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Enhanced detection of myeloperoxidase activity in deep tissues through luminescent excitation of near-infrared nanoparticles

Abstract

A previous study reported the use of luminol for the detection of myeloperoxidase (MPO) activity using optical imaging in infiltrating neutrophils under inflammatory disease conditions. The detection is based on a photon-emitting reaction between luminol and an MPO metabolite. Because of tissue absorption and scattering, however, luminol-emitted blue light can be efficiently detected from superficial inflammatory foci only. In this study we report a chemiluminescence resonance energy transfer (CRET) methodology in which luminol-generated blue light excites nanoparticles to emit light in the near-infrared spectral range, resulting in remarkable improvement of MPO detectability in vivo. CRET caused a 37-fold increase in luminescence emission over luminol alone in detecting MPO activity in lung tissues after lipopolysaccharide challenge. We demonstrated a dependence of the chemiluminescent signal on MPO activity using MPO-deficient mice. In addition, co-administration of 4-aminobenzoic acid hydrazide (4-ABAH), an irreversible inhibitor of MPO, significantly attenuated luminescent emission from inflamed lungs. Inhibition of nitric oxide synthase with a nonspecific inhibitor, L-NAME, had no effect on luminol-mediated chemiluminescence production. Pretreatment of mice with MLN120B, a selective inhibitor of IKK-2, resulted in suppression of neutrophil infiltration to the lung tissues and reduction of MPO activity. We also demonstrated that CRET can effectively detect MPO activity at deep tissue tumor foci due to tumor development–associated neutrophil infiltration. We developed a sensitive MPO detection methodology that provides a means for visualizing and quantifying oxidative stress in deep tissue. This method is amenable to rapid evaluation of anti-inflammatory agents in animal models.

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Figure 1: Detection of pulmonary inflammation with Luminol-R.
Figure 2: Confirmation that CRET substantially improves the in vivo detectability of MPO activity.
Figure 3: Luminol-R-emitted luminescent signal is dependent on MPO activity.
Figure 4: Effect of MLN120B treatment on MPO activity.
Figure 5: Imaging MPO activity in MDA-MB-231-luc2 tumor metastases.

References

  1. Haegens, A., Vernooy, J.H., Heeringa, P., Mossman, B.T. & Wouters, E.F. Myeloperoxidase modulates lung epithelial responses to pro-inflammatory agents. Eur. Respir. J. 31, 252–260 (2008).

    Article  CAS  Google Scholar 

  2. Winterbourn, C.C. Reconciling the chemistry and biology of reactive oxygen species. Nat. Chem. Biol. 4, 278–286 (2008).

    Article  CAS  Google Scholar 

  3. Davies, M.J., Hawkins, C.L., Pattison, D.I. & Rees, M.D. Mammalian heme peroxidases: from molecular mechanisms to health implications. Antioxid. Redox Signal. 10, 1199–1234 (2008).

    Article  CAS  Google Scholar 

  4. O'Donnell, C. et al. 3-chlorotyrosine in sputum of COPD patients: relationship with airway inflammation. COPD 7, 411–417 (2010).

    Article  Google Scholar 

  5. Hazen, S.L. & Heinecke, J.W. 3-chlorotyrosine, a specific marker of myeloperoxidase-catalyzed oxidation, is markedly elevated in low density lipoprotein isolated from human atherosclerotic intima. J. Clin. Invest. 99, 2075–2081 (1997).

    Article  CAS  Google Scholar 

  6. Malle, E., Furtmüller, P.G., Sattler, W. & Obinger, C. Myeloperoxidase: a target for new drug development? Br. J. Pharmacol. 152, 838–854 (2007).

    Article  CAS  Google Scholar 

  7. Kettle, A.J., Gedye, C.A. & Winterbourn, C.C. Mechanism of inactivation of myeloperoxidase by 4-aminobenzoic acid hydrazide. Biochem. J. 321, 503–508 (1997).

    Article  CAS  Google Scholar 

  8. Gross, S. et al. Bioluminescence imaging of myeloperoxidase activity in vivo. Nat. Med. 15, 455–461 (2009).

    Article  CAS  Google Scholar 

  9. Altinoğlu, E.I. & Adair, J.H. Near infrared imaging with nanoparticles. Wiley Interdiscip. Rev. Nanomed. Nanobiotechnol. 2, 461–477 (2010).

    Article  Google Scholar 

  10. So, M.-K., Xu, C., Loening, A.M., Gambhir, S.S. & Rao, J. Self-illuminating quantum dot conjugates for in vivo imaging. Nat. Biotechnol. 24, 339–343 (2006).

    Article  CAS  Google Scholar 

  11. Wang, H.Q. et al. Influence of quantum dot's quantum yield to chemiluminescent resonance energy transfer. Anal. Chim. Acta 610, 68–73 (2008).

    Article  CAS  Google Scholar 

  12. Ansaldi, D. et al. Imaging pulmonary NF-κB activation and therapeutic effects of MLN120B and TDZD-8. PLoS ONE 6, e25093 (2011).

    Article  CAS  Google Scholar 

  13. Orita, T., Shimozaki, K., Murakami, H. & Nagata, S. Binding of NF-Y transcription factor to one of the cis-elements in the myeloperoxidase gene promoter that responds to granulocyte colony-stimulating factor. J. Biol. Chem. 272, 23216–23223 (1997).

