1,720,976 research outputs found
Large Population Of Homogeneous Spheroids Models For Testing Toxicity And Diffusion Of Nanomaterials
No full textThe toxicity and the therapeutic effect of drugs need to be assessed on relevant cell models before they are tested on animal models or humans. Analogously, the safety of nanomaterials intended to be used in food should be tested for toxicity on relevant and reliable models. In recent years, the use of 3D cell models has been recognized as a step forward in complexity and relevance in biological cell assays. These models can mimic the complexity and the biological barriers of tissues better than conventional 2D systems.
Nowadays, protocols are available to produce 3D cell models for many cell types. One of the common problems in 3D cell models is the variability in shape and size of the 3D structures, leading to a scarce reliability of the assays. We have developed and improved methods for the culturing of 3D spheroids in microwells arrays that could also mimic the mechanical properties of tissues. Large populations of up to several hundreds of homogeneous 3D tumor spheroids can be produced reliably and their growth and molecular properties can be characterized in situ during and after growth or treatment with drugs or nanostructures. It is possible to characterize the drug distribution in the spheroids in order to gain insight into their penetration properties and persistence.
Recently, we obtained 3D heterotypic cell spheroids where 3D cell structures are made of different cell types so that more complex and reliable tissue models can be obtained. Additionally, it is possible to exploit the 3D bioprinting technology to obtain complex cell mixtures and achieve 3D heterotypic cell models with a range of compositions and treat them with drugs or nanostructures in the same environment. We trust that this technology will prove useful in reliably and quantitatively testing the toxicity and other biological effects of food-relevant nanomaterials as well as other nanostructures
A RAD51/BRCA2 small molecule inhibitor enhances the antineoplastic effect of the PARPi talazoparib in pancreatic cancer
No full textIntroduction: PARP inhibitors (PARPi) are drugs showing efficacy in HR-defective cancers. The RAD51/BRCA2 interaction is a key step in HR, which is the main DNA double strand break (DSB) repair pathway. As this mechanism is activated following single strand break (SSB) repair inhibition by PARPi, the simultaneous impairment of HR and PARP should lead to increased antineoplastic power of PARPi. We characterized the mechanism of action of ARN24922 (1), which is a new promising RAD51/BRCA2 inhibitor, developed in house. Furthermore, we evaluated its potential to increase the effect of talazoparib (TLZ, a potent PARPi) on 2D and 3D models of pancreatic cancer (PAC).
Materials and methods: We evaluated the potential of 1 in inhibiting HR activity through the mClover HR assay, which is based on the insertion of a mClover-containing sequence into a Cas9-generated DSB in the LaminA gene; the subsequent DSB repair leads to cell fluorescence. To confirm HR inhibition, we examined RAD51 nuclear foci after the induction of DNA damage by cisplatin (CPL) and RAD51/BRCA2 inhibition by 1. Finally, we studied the antineoplastic effect of the 1-TLZ combination in different PAC cell lines (BxPC-3, HPAC, AsPC-1) and in 3D models of PAC (PT291 and PDM41 organoids), with each of the models being characterized by functional BRCA genes and HR.
Results and discussion: Preliminarily, by applying the mClover HR assay, we found that 1 produced an 80% HR inhibition. Moreover, 30 μM 1 caused a significant reduction of RAD51 nuclear foci in CPL exposed cells. This finding is in agreement with the results obtained in the HR assay, supporting the expected mechanism of action of 1. In 2D PAC cultures, we found that 50 μM 1 significantly increased the anti-proliferative effect of 2 μM TLZ. The cell death mechanism involved in this effect was studied in BxPC-3 cells and was characterized as apoptosis. When 1 was co-administered with TLZ, in 3D PAC models, a significant increase in the antineoplastic activity of TLZ was also observed. In addition, in PT291 organoid, 1 was found to increase the DNA damage signatures caused by TLZ, as shown by immunofluorescence detection of γ-H2AX in cell nuclei.
Conclusions: The obtained data support the idea that the inhibition of RAD51/BRCA2 interaction can extend the use TLZ to HR proficient tumor forms, highlighting a promising anticancer strategy for PAC
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Epigallocatechin-3-Gallate (EGCG) Mitigates Endothelial and Circulating Cells Alterations Following PLLA Electrospun Mat Placement
Full text linkBackground. Synthetic vascular graft calcification is a serious complication of graft placement. Here, we analysed migration and osteogenic genes of human umbilical vein endothelial cells (HUVEC) cultured with a poly-L-lactic acid (PLLA) electrospun mat. The role of epigallo-catechin-3-gallate (EGCG) in pathogenic processes involving HUVEC and peripheral blood mononuclear cells (PBMCs) was also tested. Methods. HUVEC were cultured in indirect contact with PLLA for 48 h, with or without EGCG, and processed for mRNA expression. HUVEC proliferation, migration and osteogenic differentiation were evaluated after EGCG treatment. EGCG was also administrated to human PBMCs, to analyse proliferation and migration toward HUVEC cultured with PLLA. Results. HUVEC cultured with PLLA exhibited increased expression of SLUG, VIMENTIN, MMP-9 (migration, vascular remodelling) and RUNX-2 (osteogenic transcription factor). EGCG at 25 μM significantly reduced HUVEC migration, osteogenic differentiation, without affecting cell viability, and mitigated PLLA influence on SLUG, MMP-9, VIMENTIN and RUNX-2 expression. EGCG affected PBMC proliferation and migration toward PLLA in a transwell co-culture system with HUVEC. Conclusion. Our study suggests the pro-calcific effect of PLLA, proposing EGCG as an anti-inflammatory modulatory approach. Research efforts need to deepen PLLA-vascular wall interactions for preventing vascular graft failure
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Proteomic profile of BxPC-3 cells after treatment with BRC4
No full textBRCA2 and RAD51 are two proteins that play a central role in homologous recombination (HR) and DNA double strand break (DSB) repair. BRCA2 assists RAD51 fibrillation and defibrillation through binding with its eight BRC repeats, with BRC4 being one of the most efficient and best characterized. RAD51 inactivation by small molecules has been proposed as a strategy to impair BRCA2/RAD51 binding and, ultimately, the HR pathway, with the aim of making cancer cells more sensitive to PARP inhibitors (PARPi). This strategy, which mimics a synthetic lethality (SL) approach, has been successfully performed in vitro by using the myristoylated derivative of BRC4 (myr-BRC4), designed for a more efficient cell entry. The present study applies a method to obtain a proteomic fingerprint after cellular treatment with the myr-BRC4 peptide using a mass spectroscopy (MS) proteomic approach. (Data are available via ProteomeXchange with identifier PXD042696.) We performed a comparative proteomic profiling of the myr-BRC4 treated vs. untreated BxPC-3 pancreatic cancer cells and evaluated the differential expression of proteins. Among the identified proteins, we focused our attention on proteins shared by both the RAD51 and the BRCA2 interactomes, and on those whose reduction showed high statistical significance. Three downregulated proteins were identified (FANCI, FANCD2, and RPA3), and protein downregulation was confirmed through immunoblotting analysis, validating the MS approach. Our results suggest that, being a direct consequence of myr-BRC4 treatment, the detection of FANCD2, FANCI, and RPA3 downregulation could be used as an indicator for monitoring HR impairment
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
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