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Sudden cardiac death, arising from abnormal electrical conduction, occurs frequently in patients with coronary heart disease. Myocardial ischemia simultaneously induces arrhythmia and massive myocardial leukocyte changes. In this study, we optimized a mouse model in which hypokalemia combined with myocardial infarction triggered spontaneous ventricular tachycardia in ambulatory mice, and we showed that major leukocyte subsets have opposing effects on cardiac conduction. Neutrophils increased ventricular tachycardia via lipocalin-2 in mice, whereas neutrophilia associated with ventricular tachycardia in patients. In contrast, macrophages protected against arrhythmia. Depleting recruited macrophages in Ccr2−/− mice or all macrophage subsets with Csf1 receptor inhibition increased both ventricular tachycardia and fibrillation. Higher arrhythmia burden and mortality in Cd36−/− and Mertk−/− mice, viewed together with reduced mitochondrial integrity and accelerated cardiomyocyte death in the absence of macrophages, indicated that receptor-mediated phagocytosis protects against lethal electrical storm. Thus, modulation of leukocyte function provides a potential therapeutic pathway for reducing the risk of sudden cardiac death.
@article{2023-SCIENCE,author={Hulsmans, M. and Schloss, M.J. and Lee, I.H. and Bapat, A. and Iwamoto, Y. and Vinegoni, C. and Paccalet, A. and Yamazoe M. and Grune, J. and Pabel, S. and Momin, N. and Seung, H. and Kumowski, N. and Pulous, F.E. and Keller, D. and Bening C. and Green, U. and Lennerz, J.K. and Mitchell, R.N. and Lewis, A. and Casadei, B. and Iborra-Egea, O. and Bayes-Genis, A. and Sossalla, S. and Ong, C.S. and Pierson, R.N. and Aster, J.C. and Rohde, D. and Wojtkiewicz, G.R. and Weissleder, R. and Swirski, F.K. and Tellides, G. and Tolis, G. and Melnitchouk, S. and Milan, D.J. and Ellinor, P.T. and Naxerova, K. and Nahrendorf, M.},title={Recruited macrophages elicit atrial fibrillation},journal={Science},alternatejournal={Sci.},year={2023},volume={381},number={6654},pages={231–239},doi={10.1126/science.abq3061},}
37440641
10.1126/science.abq3061
2022
Nat. Card. Res.
Neutrophils incite and macrophages avert electrical storm after myocardial infarction
Grune, J.,
Lewis, A.J.M.,
Yamazoe, M.,
Hulsmans, M.,
Rohde, D.,
Xiao, L.,
Zhang, S.,
Ott, C.,
Calcagno, D.M.,
Zhou, Y.,
Timm, K.,
Shanmuganathan, M.,
Pulous, F.E.,
Schloss, M.J.,
Foy, B.H.,
Capen, D.,
Vinegoni, C.,
Wojtkiewicz, G.R.,
Iwamoto, I.,
Grune, T.,
Brown, D.,
Higgins, J.,
Ferreira, V.M.,
Herring, N.,
Channon, K.M.,
Neubauer, S.,
Study, Oxford Acute Myocardial Infarction (OxAMI),
Sosnovik, D.E.,
Milan, D.J.,
Swirski, F.K.,
King, K.R.,
Aguirre, A.D.,
Ellinor, P.T.,
and Nahrendorf#, M.
Sudden cardiac death, arising from abnormal electrical conduction, occurs frequently in patients with coronary heart disease. Myocardial ischemia simultaneously induces arrhythmia and massive myocardial leukocyte changes. In this study, we optimized a mouse model in which hypokalemia combined with myocardial infarction triggered spontaneous ventricular tachycardia in ambulatory mice, and we showed that major leukocyte subsets have opposing effects on cardiac conduction. Neutrophils increased ventricular tachycardia via lipocalin-2 in mice, whereas neutrophilia associated with ventricular tachycardia in patients. In contrast, macrophages protected against arrhythmia. Depleting recruited macrophages in Ccr2−/− mice or all macrophage subsets with Csf1 receptor inhibition increased both ventricular tachycardia and fibrillation. Higher arrhythmia burden and mortality in Cd36−/− and Mertk−/− mice, viewed together with reduced mitochondrial integrity and accelerated cardiomyocyte death in the absence of macrophages, indicated that receptor-mediated phagocytosis protects against lethal electrical storm. Thus, modulation of leukocyte function provides a potential therapeutic pathway for reducing the risk of sudden cardiac death.
@article{2022-NCR-2,author={Grune, J. and Lewis, A.J.M. and Yamazoe, M. and Hulsmans, M. and Rohde, D. and Xiao, L. and Zhang, S. and Ott, C. and Calcagno, D.M. and Zhou, Y. and Timm, K. and Shanmuganathan, M. and Pulous, F.E. and Schloss, M.J. and Foy, B.H. and Capen, D. and Vinegoni, C. and Wojtkiewicz, G.R. and Iwamoto, I. and Grune, T. and Brown, D. and Higgins, J. and Ferreira, V.M. and Herring, N. and Channon, K.M. and Neubauer, S. and Oxford Acute Myocardial Infarction (OxAMI) Study and Sosnovik, D.E. and Milan, D.J. and Swirski, F.K. and King, K.R. and Aguirre, A.D. and Ellinor, P.T. and Nahrendorf, M.},title={Neutrophils incite and macrophages avert electrical storm after myocardial infarction},journal={Nature Cardiovascular Research},alternatejournal={Nat. Card. Res.},year={2022},volume={1},number={1},pages={649–664},doi={10.1038/s44161-022-00094-w},}
36034743
PMC9410341
10.1038/s44161-022-00094-w
Nat. Biotechnol.
Spatiotemporal multiplexed immunofluorescence imaging of living cells and tissues with bioorthogonal cycling of fluorescent probes
Ko, J. A.,
Wilkovitsch, M.,
Oh, J.,
Kohler, R. H.,
Bolli, E.,
Pittet, M. J.,
Vinegoni, C.,
Sykes, D. B.,
Mikula, H.,
Weissleder#, R.,
and Carlson#, J. C. T.
Cells in complex organisms undergo frequent functional changes, but few methods allow comprehensive longitudinal profiling of living cells. Here we introduce scission-accelerated fluorophore exchange (SAFE), a method for multiplexed temporospatial imaging of living cells with immunofluorescence. SAFE uses a rapid bioorthogonal click chemistry to remove immunofluorescent signals from the surface of labeled cells, cycling the nanomolar-concentration reagents in seconds and enabling multiple rounds of staining of the same samples. It is non-toxic and functional in both dispersed cells and intact living tissues. We demonstrate multiparameter (n >= 14), non-disruptive imaging of murine peripheral blood mononuclear and bone marrow cells to profile cellular differentiation. We also show longitudinal multiplexed imaging of bone marrow progenitor cells as they develop into neutrophils over 6 days and real-time multiplexed cycling of living mouse hepatic tissues. We anticipate that SAFE will find broad utility for investigating physiologic dynamics in living systems. Live cells and tissues are imaged over long time periods using a fast, non-toxic click chemistry.
@article{2022-NB,author={Ko, J. A. and Wilkovitsch, M. and Oh, J. and Kohler, R. H. and Bolli, E. and Pittet, M. J. and Vinegoni, C. and Sykes, D. B. and Mikula, H. and Weissleder<sup>#</sup>, R. and Carlson<sup>#</sup>, J. C. T.},title={Spatiotemporal multiplexed immunofluorescence imaging of living cells and tissues with bioorthogonal cycling of fluorescent probes},journal={Nature Biotechnology},alternatejournal={Nat. Biotechnol.},year={1998},pages={16},issn={1087-0156},doi={10.1038/s41587-022-01339-6},pmid={35654978}}
35654978
10.1038/s41587-022-01339-6
Nat. Neurosci.
Cerebrospinal fluid can exit into the skull bone marrow and instruct cranial hematopoiesis in mice with bacterial meningitis
Pulous, F. E.,
Cruz-Hernandez, J. C.,
Yang, C. B.,
Kaya, Z.,
Paccalet, A.,
Wojtkiewicz, G.,
Capen, D.,
Brown, D.,
Wu, J. W.,
Schloss, M. J.,
Vinegoni, C.,
Richter, D.,
Yamazoe, M.,
Hulsmans, M.,
Momin, N.,
Grune, J.,
Rohde, D.,
McAlpine, C. S.,
Panizzi, P.,
Weissleder, R.,
Kim, D. E.,
Swirski, F. K.,
Lin, C. P.,
Moskowitz, M. A.,
and Nahrendorf#, M.
Interactions between the immune and central nervous systems strongly influence brain health. Although the blood-brain barrier restricts this crosstalk, we now know that meningeal gateways through brain border tissues facilitate intersystem communication. Cerebrospinal fluid (CSF), which interfaces with the glymphatic system and thereby drains the brain’s interstitial and perivascular spaces, facilitates outward signaling beyond the blood-brain barrier. In the present study, we report that CSF can exit into the skull bone marrow. Fluorescent tracers injected into the cisterna magna of mice migrate along perivascular spaces of dural blood vessels and then travel through hundreds of sub-millimeter skull channels into the calvarial marrow. During meningitis, bacteria hijack this route to invade the skull’s hematopoietic niches and initiate cranial hematopoiesis ahead of remote tibial sites. As skull channels also directly provide leukocytes to meninges, the privileged sampling of brain-derived danger signals in CSF by regional marrow may have broad implications for inflammatory neurological disorders. This manuscript describes a new cerebral spinal fluid exit route via hundreds of skull channels, with the cranial bone marrow as a destination. In meningitis, bacteria hijack this path and alert hematopoietic stem cells residing in the skull marrow.
@article{2022-NN,author={Pulous, F. E. and Cruz-Hernandez, J. C. and Yang, C. B. and Kaya, Z. and Paccalet, A. and Wojtkiewicz, G. and Capen, D. and Brown, D. and Wu, J. W. and Schloss, M. J. and Vinegoni, C. and Richter, D. and Yamazoe, M. and Hulsmans, M. and Momin, N. and Grune, J. and Rohde, D. and McAlpine, C. S. and Panizzi, P. and Weissleder, R. and Kim, D. E. and Swirski, F. K. and Lin, C. P. and Moskowitz, M. A. and Nahrendorf<sup>#</sup>, M.},title={Cerebrospinal fluid can exit into the skull bone marrow and instruct cranial hematopoiesis in mice with bacterial meningitis},journal={Nature Neuroscience},alternatejournal={Nat. Neurosci.},year={2022},volume={25},number={5},pages={567-+},issn={1097-6256},doi={10.1038/s41593-022-01060-2},pmcid={PMC9081225},pmid={35501382}}
35501382
PMC9081225
10.1038/s41593-022-01060-2
Nat. Card. Res.
Bone marrow endothelial dysfunction promotes myeloid cell expansion in cardiovascular disease
Abnormal hematopoiesis advances cardiovascular disease by generating excess inflammatory leukocytes that attack the arteries and the heart. The bone marrow niche regulates hematopoietic stem cell proliferation and hence the systemic leukocyte pool, but whether cardiovascular disease affects the hematopoietic organ’s microvasculature is unknown. Here we show that hypertension, atherosclerosis and myocardial infarction (MI) instigate endothelial dysfunction, leakage, vascular fibrosis and angiogenesis in the bone marrow, altogether leading to overproduction of inflammatory myeloid cells and systemic leukocytosis. Limiting angiogenesis with endothelial deletion of Vegfr2 (encoding vascular endothelial growth factor (VEGF) receptor 2) curbed emergency hematopoiesis after MI. We noted that bone marrow endothelial cells assumed inflammatory transcriptional phenotypes in all examined stages of cardiovascular disease. Endothelial deletion of Il6 or Vcan (encoding versican), genes shown to be highly expressed in mice with atherosclerosis or MI, reduced hematopoiesis and systemic myeloid cell numbers in these conditions. Our findings establish that cardiovascular disease remodels the vascular bone marrow niche, stimulating hematopoiesis and production of inflammatory leukocytes.
Astrocytic interleukin-3 programs microglia and limits Alzheimer’s disease
McAlpine, C. S.,
Park, J.,
Griciuc, A.,
Kim, E.,
Choi, S. H.,
Iwamoto, Y.,
Kiss, M. G.,
Christie, K. A.,
Vinegoni, C.,
Poller, W. C.,
Mindur, J. E.,
Chan, C. T.,
He, S.,
Janssen, H.,
Wong, L. P.,
Downey, J.,
Singh, S.,
Anzai, A.,
Kahles, F.,
Jorfi, M.,
Feruglio, P. F.,
Sadreyev, R. I.,
Weissleder, R.,
Kleinstiver, B. P.,
Nahrendorf, M.,
Tanzi, R. E.,
and Swirski#, F. K.
Communication within the glial cell ecosystem is essential for neuronal and brain health(1-3). The influence of glial cells on the accumulation and clearance of beta-amyloid (A beta) and neurofibrillary tau in the brains of individuals with Alzheimer’s disease (AD) is poorly understood, despite growing awareness that these are therapeutically important interactions(4,5). Here we show, in humans and mice, that astrocyte-sourced interleukin-3 (IL-3) programs microglia to ameliorate the pathology of AD. Upon recognition of A beta deposits, microglia increase their expression of IL-3R alpha-the specific receptor for IL-3 (also known as CD123)-making them responsive to IL-3. Astrocytes constitutively produce IL-3, which elicits transcriptional, morphological, and functional programming of microglia to endow them with an acute immune response program, enhanced motility, and the capacity to cluster and clear aggregates of A beta and tau. These changes restrict AD pathology and cognitive decline. Our findings identify IL-3 as a key mediator of astrocyte-microglia cross-talk and a node for therapeutic intervention in AD.
@article{2021-NATURE,author={McAlpine, C. S. and Park, J. and Griciuc, A. and Kim, E. and Choi, S. H. and Iwamoto, Y. and Kiss, M. G. and Christie, K. A. and Vinegoni, C. and Poller, W. C. and Mindur, J. E. and Chan, C. T. and He, S. and Janssen, H. and Wong, L. P. and Downey, J. and Singh, S. and Anzai, A. and Kahles, F. and Jorfi, M. and Feruglio, P. F. and Sadreyev, R. I. and Weissleder, R. and Kleinstiver, B. P. and Nahrendorf, M. and Tanzi, R. E. and Swirski<sup>#</sup>, F. K.},title={Astrocytic interleukin-3 programs microglia and limits Alzheimer’s disease},journal={Nature},alternatejournal={Nature},year={2021},volume={595},number={7869},pages={701-+},issn={0028-0836},doi={10.1038/s41586-021-03734-6},pmcid={PMC8934148},pmid={34262178}}
34262178
PMC8934148
10.1038/s41586-021-03734-6
2020
J. Am. Coll. Cardiol.
Diminished Reactive Hematopoiesis and Cardiac Inflammation in a Mouse Model of Recurrent Myocardial Infarction
Cremer, S.,
Schloss, M. J.,
Vinegoni, C.,
Foy, B. H.,
Zhang, S.,
Rohde, D.,
Hulsmans, M.,
Feruglio, P. F.,
Schmidt, S.,
Wojtkiewicz, G.,
Higgins, J. M.,
Weissleder, R.,
Swirski, F. K.,
and Nahrendorf#, M.
Journal of the American College of Cardiology 2020
BACKGROUND Recurrent myocardial infarction (MI) is common in patients with coronary artery disease and is associated with high mortality. Long-term reprogramming of myeloid progenitors occurs in response to inflammatory stimuli and alters the organism’s response to secondary inflammatory challenges. OBJECTIVES This study examined the effect of recurrent MI on bone marrow response and cardiac inflammation. METHODS The investigators developed a surgical mouse model in which 2 subsequent MIs affected different left ventricular regions in the same mouse. Recurrent MI was induced by ligating the left circumflex artery followed by the left anterior descending coronary artery branch. The study characterized the resulting ischemia by whole-heart fluorescent coronary angiography after optical organ clearing and by cardiac magnetic resonance imaging. RESULTS A first MI-induced bone marrow "memory" via a drculating signal, reducing hematopoietic maintenance factor expression in bone marrow macrophages. This dampened the organism’s reaction to subsequent events. Despite a similar extent of injury according to troponin levels, recurrent MI caused reduced emergency hematopoiesis and less leukocytosis than a first MI. Consequently, fewer leukocytes migrated to the ischemic myocardium. The hematopoietic response to lipopolysaccharide was also mitigated after a previous ML The increase of white blood count in 28 patients was lower after recurrent MI compared with their first MI. CONCLUSIONS The data suggested that hematopoietic and innate immune responses are shaped by a preceding MI. (C) 2020 by the American College of Cardiology Foundation.
@article{2020-JAMCC,author={Cremer, S. and Schloss, M. J. and Vinegoni, C. and Foy, B. H. and Zhang, S. and Rohde, D. and Hulsmans, M. and Feruglio, P. F. and Schmidt, S. and Wojtkiewicz, G. and Higgins, J. M. and Weissleder, R. and Swirski, F. K. and Nahrendorf<sup>#</sup>, M.},title={Diminished Reactive Hematopoiesis and Cardiac Inflammation in a Mouse Model of Recurrent Myocardial Infarction},journal={Journal of the American College of Cardiology},alternatejournal={J. Am. Coll. Cardiol.},year={2020},volume={75},number={8},pages={901-915},issn={0735-1097},doi={10.1016/j.jacc.2019.12.056},pmcid={PMC7254576},pmid={32130926}}
32130926
PMC7254576
10.1016/j.jacc.2019.12.056
J. Biomed. Opt.
Extended dynamic range imaging for noise mitigation in fluorescence anisotropy imaging
Feruglio†, P. F.,
Vinegoni#†, C.,
and Weissleder, R.
Significance: Fluorescence polarization (FP) and fluorescence anisotropy (FA) microscopy are powerful imaging techniques that allow to translate the common IT assay capabilities into the in vitro and in vivo cellular domain. As a result, they have found potential for mapping drug- protein or protein-protein interactions. Unfortunately, these imaging modalities are ratiometric in nature and as such they suffer from excessive noise even under regular imaging conditions, preventing accurate image-feature analysis of fluorescent molecules behaviors. Aim: We present a high dynamic range (HDR)-based FA imaging modality for improving image quality in FA microscopy. Approach: The method exploits ad hoc acquisition schemes to extend the dynamic range of individual FP channels, allowing to obtain FA images with increased signal-to-noise ratio. Results: A direct comparison between FA images obtained with our method and the standard, clearly indicates how an HDR-based FA imaging approach allows to obtain high-quality images, with the ability to correctly resolve image features at different values of FA and over a substantially higher range of fluorescence intensities. Conclusion: The method presented is shown to outperform standard FA imaging microscopy narrowing the spread of the propagated error and yielding higher quality images. The method can be effectively and routinely used on any commercial imaging system and could be also translated to other microscopy ratiometric imaging modalities. (C) The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License.
@article{2020-JBO,author={Feruglio<sup>†</sup>, P. F. and Vinegoni<sup>#†</sup>, C. and Weissleder, R.},title={Extended dynamic range imaging for noise mitigation in fluorescence anisotropy imaging},journal={Journal of Biomedical Optics},alternatejournal={J. Biomed. Opt.},year={2020},volume={25},number={8},pages={13},issn={1083-3668},doi={10.1117/1.Jbo.25.8.086003},pmcid={PMC7439791},pmid={32820624}}
32820624
PMC7439791
10.1117/1.Jbo.25.8.086003
Sci. Transl. Med.
CytoPAN-Portable cellular analyses for rapid point-of-care cancer diagnosis
Min, J.,
Chin, L. K.,
Oh, J.,
Landeros, C.,
Vinegoni, C.,
Lee, J.,
Lee, S. J.,
Park, J. Y.,
Liu, A. Q.,
Castro, C. M.,
Lee, H.,
Im, H.,
and Weissleder#, R.
Rapid, automated, point-of-care cellular diagnosis of cancer remains difficult in remote settings due to lack of specialists and medical infrastructure. To address the need for same-day diagnosis, we developed an automated image cytometry system (CytoPAN) that allows rapid breast cancer diagnosis of scant cellular specimens obtained by fine needle aspiration (FNA) of palpable mass lesions. The system is devoid of moving parts for stable operations, harnesses optimized antibody kits for multiplexed analysis, and offers a user-friendly interface with automated analysis for rapid diagnoses. Through extensive optimization and validation using cell lines and mouse models, we established breast cancer diagnosis and receptor subtyping in 1 hour using as few as 50 harvested cells. In a prospective patient cohort study (n = 68), we showed that the diagnostic accuracy was 100% for cancer detection and the receptor subtyping accuracy was 96% for human epidermal growth factor receptor 2 and 93% for hormonal receptors (ER/PR), two key biomarkers associated with breast cancer. A combination of FNA and CytoPAN offers faster, less invasive cancer diagnoses than the current standard (core biopsy and histopathology). This approach should enable the ability to more rapidly diagnose breast cancer in global and remote settings.
@article{2020-STM,author={Min, J. and Chin, L. K. and Oh, J. and Landeros, C. and Vinegoni, C. and Lee, J. and Lee, S. J. and Park, J. Y. and Liu, A. Q. and Castro, C. M. and Lee, H. and Im, H. and Weissleder<sup>#</sup>, R.},title={CytoPAN-Portable cellular analyses for rapid point-of-care cancer diagnosis},journal={Science Translational Medicine},alternatejournal={Sci. Transl. Med.},year={2020},volume={12},number={555},pages={14},issn={1946-6234},doi={10.1126/scitranslmed.aaz9746},pmcid={PMC8217912},pmid={32759277}}
32759277
PMC8217912
10.1126/scitranslmed.aaz9746
Sci. Rep.
Fluorescence microscopy tensor imaging representations for large-scale dataset analysis
Vinegoni#†, C.,
Feruglio†, P. F.,
Courties, G.,
Schmidt, S.,
Hulsmans, M.,
Lee, S.,
Wang, R.,
Sosnovik, D.,
Nahrendorf, M.,
and Weissleder, R.
Understanding complex biological systems requires the system-wide characterization of cellular and molecular features. Recent advances in optical imaging technologies and chemical tissue clearing have facilitated the acquisition of whole-organ imaging datasets, but automated tools for their quantitative analysis and visualization are still lacking. We have here developed a visualization technique capable of providing whole-organ tensor imaging representations of local regional descriptors based on fluorescence data acquisition. This method enables rapid, multiscale, analysis and virtualization of large-volume, high-resolution complex biological data while generating 3D tractographic representations. Using the murine heart as a model, our method allowed us to analyze and interrogate the cardiac microvasculature and the tissue resident macrophage distribution and better infer and delineate the underlying structural network in unprecedented detail.
@article{2020-SCIREP,author={Vinegoni<sup>#†</sup>, C. and Feruglio<sup>†</sup>, P. F. and Courties, G. and Schmidt, S. and Hulsmans, M. and Lee, S. and Wang, R. and Sosnovik, D. and Nahrendorf, M. and Weissleder, R.},title={Fluorescence microscopy tensor imaging representations for large-scale dataset analysis},journal={Scientific Reports},alternatejournal={Sci. Rep.},year={2020},volume={10},number={1},pages={15},issn={2045-2322},doi={10.1038/s41598-020-62233-2},pmcid={PMC7101442},pmid={32221334}}
32221334
PMC7101442
10.1038/s41598-020-62233-2
2019
Neuro-Oncology
Characterization of single microvesicles in plasma from glioblastoma patients
Fraser, K.,
Jo, A.,
Giedt, J.,
Vinegoni, C.,
Yang, K. S.,
Peruzzi, P.,
Chiocca, E. A.,
Breakefield, X. O.,
Lee, H.,
and Weissleder#, R.
