scholarly journals Correlative Fluorescence- and Electron Microscopy of Whole Breast Cancer Cells Reveals Different Distribution of ErbB2 Dependent on Underlying Actin

2020 ◽  
Vol 8 ◽  
Author(s):  
Indra Navina Dahmke ◽  
Patrick Trampert ◽  
Florian Weinberg ◽  
Zahra Mostajeran ◽  
Franziska Lautenschläger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Fluorescence And Electron Microscopy

scholarly journals A novel immuno-gold labeling protocol for nanobody-based detection of HER2 in breast cancer cells using immuno-electron microscopy

2017 ◽  
Vol 199 (1) ◽  
pp. 1-11 ◽  
Author(s):  
M. Kijanka ◽  
E.G. van Donselaar ◽  
W.H. Müller ◽  
B. Dorresteijn ◽  
D. Popov-Čeleketić ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Immuno Electron Microscopy

scholarly journals Liquid-phase electron microscopy of molecular drug response in breast cancer cells reveals irresponsive cell subpopulations related to lack of HER2 homodimers

2017 ◽  
Vol 28 (23) ◽  
pp. 3193-3202 ◽  
Author(s):  
Diana B. Peckys ◽  
Ulrike Korf ◽  
Stefan Wiemann ◽  
Niels de Jonge
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Drug Resistance ◽  
Liquid Phase ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Plasma Membranes ◽  
Drug Binding ◽  
Resting Cells ◽  
Cell Heterogeneity

The development of drug resistance in cancer poses a major clinical problem. An example is human epidermal growth factor receptor 2 (HER2) overexpressing breast cancer often treated with anti-HER2 antibody therapies, such as trastuzumab. Because drug resistance is rooted mainly in tumor cell heterogeneity, we examined the drug effect in different subpopulations of SKBR3 breast cancer cells and compared the results with those of a drug-resistant cell line, HCC1954. Correlative light microscopy and liquid-phase scanning transmission electron microscopy were used to quantitatively analyze HER2 responses upon drug binding, whereby many tens of whole cells were imaged. Trastuzumab was found to selectively cross-link and down-regulate HER2 homodimers from the plasma membranes of bulk cancer cells. In contrast, HER2 resided mainly as monomers in rare subpopulations of resting and cancer stem cells (CSCs), and these monomers were not internalized after drug binding. The HER2 distribution was hardly influenced by trastuzumab for the HCC1954 cells. These findings show that resting cells and CSCs are irresponsive to the drug and thus point toward a molecular explanation behind the origin of drug resistance. This analytical method is broadly applicable to study membrane protein interactions in the intact plasma membrane, while accounting for cell heterogeneity.

scholarly journals EGFR Expression in HER2-Driven Breast Cancer Cells

2020 ◽  
Vol 21 (23) ◽  
pp. 9008
Author(s):  
Florian Weinberg ◽  
Diana B. Peckys ◽  
Niels de Jonge
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Plasma Membrane ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Simultaneous Detection ◽  
Orphan Receptor ◽  
Membrane Expression ◽  
Intact Cells ◽  
Egfr Expression

The epidermal growth factor receptor HER2 is overexpressed in 20% of breast cancer cases. HER2 is an orphan receptor that is activated ligand-independently by homodimerization. In addition, HER2 is able to heterodimerize with EGFR, HER3, and HER4. Heterodimerization has been proposed as a mechanism of resistance to therapy for HER2 overexpressing breast cancer. Here, a method is presented for the simultaneous detection of individual EGFR and HER2 receptors in the plasma membrane of breast cancer cells via specific labeling with quantum dot nanoparticles (QDs). Correlative fluorescence microscopy and liquid phase electron microscopy were used to analyze the plasma membrane expression levels of both receptors in individual intact cells. Fluorescent single-cell analysis of SKBR3 breast cancer cells dual-labeled for EGFR and HER2 revealed a heterogeneous expression for receptors within both the cell population as well as within individual cells. Subsequent electron microscopy of individual cells allowed the determination of individual receptors label distributions. QD-labeled EGFR was observed with a surface density of (0.5–5) × 101 QDs/µm2, whereas labeled HER2 expression was higher ranging from (2–10) × 102 QDs/µm2. Although most SKBR3 cells expressed low levels of EGFR, an enrichment was observed at large plasma membrane protrusions, and amongst a newly discovered cellular subpopulation termed EGFR-enriched cells.