    Article  CAS  Google Scholar 

  14. Wu, X. et al. Immunofluorescent labeling of cancer marker Her2 and other cellular targets with semiconductor quantum dots. Nat. Biotechnol. 21, 41–46 (2003).

    Article  CAS  Google Scholar 

  15. Medintz, I.L., Uyeda, H.T., Goldman, E.R. & Mattoussi, H. Quantum dot bioconjugates for imaging, labeling and sensing. Nat. Mater. 4, 435–446 (2005).

    Article  CAS  Google Scholar 

  16. Chan, W.C. & Nie, S. Quantum dot bioconjugates for ultrasensitive nonisotopic detection. Science 281, 2016–2018 (1998).

    Article  CAS  Google Scholar 

  17. Michel, O. et al. Severity of asthma is related to endotoxin in house dust. Am. J. Respir. Crit. Care Med. 154, 1641–1646 (1996).

    Article  CAS  Google Scholar 

  18. Linden, M. et al. Airway inflammation in smokers with nonobstructive and obstructive chronic bronchitis. Am. Rev. Respir. Dis. 148, 1226–1232 (1993).

    Article  CAS  Google Scholar 

  19. Homayoun, H., Khavandgar, S. & Dehpour, A.R. The involvement of endogenous opioids and nitricoxidergic pathway in the anticonvulsant effects of foot-shock stress in mice. Epilepsy Res. 49, 131–142 (2002).

    Article  CAS  Google Scholar 

  20. Nuchprayoon, I. et al. PEBP2/CBF, the murine homolog of the human myeloid AML1 and PEBP2β/CBFβ proto-oncoproteins, regulates the murine myeloperoxidase and neutrophil elastase genes in immature myeloid cells. Mol. Cell Biol. 14, 5558–5568 (1994).

    Article  CAS  Google Scholar 

  21. Shen, W., Gan, J., Xu, S., Jiang, G. & Wu, H. Penehyclidine hydrochloride attenuates LPS-induced acute lung injury involvement of NF-κB pathway. Pharmacol. Res. 60, 296–302 (2009).

    Article  CAS  Google Scholar 

  22. Islam, M.S. et al. Anti-inflammatory effects of phytosteryl ferulates in colitis induced by dextran sulphate sodium in mice. Br. J. Pharmacol. 154, 812–824 (2008).

    Article  CAS  Google Scholar 

  23. Nagashima, K. et al. Rapid TNFR1-dependent lymphocyte depletion in vivo with a selective chemical inhibitor of IKKβ. Blood 107, 4266–4273 (2006).

    Article  CAS  Google Scholar 

  24. Schultz, J. & Kaminker, K. Myeloperoxidase of the leucocyte of normal human blood. I. Content and localization. Arch. Biochem. Biophys. 96, 465–467 (1962).

    Article  CAS  Google Scholar 

  25. Bos, A., Wever, R. & Roos, D. Characterization and quantification of the peroxidase in human monocytes. Biochim. Biophys. Acta 525, 37–44 (1978).

    Article  CAS  Google Scholar 

  26. Gregory, A.D. & Houghton, A.M. Tumor-associated neutrophils: new targets for cancer therapy. Cancer Res. 71, 2411–2416 (2011).

    Article  CAS  Google Scholar 

  27. Irie, S. The treatment of alopecia areata with 3-aminophthal-hydrazide. Curr. Ther. Res. Clin. Exp. 2, 107–110 (1960).

    CAS  PubMed  Google Scholar 

  28. Sanders, J.M., Chen, L.J., Burka, L.T. & Matthews, H.B. Metabolism and disposition of luminol in the rat. Xenobiotica 30, 263–272 (2000).

    Article  CAS  Google Scholar 

  29. Larson, D.R. et al. Water-soluble quantum dots for multiphoton fluorescence imaging in vivo. Science 300, 1434–1436 (2003).

    Article  CAS  Google Scholar 

  30. Stroh, M. et al. Quantum dots spectrally distinguish multiple species within the tumor milieu in vivo. Nat. Med. 11, 678–682 (2005).

    Article  CAS  Google Scholar 

  31. Gao, J. et al. In vivo tumor-targeted fluorescence imaging using near-infrared non-cadmium quantum dots. Bioconjug. Chem. 21, 604–609 (2010).

    Article  CAS  Google Scholar 

  32. Hahn, M.A., Singh, A.K., Sharma, P., Brown, S.C. & Moudgil, B.M. Nanoparticles as contrast agents for in-vivo bioimaging: current status and future perspectives. Anal. Bioanal. Chem. 399, 3–27 (2011).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank H. Xu for spectrometer analysis, E. Lim and K. Wong for technical assistance and S. Ray and R. Singh for valuable discussion.

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Experimental design and concepts were devised by N.Z. and K.P.F., D.A. conducted all the experiments except the Mpo−/− study, which was performed by D.A., A.P. and K.P.F., D.A. and N.Z. performed the data analysis. N.Z. wrote the manuscript.

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Correspondence to Ning Zhang.

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The authors declare no competing financial interests.

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Zhang, N., Francis, K., Prakash, A. et al. Enhanced detection of myeloperoxidase activity in deep tissues through luminescent excitation of near-infrared nanoparticles. Nat Med 19, 500–505 (2013). https://doi.org/10.1038/nm.3110

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