MADNet: microarray database network web server

MADNet is a user-friendly data mining and visualization tool for rapid analysis of diverse high-throughput biological data such as microarray, phage display or even metagenome experiments. It presents biological information in the context of metabolic and signalling pathways, transcription factors and drug targets through minimal user input, consisting only of the file with the experimental data. These data are integrated with information stored in various biological databases such as NCBI nucleotide and protein databases, metabolic and signalling pathway databases (KEGG), transcription regulation (TRANSFAC©) and drug target database (DrugBank). MADNet is freely available for academic use at http://www.bioinfo.hr/madnet

ELF5 modulates the estrogen receptor cistrome in breast cancer.

Acquired resistance to endocrine therapy is responsible for half of the therapeutic failures in the treatment of breast cancer. Recent findings have implicated increased expression of the ETS transcription factor ELF5 as a potential modulator of estrogen action and driver of endocrine resistance, and here we provide the first insight into the mechanisms by which ELF5 modulates estrogen sensitivity. Using chromatin immunoprecipitation sequencing we found that ELF5 binding overlapped with FOXA1 and ER at super enhancers, enhancers and promoters, and when elevated, caused FOXA1 and ER to bind to new regions of the genome, in a pattern that replicated the alterations to the ER/FOXA1 cistrome caused by the acquisition of resistance to endocrine therapy. RNA sequencing demonstrated that these changes altered estrogen-driven patterns of gene expression, the expression of ER transcription-complex members, and 6 genes known to be involved in driving the acquisition of endocrine resistance. Using rapid immunoprecipitation mass spectrometry of endogenous proteins, and proximity ligation assays, we found that ELF5 interacted physically with members of the ER transcription complex, such as DNA-PKcs. We found 2 cases of endocrine-resistant brain metastases where ELF5 levels were greatly increased and ELF5 patterns of gene expression were enriched, compared to the matched primary tumour. Thus ELF5 alters ER-driven gene expression by modulating the ER/FOXA1 cistrome, by interacting with it, and by modulating the expression of members of the ER transcriptional complex, providing multiple mechanisms by which ELF5 can drive endocrine resistance

Correlation between radiological assessment of acute ankle fractures and syndesmotic injury on MRI

Item does not contain fulltextOBJECTIVE: Owing to the shortcomings of clinical examination and radiographs, injury to the syndesmotic ligaments is often misdiagnosed. When there is no indication requiring that the fractured ankle be operated on, the syndesmosis is not tested intra-operatively, and rupture of this ligamentous complex may be missed. Subsequently the patient is not treated properly leading to chronic complaints such as instability, pain, and swelling. We evaluated three fracture classification methods and radiographic measurements with respect to syndesmotic injury. MATERIALS AND METHODS: Prospectively the radiographs of 51 consecutive ankle fractures were classified according to Weber, AO-M�ller, and Lauge-Hansen. Both the fracture type and additional measurements of the tibiofibular clear space (TFCS), tibiofibular overlap (TFO), medial clear space (MCS), and superior clear space (SCS) were used to assess syndesmotic injury. MRI, as standard of reference, was performed to evaluate the integrity of the distal tibiofibular syndesmosis. The sensitivity and specificity for detection of syndesmotic injury with radiography were compared to MRI. RESULTS: The Weber and AO-M�ller fracture classification system, in combination with additional measurements, detected syndesmotic injury with a sensitivity of 47\% and a specificity of 100\%, and Lauge-Hansen with both a sensitivity and a specificity of 92\%. TFCS and TFO did not correlate with syndesmotic injury, and a widened MCS did not correlate with deltoid ligament injury. CONCLUSION: Syndesmotic injury as predicted by the Lauge-Hansen fracture classification correlated well with MRI findings. With MRI the extent of syndesmotic injury and therefore fracture stage can be assessed more accurately compared to radiographs

Cryopreservation of human cancers conserves tumour heterogeneity for single-cell multi-omics analysis

Background: High throughput single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for exploring cellular heterogeneity among complex human cancers. scRNA-Seq studies using fresh human surgical tissue are logistically difficult, preclude histopathological triage of samples, and limit the ability to perform batch processing. This hindrance can often introduce technical biases when integrating patient datasets and increase experimental costs. Although tissue preservation methods have been previously explored to address such issues, it is yet to be examined on complex human tissues, such as solid cancers and on high throughput scRNA-Seq platforms. Methods: Using the Chromium 10X platform, we sequenced a total of ~ 120,000 cells from fresh and cryopreserved replicates across three primary breast cancers, two primary prostate cancers and a cutaneous melanoma. We performed detailed analyses between cells from each condition to assess the effects of cryopreservation on cellular heterogeneity, cell quality, clustering and the identification of gene ontologies. In addition, we performed single-cell immunophenotyping using CITE-Seq on a single breast cancer sample cryopreserved as solid tissue fragments. Results: Tumour heterogeneity identified from fresh tissues was largely conserved in cryopreserved replicates. We show that sequencing of single cells prepared from cryopreserved tissue fragments or from cryopreserved cell suspensions is comparable to sequenced cells prepared from fresh tissue, with cryopreserved cell suspensions displaying higher correlations with fresh tissue in gene expression. We showed that cryopreservation had minimal impacts on the results of downstream analyses such as biological pathway enrichment. For some tumours, cryopreservation modestly increased cell stress signatures compared to freshly analysed tissue. Further, we demonstrate the advantage of cryopreserving whole-cells for detecting cell-surface proteins using CITE-Seq, which is impossible using other preservation methods such as single nuclei-sequencing. Conclusions: We show that the viable cryopreservation of human cancers provides high-quality single-cells for multiomics analysis. Our study guides new experimental designs for tissue biobanking for future clinical single-cell RNA sequencing studies

