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In vivo ultrasound-induced luminescence molecular imaging

Nature Photonics volume 18, pages 334–343 (2024)Cite this article

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Abstract

Optical imaging is crucial to study biological or pathological processes and diagnose diseases. However, poor tissue penetration typically limits conventional optical imaging. Here we report an imaging technique that uses ultrasound to activate luminescent molecules or nanoparticles through two-step intraparticle energy conversion. Ultrasonic activation can convert mechanical fluctuations into chemical energy via the piezoelectric effect and then induce luminescence via the chemiluminescent effect. We demonstrate two modalities for ultrasound-induced luminescence imaging: one achieves delayed imaging after cessation of the ultrasonic excitation, and the other enables real-time imaging during the ultrasonic excitation. Our imaging modality offers an improvement in luminescence intensity of up to 2,000-fold compared with sonoluminescence of H2O, a 10-fold improved of signal-to-noise ratio compared with fluorescence imaging, a spatial resolution of 1.46 mm and tissue penetration of up to 2.2 cm. We demonstrate its applicability for imaging subcutaneous and orthotopic tumours, mapping lymph nodes and screening peritoneal metastatic tumours. Furthermore, we design analyte-activatable luminescence probes based on resonance energy transfer, which can assess drug-induced hepatotoxicity and distinguish the responsivity of tumours after drug treatment. We expect that our technique will enable further preclinical and clinical applications, such as the study of histopathological lesions in living animals, the early detection of tumours, the profiling of biological molecules and the monitoring of cancer treatment or prognosis, among others.

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Fig. 1: Synthesis and measurement of luminescent nanoparticles for ultrasound-induced luminescence imaging.
Fig. 2: Study of the delayed ultrasound-induced luminescence imaging performance for TD NPs.
Fig. 3: Mechanism for ultrasound-induced luminescence.
Fig. 4: Delayed ultrasound-induced luminescence imaging in vivo.
Fig. 5: Activatable ultrasound-induced luminescence probes for imaging of enzymes and ONOO−.
Fig. 6: TD-Grz-BHQ activatable ultrasound-induced luminescence imaging of granzyme B in vivo.

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Data availability

All of the data that support the findings of this study are reported in the main text and Supplementary Information. The data that support the findings of this study are available from the corresponding author upon reasonable request, and data for all figures can be found at https://doi.org/10.6084/m9.figshare.24579853. Source data are provided with this paper.

References

  1. Chen, Y.-S. et al. Ultra-high-frequency radio-frequency acoustic molecular imaging with saline nanodroplets in living subjects. Nat. Nanotechnol. 16, 717–724 (2021).

    Article  ADS  Google Scholar 

  2. Jokerst, J. V. & Gambhir, S. S. Molecular imaging with theranostic nanoparticles. Acc. Chem. Res. 44, 1050–1060 (2011).

    Article  Google Scholar 

  3. Klinkhammer, B. M. et al. Non-invasive molecular imaging of kidney diseases. Nat. Rev. Nephrol. 17, 688–703 (2021).

    Article  Google Scholar 

  4. Lavine, K. J. & Liu, Y. The dynamic cardiac cellular landscape: visualization by molecular imaging. Nat. Rev. Cardiol. 19, 345–347 (2022).

    Article  Google Scholar 

  5. Liu, J.-n, Bu, W. & Shi, J. Chemical design and synthesis of functionalized probes for imaging and treating tumor hypoxia. Chem. Rev. 117, 6160–6224 (2017).

    Article  Google Scholar 

  6. Wang, Z. et al. Two-way magnetic resonance tuning and enhanced subtraction imaging for non-invasive and quantitative biological imaging. Nat. Nanotechnol. 15, 482–490 (2020).

    Article  ADS  Google Scholar 

  7. Jiang, Y. & Pu, K. Molecular probes for autofluorescence-free optical imaging. Chem. Rev. 121, 13086–13131 (2021).

    Article  Google Scholar 

  8. Zhao, Z. et al. Ultra-bright Raman dots for multiplexed optical imaging. Nat. Commun. 12, 1305 (2021).

    Article  ADS  Google Scholar 

  9. Kim, E. H., Chin, G., Rong, G., Poskanzer, K. E. & Clark, H. A. Optical probes for neurobiological sensing and imaging. Acc. Chem. Res. 51, 1023–1032 (2018).

    Article  Google Scholar 

  10. Li, Y., Gecevicius, M. & Qiu, J. Long persistent phosphors—from fundamentals to applications. Chem. Soc. Rev. 45, 2090–2136 (2016).

    Article  Google Scholar 

  11. Weber, J., Beard, P. C. & Bohndiek, S. E. Contrast agents for molecular photoacoustic imaging. Nat. Methods 13, 639–650 (2016).

