scholarly journals Biological Atomic Force Microscopy for Imaging Gold-Labeled Liposomes on Human Coronary Artery Endothelial Cells

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Ana-María Zaske ◽  
Delia Danila ◽  
Michael C. Queen ◽  
Eva Golunski ◽  
Jodie L. Conyers
Keyword(s):  
Atomic Force Microscopy ◽  
Endothelial Cells ◽  
Coronary Artery ◽  
Cell Membrane ◽  
Fluorescent Microscopy ◽  
Human Coronary Artery ◽  
Force Microscopy ◽  
Cell Internalization ◽  
Atomic Force

Although atomic force microscopy (AFM) has been used extensively to characterize cell membrane structure and cellular processes such as endocytosis and exocytosis, the corrugated surface of the cell membrane hinders the visualization of extracellular entities, such as liposomes, that may interact with the cell. To overcome this barrier, we used 90 nm nanogold particles to label FITC liposomes and monitor their endocytosis on human coronary artery endothelial cells (HCAECs) in vitro. We were able to study the internalization process of gold-coupled liposomes on endothelial cells, by using AFM. We found that the gold-liposomes attached to the HCAEC cell membrane during the first 15–30 min of incubation, liposome cell internalization occurred from 30 to 60 min, and most of the gold-labeled liposomes had invaginated after 2 hr of incubation. Liposomal uptake took place most commonly at the periphery of the nuclear zone. Dynasore monohydrate, an inhibitor of endocytosis, obstructed the internalization of the gold-liposomes. This study showed the versatility of the AFM technique, combined with fluorescent microscopy, for investigating liposome uptake by endothelial cells. The 90 nm colloidal gold nanoparticles proved to be a noninvasive contrast agent that efficiently improves AFM imaging during the investigation of biological nanoprocesses.

scholarly journals Cell penetration efficiency analysis of different atomic force microscopy nanoneedles into living cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marcos Penedo ◽  
Tetsuya Shirokawa ◽  
Mohammad Shahidul Alam ◽  
Keisuke Miyazawa ◽  
Takehiko Ichikawa ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Cell Membrane ◽  
Cell Biology ◽  
Cell Damage ◽  
Efficiency Analysis ◽  
Living Cells ◽  
Biomolecular Recognition ◽  
Cell Penetration ◽  
Force Microscopy ◽  
Atomic Force

AbstractOver the last decade, nanoneedle-based systems have demonstrated to be extremely useful in cell biology. They can be used as nanotools for drug delivery, biosensing or biomolecular recognition inside cells; or they can be employed to select and sort in parallel a large number of living cells. When using these nanoprobes, the most important requirement is to minimize the cell damage, reducing the forces and indentation lengths needed to penetrate the cell membrane. This is normally achieved by reducing the diameter of the nanoneedles. However, several studies have shown that nanoneedles with a flat tip display lower penetration forces and indentation lengths. In this work, we have tested different nanoneedle shapes and diameters to reduce the force and the indentation length needed to penetrate the cell membrane, demonstrating that ultra-thin and sharp nanoprobes can further reduce them, consequently minimizing the cell damage.

Quantification of fenestrations in liver sinusoidal endothelial cells by atomic force microscopy

Micron ◽  
2017 ◽  
Vol 101 ◽  
pp. 48-53 ◽  
Author(s):  
Bartlomiej Zapotoczny ◽  
Karolina Szafranska ◽  
Edyta Kus ◽  
Stefan Chlopicki ◽  
Marek Szymonski
Keyword(s):  
Atomic Force Microscopy ◽  
Endothelial Cells ◽  
Liver Sinusoidal Endothelial Cells ◽  
Sinusoidal Endothelial Cells ◽  
Force Microscopy ◽  
Atomic Force

scholarly journals Regulation of capillary tubules and lipid formation in vascular endothelial cells and macrophages via extracellular vesicle-mediated microRNA-4306 transfer

2018 ◽  
Vol 47 (1) ◽  
pp. 453-469 ◽  
Author(s):  
Ying Yang ◽  
Hui Luo ◽  
Can Zhou ◽  
Rongyi Zhang ◽  
Si Liu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Vascular Endothelial Cells ◽  
Density Lipoprotein ◽  
Extracellular Vesicle ◽  
Luciferase Reporter ◽  
Vascular Endothelial ◽  
Human Coronary Artery ◽  
Lipid Formation

Objective This study aimed to examine regulation of capillary tubules and lipid formation in vascular endothelial cells and macrophages via extracellular vesicle-mediated microRNA (miRNA)-4306 transfer Methods Whole blood samples (12 mL) were collected from 53 patients, and miR-4306 levels in extracellular vesicles (EVs) were analyzed by reverse transcription-polymerase chain reaction. Human coronary artery vascular endothelial cells (HCAECs) and human monocyte-derived macrophages (HMDMs) were transfected with a scrambled oligonucleotide, an miR-4306 mimic, or an anti-miR-4306 inhibitor. The direct effect of miR-4306 on the target gene was analyzed by a dual-luciferase reporter assay. Results EV-contained miR-4306 released from HMDMs was significantly upregulated in coronary artery disease. Oxidized low-density lipoprotein (ox-LDL)-stimulated HMDM-derived EVs inhibited proliferation, migration, and angiogenesis abilities of HCAECs in vitro. However, ox-LDL-stimulated HCAEC-derived EVs enhanced lipid formation of HMDMs. The possible mechanism of these findings was partly due to EV-mediated miR-4306 upregulation of the Akt/nuclear factor kappa B signaling pathway. Conclusions Paracrine cellular crosstalk between HCAECs and HMDMs probably supports the pro-atherosclerotic effects of EVs under ox-LDL stress.

