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Fluorescent lifetime imaging microscopy using Europium complexes improves atherosclerotic plaques discrimination

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Abstract

The objective of this study is to characterize arterial tissue with and without atherosclerosis by fluorescence lifetime imaging microscopy (FLIM) using Europium Chlortetracycline complex (EuCTc) as fluorescent marker. For this study, twelve rabbits were randomly divided into a control group (CG) and an experimental group (EG), where they were fed a normal and hypercholesterolemic diet, respectively, and were treated for 60 days. Cryosections of the aortic arch specimens were cut in a vertical plane, mounted on glass slides, and stained with Europium (Eu), Chlortetracycline (CTc), Europium Chlortetracycline (EuCTc), and Europium Chlortetracycline Magnesium (EuCTcMg) solutions. FLIM images were obtained with excitation at 405 nm. The average autofluorescence lifetime within plaque depositions was ~1.36 ns. Reduced plaque autofluorescence lifetimes of 0.23 and 0.31 ns were observed on incubation with EuCTc and EuCTcMg respectively. It was observed a quenching of collagen, cholesterol and TG emission spectra increasing EuCTc concentration. The drastic reduction in fluorescence lifetimes is due to a resonant energy transfer between collagen, triglycerides, cholesterol and europium complexes, quenching fluorescence.

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Acknowledgments

The authors acknowledge the National Institute of Science and Technology on Photonics Applied to Cell Biology (INFABIC) at the State University of Campinas for granting access to equipment and for providing assistance in this study. INFABIC is co-funded by Fundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP) (08/57906-3) and Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq) (573913/2008-0), and the FAPESP 2014/06960-9 (Sources of Funding).

Funding

This study was funded by Fapesp (Grant Number 2014/06960-9).

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Authors and Affiliations

  1. Center of Lasers and Applications, Institute of Energy and Nuclear Research, São Paulo University, São Paulo, Brazil

    Letícia Bonfante Sicchieri & Lilia Coronato Courrol

  2. Departments of Pathology, Faculty of Medical Sciences, State University of Campinas, Campinas, São Paulo, Brazil

    Rodrigo de Andrade Natal

  3. Department of Exact and Earth Sciences, Federal University of São Paulo, Diadema, São Paulo, Brazil

    Lilia Coronato Courrol

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  1. Letícia Bonfante Sicchieri
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  3. Lilia Coronato Courrol
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Correspondence to Lilia Coronato Courrol.

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None of the authors have conflict of interest in regards to the content of this manuscript.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. Ethics Committee of UNIFESP (Protocol no. 0327/12).

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Sicchieri, L.B., de Andrade Natal, R. & Courrol, L.C. Fluorescent lifetime imaging microscopy using Europium complexes improves atherosclerotic plaques discrimination. Int J Cardiovasc Imaging 32, 1595–1604 (2016). https://doi.org/10.1007/s10554-016-0936-7

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  • DOI: https://doi.org/10.1007/s10554-016-0936-7

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