RESUMEN
Light is a powerful investigational tool in biomedicine, at all levels of structural organization. Its multitude of features (intensity, wavelength, polarization, interference, coherence, timing, non-linear absorption, and even interactions with itself) able to create contrast, and thus images that detail the makeup and functioning of the living state can and should be combined for maximum effect, especially if one seeks simultaneously high spatiotemporal resolution and discrimination ability within a living organism. The resulting high relevance should be directed towards a better understanding, detection of abnormalities, and ultimately cogent, precise, and effective intervention. The new optical methods and their combinations needed to address modern surgery in the operating room of the future, and major diseases such as cancer and neurodegeneration are reviewed here, with emphasis on our own work and highlighting selected applications focusing on quantitation, early detection, treatment assessment, and clinical relevance, and more generally matching the quality of the optical detection approach to the complexity of the disease. This should provide guidance for future advanced theranostics, emphasizing a tighter coupling-spatially and temporally-between detection, diagnosis, and treatment, in the hope that technologic sophistication such as that of a Mars rover can be translationally deployed in the clinic, for saving and improving lives.
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Imagen Óptica , Investigación Biomédica Traslacional , Experimentación Animal , Animales , Diagnóstico por Imagen/métodos , Diagnóstico por Imagen/normas , Manejo de la Enfermedad , Humanos , Microscopía/métodos , Imagen Molecular/métodos , Imagen Multimodal/métodos , Imagen Multimodal/normas , Imagen Óptica/métodos , Imagen Óptica/normas , Investigación , Investigación Biomédica Traslacional/métodosRESUMEN
Advances in image-guided therapy enable physicians to obtain real-time information on neurological disorders such as brain tumors to improve resection accuracy. Image guidance data include the location, size, shape, type, and extent of tumors. Recent technological advances in neurophotonic engineering have enabled the development of techniques for minimally invasive neurosurgery. Incorporation of these methods in intraoperative imaging decreases surgical procedure time and allows neurosurgeons to find remaining or hidden tumor or epileptic lesions. This facilitates more complete resection and improved topology information for postsurgical therapy (i.e., radiation). We review the clinical application of recent advances in neurophotonic technologies including Raman spectroscopy, thermal imaging, optical coherence tomography, and fluorescence spectroscopy, highlighting the importance of these technologies in live intraoperative tissue mapping during neurosurgery. While these technologies need further validation in larger clinical trials, they show remarkable promise in their ability to help surgeons to better visualize the areas of abnormality and enable safe and successful removal of malignancies.
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We investigate the potential of mobile smartphone-based multispectral imaging for the quantitative diagnosis and management of skin lesions. Recently, various mobile devices such as a smartphone have emerged as healthcare tools. They have been applied for the early diagnosis of nonmalignant and malignant skin diseases. Particularly, when they are combined with an advanced optical imaging technique such as multispectral imaging and analysis, it would be beneficial for the early diagnosis of such skin diseases and for further quantitative prognosis monitoring after treatment at home. Thus, we demonstrate here the development of a smartphone-based multispectral imaging system with high portability and its potential for mobile skin diagnosis. The results suggest that smartphone-based multispectral imaging and analysis has great potential as a healthcare tool for quantitative mobile skin diagnosis.
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Changes in the pattern and distribution of both melanocytes (pigment producing) and vasculature (hemoglobin containing) are important in distinguishing melanocytic proliferations. The ability to accurately measure melanin distribution at different depths and to distinguish it from hemoglobin is clearly important when assessing pigmented lesions (benign versus malignant). We have developed a multimode hyperspectral dermoscope (SkinSpect™) able to more accurately image both melanin and hemoglobin distribution in skin. SkinSpect uses both hyperspectral and polarization-sensitive measurements. SkinSpect's higher accuracy has been obtained by correcting for the effect of melanin absorption on hemoglobin absorption in measurements of melanocytic nevi. In vivo human skin pigmented nevi (N=20) were evaluated with the SkinSpect, and measured melanin and hemoglobin concentrations were compared with spatial frequency domain spectroscopy (SFDS) measurements. We confirm that both systems show low correlation of hemoglobin concentrations with regions containing different melanin concentrations (R=0.13 for SFDS, R=0.07 for SkinSpect).