    Article  Google Scholar 

  12. Pei, P. et al. X-ray-activated persistent luminescence nanomaterials for NIR-II imaging. Nat. Nanotechnol. 16, 1011–1018 (2021).

    Article  ADS  Google Scholar 

  13. Pratt, E. C. et al. Prospective testing of clinical Cerenkov luminescence imaging against standard-of-care nuclear imaging for tumour location. Nat. Biomed. Eng. 6, 559–568 (2022).

    Article  Google Scholar 

  14. Wu, X. et al. Tether-free photothermal deep-brain stimulation in freely behaving mice via wide-field illumination in the near-infrared-II window. Nat. Biomed. Eng. 6, 754–770 (2022).

    Article  Google Scholar 

  15. Yang, M. et al. Chemiluminescence for bioimaging and therapeutics: recent advances and challenges. Chem. Soc. Rev. 49, 6800–6815 (2020).

    Article  Google Scholar 

  16. Diao, S. et al. Fluorescence imaging in vivo at wavelengths beyond 1500 nm. Angew. Chem. Int. Ed. 54, 14758–14762 (2015).

    Article  Google Scholar 

  17. He, S., Song, J., Qu, J. & Cheng, Z. Crucial breakthrough of second near-infrared biological window fluorophores: design and synthesis toward multimodal imaging and theranostics. Chem. Soc. Rev. 47, 4258–4278 (2018).

    Article  Google Scholar 

  18. Miao, Q. et al. Molecular afterglow imaging with bright, biodegradable polymer nanoparticles. Nat. Biotechnol. 35, 1102–1110 (2017).

    Article  Google Scholar 

  19. 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  Google Scholar 

  20. Liu, H. et al. Intraoperative imaging of tumors using Cerenkov luminescence endoscopy: a feasibility experimental study. J. Nucl. Med. 53, 1579–1584 (2012).

    Article  Google Scholar 

  21. Farhadi, A., Ho, G. H., Sawyer, D. P., Bourdeau, R. W. & Shapiro, M. G. Ultrasound imaging of gene expression in mammalian cells. Science 365, 1469–1475 (2019).

    Article  ADS  Google Scholar 

  22. Gilad, A. A. & Shapiro, M. G. Molecular imaging in synthetic biology, and synthetic biology in molecular imaging. Mol. Imaging Biol. 19, 373–378 (2017).

    Article  Google Scholar 

  23. Heiles, B., Terwiel, D. & Maresca, D. The advent of biomolecular ultrasound imaging. Neuroscience 474, 122–133 (2021).

    Article  Google Scholar 

  24. Frenzel, H. & Schultes, H. Luminescenz im ultraschallbeschickten Wasser. Z. Phys. Chem. 27, 421–424 (1934).

    Article  Google Scholar 

  25. Didenko, Y. T., McNamara III, W. B. & Suslick, K. S. Molecular emission from single-bubble sonoluminescence. Nature 407, 877–879 (2000).

    Article  ADS  Google Scholar 

  26. Doktycz, S. J. & Suslick, K. S. Interparticle collisions driven by ultrasound. Science 247, 1067–1069 (1990).

    Article  ADS  Google Scholar 

  27. Barber, B. P. & Putterman, S. J. Observation of synchronous picosecond sonoluminescence. Nature 352, 318–320 (1991).

    Article  ADS  Google Scholar 

  28. Xing, D., He, L., Tang, Y. & Tan, S. Sonoluminescence imaging in vivo. In Proc. SPIE 4162, Controlling Tissue Optical Properties: Applications in Clinical Study (ed. Tuchin, V. V.) 86–92 (SPIE, 2000).

  29. He, L., Xing, D., Yan, X. & Ueda, K.-I. Chemiluminescence detection from sonodynamic action in vitro and in vivo. In Proc. SPIE 4597, Biophotonics Instrumentation and Analysis (eds Chiou, A. E. T. et al.) 43–50 (SPIE, 2001).

  30. He, L., Xing, D., Yao, Y., Yan, X. & Ueda, K.-I. Tumor detection with sonodynamic chemiluminescence from ATX-70 and FCLA under ultrasonic excitation. In Proc. SPIE 4612, Optical Methods for Tumor Treatment and Detection: Mechanisms and Techniques in Photodynamic Therapy XI (ed. Dougherty, T. J.) 196–203 (SPIE, 2002).

  31. He, Y., Tang, Y., Tan, S. & Xing, D. FCLA-enhanced sonoluminescence imaging in vivo. In Proc. SPIE 3863, 1999 International Conference on Biomedical Optics (eds Luo, Q. et al.) 129–133 (SPIE, 1999).

  32. He, Y., Xing, D., Tan, S., Tang, Y. & Ueda, K.-I. In vivo sonoluminescence imaging with the assistance of FCLA. Phys. Med. Biol. 47, 1535–1541 (2002).

