scholarly journals Quantitative Capillary Electrophoresis for Analysis of Extracellular Vesicles (EVqCE)

Separations ◽  
2021 ◽  
Vol 8 (8) ◽  
pp. 110
Author(s):  
Yuchu Dou ◽  
Lixuan Ren ◽  
Prabir Kumar Kulabhusan ◽  
Emil A. Zaripov ◽  
Maxim V. Berezovski
Keyword(s):  
Capillary Electrophoresis ◽  
Nucleic Acids ◽  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Dynamic Range ◽  
Fluorescent Detection ◽  
Average Mass ◽  
Promising Tool ◽  
Aggressive Cancer ◽  
Capillary Zone

Extracellular Vesicles (EVs) gained significant interest within the last decade as a new source of biomarkers for the early detection of diseases and a promising tool for therapeutic applications. In this work, we present Extracellular Vesicles Quantitative Capillary Electrophoresis (EVqCE) to measure an average mass of RNA in EVs, determine EV concentrations and the degree of EV degradation after sample handling. We used EVqCE to analyze EVs isolated from conditioned media of three cancer cell lines. EVqCE employs capillary zone electrophoresis with laser-induced fluorescent detection to separate intact EVs from free nucleic acids. After lysis of EVs with a detergent, the encapsulated nucleic acids are released. Therefore, the initial concentration of intact EVs is calculated based on a nucleic acid peak gain. EVqCE works in a dynamic range of EV concentrations from 108 to 1010 particles/mL. The quantification process can be completed in less than one hour and requires minimum optimization. Furthermore, the average mass of RNA was found to be in the range of 200–400 ag per particle, noting that more aggressive cancer cells have less RNA in EVs (200 ag per particle) than non-aggressive cancer cells (350 ag per particle). EVqCE works well for the degradation analysis of EVs. Sonication for 10 min at 40 kHz caused 85% degradation of EVs, 10 freeze-thaw cycles (from −80 °C to 22 °C) produced 40%, 14-day storage at 4 °C made 32%, and vortexing for 5 min caused 5% degradation. Presently, EVqCE cannot separate and distinguish individual EV populations (exosomes, microvesicles, apoptotic bodies) from each other. Still, it is tolerant to the presence of non-EV particles, protein-lipid complexes, and protein aggregates.

627: Multitarget Gene Inhibition by Synthetic Nucleic Acids in Bladder Cancer Cells

2006 ◽  
Vol 175 (4S) ◽  
pp. 202-202 ◽  
Author(s):  
Yvonne Burmeister ◽  
Kai Kraemer ◽  
Susanne Fuessel ◽  
Matthias Kotzsch ◽  
Axel Meye ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Nucleic Acids ◽  
Cancer Cells ◽  
Bladder Cancer Cells ◽  
Gene Inhibition

scholarly journals An LED-Driven AuNPs-PDMS Microfluidic Chip and Integrated Device for the Detection of Digital Loop-Mediated Isothermal DNA Amplification

Micromachines ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 177 ◽  
Author(s):  
Zengming Zhang ◽  
Shuhao Zhao ◽  
Fei Hu ◽  
Guangpu Yang ◽  
Juan Li ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Microfluidic Chip ◽  
Cross Flow ◽  
Flow Channel ◽  
Isothermal Amplification ◽  
Fluorescent Detection ◽  
Lamp Method ◽  
Biological Studies ◽  
Micro Cavity ◽  
Integrated Device

The sensitive quantification of low-abundance nucleic acids holds importance for a range of clinical applications and biological studies. In this study, we describe a facile microfluidic chip for absolute DNA quantifications based on the digital loop-mediated isothermal amplification (digital LAMP) method. This microfluidic chip integrates a cross-flow channel for droplet generation with a micro-cavity for droplet tiling. DNA templates in the LAMP reagent were divided into ~20,000 water-in-oil droplets at the cross-flow channel. The droplets were then tiled in the micro-cavity for isothermal amplification and fluorescent detection. Different from the existing polydimethylsiloxane (PDMS) microfluidic chips, this study incorporates gold nanoparticles (AuNPs) into PDMS substrate through silica coating and dodecanol modification. The digital LAMP chip prepared by AuNPs-PDMS combines the benefits of the microstructure manufacturing performance of PDMS with the light-to-heat conversion advantages of AuNPs. Upon illumination with a near infrared (NIR) LED, the droplets were stably and efficiently heated by the AuNPs in PDMS. We further introduce an integrated device with a NIR heating unit and a fluorescent detection unit. The system could detect HBV (hepatitis B virus)-DNA at a concentration of 1 × 101 to 1 × 104 copies/μL. The LED-driven digital LAMP chip and the integrated device; therefore, demonstrate high accuracy and excellent performance for the absolute quantification of low-abundance nucleic acids, showing the advantages of integration, miniaturization, cost, and power consumption.