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Dermoscopía/métodos , Melaninas/química , Nevo/diagnóstico por imagen , Neoplasias Cutáneas/diagnóstico por imagen , Piel/diagnóstico por imagen , Análisis Espectral/métodos , Algoritmos , Diseño de Equipo , Humanos , Interpretación de Imagen Asistida por Computador/métodos , Melaninas/análisis , Nevo/irrigación sanguínea , Nevo/química , Imagen Óptica/métodos , Fantasmas de Imagen , Piel/irrigación sanguínea , Piel/química , Neoplasias Cutáneas/irrigación sanguínea , Neoplasias Cutáneas/químicaRESUMEN
Attempts to understand the changes in the structure and physiology of human skin abnormalities by non-invasive optical imaging are aided by spectroscopic methods that quantify, at the molecular level, variations in tissue oxygenation and melanin distribution. However, current commercial and research systems to map hemoglobin and melanin do not correlate well with pathology for pigmented lesions or darker skin. We developed a multimode dermoscope that combines polarization and hyperspectral imaging with an efficient analytical model to map the distribution of specific skin bio-molecules. This corrects for the melanin-hemoglobin misestimation common to other systems, without resorting to complex and computationally intensive tissue optical models. For this system's proof of concept, human skin measurements on melanocytic nevus, vitiligo, and venous occlusion conditions were performed in volunteers. The resulting molecular distribution maps matched physiological and anatomical expectations, confirming a technologic approach that can be applied to next generation dermoscopes and having biological plausibility that is likely to appeal to dermatologists.
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Imagen Óptica/métodos , Piel/patología , Humanos , Procesamiento de Imagen Asistido por Computador , Nevo Pigmentado/diagnóstico , Análisis Espectral/métodos , Vitíligo/diagnósticoRESUMEN
A new approach for generating high-speed multispectral confocal images has been developed. The central concept is that spectra can be acquired for each pixel in a confocal spatial scan by using a fast spectrometer based on optical fiber delay lines. This approach merges fast spectroscopy with standard spatial scanning to create datacubes in real time. The spectrometer is based on a serial array of reflecting spectral elements, delay lines between these elements, and a single element detector. The spatial, spectral, and temporal resolution of the instrument is described and illustrated by multispectral images of laser-induced autofluorescence in biological tissues.
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Procesamiento de Imagen Asistido por Computador/métodos , Microscopía Confocal/métodos , Imagen Óptica/métodos , Animales , Tecnología de Fibra Óptica , Colorantes Fluorescentes , Ratones , Microscopía Confocal/instrumentación , Neoplasias Experimentales , Imagen Óptica/instrumentación , RatasRESUMEN
The HER2+ tumor-targeted nanoparticle, HerDox, exhibits tumor-preferential accumulation and tumor-growth ablation in an animal model of HER2+ cancer. HerDox is formed by non-covalent self-assembly of a tumor targeted cell penetration protein with the chemotherapy agent, doxorubicin, via a small nucleic acid linker. A combination of electrophilic, intercalation, and oligomerization interactions facilitate self-assembly into round 10-20 nm particles. HerDox exhibits stability in blood as well as in extended storage at different temperatures. Systemic delivery of HerDox in tumor-bearing mice results in tumor-cell death with no detectable adverse effects to non-tumor tissue, including the heart and liver (which undergo marked damage by untargeted doxorubicin). HER2 elevation facilitates targeting to cells expressing the human epidermal growth factor receptor, hence tumors displaying elevated HER2 levels exhibit greater accumulation of HerDox compared to cells expressing lower levels, both in vitro and in vivo. Fluorescence intensity imaging combined with in situ confocal and spectral analysis has allowed us to verify in vivo tumor targeting and tumor cell penetration of HerDox after systemic delivery. Here we detail our methods for assessing tumor targeting via multimode imaging after systemic delivery.
