SFMetrics: An analysis tool for scanning force microscopy images of biomolecules

Scanning force microscopy (SFM) allows direct, rapid and high-resolution visualization of single molecular complexes; irregular shapes and differences in sizes are immediately revealed by the scanning tip in three-dimensional images. However, high-throughput analysis of SFM data is limited by the lack of versatile software tools accessible to SFM users. Most existing SFM software tools are aimed at broad general use: from material-surface analysis to visualization of biomolecules. Results: We present SFMetrics as a metrology toolbox for SFM, specifically aimed at biomolecules like DNA and proteins, which features (a) semi-automatic high-throughput analysis of individual molecules; (b) ease of use working within MATLAB environment or as a stand-alone application; (c) compatibility with MultiMode (Bruker), NanoWizard (JPK instruments), Asylum (Asylum research), ASCII, and TIFF files, that can be adjusted with minor modifications to other formats. Conclusion: Assembled in a single user interface, SFMetrics serves as a semi-automatic analysis tool capable of measuring several geometrical properties (length, volume and angles) from DNA and protein complexes, but is also applicable to other samples with irregular shapes. &Copy; Malm et al.; licensee BioMed Central

Comparative Study of Insertion of Light Elements N, O in Ternary Compounds Ti3AlN and Ti3Al(O)

International audienceThe region close to Ti3AlX (X= N, O) composition of the Ti-Al-O and Ti-Al-N phase diagrams has been experimentally addressed. Alloys with the compositions TiJAIN and Ti~AIO have been processed, Phase analysis and microstructural characterization have been carried out. Both alloys are found to be multi-phased. Under-stoechiornetric Ti3A10X and Ti3AlNX are found to be hexagonal and tetragonal (perovskite-like structure), respectively. Additional information concerning the crystallographic site of insertion of nitrogen and oxygen has been obtained by Electron Energy Loss Spectrometry (EELS). In the Ti-Al-N perovskite-like compound, with an average composition of Ti3AlNo,45.the tetragonal structure stems from the long range nitrogen-vacancy ordering

Le récepteur CCK2 dans les cancers : ciblage diagnostique et thérapeutique grâce à la vectorisation de nanoparticules magnétiques

Le récepteur CCK2 est un récepteur à 7 domaines transmembranaires couplé aux protéines G. D'un point de vue physiologique, le récepteur CCK2 joue un rôle central dans la régulation de la digestion, et agit également au niveau du système nerveux central. D'un point de vue pathologique, il est impliqué dans la carcinogenèse digestive, et il est surexprimé dans les tumeurs neuroendocrines. Il constitue donc une cible diagnostique et thérapeutique potentielle pour ces cancers. Dans un premier temps, nous avons recherché l'existence de variants de ce récepteur dans les tissus tumoraux issus de patients atteints de tumeurs qui expriment le RCCK2. Cette étude a permis de mettre en évidence un nouveau variant d'épissage, délété de l'exon 2, qui code pour une protéine possédant 5 domaines transmembranaires. Dans la cellule, ce variant est localisé uniquement dans le réticulum endoplasmique. Lorsqu'il est co-transfecté avec le récepteur sauvage, il exerce un rôle de dominant négatif sur l'expression membranaire du RCCK2, en entraînant sa rétention dans le réticulum endoplasmique. Ce phénomène se traduit par une diminution de l'activité biologique du récepteur sauvage. Dans un deuxième temps, nous avons développé un outil pour cibler les tumeurs qui surexpriment le récepteur CCK2, basé sur l'utilisation de nanoparticules magnétiques qui sont utilisées comme agent de contraste en IRM. Pour cela nous avons greffé le ligand gastrine du RCCK2 à la surface des nanoparticules. Les nanoparticules vectorisées s'accumulent dans les cellules de façon dépendante du récepteur. L'internalisation des nanoparticules requière l'intervention de la beta-arrestine 2, de la clathrine et de la dynamine. Nous avons montré que la présence de la nanoparticule ne modifiait pas les mécanismes d'internalisation du ligand dans les cellules, en revanche elle modifie la cinétique le recyclage lent (minoritaire) du RCCK2. L'accumulation tumorale des nanoparticules vectorisées a également été analysée in vivo suite à leur injection chez des souris nudes transplantées avec des xénogreffes de cellules tumorales qui expriment le RCCK2. Enfin, nous avons contribué à montrer qu'un autre récepteur à 7 domaines transmembranaires, celui du GIP (glucose-dependent insulinotropic polypeptide), est surexprimé dans certains types de tumeurs neuroendocrines. En outre ce récepteur est surexprimé dans 90% des tumeurs endocrines qui échappent au diagnostic par la somatostatine radiomarquée. Ces résultats posent la question du rôle du récepteur GIP dans le contexte tumoral et en font une nouvelle cible potentielle en cancérologie.The CCK2 receptor belongs to the family of seven transmembrane domain G protein coupled receptors. From a physiological point of view, the CCK2 receptor exerts a central role in digestion regulation, and also acts on the central nervous system. From a pathological point of view, it was reported to be involved in digestive cancer development and overexpressed in neuroendocrine tumors. CCK2R is a potential diagnostic and therapeutic target of these cancers. Firstly, we searched for receptor variants in tumors overexpressing CCK2R. We discovered a new splice variant of the CCK2R deleted of exon 2 and coding for a putative five-transmembrane domain receptor. Ectopic expression cells revealed that this variant lacks biological activity due to its sequestration in the endoplasmic reticulum. When co-expressed with the intact CCK2R, this variant diminished membrane density of the CCK2R and CCK2R-mediated activity, acting as a dominant negative on membrane density of the wild-type receptor. Secondly, we developed CCK2R positive neuroendocrine tumor targeting with magnetic nanoparticles. We grafted a synthetic replicate of the CCK2R ligand, gastrin, on the nanoparticles. Targeted nanoparticles uptake is receptor dependant, and requires involvement of beta-arrestine 2, clathrine and dynamine. We demonstrated that the nanoparticle did not modify ligand internalization in cells, but changes the kinetic of CCK2R intracellular trafic. Tumor accumulation of the targeted nanoparticles was assessed in vivo in mouse bearing tumor xenografts overexpressing CCK2R. Finally, we collaborated on a project demonstrating that another G protein coupled receptor, the GIP receptor (glucose-dependent insulinotropic polypeptide) was overexpressed in neuroendocrine tumors with a high density and incidence. Interestingly this receptor was detected in most somatostatin receptor-negative tumors. These results underlined a likely role of GIPR in tumoral carcinogenesis, and potential target for clinical applications in particular for in vivo scintigraphy and targeted radiotherapy

