In Vivo Metabolic Fingerprinting of Neutral Lipids with Hyperspectral Stimulated Raman Scattering Microscopy

Abstract

Metabolic fingerprinting provides valuable information on the physiopathological states of cells and tissues. Traditional imaging mass spectrometry and magnetic resonance imaging are unable to probe the spatial-temporal dynamics of metabolites at the subcellular level due to either lack of spatial resolution or inability to perform live cell imaging. Here we report a complementary metabolic imaging technique that is based on hyperspectral stimulated Raman scattering (hsSRS). We demonstrated the use of hsSRS imaging in quantifying two major neutral lipids: cholesteryl ester and triacylglycerol in cells and tissues. Our imaging results revealed previously unknown changes of lipid composition associated with obesity and steatohepatitis. We further used stable-isotope labeling to trace the metabolic dynamics of fatty acids in live cells and live Caenorhabditis elegans with hsSRS imaging. We found that unsaturated fatty acid has preferential uptake into lipid storage while saturated fatty acid exhibits toxicity in hepatic cells. Simultaneous metabolic fingerprinting of deuterium-labeled saturated and unsaturated fatty acids in living C. elegans revealed that there is a lack of interaction between the two, unlike previously hypothesized. Our findings provide new approaches for metabolic tracing of neutral lipids and their precursors in living cells and organisms, and could potentially serve as a general approach for metabolic fingerprinting of other metabolites.

Dispersion-based stimulated Raman scattering spectroscopy, holography, and optical coherence tomography

“…Vibrational spectroscopy provides the means to non-destructively assess the molecular composition of subcellular structures [1][2][3], trace metabolic and functional processes [4,5], and identify disease [6][7][8]. A variety of different methods have been demonstrated for extracting the molecular information, each with different characteristics (see Ref [9] for a recent review).…”

“…A variety of different methods have been demonstrated for extracting the molecular information, each with different characteristics (see Ref [9] for a recent review). In this work we focus on stimulated Raman scattering (SRS), which provides several advantages over other methods when it is applied to microscopy; for example, under appropriate conditions it has orders of magnitude better sensitivity over spontaneous Raman scattering [3,10, [4,5]. However, due to the different sources of noise and technical constraints (discussed below), most SRS methods are limited to point scanning at a few Raman frequencies (most often, only one).…”

New Gateways to the Platinum Group Metal-Catalyzed Direct Deuterium-Labeling Method Utilizing Hydrogen as a Catalyst Activator

“…[49][50][51][52][53][54][55][56][57][58][59][60][61][62][63][64][65][66][67][68] Although Pt/C has been traditionally used in D 2 O for the preparation of deuterium-labeled saturated fatty acids, harsh hydrothermal reaction conditions and repetitive deuteration processes are required, and the D contents and scope of the substrates were unsatisfactory using previously reported methods. 69) Since the deuteration of alkanes, such Various fatty acids are effectively deuterium-labeled in the presence of Pt/C in an i-PrOD-d 8 /D 2 O mixed solvent (Chart 13).…”

Spectral focusing dual-comb coherent anti-Stokes Raman spectroscopic imaging

“…However, real-time identifying and mapping for multiple or possibly unknown molecules requires broad spectral breadth, desired resolution and high-speed acquisition, and has been a long-term pursuit in the field [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23].…”

“…Hyperspectral SRS and CARS microscopy are now within reach using frequency-swept narrowband laser beams for acquiring images at a series of Raman shifts, but suffer from limitations in laser tuning speed and are not applicable to highly dynamic objects [7][8][9][10][11]. Broadband CARS microscopy using parallel-detection can simultaneously excite multiple Raman bands and reach a speed of 3.5 ms for one pixel [5].…”