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Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/enzimología , Doxorrubicina/administración & dosificación , Nanopartículas/administración & dosificación , Receptor ErbB-2/metabolismo , Animales , Antibióticos Antineoplásicos/administración & dosificación , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Doxorrubicina/química , Doxorrubicina/farmacocinética , Estabilidad de Medicamentos , Femenino , Humanos , Ratones , Ratones Desnudos , Microscopía Fluorescente/métodos , Nanopartículas/química , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The tumor-targeted corrole particle, HerGa, displays preferential toxicity to tumors in vivo and can be tracked via fluorescence for simultaneous detection, imaging, and treatment. We have recently uncovered an additional feature of HerGa in that its cytotoxicity is enhanced by light irradiation. In the present study, we have elucidated the cellular mechanisms for HerGa photoexcitation-mediated cell damage using fluorescence optical imaging. In particular, we found that light irradiation of HerGa produces singlet oxygen, causing mitochondrial damage and cytochrome c release, thus promoting apoptotic cell death. An understanding of the mechanisms of cell death induced by HerGa, particularly under conditions of light-mediated excitation, may direct future efforts in further customizing this nanoparticle for additional therapeutic applications and enhanced potency.
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Antineoplásicos/administración & dosificación , Nanopartículas/administración & dosificación , Neoplasias/tratamiento farmacológico , Porfirinas/administración & dosificación , Oxígeno Singlete , Antineoplásicos/efectos de la radiación , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citocromos c/metabolismo , Humanos , Luz , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiología , Nanopartículas/efectos de la radiación , Neoplasias/metabolismo , Porfirinas/efectos de la radiaciónRESUMEN
This study presents a new multimodal imaging approach that includes high-frequency ultrasound, fluorescence intensity, confocal, and spectral imaging to improve the preclinical evaluation of new therapeutics in vivo. Here we use this approach to assess in vivo the therapeutic efficacy of the novel chemotherapy construct, HerDox during and after treatment. HerDox is comprised of doxorubicin non-covalently assembled in a viral-like particle targeted to HER2+ tumor cells, causing tumor cell death at over 10-fold lower dose compared to the untargeted drug, while sparing the heart. Whereas our initial proof-of-principle studies on HerDox used tumor growth/shrinkage rates as a measure of therapeutic efficacy, here we show that multimodal imaging deployed during and after treatment can supplement traditional modes of tumor monitoring to further characterize the particle in tissues of treated mice. Specifically, we show here that tumor cell apoptosis elicited by HerDox can be monitored in vivo during treatment using high frequency ultrasound imaging, while in situ confocal imaging of excised tumors shows that HerDox indeed penetrated tumor tissue and can be detected at the subcellular level, including in the nucleus, via Dox fluorescence. In addition, ratiometric spectral imaging of the same tumor tissue enables quantitative discrimination of HerDox fluorescence from autofluorescence in situ. In contrast to standard approaches of preclinical assessment, this new method provides multiple/complementary information that may shorten the time required for initial evaluation of in vivo efficacy, thus potentially reducing the time and cost for translating new drug molecules into the clinic.
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Doxorrubicina/química , Doxorrubicina/farmacología , Imagen Molecular/métodos , Nanopartículas , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Doxorrubicina/metabolismo , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Ratones , Terapia Molecular Dirigida , Receptor ErbB-2/metabolismo , Espectrometría de Fluorescencia , Resultado del Tratamiento , UltrasonidoRESUMEN
AIM: This study tests the hypothesis that DNA intercalation and electrophilic interactions can be exploited to noncovalently assemble doxorubicin in a viral protein nanoparticle designed to target and penetrate tumor cells through ligand-directed delivery. We further test whether this new paradigm of doxorubicin targeting shows therapeutic efficacy and safety in vitro and in vivo. MATERIALS & METHODS: We tested serum stability, tumor targeting and therapeutic efficacy in vitro and in vivo using biochemical, microscopy and cytotoxicity assays. RESULTS: Self-assembly formed approximately 10-nm diameter serum-stable nanoparticles that can target and ablate HER2+ tumors at >10× lower dose compared with untargeted doxorubicin, while sparing the heart after intravenous delivery. The targeted nanoparticle tested here allows doxorubicin potency to remain unaltered during assembly, transport and release into target cells,while avoiding peripheral tissue damage and enabling lower, and thus safer, drug dose for tumor killing. CONCLUSION: This nanoparticle may be an improved alternative to chemical conjugates and signal-blocking antibodies for tumor-targeted treatment.