Study of NSCLC cell migration promoted by NSCLC-Derived Extracellular Vesicle using atomic force microscopy

2021 ◽  
Author(s):  
Shuwei Wang ◽  
Jiajia Wang ◽  
Tuoyu Ju ◽  
Kaige Qu ◽  
Fan Yang ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Cell Migration ◽  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Extracellular Vesicle ◽  
Cancer Microenvironment ◽  
Biological Processes ◽  
Force Microscopy ◽  
Atomic Force ◽  
Cellular Components

Extracellular Vesicles (EVs) secreted by cancer cells have a key role in the cancer microenvironment and progression. Previous studies have mainly focused on molecular functions, cellular components and biological processes...

Polymer-Coated Extracellular Vesicles for Selective Codelivery of Chemotherapeutics and siRNA to Cancer Cells

2021 ◽  
Vol 4 (2) ◽  
pp. 1294-1306
Author(s):  
Yong-Yu Jhan ◽  
Guillermo Palou Zuniga ◽  
Kanwar Abhay Singh ◽  
Akhilesh K. Gaharwar ◽  
Daniel L. Alge ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Extracellular Vesicles

scholarly journals Trastuzumab Modulates the Protein Cargo of Extracellular Vesicles Released by ERBB2+ Breast Cancer Cells

Membranes ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 199
Author(s):  
Silvia Marconi ◽  
Sara Santamaria ◽  
Martina Bartolucci ◽  
Sara Stigliani ◽  
Cinzia Aiello ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Breast Cancer Cells ◽  
Culture Supernatant ◽  
High Resolution Mass Spectrometry ◽  
Recombinant Antibody ◽  
Target Cells ◽  
Cellular Processes ◽  
Erbb2 Signaling

Cancers overexpressing the ERBB2 oncogene are aggressive and associated with a poor prognosis. Trastuzumab is an ERBB2 specific recombinant antibody employed for the treatment of these diseases since it blocks ERBB2 signaling causing growth arrest and survival inhibition. While the effects of Trastuzumab on ERBB2 cancer cells are well known, those on the extracellular vesicles (EVs) released from these cells are scarce. This study focused on ERBB2+ breast cancer cells and aimed to establish what type of EVs they release and whether Trastuzumab affects their morphology and molecular composition. To these aims, we performed immunoelectron microscopy, immunoblot, and high-resolution mass spectrometry analyses on EVs purified by differential centrifugation of culture supernatant. Here, we show that EVs released from ERBB2+ breast cancer cells are polymorphic in size and appearance and that ERBB2 is preferentially associated with large (120 nm) EVs. Moreover, we report that Trastuzumab (Tz) induces the expression of a specific glycosylated 50 kDa isoform of the CD63 tetraspanin and modulates the expression of 51 EVs proteins, including TOP1. Because these proteins are functionally associated with organelle organization, cytokinesis, and response to lipids, we suggest that Tz may influence these cellular processes in target cells at distant sites via modified EVs.

Cancer stem cell marker DCLK1 reprograms small extracellular vesicles toward migratory phenotype in gastric cancer cells

PROTEOMICS ◽  
2021 ◽  
pp. 2000098
Author(s):  
Annalisa L.E. Carli ◽  
Shoukat Afshar‐Sterle ◽  
Alin Rai ◽  
Haoyun Fang ◽  
Ryan O'Keefe ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Stem Cell ◽  
Cancer Stem Cell ◽  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Stem Cell Marker ◽  
Cancer Stem Cell Marker ◽  
Cell Marker ◽  
Gastric Cancer Cells ◽  
Migratory Phenotype

scholarly journals An Electrochemical Strategy for the Simultaneous Detection of Doxorubicin and Simvastatin for Their Potential Use in the Treatment of Cancer