Targeting the survival kinase DYRK1B: A novel approach to overcome radiotherapy-related treatment resistance

BACKGROUND Cancer cell survival under stress conditions is a prerequisite for the development of treatment resistance. The survival kinase DYRK1B is a key regulator of stress survival pathways and might thereby also contribute to radiation resistance. Here we investigate the strategy of targeting DYRK1B in combination with ionizing radiation (IR) to enhance tumor cell killing under stress conditions. METHODS DYRK1B expression, ROS formation and DNA damage were investigated under serum-starvation (0.1% FBS), hypoxia (0.2%, 1% O$_{2}$) and IR. The combined treatment modality of IR and DYRK1B inhibition was investigated in 2D and in spheroids derived from the colorectal cancer cell line SW620, and in primary patient-derived colorectal carcinoma (CRC) organoids. RESULTS Expression of DYRK1B was upregulated under starvation and hypoxia, but not in response to IR. The small molecule DYRK1B inhibitor AZ191 and shRNA-mediated DYRK1B knockdown significantly reduced proliferative activity and clonogenicity of SW620 tumor cells alone and in combination with IR under serum-starved conditions, which correlated with increased ROS levels and DNA damage. Furthermore, AZ191 successfully targeted the hypoxic core of tumor spheroids while IR preferentially targeted normoxic cells in the rim of the spheroids. A combined treatment effect was also observed in CRC-organoids but not in healthy tissue-derived organoids. CONCLUSION Combined treatment with the DYRK1B inhibitor AZ191 and IR resulted in (supra-) additive tumor cell killing in colorectal tumor cell systems and in primary CRC organoids. Mechanistic investigations support the rational to target the stress-enhanced survival kinase DYRK1B in combination with irradiation to overcome hypoxia- and starvation-induced treatment resistances

Structural and torsional properties of the RAD51-dsDNA nucleoprotein filament

Human RAD51 is a key protein in the repair of DNA by homologous recombination. Its assembly onto DNA, which induces changes in DNA structure, results in the formation of a nucleoprotein filament that forms the basis of strand exchange. Here, we determine the structural and mechanical properties of RAD51-dsDNA filaments. Our measurements use two recently developed magnetic tweezers assays, freely orbiting magnetic tweezers and magnetic torque tweezers, designed to measure the twist and torque of individual molecules. By directly monitoring changes in DNA twist on RAD51 binding, we determine the unwinding angle per RAD51 monomer to be 45°, in quantitative agreement with that of its bacterial homolog, RecA. Measurements of the torque that is built up when RAD51-dsDNA filaments are twisted show that under conditions that suppress ATP hydrolysis the torsional persistence length of the RAD51-dsDNA filament exceeds that of its RecA counterpart by a factor of three. Examination of the filament's torsional stiffness for different combinations of divalent ions and nucleotide cofactors reveals that the Ca2+ ion, apart from suppressing ATPase activity, plays a key role in increasing the torsional stiffness of the filament. These quantitative measurements of RAD51-imposed DNA distortions and accumulated mechanical stress suggest a finely tuned interplay between chemical and mechanical interactions within the RAD51 nucleoprotein filament

