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Uptake of fatty acids by a single endothelial cell investigated by Raman spectroscopy supported by AFM

Abstract

Raman spectroscopy is an attractive tool in identification cells, especially when studying cellular processes, including lipid droplets (LDs) formation. In this work, confocal Raman imaging was used to study the formation of LDs in vitro in a single endothelial cell upon incubation with polyunsaturated fatty acids (10 or 25 µM) including arachidonic acid (AA) and its deuterated analog (AA-d8), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Based on Raman spectra taken from a single endothelial cells, it was possible to investigate biochemical changes induced by addition of polyunsaturated fatty acids. In particular, content of lipids in formed LDs as well as unsaturation degree was identified by Raman spectroscopy by marker bands at 1660 cm−1 due to C=C stretching and ~3015 cm−1 due to the stretching mode of =C–H associated with C=C double bonds (except for a deuterated form where these bands are shifted respectively). To establish if exogenous fatty acid was taken up by the cell and stored in LDs, a deuterium labelled polyunsaturated fatty acids was used. AA-d8 shows characteristic bands around 2200–2300 cm−1 assigned to =C-D stretching modes. We established uptake of AA and accumulation of EPA into newly formed LD’s in the endothelial cells. In contrast, no accumulation of DHA in LD’s was observed even though LD’s were formed upon DHA incubation. Furthermore, using AFM we demonstrated that the presence of LDs in the endothelium affected endothelial stiffness which could have pathophysiological significance. In summary, the results suggest that formation of LDs in endothelium involves exogenous and endogenous polyunsaturated fatty acids, and their relative contribution to LD’s formation seem distinct for AA, EPA and DHA.

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Supplementary files

Publication details

The article was received on 22 Jun 2017, accepted on 21 Dec 2017 and first published on 04 Jan 2018


Article type: Paper
DOI: 10.1039/C7AN01043E
Citation: Analyst, 2018, Accepted Manuscript
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    Uptake of fatty acids by a single endothelial cell investigated by Raman spectroscopy supported by AFM

    K. Majzner, S. Tott, L. Roussille, V. Deckert, S. Chlopicki and M. Baranska, Analyst, 2018, Accepted Manuscript , DOI: 10.1039/C7AN01043E

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