Raman Imaging Providing Insights into Chemical Composition of Lipid Droplets of Different Size and Origin: In Hepatocytes and Endothelium

Abstract

In this work, 3D linear Raman spectroscopy was used to study lipid droplets (LDs) ex vivo in liver tissue and also in vitro in a single endothelial cell. Spectroscopic measurements combined with fluorescence microscopy and/or histochemical staining gave complex chemical information about LD composition and enabled detailed investigations of the changes occurring in various pathological states. Lipid analysis in fatty liver tissue was performed using a dietary mouse model of liver steatosis, induced by a high fat diet (HFD). HFD is characterized by a high percentage of calories from saturated fat (60%) and reflects closely the detrimental effects of dietary habits responsible for increased morbidity due to obesity and its complications in well-developed Western societies. Such diets lead to obesity, hyperlipidemia, insulin resistance, and steatosis that may also be linked to endothelial dysfunction. In the present work, Raman spectroscopy was applied to characterized chemical composition of lipid droplets in hepatocytes from mice fed HFD and in the endothelium treated with exogenous unsaturated free fatty acid (arachidonic acid). The results demonstrate the usefulness of Raman spectroscopy to characterize intracellular lipid distribution in 2D and 3D images and can be used to determine the degree of saturation. Raman spectroscopy shows the potential to be a valuable tool for studying the role of LDs in physiology and pathology. The method is generally applicable for the determination of LDs of different size, origin, and composition. Moreover, for the first time, the process of LD formation in the endothelium was detected and visualized in 3D.

Morphology and composition of oxidized InAs nanowires studied by combined Raman spectroscopy and transmission electron microscopy

“…Investigating the fundamental oxidation process of the nanowires is therefore highly relevant. To this end, locally accelerating the reaction by focused laser irradiation has been demonstrated and can be combined with micro-Raman spectroscopy to provide information about the vibrational modes of individual semiconductor nanowires [8][9][10][11] and also conveying information about the elemental composition [12,13], crystal properties [14], and defects [15] as well as free carrier densities and mobilities [16]. Also, at high intensities, focused laser irradiation has been shown to enable controlled local cutting or chemical modification of InAs and GaAs nanowires by local oxidation and formation of crystalline arsenic [17][18][19][20][21] and enables welding of metal nanowires [22].…”

Spectroscopic studies of anthracyclines: Structural characterization and in vitro tracking

“…Raman spectroscopy coupled with confocal microscopy has a great potential for biomedical and biopharmaceutical applications due to ability of noninvasive, label-free detection of cellular changes at single cell level and imaging biomolecule distribution in an individual cell [41,42]. The high information content of Raman spectra can be used to characterize drug-cell interactions [43,44] and therefore serve as a suitable tool for drug screening.…”

Complementary analysis of tissue homogenates composition obtained by Vis and NIR laser excitations and Raman spectroscopy

“…785 cm À1 confirming the assignment of the 565 and 1093 cm À1 bands to nucleic acid. The 785 cm À1 band of weak intensity was found in Raman maps of the nucleus of endothelial cells irradiated by a 488 nm excitation line but not in liver tissue imaged by using the 532 nm laser [29]. This band is easily recognized in NIR Raman spectra of brain tissue, see Table 4.…”

“…Additionally, since Raman features depend on the wavelength of the laser excitation as well as the contribution of resonance Raman (RR) effect of chromophoric components, we recorded Raman spectra of the tissues with the use of the most common laser lines in the radiation range from 488 to 785 nm. Although most Raman studies on animal and human tissues have been mainly carried out using the NIR laser excitation (785 nm) due to the RR effect of heme-based metalloproteins and fluorescence background [3,4,6,11,12,18], some investigations have been also performed with an excitation in the Vis region of radiation [13,15,21,[26][27][28][29]. It is possible since recent technological developments allow for tight laser focusing (creating a small voxel), and consequently eliminating the contribution of fluorescent components to Raman spectrum registered from a larger volume of the laser spot [8,13,15,21].…”