Abstract
Recent clinical trials demonstrated strong association between lipid abnormalities and progression of diabetic retinopathy (DR); however, whether circulating lipid levels or retinal lipid metabolism, or both, contributes to the pathogenesis of DR is not well understood. Limited amounts of retinal tissue available from animal models, such as mouse models of DR, have proved. Limited amount of retinal tissue was especially challenging for cholesterol and oxysterol detection as it precluded identification of individual isomers of each nonesterified sterol class. To measure cholesterol and oxysterols from limited retinal tissue samples, we developed extremely sensitive electrospray ionization liquid chromatography high-resolution/accurate mass measurements on an LTQ Orbitrap Velos mass spectrometer that are able to resolve sterols and oxysterols separated by reverse-phase HPLC using a gradient of 85–100% methanol containing 0.1% formic acid, with subsequent detection in positive ionization mode. This methodology will aid in our understanding of diabetes-induced changes in retinal cholesterol and oxysterol metabolism.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Busik JV (2021) Lipid metabolism dysregulation in diabetic retinopathy. J Lipid Res 62(100017). https://doi.org/10.1194/jlr.TR120000981
Hammer SS, Vieira CP, McFarland D, Sandler M, Levitsky Y, Dorweiler TF et al (2021) Fasting and fasting-mimicking treatment activate SIRT1/LXRalpha and alleviate diabetes-induced systemic and microvascular dysfunction. Diabetologia 64(7):1674–1689. https://doi.org/10.1007/s00125-021-05431-5
DeBarber AE, Sandlers Y, Pappu AS, Merkens LS, Duell PB, Lear SR et al (2011) Profiling sterols in cerebrotendinous xanthomatosis: utility of Girard derivatization and high resolution exact mass LC-ESI-MS(n) analysis. J Chromatogr B Analyt Technol Biomed Life Sci 879(17-18):1384–1392. https://doi.org/10.1016/j.jchromb.2010.11.019
Curtis TM, McLaughlin D, O’Hare M, Kur J, Barabas P, Revolta G et al (2018) Isolation of retinal arterioles for ex vivo cell physiology studies. J Vis Exp 137. https://doi.org/10.3791/57944
Chintalapudi SR, Patel NN, Goldsmith ZK, Djenderedjian L, Wang XD, Marion TN et al (2017) Isolation of primary murine retinal ganglion cells (RGCs) by flow cytometry. J Vis Exp 125. https://doi.org/10.3791/55785
Winkler BS (1972) The electroretinogram of the isolated rat retina. Vision Res 12(6):1183–1198. https://doi.org/10.1016/0042-6989(72)90106-x
Lydic TA, Busik JV, Reid GE (2014) A monophasic extraction strategy for the simultaneous lipidome analysis of polar and nonpolar retina lipids. J Lipid Res 55(8):1797–1809. https://doi.org/10.1194/jlr.D050302
Machacek M, Saunders H, Zhang Z, Tan EP, Li J, Li T et al (2019) Elevated O-GlcNAcylation enhances pro-inflammatory Th17 function by altering the intracellular lipid microenvironment. J Biol Chem 294(22):8973–8990. https://doi.org/10.1074/jbc.RA119.008373
Vieira CP, Fortmann SD, Hossain M, Longhini AL, Hammer SS, Asare-Bediako B et al (2020) Selective LXR agonist DMHCA corrects retinal and bone marrow dysfunction in type 2 diabetes. JCI Insight 5(13). https://doi.org/10.1172/jci.insight.137230
Agrawal S, Kumar S, Sehgal R, George S, Gupta R, Poddar S et al (2019) El-MAVEN: a fast, robust, and user-friendly mass spectrometry data processing engine for metabolomics. Methods Mol Biol 1978:301–321. https://doi.org/10.1007/978-1-4939-9236-2_19
McDonald JG, Thompson BM, McCrum EC, Russell DW (2007) Extraction and analysis of sterols in biological matrices by high performance liquid chromatography electrospray ionization mass spectrometry. Methods Enzymol. 432:145–170. https://doi.org/10.1016/S0076-6879(07)32006-5
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Lydic, T.A., Busik, J.V. (2023). Diabetes Retinopathy: New Ways to Detect and Treat. In: Moore, A., Wang, P. (eds) Type-1 Diabetes. Methods in Molecular Biology, vol 2592. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2807-2_6
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2807-2_6
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2806-5
Online ISBN: 978-1-0716-2807-2
eBook Packages: Springer Protocols