    Article  Google Scholar 

  33. Wang, L. V. & Shen, Q. Sonoluminescent tomography of strongly scattering media. Opt. Lett. 23, 561–563 (1998).

    Article  ADS  Google Scholar 

  34. Xu, C. et al. Nanoparticles with ultrasound-induced afterglow luminescence for tumour-specific theranostics. Nat. Biomed. Eng. 7, 298–312 (2022).

    Article  Google Scholar 

  35. Wang, W. et al. Ultrasound triggered organic mechanoluminescence materials. Adv. Drug Deliv. Rev. 186, 114343 (2022).

    Article  Google Scholar 

  36. Wang, W. et al. Ultrasound-triggered in situ photon emission for noninvasive optogenetics. J. Am. Chem. Soc. 145, 1097–1107 (2023).

    Article  Google Scholar 

  37. Wu, X. et al. Sono-optogenetics facilitated by a circulation-delivered rechargeable light source for minimally invasive optogenetics. Proc. Natl Acad. Sci. USA 116, 26332–26342 (2019).

    Article  ADS  Google Scholar 

  38. Zhang, G. et al. A selective and sensitive chemiluminescence reaction of 4,4′(5′)-bis[2-(9-anthryloxy)ethylthio]tetrathiafulvalene with singlet oxygen. Chem. Commun., 2072–2073 (2004).

  39. Dang, J. & Zhang, Q. Gas-phase reaction of benzo[a]anthracene with hydroxyl radical in the atmosphere: products, oxidation mechanism, and kinetics. J. Mol. Model. 24, 320 (2018).

    Article  Google Scholar 

  40. Li, H. et al. Activity-based NIR fluorescent probes based on the versatile hemicyanine scaffold: design strategy, biomedical applications, and outlook. Chem. Soc. Rev. 51, 1795–1835 (2022).

    Article  Google Scholar 

  41. Wu, X., Li, H., Lee, E. & Yoon, J. Sensors for in situ real-time fluorescence imaging of enzymes. Chem 6, 2893–2901 (2020).

    Article  Google Scholar 

  42. Zhou, L., Zhang, P., Wang, H., Wang, D. & Li, Y. Smart nanosized drug delivery systems inducing immunogenic cell death for combination with cancer immunotherapy. Acc. Chem. Res. 53, 1761–1772 (2020).

    Article  Google Scholar 

  43. Nguyen, A. et al. Granzyme B nanoreporter for early monitoring of tumor response to immunotherapy. Sci. Adv. 6, eabc2777 (2020).

    Article  ADS  Google Scholar 

  44. Larimer, B. M. et al. Granzyme B PET imaging as a predictive biomarker of immunotherapy response. Cancer Res. 77, 2318–2327 (2017).

    Article  Google Scholar 

  45. He, S., Li, J., Lyu, Y., Huang, J. & Pu, K. Near-infrared fluorescent macromolecular reporters for real-time imaging and urinalysis of cancer immunotherapy. J. Am. Chem. Soc. 142, 7075–7082 (2020).

    Article  Google Scholar 

  46. Ngwa, W. et al. Using immunotherapy to boost the abscopal effect. Nat. Rev. Cancer 18, 313–322 (2018).

    Article  Google Scholar 

  47. Zhao, Y. et al. ICAM-1 orchestrates the abscopal effect of tumor radiotherapy. Proc. Natl Acad. Sci. USA 118, e2010333118 (2021).

    Article  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Key R&D Program of China (grant no. 2019YFA0210100 (to X.-B.Z.)) and the National Natural Science Foundation of China (grant nos U21A20287 (to G.S.) and 22234003 and 21890744 (to X.-B.Z.)).

Author information

Authors and Affiliations

  1. State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, China

    Youjuan Wang, Jing Guo, Shiyi Liao, Zhe Li, Shuai Xu, Baoli Yin, Yongchao Liu, Yurong Feng, Qiming Rong, Guosheng Song, Xiao-Bing Zhang & Weihong Tan

  2. Department of Chemistry, National University of Singapore, Singapore, Singapore

    Zhigao Yi & Xiaogang Liu

  3. The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, China

    Weihong Tan

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Contributions

J.G. and Z.L. synthesized the TD molecules. S.X. and Y.F. synthesized the HBA-COOH molecules. Y.L. synthesized the BODIPY, BODIPY-Br and HD molecules. Z.L. and B.Y. synthesized the BTz-IC-H, BTz-IC-F and BTz-IC-Cl molecules. Y.W. and S.L. conducted all experiments in solution. Y.W., S.L. and Q.R. built all of the animal models and conducted all of the animal experiments. Y.W., Z.L. and S.L. synthesized all nanoparticles. Y.W. collected the raw data in all experiments and designed the schematic diagram. G.S. conceived the idea for this project. G.S. and Y.W. designed the research and all experiments. Y.W. and G.S. analysed all of the data and interpreted the results. Y.W., G.S., X.-B.Z. and W.T. wrote the paper. G.S., X.-B.Z., W.T. and Y.W. developed the discussion. G.S. supervised all experiments. Z.Y. and X.L. helped to analyse the luminescence mechanism and wrote the paper. All authors provided critical feedback on the research and the paper.

