Label-free nonlinear optical signatures of extracellular vesicles in liquid and tissue biopsies of human breast cancer.
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
06 Mar 2024
06 Mar 2024
Historique:
received:
20
11
2023
accepted:
26
02
2024
medline:
7
3
2024
pubmed:
7
3
2024
entrez:
6
3
2024
Statut:
epublish
Résumé
Extracellular vesicles (EVs) have been implicated in metastasis and proposed as cancer biomarkers. However, heterogeneity and small size makes assessments of EVs challenging. Often, EVs are isolated from biofluids, losing spatial and temporal context and thus lacking the ability to access EVs in situ in their native microenvironment. This work examines the capabilities of label-free nonlinear optical microscopy to extract biochemical optical metrics of EVs in ex vivo tissue and EVs isolated from biofluids in cases of human breast cancer, comparing these metrics within and between EV sources. Before surgery, fresh urine and blood serum samples were obtained from human participants scheduled for breast tumor surgery (24 malignant, 6 benign) or healthy participants scheduled for breast reduction surgery (4 control). EVs were directly imaged both in intact ex vivo tissue that was removed during surgery and in samples isolated from biofluids by differential ultracentrifugation. Isolated EVs and freshly excised ex vivo breast tissue samples were imaged with custom nonlinear optical microscopes to extract single-EV optical metabolic signatures of NAD(P)H and FAD autofluorescence. Optical metrics were significantly altered in cases of malignant breast cancer in biofluid-derived EVs and intact tissue EVs compared to control samples. Specifically, urinary isolated EVs showed elevated NAD(P)H fluorescence lifetime in cases of malignant cancer, serum-derived isolated EVs showed decreased optical redox ratio in stage II cancer, but not earlier stages, and ex vivo breast tissue showed an elevated number of EVs in cases of malignant cancer. Results further indicated significant differences in the measured optical metabolic signature based on EV source (urine, serum and tissue) within individuals.
Identifiants
pubmed: 38448508
doi: 10.1038/s41598-024-55781-4
pii: 10.1038/s41598-024-55781-4
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
5528Subventions
Organisme : U.S. Department of Health and Human Services | NIH | National Institute of Biomedical Imaging and Bioengineering (NIBIB)
ID : T32EB019944
Organisme : U.S. Department of Health and Human Services | NIH | National Institute of Biomedical Imaging and Bioengineering (NIBIB)
ID : P41EB031772
Organisme : U.S. Department of Health and Human Services | NIH | National Cancer Institute (NCI)
ID : R01CA213149
Organisme : U.S. Department of Health and Human Services | NIH | National Cancer Institute (NCI)
ID : R01CA241618
Informations de copyright
© 2024. The Author(s).
Références
Van Niel, G., d’Angelo, G. & Raposo, G. Shedding light on the cell biology of extracellular vesicles. Nat. Rev. Mol. Cell. Biol. 19, 213–228 (2018).
pubmed: 29339798
doi: 10.1038/nrm.2017.125
Becker, A. et al. Extracellular vesicles in cancer: cell-to-cell mediators of metastasis. Cancer Cell 30, 836–848 (2016).
pubmed: 27960084
pmcid: 5157696
doi: 10.1016/j.ccell.2016.10.009
Cooks, T. et al. Mutant p53 cancers reprogram macrophages to tumor supporting macrophages via exosomal miR-1246. Nat. Commun. 9, 771 (2018).
pubmed: 29472616
pmcid: 5823939
doi: 10.1038/s41467-018-03224-w
Park, J. E. et al. Hypoxia-induced tumor exosomes promote M2-like macrophage polarization of infiltrating myeloid cells and microRNA-mediated metabolic shift. Oncogene 38, 5158–5173 (2019).
pubmed: 30872795
doi: 10.1038/s41388-019-0782-x
Nazarenko, I. et al. Cell surface tetraspanin Tspan8 contributes to molecular pathways of exosome-induced endothelial cell activation. Cancer Res. 70, 1668–1678 (2010).
pubmed: 20124479
doi: 10.1158/0008-5472.CAN-09-2470
Huang, M. et al. New insights into the regulatory roles of extracellular vesicles in tumor angiogenesis and their clinical implications. Front. Cell. Dev. Biol. 9, 791882 (2021).
pubmed: 34966744
pmcid: 8710745
doi: 10.3389/fcell.2021.791882
Bertolini, I. et al. Small extracellular vesicle regulation of mitochondrial dynamics reprograms a hypoxic tumor microenvironment. Dev. Cell. 55, 163–177 (2020).
