Raman spectroscopy analysis of lipid droplets content, distribution and saturation level in Non‐Alcoholic Fatty Liver Disease in mice

Abstract

Non-Alcoholic Fatty Liver Disease (NAFLD) is a common liver disorder, characterized by an excessive lipids deposition within the hepatic tissue. Due to the lack of clear-cut symptoms and optimal diagnostic method, the actual prevalence of NAFLD and its pathogenesis remains unclear, especially in the early stages of progression. In the presented work confocal Raman microspectroscopy was used to investigate alterations in the chemical composition of the NAFLD-affected liver. We have investigated two NAFLD models, representative for macrovesicular and microvesicular steatosis, induced by High Fat Diet (60 kcal %) and Low Carbohydrate High Protein Diet (LCHP), respectively. In both models we confirmed the development of NAFLD, manifested by the presence of lipid droplets (LDs), but of different sizes. Model of macrovesicular steatosis was characterized by large LDs, whereas in the microvesicular steatosis model small droplets were found. In both models, however, we observed a significant decrease in the degree of unsaturation of lipids, in comparison to the control. In addition, for both models, the impact of medical treatment with selected drugs (perindopril and nicotinic acid, respectively) was tested, indicating a significant influence of medicine not only on the occurrence and size of the droplets, but also on their composition. In both cases the drug treatment resulted in an increase of the degree of unsaturation of lipids forming droplets. Confocal Raman microspectroscopy was proven to be a powerful tool providing detailed insight into selected areas of hepatic tissue, following the NAFLD pathogenesis and diagnostic potential of the applied drugs.

Hepatic Steatosis Accompanies Pulmonary Alveolar Proteinosis

“…The potential lipotoxic pathology was probed using (i) ORO staining for identifying neutral lipid droplets accumulation, (ii) CARS microscopy to provide semiquantitative spatial characterisation of lipid droplets, (iii) SHG to identify any collagen associated with fibrotic changes and (iv) TEM for a detailed intracellular view of lipid droplet distribution. Comparable patterns of ORO staining in wild type mouse liver have been reported previously (8).…”

“…Following GM-CSF ablation, however, the liver neutral lipid content was much more heavily stained with ORO ( Figure 4D) with significant intracellular deposition and with a clear steatosis that shared appearance characteristics of both microvesicular steatosis and the more common macrovesicular steatosis (8).…”

“…It provides a non-invasive, non-destructive and label-free modality to selectively image lipids in intact cells and tissue without the use of complex procedures for sample preparation and histopathological examination. Examples of its use include in differential diagnosis of lung carcinoma (28) and for quantification of hepatic lipid in liver tissue (8). For liver has been shown that CARS can successfully evaluate hepatic microvesicular steatosis, by detection and quantification of lipid droplets, their number and size (29).…”

Demonstration of Carbon Catabolite Repression in Naphthalene Degrading Soil Bacteria via Raman Spectroscopy Based Stable Isotope Probing

“…[13][14][15][16] The acquisition of a biomolecular fingerprint consisting of signatures from DNA, RNA, carbohydrates, lipids and proteins present in cells and tissues at the same time gives the technique an enormous advantage. [17][18][19][20] In recent years, Raman spectroscopy has been used to monitor the uptake and distribution of specific sources into different biomolecules by means of stable isotope labeling. 11 Stable isotope probing (SIP) has become a valuable tool in microbial ecology used to identify the uptake of mostly 13 C, 2 H, 15 N from labeled substrates like methane, naphthalene etc.…”

Vascular diseases investigated ex vivo by using Raman, FT-IR and complementary methods

“…144 The Raman spectra of the lipid fraction contain bands connected 145 mainly to cholesterol esters and cholesterol, observed at 1674 cm À1 146 (C5 5C stretching mode), 1443 cm À1 (C-H bending mode), 1740 cm À1 147 (C5 5O stretching mode), 2885 cm À1 at (C-H stretching mode), and 148 704 cm À1 (vibration modes of steroid rings) [40,41]. Raman confocal 149 mapping additionally provides a unique advantage in obtaining 150 information of the tissue composition directly from a chosen 151 location (i.e.…”