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Antibióticos Antineoplásicos/administración & dosificación , ADN/química , Doxorrubicina/administración & dosificación , Sistemas de Liberación de Medicamentos , Nanopartículas/química , Proteínas Virales/química , Animales , Antibióticos Antineoplásicos/metabolismo , Antibióticos Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Doxorrubicina/metabolismo , Doxorrubicina/uso terapéutico , Estabilidad de Medicamentos , Femenino , Humanos , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Receptor ErbB-2/metabolismo , Suero/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We recently reported that a targeted, brightly fluorescent gallium corrole (HerGa) is highly effective for breast tumor detection and treatment. Unlike structurally similar porphryins, HerGa exhibits tumor-targeted toxicity without the need for photoexcitation. We have now examined whether photoexcitation further modulates HerGa toxicity, using multimode optical imaging of live cells, including two-photon excited fluorescence, differential interference contrast (DIC), spectral, and lifetime imaging. Using two-photon excited fluorescence imaging, we observed that light at specific wavelengths augments the HerGa-mediated mitochondrial membrane potential disruption of breast cancer cells in situ. In addition, DIC, spectral, and fluorescence lifetime imaging enabled us to both validate cell damage by HerGa photoexcitation and investigate HerGa internalization, thus allowing optimization of light dose and timing. Our demonstration of HerGa phototoxicity opens the way for development of new methods of cancer intervention using tumor-targeted corroles.
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Colorantes Fluorescentes/farmacología , Galio/farmacología , Mitocondrias/efectos de los fármacos , Mitocondrias/efectos de la radiación , Porfirinas/farmacología , Antineoplásicos/química , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Línea Celular Tumoral , Supervivencia Celular , Colorantes Fluorescentes/química , Colorantes Fluorescentes/farmacocinética , Galio/química , Galio/farmacocinética , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de la radiación , Microscopía Fluorescente/métodos , Mitocondrias/patología , Compuestos Organometálicos/química , Procesos Fotoquímicos/efectos de los fármacos , Procesos Fotoquímicos/efectos de la radiación , Porfirinas/química , Porfirinas/farmacocinéticaRESUMEN
The aim of this study was to proof applicability of hyperspectral imaging for the analysis and classification of human mucosal surfaces in vivo. The larynx as a prototypical anatomically well-defined surgical test area was analyzed by microlaryngoscopy with a polychromatic lightsource and a synchronous triggered monochromatic CCD-camera. Image stacks (5 benign, 7 malignant tumors) were analyzed by established software (principal component analysis PCA, hyperspectral classification, spectral profiles). Hyperspectral image datacubes were analyzed and classified by conventional software. In PCA, images at 590-680 nm loaded most onto the first PC which typically contained 95% of the total information. Hyperspectral classification clustered the data highlighting altered mucosa. The spectral profiles clearly differed between the different groups. Hyperspectral imaging can be applied to mucosal surfaces. This approach opens the way to analyze spectral characteristics of histologically different lesions in order to build up a spectral library and to allow non-touch optical biopsy.