Background. Extracellular vesicles (EV) are shed by tumor cells but little is known about their individual molecular phenotypes and heterogeneity. While exosomes have received considerable attention, much less is known about larger microvesicles. Here we profile single microvesicles (MV) and exosomes from glioblastoma (GB) cells and MV from the plasma of patients. Methods. EV secreted from mouse glioma GL261 and human primary GBM8 cell lines as well as from the plasma of 8 patients with diagnoses of GB and 2 healthy controls were isolated and processed for single vesicle analysis. EV were immobilized on glass slides and the heterogeneity of vesicle and tumor markers were analyzed at the single vesicle level. Results. We show that (i) MV are abundant, (ii) only a minority of MV expresses putative MV markers, and (iii) MV share tetraspanin biomarkers previously thought to be diagnostic of exosomes. Using MV capture and staining techniques that allow differentiation of host cell and GB-derived MV we further demonstrate that (i) tumoral MV often present as <10% of all MV in GB patient plasma, and (ii) there is extensive heterogeneity in tumor marker expression in these tumor-derived MV. Conclusion. These results indicate that single MV analysis is likely necessary to identify rare tumoral MV populations and the single vesicle analytical technique used here can be applied to both MV and exosome fractions without the need for their separation from each other. These studies form the basis for using single EV analyses for cancer diagnostics.
@article{2019-NO,author={Fraser, K. and Jo, A. and Giedt, J. and Vinegoni, C. and Yang, K. S. and Peruzzi, P. and Chiocca, E. A. and Breakefield, X. O. and Lee, H. and Weissleder<sup>#</sup>, R.},title={Characterization of single microvesicles in plasma from glioblastoma patients},journal={Neuro-Oncology},alternatejournal={Neuro-Oncology},year={2019},volume={21},number={5},pages={606-615},issn={1522-8517},doi={10.1093/neuonc/noy187},pmcid={6502493},pmid={30561734}}
30561734
6502493
10.1093/neuonc/noy187
Adv. Drug Deliv. Rev.
Fluorescence anisotropy imaging in drug discovery
Vinegoni#, C.,
Feruglio, P. F.,
Gryczynski, I.,
Mazitschek, R.,
and Weissleder, R.
Non-invasive measurement of drug-target engagement can provide critical insights in the molecular pharmacology of small molecule drugs. Fluorescence polarization/fluorescence anisotropy measurements are commonly employed in protein/cell screening assays. However, the expansion of such measurements to the in vivo setting has proven difficult until recently. With the advent of high-resolution fluorescence anisotropy microscopy it is now possible to perform kinetic measurements of intracellular drug distribution and target engagement in commonly used mouse models. In this review we discuss the background, current advances and future perspectives in intravital fluorescence anisotropy measurements to derive pharmacokinetic and pharmacodynamic measurements in single cells and whole organs. (C) 2018 Elsevier B.V. All rights reserved.
@article{2019-ADDR,author={Vinegoni<sup>#</sup>, C. and Feruglio, P. F. and Gryczynski, I. and Mazitschek, R. and Weissleder, R.},title={Fluorescence anisotropy imaging in drug discovery},journal={Advanced Drug Delivery Reviews},alternatejournal={Adv. Drug Deliv. Rev.},year={2019},volume={151},pages={262-288},issn={0169-409X},doi={10.1016/j.addr.2018.01.019},pmcid={6072632},pmid={29410158}}
29410158
6072632
10.1016/j.addr.2018.01.019
IEEE J.S.T.Q.E.
High Dynamic Range Fluorescence Imaging
Vinegoni#, C.,
Feruglio, P. F.,
and Weissleder, R.
Ieee Journal of Selected Topics in Quantum Electronics 2019
Fluorescence acquisition and image display over a high dynamic range is highly desirable. However, the limited dynamic range of current photodetectors and imaging charge-coupled devices impose a limit on the fluorescence intensities that can be simultaneously captured during a single image acquisition. This is particularly troublesome when imaging biological samples, where protein expression fluctuates considerably. As a result, biological images will often contain regions with signal that is either saturated or hidden within background noise, causing information loss. In this paper, we summarize recent work from our group and others, to extended conventional to high dynamic range fluorescence imaging. These strategies have many biological applications, such as mapping of neural connections, vascular imaging, biodistribution studies or pharmacologic imaging at the single cell and organ level.
@article{2019-JSTQE,author={Vinegoni<sup>#</sup>, C. and Feruglio, P. F. and Weissleder, R.},title={High Dynamic Range Fluorescence Imaging},journal={Ieee Journal of Selected Topics in Quantum Electronics},alternatejournal={IEEE J.S.T.Q.E.},year={2019},volume={25},number={1},pages={7},issn={1077-260X},doi={10.1109/jstqe.2018.2881608},pmcid={PMC6785194},pmid={31598059}}
31598059
PMC6785194
10.1109/jstqe.2018.2881608
2018
Nat. Neurosci.
Direct vascular channels connect skull bone marrow and the brain surface enabling myeloid cell migration
Herisson, F.,
Frodermann, V.,
Courties, G.,
Rohde, D.,
Sun, Y.,
Vandoorne, K.,
Wojtkiewicz, G. R.,
Masson, G. S.,
Vinegoni, C.,
Kim, J.,
Kim, D. E.,
Weissleder, R.,
Swirski, F. K.,
Moskowitz, M. A.,
and Nahrendorf#, M.
Innate immune cells recruited to inflammatory sites have short life spans and originate from the marrow, which is distributed throughout the long and flat bones. While bone marrow production and release of leukocyte increases after stroke, it is currently unknown whether its activity rises homogeneously throughout the entire hematopoietic system. To address this question, we employed spectrally resolved in vivo cell labeling in the murine skull and tibia. We show that in murine models of stroke and aseptic meningitis, skull bone marrow-derived neutrophils are more likely to migrate to the adjacent brain tissue than cells that reside in the tibia. Confocal microscopy of the skull-dura interface revealed myeloid cell migration through microscopic vascular channels crossing the inner skull cortex. These observations point to a direct local interaction between the brain and the skull bone marrow through the meninges.
@article{2018-NATNEURO,author={Herisson, F. and Frodermann, V. and Courties, G. and Rohde, D. and Sun, Y. and Vandoorne, K. and Wojtkiewicz, G. R. and Masson, G. S. and Vinegoni, C. and Kim, J. and Kim, D. E. and Weissleder, R. and Swirski, F. K. and Moskowitz, M. A. and Nahrendorf<sup>#</sup>, M.},title={Direct vascular channels connect skull bone marrow and the brain surface enabling myeloid cell migration},journal={Nature Neuroscience},alternatejournal={Nat. Neurosci.},year={2018},volume={21},number={9},pages={1209-+},issn={1097-6256},doi={10.1038/s41593-018-0213-2},pmcid={PMC6148759 },pmid={30150661}}
30150661
PMC6148759
10.1038/s41593-018-0213-2
J. Exp. Med.
Cardiac macrophages promote diastolic dysfunction
Hulsmans, M.,
Sager, H. B.,
Roh, J. D.,
Valero-Munoz, M.,
Houstis, N. E.,
Iwamoto, Y.,
Sun, Y.,
Wilson, R. M.,
Wojtkiewicz, G.,
Tricot, B.,
Osborne, M. T.,
Hung, J.,
Vinegoni, C.,
Naxerova, K.,
Sosnovik, D. E.,
Zile, M. R.,
Bradshaw, A. D.,
Liao, R.,
Tawakol, A.,
Weissleder, R.,
Rosenzweig, A.,
Swirski, F. K.,
Sam, F.,
and Nahrendorf#, M.
Macrophages populate the healthy myocardium and, depending on their phenotype, may contribute to tissue homeostasis or disease. Their origin and role in diastolic dysfunction, a hallmark of cardiac aging and heart failure with preserved ejection fraction, remain unclear. Here we show that cardiac macrophages expand in humans and mice with diastolic dysfunction, which in mice was induced by either hypertension or advanced age. A higher murine myocardial macrophage density results from monocyte recruitment and increased hematopoiesis in bone marrow and spleen. In humans, we observed a parallel constellation of hematopoietic activation: circulating myeloid cells are more frequent, and splenic F-18-FDGPET/CT imaging signal correlates with echocardiographic indices of diastolic dysfunction. While diastolic dysfunction develops, cardiac macrophages produce IL-10, activate fibroblasts, and stimulate collagen deposition, leading to impaired myocardial relaxation and increased myocardial stiffness. Deletion of IL-10 in macrophages improves diastolic function. These data imply expansion and phenotypic changes of cardiac macrophages as therapeutic targets for cardiac fibrosis leading to diastolic dysfunction.
@article{2018-JEM,author={Hulsmans, M. and Sager, H. B. and Roh, J. D. and Valero-Munoz, M. and Houstis, N. E. and Iwamoto, Y. and Sun, Y. and Wilson, R. M. and Wojtkiewicz, G. and Tricot, B. and Osborne, M. T. and Hung, J. and Vinegoni, C. and Naxerova, K. and Sosnovik, D. E. and Zile, M. R. and Bradshaw, A. D. and Liao, R. and Tawakol, A. and Weissleder, R. and Rosenzweig, A. and Swirski, F. K. and Sam, F. and Nahrendorf<sup>#</sup>, M.},title={Cardiac macrophages promote diastolic dysfunction},journal={Journal of Experimental Medicine},alternatejournal={J. Exp. Med.},year={2018},volume={215},number={2},pages={423-440},issn={0022-1007},doi={10.1084/jem.20171274},pmcid={PMC3366498},pmid={29339450}}
29339450
PMC3366498
10.1084/jem.20171274
Circ.Res.
Imaging the Vascular Bone Marrow Niche During Inflammatory Stress
Vandoorne, K.,
Rohde, D.,
Kim, H. Y.,
Courties, G.,
Wojtkiewicz, G.,
Honold, L.,
Hoyer, F. F.,
Frodermann, V.,
Nayar, R.,
Herisson, F.,
Jung, Y.,
Desogere, P. A.,
Vinegoni, C.,
Caravan, P.,
Weissleder, R.,
Sosnovik, D. E.,
Lin, C. P.,
Swirski, F. K.,
and Nahrendorf#, M.
Rationale: Inflammatory stress induced by exposure to bacterial lipopolysaccharide causes hematopoietic stem cell expansion in the bone marrow niche, generating a cellular immune response. As an integral component of the hematopoietic stem cell niche, the bone marrow vasculature regulates the production and release of blood leukocytes, which protect the host against infection but also fuel inflammatory diseases. Objective: We aimed to develop imaging tools to explore vascular changes in the bone marrow niche during acute inflammation. Methods and Results: Using the TLR (Toll-like receptor) ligand lipopolysaccharide as a prototypical danger signal, we applied multiparametric, multimodality and multiscale imaging to characterize how the bone marrow vasculature adapts when hematopoiesis boosts leukocyte supply. In response to lipopolysaccharide, ex vivo flow cytometry and histology showed vascular changes to the bone marrow niche. Specifically, proliferating endothelial cells gave rise to new vasculature in the bone marrow during hypoxic conditions. We studied these vascular changes with complementary intravital microscopy and positron emission tomography/magnetic resonance imaging. Fluorescence and positron emission tomography integrin alpha V beta 3 imaging signal increased during lipopolysaccharide-induced vascular remodeling. Vascular leakiness, quantified by albumin-based in vivo microscopy and magnetic resonance imaging, rose when neutrophils departed and hematopoietic stem and progenitor cells proliferated more vigorously. Conclusions: Introducing a tool set to image bone marrow either with cellular resolution or noninvasively within the entire skeleton, this work sheds light on angiogenic responses that accompany emergency hematopoiesis. Understanding and monitoring bone marrow vasculature may provide a key to unlock therapeutic targets regulating systemic inflammation.
@article{2018-CIRCRES,author={Vandoorne, K. and Rohde, D. and Kim, H. Y. and Courties, G. and Wojtkiewicz, G. and Honold, L. and Hoyer, F. F. and Frodermann, V. and Nayar, R. and Herisson, F. and Jung, Y. and Desogere, P. A. and Vinegoni, C. and Caravan, P. and Weissleder, R. and Sosnovik, D. E. and Lin, C. P. and Swirski, F. K. and Nahrendorf<sup>#</sup>, M.},title={Imaging the Vascular Bone Marrow Niche During Inflammatory Stress},journal={Circulation Research},alternatejournal={Circ.Res.},year={2018},volume={123},number={4},pages={415-427},issn={0009-7330},doi={10.1161/circresaha.118.313302},pmcid={6202141},pmid={29980569}}
29980569
6202141
10.1161/circresaha.118.313302
Sci. Rep.
The anti-tumor diterpene oridonin is a direct inhibitor of Nucleolin in cancer cells
Vasaturo, M.,
Cotugno, R.,
Fiengo, L.,
Vinegoni, C.,
Dal Piaz, F.,
and De Tommasi#, N.
The bioactive plant diterpene oridonin displays important pharmacological activities and is widely used in traditional Chinese medicine; however, its molecular mechanism of action is still incompletely described. In vitro and in vivo data have demonstrated anti-tumor activity of oridonin and its ability to interfere with several cell pathways; however, presently only the molecular chaperone HSP70 has been identified as a direct potential target of this compound. Here, using a combination of different proteomic approaches, innovative Cellular Thermal Shift Assay (CETSA) experiments, and classical biochemical methods, we demonstrate that oridonin interacts with Nucleolin, effectively modulating the activity of this multifunctional protein. The ability of oridonin to target Nucleolin and/or HSP70 could account for the bioactivity profile of this plant diterpene. Recently, Nucleolin has attracted attention as a druggable target, as its diverse functions are implicated in pathological processes such as cancer, inflammation, and viral infection. However, up to now, no small molecule as Nucleolin binders has been reported, thus our finding represents the first evidence of Nucleolin modulation by a small inhibitor.
@article{2018-SCIREP,author={Vasaturo, M. and Cotugno, R. and Fiengo, L. and Vinegoni, C. and Dal Piaz, F. and De Tommasi<sup>#</sup>, N.},title={The anti-tumor diterpene oridonin is a direct inhibitor of Nucleolin in cancer cells},journal={Scientific Reports},alternatejournal={Sci. Rep.},year={2018},volume={8},pages={13},issn={2045-2322},doi={10.1038/s41598-018-35088-x},pmcid={6233161},pmid={30425290}}
30425290
6233161
10.1038/s41598-018-35088-x
2017
Nat. Chem. Biol.
Quantitating drug-target engagement in single cells in vitro and in vivo
Dubach, J. M.,
Kim, E.,
Yang, K.,
Cuccarese, M.,
Giedt, R. J.,
Meirnetis, L. G.,
Vinegoni#, C.,
and Weissleder#, R.
Quantitation of drug target engagement in single cells has proven to be difficult, often leaving unanswered questions in the drug development process. We found that intracellular target engagement of unlabeled new therapeutics can be quantitated using polarized microscopy combined with competitive binding of matched fluorescent companion imaging probes. We quantitated the dynamics of target engagement of covalent BTK inhibitors, as well as reversible PARP inhibitors, in populations of single cells using a single companion imaging probe for each target. We then determined average in vivo tumor concentrations and found marked population heterogeneity following systemic delivery, revealing single cells with low target occupancy at high average target engagement in vivo.
@article{2017-NCBIOL,author={Dubach, J. M. and Kim, E. and Yang, K. and Cuccarese, M. and Giedt, R. J. and Meirnetis, L. G. and Vinegoni<sup>#</sup>, C. and Weissleder<sup>#</sup>, R.},title={Quantitating drug-target engagement in single cells in vitro and in vivo},journal={Nature Chemical Biology},alternatejournal={Nat. Chem. Biol.},year={2017},volume={13},number={2},pages={168-173},issn={1552-4450},doi={10.1038/nchembio.2248},pmcid={PMC5630128},pmid={27918558}}
27918558
PMC5630128
10.1038/nchembio.2248
Cell
Macrophages Facilitate Electrical Conduction in the Heart
Hulsmans, M.,
Clauss, S.,
Xiao, L.,
Aguirre, A. D.,
King, K. R.,
Hanley, A.,
Hucker, W. J.,
Wulfers, E. M.,
Seemann, G.,
Courties, G.,
Iwamoto, Y.,
Sun, Y.,
Savol, A. J.,
Sager, H. B.,
Lavine, K. J.,
Fishbein, G. A.,
Capen, D. E.,
Da Silva, N.,
Miquerol, L.,
Wakimoto, H.,
Seidman, C. E.,
Seidman, J. G.,
Sadreyev, R. I.,
Naxerova, K.,
Mitchell, R. N.,
Brown, D.,
Libby, P.,
Weissleder, R.,
Swirski, F. K.,
Kohl, P.,
Vinegoni, C.,
Milan, D. J.,
Ellinor, P. T.,
and Nahrendorf#, M.
Organ-specific functions of tissue-resident macrophages in the steady-state heart are unknown. Here, we show that cardiac macrophages facilitate electrical conduction through the distal atrioventricular node, where conducting cells densely intersperse with elongated macrophages expressing connexin 43. When coupled to spontaneously beating cardiomyocytes via connexin-43-containing gap junctions, cardiac macrophages have a negative resting membrane potential and depolarize in synchrony with cardiomyocytes. Conversely, macrophages render the resting membrane potential of cardiomyocytes more positive and, according to computational modeling, accelerate their repolarization. Photostimulation of channelrhodopsin-2-expressing macrophages improves atrioventricular conduction, whereas conditional deletion of connexin 43 in macrophages and congenital lack of macrophages delay atrioventricular conduction. In the Cd11b(DTR) mouse, macrophage ablation induces progressive atrioventricular block. These observations implicate macrophages in normal and aberrant cardiac conduction.
@article{2017-CELL,author={Hulsmans, M. and Clauss, S. and Xiao, L. and Aguirre, A. D. and King, K. R. and Hanley, A. and Hucker, W. J. and Wulfers, E. M. and Seemann, G. and Courties, G. and Iwamoto, Y. and Sun, Y. and Savol, A. J. and Sager, H. B. and Lavine, K. J. and Fishbein, G. A. and Capen, D. E. and Da Silva, N. and Miquerol, L. and Wakimoto, H. and Seidman, C. E. and Seidman, J. G. and Sadreyev, R. I. and Naxerova, K. and Mitchell, R. N. and Brown, D. and Libby, P. and Weissleder, R. and Swirski, F. K. and Kohl, P. and Vinegoni, C. and Milan, D. J. and Ellinor, P. T. and Nahrendorf<sup>#</sup>, M.},title={Macrophages Facilitate Electrical Conduction in the Heart},journal={Cell},alternatejournal={Cell},year={2017},volume={169},number={3},pages={510-+},issn={0092-8674},doi={10.1016/j.cell.2017.03.050},pmcid={PMC5474950},pmid={28431249}}
28431249
PMC5474950
10.1016/j.cell.2017.03.050
J. Biomed. Opt.
Motion characterization scheme to minimize motion artifacts in intravital microscopy
Lee, S.,
Courties, G.,
Nahrendorf, M.,
Weissleder, R.,
and Vinegoni#, C.
Respiratory- and cardiac-induced motion artifacts pose a major challenge for in vivo optical imaging, limiting the temporal and spatial imaging resolution in fluorescence laser scanning microscopy. Here, we present an imaging platform developed for in vivo characterization of physiologically induced axial motion. The motion characterization system can be straightforwardly implemented on any conventional laser scanning microscope and can be used to evaluate the effectiveness of different motion stabilization schemes. This method is particularly useful to improve the design of novel tissue stabilizers and to facilitate stabilizer positioning in real time, therefore facilitating optimal tissue immobilization and minimizing motion induced artifacts. (C) 2017 Society of Photo-Optical Instrumentation Engineers (SPIE).
@article{2017-JBO,author={Lee, S. and Courties, G. and Nahrendorf, M. and Weissleder, R. and Vinegoni<sup>#</sup>, C.},title={Motion characterization scheme to minimize motion artifacts in intravital microscopy},journal={Journal of Biomedical Optics},alternatejournal={J. Biomed. Opt.},year={2017},volume={22},number={3},pages={7},issn={1083-3668},doi={10.1117/1.Jbo.22.3.036005},pmcid={PMC5333764},pmid={28253383}}
28253383
PMC5333764
10.1117/1.Jbo.22.3.036005
Theranostics
Design and Development of Fluorescent Vemurafenib Analogs for In Vivo Imaging
Mikula, H.,
Stapleton, S.,
Kohler, R. H.,
Vinegoni, C.,
and Weissleder#, R.
Herein we describe fluorescent derivatives of vemurafenib to probe therapeutic BRAF inhibition in live cells and in vivo. The compounds were evaluated and compared by determining target binding, inhibition of mutant BRAF melanoma cell lines and live cell imaging. We show that vemurafenib-BODIPY is a superior imaging drug to visualize the targets of vemurafenib in live cells and in vivo in non-resistant and resistant melanoma tumors.
@article{2017-THERA,author={Mikula, H. and Stapleton, S. and Kohler, R. H. and Vinegoni, C. and Weissleder<sup>#</sup>, R.},title={Design and Development of Fluorescent Vemurafenib Analogs for In Vivo Imaging},journal={Theranostics},alternatejournal={Theranostics},year={2017},volume={7},number={5},pages={1257-1265},issn={1838-7640},doi={10.7150/thno.18238},pmcid={PMC5399591},pmid={28435463}}
28435463
PMC5399591
10.7150/thno.18238
ACS Nano
Transparent Electrophysiology Microelectrodes and Interconnects from Meta Nanomesh
Seo, K. J.,
Qiang, Y.,
Bilgin, I.,
Kar, S.,
Vinegoni, C.,
Weissleder, R.,
and Fang#, H.
Mapping biocurrents at both microsecond and single-cell resolution requires the combination of optical imaging with innovative electrophysiological sensing techniques. Here, we present transparent electrophysiology electrodes and interconnects made of gold (Au) nanomesh on flexible substrates to achieve such measurements. Compared to previously demonstrated indium tin oxide (ITO) and graphene electrodes, the ones from Au nanomesh possess superior properties including low electrical impedance, high transparency, good cell viability, and superb flexibility. Specifically, we demonstrated a 15 nm thick Au nanomesh electrode with 8.14 Omega.cm(2) normalized impedance, >65% average transmittance over a 300-1100 nm window, and stability up to 300 bending cycles. Systematic sheet resistance measurements, electrochemical impedance studies, optical characterization, mechanical bending tests, and cell studies highlight the capabilities of the Au nanomesh as a transparent electrophysiology electrode and interconnect material. Together, these results demonstrate applicability of using nanomesh under biological conditions and broad applications in biology and medicine.
@article{2017-ACSN,author={Seo, K. J. and Qiang, Y. and Bilgin, I. and Kar, S. and Vinegoni, C. and Weissleder, R. and Fang<sup>#</sup>, H.},title={Transparent Electrophysiology Microelectrodes and Interconnects from Meta Nanomesh},journal={Acs Nano},alternatejournal={ACS Nano},year={2017},volume={11},number={4},pages={4365-4372},issn={1936-0851},doi={10.1021/acsnano.7b01995},pmid={28391679}}
28391679
10.1021/acsnano.7b01995
Nat. Protoc.
Measurement of drug-target engagement in live cells by two-photon fluorescence anisotropy imaging
Vinegoni#, C.,
Fumene Feruglio, P.,
Brand, C.,
Lee, S.,
Nibbs, A. E.,
Stapleton, S.,
Shah, S.,
Gryczynski, I.,
Reiner, T.,
Mazitschek, R.,
and Weissleder, R.
The ability to directly image and quantify drug-target engagement and drug distribution with subcellular resolution in live cells and whole organisms is a prerequisite to establishing accurate models of the kinetics and dynamics of drug action. Such methods would thus have far-reaching applications in drug development and molecular pharmacology. We recently presented one such technique based on fluorescence anisotropy, a spectroscopic method based on polarization light analysis and capable of measuring the binding interaction between molecules. Our technique allows the direct characterization of target engagement of fluorescently labeled drugs, using fluorophores with a fluorescence lifetime larger than the rotational correlation of the bound complex. Here we describe an optimized protocol for simultaneous dual-channel two-photon fluorescence anisotropy microscopy acquisition to perform drug-target measurements. We also provide the necessary software to implement stream processing to visualize images and to calculate quantitative parameters. The assembly and characterization part of the protocol can be implemented in 1 d. Sample preparation, characterization and imaging of drug binding can be completed in 2 d. Although currently adapted to an Olympus FV1000MPEPE microscope, the protocol can be extended to other commercial or custom-built microscopes.