Corresponding authors

Correspondence to Guosheng Song, Xiao-Bing Zhang or Weihong Tan.

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Nature Photonics thanks Kanyi Pu and the other anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data

Extended Data Fig. 1

Schematic diagram of set up apparatus for the mode of “delayed ultrasound-induced luminescence imaging” and “real-time ultrasound-induced luminescence imaging”.

Source data

Extended Data Fig. 2 Design of composite nanoparticles to enhance ultrasound-induced luminescence, in the mode of “delayed ultrasound-induced luminescence imaging”.

a, Schematic diagram for design of composite nanoparticles by luminescent molecule (PFODBT) and chemiluminescent substrates. b, TEM image revealed spherical PFODBT@HBA-NPs. c, Ultrasound-induced luminescence intensity of various chemiluminescent substrates doped PFODBT-NPs (50 μg mL–1, 400 µL) pre-excited with ultrasound (30 kHz, 4.5 W cm–2) for 45 s. d, Ultrasound-induced luminescence intensity of PFODBT@HBA-NPs with different ratios of HBA-COOH: PFODBT, nanoparticle solutions (50 μg/mL, 200 µL) were pre-excited with ultrasound (30 kHz, 4.5 W/cm2) for 15 s. e, Ultrasound-induced luminescence intensity of PFODBT@HBA-NPs with different surfactants, those nanoparticles (50 μg mL–1, 400 μL) were pre-excited with ultrasound (30 kHz, 4.5 W cm–2) for 30 s. f, Ultrasound-induced luminescence emission of PFODBT@HBA-NPs (PFODBT: HBA-COOH = 1: 1, 25 μg mL–1, 200 µL) in different channels, pre-excited with ultrasound (30 kHz, 4.5 W cm–2) for 15 s. g, Ultrasound-induced luminescence intensity of PFODBT@HBA-NPs (6.25 μg mL–1, 200 µL) after different time of ultrasonic excitation (30 kHz, 4.5 W cm–2). h, Ultrasound-induced luminescence intensity of various concentrations of PFODBT@HBA-NPs pre-excited with ultrasound (30 kHz, 4.5 W cm–2) for 15 s. Data for d–g are presented as mean values ± s.d. (n = 3).

Source data

Extended Data Fig. 3 Mechanism for ultrasound-induced luminescence of PFODBT@HBA-NPs.

a, Ultrasound-induced luminescence intensity of HBA-COOH-NPs, PFODBT-NPs, PFODBT@HBA-NPs (50 μg mL–1, 400 µL), pre-excited with ultrasound (30 kHz, 4.5 W cm–2) for 45 s. Data are presented as mean values ± s.d. (n = 3). b, Luminescence intensity of PFODBT@HBA-NPs (12.5 μg mL–1, 100 µL) excited by heating at various temperatures (for example, 10, 20, and 40 oC) or ultrasonic excitation (30 kHz, 4.5 W cm–2 for 15 s). c, Temperatures of PFODBT@HBA-NPs after receiving different times of ultrasonic excitation (30 kHz, 6.5 W cm–2), as measured by an infrared thermal camera. d, Reproducible voltage output of PFODBT when the ultrasonic transducer (30 kHz) was turned on and off. e, ESR spectra of 1O2 generated from PFODBT-NPs (200 µg mL–1, 100 µL) and trapping by 4-oxo-2,2,6,6-tetramethylpiperidine (TEMP) (1 M, 100 µL) before or after ultrasonic excitation. f, ESR spectra of •OH generated from PFODBT-NPs (200 μg mL–1, 100 μL) and trapped by dimethyl-1-pyrroline N-oxide (DMPO) (1 M, 100 μL) in water before or after ultrasonic excitation. g, Schematic diagram of ultrasound-induced luminescence mechanism for PFODBT@HBA-NPs.

Supplementary information

Supplementary Information

Supplementary Tables 1 and 2, Methods, Results and discussion, and Figs. 1–58.

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Source Data Extended Data Fig. 1

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Wang, Y., Yi, Z., Guo, J. et al. In vivo ultrasound-induced luminescence molecular imaging. Nat. Photon. 18, 334–343 (2024). https://doi.org/10.1038/s41566-024-01387-1

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