Effect of laminin, polylysine and cell medium components on the attachment of human hepatocellular carcinoma cells to cellulose nanofibrils analyzed by surface plasmon resonance

The development of in vitro cell models that mimic cell behavior in organs and tissues is an approach that may have remarkable impact on drug testing and tissue engineering applications in the future. Plant based, chemically unmodified cellulose nanofibrils (CNF) hydrogel is a natural, abundant, and biocompatible material that has attracted great attention for biomedical applications, in particular for threedimensional cell cultures. However, the mechanisms of cell-CNF interactions and factors that affect these interactions are not yet fully understood. In this work, multi-parametric surface plasmon resonance (SPR) was used to study how the adsorption of human hepatocellular carcinoma (HepG2) cells on CNF films is affected by the different proteins and components of the cell medium. Both human recombinant laminin 521 (LN-521, a natural protein of the extracellular matrix) and poly -L-lysine (PLL) adsorbed on CNF films and enhanced the attachment of HepG2 cells. Cell medium components (glucose and amino acids) and serum proteins (fetal bovine serum, FBS) also adsorbed on both bare CNF and on protein-coated CNF substrates. However, the adsorption of FBS hindered the attachment of HepG2 cells to LN-521and PLLcoated CNF substrates, suggesting that serum proteins blocked the formation of laminin-integrin bonds and decreased favorable PLL-cell electrostatic interactions. This work sheds light on the effect of different factors on cell attachment to CNF, paving the way for the utilization and optimization of CNF-based materials for different tissue engineering applications. (C) 2020 The Authors. Published by Elsevier Inc.Peer reviewe

Quantified forces between HepG2 hepatocarcinoma and WA07 pluripotent stem cells with natural biomaterials correlate with in vitro cell behavior

In vitro cell culture or tissue models that mimic in vivo cellular response have potential in tissue engineering and regenerative medicine, and are a more economical and accurate option for drug toxicity tests than animal experimentation. The design of in vivo-like cell culture models should take into account how the cells interact with the surrounding materials and how these interactions affect the cell behavior. Cell-material interactions are furthermore important in cancer metastasis and tumor progression, so deeper understanding of them can support the development of new cancer treatments. Herein, the colloidal probe microscopy technique was used to quantify the interactions of two cell lines (human pluripotent stem cell line WA07 and human hepatocellular carcinoma cell line HepG2) with natural, xeno-free biomaterials of different chemistry, morphology, and origin. Key components of extracellular matrices -human collagens I and IV, and human recombinant laminin-521-, as well as wood-derived, cellulose nanofibrils -with evidenced potential for 3D cell culture and tissue engineering- were analysed. Both strength of adhesion and force curve profiles depended on biomaterial nature and cell characteristics. The successful growth of the cells on a particular biomaterial required cell-biomaterial adhesion energies above 0.23 nJ/m. The information obtained in this work supports the development of new materials or hybrid scaffolds with tuned cell adhesion properties for tissue engineering, and provides a better understanding of the interactions of normal and cancerous cells with biomaterials in the human body.Peer reviewe

Quantifying the interactions between biomimetic biomaterials - collagen I, collagen IV, laminin 521 and cellulose nanofibrils - by colloidal probe microscopy

Biomaterials of different nature have been and are widely studied for various biomedical applications. In many cases, biomaterial assemblies are designed to mimic biological systems. Although biomaterials have been thoroughly characterized in many aspects, not much quantitative information on the molecular level interactions between different biomaterials is available. That information is very important, on the one hand, to understand the properties of biological systems and, on the other hand, to develop new composite biomaterials for special applications. This work presents a systematic, quantitative analysis of self- and cross-interactions between films of collagen I (Col I), collagen IV (Col IV), laminin (LN-521), and cellulose nanofibrils (CNF), that is, biomaterials of different nature and structure that either exist in biological systems (e.g., extracellular matrices) or have shown potential for 3D cell culture and tissue engineering. Direct surface forces and adhesion between biomaterials-coated spherical micro-particles and flat substrates were measured in phosphate-buffered saline using an atomic force microscope and the colloidal probe technique. Different methods (Langmuir-Schaefer deposition, spin-coating, or adsorption) were applied to completely coat the flat substrates and the spherical micro particles with homogeneous biomaterial films. The adhesion between biomaterials films increased with the time that the films were kept in contact. The strongest adhesion was observed between Col IV films, and between Col IV and LN-521 films after 30 s contact time. In contrast, low adhesion was measured between CNF films, as well as between CNF and LN-521 films. Nevertheless, a good adhesion between CNF and collagen films (especially Col I) was observed. These results increase our understanding of the structure of biological systems and can support the design of new matrices or scaffolds where different biomaterials are combined for diverse biological or medical applications.Peer reviewe

Synthesis of an Azide- and Tetrazine-Functionalized [60]Fullerene and Its Controlled Decoration with Biomolecules

Bingel cyclopropanation between Buckminster fullerene and a heteroarmed malonate was utilized to produce a hexakis-functionalized C-60 core, with azide and tetrazine units. This orthogonally bifunctional C-60 scaffold can be selectively one-pot functionalized by two pericyclic click reactions, that is, inverse electron-demand Diels-Alder and azide-alkyne cycloaddition, which with appropriate ligands (monosaccharides, a peptide and oligonucleotides tested) allows one to control the assembly of heteroantennary bioconjugates.Peer reviewe