@article{2017-NP,author={Vinegoni<sup>#</sup>, C. and Fumene Feruglio, P. and Brand, C. and Lee, S. and Nibbs, A. E. and Stapleton, S. and Shah, S. and Gryczynski, I. and Reiner, T. and Mazitschek, R. and Weissleder, R.},title={Measurement of drug-target engagement in live cells by two-photon fluorescence anisotropy imaging},journal={Nature Protocols},alternatejournal={Nat. Protoc.},year={2017},volume={12},number={7},pages={1472-1497},issn={1754-2189},doi={10.1038/nprot.2017.043},pmcid={PMC5928516},pmid={28686582}}
28686582
PMC5928516
10.1038/nprot.2017.043
2016
Technology
Two-photon imaging of pancreatic beta cells in real time in vivo
Clardy, S. M.,
Kohler, R.,
Vinegoni, C.,
Iwamoto, Y.,
Keliher, E.,
and Weissleder#, R.
Here we present the first generation of two-photon beta cell specific in vivo imaging probes based on GLP1R targeting peptides. Among the three compounds of potential interest, we found quite unexpectedly that a squarine-rotaxane conjugate (2PEx-647) had near ideal in vivo imaging characteristics.
@article{2016-TECHNOL,author={Clardy, S. M. and Kohler, R. and Vinegoni, C. and Iwamoto, Y. and Keliher, E. and Weissleder<sup>#</sup>, R.},title={Two-photon imaging of pancreatic beta cells in real time in vivo},journal={Technology},alternatejournal={Technology},year={2016},volume={4},pages={130-34},doi={10.1142/S2339547816200028},}
10.1142/S2339547816200028
Sci. Rep.
Computational imaging reveals mitochondrial morphology as a biomarker of cancer phenotype and drug response
Giedt, R. J.,
Fumene Feruglio, P.,
Pathania, D.,
Yang, K. S.,
Kilcoyne, A.,
Vinegoni, C.,
Mitchison, T. J.,
and Weissleder#, R.
Mitochondria, which are essential organelles in resting and replicating cells, can vary in number, mass and shape. Past research has primarily focused on short-term molecular mechanisms underlying fission/fusion. Less is known about longer-term mitochondrial behavior such as the overall makeup of cell populations’ morphological patterns and whether these patterns can be used as biomarkers of drug response in human cells. We developed an image-based analytical technique to phenotype mitochondrial morphology in different cancers, including cancer cell lines and patient-derived cancer cells. We demonstrate that (i) cancer cells of different origins, including patient-derived xenografts, express highly diverse mitochondrial phenotypes; (ii) a given phenotype is characteristic of a cell population and fairly constant over time; (iii) mitochondrial patterns correlate with cell metabolic measurements and (iv) therapeutic interventions can alter mitochondrial phenotypes in drug-sensitive cancers as measured in pre-versus post-treatment fine needle aspirates in mice. These observations shed light on the role of mitochondrial dynamics in the biology and drug response of cancer cells. On the basis of these findings, we propose that image-based mitochondrial phenotyping can provide biomarkers for assessing cancer phenotype and drug response.
@article{2016-SCIREP,author={Giedt, R. J. and Fumene Feruglio, P. and Pathania, D. and Yang, K. S. and Kilcoyne, A. and Vinegoni, C. and Mitchison, T. J. and Weissleder<sup>#</sup>, R.},title={Computational imaging reveals mitochondrial morphology as a biomarker of cancer phenotype and drug response},journal={Scientific Reports},alternatejournal={Sci. Rep.},year={2016},volume={6},pages={10},issn={2045-2322},doi={10.1038/srep32985},pmcid={PMC5017129},pmid={27609668}}
27609668
PMC5017129
10.1038/srep32985
Nat. Commun.
Tyrosine kinase-mediated axial motility of basal cells revealed by intravital imaging
Roy, J.,
Kim, B.,
Hill, E.,
Visconti, P.,
Krapf, D.,
Vinegoni, C.,
Weissleder, R.,
Brown, D.,
and Breton#, S.
Epithelial cells are generally considered to be static relative to their neighbours. Basal cells in pseudostratified epithelia display a single long cytoplasmic process that can cross the tight junction barrier to reach the lumen. Using in vivo microscopy to visualize the epididymis, a model system for the study of pseudostratified epithelia, we report here the surprising discovery that these basal cell projections-which we call axiopodia-periodically extend and retract over time. We found that axiopodia extensions and retractions follow an oscillatory pattern. This movement, which we refer to as periodic axial motility (PAM), is controlled by c-Src and MEK1/2-ERK1/2. Therapeutic inhibition of tyrosine kinase activity induces a retraction of these projections. Such unexpected cell motility may reflect a novel mechanism by which specialized epithelial cells sample the luminal environment.
@article{2016-NCOMM-1,author={Roy, J. and Kim, B. and Hill, E. and Visconti, P. and Krapf, D. and Vinegoni, C. and Weissleder, R. and Brown, D. and Breton<sup>#</sup>, S.},title={Tyrosine kinase-mediated axial motility of basal cells revealed by intravital imaging},journal={Nature Communications},alternatejournal={Nat. Commun.},year={2016},volume={7},pages={11},doi={10.1038/ncomms10666},pmcid={PMC4754344},pmid={26868824}}
26868824
PMC4754344
10.1038/ncomms10666
Sci. Transl. Med.
RNAi targeting multiple cell adhesion molecules reduces immune cell recruitment and vascular inflammation after myocardial infarction
Sager, H. B.,
Dutta, P.,
Dahlman, J. E.,
Hulsmans, M.,
Courties, G.,
Sun, Y.,
Heidt, T.,
Vinegoni, C.,
Borodovsky, A.,
Fitzgerald, K.,
Wojtkiewicz, G. R.,
Iwamoto, Y.,
Tricot, B.,
Khan, O. F.,
Kauffman, K. J.,
Xing, Y. P.,
Shaw, T. E.,
Libby, P.,
Langer, R.,
Weissleder, R.,
Swirski, F. K.,
Anderson, D. G.,
and Nahrendorf#, M.
Myocardial infarction (MI) leads to a systemic surge of vascular inflammation in mice and humans, resulting in secondary ischemic complications and high mortality. We show that, in ApoE(-/-) mice with coronary ligation, increased sympathetic tone up-regulates not only hematopoietic leukocyte production but also plaque endothelial expression of adhesion molecules. To counteract the resulting arterial leukocyte recruitment, we developed nanoparticle-based RNA interference (RNAi) that effectively silences five key adhesion molecules. Simultaneously encapsulating small interfering RNA (siRNA)-targeting intercellular cell adhesion molecules 1 and 2 (Icam1 and Icam2), vascular cell adhesion molecule 1 (Vcam1), and E-and P-selectins (Sele and Selp) into polymeric endothelial-avid nanoparticles reduced post-MI neutrophil and monocyte recruitment into atherosclerotic lesions and decreased matrix-degrading plaque protease activity. Five-gene combination RNAi also curtailed leukocyte recruitment to ischemic myocardium. Therefore, targeted multigene silencing may prevent complications after acute MI.
@article{2016-STM,author={Sager, H. B. and Dutta, P. and Dahlman, J. E. and Hulsmans, M. and Courties, G. and Sun, Y. and Heidt, T. and Vinegoni, C. and Borodovsky, A. and Fitzgerald, K. and Wojtkiewicz, G. R. and Iwamoto, Y. and Tricot, B. and Khan, O. F. and Kauffman, K. J. and Xing, Y. P. and Shaw, T. E. and Libby, P. and Langer, R. and Weissleder, R. and Swirski, F. K. and Anderson, D. G. and Nahrendorf<sup>#</sup>, M.},title={RNAi targeting multiple cell adhesion molecules reduces immune cell recruitment and vascular inflammation after myocardial infarction},journal={Science Translational Medicine},alternatejournal={Sci. Transl. Med.},year={2016},volume={8},number={342},pages={11},issn={1946-6234},doi={10.1126/scitranslmed.aaf1435},pmcid={PMC5125383},pmid={27280687}}
27280687
PMC5125383
10.1126/scitranslmed.aaf1435
IEEE J.S.T.Q.E.
Two-Photon Fluorescence Anisotropy Microscopy for Imaging and Direct Measurement of Intracellular Drug Target Engagement
Vinegoni#, C.,
Dubach, J. M.,
Feruglio, P. F.,
and Weissleder, R.
Ieee Journal of Selected Topics in Quantum Electronics 2016
Small molecule therapeutic drugs must reach their intended cellular targets (pharmacokinetics) and engage them to modulate therapeutic effects (pharmacodynamics). These processes are often difficult to measure in vivo due to their complexities and occurrence within single cells. It has been particularly difficult to directly measure cellular drug target binding. Fluorescence polarization is commonly used in pharmacological screening assays to measure drug-protein or protein-protein interactions. We hypothesized that fluorescence polarization imaging could be adapted and used with fluorescently labeled drugs to measure drug target engagement in vivo. Here, we summarize recent results using two photon fluorescence anisotropy microscopy. Our imaging technique offers quantitative pharmacological binding information of diverse molecular interactions at themicroscopic level, differentiating between bound, and unbound states. Used in combination with other recent advances in the development of novel fluorescently labeled drugs, we expect that the described imaging modality will provide a window into the distribution and efficacy of drugs in real time and in vivo at the cellular and subcellular level.
@article{2016-IEEE,author={Vinegoni<sup>#</sup>, C. and Dubach, J. M. and Feruglio, P. F. and Weissleder, R.},title={Two-Photon Fluorescence Anisotropy Microscopy for Imaging and Direct Measurement of Intracellular Drug Target Engagement},journal={Ieee Journal of Selected Topics in Quantum Electronics},alternatejournal={IEEE J.S.T.Q.E.},year={2016},volume={22},number={3},pages={7},issn={1077-260X},doi={10.1109/jstqe.2015.2501384},pmcid={PMC4946648},pmid={27440991}}
27440991
PMC4946648
10.1109/jstqe.2015.2501384
Nat. Commun.
Real-time high dynamic range laser scanning microscopy
Vinegoni#†, C.,
Swisher†, C. L.,
Fumene Feruglio†, P.,
Giedt, R. J.,
Rousso, D. L.,
Stapleton, S.,
and Weissleder, R.
In conventional confocal/multiphoton fluorescence microscopy, images are typically acquired under ideal settings and after extensive optimization of parameters for a given structure or feature, often resulting in information loss from other image attributes. To overcome the problem of selective data display, we developed a new method that extends the imaging dynamic range in optical microscopy and improves the signal-to-noise ratio. Here we demonstrate how real-time and sequential high dynamic range microscopy facilitates automated three-dimensional neural segmentation. We address reconstruction and segmentation performance on samples with different size, anatomy and complexity. Finally, in vivo real-time high dynamic range imaging is also demonstrated, making the technique particularly relevant for longitudinal imaging in the presence of physiological motion and/or for quantification of in vivo fast tracer kinetics during functional imaging.
@article{2016-NCOMM-2,author={Vinegoni<sup>#†</sup>, C. and Swisher<sup>†</sup>, C. L. and Fumene Feruglio<sup>†</sup>, P. and Giedt, R. J. and Rousso, D. L. and Stapleton, S. and Weissleder, R.},title={Real-time high dynamic range laser scanning microscopy},journal={Nature Communications},alternatejournal={Nat. Commun.},year={2016},volume={7},pages={13},issn={2041-1723},doi={10.1038/ncomms11077},pmcid={PMC4821995},pmid={27032979}}
27032979
PMC4821995
10.1038/ncomms11077
2015
Sci. Rep.
Rapid, high efficiency isolation of pancreatic beta-cells
Clardy, S. M.,
Mohan, J. F.,
Vinegoni, C.,
Keliher, E. J.,
Iwamoto, Y.,
Benoist, C.,
Mathis, D.,
and Weissleder#, R.
The ability to isolate pure pancreatic beta-cells would greatly aid multiple areas of diabetes research. We developed a fluorescent exendin-4-like neopeptide conjugate for the rapid purification and isolation of functional mouse pancreatic beta-cells. By targeting the glucagon-like peptide-1 receptor with the fluorescent conjugate, beta-cells could be quickly isolated by flow cytometry and were >99% insulin positive. These studies were confirmed by immunostaining, microscopy and gene expression profiling on isolated cells. Gene expression profiling studies of cytofluorometrically sorted beta-cells from 4 and 12 week old NOD mice provided new insights into the genetic programs at play of different stages of type-1 diabetes development. The described isolation method should have broad applicability to the beta-cell field.
@article{2015-SREP,author={Clardy, S. M. and Mohan, J. F. and Vinegoni, C. and Keliher, E. J. and Iwamoto, Y. and Benoist, C. and Mathis, D. and Weissleder<sup>#</sup>, R.},title={Rapid, high efficiency isolation of pancreatic beta-cells},journal={Scientific Reports},alternatejournal={Sci. Rep.},year={2015},volume={5},pages={9},issn={2045-2322},doi={10.1038/srep13681},pmcid={PMC4557033},pmid={26330153}}
26330153
PMC4557033
10.1038/srep13681
Cell Stem Cell
Myocardial Infarction Activates CCR2(+) Hematopoietic Stem and Progenitor Cells
Dutta, P.,
Sager, H. B.,
Stengel, K. R.,
Naxerova, K.,
Courties, G.,
Saez, B.,
Silberstein, L.,
Heidt, T.,
Sebas, M.,
Sun, Y.,
Wojtkiewicz, G.,
Fumene Feruglio, P.,
King, K.,
Baker, J. N.,
Laan, A. M.,
Borodovsky, A.,
Fitzgerald, K.,
Hulsmans, M.,
Hoyer, F.,
Iwamoto, Y.,
Vinegoni, C.,
Brown, D.,
Di Carli, M.,
Libby, P.,
Hiebert, S. W.,
Scadden, D. T.,
Swirski, F. K.,
Weissleder, R.,
and Nahrendorf#, M.
Following myocardial infarction (MI), myeloid cells derived from the hematopoietic system drive a sharp increase in systemic leukocyte levels that correlates closely with mortality. The origin of these myeloid cells, and the response of hematopoietic stem and progenitor cells (HSPCs) to MI, however, is unclear. Here, we identify a CCR2(+)CD150(+)CD48(-) LSK hematopoietic subset as the most upstream contributor to emergency myelopoiesis after ischemic organ injury. This subset has 4-fold higher proliferation rates than CCR2(-)CD150(+)CD48(-) LSK cells, displays a myeloid differentiation bias, and dominates the migratory HSPC population. We further demonstrate that the myeloid translocation gene 16 (Mtg16) regulates CCR2(+) HSPC emergence. Mtg16(-/-) mice have decreased levels of systemic monocytes and infarct-associated macrophages and display compromised tissue healing and post-MI heart failure. Together, these data provide insights into regulation of emergency hematopoiesis after ischemic injury and identify potential therapeutic targets to modulate leukocyte output after MI.
@article{2015-CSC,author={Dutta, P. and Sager, H. B. and Stengel, K. R. and Naxerova, K. and Courties, G. and Saez, B. and Silberstein, L. and Heidt, T. and Sebas, M. and Sun, Y. and Wojtkiewicz, G. and Fumene Feruglio, P. and King, K. and Baker, J. N. and van der Laan, A. M. and Borodovsky, A. and Fitzgerald, K. and Hulsmans, M. and Hoyer, F. and Iwamoto, Y. and Vinegoni, C. and Brown, D. and Di Carli, M. and Libby, P. and Hiebert, S. W. and Scadden, D. T. and Swirski, F. K. and Weissleder, R. and Nahrendorf<sup>#</sup>, M.},title={Myocardial Infarction Activates CCR2(+) Hematopoietic Stem and Progenitor Cells},journal={Cell Stem Cell},alternatejournal={Cell Stem Cell},year={2015},volume={16},number={5},pages={477-487},issn={1934-5909},doi={10.1016/j.stem.2015.04.008},pmcid={PMC4426344},pmid={25957903}}
25957903
PMC4426344
10.1016/j.stem.2015.04.008
Drug Discov. Today
Advances in measuring single-cell pharmacology in vivo
Vinegoni#, C.,
Dubach, J. M.,
Thurber, G. M.,
Miller, M. A.,
Mazitschek, R.,
and Weissleder, R.
Measuring key pharmacokinetic and pharmacodynamic parameters in vivo at the single cell level is likely to enhance drug discovery and development. In this review, we summarize recent advances in this field and highlight current and future capabilities.
@article{2015-DDT,author={Vinegoni<sup>#</sup>, C. and Dubach, J. M. and Thurber, G. M. and Miller, M. A. and Mazitschek, R. and Weissleder, R.},title={Advances in measuring single-cell pharmacology in vivo},journal={Drug Discovery Today},alternatejournal={Drug Discov. Today},year={2015},volume={20},number={9},pages={1087-1092},issn={1359-6446},doi={10.1016/j.drudis.2015.05.011},pmcid={PMC4567932},pmid={26024776}}
26024776
PMC4567932
10.1016/j.drudis.2015.05.011
Nat. Protoc.
Imaging the beating heart in the mouse using intravital microscopy techniques
Vinegoni#†, C.,
Aguirre†, A. D.,
Lee, S.,
and Weissleder, R.
Real-time microscopic imaging of moving organs at single-cell resolution represents a major challenge in studying complex biology in living systems. Motion of the tissue from the cardiac and respiratory cycles severely limits intravital microscopy by compromising ultimate spatial and temporal imaging resolution. However, significant recent advances have enabled single-cell resolution imaging to be achieved in vivo. In this protocol, we describe experimental procedures for intravital microscopy based on a combination of thoracic surgery, tissue stabilizers and acquisition gating methods, which enable imaging at the single-cell level in the beating heart in the mouse. Setup of the model is typically completed in 1 h, which allows 2 h or more of continuous cardiac imaging. This protocol can be readily adapted for the imaging of other moving organs, and it will therefore broadly facilitate in vivo high-resolution microscopy studies.
@article{2015-NPROT,author={Vinegoni<sup>#†</sup>, C. and Aguirre<sup>†</sup>, A. D. and Lee, S. and Weissleder, R.},title={Imaging the beating heart in the mouse using intravital microscopy techniques},journal={Nature Protocols},alternatejournal={Nat. Protoc.},year={2015},volume={10},number={11},pages={1802-1819},issn={1754-2189},doi={10.1038/nprot.2015.119},pmcid={PMC5380003},pmid={26492138}}
26492138
PMC5380003
10.1038/nprot.2015.119
Front. Physiol.
New techniques for motion-artifact-free in vivo cardiac microscopy
Vinegoni#†, C.,
Lee†, S.,
Aguirre, A. D.,
and Weissleder, R.
Intravital imaging microscopy (i.e., imaging in live animals at microscopic resolution) has become an indispensable tool for studying the cellular micro-dynamics in cancer, immunology and neurobiology. High spatial and temporal resolution, combined with large penetration depth and multi-reporter visualization capability make fluorescence intravital microscopy compelling for heart imaging. However, tissue motion caused by cardiac contraction and respiration critically limits its use. As a result, in vitro cell preparations or non-contracting explanted heart models are more commonly employed. Unfortunately, these approaches fall short of understanding the more complex host physiology that may be dynamic and occur over longer periods of time. In this review, we report on novel technologies, which have been recently developed by our group and others, aimed at overcoming motion-induced artifacts and capable of providing in vivo subcellular resolution imaging in the beating mouse heart. The methods are based on mechanical stabilization, image processing algorithms, gated/triggered acquisition schemes or a combination of both. We expect that in the immediate future all these methodologies will have considerable applications in expanding our understanding of the cardiac biology, elucidating cardiomyocyte function and interactions within the organism in vivo, and ultimately improving the treatment of cardiac diseases.
@article{2015-FPHYS,author={Vinegoni<sup>#†</sup>, C. and Lee<sup>†</sup>, S. and Aguirre, A. D. and Weissleder, R.},title={New techniques for motion-artifact-free in vivo cardiac microscopy},journal={Frontiers in Physiology},alternatejournal={Front. Physiol.},year={2015},volume={6},pages={12},issn={1664-042X},doi={10.3389/fphys.2015.00147},pmcid={PMC4428079},pmid={26029116}}
26029116
PMC4428079
10.3389/fphys.2015.00147
2014
P.N.A.S.
Intravital imaging of cardiac function at the single-cell level
Aguirre, A. D.,
Vinegoni#, C.,
Sebas, M.,
and Weissleder#, R.
Proceedings of the National Academy of Sciences of the United States of America 2014
Knowledge of cardiomyocyte biology is limited by the lack of methods to interrogate single-cell physiology in vivo. Here we show that contracting myocytes can indeed be imaged with optical microscopy at high temporal and spatial resolution in the beating murine heart, allowing visualization of individual sarcomeres and measurement of the single cardiomyocyte contractile cycle. Collectively, this has been enabled by efficient tissue stabilization, a prospective real-time cardiac gating approach, an image processing algorithm for motion-artifact-free imaging throughout the cardiac cycle, and a fluorescent membrane staining protocol. Quantification of cardiomyocyte contractile function in vivo opens many possibilities for investigating myocardial disease and therapeutic intervention at the cellular level.
@article{2014-PNAS,author={Aguirre, A. D. and Vinegoni<sup>#</sup>, C. and Sebas, M. and Weissleder<sup>#</sup>, R.},title={Intravital imaging of cardiac function at the single-cell level},journal={Proceedings of the National Academy of Sciences of the United States of America},alternatejournal={P.N.A.S.},year={2014},volume={111},number={31},pages={11257-11262},issn={0027-8424},doi={10.1073/pnas.1401316111},pmcid={PMC4128110},pmid={25053815}}
25053815
PMC4128110
10.1073/pnas.1401316111
Nat. Commun.
In vivo imaging of specific drug-target binding at subcellular resolution
Dubach†, J. M.,
Vinegoni#†, C.,
Mazitschek, R.,
Fumene Feruglio, P.,
Cameron, L. A.,
and Weissleder, R.
The possibility of measuring binding of small-molecule drugs to desired targets in live cells could provide a better understanding of drug action. However, current approaches mostly yield static data, require lysis or rely on indirect assays and thus often provide an incomplete understanding of drug action. Here, we present a multiphoton fluorescence anisotropy microscopy live cell imaging technique to measure and map drug-target interaction in real time at subcellular resolution. This approach is generally applicable using any fluorescently labelled drug and enables high-resolution spatial and temporal mapping of bound and unbound drug distribution. To illustrate our approach we measure intracellular target engagement of the chemotherapeutic Olaparib, a poly(ADP-ribose) polymerase inhibitor, in live cells and within a tumour in vivo. These results are the first generalizable approach to directly measure drug-target binding in vivo and present a promising tool to enhance understanding of drug activity.
@article{2014-NC,author={Dubach<sup>†</sup>, J. M. and Vinegoni<sup>#†</sup>, C. and Mazitschek, R. and Fumene Feruglio, P. and Cameron, L. A. and Weissleder, R.},title={In vivo imaging of specific drug-target binding at subcellular resolution},journal={Nature Communications},alternatejournal={Nat. Commun.},year={2014},volume={5},pages={9},issn={2041-1723},doi={10.1038/ncomms4946},pmcid={PMC4362617},pmid={24867710}}
24867710
PMC4362617
10.1038/ncomms4946
Opt. Lett.
Steady state anisotropy two-photon microscopy resolves multiple, spectrally similar fluorophores, enabling in vivo multilabel imaging
Dubach†, J. M.,
Vinegoni#†, C.,
and Weissleder, R.