Chromosomal transformation in Bacillus subtilis is a non-polar recombination reaction

Natural chromosomal transformation is one of the primary driving forces of bacterial evolution. This reaction involves the recombination of the internalized linear single-stranded (ss) DNA with the homologous resident duplex via RecA-mediated integration in concert with SsbA and DprA or RecO. We show that sequence divergence prevents Bacillus subtilis chromosomal transformation in a log-linear fashion, but it exerts a minor effect when the divergence is localized at a discrete end. In the nucleotide bound form, RecA shows no apparent preference to initiate recombination at the 3′- or 5′-complementary end of the linear duplex with circular ssDNA, but nucleotide hydrolysis is required when heterology is present at both ends. RecA·dATP initiates pairing of the linear 5′ and 3′ complementary ends, but only initiation at the 5′-end remains stably paired in the absence of SsbA. Our results suggest that during gene transfer RecA·ATP, in concert with SsbA and DprA or RecO, shows a moderate preference for the 3′-end of the duplex. We show that RecA-mediated recombination initiated at the 3′- or 5′-complementary end might have significant implication on the ecological diversification of bacterial species with natural transformation

End of life care in nursing homes in Spain: exploratory analysis and evidences of validity of a new scale.

This document is the accepted manuscript version of the following article: Maria Remedios Sánchez-GarcÍa, Jose Antonio Gutiérrez-Romero, Manuel Fernández Alcántara, César Hueso-Montoro, Claire Goodman, and Rafael Montoya-Juárez, ‘End of life care in nursing homes in Spain: Exploratory analysis and evidences of validity of a new scale’, Applied Nursing Research, Vol. 37: 6-12, October 2017. Under embargo until 5 July 2018. The final, definite version is available online at DOI: https://doi.org/10.1016/j.apnr.2017.07.001.Quality end-of-life care is a central issue in nursing homes, requiring the assessment of individual and family needs by health professionals. Although previous instruments have been developed, they usually rely on family reports and have been adapted from other clinical contexts (hospital or primary care). It is important to consider how health care professionals working in nursing homes perceive what is necessary to achieve quality end-of-life care. In this study, the objective was to develop an instrument to assess quality of end-of-life care in the context of Spanish care homes. A 24 item scale Nursing Home End of Life Care Scale (NHEOLC) was developed through a systematic evaluation of existing tools combined with an iterative process of consultation with group experts in end of life care in long term care settings. A total of 307 health care professionals agreed to participate in the study and completed the scale. The scale was grouped in six dimensions: physical, psychological aspects and spiritual aspects of care, family care, bereavement, and patient/family preferences management. The results suggest an adequate factorial structure of the scale and good internal consistency for the total score and the subscales. In addition, the results showed significant differences depending on the size of the nursing home, the category of health professionals, and their own perceptions of his work regarding end-of-life care.Peer reviewe

Nanocrystalline cellulose reinforced poly(ethylene oxide) electrolytes for lithium-metal batteries with excellent cycling stability

Polyethylene oxide (PEO) based polymer electrolytes are still the state of the art for commercial lithium-metal batteries (LMBs) despite their remaining challenges such as the limited ionic conductivity at ambient temperature. Accordingly, the realization of thin electrolyte membranes and, thus, higher conductance is even more important, but this requires a sufficiently high mechanical strength. Herein, the incorporation of nanocrystalline cellulose into PEO-based electrolyte membranes is investigated with a specific focus on the electrochemical properties and the compatibility with lithium-metal and LiFePO$_4$-based electrodes. The excellent cycling stability of symmetric Li||Li cells, including the complete stripping of lithium from one electrode to the other, and Li||LiFePO$_4$ cells renders this approach very promising for eventually yielding thin high-performance electrolyte membranes for LMBs

Self-textured ZnO via AACVD of alkyl alkoxides: a solution-based seed-less route towards optoelectronic-grade coatings

Carbon-free, crystalline and transparent (002)-oriented ZnO films with thickness below 200 nm were deposited at 350 °C on plain glass via AACVD. ZnO films restricted to PVD-growth are achievable through a fast, cost-effective and scalable methodology

Abordaje diagnóstico del gato con enfermedad del tracto urinario

ABORDAJE DIAGNÓSTICO DEL GATO CON ENFERMEDAD DEL TRACTO URINARIOLas enfermedades del tracto urinario caudal de los gatos (ETUCG) son un grupo de patologías que afectan a la vejiga y/o uretra, manifestándose por la presencia de signos clínicos del tracto urinario caudal; estas se pueden mostrar con cuadros obstructivos o no obstructivos. Su abordaje diagnóstico implica un proceso sistemático basado en los hallazgos de los estudios epidemiológicos