Longitudinal-Bildgebung ist in präklinischen Studien eingesetzt, um den Fortschritt der Krankheit zu folgen oder messen die Wirkung einer therapeutischen. In der Onkologie bieten optische Verfahren rigorose Werkzeuge, um das Tumorwachstum zu überwachen und liefern präzise Quantifizierung des Zellwachstums oder der Genexpression zu jedem Zeitpunkt in einer solchen Studie. Anatomische Veränderungen können mit Hilfe eines hochauflösenden Technik wie MicroCT sein, aber für die Längs-Bildgebung eine niedrige Röntgendosis muss verwendet werden, um biologische Artefakte zu vermeiden. Optische und MicroCT Bilder können co-registriert sein, um eine Kombination von funktionellen und anatomischen Daten sichergestellt wird, dass ein Maximum an Informationen aus dem Tiermodell extrahiert bieten. 1. Zellpräparation <ol><li> Caliper bietet eine Reihe von Luciferase exprimierenden Zelllinien für die präklinische Forschung in Mausmodellen. </li><li> MDA-MB-231-luc-D2H2LN ist eine menschliche Brusttumor-Zelllinie exprimiert das Luciferase-Gen, das als eine optische Anzeige der Tumorgenese in vivo verwendet werden können. Diese Zelllinie ist aus einer spontanen Lymphknotenmetastasen, die von einem D3H1 Brustfettpolster Tumor entstanden ist erstellt und ist dafür bekannt, aggressiv Metastasen bilden. </li><li> Die Zellen werden als pathogen-freien gefrorenen Kultur, die sich leicht wächst in Standard-Medien mit keine Notwendigkeit für Selektionsmarker zur Verfügung gestellt. </li><li> Um zu überprüfen, Luciferaseaktivität vor der Injektion in das Tier eine 90% konfluent Kolben wird durch Trypsinierung geerntet. Luciferaseaktivität wird durch den Verzicht auf 50.000 Zellen in einer Mikrotiterplatte und darstellende Verdünnungsreihe gemessen. </li></ol> 2. Intrakardiale Injektion von Zellen in Tieren <ol><li> Vor der Injektion werden die Tiere narkotisiert mit 3% Isofluran. </li><li> 1 bis 3.000.000 Zellen sind in 50ul Volumen in den linken Ventrikel injiziert. Zell-Suspension enthält 150ug/ml D-Luciferin zu Injektionstechnik zu validieren. </li><li> Zur Bestätigung einer korrekten intrakardiale Injektion Mäuse sind in der IVIS System abgebildet werden. Mäuse, die BLI-Signal zeigen, im ganzen Körper wurden korrekt injiziert. Wenn das Signal lokalisiert wurden die Zellen nicht richtig in das Herz injiziert. </li></ol> 3. BLI Bildgebung zu überwachen Metastasen <ol><li> Die Imaging-Shuttle unterstützt die Maus für die Datenerfassung in beiden IVIS Spectrum und der QuantumFX MicroCT System. </li><li> Der Shuttle hilft bei der Erhaltung des Tieres in der gleichen Position für die Bildgebung und beinhaltet fiducial Signale in automatische Co-Registrierung zu unterstützen. Die Kammer ist der isofluorance Verdampfer verbunden Narkose aufrecht zu erhalten. </li><li> Imaging-Assistenten in der Living Image Software 4.1 kann verwendet werden, um automatisch die Belichtungszeit, Blende und Binning und sorgen für quantitative Datenerhebung werden. </li><li> Mehrere Bilder können erworben werden und im Vergleich in Längsschnittstudien für Sekunden oder Monaten, je nach der Art des Experiments. </li><li> Tag 6 nach der Injektion von Zellen, zeigt BLI Bildgebung mit IVIS Spectrum Metastasen in den Knochen in sieben von neun Mäusen. </li></ol> 4. Longitudinal Monitoring von Osteolysen mit Mikro-CT <ol><li> Mäuse, die klare Knochenmetastasen sichtbar mit BLI zeigten, wurden in den Quantum FX Mikro-CT-System abgebildet. </li><li> Die Quantum FX Mikro-CT-System liefert qualitativ hochwertige Bilder bei einer Röntgen-Dosis niedrig genug, um wahr Längs-Mikro-CT in präklinischen Studien ermöglichen. </li><li> Knochenabbau in beiden Beinen in Längsrichtung überwacht werden mit diesem System in weniger als einer Minute. </li><li> Mäuse sind narkotisiert mit 3% Isofluran und kann direkt aus dem IVIS Spectrum-System übertragen werden. </li><li> Zwei 60mm-Bilder sind zusammen, um das ganze Tier sehen zu erfassen genäht. </li><li> A 30mm FOV ist ideal für die Bildgebung des Kniegelenks in hoher Auflösung. </li><li> Ein einzelner Scan mit ausreichend hoher Auflösung zu messen und zu quantifizieren Knochenmetastasen dauert nur 18 Sekunden und liefert eine Dosis von nur 13mGy. </li><li> Die Mäuse wurden bis zu 8 mal in 36 Tagen mit dem Mikro-CT-System abgebildet, ohne die das Tumorwachstum oder der Gesundheit des Tieres. </li><li> Die Mikro-CT-Bildgebung zeigte, dass Knochen leisions sichtbar am Tag 10 wurden fortgeschritten und sehr schnell, entsprechend der Zunahme der BLI-Signal. </li></ol> 5. 3D-Co-Registrierung und Analyse von BLI und UCT <ol><li> Nachdem die Bilder haben sowohl auf der IVIS Spektrum und Quantum FX Mikro-CT-System Daten erfasst in eine 3D-Rekonstruktion co-registriert werden. </li><li> MicroCT Bilder werden als DICOM-Dateien exportiert und in das Wohnzimmer Bild 4.1 3D Multi Modalität Werkzeug. </li><li> In der Living Image Software laufen DLIT Programm dreidimensional rekonstruieren das optische Signal. </li><li> Die fiducial Tool in der Software wird verwendet, um Co-Registrierung der optischen BLI Signal an die Mikro-CT-Bild. </li><li> Koregistrierten Datensätze können quantitativ analysiert werden, um mehrere Parameter während der Progression der Erkrankung zu überwachen sind wir hier verfolgen das Wachstum der Metastase zusammen mit der Entwicklung der Osteolyse in den Knochen. </li> &lt;/ Ol&gt;

