FT-IR Spectroscopic Imaging of Endothelial Cells Response to Tumor Necrosis Factor-α: To Follow Markers of Inflammation Using Standard and High-Magnification Resolution

Abstract

Two endothelial cell lines were selected as models to investigate an effect of incubation with cytokine tumor necrosis factor type α (TNF-α) using Fourier transform infrared (FT-IR) imaging spectroscopy. Both cell lines are often used in laboratories and are typical lung vascular endothelial cells (HMLVEC) derived from the fusion of umbilical vein endothelial cells with lung adenocarcinoma cells (EA.hy926). This study was focused on alteration of spectral changes accompanying inflammation at the cellular level by applying two resolution systems of FT-IR microscopy. The standard approach, with a pixel size of ca. 5.5 μm, determined the inflammatory state of the whole cell, while a high-magnification resolution (pixel size of ca. 1.1 μm) provided information at the subcellular level. Importantly, the analysis of IR spectra recorded with different modes produced similar results overall and yielded unambiguous classification of inflamed cells. Generally, the most significant changes in the cells under the influence of TNF-α are related with lipids-their composition and concentration; however, segregation of cells into subcellular compartments provided an additional insight into proteins and nucleic acids related events. The observed spectral alterations are specific for the type of endothelial cell line.

Comparison of spectral and spatial denoising techniques in the context of High Definition FT-IR imaging hyperspectral data

“…This stems from the fact that IR imaging is a relatively high SNR technique and unless used in HD-IR mode with a thermal source it provides data of reasonable quality. The emergence of HD-IR has opened the need for efficient denoising algorithms 20 – 22 . The developments in Quantum Cascade Laser Microscopes 23 are enabling Discrete Frequency imaging and that puts even more interest in denoising techniques based on a limited number of frequencies, i.e.…”

Raman imaging highlights biochemical heterogeneity of human eosinophils versus human eosinophilic leukaemia cell line

“…The nano FT‐IR spectra collected from two adjacent control EoL‐1 cells are virtually identical and suggests that the outer membrane is built of proteins with a dominant contribution of β sheets compared to α‐helical, cf. Fig A (Staniszewska et al , ; Wiercigroch et al , ). Changes in relative intensities of the three amide I bands (at 1620, 1634, and 1644/cm) were also found in the crater‐like structure while its centre also contains free and esterified fatty acids, as revealed by bands at 1722 and 1745/cm, respectively (blue point in Fig B).…”

“…Three spectra collected from two adjacent control EoL-1 cells showed the presence of bands at 1634 and 1644/cm corresponding to b-sheet and a-helical conformations, respectively. In turn the 1620/cm absorbance was assigned to extended chains structures (Staniszewska et al, 2014;Wiercigroch et al, 2018).…”

Investigation of molecular mechanisms of experimental compounds in murine models of chronic allergic airways disease using synchrotron Fourier-transform infrared microspectroscopy

“…Using conventional methodologies, inflammation is characterised via histological assessment of immune cells infiltrating into airway tissues, changes to the composition of immune cells in bronchoalveolar lavage fluid, and key changes in inflammatory markers 61 . Several reports have identified inflammation by biochemical changes observed through the S-FTIR spectra 62 – 64 . Based on our data, however, specific peaks characteristic of inflammation were not observed in the acquired S-FTIR spectra.…”