The use of spectrally distinguishable fluorescent dyes enables imaging of multiple targets. However, in two-photon microscopy, the number of fluorescent labels with distinct emission spectra that can be effectively excited and resolved is constrained by the confined tuning range of the excitation laser and the broad and overlapping nature of fluorophore two-photon absorption spectra. This limitation effectively reduces the number of available imaging channels. Here, we demonstrate that two-photon steady state anisotropy imaging (2PSSA) offers the capability to resolve otherwise unresolvable fluorescent tracers both in live cells and in mouse tumor models. This approach expands the number of biological targets that can be imaged simultaneously, increasing the total amount of information that can be obtained through imaging. (C) 2014 Optical Society of America
@article{2014-OL,author={Dubach<sup>†</sup>, J. M. and Vinegoni<sup>#†</sup>, C. and Weissleder, R.},title={Steady state anisotropy two-photon microscopy resolves multiple, spectrally similar fluorophores, enabling in vivo multilabel imaging},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2014},volume={39},number={15},pages={4482-4485},issn={0146-9592},doi={10.1364/ol.39.004482},pmcid={PMC4341989},pmid={25078208}}
Heidt, T.,
Sager, H. B.,
Courties, G.,
Dutta, P.,
Iwamoto, Y.,
Zaltsman, A.,
Muhlen, C.,
Bode, C.,
Fricchione, G. L.,
Denninger, J.,
Lin, C. P.,
Vinegoni, C.,
Libby, P.,
Swirski, F. K.,
Weissleder, R.,
and Nahrendorf#, M.
Exposure to psychosocial stress is a risk factor for many diseases, including atherosclerosis(1,2). Although incompletely understood, interaction between the psyche and the immune system provides one potential mechanism linking stress and disease inception and progression. Known cross-talk between the brain and immune system includes the hypothalamic-pituitary-adrenal axis, which centrally drives glucocorticoid production in the adrenal cortex, and the sympathetic-adrenal-medullary axis, which controls stress-induced catecholamine release in support of the fight-or-flight reflex(3,4). It remains unknown, however, whether chronic stress changes hematopoietic stem cell activity. Here we show that stress increases proliferation of these most primitive hematopoietic progenitors, giving rise to higher levels of disease-promoting inflammatory leukocytes. We found that chronic stress induced monocytosis and neutrophilia in humans. While investigating the source of leukocytosis in mice, we discovered that stress activates upstream hematopoietic stem cells. Under conditions of chronic variable stress in mice, sympathetic nerve fibers released surplus noradrenaline, which signaled bone marrow niche cells to decrease CXCL12 levels through the beta(3)-adrenergic receptor. Consequently, hematopoietic stem cell proliferation was elevated, leading to an increased output of neutrophils and inflammatory monocytes. When atherosclerosis-prone Apoe(-/-) mice were subjected to chronic stress, accelerated hematopoiesis promoted plaque features associated with vulnerable lesions that cause myocardial infarction and stroke in humans.
@article{2014-NM,author={Heidt, T. and Sager, H. B. and Courties, G. and Dutta, P. and Iwamoto, Y. and Zaltsman, A. and von zur Muhlen, C. and Bode, C. and Fricchione, G. L. and Denninger, J. and Lin, C. P. and Vinegoni, C. and Libby, P. and Swirski, F. K. and Weissleder, R. and Nahrendorf<sup>#</sup>, M.},title={Chronic variable stress activates hematopoietic stem cells},journal={Nature Medicine},alternatejournal={Nat. Med.},year={2014},volume={20},number={7},pages={754-758},issn={1078-8956},doi={10.1038/nm.3589},pmcid={PMC4087061},pmid={24952646}}
24952646
PMC4087061
10.1038/nm.3589
Sci. Rep.
Automated motion artifact removal for intravital microscopy, without a priori information
Lee†, S.,
Vinegoni#†, C.,
Sebas, M.,
and Weissleder, R.
Intravital fluorescence microscopy, through extended penetration depth and imaging resolution, provides the ability to image at cellular and subcellular resolution in live animals, presenting an opportunity for new insights into in vivo biology. Unfortunately, physiological induced motion components due to respiration and cardiac activity are major sources of image artifacts and impose severe limitations on the effective imaging resolution that can be ultimately achieved in vivo. Here we present a novel imaging methodology capable of automatically removing motion artifacts during intravital microscopy imaging of organs and orthotopic tumors. The method is universally applicable to different laser scanning modalities including confocal and multiphoton microscopy, and offers artifact free reconstructions independent of the physiological motion source and imaged organ. The methodology, which is based on raw data acquisition followed by image processing, is here demonstrated for both cardiac and respiratory motion compensation in mice heart, kidney, liver, pancreas and dorsal window chamber.
@article{2014-SR,author={Lee<sup>†</sup>, S. and Vinegoni<sup>#†</sup>, C. and Sebas, M. and Weissleder, R.},title={Automated motion artifact removal for intravital microscopy, without a priori information},journal={Scientific Reports},alternatejournal={Sci. Rep.},year={2014},volume={4},pages={9},issn={2045-2322},doi={10.1038/srep04507},pmcid={PMC3968488},pmid={24676021}}
24676021
PMC3968488
10.1038/srep04507
IEEE J.S.T.Q.E.
Advanced Motion Compensation Methods for Intravital Optical Microscopy
Vinegoni#†, C.,
Lee†, S.,
Fumene Feruglio, P.,
and Weissleder, R.
Ieee Journal of Selected Topics in Quantum Electronics 2014
Intravital microscopy has emerged in the recent decade as an indispensible imaging modality for the study of the microdynamics of biological processes in live animals. Technical advancements in imaging techniques and hardware components, combined with the development of novel targeted probes and new mice models, have enabled us to address long-standing questions in several biology areas such as oncology, cell biology, immunology, and neuroscience. As the instrument resolution has increased, physiological motion activities have become a major obstacle that prevents imaging live animals at resolutions analogue to the ones obtained in vitro. Motion compensation techniques aim at reducing this gap and can effectively increase the in vivo resolution. This paper provides a technical review of some of the latest developments in motion compensation methods, providing organ specific solutions.
@article{2014-IEEE,author={Vinegoni<sup>#†</sup>, C. and Lee<sup>†</sup>, S. and Fumene Feruglio, P. and Weissleder, R.},title={Advanced Motion Compensation Methods for Intravital Optical Microscopy},journal={Ieee Journal of Selected Topics in Quantum Electronics},alternatejournal={IEEE J.S.T.Q.E.},year={2014},volume={20},number={2},pages={9},issn={1077-260X},doi={10.1109/jstqe.2013.2279314},pmcid={PMC3832946},pmid={24273405}}
24273405
PMC3832946
10.1109/jstqe.2013.2279314
2013
JHGD
Perinatal health and translational medicine
Figueiredo, J. L.,
Vinegoni, C.,
and De Abreu, L. C.
Throughout fetal life, tissues and organs of
the body are found at a critical period of development
which coincides with the period of rapid cell division.
When faced with lack of nutrients and hypoxia, the
first fetal adaptation is a decrease in cell division
rate. This cell division rate occurs both by direct
effect of nitrogen starvation, and by hormonal and
growth factors changes1. A reduction in the number
of cells, the change in structure and functioning of
organs, permanent change in DNA methylation, and
in gene expression have also been considered to be
molecular mechanisms responsible for fetal
programming1.
@article{2013-JHGD,author={Figueiredo, J. L. and Vinegoni, C. and De Abreu, L. C.},title={Perinatal health and translational medicine},journal={Journal of Human Growth and Development},alternatejournal={JHGD},year={2013},volume={23},number={2},pages={125-127},doi={10.7322/jhgd.61318},}
10.7322/jhgd.61318
J. Biophotonics
Noise suppressed, multifocus image fusion for enhanced intraoperative navigation
Fumene Feruglio†, P.,
Vinegoni#†, C.,
Fexon, L.,
Thurber, G.,
Sbarbati, A.,
and Weissleder, R.
Current intraoperative imaging systems are typically not able to provide sharp’ images over entire large areas or entire organs. Distinct structures such as tissue margins or groups of malignant cells are therefore often difficult to detect, especially under low signal-to-noise-ratio conditions. In this report, we introduce a noise suppressed multifocus image fusion algorithm, that provides detailed reconstructions even when images are acquired under sub-optimal conditions, such is the case for real time fluorescence intraoperative surgery. The algorithm makes use of the Anscombe transform combined with a multi-level stationary wavelet transform with individual threshold-based shrinkage. While the imaging system is integrated with a respiratory monitor triggering system, it can be easily adapted to any commercial imaging system. The developed algorithm is made available as a plugin for Osirix. ((c) 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
@article{2013-BIOPHTNCS,author={Fumene Feruglio<sup>†</sup>, P. and Vinegoni<sup>#†</sup>, C. and Fexon, L. and Thurber, G. and Sbarbati, A. and Weissleder, R.},title={Noise suppressed, multifocus image fusion for enhanced intraoperative navigation},journal={Journal of Biophotonics},alternatejournal={J. Biophotonics},year={2013},volume={6},number={4},pages={363-370},issn={1864-063X},doi={10.1002/jbio.201200086},pmcid={PMC3779878},pmid={22887724}}
22887724
PMC3779878
10.1002/jbio.201200086
Biomed. Opt. Exp.
Sequential average segmented microscopy for high signal-to-noise ratio motion-artifact-free in vivo heart imaging
Vinegoni#†, C.,
Lee†, S.,
Fumene Feruglio, P.,
Marzola, P.,
Nahrendorf, M.,
and Weissleder, R.
In vivo imaging is often severely compromised by cardiovascular and respiratory motion. Highly successful motion compensation techniques have been developed for clinical imaging (e.g. magnetic resonance imaging) but the use of more advanced techniques for intravital microscopy is largely unexplored. Here, we implement a sequential cardiorespiratory gating scheme (SCG) for averaged microscopy. We show that SCG is very efficient in eliminating motion artifacts, is highly practical, enables high signal-to-noise ratio (SNR) in vivo imaging, and yields large field of views. The technique is particularly useful for high-speed data acquisition or for imaging scenarios where the fluorescence signal is not significantly above noise or background levels. (c) 2013 Optical Society of America
@article{2013-BOE,author={Vinegoni<sup>#†</sup>, C. and Lee<sup>†</sup>, S. and Fumene Feruglio, P. and Marzola, P. and Nahrendorf, M. and Weissleder, R.},title={Sequential average segmented microscopy for high signal-to-noise ratio motion-artifact-free in vivo heart imaging},journal={Biomedical Optics Express},alternatejournal={Biomed. Opt. Exp.},year={2013},volume={4},number={10},pages={2095-2106},issn={2156-7085},doi={10.1364/boe.4.002095},pmcid={PMC3799669},pmid={24156067}}
24156067
PMC3799669
10.1364/boe.4.002095
2012
Nature
Myocardial infarction accelerates atherosclerosis
Dutta, P.,
Courties, G.,
Wei, Y.,
Leuschner, F.,
Gorbatov, R.,
Robbins, C. S.,
Iwamoto, Y.,
Thompson, B.,
Carlson, A. L.,
Heidt, T.,
Majmudar, M. D.,
Lasitschka, F.,
Etzrodt, M.,
Waterman, P.,
Waring, M. T.,
Chicoine, A. T.,
Laan, A. M.,
Niessen, H. W. M.,
Piek, J. J.,
Rubin, B. B.,
Butany, J.,
Stone, J. R.,
Katus, H. A.,
Murphy, S. A.,
Morrow, D. A.,
Sabatine, M. S.,
Vinegoni, C.,
Moskowitz, M. A.,
Pittet, M. J.,
Libby, P.,
Lin, C. P.,
Swirski, F. K.,
Weissleder, R.,
and Nahrendorf#, M.
During progression of atherosclerosis, myeloid cells destabilize lipid-rich plaques in the arterial wall and cause their rupture, thus triggering myocardial infarction and stroke. Survivors of acute coronary syndromes have a high risk of recurrent events for unknown reasons. Here we show that the systemic response to ischaemic injury aggravates chronic atherosclerosis. After myocardial infarction or stroke, Apoe(-/-) mice developed larger atherosclerotic lesions with a more advanced morphology. This disease acceleration persisted over many weeks and was associated with markedly increased monocyte recruitment. Seeking the source of surplus monocytes in plaques, we found that myocardial infarction liberated haematopoietic stem and progenitor cells from bone marrow niches via sympathetic nervous system signalling. The progenitors then seeded the spleen, yielding a sustained boost in monocyte production. These observations provide new mechanistic insight into atherogenesis and provide a novel therapeutic opportunity to mitigate disease progression.
@article{2012-NATURE,author={Dutta, P. and Courties, G. and Wei, Y. and Leuschner, F. and Gorbatov, R. and Robbins, C. S. and Iwamoto, Y. and Thompson, B. and Carlson, A. L. and Heidt, T. and Majmudar, M. D. and Lasitschka, F. and Etzrodt, M. and Waterman, P. and Waring, M. T. and Chicoine, A. T. and van der Laan, A. M. and Niessen, H. W. M. and Piek, J. J. and Rubin, B. B. and Butany, J. and Stone, J. R. and Katus, H. A. and Murphy, S. A. and Morrow, D. A. and Sabatine, M. S. and Vinegoni, C. and Moskowitz, M. A. and Pittet, M. J. and Libby, P. and Lin, C. P. and Swirski, F. K. and Weissleder, R. and Nahrendorf<sup>#</sup>, M.},title={Myocardial infarction accelerates atherosclerosis},journal={Nature},alternatejournal={Nature},year={2012},volume={487},number={7407},pages={325-329},issn={0028-0836},doi={10.1038/nature11260},pmcid={PMC3401326},pmid={22763456}}
22763456
PMC3401326
10.1038/nature11260
Cancer Res.
In Vivo Imaging of Drug-Induced Mitochondrial Outer Membrane Permeabilization at Single-Cell Resolution
Earley, S.,
Vinegoni, C.,
Dunham, J.,
Gorbatov, R.,
Fumene Feruglio, P.,
and Weissleder#, R.
Observing drug responses in the tumor microenvironment in vivo can be technically challenging. As a result, cellular responses to molecularly targeted cancer drugs are often studied in cell culture, which does not accurately represent the behavior of cancer cells growing in vivo. Using high-resolution microscopy and fluorescently labeled genetic reporters for apoptosis, we developed an approach to visualize drug-induced cell death at single-cell resolution in vivo. Stable expression of the mitochondrial intermembrane protein IMS-RP was established in human breast and pancreatic cancer cells. Image analysis was then used to quantify release of IMS-RP into the cytoplasm upon apoptosis and irreversible mitochondrial permeabilization. Both breast and pancreatic cancer cells showed higher basal apoptotic rates in vivo than in culture. To study drug-induced apoptosis, we exposed tumor cells to navitoclax (ABT-263), an inhibitor of Bcl-2, Bcl-xL, and Bcl-w, both in vitro and in vivo. Although the tumors responded to Bcl-2 inhibition in vivo, inducing apoptosis in around 20% of cancer cells, the observed response was much higher in cell culture. Together, our findings show an imaging technique that can be used to directly visualize cell death within the tumor microenvironment in response to drug treatment. Cancer Res; 72(12); 2949-56. (C)2012 AACR.
@article{2012-CNCRRES,author={Earley, S. and Vinegoni, C. and Dunham, J. and Gorbatov, R. and Fumene Feruglio, P. and Weissleder<sup>#</sup>, R.},title={In Vivo Imaging of Drug-Induced Mitochondrial Outer Membrane Permeabilization at Single-Cell Resolution},journal={Cancer Research},alternatejournal={Cancer Res.},year={2012},volume={72},number={12},pages={2949-2956},issn={0008-5472},doi={10.1158/0008-5472.Can-11-4096},pmcid={PMC3603290},pmid={22505651}}
22505651
PMC3603290
10.1158/0008-5472.Can-11-4096
P.N.A.S.
Implantable microenvironments to attract hematopoietic stem/cancer cells
Lee, J.,
Li, M.,
Milwid, J.,
Dunham, J.,
Vinegoni, C.,
Gorbatov, R.,
Iwamoto, Y.,
Wang, F. J.,
Shen, K. Y.,
Hatfield, K.,
Enger, M.,
Shafiee, S.,
McCormack, E.,
Ebert, B. L.,
Weissleder, R.,
Yarmush, M. L.,
and Parekkadan#, B.
Proceedings of the National Academy of Sciences of the United States of America 2012
The environments that harbor hematopoietic stem and progenitor cells are critical to explore for a better understanding of hematopoiesis during health and disease. These compartments often are inaccessible for controlled and rapid experimentation, thus limiting studies to the evaluation of conventional cell culture and transgenic animal models. Here we describe the manufacture and image-guided monitoring of an engineered microenvironment with user-defined properties that recruits hematopoietic progenitors into the implant. Using intravital imaging and fluorescence molecular tomography, we show in real time that the cell homing and retention process is efficient and durable for short-and long-term engraftment studies. Our results indicate that bone marrow stromal cells, precoated on the implant, accelerate the formation of new sinusoidal blood vessels with vascular integrity at the micro-capillary level that enhances the recruitment hematopoietic progenitor cells to the site. This implantable construct can serve as a tool enabling the study of hematopoiesis.
@article{2012-PNAS,author={Lee, J. and Li, M. and Milwid, J. and Dunham, J. and Vinegoni, C. and Gorbatov, R. and Iwamoto, Y. and Wang, F. J. and Shen, K. Y. and Hatfield, K. and Enger, M. and Shafiee, S. and McCormack, E. and Ebert, B. L. and Weissleder, R. and Yarmush, M. L. and Parekkadan<sup>#</sup>, B.},title={Implantable microenvironments to attract hematopoietic stem/cancer cells},journal={Proceedings of the National Academy of Sciences of the United States of America},alternatejournal={P.N.A.S.},year={2012},volume={109},number={48},pages={19638-19643},issn={0027-8424},doi={10.1073/pnas.1208384109},pmcid={PMC3786172},pmid={24086796}}
24086796
PMC3786172
10.1073/pnas.1208384109
J. Am. Coll. Cardiol.
PET/MRI of Inflammation in Myocardial Infarction
Lee, W. W.,
Marinelli, B.,
Laan, A. M.,
Sena, B. F.,
Gorbatov, R.,
Leuschner, F.,
Dutta, P.,
Iwamoto, Y.,
Ueno, T.,
Begieneman, M. P. V.,
Niessen, H. W. M.,
Piek, J. J.,
Vinegoni, C.,
Pittet, M. J.,
Swirski, F. K.,
Tawakol, A.,
Di Carli, M.,
Weissleder, R.,
and Nahrendorf#, M.
Journal of the American College of Cardiology 2012
Objectives The aim of this study was to explore post-myocardial infarction (MI) myocardial inflammation. Background Innate immune cells are centrally involved in infarct healing and are emerging therapeutic targets in cardiovascular disease; however, clinical tools to assess their presence in tissue are scarce. Furthermore, it is currently not known if the nonischemic remote zone recruits monocytes. Methods Acute inflammation was followed in mice with coronary ligation by 18-fluorodeoxyglucose ((18)FDG) positron emission tomography/magnetic resonance imaging, fluorescence-activated cell sorting, polymerase chain reaction, and histology. Results Gd-DTPA-enhanced infarcts showed high (18)FDG uptake on day 5 after MI. Cell depletion and isolation data confirmed that this largely reflected inflammation; CD11b(+) cells had 4-fold higher (18)FDG uptake than the infarct tissue from which they were isolated (p < 0.01). Surprisingly, there was considerable monocyte recruitment in the remote myocardium (approximately 10(4)/mg of myocardium, 5.6-fold increase; p < 0.01), a finding mirrored by macrophage infiltration in the remote myocardium of patients with acute MI. Temporal kinetics of cell recruitment were slower than in the infarct, with peak numbers on day 10 after ischemia. Quantitative polymerase chain reaction showed a robust increase of recruiting adhesion molecules and chemokines in the remote myocardium (e.g., 12-fold increase of monocyte chemoattractant protein-1), although levels were always lower than in the infarct. Finally, matrix metalloproteinase activity was significantly increased in noninfarcted myocardium, suggesting that monocyte recruitment to the remote zone may contribute to post-MI dilation. Conclusions This study shed light on the innate inflammatory response in remote myocardium after MI. (J Am Coll Cardiol 2012; 59: 153-63) (C) 2012 by the American College of Cardiology Foundation
@article{2012-JACC,author={Lee, W. W. and Marinelli, B. and van der Laan, A. M. and Sena, B. F. and Gorbatov, R. and Leuschner, F. and Dutta, P. and Iwamoto, Y. and Ueno, T. and Begieneman, M. P. V. and Niessen, H. W. M. and Piek, J. J. and Vinegoni, C. and Pittet, M. J. and Swirski, F. K. and Tawakol, A. and Di Carli, M. and Weissleder, R. and Nahrendorf<sup>#</sup>, M.},title={PET/MRI of Inflammation in Myocardial Infarction},journal={Journal of the American College of Cardiology},alternatejournal={J. Am. Coll. Cardiol.},year={2012},volume={59},number={2},pages={153-163},issn={0735-1097},doi={10.1016/j.jacc.2011.08.066},pmcid={PMC3257823},pmid={22222080}}
22222080
PMC3257823
10.1016/j.jacc.2011.08.066
Nat. Commun.
Real-time in vivo imaging of the beating mouse heart at microscopic resolution
Lee†, S.,
Vinegoni#†, C.,
Fumene Feruglio, P.,
Fexon, L.,
Gorbatov, R.,
Pivoravov, M.,
Sbarbati, A.,
Nahrendorf, M.,
and Weissleder, R.
Real-time imaging of moving organs and tissues at microscopic resolutions represents a major challenge in studying the complex biology of live animals. Here we present a technique based on a novel stabilizer setup combined with a gating acquisition algorithm for the imaging of a beating murine heart at the single-cell level. The method allows serial in vivo fluorescence imaging of the beating heart in live mice in both confocal and nonlinear modes over the course of several hours. We demonstrate the utility of this technique for in vivo optical sectioning and dual-channel time-lapse fluorescence imaging of cardiac ischaemia. The generic method could be adapted to other moving organs and thus broadly facilitate in vivo microscopic investigations.
@article{2012-NC,author={Lee<sup>†</sup>, S. and Vinegoni<sup>#†</sup>, C. and Fumene Feruglio, P. and Fexon, L. and Gorbatov, R. and Pivoravov, M. and Sbarbati, A. and Nahrendorf, M. and Weissleder, R.},title={Real-time in vivo imaging of the beating mouse heart at microscopic resolution},journal={Nature Communications},alternatejournal={Nat. Commun.},year={2012},volume={3},pages={8},issn={2041-1723},doi={10.1038/ncomms2060},pmcid={PMC3622400},pmid={22968700}}
22968700
PMC3622400
10.1038/ncomms2060
J. Biomed. Opt.
Improved intravital microscopy via synchronization of respiration and holder stabilization
Lee†, S.,
Vinegoni#†, C.,
Fumene Feruglio, P.,
and Weissleder, R.
A major challenge in high-resolution intravital confocal and multiphoton microscopy is physiologic tissue movement during image acquisition. Of the various physiological sources of movement, respiration has arguably the largest and most wide-ranging effect. We describe a technique for achieving stabilized microscopy imaging using a dual strategy. First, we designed a mechanical stabilizer for constraining physical motion; this served to simultaneously increase the in-focus range over which data can be acquired as well as increase the reproducibility of imaging a certain position within each confocal imaging plane. Second, by implementing a retrospective breathing-gated imaging modality, we performed selective image extraction gated to a particular phase of the respiratory cycle. Thanks to the high reproducibility in position, all gated images presented a high degree of correlation over time. The images obtained using this technique not only showed significant improvements over images acquired without the stabilizer, but also demonstrated accurate in vivo imaging during longitudinal studies. The described methodology is easy to implement with any commercial imaging system, as are used by most biological imaging laboratories, and can be used for both confocal and multiphoton laser scanning microscopy. c 2012 Society of Photo-Optical Instrumentation Engineers (SPIE). [DOI: 10.1117/1.JBO.17.9.096018]
@article{2012-JBO,author={Lee<sup>†</sup>, S. and Vinegoni<sup>#†</sup>, C. and Fumene Feruglio, P. and Weissleder, R.},title={Improved intravital microscopy via synchronization of respiration and holder stabilization},journal={Journal of Biomedical Optics},alternatejournal={J. Biomed. Opt.},year={2012},volume={17},number={9},pages={8},issn={1083-3668},doi={10.1117/1.Jbo.17.9.096018},pmcid={PMC3449295},pmid={23085919}}
23085919
PMC3449295
10.1117/1.Jbo.17.9.096018
Bioconjugate Chem.