<ol>
	<li>Jenkins, D. E., Hornig, Y. S., Oei, Y., Dusich, J., Purchio, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioluminescent+Human+Breast+Cancer+Cell+Lines+that+Permit+Rapid+and+Sensitive+in+vivo+Detection+of+Mammary+Tumors+and+Multiple+Metastases+in+Immune+Deficient+Mice.">Bioluminescent Human Breast Cancer Cell Lines that Permit Rapid and Sensitive in vivo Detection of Mammary Tumors and Multiple Metastases in Immune Deficient Mice.</a> Breast Cancer Research. 7, R444-R454 (2005).</li><li>Cowey, S., Szafran, A. A., Kappes, J., Zinn, K. R., Siegal, G. P., Desmond, R. A., Kim, H., Evans, L., Hardy, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breast+Cancer+Metastasis+to+Bone:+Evaluation+of+Bioluminescent+Imaging+and+MicroSPECT/CT+for+Detecting+Bone+Metastasis+in+Immunodeficient+Mice.">Breast Cancer Metastasis to Bone: Evaluation of Bioluminescent Imaging and MicroSPECT/CT for Detecting Bone Metastasis in Immunodeficient Mice.</a> Clinical and Experimental Metastasis. 24, 389-401 (2007).</li><li>Labrinidis, A., Hay, S., Liapis, V., Ponomarev, V., Findlay, D. M., Evdokiou, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zoledronic+Acid+Inhibits+Both+the+Osteolytic+and+Osteoblastic+Components+of+Osteosarcoma+Lesions+in+a+Mouse+Model.">Zoledronic Acid Inhibits Both the Osteolytic and Osteoblastic Components of Osteosarcoma Lesions in a Mouse Model.</a> Clinical Cancer Research. 15, (2009).</li><li>Kaijzel, E. L., Pluijm, G. v. a. n. d. e. r., Lowik, C. W. G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole-Body+Optical+Imaging+in+Animal+Models+to+Assess+Cancer+Development+and+Progression.">Whole-Body Optical Imaging in Animal Models to Assess Cancer Development and Progression.</a> Clinical Cancer Research. 13, 3490-3497 (2007).</li><li>Minn, A. J., Kang, Y., Serganova, I., Gupta, G. P., Giri, D. D., Doubrovin, M., Ponomarev, V., Gerald, W. L., Blasberg, R., Massague, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distinct+Organ-Specific+Metastatic+Potential+of+Individual+Breast+Cancer+Cells+and+Primary+Tumors.">Distinct Organ-Specific Metastatic Potential of Individual Breast Cancer Cells and Primary Tumors.</a> The Journal of Clinical Investigation. 115, (2005).</li><li>Xu, H., Christensen, A., Kuo, C., Nilson, D., Lim, E., Whalen, J., Kim, J. B., Zhang, N., Singh, R., Oldfield, S., Troy, T., Rice, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Co-registration+of+in+vivo+Optical+Tomography+and+Micro-CT+for+Longitudinal+Studies.">Co-registration of in vivo Optical Tomography and Micro-CT for Longitudinal Studies.</a> Word Molecular Imaging Congress. , (2010).</li></ol>