scholarly journals Visualization by atomic force microscopy of tobacco mosaic virus movement protein–RNA complexes formed in vitro

2001 ◽  
Vol 82 (6) ◽  
pp. 1503-1508 ◽  
Author(s):  
O. I. Kiselyova ◽  
I. V. Yaminsky ◽  
E. M. Karger ◽  
O. Yu. Frolova ◽  
Y. L. Dorokhov ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Movement Protein ◽  
Structural Conversion ◽  
Force Microscopy ◽  
Rna Transcripts ◽  
Atomic Force ◽  
Rna Complexes

The structure of complexes formed in vitro by tobacco mosaic virus (TMV)-coded movement protein (MP) with TMV RNA and short (890 nt) synthetic RNA transcripts was visualized by atomic force microscopy on a mica surface. MP molecules were found to be distributed along the chain of RNA and the structure of MP–RNA complexes depended on the molar MP:RNA ratios at which the complexes were formed. A rise in the molar MP:TMV RNA ratio from 20:1 to 60–100:1 resulted in an increase in the density of the MP packaging on TMV RNA and structural conversion of complexes from RNase-sensitive ‘beads-on-a-string’ into a ‘thick string’ form that was partly resistant to RNase. The ‘thick string’-type RNase-resistant complexes were also produced by short synthetic RNA transcripts at different MP:RNA ratios. The ‘thick string’ complexes are suggested to represent clusters of MP molecules cooperatively bound to discrete regions of TMV RNA and separated by protein-free RNA segments.

Abstract P211: N-Acetyl-Seryl-Aspartyl-Lysyl-Proline (Ac-SDKP) Promotes Tube Formation in Human Coronary Artery Endothelial Cells

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Ginette Bordcoch ◽  
Pablo Nakagawa ◽  
Cesar A Romero ◽  
Oscar A Romero
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Capillary Tube ◽  
Tube Formation ◽  
Endothelial Cell Migration ◽  
Tube Length ◽  
Human Coronary Artery ◽  
Double Phase ◽  
Angiogenic Effect

Ac-SDKP is an endogenous peptide with anti-inflammation and anti-fibrotic effects in hypertensive and cardiovascular diseases. It is cleaved from Thymosin β4 (Tβ4) and hydrolyzed by angiotensin converting enzyme (ACE). Ac-SDKP plasma concentration increases after treatment with ACE inhibitors (ACEi) and some of the beneficial effects of ACEi treatment has been ascribed to Ac-SDKP. Ac-SDKP is a mediator of angiogenesis in in-vitro and in-vivo animal models. Ac-SDKP stimulates rodents derived immortalized aortic endothelial cells migration and capillary-like structures formation (tube formation). Similarly, Ac-SDKP increases capillary density after myocardial infarction in rats. The mechanism related to angiogenesis induced by Ac-SDKP is not known. Tβ4 (Ac-SDKP precursor) promotes endothelial cell migration and angiogenesis by the activation of the VEGF/AKT pathway. Our objective is to evaluate the Ac-SDKP pro-angiogenic effect in Human Coronary Artery Endothelial Cells (HCAEC) and the mechanism that regulates the angiogenic effect of Ac-SDKP. HCAEC do not produce VEGF, thus we hypothesize that Ac-SDKP increases VEGF expression in fibroblasts and that indirectly could promote capillary tube formation in endothelial cells. We used primary culture of rat cardiac fibroblast (RCF) and we treated these cells with 10nM Ac-SDKP for 24 hours. VEGF concentration in cell supernatant was measured by ELISA. Cells were starved without serum overnight before the Ac-SDKP treatment. For capillary tube formation assay, HCAEC cells were seeded into matrigel and incubated in presence of 10nM Ac-SDKP for 12 hours, pictures were taken by double phase contrast microscope and tube length was quantified with image J software and the results were expressed as percentage of control. After Ac-SDKP treatment, VEGF concentration did not increase in the supernatant of RCF (control: 0.12±0.07 vs. Ac-SDKP: 0.14±0.09 mg/ml; p=0.7). However, Ac-SDKP treatment induced the development of tube formation in HCAECs by 7±2% respect to control (p=0.037). We conclude that Ac-SDKP induces capillary tube formation not only in rodent but also in human derived endothelial cells. The mechanism by which Ac-SDKP promotes tube formation in HCAECs is still unknown.

Sign in / Sign up

Export Citation Format

Share Document