Dissociation of 19F and fluorescence signal upon cellular uptake of dual-contrast perfluorocarbon nanoemulsions

Abstract

Objective

Perfluorocarbon nanoemulsions (PFCs) tagged with fluorescence dyes have been intensively used to confirm the in vivo 19F magnetic resonance imaging (MRI) localization of PFCs by post mortem histology or flow cytometry. However, only limited data are available on tagged PFCs and the potential dissociation of fluorescence and 19F label after cellular uptake over time.

Materials and methods

PFCs were coupled to rhodamine (Rho) or carboxyfluorescein (Cfl) and their fate was analyzed after in vitro uptake by J774, RAW and CHO cells by flow cytometry and 19F MRI. In separate in vivo experiments, the dual-labelled emulsions were intravenously applied into mice and their distribution was monitored in spleen and liver over 24 h. In a final step, time course of fluorescence and 19F signals from injected emulsions were tracked in a local inflammation model making use of a subcutaneous matrigel depot doped with LPS (lipopolysaccharide).

Results

Internalization of fluorescence-labelled PFCs was associated with a substantial whitening over 24 h in all macrophage cell lines while the 19F signal remained stable over time. In all experiments, CflPFCs were more susceptible to bleaching than RhoPFCs. After intravenous injection of RhoPFCs, the fluorescence signal in spleen and liver peaked after 30 min and 2 h, respectively, followed by a successive decrease over 24 h, whereas the 19F signal continuously increased during this observation period. Similar results were found in the matrigel/LPS model, where we observed increasing 19F signals in the inflammatory hot spot over time while the fluorescence signal of immune cells isolated from the matrigel depot 24 h after its implantation was only marginally elevated over background levels. This resulted in a massive underestimation of the true PFC deposition in the reticuloendothelial system and at inflammatory hot spots.

Conclusion

Cellular uptake of fluorescently tagged PFCs leads to a dissociation of the fluorescence and the 19F label signal over time, which critically impacts on interpretation of long-term experiments validated by histology or flow cytometry.

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References

  1. 1.

    Bulte JWM (2005) Hot spot MRI emerges from the background. Nat Biotechnol 23:945–946

    Article  CAS  PubMed  Google Scholar 

  2. 2.

    Riess JG (2005) Understanding the fundamentals of Perfluorocarbons and perfluorocarbon emulsions relevant to in vivo oxygen delivery. Artif Cells Blood Substit Biotechnol 33:47–63

    Article  CAS  Google Scholar 

  3. 3.

    O’Hagan D (2008) Understanding organofluorine chemistry. An introduction to the C–F bond. Chem Soc Rev 37:308–319

    Article  PubMed  Google Scholar 

  4. 4.

    Grapentin C, Mayenfels F, Barnert S, Süss R, Schubert R, Temme S, Jacoby C, Schrader J, Flögel U (2014) Optimization of perfluorocarbon nanoemulsions for molecular imaging by 19F MRI. Nanomed One Central Press, Machester, pp 268–286

    Google Scholar 

  5. 5.

    Srinivas M, Boehm-Sturm P, Figdor CG, de Vries IJ, Hoehn M (2012) Labeling cells for in vivo tracking using 19F MRI. Biomaterials 33:8830–8840

    Article  CAS  PubMed  Google Scholar 

  6. 6.

    Temme S, Grapentin C, Güden-Silber T, Flögel U (2016) Active targeting of perfluorocarbon nanoemulsions. Fluorine magnetic resonance imaging. CRC Press, Boca Raton, pp 97–133

    Google Scholar 

  7. 7.

    Ebner B, Behm P, Jacoby C, Burghoff S, French BA, Schrader J, Flögel U (2010) Early assessment of pulmonary inflammation by 19F MRI in vivo. Circ Cardiovasc Imaging 3:202–210

    Article  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Flögel U, Ding Z, Hardung H, Jander S, Reichmann G, Jacoby C, Schubert R, Schrader J (2008) In vivo monitoring of inflammation after cardiac and cerebral ischemia by fluorine magnetic resonance imaging. Circulation 118:140–148

    Article  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Flögel U, Burghoff S, van Lent PLEM, Temme S, Galbarz L, Ding Z, El-Tayeb A, Huels S, Bönner F, Borg N, Jacoby C, Muller CE, van den Berg WB, Schrader J (2012) Selective activation of adenosine A2A receptors on immune cells by a CD73-dependent prodrug suppresses joint inflammation in experimental rheumatoid arthritis. Sci Transl Med 4:146ra108

    Article  CAS  PubMed  Google Scholar 

  10. 10.

    van Heeswijk RB, Pellegrin M, Flögel U, Gonzales C, Aubert J-F, Mazzolai L, Schwitter J, Stuber M (2015) Fluorine MR imaging of inflammation in atherosclerotic plaque in vivo. Radiology 275:421–429

    Article  PubMed  Google Scholar 

  11. 11.