pubmed: 32780991
pmcid: 7606608
doi: 10.1016/j.devcel.2020.07.014
Yu, D. et al. Exosomes as a new frontier of cancer liquid biopsy. Mol Cancer 21(1), 56 (2022).
pubmed: 35180868
pmcid: 8855550
doi: 10.1186/s12943-022-01509-9
Gardiner, C., Ferreira, Y. J., Dragovic, R. A., Redman, C. W. G. & Sargent, I. L. Extracellular vesicle sizing and enumeration by nanoparticle tracking analysis. J. Extracell. Ves. 2(1), 19671 (2013).
doi: 10.3402/jev.v2i0.19671
Rikkert, L. G., Nieuwland, R., Terstappen, L. W. M. M. & Coumans, F. A. W. Quality of extracellular vesicle images by transmission electron microscopy is operator and protocol dependent. J. Extracel. Ves. 8(1), 1555416 (2019).
Palmieri, V. et al. Dynamic light scattering for the characterization and counting of extracellular vesicles: a powerful noninvasive tool. J. Nanoparticle Res. 16, 2583 (2014).
doi: 10.1007/s11051-014-2583-z
Sajidah, E. S. et al. Spatiotemporal tracking of small extracellular vesicle nanotopology in response to physicochemical stresses revealed by HS-AFM. J. Extracell. Ves. 11, 12275 (2022).
doi: 10.1002/jev2.12275
Ferguson, S., Yang, K. S. & Weissleder, R. Single extracellular vesicle analysis for early cancer detection. Trends Mol. Med. 28, 681–692 (2022).
pubmed: 35624008
pmcid: 9339504
doi: 10.1016/j.molmed.2022.05.003
Penders, J. et al. Single particle automated Raman trapping analysis of breast cancer cell-derived extracellular vesicles as cancer biomarkers. ACS Nano 15, 18192–18205 (2021).
pubmed: 34735133
pmcid: 9286313
doi: 10.1021/acsnano.1c07075
Liu, Z., et al. Mapping metabolic changes by noninvasive, multiparametric, high-resolution imaging using endogenous contrast. Sci. Adv. 4, eaap9302 (2018).
Park, J. et al. Exosome classification by pattern analysis of surface-enhanced Raman spectroscopy data for lung cancer diagnosis. Anal. Chem. 89, 6695–6701 (2017).
pubmed: 28541032
doi: 10.1021/acs.analchem.7b00911
Kazemzadeh, M. et al. Label-free classification of bacterial extracellular vesicles by combining nanoplasmonic sensors with machine learning. IEEE Sens. J. 22, 1128–1137 (2021).
doi: 10.1109/JSEN.2021.3131527
Jalali, M. et al. MoS2-Plasmonic nanocavities for Raman spectra of single extracellular vesicles reveal molecular progression in glioblastoma. ACS Nano 17, 12052–12071 (2023).
pubmed: 37366177
pmcid: 10339787
doi: 10.1021/acsnano.2c09222
Smith, Z. J. et al. Single exosome study reveals subpopulations distributed among cell lines with variability related to membrane content. J Extracell. Ves. 4, 28533 (2015).
doi: 10.3402/jev.v4.28533
Nolan, J. P. & Duggan, E. Analysis of individual extracellular vesicles by flow cytometry. Methods Mol. Biol. 1678, 79–92 (2018).
pubmed: 29071676
doi: 10.1007/978-1-4939-7346-0_5
Banijamali, M. et al. Characterizing single extracellular vesicles by droplet barcode sequencing for protein analysis. J. Extracell. Ves. 11, 12277 (2022).
doi: 10.1002/jev2.12277
Linnemannstöns, K. et al. Microscopic and biochemical monitoring of endosomal trafficking and extracellular vesicle secretion in an endogenous in vivo model. J. Extracell. Ves. 11, 12263 (2022).
doi: 10.1002/jev2.12263
Hyenne, V. et al. Studying the fate of tumor extracellular vesicles at high spatiotemporal resolution using the zebrafish embryo. Dev. Cell. 48, 554–572 (2019).
pubmed: 30745140
doi: 10.1016/j.devcel.2019.01.014
You, S. et al. Label-free visualization and characterization of extracellular vesicles in breast cancer. Proc. Natl. Acad. Sci. 116, 24012–24018 (2019).
pubmed: 31732668
pmcid: 6883786
doi: 10.1073/pnas.1909243116
Sorrells, J. E. et al. Label-free characterization of single extracellular vesicles using two-photon fluorescence lifetime imaging microscopy of NAD(P)H. Sci. Rep. 11, 3308 (2021).
pubmed: 33558561
pmcid: 7870923
doi: 10.1038/s41598-020-80813-0
Tu, H. et al. Concurrence of extracellular vesicle enrichment and metabolic switch visualized label-free in the tumor microenvironment. Sci. Adv. 3, e1600675 (2017).
pubmed: 28138543
pmcid: 5266479
doi: 10.1126/sciadv.1600675
Sun, Y., et al. Intraoperative visualization of the tumor microenvironment and quantification of extracellular vesicles by label-free nonlinear imaging. Sci. Adv. 4, eaau5603 (2018).