Optochemogenetics (OCG) Allows More Precise Control of Genetic Engineering in Mice with CreER regulators
Lu, X.,
Agasti, S. S.,
Vinegoni, C.,
Waterman, P.,
DePinho, R. A.,
and Weissleder#, R.
New approaches that allow precise spatiotemporal control of gene expression in model organisms at the single cell level are necessary to better dissect the role of specific genes and cell populations in development, disease, and therapy. Here, we describe a new optochemogenetic switch (OCG switch) to control CreER/loxP-mediated recombination via photoactivatable ("caged") tamoxifen analogues in individual cells in cell culture, organoid culture, and in vivo in adult mice. This approach opens opportunities to more fully exploit existing CreER transgenic mouse strains to achieve more precise temporal- and location-specific regulation of genetic events and gene expression.
@article{2012-BIOCHEM,author={Lu, X. and Agasti, S. S. and Vinegoni, C. and Waterman, P. and DePinho, R. A. and Weissleder<sup>#</sup>, R.},title={Optochemogenetics (OCG) Allows More Precise Control of Genetic Engineering in Mice with CreER regulators},journal={Bioconjugate Chemistry},alternatejournal={Bioconjugate Chem.},year={2012},volume={23},number={9},pages={1945-1951},issn={1043-1802},doi={10.1021/bc300319c},pmcid={PMC3775343},pmid={22917215}}
22917215
PMC3775343
10.1021/bc300319c
Curr. Pharm. Biotech.
Deep Tissue Optical and Optoacoustic Molecular Imaging Technologies for Pre-Clinical Research and Drug Discovery
Razansky#, D.,
Deliolanis, N. C.,
Vinegoni, C.,
and Ntziachristos, V.
For centuries, biological discoveries were based on optical imaging, in particular microscopy but also several chromophoric assays and photographic approaches. With the recent emergence of methods appropriate for bio-marker in vivo staining, such as bioluminescence, fluorescent molecular probes and proteins, as well as nanoparticle-based targeted agents, significant attention has been shifted toward in vivo interrogations of different dynamic biological processes at the molecular level. This progress has been largely supported by the development of advanced tomographic imaging technologies suitable for obtaining volumetric visualization of bio-marker distributions in small animals at a whole-body or whole-organ scale, an imaging frontier that is not accessible by the existing tissue-sectioning microscopic techniques due to intensive light scattering beyond the depth of a few hundred microns. Major examples of such recently developed optical imaging modalities are reviewed here, including bioluminescence tomography (BLT), fluorescence molecular tomography (FMT), and optical projection tomography (OPT). The pharmaceutical imaging community has quickly appropriated itself of these novel forms of optical imaging, since they come with very compelling advantages, such as quantitative three-dimensional capabilities, direct correlation to the biological cultures, easiness and cost-effectiveness of use, and the use of safe non-ionizing radiation. Some multi-modality approaches, combining light with other imaging modalities such as X-Ray CT or MRI, giving the ability to acquire both an optical contrast reconstruction along with a hi-fidelity anatomical images, are also reviewed. A separate section is devoted to the hybrid imaging techniques based on the optoacoustic phenomenon, such as multispectral optoacoustic tomography (MSOT), which are poised to leverage the traditional contrast and specificity advantages of optical spectrum by delivering an ever powerful set of capabilities, including real-time operation and high spatial resolution, not affected by the scattering nature of biological tissues.
@article{2012-CPB,author={Razansky<sup>#</sup>, D. and Deliolanis, N. C. and Vinegoni, C. and Ntziachristos, V.},title={Deep Tissue Optical and Optoacoustic Molecular Imaging Technologies for Pre-Clinical Research and Drug Discovery},journal={Current Pharmaceutical Biotechnology},alternatejournal={Curr. Pharm. Biotech.},year={2012},volume={13},number={4},pages={504-522},issn={1389-2010},doi={10.2174/138920112799436258},pmid={22216767}}
22216767
10.2174/138920112799436258
Neoplasia
Imaging Therapeutic PARP Inhibition In Vivo through Bioorthogonally Developed Companion Imaging Agents
Reiner, T.,
Lacy, J.,
Keliher, E. J.,
Yang, K. S.,
Ullal, A.,
Kohler, R. H.,
Vinegoni, C.,
and Weissleder#, R.
A number of small-molecule poly (ADP-ribose) polymerase (PARP) inhibitors are currently undergoing advanced clinical trials. Determining the distribution and target inhibitory activity of these drugs in individual subjects, however, has proven problematic. Here, we used a PARP agent for positron emission tomography-computed tomography (PET-CT) imaging (F-18-BO), which we developed based on the Olaparib scaffold using rapid bioorthogonal conjugation chemistries. We show that the bioorthogonal F-18 modification of the parent molecule is simple, highly efficient, and well tolerated, resulting in a half maximal inhibitory concentration (IC50) of 17.9 +/- 1.1 nM. Intravital imaging showed ubiquitous distribution of the drug and uptake into cancer cells, with ultimate localization within the nucleus, all of which were inhibitable. Whole-body PET-CT imaging showed tumoral uptake of the drug, which decreased significantly, after a daily dose of Olaparib. Standard F-18-fludeoxyglucose imaging, however, failed to detect such therapy-induced changes. This research represents a step toward developing a more generic approach for the rapid codevelopment of companion imaging agents based on small-molecule therapeutic inhibitors.
@article{2012-NEOPLASIA,author={Reiner, T. and Lacy, J. and Keliher, E. J. and Yang, K. S. and Ullal, A. and Kohler, R. H. and Vinegoni, C. and Weissleder<sup>#</sup>, R.},title={Imaging Therapeutic PARP Inhibition In Vivo through Bioorthogonally Developed Companion Imaging Agents},journal={Neoplasia},alternatejournal={Neoplasia},year={2012},volume={14},number={3},pages={169-+},issn={1476-5586},doi={10.1593/neo.12414},pmcid={PMC3323895},pmid={22496617}}
22496617
PMC3323895
10.1593/neo.12414
PLoS One
Mapping Molecular Agents Distributions in Whole Mice Hearts Using Born-Normalized Optical Projection Tomography
Vinegoni#†, C.,
Fumene Feruglio†, P.,
Razansky, D.,
Gorbatov, R.,
Ntziachristos, V.,
Sbarbati, A.,
Nahrendorf, M.,
and Weissleder, R.
To date there is a lack of tools to map the spatio-temporal dynamics of diverse cells in experimental heart models. Conventional histology is labor intensive with limited coverage, whereas many imaging techniques do not have sufficiently high enough spatial resolution to map cell distributions. We have designed and built a high resolution, dual channel Born-normalized near-infrared fluorescence optical projection tomography system to quantitatively and spatially resolve molecular agents distribution within whole murine heart. We validated the use of the system in a mouse model of monocytes/macrophages recruitment during myocardial infarction. While acquired, data were processed and reconstructed in real time. Tomographic analysis and visualization of the key inflammatory components were obtained via a mathematical formalism based on left ventricular modeling. We observed extensive monocyte recruitment within and around the infarcted areas and discovered that monocytes were also extensively recruited into non-ischemic myocardium, beyond that of injured tissue, such as the septum.
@article{2012-PLOSONE,author={Vinegoni<sup>#†</sup>, C. and Fumene Feruglio<sup>†</sup>, P. and Razansky, D. and Gorbatov, R. and Ntziachristos, V. and Sbarbati, A. and Nahrendorf, M. and Weissleder, R.},title={Mapping Molecular Agents Distributions in Whole Mice Hearts Using Born-Normalized Optical Projection Tomography},journal={Plos One},alternatejournal={PLoS One},year={2012},volume={7},number={4},pages={9},issn={1932-6203},doi={10.1371/journal.pone.0034427},pmcid={PMC3324534},pmid={22509302}}
22509302
PMC3324534
10.1371/journal.pone.0034427
Intravital
Motion compensation using a suctioning stabilizer for intravital microscopy
Vinegoni#†, C.,
Lee†, S.,
Gorbatov, R.,
and Weissleder, R.
Motion artifacts continue to present a major challenge to single cell imaging in cardiothoracic organs such as the beating
heart, blood vessels or lung. In this study, we present a new water-immersion suctioning stabilizer that enables minimally
invasive intravital fluorescence microscopy using water-based stick objectives. The stabilizer works by reducing major
motion excursions and can be used in conjunction with both prospective or retrospective gating approaches. We show
that the new approach offers cellular resolution in the beating murine heart without perturbing normal physiology.
In addition, because this technique allows multiple areas to be easily probed, it offers the opportunity for wide area
coverage at high resolution.
@article{2012-INTRVTL,author={Vinegoni<sup>#†</sup>, C. and Lee<sup>†</sup>, S. and Gorbatov, R. and Weissleder, R.},title={Motion compensation using a suctioning stabilizer for intravital microscopy},journal={Intravital},alternatejournal={Intravital},year={2012},volume={1},pages={115},issn={1083-3668},doi={10.4161/intv.23017},pmcid={PMC3786172},pmid={24086796}}
24086796
PMC3786172
10.4161/intv.23017
Angew. Chem.
Bioorthogonal Imaging of Aurora Kinase A in Live Cells
Yang, K. S.,
Budin, G.,
Reiner, T.,
Vinegoni, C.,
and Weissleder#, R.
Cell division (mitosis) is a tightly controlled process that is coordinated and regulated by a network of proteins localized in the nucleus. The key stages of mitosis are centrosome maturation, chromosome condensation, nuclear envelope breakdown, centrosome separation, bipolar spindle formation, chromosome separation, and finally cytokinesis. Aurora kinase A (AKA) belongs to the Aurora kinase family of serine/threonine kinases, which have been shown toplaycriticalrolesinmitoticprogression. Duringmitosis, AKA localizes to centrosomes during late S to early G2 phase. As the cell proceeds to metaphase, AKA localizes to the microtubules and near the spindle poles, where it remains until anaphase when it migrates to some extent to the spindle midzone. Finally, during cytokinesis, AKA localizes to the midbody. Whilst localized to these specific cellular regions, AKA interacts with and phosphorylates several intracellular targets, including p53, MBD3, and BRCA1, each of which are critical mediators of malignant transformation. The unique stage-specific nuclear and intracellular locations of AKA during mitosis thus make it an interesting imaging target
@article{2012-AC,author={Yang, K. S. and Budin, G. and Reiner, T. and Vinegoni, C. and Weissleder<sup>#</sup>, R.},title={Bioorthogonal Imaging of Aurora Kinase A in Live Cells},journal={Angewandte Chemie-International Edition},alternatejournal={Angew. Chem.},year={2012},volume={51},number={27},pages={6598-6603},issn={1433-7851},doi={10.1002/anie.201200994},pmcid={PMC3523717},pmid={22644700}}
22644700
PMC3523717
10.1002/anie.201200994
2011
World J.Surg.
Intraoperative Near-infrared Fluorescent Cholangiography (NIRFC) in Mouse Models of Bile Duct Injury: Reply
Figueiredo, J. L.,
Nahrendorf, M.,
Vinegoni, C.,
and Weissleder#, R.
Near-infrared (NIR) imaging has become an important minimally invasive, surgical and preclinical tool. This is so because light in the 650–900 nm window traverses tissue more effectively than light in the visible range, and also because less autofluorescence occurs in this region. Ultimately, the goal for NIR imaging is to use multiple fluorescent probes in the same way as multicolor imaging (fluorescent intravital livemicroscopy, FILM),toreport ona variety of anatomic, physiologic, or molecular events during real-time surgical intervention.
@article{2010-WJS,author={Figueiredo, J. L. and Nahrendorf, M. and Vinegoni, C. and Weissleder<sup>#</sup>, R.},title={Intraoperative Near-infrared Fluorescent Cholangiography (NIRFC) in Mouse Models of Bile Duct Injury: Reply},journal={World Journal of Surgery},alternatejournal={World J.Surg.},year={2011},volume={35},number={3},pages={694-695},issn={0364-2313},doi={10.1007/s00268-010-0728-5},pmid={20645091}}
20645091
10.1007/s00268-010-0728-5
Eur. Biophys. J.
A multimodal approach for tracing lateralisation along the olfactory pathway in the honeybee through electrophysiological recordings, morpho-functional imaging, and behavioural studies
Haase, A.,
Rigosi, E.,
Frasnelli, E.,
Trona, F.,
Tessarolo, F.,
Vinegoni, C.,
Anfora, G.,
Vallortigara, G.,
and Antolini#, R.
European Biophysics Journal with Biophysics Letters 2011
Recent studies have revealed asymmetries between the left and right sides of the brain in invertebrate species. Here we present a review of a series of recent studies from our laboratories, aimed at tracing asymmetries at different stages along the honeybee’s (Apis mellifera) olfactory pathway. These include estimates of the number of sensilla present on the two antennae, obtained by scanning electron microscopy, as well as electroantennography recordings of the left and right antennal responses to odorants. We describe investigative studies of the antennal lobes, where multi-photon microscopy was used to search for possible morphological asymmetries between the two brain sides. Moreover, we report on recently published results obtained by two-photon calcium imaging for functional mapping of the antennal lobe aimed at comparing patterns of activity evoked by different odours. Finally, possible links to the results of behavioural tests, measuring asymmetries in single-sided olfactory memory recall, are discussed.
@article{2011-EB,author={Haase, A. and Rigosi, E. and Frasnelli, E. and Trona, F. and Tessarolo, F. and Vinegoni, C. and Anfora, G. and Vallortigara, G. and Antolini<sup>#</sup>, R.},title={A multimodal approach for tracing lateralisation along the olfactory pathway in the honeybee through electrophysiological recordings, morpho-functional imaging, and behavioural studies},journal={European Biophysics Journal with Biophysics Letters},alternatejournal={Eur. Biophys. J.},year={2011},volume={40},number={11},pages={1247-1258},issn={0175-7571},doi={10.1007/s00249-011-0748-6},pmcid={PMC3366498},pmid={21956452}}
21956452
PMC3366498
10.1007/s00249-011-0748-6
Biomed. Opt. Exp.
In-vivo two-photon imaging of the honey bee antennal lobe
Haase, A.,
Rigosi, E.,
Trona, F.,
Anfora, G.,
Vallortigara, G.,
Antolini, R.,
and Vinegoni#, C.
Due to the honey bee’s importance as a simple neural model, there is a great need for new functional imaging modalities. Herein we report on the development and new finding of a combined two-photon microscope with a synchronized odor stimulus platform for in-vivo functional and morphological imaging of the honey bee’s olfactory system focusing on its primary centers, the antennal lobes (ALs). Our imaging platform allows for simultaneously obtaining both morphological measurements of the AL’s functional units, the glomeruli, and in-vivo calcium recording of their neural activities. By applying external odor stimuli to the bee’s antennae, we were able to record the characteristic glomerular odor response maps. Compared to previous works where conventional fluorescenc microscopy was used, our approach has been demonstrated to offer all the advantages of multi-photon imaging, providing substantial enhancement in both spatial and temporal resolutions while minimizing photo-damages. In addition, compared to previous full-fiel microscopy calcium recordings, a four-fold improvement in the functional signal has been achieved. Finally, the multi-photon associated extended penetration depth allows for functional imaging of profound glomeruli. (C) 2010 Optical Society of America
@article{2010-BOEX,author={Haase, A. and Rigosi, E. and Trona, F. and Anfora, G. and Vallortigara, G. and Antolini, R. and Vinegoni<sup>#</sup>, C.},title={In-vivo two-photon imaging of the honey bee antennal lobe},journal={Biomedical Optics Express},alternatejournal={Biomed. Opt. Exp.},year={2011},volume={2},number={1},pages={131-138},issn={2156-7085},doi={10.1364/boe.2.000131},pmcid={PMC3028488},pmid={21326643}}
21326643
PMC3028488
10.1364/boe.2.000131
P.N.A.S.
Accurate measurement of pancreatic islet beta-cell mass using a second-generation fluorescent exendin-4 analog
Reiner, T.,
Thurber, G.,
Gaglia, J.,
Vinegoni, C.,
Liew, C. W.,
Upadhyay, R.,
Kohler, R. H.,
Li, L.,
Kulkarni, R. N.,
Benoist, C.,
Mathis, D.,
and Weissleder#, R.
Proceedings of the National Academy of Sciences of the United States of America 2011
The hallmark of type 1 diabetes is autoimmune destruction of the insulin-producing beta-cells of the pancreatic islets. Autoimmune diabetes has been difficult to study or treat because it is not usually diagnosed until substantial beta-cell loss has already occurred. Imaging agents that permit noninvasive visualization of changes in beta-cell mass remain a high-priority goal. We report on the development and testing of a near-infrared fluorescent beta-cell imaging agent. Based on the amino acid sequence of exendin-4, we created a neopeptide via introduction of an unnatural amino acid at the K(12) position, which could subsequently be conjugated to fluorophores via bioorthogonal copper-catalyzed click-chemistry. Cell assays confirmed that the resulting fluorescent probe (E4(x12)-VT750) had a high binding affinity (similar to 3 nM). Its in vivo properties were evaluated using high-resolution intravital imaging, histology, whole-pancreas visualization, and endoscopic imaging. According to intravital microscopy, the probe rapidly bound to beta-cells and, as demonstrated by confocal microscopy, it was internalized. Histology of the whole pancreas showed a close correspondence between fluorescence and insulin staining, and there was an excellent correlation between imaging signals and beta-cell mass in mice treated with streptozotocin, a beta-cell toxin. Individual islets could also be visualized by endoscopic imaging. In short, E4(x12)-VT750 showed strong and selective binding to glucose-like peptide-1 receptors and permitted accurate measurement of beta-cell mass in both diabetic and nondiabetic mice. This near-infrared imaging probe, as well as future radioisotope-labeled versions of it, should prove to be important tools for monitoring diabetes, progression, and treatment in both experimental and clinical contexts.
@article{2011-PNAS,author={Reiner, T. and Thurber, G. and Gaglia, J. and Vinegoni, C. and Liew, C. W. and Upadhyay, R. and Kohler, R. H. and Li, L. and Kulkarni, R. N. and Benoist, C. and Mathis, D. and Weissleder<sup>#</sup>, R.},title={Accurate measurement of pancreatic islet beta-cell mass using a second-generation fluorescent exendin-4 analog},journal={Proceedings of the National Academy of Sciences of the United States of America},alternatejournal={P.N.A.S.},year={2011},volume={108},number={31},pages={12815-12820},issn={0027-8424},doi={10.1073/pnas.1109859108},pmcid={PMC3150928},pmid={21768367}}
21768367
PMC3150928
10.1073/pnas.1109859108
Behav. Brain Res.
Searching for anatomical correlates of olfactory lateralization in the honeybee antennal lobes: A morphological and behavioural study
Rigosi#, E.,
Frasnelli, E.,
Vinegoni, C.,
Antolini, R.,
Anfora, G.,
Vallortigara, G.,
and Haase, A.
The honeybee, Apis mellifera L (Hymenoptera: Apidae), has recently become a model for studying brain asymmetry among invertebrates. A strong lateralization favouring the right antenna was discovered in odour learning and short-term memory recall experiments, and a lateral shift favouring the left antenna for long-term memory recall. Corresponding morphological asymmetries have been found in the distribution of olfactory sensilla between the antennae and confirmed by electrophysiological odour response measurements in isolated right and left antennae. The aim of this study was to investigate whether a morphological asymmetry can be observed in the volume of the primary olfactory centres of the central nervous system, the antennal lobes (ALs). Precise volume measurements of a subset of their functional units, the glomeruli, were performed in both sides of the brain, exploiting the advantages of two-photon microscopy. This novel method allowed minimal invasive acquisition of volume images of the ALs, avoiding artefacts from brain extraction and dehydration. The study was completed by a series of behavioural experiments in which response asymmetry in odour recall following proboscis extension reflex conditioning was assessed for odours, chosen to stimulate strong activity in the same glomeruli as in the morphological study. The volumetric measurements found no evidence of lateralization in the investigated glomeruli within the experimental limits. Instead, in the behavioural experiments, a striking odour dependence of the lateralization was observed. The results are discussed on the basis of recent neurophysiological and ethological experiments in A. mellifera. (C) 2011 Elsevier BM. All rights reserved.
@article{2011-BBR,author={Rigosi<sup>#</sup>, E. and Frasnelli, E. and Vinegoni, C. and Antolini, R. and Anfora, G. and Vallortigara, G. and Haase, A.},title={Searching for anatomical correlates of olfactory lateralization in the honeybee antennal lobes: A morphological and behavioural study},journal={Behavioural Brain Research},alternatejournal={Behav. Brain Res.},year={2011},volume={221},number={1},pages={290-294},issn={0166-4328},doi={10.1016/j.bbr.2011.03.015},pmcid={PMC3089663},pmid={21402106}}
21402106
PMC3089663
10.1016/j.bbr.2011.03.015
Sci. Transl. Med.
Indocyanine Green Enables Near-Infrared Fluorescence Imaging of Lipid-Rich, Inflamed Atherosclerotic Plaques
Vinegoni, C.,
Botnaru, I.,
Aikawa, E.,
Calfon, M. A.,
Iwamoto, Y.,
Folco, E. J.,
Ntziachristos, V.,
Weissleder, R.,
Libby, P.,
and Jaffer#, F. A.
New high-resolution molecular and structural imaging strategies are needed to visualize high-risk plaques that are likely to cause acute myocardial infarction, because current diagnostic methods do not reliably identify at-risk subjects. Although molecular imaging agents are available for low-resolution detection of atherosclerosis in large arteries, a lack of imaging agents coupled to high-resolution modalities has limited molecular imaging of atherosclerosis in the smaller coronary arteries. Here, we have demonstrated that indocyanine green (ICG), a Food and Drug Administration-approved near-infrared fluorescence (NIRF)-emitting compound, targets atheromas within 20 min of injection and provides sufficient signal enhancement for in vivo detection of lipid-rich, inflamed, coronary-sized plaques in atherosclerotic rabbits. In vivo NIRF sensing was achieved with an intravascular wire in the aorta, a vessel of comparable caliber to human coronary arteries. Ex vivo fluorescence reflectance imaging showed high plaque target-to-background ratios in atheroma-bearing rabbits injected with ICG compared to atheroma-bearing rabbits injected with saline. In vitro studies using human macrophages established that ICG preferentially targets lipid-loaded macrophages. In an early clinical study of human atheroma specimens from four patients, we found that ICG colocalized with plaque macrophages and lipids. The atheroma-targeting capability of ICG has the potential to accelerate the clinical development of NIRF molecular imaging of high-risk plaques in humans.
@article{2011-STM,author={Vinegoni, C. and Botnaru, I. and Aikawa, E. and Calfon, M. A. and Iwamoto, Y. and Folco, E. J. and Ntziachristos, V. and Weissleder, R. and Libby, P. and Jaffer<sup>#</sup>, F. A.},title={Indocyanine Green Enables Near-Infrared Fluorescence Imaging of Lipid-Rich, Inflamed Atherosclerotic Plaques},journal={Science Translational Medicine},alternatejournal={Sci. Transl. Med.},year={2011},volume={3},number={84},pages={9},issn={1946-6234},doi={10.1126/scitranslmed.3001577},pmcid={PMC3112179},pmid={21613624}}
21613624
PMC3112179
10.1126/scitranslmed.3001577
2010
J. Biomed. Opt.
Intravascular near-infrared fluorescence molecular imaging of atherosclerosis: toward coronary arterial visualization of biologically high-risk plaques
Calfon, M. A.,
Vinegoni, C.,
Ntziachristos, V.,
and Jaffer#, F. A.