    Boehm-Sturm P, Mengler L, Wecker S, Hoehn M, Kallur T (2011) In vivo tracking of human neural stem cells with 19F magnetic resonance imaging. PLoS One 6:e29040

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. 12.

    Helfer BM, Balducci A, Sadeghi Z, Ohanlon C, Hijaz A, Flask CA, Wesa A (2013) 19F MRI tracer preserves in vitro and in vivo properties of hematopoietic stem cells. Cell Transplant 22:87–97

    Article  PubMed  Google Scholar 

  13. 13.

    Ding Z, Temme S, Quast C, Friebe D, Jacoby C, Zanger K, Bidmon H-J, Grapentin C, Schubert R, Flogel U, Schrader J (2016) Epicardium-derived cells formed after myocardial injury display phagocytic activity permitting in vivo labeling and tracking. Stem Cells Transl Med 5:639–650

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Solomon M, Muro S (2017) Lysosomal enzyme replacement therapies: historical development, clinical outcomes, and future perspectives. Adv Drug Deliv Rev 118:109–134

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. 15.

    Dupré-Crochet S, Erard M, Nüße O (2013) ROS production in phagocytes: why, when, and where? J Leukoc Biol 94:657–670

    Article  CAS  PubMed  Google Scholar 

  16. 16.

    Temme S, Jacoby C, Ding Z, Bönner F, Borg N, Schrader J, Flögel U (2014) Technical advance: monitoring the trafficking of neutrophil granulocytes and monocytes during the course of tissue inflammation by noninvasive 19F MRI. J Leukoc Biol 95:689–697

    Article  CAS  PubMed  Google Scholar 

  17. 17.

    Temme S, Grapentin C, Quast C, Jacoby C, Grandoch M, Ding Z, Owenier C, Mayenfels F, Fischer JW, Schubert R, Schrader J, Flögel U (2015) Noninvasive imaging of early venous thrombosis by 19F magnetic resonance imaging with targeted perfluorocarbon nanoemulsions. Circulation 131:1405–1414

    Article  CAS  PubMed  Google Scholar 

  18. 18.

    Vasir JK, Labhasetwar V (2007) Biodegradable nanoparticles for cytosolic delivery of therapeutics. Adv Drug Deliv Rev 59:718–728

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Skotland T, Sontum PC, Oulie I (2002) In vitro stability analyses as a model for metabolism of ferromagnetic particles (Clariscan™), a contrast agent for magnetic resonance imaging. J Pharm Biomed Anal 28:323–329

    Article  CAS  PubMed  Google Scholar 

  20. 20.

    Seleverstov O, Zabirnyk O, Zscharnack M, Bulavina L, Nowicki M, Heinrich J-M, Yezhelyev M, Emmrich F, O’Regan R, Bader A (2006) Quantum dots for human mesenchymal stem cells labeling. A size-dependent autophagy activation. Nano Lett 6:2826–2832

    Article  CAS  PubMed  Google Scholar 

  21. 21.

    Fitzpatrick JAJ, Andreko SK, Ernst LA, Waggoner AS, Ballou B, Bruchez MP (2009) Long-term persistence and spectral blue shifting of quantum dots in vivo. Nano Lett 9:2736–2741

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Ratner AV, Hurd R, Muller HH, Bradley-Simpson B, Pitts W, Shibata D, Sotak C, Young SW (1987) 19F magnetic resonance imaging of the reticuloendothelial system. Magn Reson Med 5:548–554

    Article  CAS  PubMed  Google Scholar 

  23. 23.

    Long DM, Multer FK, Greenburg AG, Peskin GW, Lasser EC, Wickham WG, Sharts CM (1978) Tumor imaging with X-rays using macrophage uptake of radiopaque fluorocarbon emulsions. Surgery 84:104–112

    CAS  PubMed  Google Scholar 

  24. 24.

    Jacoby C, Temme S, Mayenfels F, Benoit N, Krafft MP, Schubert R, Schrader J, Flögel U (2014) Probing different perfluorocarbons for in vivo inflammation imaging by 19F MRI: image reconstruction, biological half-lives and sensitivity. NMR Biomed 27:261–271

    Article  CAS  PubMed  Google Scholar 

  25. 25.

