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Fluorescent methods for the detection and investigation of low density lipoprotein low affinity binding in live cells

Keywords:

M.N. Balatskaya – Junior Research Scientist, Faculty of Medicine, Moscow State University. E-mail: m.balatskaya@gmail.com
G.V. Sharonov – Ph.D. (Phys.-Math.), Senior Research Scientist, Faculty of Medicine, Moscow State University. E-mail: sharonov@gmail.com
N.N. Mamedov – Post-gradant Student, Faculty of Medicine, Lomonosov Moscow State University. E-mail: nick_el@mail.ru
Y.P. Rubtsov – Ph.D. (Chem.), Assistant Professor, Faculty of Medicine, Lomonosov Moscow State University. E-mail: yrubtsov@gmail.com


Elevated level of low-density lipoprotein (LDL) is the main factor in the development of atherosclerosis in humans. However the relationship between the level of lipoproteins and cellular processes that lead to the formation of atherosclerotic plaques is still not clear. T-cadherin (T-cad) is one of the receptors that is responsible for the intracellular signaling in response to LDL and its gene polymorphisms correlate with metabolic syndrome and hypertension. Therefore, the determination of T-cad binding to LDL is not only of academic interest, but may be clinically significant. For observation of the localization of T-cad in live cells, we have created a fluorescent protein chimera of T- cadherin. By using this chimera, we found that besides the plasma membrane T-cadherin is concentrated in the endoplasmic reticulum (ER). These data together with the literature speak about possible involvement of T-cad in ER stress. Created T-cadherin fluorescent chimera is a unique tool for the detection of cellular and molecular mechanisms of LDL action in live cells. We have developed a method for detecting LDL binding on the surface of live cells by flow cytometry. It is shown that the method in addition to high affinity (2 ug/ml) binding to apoB/E receptor is also possible to determine the low affinity (40 ug/ml) binding of T-cad.
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May 29, 2020

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