Die Autoren sind Mitarbeiter von Caliper Life Sciences, der das Instrument in diesem Artikel vorgestellten machen.

Kleintier-Bildgebung mit MicroCT hat die Überwachung osteolytische Läsionen wurden unter pathologischen Bedingungen, wie Tumor-Entwicklung (..; Labrinidis et al, 2009 Cowey et al 2007) angewendet. Als Biolumineszenz-Bildgebung der Luciferase gekennzeichnet Tumoren weitgehend angewandt worden präklinisch, vorausgesetzt, Co-Registrierung der Biolumineszenz und MicroCT Bilder eine bessere Definition der anatomischen Lage der Biolumineszenz-Signale (Kaijzel et al. 2007). Früheren Studie mit einem Luciferase gekennzeichnet Brusttumor-Zelllinie MDA-MB-231-luc-D3H2LN zeigte Metastasen in Knochen und Entwicklung von Osteolysen folgenden intrakardiale Injektion (Jenkins et al 2005;. Minn et al, 2005). In dieser Studie untersuchten wir eine Längsschnittstudie von Tumormetastasen und osteolytische Aktivität mit Kombinatorik optischen und strahlungsarme-Dosis MicroCT Bildgebung mit der systemischen MDA-MB-231-luc-D3H2LN Metastasen-Modell. Wir führten sequentielle Bildgebung mit IVIS Spectrum und Quantum FX MicroCT Systeme und erreicht Biolumineszenz-und CT-Co-Registrierung mit unserem neu entwickelten Wohnzimmer Bild 4.1-Software (Heng et al., 2010). Biolumineszenz-Bildgebung mit IVIS bietet nicht-invasive Detektion der Tumorzellen mit hoher Empfindlichkeit, so dass die Tumoren in einem frühen Stadium erkannt werden. Imaging mit Quantum FX ermöglicht die genaue 3D-Rekonstruktion der Knochenläsionen und detaillierten anatomischen Informationen. Quantum FX-System bietet niedrige Strahlendosis Bildgebung, die Längs-Überwachung des Krankheitsverlaufs ermöglicht. Korrelation von Knochenläsionen und Tumormetastasen war in der Zusammenarbeit registrierte Bild dargestellt, und der Vorteil der Multimodalität Verfolgung von Läsion Entwicklung längs eindeutig nachgewiesen werden konnte.