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Imagen Molecular/métodos , Membrana Mucosa/citología , Humanos , Procesamiento de Imagen Asistido por Computador , Proyectos Piloto , Análisis de Componente Principal , Programas Informáticos , Análisis Espectral , Propiedades de SuperficieRESUMEN
PURPOSE: Several established optical imaging approaches have been applied, usually in isolation, to preclinical studies; however, truly useful in vivo imaging may require a simultaneous combination of imaging modalities to examine dynamic characteristics of cells and tissues. We developed a new multimode optical imaging system designed to be application-versatile, yielding high sensitivity, and specificity molecular imaging. PROCEDURES: We integrated several optical imaging technologies, including fluorescence intensity, spectral, lifetime, intravital confocal, two-photon excitation, and bioluminescence, into a single system that enables functional multiscale imaging in animal models. RESULTS: The approach offers a comprehensive imaging platform for kinetic, quantitative, and environmental analysis of highly relevant information, with micro-to-macroscopic resolution. Applied to small animals in vivo, this provides superior monitoring of processes of interest, represented here by chemo-/nanoconstruct therapy assessment. CONCLUSIONS: This new system is versatile and can be optimized for various applications, of which cancer detection and targeted treatment are emphasized here.
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Antineoplásicos/uso terapéutico , Imagen Molecular/métodos , Neoplasias/diagnóstico , Neoplasias/tratamiento farmacológico , Dispositivos Ópticos , Ensayos Antitumor por Modelo de Xenoinjerto , Animales , Línea Celular Tumoral , Estudios de Factibilidad , Humanos , Mediciones Luminiscentes , Ratones , Ratones Desnudos , Nanopartículas , Especificidad de Órganos , Ratas , Espectrometría de FluorescenciaRESUMEN
HerGa is a self-assembled tumor-targeted particle that bears both tumor detection and elimination activities in a single, two-component complex (Agadjanian et al. Proc. Natl. Acad. Sci. U.S.A.2009, 106, 6105-6110). Given its multifunctionality, HerGa (composed of the fluorescent cytotoxic corrole macrocycle, S2Ga, noncovalently bound to the tumor-targeted cell penetration protein, HerPBK10) has the potential for high clinical impact, but its mechanism of cell killing remains to be elucidated, and hence is the focus of the present study. Here we show that HerGa requires HerPBK10-mediated cell entry to induce toxicity. HerGa (but not HerPBK10 or S2Ga alone) induced mitochondrial membrane potential disruption and superoxide elevation, which were both prevented by endosomolytic-deficient mutants, indicating that cytosolic exposure is necessary for corrole-mediated cell death. A novel property discovered here is that corrole fluorescence lifetime acts as a pH indicator, broadcasting the intracellular microenvironmental pH during uptake in live cells. This feature in combination with two-photon imaging shows that HerGa undergoes early endosome escape during uptake, avoiding compartments of pH < 6.5. Cytoskeletal disruption accompanied HerGa-mediated mitochondrial changes whereas oxygen scavenging reduced both events. Paclitaxel treatment indicated that HerGa uptake requires dynamic microtubules. Unexpectedly, low pH is insufficient to induce release of the corrole from HerPBK10. Altogether, these studies identify a mechanistic pathway in which early endosomal escape enables HerGa-induced superoxide generation leading to cytoskeletal and mitochondrial damage, thus triggering downstream cell death.
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Sistemas de Liberación de Medicamentos , Inmunotoxinas/toxicidad , Porfirinas/toxicidad , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Humanos , Concentración de Iones de Hidrógeno , Microscopía Confocal , Modelos Biológicos , Neoplasias/patología , Paclitaxel/farmacologíaRESUMEN
We report a novel in vivo spectral imaging approach to cancer detection and chemotherapy assessment. We describe and characterize a ratiometric spectral imaging and analysis method and evaluate its performance for tumor detection and delineation by quantitatively monitoring the specific accumulation of targeted gallium corrole (HerGa) into HER2-positive (HER2 +) breast tumors. HerGa temporal accumulation in nude mice bearing HER2 + breast tumors was monitored comparatively by a. this new ratiometric imaging and analysis method; b. established (reflectance and fluorescence) spectral imaging; c. more commonly used fluorescence intensity imaging. We also tested the feasibility of HerGa imaging in vivo using the ratiometric spectral imaging method for tumor detection and delineation. Our results show that the new method not only provides better quantitative information than typical spectral imaging, but also better specificity than standard fluorescence intensity imaging, thus allowing enhanced in vivo outlining of tumors and dynamic, quantitative monitoring of targeted chemotherapy agent accumulation into them.