AFM Force Spectroscopy Reveals the Role of Integrins and Their Activation in Cell-Biomaterial Interactions

Transmembrane protein integrins play a key role in cell adhesion. Cell-biomaterial interactions are affected by integrin expression and conformation, which are actively controlled by cells. Although integrin structure and function have been studied in detail, quantitative analyses of integrin-mediated cell-biomaterial interactions are still scarce. Here, we have used atomic force spectroscopy to study how integrin distribution and activation (via intracellular mechanisms in living cells or by divalent cations) affect the interaction of human pluripotent stem cells (WA07) and human hepatocarcinoma cells (HepG2) with promising biomaterials.human recombinant laminin-521 (LN-521) and cellulose nanofibrils (CNF). Cell adhesion to LN-521-coated probes was remarkably influenced by cell viability, divalent cations, and integrin density in WA07 colonies, indicating that specific bonds between LN-521 and activated integrins play a significant role in the interactions between LN-521 and HepG2 and WA07 cells. In contrast, the interactions between CNF and cells were nonspecific and not influenced by cell viability or the presence of divalent cations. These results shed light on the underlying mechanisms of cell adhesion, with direct impact on cell culture and tissue engineering applications.Peer reviewe

Cytokeratin 5 determines maturation of the mammary myoepithelium

At invasion, transformed mammary epithelial cells expand into the stroma through a disrupted myoepithelial (ME) cell layer and basement membrane (BM). The intact ME cell layer has thus been suggested to act as a barrier against invasion. Here, we investigate the mechanisms behind the disruption of ME cell layer. We show that the expression of basal/ME proteins CK5, CK14, and alpha-SMA altered along increasing grade of malignancy, and their loss affected the maintenance of organotypic 3D mammary architecture. Furthermore, our data suggests that loss of CK5 prior to invasive stage causes decreased levels of Zinc finger protein SNAI2 (SLUG), a key regulator of the mammary epithelial cell lineage determination. Consequently, a differentiation bias toward luminal epithelial cell type was detected with loss of mature, alpha-SMA-expressing ME cells and reduced deposition of basement membrane protein laminin-5. Therefore, our data discloses the central role of CK5 in mammary epithelial differentiation and maintenance of normal ME layer.Peer reviewe

Evolución paleoambiental de la mitad sur de la Península Ibérica. Aplicación a la evaluación del comportamiento de los repositorios de residuos radiactivos

Esta publicación refleja los resultados obtenidos de la realización de los proyectos "EQUIP: Evidency from Quaternary Infills Palaeohydrogeology" (F 14W/ CT96/0031), financiado por la UE, "Evolución Paleoclimática de [a Mitad Sur de [a Península Ibérica" financiado por ENRESA y "Paleoclima" financiado por el Consejo de Seguridad Nuclear y ENRESA. La cuenca de Guadix-Baza, sector oriental, es una de las escasas zonas europeas donde se ha conservado un registro razonablemente completo del Pleistoceno. La cuenca de Guadix Baza, en régimen continental desde el Plioceno, funcionó bajo un régimen centrípeto, con abanicos aluviales en los bordes y una zona compleja lacustre en mosaico en su centro. Se ha podido establecer la existencia de una amplia variedad de facies aluviales y fluviales y sus interrelaciones. La sedimentación en el margen lacustre estaba constituida por arenas bioclásticas y lutitas con fósiles, mientras que en zonas más centrales predominaron lutitas yesíferas, lutitas con yeso intrasedimentario, arenas yesíferas yesos y, ocasionalmente, carbonatos que durante un período concreto depositaron los materiales que configuran el "Nivel Calcáreo de Orce", calizas y dolomías con cantidades variables de yeso y terrígenos cerca de los bordes. El trabajo de campo permitió el establecimiento de una serie estratigráfica tipo compuesta, que refleja los principales acontecimiento paleoambientales que tuvieron lugar durante el Pleistoceno. Con el fin de evitar la yesificación de la calcita y aragonito, propia del ambiente lacustre, la serie tipo de ha establecido en el registro del margen lacustre. Con ayuda del análisis paleomagnético y el análisis de la racemización de aminoácidos en conchas de moluscos y ostrácodos se ha establecido la cronología numérica de la sección estratigráfica tipo que cubre desde el límite Plioceno-Pleistoceno hasta unos 250 ka BP, cuando la erosión de la cabecera del río Fardes alcanza la cuenca abriéndola hacia el valle del Guadalquivir y cesando [a sedimentación lacustre. También se ha datado un depósito de terraza fluvial. Se han obtenido datos paleoambientales "instantáneos" mediante el estudio palinológico, el análisis paleobotánico de material silicificado (madera opalizada) o carbonizado (análisis paleoantracológico), el análisis geoquímico orgánico de algunos niveles especialmente favorables. La geoquímica orgánica de biomarcadores de algunos niveles concretos proporcionó datos sobre su origen (plantas terrestres o acuáticas) y confirmó que un conspicuo nivel carbonoso tuvo su origen en un incendio forestal. El análisis paleobiológico basado en la distribución de especies de ostrácodos, gasterópodos y pelecípodos ha permitido una primera aproximación paleoambiental, pese al evidente sesgo introducido por la presencia de fuentes salinas y materiales yesíferos en el área fuente de los abanicos aluviales del límite oriental de la cuenca. Esta interpretación se ha depurado mediante el estudio geoquímico inorgánico, isótopos estables y elementos traza de las conchas calcíticas de ostrácodos. Ello ha permitido el establecimiento de una alternancia de períodos "fríos y húmedos" y "cálidos y áridos" que permiten reconocer de las clases climáticas "mediterráneo seco" y "mediterráneo húmedo" sensu Horowitz (1989), que tienen sus correlatos en el registro paleoclimático de los grandes lagos del rift del Mar Muerto, el Mar Caspio y lagos pluviales de Norte América. La correlación de los datos isotópicos con las paleosalinidades deducidas del estudio de las inclusiones fluidas en yeso intrasedimentario ha permitido corroborar estas interpretaciones. Esto plantea un nuevo enfoque en el análisis del comportamiento de los repositorios de residuos radiactivos de alta

