Union's Horizon 2020 research and innovation programme under grant agreement No 869300. The Mediterranean Marine Turtle Working Group was established in 2017 and is continuously supported by MedPAN and the National Marine Park of Zakynthos. The work of AC was supported by the Hellenic Foundation for Research and Innovation (H.F.R.I.) under the “First Call for H.F.R.I. Research Projects to support Faculty members and Researchers and the procurement of high-cost research equipment grant” (Project Number: 2340).Peer reviewe

Isolation and H-Binding Study of Humic Substances from Greek Soil and Lignite: NICA-Donnan Model Parameters

Humic acids from soil and lignite, from different locations of Greece were isolated according to the International Humic Substances Society (IHSS) isolation protocol. An analytical proton binding study of these humic acids using NICA model gives us a wide range of parameters compared to the parameter values of IHSS reference samples

Modelling the humification process and the humic acid molecule

The structural origin of the properties of humic acids (HA) is of great importance. Natural humic acids (HA) consist of several major functional groups, predominantly carboxylic (R-COOH) and phenolic (R-OH), as well as carbonyl (C=O) and quinoid. Carboxylic and phenolic functionalities determine the H-binding, metal binding and charge properties of HA. In addition, all HA of diverse geographic and climatic origins contain stable organic radicals which are considered to play a key-role on the transformation of humic acid itself. Presently, the oxidative polymerization of polyphenols in soils is thought to be among the major processes of formation of natural humic substances. Given the acknowledged structural complexity and heterogeneity of natural HA, an approach adopted by various groups was the production of ‘‘synthetic HA”. Recently we have presented a method for performing a Humic Acid Like Polymer (HALP) at significant yields with no use of a catalyst. Herein we present a detailed study off the mechanism of the polymerization in conjunction with S.E.C. data of the evolution of the molecular size of the HALP. Finally the relation of the structures with physicochemical properties is discussed

A Water Soluble Polymer as a Working Structural Model for Humic Acids: H-binding and Spectroscopic Properties

HA occur ubiquitously in terrestrial and aquatic environments, indicating that many precursors and several formation pathways exist. Distinguishing among these pathways has been difficult because individual scientists have tended to concentrate their efforts on specific types of samples (e.g. soil, sediment or water) for which different isolation or fractionation methods are often used. Humic acids ultimately result from abiotic and biotic polymerization and polycondensation reactions involving such small precursors. The relative importance of abiotic and biotic processes in humification in soils and sediments still remains obscure even though much research has been devoted to them. For example the oxidative polymerization of phenols, which is an important precursor reaction in the formation of HA, is catalysed both by enzymes and abiotic (inorganic) catalysts. Polyphenols have been shown to be important precursors in the abiotic formation of HA. More particularly, the oxidative polymerization of polyphenols in soils is thought to be one of the major processes of formation of humic substances. According to the polyphenolic theory, humic acids are formed from simple phenols and phenolic acids via the formation of a semiquinone radical. Coupling of semiquinones originating from diphenols can lead to the formation of stable humic polymers under the catalytic action of Mn (IV) oxides. In the present case, we have focused our work on molecules bearing both carboxy and phenolic moieties as well as radical activity. In this model, protocatechuic acid (3,4-dihydroxybenzoic acid), and gallic acid (3,4,5-trihydroxybenzoic acid) are among the main low molecular weight aromatic acids formed after lignin degradation. As we have reported recently polymerization of the mono-, di-, and trihydroxyphenolic compounds may occur in alkaline media i.e. under ambient O2. i.e. under conditions where radicals are formed. In the present work, we have investigated the oxidative co-polymerization of some simple hydrobenzoic molecules which have been shown to be good models certain physicochemical properties of humic acids. As we show by controlling the pH, redox potential and the ratio of the polyphenolics, a water soluble polymer is produced that mimics certain key- physicochemical and spectroscopic properties of humic acids

Soil-foundation interaction of wind turbine towers on monopiles

Μη διαθέσιμη περίληψηNot available summarizationΠαρουσιάστηκε στο: The Seventh International Conference on Engineering Computational Technolog

Humic Acid: Aggregate, Floc, Macromolecule, Micelle, Polymer, or Supramolecule?

Two humic acids, extracted from an Alfisol forest soil and a lignite mined in Greece, were purified of ash components, size fractionated by membrane dialysis, and polarity fractionated by adsorption chromatography on XAD-8 resin to determine binding mechanisms of humic acid constituents. Humic acids, contained in 12,000 to 15,000 dialysis membranes, were successively dialyzed against 0.1 M HCl, 0.2M HF, 0.1M sodium citrate at pH 10, and 0.1 M NaOH to remove ash components and disaggregate and size fractionate with increasing pH. A part of humic acid did not permeate the dialysis membranes during the acid treatment, while the humic acid that permeated during the sodium citrate and NaOH treatments, was polarity fractionated by a decreasing pH gradient on XAD-8 resin. The resulting fractions were characterized by mass balance, elemental analyses, pH titration, and FT-IR spectrometry. Certain fractions of the lignite humic acid had characteristics very similar to molecular condensates formed by radical coupling of phenolic acid precursors of lignin. Significant differences between the composition and fractionation patterns of the soil and lignite humic acids were interpreted in terms of binding mechanisms within the humic acid precipitates

Probing Local pH in Hydrophobic Domains of Humic Acids: An EPR Based Method

Despite the focused efforts by many labs during the last 30 years, vital aspects of the structure and fundamental physicochemical aspects of Humic Acids (HA) are largely unknown. It is currently accepted that humic acids contain both hydrophilic as well as hydrophobic domains which are the locus of sequestration of xenobiotic molecules. However vital information on the detailed nature, the accessibility of the hydrophobic domains is lacking. On the other hand, HA behave as polyelectrolytes which bear variable charge modulated by pH. It is considered that this pH-dependent charge is modulated by the local pKa of the chargeable structural groups i.e. carboxylates and phenolics. Traditionally only the water accessible hydrophilic fraction of the chargeable groups can be probed by H-binding or metal binding studies. However the function of chargeable groups in hydrophobic domains, local pKa, and their geometrical location are largely unknown. In this context we have developed a novel methodology by using the Spin-Probe method with Electron Paramagnetic Resonance (EPR) spectroscopy. A series of specific spin-probes with varying hydrophobicity as well as pH sensitivity were used and studied with EPR spectroscopy. The experiments were carried out in well characterized HA samples of varying molecular weight providing information of the local environment of the spin-probes, their mobility, and local pKa values

A novel bentonite humic acid composite material BephosTM as an environmental-friendly tool in phosphate and ammonium management

The objective of the present study was to examine the feasibility of using a novel modified bentonite as adsorbent for phosphate and ammonium removal from natural eutrophic waters. The uptake of phosphate and ammonium in adsorption kinetics A critical parameter was the initial phosphate concentration aiming to represent a eutrophic natural ecosystem. Moreover, its adsorption capacity and efficiency as a restoration tool was compared with a number of materials tested on the removal of phosphate and ammonium from aqueous solutions. Bephos™ is capable for simultaneous adsorption of both phosphorus and ammonia and this is an important advantage against other materials used to restore eutrophic water bodies. Adsorption kinetics showed that more than 90% and 70% removal of phosphate and ammonium respectively from water within 30 min. Bephos™ is potent for remediation of phosphate and ammonium at low concentrations that occur in natural water ecosystems