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Sizing lipid droplets from adult and geriatric mouse liver tissue via nanoparticle tracking analysis

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Abstract

The significance of lipid droplets in lipid metabolism, cell signaling, and regulating longevity is increasingly recognized, yet the lipid droplet’s unique properties and architecture make it difficult to size and study using conventional methods. To begin to address this issue, we demonstrate the capabilities of nanoparticle tracking analysis (NTA) for sizing of lipid droplets. NTA was found to be adequate to assess lipid droplet stability over time, indicating that lipid droplet preparations are stable for up to 24 h. NTA had the ability to compare the size distributions of lipid droplets from adult and geriatric mouse liver tissue, suggesting an age-related decrease in lipid droplet size. This is the first report on the use of NTA to size intracellular organelles.

Light scattering reveals the temporal positions of individual lipid droplets, which are recorded with a camera. The two-dimensional diffusion constant of each lipid droplet is extracted from the data set, which is then used to calculate a hydrodynamic radius using the Stokes-Einstein equation.

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References

  1. Murphy DJ. The biogenesis and functions of lipid bodies in animals, plants and microorganisms. Prog Lipid Res. 2001;40:325–438.

    Article  CAS  PubMed  Google Scholar 

  2. Goldberg AA, Bourque SD, Kyryakov P, Boukh-Viner T, Gregg C, Beach A, et al. A novel function of lipid droplets in regulating longevity. Biochem Soc Trans. 2009;37:1050–5.

    Article  CAS  PubMed  Google Scholar 

  3. Mcintosh AL, Storey SM, Atshaves BP. Intracellular lipid droplets contain dynamic pools of sphingomyelin: ADRP binds phospholipids with high affinity. Lipids. 2010;45:465–77.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Londos C, Brasaemle DL, Schultz CJ, Segrest JP, Kimmel AR. Perilipins, ADRP, and other proteins that associate with intracellular neutral lipid droplets in animal cells. Semin Cell Dev Biol. 1999;10:51–8.

    Article  CAS  PubMed  Google Scholar 

  5. Chanderbhan R, Noland BJ, Scallen TJ, Vahouny GV. Sterol carrier protein2. Delivery of cholesterol from adrenal lipid droplets to mitochondria for pregnanolone synthesis. J Biol Chem. 1982;257:8928–34.

    CAS  PubMed  Google Scholar 

  6. Krahmer N, Farese RV, Walther TC. Balancing the fat: lipid droplets and human disease. EMBO Mol Med. 2013;5:905–15.

    Article  CAS  PubMed Central  Google Scholar 

  7. Anstee QM, Targher G, Day CP. Progression of NAFLD to diabetes mellitus, cardiovascular disease or cirrhosis. Nat Rev Gastroenterol Hepatol. 2013;10:330–44.

    Article  CAS  PubMed  Google Scholar 

  8. Day CP, James OFW. Steatohepatitis: a tale of two “Hits”? Gastroenterology. 1998;114:842–5.

    Article  CAS  PubMed  Google Scholar 

  9. Ding Y, Zhang S, Yang L, Na H, Zhang P, Zhang H, et al. Isolating lipid droplets from multiple species. Nat Protoc. 2012;8:43–51.

    Article  CAS  PubMed  Google Scholar 

  10. Nishimoto Y, Nakajima S, Tateya S, Saito M, Ogawa W, Tamori Y. Cell death-inducing DNA fragmentation factor A-like effector A and fat-specific protein 27β coordinately control lipid droplet size in brown adipocytes. J Biol Chem. 2017;292:10824–34.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Shi X, Li J, Zou X, Greggain J, Rødkær SV, Færgeman NJ, et al. Regulation of lipid droplet size and phospholipid composition by stearoyl-CoA desaturase. J Lipid Res. 2013;54:2504–14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Greenspan P, Mayer E, Fowler S. Nile red: a selective fluorescent stain for intracellular lipid droplets. J Cell Biol. 1985;100:965–73.

    Article  CAS  PubMed  Google Scholar 

  13. Yang HJ, Hsu CL, Yang JY, Yang WY. Monodansylpentane as a blue-fluorescent lipid-droplet marker for multi-color live-cell imaging. PLoS One. 2012;7 https://doi.org/10.1371/journal.pone.0032693.

  14. Mehlem A, Hagberg CE, Muhl L, Eriksson U, Falkevall A. Imaging of neutral lipids by oil red O for analyzing the metabolic status in health and disease. Nat Protoc. 2013;8:1149–55.