Tibiofibular syndesmosis in acute ankle fractures: additional value of an oblique MR image plane

Item does not contain fulltextOBJECTIVE: To evaluate the additional value of a 45� oblique MRI scan plane for assessing the anterior and posterior distal tibiofibular syndesmotic ligaments in patients with an acute ankle fracture. MATERIALS AND METHODS: Prospectively, data were collected for 44 consecutive patients with an acute ankle fracture who underwent a radiograph (AP, lateral, and mortise view) as well as an MRI in both the standard three orthogonal planes and in an additional 45� oblique plane. The fractures on the radiographs were classified according to Lauge-Hansen (LH). The anterior (ATIFL) and posterior (PTIFL) distal tibiofibular ligaments, as well as the presence of a bony avulsion in both the axial and oblique planes was evaluated on MRI. MRI findings regarding syndesmotic injury in the axial and oblique planes were compared to syndesmotic injury predicted by LH. Kappa and the agreement score were calculated to determine the interobserver agreement. The Wilcoxon signed rank test and McNemar's test were used to compare the two scan planes. RESULTS: The interobserver agreement (?) and agreement score [AS (\%)] regarding injury of the ATIFL and PTIFL and the presence of a fibular or tibial avulsion fracture were good to excellent in both the axial and oblique image planes (? 0.61-0.92, AS 84-95\%). For both ligaments the oblique image plane indicated significantly less injury than the axial plane (p?<?0.001). There was no significant difference in detection of an avulsion fracture in the axial or oblique plane, neither anteriorly (p?=?0.50) nor posteriorly (p?=?1.00). With syndesmotic injury as predicted by LH as comparison, the specificity in the oblique MR plane increased for both anterior (to 86\% from 7\%) and posterior (to 86\% from 48\%) syndesmotic injury when compared to the axial plane. CONCLUSION: Our results show the additional value of an 45� oblique MR image plane for detection of injury of the anterior and posterior distal tibiofibular syndesmoses in acute ankle fractures. Findings of syndesmotic injury in the oblique MRI plane were closer to the diagnosis as assumed by the Lauge-Hansen classification than in the axial plane. With more accurate information, the surgeon can better decide when to stabilize syndesmotic injury in acute ankle fractures

MicroRNA profiling of diverse endothelial cell types

<p>Abstract</p> <p>Background</p> <p>MicroRNAs are ~22-nt long regulatory RNAs that serve as critical modulators of post-transcriptional gene regulation. The diversity of miRNAs in endothelial cells (ECs) and the relationship of this diversity to epithelial and hematologic cells is unknown. We investigated the baseline miRNA signature of human ECs cultured from the aorta (HAEC), coronary artery (HCEC), umbilical vein (HUVEC), pulmonary artery (HPAEC), pulmonary microvasculature (HPMVEC), dermal microvasculature (HDMVEC), and brain microvasculature (HBMVEC) to understand the diversity of miRNA expression in ECs.</p> <p>Results</p> <p>We identified 166 expressed miRNAs, of which 3 miRNAs (miR-99b, miR-20b and let-7b) differed significantly between EC types and predicted EC clustering. We confirmed the significance of these miRNAs by RT-PCR analysis and in a second data set by Sylamer analysis. We found wide diversity of miRNAs between endothelial, epithelial and hematologic cells with 99 miRNAs shared across cell types and 31 miRNAs unique to ECs. We show polycistronic miRNA chromosomal clusters have common expression levels within a given cell type.</p> <p>Conclusions</p> <p>EC miRNA expression levels are generally consistent across EC types. Three microRNAs were variable within the dataset indicating potential regulatory changes that could impact on EC phenotypic differences. MiRNA expression in endothelial, epithelial and hematologic cells differentiate these cell types. This data establishes a valuable resource characterizing the diverse miRNA signature of ECs.</p

Droplet-based single cell RNAseq tools: A practical guide

© 2019 The Royal Society of Chemistry. Droplet based scRNA-seq systems such as Drop-seq, inDrop and Chromium 10X have been the catalyst for the wide adoption of high-throughput scRNA-seq technologies in the research laboratory. In order to understand the capabilities of these systems to deeply interrogate biology; here we provide a practical guide through all the steps involved in a typical scRNA-seq experiment. Through comparing and contrasting these three main droplet based systems (and their derivatives), we provide an overview of all critical considerations in obtaining high quality and biologically relevant data. We also discuss the limitations of these systems and how they fit into the emerging field of Genomic Cytometry

Presentació del Servei de Biblioteques als usuaris del Programa ARGÓ

Presentació realitzada per Rafa Giráldez als alumnes del programa ARGÓ el 26 de juny de 2015 a la Sala d'Actes del Rectora