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@ARTICLE{LeBihan:141262,
author = {Le Bihan, Olivier and Decossas, Marion and Gontier, Etienne
and Gerbod-Giannone, Marie-Christine and Lambert, Olivier},
title = {{V}isualization of adherent cell monolayers by
cryo-electron microscopy: {A} snapshot of endothelial
adherens junctions.},
journal = {Journal of structural biology},
volume = {192},
number = {3},
issn = {1047-8477},
address = {San Diego, Calif.},
publisher = {Elsevier},
reportid = {DZNE-2020-07584},
pages = {470-477},
year = {2015},
abstract = {Cryo-electron microscopy (cryo-EM) allows the visualization
of the cell architecture in its native state. We developed a
robust solution to adapt cryo-electron microscopy of
vitreous sections (CEMOVIS) to a monolayer of adherent cells
using a functionalized polyacrylamide hydrogel growing
substrate. We applied this method to reconstitute an
endothelial cell monolayer to visualize the morphology of
adherens junctions (AJs) which regulate permeability and
integrity of the vascular barrier. The fine morphology and
ultrastructure of AJs from cultured primary human coronary
artery endothelial cells (HCAECs) were analyzed in their
native state by using CEMOVIS. Doxycycline and
sphingosine-1-phosphate (S1P) are known as efficient
regulators of endothelial permeability. Doxycycline and S1P
treatments both led to a drastic morphological switch from
very uneven to standardized 14-17 nm wide AJs over several
microns indicative of a better membrane tethering.
Repetitive structures were occasionally noticed within the
AJ cleft reflecting a local improved structural organization
of VE-cadherin molecules. The ultrastructural stabilization
of AJs observed upon treatment likely indicates a better
adhesion and thus provides structural clues on the mechanism
by which these treatments improve the endothelial barrier
function. This method was also successfully extended to a
thick epithelial barrier model. We expect our strategy to
extend the reliable application of CEMOVIS to virtually any
adherent cultured cell systems.},
keywords = {Adherens Junctions: physiology / Antigens, CD: metabolism /
Caco-2 Cells / Cadherins: metabolism / Cell Line, Tumor /
Coronary Vessels: cytology / Cryoelectron Microscopy:
methods / Doxycycline: pharmacology / Endothelial Cells:
cytology / Humans / Lysophospholipids: pharmacology /
Sphingosine: analogs $\&$ derivatives / Sphingosine:
pharmacology / Staining and Labeling / Antigens, CD (NLM
Chemicals) / Cadherins (NLM Chemicals) / Lysophospholipids
(NLM Chemicals) / cadherin 5 (NLM Chemicals) / sphingosine
1-phosphate (NLM Chemicals) / Doxycycline (NLM Chemicals) /
Sphingosine (NLM Chemicals)},
cin = {AG Alamoudi},
ddc = {540},
cid = {I:(DE-2719)1013012},
pnm = {341 - Molecular Signaling (POF3-341)},
pid = {G:(DE-HGF)POF3-341},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:26470813},
doi = {10.1016/j.jsb.2015.10.009},
url = {https://pub.dzne.de/record/141262},
}