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Medios de Contraste , Monitoreo de Drogas/métodos , Imagen Molecular/métodos , Neoplasias/patología , Algoritmos , Animales , Medios de Contraste/farmacocinética , Femenino , Galio/farmacocinética , Ratones , Ratones Desnudos , Neoplasias/tratamiento farmacológico , Neoplasias/enzimología , Neoplasias/metabolismo , Receptor ErbB-2/biosíntesis , Espectrometría de Fluorescencia , Trasplante HeterólogoRESUMEN
We report fast, non-scanning, wide-field two-photon fluorescence excitation with spectral and lifetime detection for in vivo biomedical applications. We determined the optical characteristics of the technique, developed a Gaussian flat-field correction method to reduce artifacts resulting from non-uniform excitation such that contrast is enhanced, and showed that it can be used for ex vivo and in vivo cellular-level imaging. Two applications were demonstrated: (i) ex vivo measurements of beta-amyloid plaques in retinas of transgenic mice, and (ii) in vivo imaging of sulfonated gallium(III) corroles injected into tumors. We demonstrate that wide-field two photon fluorescence excitation with flat-field correction provides more penetration depth as well as better contrast and axial resolution than the corresponding one-photon wide field excitation for the same dye. Importantly, when this technique is used together with spectral and fluorescence lifetime detection modules, it offers improved discrimination between fluorescence from molecules of interest and autofluorescence, with higher sensitivity and specificity for in vivo applications.
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Endothelial cells (ECs) represent the major component of the embryonic pancreatic niche and play a key role in the differentiation of insulin-producing ß cells in vivo. However, it is unknown if ECs promote such differentiation in vitro. We investigated whether interaction of ECs with mouse embryoid bodies (EBs) in culture promotes differentiation of pancreatic progenitors and insulin-producing cells and the mechanisms involved. We developed a co-culture system of mouse EBs and human microvascular ECs (HMECs). An increase in the expression of the pancreatic markers PDX-1, Ngn3, Nkx6.1, proinsulin, GLUT-2, and Ptf1a was observed at the interface between EBs and ECs (EB-EC). No expression of these markers was found at the periphery of EBs cultured without ECs or those co-cultured with mouse embryonic fibroblasts (MEFs). At EB-EC interface, proinsulin and Nkx6.1 positive cells co-expressed phospho-Smad1/5/8 (pSmad1/5/8). Therefore, EBs were treated with HMEC conditioned media (HMEC-CM) suspecting soluble factors involved in bone morphogenetic protein (BMP) pathway activation. Upregulation of PDX-1, Ngn3, Nkx6.1, insulin-1, insulin-2, amylin, SUR1, GKS, and amylase as well as down-regulation of SST were detected in treated EBs. In addition, higher expression of BMP-2/-4 and their receptor (BMPR1A) were also found in these EBs. Recombinant human BMP-2 (rhBMP-2) mimicked the effects of the HMEC-CM on EBs. Noggin (NOG), a BMP antagonist, partially inhibited these effects. These results indicate that the differentiation of EBs to pancreatic progenitors and insulin-producing cells can be enhanced by ECs in vitro and that BMP pathway activation is central to this process.