    Article  CAS  PubMed  Google Scholar 

  15. Klapper M, Ehmke M, Palgunow D, Böhme M, Matthäus C, Bergner G, et al. Fluorescence-based fixative and vital staining of lipid droplets in Caenorhabditis elegans reveal fat stores using microscopy and flow cytometry approaches. J Lipid Res. 2011;52:1281–93.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Deutsch MJ, Schriever SC, Roscher AA, Ensenauer R. Digital image analysis approach for lipid droplet size quantitation of oil red O-stained cultured cells. Anal Biochem. 2014;445:87–9.

    Article  CAS  PubMed  Google Scholar 

  17. Wu A, Kolanowski JL, Boumelhem BB, Yang K, Lee R, Kaur A, et al. A carborane-containing fluorophore as a stain of cellular lipid droplets. Chem Asian J. 2017;12:1704–8.

    Article  CAS  PubMed  Google Scholar 

  18. Gao M, Su H, Li S, Lin Y, Ling X, Qin A, et al. An easily accessible aggregation-induced emission probe for lipid droplet-specific imaging and movement tracking. Chem Commun. 2017;53:921–4.

    Article  CAS  Google Scholar 

  19. Smus JP, Moura CC, McMorrow E, Tare RS, Oreffo ROC, Mahajan S. Tracking adipogenic differentiation of skeletal stem cells by label-free chemically selective imaging. Chem Sci. 2015;6:7089–96.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Cao C, Zhou D, Chen T, Streets AM, Huang Y. Label-free digital quantification of lipid droplets in single cells by stimulated raman microscopy on a microfluidic platform. Anal Chem. 2016;88:4931–9.

    Article  CAS  PubMed  Google Scholar 

  21. Kim K, Lee S, Yoon J, Heo J, Choi C, Park Y. Three-dimensional label-free imaging and quantification of lipid droplets in live hepatocytes. Sci Rep. 2016;6:1–8.

    Article  CAS  Google Scholar 

  22. Daniele JR, Heydari K, Arriaga EA, Dillin A. Identification and characterization of mitochondrial subtypes in caenorhabditis elegans via analysis of individual mitochondria by flow cytometry. Anal Chem. 2016;88:6309–16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Taylor TH, Frost NW, Bowser MT, Arriaga EA. Analysis of individual mitochondria via fluorescent immunolabeling with anti-TOM22 antibodies. Anal Bioanal Chem. 2014;406:1683–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Degtyarev M, Reichelt M, Lin K. Novel quantitative autophagy analysis by organelle flow cytometry after cell sonication. PLoS One. 2014;9 https://doi.org/10.1371/journal.pone.0087707.

  25. Malloy A, Carr B. Nanoparticle tracking analysis - the halo system. Part Syst Charact. 2006;23:197–204.

    Article  Google Scholar 

  26. Soo CY, Song Y, Zheng Y, Campbell EC, Riches AC, Gunn-Moore F, et al. Nanoparticle tracking analysis monitors microvesicle and exosome secretion from immune cells. Immunology. 2012;136:192–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Oosthuyzen W, Sime NEL, Ivy JR, Turtle EJ, Street JM, Pound J, et al. Quantification of human urinary exosomes by nanoparticle tracking analysis. J Physiol. 2013;23:5833–42.

    Article  CAS  Google Scholar 

  28. Tatischeff I, Larquet E, Falcon-Perez JM, Turpin PY, Kruglik SG (2012) Fast characterization of cell-derived extracellular vesicles by nanoparticles tracking analysis, cryo-electron microscopy, and Raman tweezers microspectroscopy. J Extracell Vesicles 1: doi: https://doi.org/10.3402/jev.v1i0.19179

  29. Gardiner C, Ferreira YJ, Dragovic RA, Redman CWG, Sargent IL. Extracellular vesicle sizing and enumeration by nanoparticle tracking analysis. J Extracell Vesicles. 2013;2 https://doi.org/10.3402/jev.v2i0.19671.

  30. Kramberger P, Ciringer M, Štrancar A, Peterka M. Evaluation of nanoparticle tracking analysis for total virus particle determination. Virol J. 2012;9:265.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. James AE, Driskell JD. Monitoring gold nanoparticle conjugation and analysis of biomolecular binding with nanoparticle tracking analysis (NTA) and dynamic light scattering (DLS). Analyst. 2013;138:1212.

    Article  CAS  PubMed  Google Scholar 

  32. Atshaves BP, Storey SM, McIntosh AL, Petrescu AD, Lyuksyutova OI, Greenberg AS, et al. Sterol carrier protein-2 expression modulates protein and lipid composition of lipid droplets. J Biol Chem. 2001;276:25324–35.

