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Unsaturated lipid bodies as a hallmark of inflammation studied by Raman 2D and 3D microscopy.

ABSTRACT: Endothelial HMEC-1 cells incubated with pro-inflammatory cytokine TNF-? for 6 and 24?hours were studied as a model of inflammation using Raman imaging. Striking changes in distribution, composition and concentration of cellular lipids were observed after exposure to TNF-? compared to the control. In particular, 3D Raman imaging revealed a significant increase in the amount of lipid entities formed under inflammation. Lipid bodies were randomly distributed in the cytoplasm and two types of droplets were assembled: more saturated one, in spectral characteristics resembling phosphatidylcholine and saturated cholesteryl esters, observed also in the control, and highly unsaturated one, containing also cholesterols, being a hallmark of inflamed cells. The statistical analysis showed that the number of lipid bodies was significantly dependent on the exposure time to TNF-?. Overall, observed formation of unsaturated lipid droplets can be directly correlated with the increase in production of prostacyclins - endogenous inflammation mediators.

SUBMITTER: Czamara K

PROVIDER: S-EPMC5241649 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Unsaturated lipid bodies as a hallmark of inflammation studied by Raman 2D and 3D microscopy.

Czamara K K Majzner K K Selmi A A Baranska M M Ozaki Y Y Kaczor A A

Scientific reports 20170118

Endothelial HMEC-1 cells incubated with pro-inflammatory cytokine TNF-α for 6 and 24 hours were studied as a model of inflammation using Raman imaging. Striking changes in distribution, composition and concentration of cellular lipids were observed after exposure to TNF-α compared to the control. In particular, 3D Raman imaging revealed a significant increase in the amount of lipid entities formed under inflammation. Lipid bodies were randomly distributed in the cytoplasm and two types of drople ...[more]

PMID: 28098251

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Project description:BACKGROUND:Oils and bioproducts extracted from cultivated algae can be used as sustainable feedstock for fuels, nutritional supplements, and other bio-based products. Discovery and isolation of new algal species and their subsequent optimization are needed to achieve economical feasibility for industrial applications. Here we describe and validate a workflow for in situ analysis of algal lipids through confocal Raman microscopy. We demonstrate its effectiveness to characterize lipid content of algal strains isolated from the environment as well as algal cells screened for increased lipid accumulation through UV mutagenesis combined with Fluorescence Activated Cell Sorting (FACS). RESULTS:To establish and validate our workflow, we refined an existing Raman platform to obtain better discrimination in chain length and saturation of lipids through ratiometric analyses of mixed fatty acid lipid standards. Raman experiments were performed using two different excitation lasers (? = 532 and 785 nm), with close agreement observed between values obtained using each laser. Liquid chromatography coupled with mass spectrometry (LC-MS) experiments validated the obtained Raman spectroscopic results. To demonstrate the utility and effectiveness of the improved Raman platform, we carried out bioprospecting for algal species from soil and marine environments in both temperate and subtropical geographies to obtain algal isolates from varied environments. Further, we carried out two rounds of mutagenesis screens on the green algal model species, Chlamydomonas reinhardtii, to obtain cells with increased lipid content. Analyses on both environmental isolates and screened cells were conducted which determined their respective lipids. Different saturation states among the isolates as well as the screened C. reinhardtii strains were observed. The latter indicated the presence of cell-to cell variations among cells grown under identical condition. In contrast, non-mutagenized C. reinhardtii cells showed no significant heterogeneity in lipid content. CONCLUSIONS:We demonstrate the utility of confocal Raman microscopy for lipid analysis on novel aquatic and soil microalgal isolates and for characterization of lipid-expressing cells obtained in a mutagenesis screen. Raman microscopy enables quantitative determination of the unsaturation level and chain lengths of microalgal lipids, which are key parameters in selection and engineering of microalgae for optimal production of biofuels.

Dataset Information

Unsaturated lipid bodies as a hallmark of inflammation studied by Raman 2D and 3D microscopy.

Publications

Unsaturated lipid bodies as a hallmark of inflammation studied by Raman 2D and 3D microscopy.

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