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Proteínas Morfogenéticas Óseas/metabolismo , Diferenciación Celular/fisiología , Células Madre Embrionarias/fisiología , Células Endoteliales/fisiología , Células Secretoras de Insulina/fisiología , Páncreas/citología , Transducción de Señal/fisiología , Animales , Biomarcadores/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Cuerpos Embrioides/citología , Cuerpos Embrioides/fisiología , Células Madre Embrionarias/citología , Células Endoteliales/citología , Humanos , Insulina/metabolismo , Células Secretoras de Insulina/citología , Ratones , Células Madre/citología , Células Madre/fisiologíaRESUMEN
Noninvasive monitoring of ß-amyloid (Aß) plaques, the neuropathological hallmarks of Alzheimer's disease (AD), is critical for AD diagnosis and prognosis. Current visualization of Aß plaques in brains of live patients and animal models is limited in specificity and resolution. The retina as an extension of the brain presents an appealing target for a live, noninvasive optical imaging of AD if disease pathology is manifested there. We identified retinal Aß plaques in postmortem eyes from AD patients (n=8) and in suspected early stage cases (n=5), consistent with brain pathology and clinical reports; plaques were undetectable in age-matched non-AD individuals (n=5). In APP(SWE)/PS1(∆E9) transgenic mice (AD-Tg; n=18) but not in non-Tg wt mice (n=10), retinal Aß plaques were detected following systemic administration of curcumin, a safe plaque-labeling fluorochrome. Moreover, retinal plaques were detectable earlier than in the brain and accumulated with disease progression. An immune-based therapy effective in reducing brain plaques, significantly reduced retinal Aß plaque burden in immunized versus non-immunized AD mice (n=4 mice per group). In live AD-Tg mice (n=24), systemic administration of curcumin allowed noninvasive optical imaging of retinal Aß plaques in vivo with high resolution and specificity; plaques were undetectable in non-Tg wt mice (n=11). Our discovery of Aß specific plaques in retinas from AD patients, and the ability to noninvasively detect individual retinal plaques in live AD mice establish the basis for developing high-resolution optical imaging for early AD diagnosis, prognosis assessment and response to therapies.
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Enfermedad de Alzheimer/patología , Colorantes , Curcumina , Imagenología Tridimensional/métodos , Placa Amiloide/patología , Retina/patología , Anciano , Anciano de 80 o más Años , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones Transgénicos , Microscopía Fluorescente , Persona de Mediana Edad , Espectrometría de FluorescenciaRESUMEN
Prevention and early detection of breast cancer are the major prophylactic measures taken to reduce the breast cancer related mortality and morbidity. Clinical management of breast cancer largely relies on the efficacy of the breast-conserving surgeries and the subsequent radiation therapy. A key problem that limits the success of these surgeries is the lack of accurate, real-time knowledge about the positive tumor margins in the surgically excised tumors in the operating room. This leads to tumor recurrence and, hence, the need for repeated surgeries. Current intraoperative techniques such as frozen section pathology or touch imprint cytology severely suffer from poor sampling and non-optimal detection sensitivity. Even though histopathology analysis can provide information on positive tumor margins post-operatively (~2-3 days), this information is of no immediate utility in the operating rooms. In this article, we propose a novel image analysis method for tumor margin assessment based on nuclear morphometry and tissue topology and demonstrate its high sensitivity/specificity in preclinical animal model of breast carcinoma. The method relies on imaging nuclear-specific fluorescence in the excised surgical specimen and on extracting nuclear morphometric parameters (size, number, and area fraction) from the spatial distribution of the observed fluorescence in the tissue. We also report the utility of tissue topology in tumor margin assessment by measuring the fractal dimension in the same set of images. By a systematic analysis of multiple breast tissues specimens, we show here that the proposed method is not only accurate (~97% sensitivity and 96% specificity) in thin sections, but also in three-dimensional (3D) thick tissues that mimic the realistic lumpectomy specimens. Our data clearly precludes the utility of nuclear size as a reliable diagnostic criterion for tumor margin assessment. On the other hand, nuclear area fraction addresses this issue very effectively since it is a combination of both nuclear size and count in any given region of the analyzed image, and thus yields high sensitivity and specificity (~97%) in tumor detection. This is further substantiated by an independent parameter, fractal dimension, based on the tissue topology. Although the basic definition of cancer as an uncontrolled cell growth entails a high nuclear density in tumor regions, a simple but systematic exploration of nuclear distribution in thick tissues by nuclear morphometry and tissue topology as performed in this study has never been carried out, to the best of our knowledge. We discuss the practical aspects of implementing this imaging approach in automated tissue sampling scenario where the accuracy of tumor margin assessment can be significantly increased by scanning the entire surgical specimen rather than sampling only a few sections as in current histopathology analysis.