Folate receptor-mediated delivery of 1-MDT-loaded mesoporous silica magnetic nanoparticles to target breast cancer cells

Nanomedicine ◽  
2021 ◽  
Author(s):  
Nastaran Hashemzadeh ◽  
Mitra Dolatkhah ◽  
Ayuob Aghanejad ◽  
Mohammad Barzegar-Jalali ◽  
Yadollah Omidi ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Magnetic Nanoparticles ◽  
Mesoporous Silica ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Near Infrared ◽  
Folate Receptor ◽  
Superparamagnetic Iron Oxide ◽  
Transmission Electron

Aims: The efficiency of mesoporous silica magnetic nanoparticles (MSMNP) as a targeted drug-delivery system was investigated. Methods: The superparamagnetic iron oxide nanoparticles (NP) were synthesized, coated with mesoporous silica and conjugated with polyethylene glycol and methotrexate. Next, 1-methyl D tryptophan was loaded into the prepared nanosystems (NS). They were characterized using transmission electron microscopy, scanning electron microscopy, dynamic light scattering, vibrating sample magnetometer, x-ray powder diffraction, Fourier transform-infrared spectroscopy and the Brunauer–Emmett–Teller method and their biological impacts on breast cancer cells were evaluated. Results: The prepared NSs displayed suitable properties and showed enhanced internalization by folate-receptor-expressing cells, exerting efficient cytotoxicity, which was further enhanced by the near-infrared radiation irradiation. Conclusions: On the basis of our findings, the engineered NS is a promising multifunctional nanomedicine/theranostic for solid tumors.

scholarly journals Local variations of HER2 dimerization in breast cancer cells discovered by correlative fluorescence and liquid electron microscopy

2015 ◽  
Vol 1 (6) ◽  
pp. e1500165 ◽  
Author(s):  
Diana B. Peckys ◽  
Ulrike Korf ◽  
Niels de Jonge
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Fluorescence Microscopy ◽  
Cancer Cells ◽  
Single Molecule ◽  
Breast Cancer Cells ◽  
Pair Correlation ◽  
Resting Cells ◽  
Membrane Topography ◽  
Cancer Aggressiveness

The formation of HER2 homodimers plays an important role in breast cancer aggressiveness and progression; however, little is known about its localization. We have studied the intra- and intercellular variation of HER2 at the single-molecule level in intact SKBR3 breast cancer cells. Whole cells were visualized in hydrated state with correlative fluorescence microscopy and environmental scanning electron microscopy (ESEM). The locations of individual HER2 receptors were detected using an anti-HER2 affibody in combination with a quantum dot (QD), a fluorescent nanoparticle. Fluorescence microscopy revealed considerable differences of HER2 membrane expression between individual cells and between different membrane regions of the same cell (that is, membrane ruffles and flat areas). Subsequent ESEM of the corresponding cellular regions provided images of individually labeled HER2 receptors. The high spatial resolution of 3 nm and the close proximity between the QD and the receptor allowed quantifying the stoichiometry of HER2 complexes, distinguishing between monomers, dimers, and higher-order clusters. Downstream data analysis based on calculating the pair correlation function from receptor positions showed that cellular regions exhibiting membrane ruffles contained a substantial fraction of HER2 in homodimeric state. Larger-order clusters were also present. Membrane areas with homogeneous membrane topography, on the contrary, displayed HER2 in random distribution. Second, HER2 homodimers appeared to be absent from a small subpopulation of cells exhibiting a flat membrane topography, possibly resting cells. Local differences in homodimer presence may point toward functional differences with possible relevance for studying metastasis and drug response.

scholarly journals Correlative Fluorescence- and Electron Microscopy of Whole Breast Cancer Cells Reveals Different Distribution of ErbB2 Dependent on Underlying Actin

2020 ◽  
Author(s):  
Indra Navina Dahmke ◽  
Patrick Trampert ◽  
Florian Weinberg ◽  
Zahra Mostajeran ◽  
Franziska Lautenschläger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Electron Microscopy ◽  
Epidermal Growth Factor Receptor ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Cancer Cells ◽  
Actin Filaments ◽  
Breast Cancer Cells ◽  
Growth Factor Receptor ◽  
Epidermal Growth

AbstractEpidermal growth factor receptor 2 (ErbB2) is found overexpressed in several cancers, such as gastric, and breast cancer, and is, therefore, an important therapeutic target. ErbB2 plays a central role in cancer cell invasiveness, and is associated with cytoskeletal reorganization. In order to study the spatial correlation of single ErbB2 proteins and actin filaments, we applied correlative fluorescence microscopy, and scanning transmission electron microscopy (STEM) to image specifically labeled SKBR3 breast cancer cells. The breast cancer cells were grown on microchips, transformed to express an actin-green fluorescent protein (GFP) fusion protein, and labeled with quantum dot (QD) nanoparticles attached to specific anti-ErbB2 Affibodies. Fluorescence microscopy was performed to identify cellular regions with spatially correlated actin and ErbB2 expression. For STEM of the intact plasma membrane of whole cells, the cells were fixed and covered with graphene. Spatial distribution patterns of ErbB2 in the actin rich ruffled membrane regions were examined, and compared to adjacent actin-low regions of the same cell, revealing an association of putative signaling active ErbB2 homodimers with actin-rich regions. ErbB2 homodimers were found absent from actin-low membrane regions, as well as after treatment of cells with Cytochalasin D, which breaks up larger actin filaments. In both latter data sets, a significant inter-label distance of 36 nm was identified, possibly indicating an indirect attachment to helical actin filaments via the formation of heterodimers of ErbB2 with epidermal growth factor receptor (EGFR). The possible attachment to actin filaments was further explored by identifying linear QD-chains in actin-rich regions, which also showed an inter-label distance of 36 nm.

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