    Flaim SF (1994) Pharmacokinetics and side effects of perfluorocarbon-based blood substitutes. Artif Cells Blood Substit Immobil Biotechnol 22:1043–1054

    Article  CAS  PubMed  Google Scholar 

  26. 26.

    Krafft MP, Riess JG, Weers JG (1998) The design and engineering of oxygen-delivering fluorocarbonemulsions. In: Benita S (ed) Submicron emulsions in drug targeting and delivery. Harwood, Amsterdam, pp 235–333

    Google Scholar 

  27. 27.

    Riess JG (2001) Oxygen carriers (“blood substitutes”)–raison d’etre, chemistry, and some physiology. Chem Rev 101:2797–2920

    Article  CAS  PubMed  Google Scholar 

  28. 28.

    Tsuda Y, Yamanouchi K, Yokoyama K, Suyama T, Watanabe M, Ohyanagi H, Saitoh Y (1988) Discussion and considerations for the excretion mechanism of perfluorochemical emulsion. Biomater Artif Cells Artif Organs 16:473–483

    Article  CAS  PubMed  Google Scholar 

  29. 29.

    Yokoyama K, Yamanouchi K, Murashima R (1975) Excretion of Perfluorochemicals after intravenous injection of their emulsion. Chem Pharm Bull (Tokyo) 23:1368–1373

    Article  CAS  Google Scholar 

  30. 30.

    Riess JG (1992) Overview of progress in the fluorocarbon approach to in vivo oxygen delivery. Biomater Artif Cells Immobil Biotechnol 20:183–202

    CAS  Google Scholar 

  31. 31.

    Bönner F, Jacoby C, Temme S, Borg N, Ding Z, Schrader J, Flögel U (2014) Multifunctional MR monitoring of the healing process after myocardial infarction. Basic Res Cardiol 109:430

    Article  CAS  PubMed  Google Scholar 

  32. 32.

    Skajaa T, Zhao Y, van den Heuvel DJ, Gerritsen HC, Cormode DP, Koole R, van Schooneveld MM, Post JA, Fisher EA, Fayad ZA, de Mello Donega C, Meijerink A, Mulder WJM (2010) Quantum dot and Cy5.5 labeled nanoparticles to investigate lipoprotein biointeractions via Förster resonance energy transfer. Nano Lett 10:5131–5138

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Lunov O, Syrovets T, Röcker C, Tron K, Ulrich Nienhaus G, Rasche V, Mailänder V, Landfester K, Simmet T (2010) Lysosomal degradation of the carboxydextran shell of coated superparamagnetic iron oxide nanoparticles and the fate of professional phagocytes. Biomaterials 31:9015–9022

    Article  CAS  PubMed  Google Scholar 

  34. 34.

    Gálisová A, Herynek V, Swider E, Sticová E, Pátiková A, Kosinová L, Kříž J, Hájek M, Srinivas M, Jirák D (2018) A trimodal imaging platform for tracking viable transplanted pancreatic islets in vivo: F-19 MR, fluorescence, and bioluminescence imaging. Mol Imaging Biol. https://doi.org/10.1007/s11307-018-1270-3

    Article  PubMed Central  Google Scholar 

  35. 35.

    Janjic JM, Srinivas M, Kadayakkara DKK, Ahrens ET (2008) Self-delivering nanoemulsions for dual fluorine-19 MRI and fluorescence detection. J Am Chem Soc 130:2832–2841

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We like to thank Bodo Steckel and Sabine Barnert for excellent technical assistance. This work was supported by the Deutsche Forschungsgemeinschaft (DFG) grants ST 1209/1-1, FL 303/6-1 and the Sonderforschungsbereich SFB 1116.

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Authors

Contributions

Study conception and design: PB, JS, RS, ST, UF. Acquisition of data: PB, WK, VF, ST. Analysis and interpretation of data: PB, WK, VF, ST, UF. Drafting of manuscript: PB, ST, UF. Critical revision: UF, ST, JS, RS.

Corresponding author

Correspondence to Ulrich Flögel.

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The authors declare that they have no conflict of interest.

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All procedures performed in studies involving animals were in accordance with the ethical standards of the institution at which the studies were conducted.

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Bouvain, P., Flocke, V., Krämer, W. et al. Dissociation of 19F and fluorescence signal upon cellular uptake of dual-contrast perfluorocarbon nanoemulsions. Magn Reson Mater Phy 32, 133–145 (2019). https://doi.org/10.1007/s10334-018-0723-7

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Keywords

  • 19F MRI
  • Perfluorocarbons
  • Inflammation
  • Fluorescence
  • Contrast media