Park, J. et al. In vivo label-free optical signatures of chemotherapy response in human pancreatic ductal adenocarcinoma patient-derived xenografts. Commun. Biol. 6, 980 (2023).
pubmed: 37749184
pmcid: 10520051
doi: 10.1038/s42003-023-05368-y
Park, J. et al. Label-free optical redox ratio from urinary extracellular vesicles as a screening biomarker for bladder cancer. Am. J. Canc. Res. 12, 2068 (2022).
Borisov, A. V. et al. A criterion of colorectal cancer diagnosis using exosome fluorescence-lifetime imaging. Diagnostics 12, 1792 (2022).
pubmed: 35892503
pmcid: 9394250
doi: 10.3390/diagnostics12081792
Ostrander, J. H. et al. Optical redox ratio differentiates breast cancer cell lines based on estrogen receptor status. Cancer Res. 70, 4759–4766 (2010).
pubmed: 20460512
doi: 10.1158/0008-5472.CAN-09-2572
Podsednik, A., Jiang, J., Jacob, A., Li, L. Z. & Xu, H. N. Optical redox imaging of treatment responses to Nampt inhibition and combination therapy in triple-negative breast cancer cells. Int. J. Mol. Sci. 22, 5563 (2021).
pubmed: 34070254
pmcid: 8197351
doi: 10.3390/ijms22115563
Skala, M. C. et al. In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia. Proc. Natl. Acad. Sci. 104, 19494–19499 (2007).
pubmed: 18042710
pmcid: 2148317
doi: 10.1073/pnas.0708425104
Xu, H. N., Tchou, J., Feng, M., Zhao, H. & Li, L. Z. Optical redox imaging indices discriminate human breast cancer from normal tissues. J. Biomed. Opt. 21, 114003 (2016).
pubmed: 27896360
pmcid: 5136669
doi: 10.1117/1.JBO.21.11.114003
Yaseen, M. A. et al. Fluorescence lifetime microscopy of NADH distinguishes alterations in cerebral metabolism in vivo. Biomed. Opt. Express 8(5), 2368–2385 (2017).
pubmed: 28663879
pmcid: 5480486
doi: 10.1364/BOE.8.002368
Blacker, T. S. et al. Separating NADH and NADPH fluorescence in live cells and tissues using FLIM. Nat. Commun. 5, 3936 (2014).
pubmed: 24874098
doi: 10.1038/ncomms4936
Lakowicz, J. R., Szmacinski, H., Nowaczyk, K. & Johnson, M. L. Fluorescence lifetime imaging of free and protein-bound NADH. Proc. Natl. Acad. Sci. 89(4), 1271–1275 (1992).
pubmed: 1741380
pmcid: 48431
doi: 10.1073/pnas.89.4.1271
Ranjit, S., Malacrida, L., Stakic, M. & Gratton, E. Determination of the metabolic index using the fluorescence lifetime of free and bound nicotinamide adenine dinucleotide using the phasor approach. J. Biophotonics 12(11), e201900156 (2019).
pubmed: 31194290
pmcid: 6842045
doi: 10.1002/jbio.201900156
Lagarto, J. L. et al. Characterization of NAD(P)H and FAD autofluorescence signatures in a Langendorff isolated-perfused rat heart model. Biomed. Opt. Express 9(10), 4931–4978 (2018).
doi: 10.1364/BOE.9.004961
You, S. et al. Intravital imaging by simultaneous label-free autofluorescence-multiharmonic microscopy. Nat. Commun. 9, 2125 (2018).
pubmed: 29844371
pmcid: 5974075
doi: 10.1038/s41467-018-04470-8
Angulo, M. A., Royo, F. & Falcón-Pérez, J. M. Metabolic nano-machines: Extracellular vesicles containing active enzymes and their contribution to liver diseases. Curr. Pathobiol. Rep. 7, 119–127 (2019).
doi: 10.1007/s40139-019-00197-3
Aksamitiene, E., Hoek, J. B. & Kiyatkin, A. Multistrip Western blotting: a tool for comparative quantitative analysis of multiple proteins. Methods Mol. Biol. 1312, 197–226 (2015).