New imaging methods are urgently needed to identify high-risk atherosclerotic lesions prior to the onset of myocardial infarction, stroke, and ischemic limbs. Molecular imaging offers a new approach to visualize key biological features that characterize high-risk plaques associated with cardiovascular events. While substantial progress has been realized in clinical molecular imaging of plaques in larger arterial vessels (carotid, aorta, iliac), there remains a compelling, unmet need to develop molecular imaging strategies targeted to high-risk plaques in human coronary arteries. We present recent developments in intravascular near-IR fluorescence catheter-based strategies for in vivo detection of plaque inflammation in coronary-sized arteries. In particular, the biological, light transmission, imaging agent, and engineering principles that underlie a new intravascular near-IR fluorescence sensing method are discussed. Intravascular near-IR fluorescence catheters appear highly translatable to the cardiac catheterization laboratory, and thus may offer a new in vivo method to detect high-risk coronary plaques and to assess novel atherosclerosis biologics. (C) 2010 Society of Photo-Optical Instrumentation Engineers. [DOI: 10.1117/1.3280282]
@article{2010-JBO,author={Calfon, M. A. and Vinegoni, C. and Ntziachristos, V. and Jaffer<sup>#</sup>, F. A.},title={Intravascular near-infrared fluorescence molecular imaging of atherosclerosis: toward coronary arterial visualization of biologically high-risk plaques},journal={Journal of Biomedical Optics},alternatejournal={J. Biomed. Opt.},year={2010},volume={15},number={1},pages={6},issn={1083-3668},doi={10.1117/1.3280282},pmcid={PMC3188610},pmid={20210433}}
20210433
PMC3188610
10.1117/1.3280282
Phys. Med. Biol.
Block matching 3D random noise filtering for absorption optical projection tomography
Fumene Feruglio†, P.,
Vinegoni#†, C.,
Gros, J.,
Sbarbati, A.,
and Weissleder, R.
Absorption and emission optical projection tomography (OPT), alternatively referred to as optical computed tomography (optical-CT) and optical-emission computed tomography (optical-ECT), are recently developed three-dimensional imaging techniques with value for developmental biology and ex vivo gene expression studies. The techniques’ principles are similar to the ones used for x-ray computed tomography and are based on the approximation of negligible light scattering in optically cleared samples. The optical clearing is achieved by a chemical procedure which aims at substituting the cellular fluids within the sample with a cell membranes’ index matching solution. Once cleared the sample presents very low scattering and is then illuminated with a light collimated beam whose intensity is captured in transillumination mode by a CCD camera. Different projection images of the sample are subsequently obtained over a 360 degrees full rotation, and a standard backprojection algorithm can be used in a similar fashion as for x-ray tomography in order to obtain absorption maps. Because not all biological samples present significant absorption contrast, it is not always possible to obtain projections with a good signal-to-noise ratio, a condition necessary to achieve high-quality tomographic reconstructions. Such is the case for example, for early stage’s embryos. In this work we demonstrate how, through the use of a random noise removal algorithm, the image quality of the reconstructions can be considerably improved even when the noise is strongly present in the acquired projections. Specifically, we implemented a block matching 3D (BM3D) filter applying it separately on each acquired transillumination projection before performing a complete three-dimensional tomographical reconstruction. To test the efficiency of the adopted filtering scheme, a phantom and a real biological sample were processed. In both cases, the BM3D filter led to a signal-to-noise ratio increment of over 30 dB on severe noise-affected reconstructions revealing original-noise-hidden-image details. These results show the utility of the BM3D approach for OPT under typical conditions of very low light absorption, suggesting its implementation as an efficient alternative to other filtering schemes such as for example the median filter.
@article{2010-PMB,author={Fumene Feruglio<sup>†</sup>, P. and Vinegoni<sup>#†</sup>, C. and Gros, J. and Sbarbati, A. and Weissleder, R.},title={Block matching 3D random noise filtering for absorption optical projection tomography},journal={Physics in Medicine and Biology},alternatejournal={Phys. Med. Biol.},year={2010},volume={55},number={18},pages={5401-5415},issn={0031-9155},doi={10.1088/0031-9155/55/18/009},pmcid={PMC2934766},pmid={20736500}}
20736500
PMC2934766
10.1088/0031-9155/55/18/009
Curr. Biol.
WNT5A/JNK and FGF/MAPK Pathways Regulate the Cellular Events Shaping the Vertebrate Limb Bud
Gros, J.,
Hu, J. K. H.,
Vinegoni, C.,
Fumene Feruglio, P.,
Weissleder, R.,
and Tabin#, C. J.
Background The vertebrate limb is a classical model for understanding patterning of three-dimensional structures during embryonic development Although decades of research have elucidated the tissue and molecular interactions within the limb bud required for patterning and morphogenesis of the limb, the cellular and molecular events that shape the limb bud itself have remained largely unknown Results We show that the mesenchymal cells of the early limb bud are not disorganized within the ectoderm as previously thought but are instead highly organized and polarized Using time-lapse video microscopy, we demonstrate that cells move and divide according to this orientation The combination of oriented cell divisions and movements drives the proximaldistal elongation of the limb bud necessary to set the stage for subsequent morphogenesis These cellular events are regulated by the combined activities of the WNT and FGF pathways We show that WNT5A/JNK is necessary for the proper orientation of cell movements and cell division In contrast, the FGF/MAPK signaling pathway, emanating from the apical ectodermal ridge, does not regulate cell orientation in the limb bud but instead establishes a gradient of cell velocity enabling continuous rearrangement of the cells at the distal tip of the limb Conclusions Together these data shed light on the cellular basis of vertebrate limb bud morphogenesis and uncover new layers to the sequential signaling pathways acting during vertebrate limb development
@article{2010-CB,author={Gros, J. and Hu, J. K. H. and Vinegoni, C. and Fumene Feruglio, P. and Weissleder, R. and Tabin<sup>#</sup>, C. J.},title={WNT5A/JNK and FGF/MAPK Pathways Regulate the Cellular Events Shaping the Vertebrate Limb Bud},journal={Current Biology},alternatejournal={Curr. Biol.},year={2010},volume={20},number={22},pages={1993-2002},issn={0960-9822},doi={10.1016/j.cub.2010.09.063},pmcid={PMC3188610},pmid={20210433}}
20210433
PMC3188610
10.1016/j.cub.2010.09.063
P.N.A.S.
Hybrid PET-optical imaging using targeted probes
Nahrendorf†#, M.,
Keliher†, E.,
Marinelli, B.,
Waterman, P.,
Fumene Feruglio, P.,
Fexon, L.,
Pivovarov, M.,
Swirski, F. K.,
Pittet, M. J.,
Vinegoni, C.,
and Weissleder#, R.
Proceedings of the National Academy of Sciences of the United States of America 2010
Fusion imaging of radionuclide-based molecular (PET) and structural data [x-ray computed tomography (CT)] has been firmly established. Here we show that optical measurements [fluorescence-mediated tomography (FMT)] show exquisite congruence to radionuclide measurements and that information can be seamlessly integrated and visualized. Using biocompatible nanoparticles as a generic platform (containing a F-18 isotope and a far red fluorochrome), we show good correlations between FMT and PET in probe concentration (r(2) > 0.99) and spatial signal distribution (r(2) > 0.85). Using a mouse model of cancer and different imaging probes to measure tumoral proteases, macrophage content and integrin expression simultaneously, we demonstrate the distinct tumoral locations of probes in multiple channels in vivo. The findings also suggest that FMT can serve as a surrogate modality for the screening and development of radionuclide-based imaging agents.
@article{2010-PNAS,author={Nahrendorf<sup>†#</sup>, M. and Keliher<sup>†</sup>, E. and Marinelli, B. and Waterman, P. and Fumene Feruglio, P. and Fexon, L. and Pivovarov, M. and Swirski, F. K. and Pittet, M. J. and Vinegoni, C. and Weissleder<sup>#</sup>, R.},title={Hybrid PET-optical imaging using targeted probes},journal={Proceedings of the National Academy of Sciences of the United States of America},alternatejournal={P.N.A.S.},year={2010},volume={107},number={17},pages={7910-7915},issn={0027-8424},doi={10.1073/pnas.0915163107},pmcid={PMC2867879},pmid={20385821}}
20385821
PMC2867879
10.1073/pnas.0915163107
Opt. Lett.
Imaging of molecular probe activity with Born-normalized fluorescence optical projection tomography
Vinegoni#, C.,
Fumene Feruglio, P.,
Cortez-Retamozo, V.,
Razansky, D.,
Medoff, B. D.,
Ntziachristos, V.,
Sbarbati, A.,
Pittet, M.,
and Weissleder, R.
Optical projection tomography is a new ex vivo imaging technique that allows imaging of whole organs in three dimensions at high spatial resolutions. In this Letter we demonstrate its capability to tomographically visualize molecular activity in whole organs of mice. In particular, eosinophil activity in asthmatic lungs is resolved using a Born-normalized fluorescence optical projection tomography and employing a near-IR molecular probe. The possibility to achieve molecularly sensitive imaging contrast in optical projection tomography by means of targeted and activatable imaging reporter agents adds a new range of capabilities for investigating molecular signatures of pathophysiological processes and a wide variety of diseases and their development. (C) 2010 Optical Society of America
@article{2010-OL,author={Vinegoni<sup>#</sup>, C. and Fumene Feruglio, P. and Cortez-Retamozo, V. and Razansky, D. and Medoff, B. D. and Ntziachristos, V. and Sbarbati, A. and Pittet, M. and Weissleder, R.},title={Imaging of molecular probe activity with Born-normalized fluorescence optical projection tomography},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2010},volume={35},number={7},pages={1088-1090},issn={0146-9592},doi={10.1364/ol.35.001088},pmcid={PMC2900933},pmid={20364226}}
20364226
PMC2900933
10.1364/ol.35.001088
2009
BioOptics World
Unprecedent in vivo views at the mesoscopic scale
Razansky, R.,
Vinegoni, C.,
and Ntziachristos#, V.
New optical and optoacustic methods are solving the bioscientist’s conundrum of imaging deep into living tissue at high resolution.
@article{2009-BOW,author={Razansky, R. and Vinegoni, C. and Ntziachristos<sup>#</sup>, V.},title={Unprecedent in vivo views at the mesoscopic scale},journal={BioOptics World},alternatejournal={BioOptics World},year={2009},volume={2},pages={22},}
Nat. Photonics
Multispectral opto-acoustic tomography of deep-seated fluorescent proteins in vivo
Razansky#, D.,
Distel, M.,
Vinegoni, C.,
Ma, R.,
Perrimon, N.,
Koster, R. W.,
and Ntziachristos#, V.
Fluorescent proteins have become essential reporter molecules for studying life at the cellular and sub-cellular level, re-defining the ways in which we investigate biology. However, because of intense light scattering, most organisms and tissues remain inaccessible to current fluorescence microscopy techniques at depths beyond several hundred micrometres. We describe a multispectral opto-acoustic tomography technique capable of high-resolution visualization of fluorescent proteins deep within highly light-scattering living organisms. The method uses multiwavelength illumination over multiple projections combined with selective-plane opto-acoustic detection for artifact-free data collection. Accurate image reconstruction is enabled by making use of wavelength-dependent light propagation models in tissue. By performing whole-body imaging of two biologically important and optically diffuse model organisms, Drosophila melanogaster pupae and adult zebrafish, we demonstrate the facility to resolve tissue-specific expression of eGFP and mCherrry fluorescent proteins for precise morphological and functional observations in vivo.
@article{2009-NATPHOT,author={Razansky<sup>#</sup>, D. and Distel, M. and Vinegoni, C. and Ma, R. and Perrimon, N. and Koster, R. W. and Ntziachristos<sup>#</sup>, V.},title={Multispectral opto-acoustic tomography of deep-seated fluorescent proteins in vivo},journal={Nature Photonics},alternatejournal={Nat. Photonics},year={2009},volume={3},number={7},pages={412-417},issn={1749-4885},doi={10.1038/nphoton.2009.98},}
10.1038/nphoton.2009.98
Phys. Med. Biol.
Imaging of mesoscopic-scale organisms using selective-plane optoacoustic tomography
Razansky#, D.,
Vinegoni, C.,
and Ntziachristos, V.
Mesoscopic-scale living organisms (i.e. 1 mm to 1 cm sized) remain largely inaccessible by current optical imaging methods due to intensive light scattering in tissues. Therefore, imaging of many important model organisms, such as insects, fishes, worms and similarly sized biological specimens, is currently limited to embryonic or other transparent stages of development. This makes it difficult to relate embryonic cellular and molecular mechanisms to consequences in organ function and animal behavior in more advanced stages and adults. Herein, we have developed a selective-plane illumination optoacoustic tomography technique for in vivo imaging of optically diffusive organisms and tissues. The method is capable of whole-body imaging at depths from the sub-millimeter up to centimeter range with a scalable spatial resolution in the order of magnitude of a few tenths of microns. In contrast to pure optical methods, the spatial resolution here is not determined nor limited by light diffusion; therefore, such performance cannot be achieved by any other optical imaging technology developed so far. The utility of the method is demonstrated on several whole-body models and small-animal extremities.
@article{2009-PMB,author={Razansky<sup>#</sup>, D. and Vinegoni, C. and Ntziachristos, V.},title={Imaging of mesoscopic-scale organisms using selective-plane optoacoustic tomography},journal={Physics in Medicine and Biology},alternatejournal={Phys. Med. Biol.},year={2009},volume={54},number={9},pages={2769-2777},issn={0031-9155},doi={10.1088/0031-9155/54/9/012},pmid={19369709}}
19369709
10.1088/0031-9155/54/9/012
Opt. Express
Diffractionless beam in free space with adiabatic changing refractive index in a single mode tapered slab waveguide
We propose a novel design to produce a free space diffractionless beam by adiabatically reducing the difference of the refractive index between the core and the cladding regions of a single mode tapered slab waveguide. To ensure only one propagating eigenmode in the adiabatic transition, the correlation of the waveguide core width and the refractive index is investigated. Under the adiabatic condition, we demonstrate that our waveguide can emit a diffractionless beam in free space up to 500 micrometers maintaining 72% of its original peak intensity. The proposed waveguide could find excellent applications for imaging purposes where an extended depth of field is required. (C) 2009 Optical Society of America
@article{2009-OE-2,author={Tsai<sup>#</sup>, C. C. and Vinegoni, C. and Weissleder, R.},title={Diffractionless beam in free space with adiabatic changing refractive index in a single mode tapered slab waveguide},journal={Optics Express},alternatejournal={Opt. Express},year={2009},volume={17},number={24},pages={21723-21731},issn={1094-4087},doi={10.1364/oe.17.021723},pmcid={PMC2805120},pmid={19997414}}
19997414
PMC2805120
10.1364/oe.17.021723
Opt. Express
High throughput transmission optical projection tomography using low cost graphics processing unit
Vinegoni#, C.,
Fexon, L.,
Fumene Feruglio, P.,
Pivovarov, M.,
Figueiredo, J. L.,
Nahrendorf, M.,
Pozzo, A.,
Sbarbati, A.,
and Weissleder, R.
We implement the use of a graphics processing unit (GPU) in order to achieve real time data processing for high-throughput transmission optical projection tomography imaging. By implementing the GPU we have obtained a 300 fold performance enhancement in comparison to a CPU workstation implementation. This enables to obtain on-the-fly reconstructions enabling for high throughput imaging. (C) 2009 Optical Society of America
@article{2009-OE-1,author={Vinegoni<sup>#</sup>, C. and Fexon, L. and Fumene Feruglio, P. and Pivovarov, M. and Figueiredo, J. L. and Nahrendorf, M. and Pozzo, A. and Sbarbati, A. and Weissleder, R.},title={High throughput transmission optical projection tomography using low cost graphics processing unit},journal={Optics Express},alternatejournal={Opt. Express},year={2009},volume={17},number={25},pages={22320-22332},issn={1094-4087},doi={10.1364/oe.17.022320},pmcid={PMC2805020},pmid={20052155}}
20052155
PMC2805020
10.1364/oe.17.022320
J Vis Exp
Born normalization for fluorescence optical projection tomography for whole heart imaging
Vinegoni#, C.,
Razansky, D.,
Figueiredo, J. L.,
Fexon, L.,
Pivoravov, M.,
Nahrendorf, M.,
Ntziachristos, V.,
and Weissleder, R.
Optical projection tomography is a three-dimensional imaging technique that has been recently introduced as an imaging tool primarily in developmental biology and gene expression studies. The technique renders biological sample optically transparent by first dehydrating them and then placing in a mixture of benzyl alcohol and benzyl benzoate in a 2:1 ratio (BABB or Murray s Clear solution). The technique renders biological samples optically transparent by first dehydrating them in graded ethanol solutions then placing them in a mixture of benzyl alcohol and benzyl benzoate in a 2:1 ratio (BABB or Murray s Clear solution) to clear. After the clearing process the scattering contribution in the sample can be greatly reduced and made almost negligible while the absorption contribution cannot be eliminated completely. When trying to reconstruct the fluorescence distribution within the sample under investigation, this contribution affects the reconstructions and leads, inevitably, to image artifacts and quantification errors.. While absorption could be reduced further with a permanence of weeks or months in the clearing media, this will lead to progressive loss of fluorescence and to an unrealistically long sample processing time. This is true when reconstructing both exogenous contrast agents (molecular contrast agents) as well as endogenous contrast (e.g. reconstructions of genetically expressed fluorescent proteins).
@article{2009-JOVE-1,author={Vinegoni<sup>#</sup>, C. and Razansky, D. and Figueiredo, J. L. and Fexon, L. and Pivoravov, M. and Nahrendorf, M. and Ntziachristos, V. and Weissleder, R.},title={Born normalization for fluorescence optical projection tomography for whole heart imaging},journal={Journal of Visualized Experiments},alternatejournal={J Vis Exp},year={2009},volume={2},number={28},pages={1389},doi={10.3791/1389},pmcid={PMC2794886},pmid={19578329}}
19578329
PMC2794886
10.3791/1389
Opt. Lett.
Normalized Born ratio for fluorescence optical projection tomography
Vinegoni#, C.,
Razansky, D.,
Figueiredo, J. L.,
Nahrendorf, M.,
Ntziachristos, V.,
and Weissleder, R.
We present a normalized Born approach for fluorescence optical projection tomography that takes into account tissue absorption properties. This approach can be particularly useful to study fluorochrome distribution within tissue. We use the algorithm to three-dimensionally reconstruct and characterize a fluorescein isothiocyanate containing absorptive phantom and an infarcted mouse heart previously injected with a fluorescent molecular probe. (C) 2009 Optical Society of America
@article{2009-OL-2,author={Vinegoni<sup>#</sup>, C. and Razansky, D. and Figueiredo, J. L. and Nahrendorf, M. and Ntziachristos, V. and Weissleder, R.},title={Normalized Born ratio for fluorescence optical projection tomography},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2009},volume={34},number={3},pages={319-321},issn={0146-9592},doi={10.1364/ol.34.000319},pmcid={PMC2771918},pmid={19183644}}
19183644
PMC2771918
10.1364/ol.34.000319
J Vis Exp
Mesoscopic Fluorescence Tomography for In-vivo Imaging of Developing Drosophila
Vinegoni#, C.,
Razansky, D.,
Pitsouli, C.,
Perrimon, N.,
Ntziachristos, V.,
and Weissleder, R.
Visualizing developing organ formation as well as progession and treatment of disease often heavily relies on the ability to optically interrogate molecular and functional changes in intact living organisms. Most existing optical imaging methods are inadequate for imaging at dimensions that lie between the penetration limits of modern optical microscopy (0.5-1mm) and the diffusion-imposed limits of optical macroscopy (>1cm) [1]. Thus, many important model organisms, e.g. insects, animal embryos or small animal extremities, remain inaccessible for in-vivo optical imaging. Although there is increasing interest towards the development of nanometer-resolution optical imaging methods, there have not been many successful efforts in improving the imaging penetration depth. The ability to perform in-vivo imaging beyond microscopy limits is in fact met with the difficulties associated with photon scattering present in tissues. Recent efforts to image entire embryos for example [2,3] require special chemical treatment of the specimen, to clear them from scattering, a procedure that makes them suitable only for post-mortem imaging. These methods however evidence the need for imaging larger specimens than the ones usually allowed by two-photon or confocal microscopy, especially in developmental biology and in drug discovery. We have developed a new optical imaging technique named Mesoscopic Fluorescence Tomography [4], which appropriate for non-invasive in-vivo imaging at dimensions of 1mm-5mm. The method exchanges resolution for penetration depth, but offers unprecedented tomographic imaging performance and it has been developed to add time as a new dimension in developmental biology observations (and possibly other areas of biological research) by imparting the ability to image the evolution of fluorescence-tagged responses over time. As such it can accelerate studies of morphological or functional dependencies on gene mutations or external stimuli, and can importantly, capture the complete picture of development or tissue function by allowing longitudinal time-lapse visualization of the same, developing organism. The technique utilizes a modified laboratory microscope and multi-projection illumination to collect data at 360-degree projections. It applies the Fermi simplification to Fokker-Plank solution of the photon transport equation, combined with geometrical optics principles in order to build a realistic inversion scheme suitable for mesoscopic range. This allows in-vivo whole-body visualization of non-transparent three-dimensional structures in samples up to several millimeters in size. We have demonstrated the in-vivo performance of the technique by imaging three-dimensional structures of developing Drosophila tissues in-vivo and by following the morphogenesis of the wings in the opaque Drosophila pupae in real time over six consecutive hours.
@article{2009-JOVE-2,author={Vinegoni<sup>#</sup>, C. and Razansky, D. and Pitsouli, C. and Perrimon, N. and Ntziachristos, V. and Weissleder, R.},title={Mesoscopic Fluorescence Tomography for In-vivo Imaging of Developing Drosophila},journal={Journal of Visualized Experiments},alternatejournal={J Vis Exp},year={2009},volume={2},number={30},pages={1510},doi={10.3791/1510},pmcid={PMC2736679},pmid={19696720}}
19696720
PMC2736679
10.3791/1510
Opt. Lett.
Transillumination fluorescence imaging in mice using biocompatible upconverting nanoparticles
Vinegoni#†, C.,
Razansky†, D.,
Hilderbrand, S. A.,
Shao, F. W.,
Ntziachristos, V.,
and Weissleder, R.
We report on a systematic study of upconverting fluorescence signal generation within turbid phantoms and real tissues. An accurate three-point Green’s function, describing the forward model of photon propagation, is established and experimentally validated. We further demonstrate, for the first time to our knowledge, autofluorescence-free transillumination imaging of mice that have received biocompatible upconverting nanoparticles. The method holds great promise for artifact-free whole-body visualization of optical molecular probes. (C) 2009 Optical Society of America
@article{2009-OL-1,author={Vinegoni<sup>#†</sup>, C. and Razansky<sup>†</sup>, D. and Hilderbrand, S. A. and Shao, F. W. and Ntziachristos, V. and Weissleder, R.},title={Transillumination fluorescence imaging in mice using biocompatible upconverting nanoparticles},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2009},volume={34},number={17},pages={2566-2568},issn={0146-9592},doi={10.1364/ol.34.002566},pmcid={PMC2749971},pmid={19724491}}
19724491
PMC2749971
10.1364/ol.34.002566
2008
J. Clin. Invest.
Real-time assessment of inflammation and treatment response in a mouse model of allergic airway inflammation
Cortez-Retamozo, V.,
Swirski, F. K.,
Waterman, P.,
Yuan, H.,
Figueiredo, J. L.,
Newton, A. P.,
Upadhyay, R.,
Vinegoni, C.,
Kohler, R.,
Blois, J.,
Smith, A.,
Nahrendorf, M.,
Josephson, L.,
Weissleder, R.,
and Pittet#, M. J.
Eosinophils are multifunctional leukocytes that degrade and remodel tissue extracellular matrix through production of proteolytic enzymes, release of proinflammatory factors to initiate and propagate inflammatory responses, and direct activation of mucus secretion and smooth muscle cell constriction. Thus, eosinophils are central effector cells during allergic airway inflammation and an important clinical therapeutic target. Here we describe the use of an injectable MMP-targeted optical sensor that specifically and quantitatively resolves eosinophil activity in the lungs of mice with experimental allergic airway inflammation. Through the use of real-time molecular imaging methods, we report the visualization of eosinophil. responses in vivo and at different scales. Eosinophil responses were seen at single-cell resolution in conducting airways using near-infrared fluorescence fiberoptic bronchoscopy, in lung parenchyma using intravital microscopy, and in the whole body using fluorescence-mediated molecular tomography. Using these real-time imaging methods, we confirmed the immunosuppressive effects of the glucocorticoid drug dexamethasone in the mouse model of allergic airway inflammation and identified a viridin-derived prodrug that potently inhibited the accumulation and enzyme activity of eosinophils in the lungs. The combination of sensitive enzyme-targeted sensors with noninvasive molecular imaging approaches permitted evaluation of airway inflammation severity and was used as a model to rapidly screen for new drug effects. Both fluorescence-mediated tomography and fiberoptic bronchoscopy techniques have the potential to be translated into the clinic.