Prevalence of submicroscopic malaria infection in immigrants living in Spain

BACKGROUND: The importance of submicroscopic malaria infections in high-transmission areas could contribute to maintain the parasite cycle. Regarding non-endemic areas, its importance remains barely understood because parasitaemia in these afebrile patients is usually below the detection limits for microscopy, hence molecular techniques are often needed for its diagnosis. In addition to this, the lack of standardized protocols for the screening of submicroscopic malaria in immigrants from endemic areas may underestimate the infection with Plasmodium spp. The aim of this study was to assess the prevalence of submicroscopic malaria in afebrile immigrants living in a non-endemic area. METHODS: A prospective, observational, multicentre study was conducted. Afebrile immigrants were included, microscopic observation of Giemsa-stained thin and thick blood smears, and two different molecular techniques detecting Plasmodium spp. were performed. Patients with submicroscopic malaria were defined as patients with negative blood smears and detection of DNA of Plasmodium spp. with one or both molecular techniques. Demographic, clinical, analytical and microbiological features were recorded and univariate analysis by subgroups was carried out with STATA v15. RESULTS: A total of 244 afebrile immigrants were included in the study. Of them, 14 had a submicroscopic malaria infection, yielding a prevalence of 5.7% (95% confidence interval 3.45-9.40). In 71.4% of the positive PCR/negative microscopy cases, Plasmodium falciparum alone was the main detected species (10 out of the 14 patients) and in 4 cases (28.6%) Plasmodium vivax or Plasmodium ovale were detected. One patient had a mixed infection including three different species. CONCLUSIONS: The prevalence of submicroscopic malaria in afebrile immigrants was similar to that previously described in Spain. Plasmodium vivax and P. ovale were detected in almost a third of the submicroscopic infections. Screening protocols for afebrile immigrants with molecular techniques could be useful for a proper management of these patients.This work was funded by projects PI14/01671, PI17/01791 and PI14CIII/00014, from the Instituto de Salud Carlos III (Ministry of Economy, Industry and Competitiveness) and cofounded by the European Regional Development Fund, and approved by the Ethics Committee of our Institution. There was no funding from the PCR manufacturers; they did not play any role in data analysis or in the reporting of the results.S

Measurements of Higgs boson production and couplings in diboson final states with the ATLAS detector at the LHC

Measurements are presented of production properties and couplings of the recently discovered Higgs boson using the decays into boson pairs, $H\rightarrow\gamma\gamma$, $H\rightarrow ZZ^{*}\rightarrow 4 \ell$ and $H\rightarrow W W \rightarrow \ell\nu\ell\nu$. The results are based on the complete pp collision data sample recorded by the ATLAS experiment at the CERN Large Hadron Collider at centre-of-mass energies of 7 TeV and 8 TeV, corresponding to an integrated luminosity of about 25 fb$^{-1}$. Evidence for Higgs boson production through vector-boson fusion is reported. Results of combined fits probing Higgs boson couplings to fermions and bosons, as well as anomalous contributions to loop-induced production and decay modes, are presented. All measurements are consistent with expectations for the Standard Model Higgs boson.Peer Reviewe

Observation of an Excited Bc±B_c^\pm Meson State with the ATLAS Detector

A search for excited states of the Bc± meson is performed using 4.9  fb-1 of 7 TeV and 19.2  fb-1 of 8 TeV pp collision data collected by the ATLAS experiment at the LHC. A new state is observed through its hadronic transition to the ground state, with the latter detected in the decay Bc±→J/ψπ±. The state appears in the m(Bc±π+π-)-m(Bc±)-2m(π±) mass difference distribution with a significance of 5.2 standard deviations. The mass of the observed state is 6842±4±5  MeV, where the first error is statistical and the second is systematic. The mass and decay of this state are consistent with expectations for the second S-wave state of the Bc± meson, Bc±(2S).Peer Reviewe