Biosensors ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 15
Author(s):  
Iulia Rus ◽  
Mihaela Tertiș ◽  
Cristina Barbălată ◽  
Alina Porfire ◽  
Ioan Tomuță ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Electrolyte Solution ◽  
Electrode Material ◽  
Dynamic Range ◽  
Limit Of Detection ◽  
Simultaneous Detection ◽  
Linear Sweep Voltammetry ◽  
Pharmaceutical Formulations ◽  
Promising Tool ◽  
Scan Rate

The aim of this study was to develop a disposable, simple, fast, and sensitive sensor for the simultaneous electrochemical detection of doxorubicin (DOX) and simvastatin (SMV), which could be used in preclinical studies for the development of new pharmaceutical formulations for drug delivery. Firstly, the electrochemical behavior of each molecule was analyzed regarding the influence of electrode material, electrolyte solution, and scan rate. After this, the proper electrode material, electrolyte solution, and scan rate for both active substances were chosen, and a linear sweep voltammetry procedure was optimized for simultaneous detection. Two chronoamperometry procedures were tested, one for the detection of DOX in the presence of SMV, and the other one for the detection of DOX and SMV together. Finally, calibration curves for DOX and SMV in the presence of each other were obtained using both electrochemical methods and the results were compared. The use of amperometry allowed for a better limit of detection (DOX: 0.1 μg/mL; SMV: 0.7 μg/mL) than the one obtained in voltammetry (1.5 μg/mL for both drugs). The limits of quantification using amperometry were 0.5 μg/mL for DOX (dynamic range: 0.5–65 μg/mL) and 2 μg/mL for SMV (dynamic range: 2–65 μg/mL), while using voltammetry 1 μg/mL was obtained for DOX (dynamic range: 1–100 μg/mL) and 5 μg/mL for SMV (dynamic range: 5–100 μg/mL). This detection strategy represents a promising tool for the analysis of new pharmaceutical formulations for targeted drug delivery containing both drugs, whose association was proven to bring benefits in the treatment of cancer.

scholarly journals Ionizing radiation affects the composition of the proteome of extracellular vesicles released by head-and-neck cancer cells in vitro

2019 ◽  
Vol 60 (3) ◽  
pp. 289-297 ◽  
Author(s):  
Agata Abramowicz ◽  
Anna Wojakowska ◽  
Lukasz Marczak ◽  
Malgorzata Lysek-Gladysinska ◽  
Mateusz Smolarz ◽  
...  
Keyword(s):  
Head And Neck Cancer ◽  
Ionizing Radiation ◽  
Head And Neck ◽  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Neck Cancer

scholarly journals Snail Overexpression Alters the microRNA Content of Extracellular Vesicles Released from HT29 Colorectal Cancer Cells and Activates Pro-Inflammatory State In Vivo

Cancers ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 172
Author(s):  
Izabela Papiewska-Pająk ◽  
Patrycja Przygodzka ◽  
Damian Krzyżanowski ◽  
Kamila Soboska ◽  
Izabela Szulc-Kiełbik ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Extracellular Vesicles ◽  
Cancer Progression ◽  
Epithelial Mesenchymal Transition ◽  
Mesenchymal Transition ◽  
Colorectal Cancer Cells ◽  
Microrna Profile ◽  
Inflammatory State ◽  
Metastatic Process

During metastasis, cancer cells undergo phenotype changes in the epithelial-mesenchymal transition (EMT) process. Extracellular vesicles (EVs) released by cancer cells are the mediators of intercellular communication and play a role in metastatic process. Knowledge of factors that influence the modifications of the pre-metastatic niche for the migrating carcinoma cells is important for prevention of metastasis. We focus here on how cancer progression is affected by EVs released from either epithelial-like HT29-cells or from cells that are in early EMT stage triggered by Snail transcription factor (HT29-Snail). We found that EVs released from HT29-Snail, as compared to HT29-pcDNA cells, have a different microRNA profile. We observed the presence of interstitial pneumonias in the lungs of mice injected with HT29-Snail cells and the percent of mice with lung inflammation was higher after injection of HT29-Snail-EVs. Incorporation of EVs released from HT29-pcDNA, but not released from HT29-Snail, leads to the increased secretion of IL-8 from macrophages. We conclude that Snail modifications of CRC cells towards more invasive phenotype also alter the microRNA cargo of released EVs. The content of cell-released EVs may serve as a biomarker that denotes the stage of CRC and EVs-specific microRNAs may be a target to prevent cancer progression.

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