pubmed: 26044004
doi: 10.1007/978-1-4939-2694-7_23
Théry, C. et al. Minimal information for studies of extracellular vesicles 2018 (MISEV2018): A position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines. J. Extracell. Ves. 7(1), 1535750 (2018).
doi: 10.1080/20013078.2018.1535750
Digman, M. A., Caiolfa, V. R., Zamai, M. & Gratton, E. The phasor approach to fluorescence lifetime imaging analysis. Biophys. J. 94, L14–L16 (2008).
pubmed: 17981902
doi: 10.1529/biophysj.107.120154
Sharick, J. T. et al. Protein-bound NAD(P)H lifetime is sensitive to multiple fates of glucose carbon. Sci. Rep. 8, 5456 (2018).
pubmed: 29615678
pmcid: 5883019
doi: 10.1038/s41598-018-23691-x
Hanahan, D. Hallmarks of cancer: New dimensions. Cancer Discov. 12, 31–46 (2022).
pubmed: 35022204
doi: 10.1158/2159-8290.CD-21-1059
Ward, P. S. & Thompson, C. B. Metabolic reprogramming: A cancer hallmark even Warburg did not anticipate. Cancer Cell 21, 297–308 (2012).
pubmed: 22439925
pmcid: 3311998
doi: 10.1016/j.ccr.2012.02.014
Svenningsen, P., Sabaratnam, R. & Jensen, B. L. Urinary extracellular vesicles: Origin, role as intercellular messengers and biomarkers; efficient sorting and potential treatment options. Acta Physiol. 228, e13346 (2020).
doi: 10.1111/apha.13346
Lourenço, C., Constâncio, V., Henrique, R., Carvalho, Â. & Jerónimo, C. Urinary extracellular vesicles as potential biomarkers for urologic cancers: An overview of current methods and advances. Cancers 13, 1529 (2021).
pubmed: 33810357
pmcid: 8036842
doi: 10.3390/cancers13071529
Li, Y., Zhang, Y., Qiu, F. & Qiu, Z. Proteomic identification of exosomal LRG1: A potential urinary biomarker for detecting NSCLC. Electrophoresis 32, 1976–1983 (2011).
pubmed: 21557262
doi: 10.1002/elps.201000598
Hoshino, A. et al. Extracellular vesicle and particle biomarkers define multiple human cancers. Cell 182, 1044–1061 (2020).
pubmed: 32795414
pmcid: 7522766
doi: 10.1016/j.cell.2020.07.009
Fang, S. et al. Clinical application of a microfluidic chip for immunocapture and quantification of circulating exosomes to assist breast cancer diagnosis and molecular classification. PloS One 12, e0175050 (2017).
pubmed: 28369094
pmcid: 5378374
doi: 10.1371/journal.pone.0175050
Hinestrosa, J. P. et al. Early-stage multi-cancer detection using an extracellular vesicle protein-based blood test. Commun. Med. 2, 29 (2022).
pubmed: 35603292
pmcid: 9053211
doi: 10.1038/s43856-022-00088-6
Lee, J. U., Kim, W. H., Lee, H. S., Park, K. H. & Sim, S. J. Quantitative and specific detection of exosomal miRNAs for accurate diagnosis of breast cancer using a surface-enhanced Raman scattering sensor based on plasmonic head-flocked gold nanopillars. Small 15, 1804968 (2019).
doi: 10.1002/smll.201804968
King, H. W., Michael, M. Z. & Gleadle, J. M. Hypoxic enhancement of exosome release by breast cancer cells. BMC Cancer 12, 421 (2012).
pubmed: 22998595
pmcid: 3488584
doi: 10.1186/1471-2407-12-421
Shcheslavskiy, V. I., Saltiel, S. M., Faustov, A., Petrov, G. I. & Yakovlev, V. V. Third-harmonic Rayleigh scattering: theory and experiment. JOSA B 22, 2402–2408 (2005).
doi: 10.1364/JOSAB.22.002402
Minciacchi, V. R. et al. Large oncosomes contain distinct protein cargo and represent a separate functional class of tumor-derived extracellular vesicles. Oncotarget 6, 11327 (2015).
pubmed: 25857301
pmcid: 4484459
doi: 10.18632/oncotarget.3598
Schürz, M. et al. EVAnalyzer: High content imaging for rigorous characterization of single extracellular vesicles using standard laboratory equipment and a new open-source ImageJ/Fiji plugin. J. Extracell. Ves. 11, 12282 (2022).
doi: 10.1002/jev2.12282
Sorrells J. Nonlinear optical imaging of breast cancer EVs. Open Science Framework, 2023. https://osf.io/d37ac/?view_only=523ce62c46aa4f3b8524c1b13a2c4255 .