@article{2008-JCI,author={Cortez-Retamozo, V. and Swirski, F. K. and Waterman, P. and Yuan, H. and Figueiredo, J. L. and Newton, A. P. and Upadhyay, R. and Vinegoni, C. and Kohler, R. and Blois, J. and Smith, A. and Nahrendorf, M. and Josephson, L. and Weissleder, R. and Pittet<sup>#</sup>, M. J.},title={Real-time assessment of inflammation and treatment response in a mouse model of allergic airway inflammation},journal={Journal of Clinical Investigation},alternatejournal={J. Clin. Invest.},year={2008},volume={118},number={12},pages={4058-4066},issn={0021-9738},doi={10.1172/jci36335},pmid={18923605}}
18923605
10.1172/jci36335
Circulation
Real-Time Catheter Molecular Sensing of Inflammation in Proteolytically Active Atherosclerosis
Jaffer, F. A.,
Vinegoni, C.,
John, M. C.,
Aikawa, E.,
Gold, H. K.,
Finn, A. V.,
Ntziachristos, V.,
Libby, P.,
and Weissleder#, R.
Background-To enable intravascular detection of inflammation in atherosclerosis, we developed a near-infrared fluorescence ( NIRF) catheter-based strategy to sense cysteine protease activity during vascular catheterization. Methods and Results-The NIRF catheter design was based on a clinical coronary artery guidewire. In phantom studies of NIRF plaques, blood produced only a mild (<30%) attenuation of the fluorescence signal compared with saline, affirming the favorable optical properties of the NIR window. Catheter evaluation in vivo used atherosclerotic rabbits (n=11). Rabbits received an injection of a cysteine protease-activatable NIRF imaging agent (Prosense750; excitation/emission, 750/770 nm) or saline. Catheter pullbacks through the blood-filled iliac artery detected NIRF signals 24 hours after injection of the probe. In the protease agent group, the in vivo peak plaque target-to-background ratio was 558% greater than controls (6.8 +/- 1.9 versus 1.3 +/- 0.3, mean +/- SEM; P<0.05). Ex vivo fluorescence reflectance imaging corroborated these results (target-to-background ratio, 10.3 +/- 1.8 for agent versus 1.8 +/- 0.3 for saline group; P<0.01). In the protease group only, saline flush-modulated NIRF signal profiles further distinguished atheromata from normal segments in vivo (P<0.01). Good correlation between the in vivo and ex vivo plaque target-to-background ratio was present (r=0.82, P<0.01). Histopathological analyses demonstrated strong NIRF signal in plaques only from the protease agent group. NIRF signals colocalized with immunoreactive macrophages and the cysteine protease cathepsin B. Conclusions-An intravascular fluorescence catheter can detect cysteine protease activity in vessels the size of human coronary arteries in real time with an activatable NIRF agent. This strategy could aid in the detection of inflammation and high-risk plaques in small arteries. (Circulation. 2008; 118: 1802-1809.)
@article{2008-CRCLT,author={Jaffer, F. A. and Vinegoni, C. and John, M. C. and Aikawa, E. and Gold, H. K. and Finn, A. V. and Ntziachristos, V. and Libby, P. and Weissleder<sup>#</sup>, R.},title={Real-Time Catheter Molecular Sensing of Inflammation in Proteolytically Active Atherosclerosis},journal={Circulation},alternatejournal={Circulation},year={2008},volume={118},number={18},pages={1802-1809},issn={0009-7322},doi={10.1161/circulationaha.108.785881},pmcid={PMC2729441},pmid={18852366}}
18852366
PMC2729441
10.1161/circulationaha.108.785881
Opt. Lett.
Polarization-sensitive optoacoustic tomography of optically diffuse tissues
Razansky#, D.,
Vinegoni, C.,
and Ntziachristos, V.
Polarization is indicative of material anisotropy, a property that reveals structural orientation information of molecules inside the material. Herein we investigate whether polarization can be detected optoacoustically in scattering and absorbing tissues. Using a laboratory prototype of polarization-sensitive optoacoustic tomography, we demonstrate high-resolution reconstructions of dichroism. contrast deep in optically diffusive tissue-mimicking phantoms. The technique is expected to enable highly accurate imaging of polarization contrast in tissues, far beyond the current capabilities of pure optical polarization-imaging approaches. (C) 2008 Optical Society of America
@article{2008-OL,author={Razansky<sup>#</sup>, D. and Vinegoni, C. and Ntziachristos, V.},title={Polarization-sensitive optoacoustic tomography of optically diffuse tissues},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2008},volume={33},number={20},pages={2308-2310},issn={0146-9592},doi={10.1364/ol.33.002308},pmid={18923605}}
18923605
10.1364/ol.33.002308
Nat. Methods
In vivo imaging of Drosophila melanogaster pupae with mesoscopic fluorescence tomography
Vinegoni#†, C.,
Pitsouli†, C.,
Razansky†, D.,
Perrimon, N.,
and Ntziachristos, V.
We report a technique for fluorescence tomography that operates beyond the penetration limits of tissue-sectioning fluorescence microscopy. The method uses multi-projection illumination and photon transport description in opaque tissues. We demonstrate whole-body three-dimensional visualization of the morphogenesis of GFP-expressing salivary glands and wing imaginal discs in living Drosophila melanogaster pupae in vivo and over time.
@article{2008-NM,author={Vinegoni<sup>#†</sup>, C. and Pitsouli<sup>†</sup>, C. and Razansky<sup>†</sup>, D. and Perrimon, N. and Ntziachristos, V.},title={In vivo imaging of Drosophila melanogaster pupae with mesoscopic fluorescence tomography},journal={Nature Methods},alternatejournal={Nat. Methods},year={2008},volume={5},number={1},pages={45-47},issn={1548-7091},doi={10.1038/nmeth1149},pmid={18066071}}
18066071
10.1038/nmeth1149
2007
Opt. Lett.
Multispectral photoacoustic imaging of fluorochromes in small animals
Razansky, D.,
Vinegoni, C.,
and Ntziachristos#, V.
Fluorochromes have become essential reporter molecules in biological research. We show that the depth-resolved distribution of fluorochromes in small animals can be imaged with 25 fmol sensitivity and 150 Am spatial resolution by means of multispectral photoacoustic imaging. The major advantage of the multispectral approach is the sensitive differentiation of chromophores and fluorochromes of interest based on self-reference measurements, as evidenced in this study by resolving a commonly used fluorochrome (Alexa Fluor 750) in mouse. The suggested method is well suited for enhancing visualization of functional and molecular information in vivo and longitudinally. (c) 2007 Optical Society of America.
@article{2007-OL,author={Razansky, D. and Vinegoni, C. and Ntziachristos<sup>#</sup>, V.},title={Multispectral photoacoustic imaging of fluorochromes in small animals},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2007},volume={32},number={19},pages={2891-2893},issn={0146-9592},doi={10.1364/ol.32.002891},pmid={17909608}}
17909608
10.1364/ol.32.002891
Microsc. Res. Tech.
Imaging cellular responses to mechanical stimuli within three-dimensional tissue constructs
Tan, W.,
Vinegoni, C.,
Norman, J. J.,
Desai, T. A.,
and Boppart#, S. A.
The cellular response to environmental cues is complex, involving both structural and functional changes within the cell. Our understanding of this response is facilitated by microscopy techniques, but has been limited by our ability to image cell structure and function deep in highly-scattering tissues or 3D constructs. A novel multimodal microscopy technique that combines coherent and incoherent imaging for simultaneous visualization of structural and functional properties of cells and engineered tissues is demonstrated. This microscopic technique allows for the simultaneous acquisition of optical coherence microscopy and multiphoton microscopy data with particular emphasis for applications in cell biology and tissue engineering. The capability of this technique is shown using representative 3D cell and tissue engineering cultures consisting of primary fibroblasts from transgenic green fluorescent protein (GFP) mice and GFP-vinculin transfected fibroblasts. Imaging is performed following static and dynamic mechanically-stimulating culture conditions. The microscopy technique presented here reveals unique complementary data on the structure and function of cells and their adhesions and interactions with the surrounding microenvironment.
@article{2007-MRT,author={Tan, W. and Vinegoni, C. and Norman, J. J. and Desai, T. A. and Boppart<sup>#</sup>, S. A.},title={Imaging cellular responses to mechanical stimuli within three-dimensional tissue constructs},journal={Microscopy Research and Technique},alternatejournal={Microsc. Res. Tech.},year={2007},volume={70},number={4},pages={361-371},issn={1059-910X},doi={10.1002/jemt.20420},pmid={17262787}}
To achieve high-spectral-resolution multiplex coherent anti-Stokes Raman scattering (CARS), one typically uses a narrowband pump pulse and a broadband Stokes pulse. This is to ensure a correspondence between anti-Stokes and vibrational frequencies. We obtain high-resolution CARS spectra of isopropanol, using a broadband chirped pump pulse and a broadband Stokes pulse, by detecting the anti-Stokes pulse with spectral interferometry. With the temporally resolved anti-Stokes signal, we can remove the chirp of the anti-Stokes pulse and restore high spectral resolution while also rejecting nonresonant scattering. (c) 2006 Optical Society of America.
@article{2006-OL-2,author={Jones, G. W. and Marks, D. L. and Vinegoni, C. and Boppart<sup>#</sup>, S. A.},title={High-spectral-resolution coherent anti-Stokes Raman scattering with interferometric ally detected broadband chirped pulses},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2006},volume={31},number={10},pages={1543-1545},issn={0146-9592},doi={10.1364/ol.31.001543},pmid={16642166}}
16642166
10.1364/ol.31.001543
Appl. Phys. Lett.
Integrated structural and functional optical imaging combining spectral-domain optical coherence and multiphoton microscopy
Vinegoni, C.,
Ralston, T.,
Tan, W.,
Luo, W.,
Marks, D. L.,
and Boppart#, S. A.
An integrated microscope that combines different optical techniques for simultaneous imaging is demonstrated. The microscope enables spectral-domain optical coherence microscopy based on optical backscatter, and multiphoton microscopy for the detection of two-photon fluorescence and second harmonic generation signals. The unique configuration of this integrated microscope allows for the simultaneous acquisition of both anatomical (structural) and functional imaging information with particular emphasis for applications in the fields of tissue engineering and cell biology. In addition, the contemporary analysis of the spectroscopic features can enhance contrast by differentiating among different tissue components. (c) 2006 American Institute of Physics.
@article{2006-APL,author={Vinegoni, C. and Ralston, T. and Tan, W. and Luo, W. and Marks, D. L. and Boppart<sup>#</sup>, S. A.},title={Integrated structural and functional optical imaging combining spectral-domain optical coherence and multiphoton microscopy},journal={Applied Physics Letters},alternatejournal={Appl. Phys. Lett.},year={2006},volume={88},number={5},pages={3},issn={0003-6951},doi={10.1063/1.2171477},}
The spectroscopic content within optical coherence tomography (OCT) data can provide a wealth of information. Spectroscopic OCT methods are frequently limited by time-frequency trade-offs that limit high spectral and spatial resolution simultaneously. We present spectroscopic spectral-domain optical coherence microscopy performed with a multimodality microscope. Restricting the spatial extent of the signal by using high-numerical-aperture optics makes high-resolution spectroscopic information accessible, facilitated with spectral-domain detection. Simultaneous acquisition of multiphoton microscopy images is used to validate tissue structure and localization of nuclei within individual cells. (c) 2006 Optical Society of America.
@article{2006-OL-1,author={Xu, C. Y. and Vinegoni, C. and Ralston, T. S. and Luo, W. and Tan, W. and Boppart<sup>#</sup>, S. A.},title={Spectroscopic spectral-domain optical coherence microscopy},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2006},volume={31},number={8},pages={1079-1081},issn={0146-9592},doi={10.1364/ol.31.001079},pmid={16625909}}
Molecular contrast in optical coherence tomography (OCT) is demonstrated by use of coherent anti-Stokes Raman scattering (CARS) for molecular sensitivity. Femtosecond laser pulses are focused into a sample by use of a low-numerical-aperture lens to generate CARS photons, and the backreflected CARS signal is interferometrically measured. With the chemical selectivity provided by CARS and the advanced imaging capabilities of OCT, this technique may be useful for molecular contrast imaging in biological tissues. CARS can be generated and interferometrically measured over at least 600 mum of the depth of field of a low-numerical-aperture objective. (C) 2005 Optical Society of America.
@article{2005-OL,author={Bredfeldt, J. S. and Vinegoni, C. and Marks, D. L. and Boppart<sup>#</sup>, S. A.},title={Molecularly sensitive optical coherence tomography},journal={Optics Letters},alternatejournal={Opt. Lett.},year={2005},volume={30},number={5},pages={495-497},issn={0146-9592},doi={10.1364/ol.30.000495},pmid={15789714}}
For decades, Raman scattering has been a useful tool for probing the molecular contents of gases, liquids and biological media.1 Its strength is that it can identify molecules non-invasively based on their resonant vibrational frequencies. The technique uses optical frequency radiation, which is relatively easy to create, manipulate and detect.
Unfortunately, the Raman effect is rather weak: Typically only one in a million photons is converted to the detected Raman radiation, so it is too slow to be used as a clinical diagnostic tool or to monitor fast biological processes. Using pulsed laser illumination, Coherent Anti-Stokes Raman Scattering (CARS) can stimulate the production of many
more Raman photons by simultaneously exciting the molecular vibrations and detecting the emitted stimulated Raman scattering.
@article{2005-OPN,author={McAlpine, C. S. and Marks, D. and Vinegoni, C. and Bredfeldt, J. and Boppart<sup>#</sup>, S.A.},title={Nonlinear Interferometric Vibrational Imaging. efficient detection of Coherent Anti-Stokes Raman Scattering},journal={Optics & Photonics News},alternatejournal={Opt Photonics News
},year={2005},volume={16},pages={23},doi={10.1364/OPN.16.12.000023},}
10.1364/OPN.16.12.000023
2004
Appl. Phys. Lett.
Interferometric differentiation between resonant coherent anti-Stokes Raman scattering and nonresonant four-wave-mixing processes
Marks, D. L.,
Vinegoni, C.,
Bredfeldt, J. S.,
and Boppart#, S. A.
A major impediment to the use of coherent anti-Stokes Raman scattering (CARS) to identify biological molecules is that the illumination levels required to produce a measurable signal often also produce significant nonresonant background from the medium, especially from water. We present a method of nonlinear interferometry to differentiate between which components of the anti-Stokes signal are resonant and nonresonant. The technique takes advantage of the persistence of intermediate states involved in the resonant process. This method is applicable to most existing pulsed CARS illumination methods and provides for identification of resonant CARS. We demonstrate the method by examining the signals produced by acetone, which exhibits resonance, and water, which does not. (C) 2004 American Institute of Physics.
@article{2004-APL,author={Marks, D. L. and Vinegoni, C. and Bredfeldt, J. S. and Boppart<sup>#</sup>, S. A.},title={Interferometric differentiation between resonant coherent anti-Stokes Raman scattering and nonresonant four-wave-mixing processes},journal={Applied Physics Letters},alternatejournal={Appl. Phys. Lett.},year={2004},volume={85},number={23},pages={5787-5789},issn={0003-6951},doi={10.1063/1.1829162},}
10.1063/1.1829162
Opt. Express
Nonlinear optical contrast enhancement for optical coherence tomography
Vinegoni, C.,
Bredfeldt, J. S.,
Marks, D. L.,
and Boppart#, S. A.
We present a new interferometric technique for measuring Coherent Anti-Stokes Raman Scattering ( CARS) and Second Harmonic Generation (SHG) signals. Heterodyne detection is employed to increase the sensitivity in both CARS and SHG signal detection, which can also be extended to different coherent processes. The exploitation of the mentioned optical nonlinearities for molecular contrast enhancement in Optical Coherence Tomography (OCT) is presented. (C) 2004 Optical Society of America.
@article{2004-OE,author={Vinegoni, C. and Bredfeldt, J. S. and Marks, D. L. and Boppart<sup>#</sup>, S. A.},title={Nonlinear optical contrast enhancement for optical coherence tomography},journal={Optics Express},alternatejournal={Opt. Express},year={2004},volume={12},number={2},pages={331-341},issn={1094-4087},doi={10.1364/opex.12.000331},pmid={19471542}}
19471542
10.1364/opex.12.000331
J. Lightwave Tech.
The statistics of polarization-dependent loss in a recirculating loop
Vinegoni#, C.,
Karlsson, M.,
Petersson, M.,
and Sunnerud, H.
In this paper, we study the statistical distribution of the accumulated polarization-dependent loss (PDL) in a recirculating loop. The distribution is studied both via numerical simulations and with analytical theory and very good agreement is found between the two. In addition, we have experimentally obtained the probability density distribution for the PDL and, even in this case, we find good agreement with the predicted one. The mean accumulated PDL is found to grow linearly with the number of circulations N in contraposition to a straight-line system. Moreover, the statistical distribution tends to become uniformly distributed as N is increasing. Finally, the statistics of the PDL for a recirculating loop, when considering small values of PDL, is found to be equal to the statistics of the differential group delay for recirculating loops.
@article{2004-JLT,author={Vinegoni<sup>#</sup>, C. and Karlsson, M. and Petersson, M. and Sunnerud, H.},title={The statistics of polarization-dependent loss in a recirculating loop},journal={Journal of Lightwave Technology},alternatejournal={J. Lightwave Tech.},year={2004},volume={22},number={4},pages={968-976},issn={0733-8724},doi={10.1109/jlt.2004.824861},}
10.1109/jlt.2004.824861
2003
IEEE P.T.L.
Statistics of PMD in recirculating loops
Petersson#, M.,
Vinegoni, C.,
Sunnerud, H.,
and Karlsson, M.
The statistical distribution of polarization-mode dispersion in a recirculating loop is investigated. Numerical simulations are performed in both Jones and Stokes space and are verified by experiments. The probability distribution of the differential group delay (DGD) is obtained numerically, theoretically, and experimentally. As the number of circulations increase, the probability density function of the DGD approaches a uniform distribution.
@article{2003-PTL-1,author={Petersson<sup>#</sup>, M. and Vinegoni, C. and Sunnerud, H. and Karlsson, M.},title={Statistics of PMD in recirculating loops},journal={Ieee Photonics Technology Letters},alternatejournal={IEEE P.T.L.},year={2003},volume={15},number={11},pages={1543-1545},issn={1041-1135},doi={10.1109/lpt.2003.818681},}
10.1109/lpt.2003.818681
IEEE P.T.L.
Distributed measurements of chromatic dispersion and nonlinear coefficient in low-PMD dispersion-shifted fibers
Vinegoni, C.,
Chen#, H. X.,
Leblanc, M.,
Schinn, G. W.,
Wegmuller, M.,
and Gisin, N.
We report on the investigation of distributed chromatic dispersion (CD) and distributed nonlinear coefficient (NLC) measurements based on phase mismatched four-wave mixing in dispersion-shifted fibers (DSFs). Experimental results of the distributed CD maps for low Polarization-mode dispersion (PMD) DSF fibers are discussed. We also report how nonnegligible values of PMD can adversely affect the distributed CD measurements. A new method to measure the distributed NLC map in low-PMD DSF fibers is also proposed and demonstrated experimentally.
@article{2003-PTL-2,author={Vinegoni, C. and Chen<sup>#</sup>, H. X. and Leblanc, M. and Schinn, G. W. and Wegmuller, M. and Gisin, N.},title={Distributed measurements of chromatic dispersion and nonlinear coefficient in low-PMD dispersion-shifted fibers},journal={Ieee Photonics Technology Letters},alternatejournal={IEEE P.T.L.},year={2003},volume={15},number={5},pages={739-741},issn={1041-1135},doi={10.1109/lpt.2003.810249},}
10.1109/lpt.2003.810249
2002
IEEE P.T.L.
Emulator of first- and second-order polarization-mode dispersion
Wegmuller, M.,
Demma, S.,
Vinegoni, C.,
and Gisin#, N.
Contrary to approaches which try to mimic a standard fiber as closely as possible, the emulator presented here gives constant (but user adjustable) values for differential group delay (DGD) and ratio of first- to second-order polarization-mode dispersion (PMD). Once it is set, the ratio is conserved while the DGD can be easily varied within a range of 0-300 ps. This allows to investigate the low-probability events of large DGD and second-order PMD important for system outage.
@article{2002-PTL,author={Wegmuller, M. and Demma, S. and Vinegoni, C. and Gisin<sup>#</sup>, N.},title={Emulator of first- and second-order polarization-mode dispersion},journal={Ieee Photonics Technology Letters},alternatejournal={IEEE P.T.L.},year={2002},volume={14},number={5},pages={630-632},issn={1041-1135},doi={10.1109/68.998707},}
10.1109/68.998707
2001
IEEE P.T.L.
Measurements of the nonlinear coefficient of standard SMF, DSF, and DCF fibers using a self-aligned interferometer and a Faraday mirror
Using a method based on the detection of the Kerr phase shift by a self-aligned interferometer, we present measurements of the nonlinear coefficient n(2)/A(eff) for standard single-mode fiber (SMF), dispersion-shifted fibers, and dispersion compensating fibers. The presence of a Faraday, mirror in the interferometer makes the setup very robust, and different test fibers can be measured without any further readjustments. Interlaboratory comparisons show that the values found with our method are in good agreement with the other ones. Further, analysis of a SMF fiber with large chromatic dispersion shows a good reproducibility of the n(2)/A(eff) measurements as a function of fiber length.
@article{2001-PTL-1,author={Vinegoni<sup>#</sup>, C. and Wegmuller, M. and Gisin, N.},title={Measurements of the nonlinear coefficient of standard SMF, DSF, and DCF fibers using a self-aligned interferometer and a Faraday mirror},journal={Ieee Photonics Technology Letters},alternatejournal={IEEE P.T.L.},year={2001},volume={13},number={12},pages={1337-1339},issn={1041-1135},doi={10.1109/68.969900},}
10.1109/68.969900
IEEE P.T.L.
Analysis of the polarization evolution in a ribbon cable using high-resolution coherent OFDR
Wegmuller, M.,
Legre, M.,
Oberson, P.,
Guinnard, O.,
Guinnard, L.,
Vinegoni, C.,
and Gisin#, N.
Exploiting the inherent polarization dependence and good spatial resolution of optical frequency domain reflectometry (OFDR), the beatlength in a ribbon fiber can be straightforwardly measured. The results clearly show the different amount of polarization ordering for inner and outer ribbon fibers due to the stress-induced birefringence from the common outer coating.
@article{2001-PTL-2,author={Wegmuller, M. and Legre, M. and Oberson, P. and Guinnard, O. and Guinnard, L. and Vinegoni, C. and Gisin<sup>#</sup>, N.},title={Analysis of the polarization evolution in a ribbon cable using high-resolution coherent OFDR},journal={Ieee Photonics Technology Letters},alternatejournal={IEEE P.T.L.},year={2001},volume={13},number={2},pages={145-147},issn={1041-1135},doi={10.1109/68.910516},}
10.1109/68.910516
2000
Phys. Rev. B
Optical absorption and photoluminescence properties of alpha-Si1-xNx : H films deposited by plasma-enhanced CVD
Optical absorption and photoluminescence (PL) measurements were performed on a series of hydrogenated silicon-nitrogen thin films deposited by plasma enhanced chemical vapor deposition covering a wide compositional range with the aim to find how absorption and emission processes are correlated. Our analysis indicates that the observed broadening of the PL spectra is linked with the existence of band tail states due to topological disorder, as confirmed by the spectral dependence of the PL decay. Moreover, nitrogen alloying reduces the temperature quenching of luminescence and increases the radiative recombination rate. Such phenomena are ascribed to the formation of wide tails in the density of states of N-rich samples and, in turn, to a lower carrier diffusion.