    Article  CAS  PubMed  Google Scholar 

  33. Storey SM, AL MI, Senthivinayagam S, Moon KC, Atshaves BP. The phospholipid monolayer associated with perilipin-enriched lipid droplets is a highly organized rigid membrane structure. AJP Endocrinol Metab. 2011;301:E991–E1003.

    Article  CAS  Google Scholar 

  34. Chandrasekhar S. Brownian motion, dynamical friction, and stellar dynamics. Rev Mod Phys. 1949;21:383–8.

    Article  Google Scholar 

  35. Einstein A. On the motion of small particles suspended in a stationary liquid, as required by the molecular kinetic theory of heat. Ann Phys. 1905;322:549–60.

    Article  Google Scholar 

  36. Marx E, Mulholland GW. Size and refractive index determination of single polystyrene spheres. J Res Natl Bur Stand. 1983;88(5):321.

    Article  CAS  Google Scholar 

  37. Haseda K, Kanematsu K, Noguchi K, Saito H, Umeda N, Ohta Y. Significant correlation between refractive index and activity of mitochondria: single mitochondrion study. Biomed Opt Express. 2015;6:859.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Gouw TH, Vlugter JC. Physical properties of triglycerides. !. Density and refractive index. Eur J Lipid Sci Technol. 1966;68:544–69.

    CAS  Google Scholar 

  39. Suzuki M, Shinohara Y, Ohsaki Y, Fujimoto T (2011) Lipid droplets: Size matters. J Electron Microsc 60: https://doi.org/10.1093/jmicro/dfr016

  40. Najt CP, Senthivinayagam S, Aljazi MB, Fader KA, Olenic SD, Brock JRL, et al. Liver-specific loss of perilipin 2 alleviates diet-induced hepatic steatosis, inflammation, and fibrosis. Am J Physiol Gastrointest Liver Physiol. 2016;310:G726–38.

    Article  PubMed  PubMed Central  Google Scholar 

  41. McManaman JL, Bales ES, Orlicky DJ, Jackman M, MacLean PS, Cain S, et al. Perilipin-2-null mice are protected against diet-induced obesity, adipose inflammation, and fatty liver disease. J Lipid Res. 2013;54:1346–59.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Zhang S, Wang Y, Cui L, Deng Y, Xu S, Yu J, et al. Morphologically and functionally distinct lipid droplet subpopulations. Sci Rep. 2016;6:29539.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Cohen B-C, Shamay A, Argov-Argaman N. Regulation of lipid droplet size in mammary epithelial cells by remodeling of membrane lipid composition-a potential mechanism. PLoS One. 2015;10:e0121645.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Bonda-Ostaszewska E, Włostowski T, Krasowska A, Kozłowski P. Seasonal and photoperiodic effects on lipid droplet size and lipid peroxidation in the brown adipose tissue of bank voles (Myodes glareolus). Acta Theriol (Warsz). 2012;57:289–94.

    Article  Google Scholar 

  45. Slawik M, Vidal-Puig AJ. Lipotoxicity, overnutrition, and energy metabolism in aging. Aging Res Rev. 2006;5:144–64.

    Article  CAS  Google Scholar 

  46. Petersen KF, Befroy D, Dufour S, Dziura J, Ariyan C, Rothman DL, et al. Mitochondrial dysfunction in the elderly: Possible role in insulin resistance. Science. 2003;300(80):1140–2.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Daniele JR, Esping DJ, Garcia G, Parsons LS, Arriaga EA, Dillin A. High-throughput characterization of region-specific mitochondrial function and morphology. Sci Rep. 2017;7:6749.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Funding

This work was supported by NIH AG020866. KAM acknowledges support through a University of Minnesota Doctoral Dissertation Fellowship and National Institutes of Health (NIH) AG029796. CPN acknowledges support from NIH DK007203. NML acknowledges support from NIH GM008700. DGM acknowledges support from NIH DK114401 and NIH AG055452. A portion of this work was carried out in the Minnesota Nano Center, which receives partial support from the National Science Foundation (NSF) through the NNCI program. A portion of this work was carried out in the Characterization Facility, University of Minnesota, which receives partial support from NSF through the MRSEC program.

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Correspondence to Edgar A. Arriaga.

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All mice were housed in a designated clean facility and treated in accordance with protocols approved by the University of Minnesota Institutional Animal Care and Use Committee.

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The authors have no potential conflicts of interest to disclose.

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Muratore, K.A., Najt, C.P., Livezey, N.M. et al. Sizing lipid droplets from adult and geriatric mouse liver tissue via nanoparticle tracking analysis. Anal Bioanal Chem 410, 3629–3638 (2018). https://doi.org/10.1007/s00216-018-1016-8

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  • DOI: https://doi.org/10.1007/s00216-018-1016-8

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