@article{2000-PRB,author={Giorgis<sup>#</sup>, F. and Vinegoni, C. and Pavesi, L.},title={Optical absorption and photoluminescence properties of alpha-Si1-xNx : H films deposited by plasma-enhanced CVD},journal={Physical Review B},alternatejournal={Phys. Rev. B},year={2000},volume={61},number={7},pages={4693-4698},issn={2469-9950},doi={10.1103/PhysRevB.61.4693},}
10.1103/PhysRevB.61.4693
Surf. Coat. Technol.
Morphological and optical characterization of GaN prepared by pulsed laser deposition
Vinegoni#, C.,
Cazzanelli, M.,
Trivelli, A.,
Mariotto, G.,
Castro, J.,
Lunney, J. G.,
and Levy, J.
GaN films were grown by pulsed laser deposition (PLD on different crystalline substrates using a KrF excimer laser to ablate a hexagonal phase GaN target in a reactive atmosphere of ammonia. Films with small homogeneously distributed granular structures over the entire sample surface were obtained, The microstructure and surface morphology of the deposited layers were characterized by X-ray diffraction (XRD, atomic force microscopy (AFM) and Raman spectroscopy (RS). XRD reveals that the structure of the GaN layer is predominantly wurtzite. AFM images reveal that all the deposited layers have a relatively smooth surface, while RS confirmed the predominant presence of hexagonal GaN with a high polycrystalline character. Analysis of the results obtained for samples grown under different conditions, such as the substrate temperatures in the growth chamber as well as different substrates used, helps to define better the experimental conditions of the growth process of PLD-GaN films. (C) 2000 Elsevier Science S.A. All rights reserved.
@article{2000-SCT,author={Vinegoni<sup>#</sup>, C. and Cazzanelli, M. and Trivelli, A. and Mariotto, G. and Castro, J. and Lunney, J. G. and Levy, J.},title={Morphological and optical characterization of GaN prepared by pulsed laser deposition},journal={Surface & Coatings Technology},alternatejournal={Surf. Coat. Technol.},year={2000},volume={124},number={2-3},pages={272-277},issn={0257-8972},doi={10.1016/s0257-8972(99)00657-x},}
10.1016/s0257-8972(99)00657-x
Electron. Lett.
Determination of nonlinear coefficient n(2)/A(eff) using self-aligned interferometer and Faraday mirror
A method for measuring the nonlinear coefficient n(2)/A(eff) in telecom fibres at 1550nm is presented. The method is based on determining the Kerr phase shift detected by a self-aligned interferometer incorporating a Faraday mirror. This makes the setup very robust, and different test fibres can be measured without the need for any further readjustments.
@article{2000-EL,author={Vinegoni<sup>#</sup>, C. and Wegmuller, M. and Gisin, N.},title={Determination of nonlinear coefficient n(2)/A(eff) using self-aligned interferometer and Faraday mirror},journal={Electronics Letters},alternatejournal={Electron. Lett.},year={2000},volume={36},number={10},pages={886-888},issn={0013-5194},doi={10.1049/el:20000668},}
10.1049/el:20000668
Opt. Commun.
All optical switching in a highly birefringent and a standard telecom fiber using a Faraday mirror stabilization scheme
Vinegoni#, C.,
Wegmuller, M.,
Huttner, B.,
and Gisin, N.
All-optical switching at 1.5 mu m based on induced nonlinear polarization rotation is demonstrated in both a polarization maintaining and a standard telecom fiber. Excellent switching stability is obtained in both cases by removing any detrimental temperature or pressure induced changes of the output polarization state with a Farady mirror stabilization scheme. (C) 2000 Elsevier Science B.V. All rights reserved.
@article{2000-OC,author={Vinegoni<sup>#</sup>, C. and Wegmuller, M. and Huttner, B. and Gisin, N.},title={All optical switching in a highly birefringent and a standard telecom fiber using a Faraday mirror stabilization scheme},journal={Optics Communications},alternatejournal={Opt. Commun.},year={2000},volume={182},number={4-6},pages={335-341},issn={0030-4018},doi={10.1016/s0030-4018(00)00845-2},}
10.1016/s0030-4018(00)00845-2
J. Opt. A
Measurement of nonlinear polarization rotation in a highly birefringent optical fibre using a Faraday mirror
Vinegoni#, C.,
Wegmuller, M.,
Huttner, B.,
and Gisin, N.
We present both a theoretical and experimental analysis of nonlinear polarization rotation in an optical fibre. Starting from the coupled nonlinear Schrodinger equations an analytical solution for the evolution of the state of polarization, valid for fibres with large linear birefringence and quasi cw input light with arbitrary polarization, is given. It allows us to model straightforwardly go-and-return paths as in interferometers with standard or Faraday mirrors. In the experiment all the fluctuations in the linear birefringence, including temperature- and pressure-induced ones, are successfully removed in a passive way by using a double pass of the fibre under test with a Faraday mirror at the end of the fibre. This allows us to use long fibres and relatively low input powers. The match between the experimental data and our model is excellent, except at higher intensities where deviations due to modulation instability start to appear.
@article{2000-JOA,author={Vinegoni<sup>#</sup>, C. and Wegmuller, M. and Huttner, B. and Gisin, N.},title={Measurement of nonlinear polarization rotation in a highly birefringent optical fibre using a Faraday mirror},journal={Journal of Optics a-Pure and Applied Optics},alternatejournal={J. Opt. A},year={2000},volume={2},number={4},pages={314-318},issn={1464-4258},doi={10.1088/1464-4258/2/4/313},}
10.1088/1464-4258/2/4/313
J. Lightwave Tech.
Distributed gain measurements in Er-doped fibers with high resolution and accuracy using an optical frequency domain reflectometer
Wegmuller#, M.,
Oberson, P.,
Guinnard, O.,
Huttner, B.,
Guinnard, L.,
Vinegoni, C.,
and Gisin, N.
For critical Erbium-doped fiber amplifier (EDFA) design, e.g., gain tilt optimization in WDM booster amplifiers, knowledge of the gain distribution within the active fiber can present a valuable information. Among the different techniques to evaluate the distributed gain in active fibers, the technique of optical frequency domain reflectometry seems most promising as it is a nondestructive measurement method well matched to the task due to its dynamic range, resolution, and range, Moreover, background light from ASE or residual pump light is strongly rejected due to the coherent detection scheme employed. Using different Erbium-doped fibers with strongly varying doping levels and confinements, we demonstrate the excellent accuracy and reproducibility of the technique.
@article{2000-JLT,author={Wegmuller<sup>#</sup>, M. and Oberson, P. and Guinnard, O. and Huttner, B. and Guinnard, L. and Vinegoni, C. and Gisin, N.},title={Distributed gain measurements in Er-doped fibers with high resolution and accuracy using an optical frequency domain reflectometer},journal={Journal of Lightwave Technology},alternatejournal={J. Lightwave Tech.},year={2000},volume={18},number={12},pages={2127-2132},issn={0733-8724},doi={10.1109/50.908823},}
10.1109/50.908823
1999
J. Solid State Chem.
Low-temperature polymorphism in tungsten trioxide powders and its dependence on mechanical treatments
Cazzanelli, E.,
Vinegoni, C.,
Mariotto#, G.,
Kuzmin, A.,
and Purans, J.
The polymorphism of WO, powder samples, resulting from mild mechanical treatments and from temperature changes between 30 K and room temperature, has been investigated by using Raman spectroscopy and X-ray diffraction. A transition from the monoclinic (I) gamma-phase to the triclinic delta-phase after moderate mechanical treatments has been observed for untreated powder, just what happens when the same samples are rapidly cooled to low-temperature. Evidences of the low temperature monoclinic (II) polar epsilon-phase have been found at room temperature in samples after a stronger milling treatment. The sequence of the low-temperature phase transitions appears to be strongly dependent on the mechanical history of the powders. A new low-temperature N-phase has been observed below about 200 K in different samples: it is the main phase in commercial untreated powders, having the monoclinic (I) gamma-phase at room temperature, but constitutes only a small fraction in moderately treated powders, having the triclinic delta-phase at room temperature. (C) 1999 Academic Press.
@article{1999-JSSC,author={Cazzanelli, E. and Vinegoni, C. and Mariotto<sup>#</sup>, G. and Kuzmin, A. and Purans, J.},title={Low-temperature polymorphism in tungsten trioxide powders and its dependence on mechanical treatments},journal={Journal of Solid State Chemistry},alternatejournal={J. Solid State Chem.},year={1999},volume={143},number={1},pages={24-32},issn={0022-4596},doi={10.1006/jssc.1998.8061},}
10.1006/jssc.1998.8061
Mater. Sci. B
Luminescent properties of GaN thin films prepared by pulsed laser deposition
Cazzanelli, M.,
Vinegoni, C.,
Cole, D.,
Lunney, J. G.,
Middleton, P. G.,
Trager-Cowan, C.,
O’Donnell, K. P.,
and Pavesi#, L.
Materials Science and Engineering B-Solid State Materials for Advanced Technology 1999
The luminescent properties of GaN thin films grown by pulsed laser deposition have been studied to understand the nature of the luminescent centres and the recombination dynamics. The films were grown on heated sapphire substrates using KrF excimer laser ablation of GaN in a reactive atmosphere of nitrogen. At low temperature the continuous wave (CW) blue luminescence of the samples grown shows two sharp lines attributed to excitonic recombination localized at extended defects. An analysis of the temperature dependence of photoluminescence (PL) lifetimes assesses the relative contributions of radiative and non-radiative recombination in the centres responsible for these emissions. The measurement of room temperature nanosecond radiative lifetimes for these lines supports the excitonic attribution of the luminescence. (C) 1999 Elsevier Science S.A. All rights reserved.
@article{1999-MSEB,author={Cazzanelli, M. and Vinegoni, C. and Cole, D. and Lunney, J. G. and Middleton, P. G. and Trager-Cowan, C. and O’Donnell, K. P. and Pavesi<sup>#</sup>, L.},title={Luminescent properties of GaN thin films prepared by pulsed laser deposition},journal={Materials Science and Engineering B-Solid State Materials for Advanced Technology},alternatejournal={Mater. Sci. B},year={1999},volume={59},number={1-3},pages={137-140},issn={0921-5107},doi={10.1016/s0921-5107(98)00333-x},}
10.1016/s0921-5107(98)00333-x
J. Appl. Phys.
Temperature dependence of the photoluminescence of all-porous-silicon optical microcavities
Photoluminescence measurements in all-porous-silicon optical microcavities (PSM) are reported over a wide temperature range. Both continuous wave and time resolved measurements have been performed. The microcavity is formed by an all porous silicon Fabry-Perot filter made by two distributed Bragg reflectors separated by a lambda-thick PS cavity layer. The luminescence properties of PSM are changed with respect to those of PS: a temperature independent narrowing in the emission line shape, a different temperature dependence of the emission intensity, and a fractional shortening of the luminescence decay time over the 50-300 K temperature interval are achieved. The PSM luminescence properties are explained by the spatial redistribution of the spontaneous emission, by an effective refractive index probed by the photon mode confined in the cavity layer and by the coupling between the singlet exciton state and the photon mode confined in the cavity layer. The saturation of the absorption of the distributed Bragg reflector is also addressed. (C) 1999 American Institute of Physics. [S0021-8979(99)04503-X].
@article{1999-JAP,author={Cazzanelli, M. and Vinegoni, C. and Pavesi<sup>#</sup>, L.},title={Temperature dependence of the photoluminescence of all-porous-silicon optical microcavities},journal={Journal of Applied Physics},alternatejournal={J. Appl. Phys.},year={1999},volume={85},number={3},pages={1760-1764},issn={0021-8979},doi={10.1063/1.369320},}
10.1063/1.369320
Ionics
Color Centres and Polymorphism in Pure WO(3) and Mixed (1-x)WO(3-y)center dot xReO(2) Powders
Cazzanelli#, E.,
Mariotto, G.,
Vinegoni, C.,
Kuzmin, A.,
and Purans, J.
The relationship between structural transformations and colour centres creation is discussed for deeply coloured hydrogen tungsten bronzes and for pure WO(3) powders, acquiring less intense colour after mechanical treatments of variable duration. A comparative study on coloration is made also for mixed compounds (1-x)WO(3-y)center dot xReO(2), where an evidence of a resonance effect for a particular Raman band at 970 cm(-1), attributed to the color centres, is observed. Besides, it is found that even moderate milling treatments result in a quite different structural evolution of tungsten trioxide upon cooling.
@article{1999-INCS,author={Cazzanelli<sup>#</sup>, E. and Mariotto, G. and Vinegoni, C. and Kuzmin, A. and Purans, J.},title={Color Centres and Polymorphism in Pure WO(3) and Mixed (1-x)WO(3-y)center dot xReO(2) Powders},journal={Ionics},alternatejournal={Ionics},year={1999},volume={5},number={5-6},pages={335-344},issn={0947-7047},doi={10.1007/bf02375997},}
10.1007/bf02375997
Solid State Ion.
Raman study of the phase transitions sequence in pure WO3 at high temperature and in HxWO3 with variable hydrogen content
Cazzanelli#, E.,
Vinegoni, C.,
Mariotto, G.,
Kuzmin, A.,
and Purans, J.
An extensive investigation of the temperature dependence of Raman spectra has been carried out on WO3 powders from room temperature to 800 degrees C. In particular the orthorhombic-to-tetragonal phase transition occurring at about 740 degrees C has been studied for the first time. The Raman active mode at 710 cm(-1) of the orthorhombic phase disappears from the spectrum at temperature below the phase transition point and the Raman activity in the tetragonal phase results very low. A comparative study of hydrogenated tungsten bronzes HxWO3 (x less than or equal to 0.23), where the same transition sequence is driven by an increase of the proton concentration from x = 0 to 0.23, reveals similar behaviour of the high frequency Raman active modes, which are better resolved due to the weaker anharmonic interactions. (C) 1999 Elsevier Science B.V. All rights reserved.
@article{1999-SSI,author={Cazzanelli<sup>#</sup>, E. and Vinegoni, C. and Mariotto, G. and Kuzmin, A. and Purans, J.},title={Raman study of the phase transitions sequence in pure WO3 at high temperature and in HxWO3 with variable hydrogen content},journal={Solid State Ionics},alternatejournal={Solid State Ion.},year={1999},volume={123},number={1-4},pages={67-74},issn={0167-2738},doi={10.1016/s0167-2738(99)00101-0},}
10.1016/s0167-2738(99)00101-0
Phys. Rev. B
Luminescence processes in amorphous hydrogenated silicon-nitride nanometric multilayers
Giorgis#, F.,
Pirri, C. F.,
Vinegoni, C.,
and Pavesi, L.
Radiative recombinations in amorphous Si1-xNx:H-based nanometric multilayer structures have been studied by stationary and time-resolved photoluminescence measurements at room temperature. Such structures show higher photoluminescence efficiency than reference bulk amorphous silicon nitride films. The enhancement is attributed to the localization of electron-hole Fairs induced by the multilayer structure. Fast monomolecular recombination processes are shown, with lifetimes dependent on the emission energy. Analysis of photoluminescence and absorption spectra shows a low density of defects and no intermixing at the multilayer heterointerfaces. Such features open the potential for the investigated structures to function as fast optoelectronic devices. [S0163-1829(99)13635-X].
@article{1999-PRB,author={Giorgis<sup>#</sup>, F. and Pirri, C. F. and Vinegoni, C. and Pavesi, L.},title={Luminescence processes in amorphous hydrogenated silicon-nitride nanometric multilayers},journal={Physical Review B},alternatejournal={Phys. Rev. B},year={1999},volume={60},number={16},pages={11572-11576},issn={2469-9950},doi={10.1103/PhysRevB.60.11572},}
10.1103/PhysRevB.60.11572
Diam. Relat. Mat.
Raman spectroscopy and scanning electron microscopy investigation of annealed amorphous carbon-germanium films deposited by d.c. magnetron sputtering
Mariotto, G.,
Vinegoni, C.,
Jacobsohn, L. G.,
and Freire#, F. L.
The precipitation of germanium nanocrystals in amorphous carbon-germanium films allows for the development of innovative devices, but the accurate control of both size and size distribution of Ge quantum dots in these matrices still constitutes a challenging step. In this paper, both the structure and morphology of amorphous carbon-germanium films (a-Ge1-xCx), deposited by d.c. magnetron sputtering onto silicon substrates and annealed in vacuum at temperatures up to 550 degrees C, are investigated by Raman spectroscopy and scanning electron microscopy. The main features of Raman spectra obtained from carbon-rich films (x > 0.43) are the D and G bands, characteristic of graphitic carbon films. The ratio between the intensities of the bands, I-D/I-G, increases with the annealing temperature, suggesting a progressive increase of the graphitic domains within the films. Raman spectra obtained in the low frequency region from both as-deposited and annealed germanium-rich films (x < 0.43) show broad bands associated with transverse acoustic and transverse optic Ge-Ge modes. Cre-Ge optic modes merge up into a well-shaped peak at 300 cm(-1) in the germanium-richest sample, and underwent annealing treatment at 550 degrees C, thus indicating the precipitation of crystalline Ge. Scanning electron microscopy analysis shows an apparently uniform nucleation of Ge crystallites at the sample surface. Microprobe Raman scattering results suggest the formation of a nearly homogeneous distribution of Ge nanocrystals in germanium-rich films annealed at 550 degrees C. (C) 1999 Elsevier Science S.A. All rights reserved.
@article{1999-DRM,author={Mariotto, G. and Vinegoni, C. and Jacobsohn, L. G. and Freire<sup>#</sup>, F. L.},title={Raman spectroscopy and scanning electron microscopy investigation of annealed amorphous carbon-germanium films deposited by d.c. magnetron sputtering},journal={Diamond and Related Materials},alternatejournal={Diam. Relat. Mat.},year={1999},volume={8},number={2-5},pages={668-672},issn={0925-9635},doi={10.1016/s0925-9635(98)00328-8},}
10.1016/s0925-9635(98)00328-8
Appl. Phys. Lett.
Resonant second harmonic generation in ZnSe bulk microcavity
Pellegrini, V.,
Colombelli, R.,
Carusotto, I.,
Beltram, F.,
Rubini, S.,
Lantier, R.,
Franciosi, A.,
Vinegoni, C.,
and Pavesi#, L.
Room-temperature resonant second harmonic generation is demonstrated in a ZnSe bulk microcavity with Si3N4/SiO2 Bragg reflectors. The resonance occurs at the second harmonic wavelength in the blue-green spectral region and yields an enhancement of one order of magnitude in the second harmonic process. Tunability of the resonant effect between 480 and 500 nm is achieved by varying the angle of incidence. (C) 1999 American Institute of Physics. [S0003-6951(99)01514-4].
@article{1999-APL,author={Pellegrini, V. and Colombelli, R. and Carusotto, I. and Beltram, F. and Rubini, S. and Lantier, R. and Franciosi, A. and Vinegoni, C. and Pavesi<sup>#</sup>, L.},title={Resonant second harmonic generation in ZnSe bulk microcavity},journal={Applied Physics Letters},alternatejournal={Appl. Phys. Lett.},year={1999},volume={74},number={14},pages={1945-1947},issn={0003-6951},doi={10.1063/1.123736},}
10.1063/1.123736
1998
Appl. Phys. Lett.
Photoluminescence of localized excitons in pulsed-laser-deposited GaN
Cazzanelli#, M.,
Cole, D.,
Donegan, J. F.,
Lunney, J. G.,
Middleton, P. G.,
O’Donnell, K. P.,
Vinegoni, C.,
and Pavesi, L.
Continuous-wave photoluminescence (PL) and time-resolved photoluminescence of gallium nitride layers grown by pulsed laser deposition are compared. The temperature dependence of the photoluminescence decay time and the PL-integrated intensity allows a determination of radiative and nonradiative time constants of GaN. We find that luminescence peaks centered at 3.360 and 3.305 eV at low temperature can be attributed to recombination of excitons localized at extended defects. The photoluminescence radiative lifetime at room temperature is on the order of tens of ns. (C) 1998 American Institute of Physics. [S0003-6951(98)00549-X].
@article{1998-APL,author={Cazzanelli<sup>#</sup>, M. and Cole, D. and Donegan, J. F. and Lunney, J. G. and Middleton, P. G. and O’Donnell, K. P. and Vinegoni, C. and Pavesi, L.},title={Photoluminescence of localized excitons in pulsed-laser-deposited GaN},journal={Applied Physics Letters},alternatejournal={Appl. Phys. Lett.},year={1998},volume={73},number={23},pages={3390-3392},issn={0003-6951},doi={10.1063/1.122776},}
10.1063/1.122776
J. Lumines.
Radiative emission properties of a-SiN : H based nanometric multilayers for light emitting devices
Giorgis#, F.,
Pirri, C. F.,
Vinegoni, C.,
and Pavesi, L.
Optical and photoluminescence characterizations were performed on nanometric multilayer structures based on amorphous silicon nitrogen alloys. Evidences are shown that the radiative efficiency of multilayers increases with respect to single-layer structures. This is ascribed to a strong electron-hole pair localization and a low heterointerfaces defect density. Time-resolved photoluminescence measurements yield fast recombination with an energy-dependent lifetime due to hopping processes. Finally, the performance of an electroluminescent device based on multilayers is presented. (C) 1999 Elsevier Science B.V. All rights reserved.
@article{1998-JL,author={Giorgis<sup>#</sup>, F. and Pirri, C. F. and Vinegoni, C. and Pavesi, L.},title={Radiative emission properties of a-SiN : H based nanometric multilayers for light emitting devices},journal={Journal of Luminescence},alternatejournal={J. Lumines.},year={1998},volume={80},number={1-4},pages={423-427},issn={0022-2313},doi={10.1016/s0022-2313(98)00141-0},}
10.1016/s0022-2313(98)00141-0
J. Appl. Phys.
X-ray diffraction, extended x-ray absorption fine structure and Raman spectroscopy studies of WO3 powders and, (1-x)WO3-y center dot xReO(2) mixtures
Kuzmin#, A.,
Purans, J.,
Cazzanelli, E.,
Vinegoni, C.,
and Mariotto, G.
Pure ground tungsten trioxide WO3 and (1-x) WO3-y. xReO(2) mixtures were studied by x-ray absorption spectroscopy, x-ray powder diffraction and Raman spectroscopy in comparison with hydrogen bronzes HxWO3 and hydrogenated calcium tungstate CaWO4:H. It was found that a grinding of pure WO3 leads to a decrease of the crystallites size and a development of the bluish coloration. The color change was found to be reversible under moderate heat treatment or after storage in oxidizing atmosphere and is attributed to the reduced W5+ ions, located at the surface of freshly ground powder. The (1-x) WO3-y. xReO(2) mixtures were found to be composed of monoclinic/orthorhombic WO3 and orthorhombic ReO2 phases with a grain boundary containing reduced W5.7+ ions which are mainly responsible for the compound color at low rhenium ion concentrations. In both cases, the W(6-z+) (0<z less than or equal to 1) color centers are responsible for strong optical absorption resulting in the dramatic decrease of the total Raman intensity. The structural models of free surface in pure ground WO3 and bulk WO3/ReO2 intragrain boundary in (1-x) WO3-y. xReO(2) mixtures are proposed and discussed. (C) 1998 American Institute of Physics. [S0021-8979(98)07322-8].
@article{1998-JAP,author={Kuzmin<sup>#</sup>, A. and Purans, J. and Cazzanelli, E. and Vinegoni, C. and Mariotto, G.},title={X-ray diffraction, extended x-ray absorption fine structure and Raman spectroscopy studies of WO3 powders and, (1-x)WO3-y center dot xReO(2) mixtures},journal={Journal of Applied Physics},alternatejournal={J. Appl. Phys.},year={1998},volume={84},number={10},pages={5515-5524},issn={0021-8979},doi={10.1063/1.368596},}