monitoring tumor metastases and osteolytic lesions with bioluminescence and micro ct imaging

Nach intrakardialen Lieferung von MDA-MB-231-luc-D3H2LN Zellen Nu / Nu-Mäusen, systemische Metastasen in den injizierten Tieren entwickelt. Biolumineszenz-Bildgebung mit IVIS Spectrum wurde eingesetzt, um den Vertrieb und die Entwicklung der Tumorzellen nach der Lieferung Verfahrens einschließlich DLIT Rekonstruktion des Tumors Signal und seine Lage Maßnahme zu überwachen. Entwicklung von Metastasen an den Knochen Gewebe löst osteolytische Aktivität und Läsionen an Tibia und Femur wurden längs mit Mikro-CT untersucht. Imaging wurde mit einem Quantum FX Mikro-CT-System mit schnellen Bildgebung und niedrige Röntgendosis. Die geringe Strahlendosis ermöglicht mehreren Imaging-Sitzungen mit einer kumulativen X-ray Dosierung weit unter LD50 durchgeführt werden. Eine Maus Imaging-Shuttle-Gerät wurde verwendet, um nacheinander Bild der Mäuse mit beiden IVIS Spectrum und Quantum FX Voraussetzung für präzise Positionierung Tier sowohl in der Biolumineszenz-und CT-Bilder. Die optischen und CT-Datensätze wurden in 3-dimentions co-registriert die Living Bild 4.1-Software. Diese Multi-Mode-Ansatz ermöglicht eine engmaschige Überwachung des Tumorwachstums und der Entwicklung gleichzeitig mit osteolytischen Aktivität.

Watch this Scientific Journal Video about Monitoring Tumor Metastases and Osteolytic Lesions with Bioluminescence and Micro CT Imaging at JoVE.com

Monitoring Tumor Metastases and Osteolytic Lesions with Bioluminescence and Micro CT Imaging

Ein experimentelles Mausmodell von Knochenmetastasen wurde nach intrakardialen Lieferung von Luciferase exprimierenden Mammatumorzellen gegründet. Tumorentstehung und führte Osteolyse wurden in Längsrichtung mit Biolumineszenz-und Mikro-CT-Bildgebung überwacht.

Überwachung Tumormetastasen und Osteolysen mit Biolumineszenz und Micro-CT Imaging

Following intracardiac delivery of MDA-MB-231-luc-D3H2LN cells to Nu/Nu mice, systemic metastases developed in the injected animals. Bioluminescence ...

monitoring-tumor-metastases-osteolytic-lesions-with-bioluminescence

Research

JoVE Journal

Medicine

24.5K Aufrufe.  Caliper Life Sciences.  中文 

Serie: Monitoring Tumor Metastases and Osteolytic Lesions with Bioluminescence and Micro CT Imaging

In vivo Bioluminescence Imaging of Tumor Hypoxia Dynamics of Breast Cancer Brain Metastasis in a Mouse Model

It is well recognized that tumor hypoxia plays an important role in promoting malignant progression and affecting therapeutic response negatively. There is little knowledge about in situ, in vivo, tumor hypoxia during intracranial development of malignant brain tumors because of lack of efficient means to monitor it in these deep-seated orthotopic tumors. Bioluminescence imaging (BLI), based on the detection of light emitted by living cells expressing a luciferase gene, has been rapidly adopted for cancer research, in particular, to evaluate tumor growth or tumor size changes in response to treatment in preclinical animal studies. Moreover, by expressing a reporter gene under the control of a promoter sequence, the specific gene expression can be monitored non-invasively by BLI. Under hypoxic stress, signaling responses are mediated mainly via the hypoxia inducible factor-1&alpha; (HIF-1&alpha;) to drive transcription of various genes. Therefore, we have used a HIF-1&alpha; reporter construct, 5HRE-ODD-luc, stably transfected into human breast cancer MDA-MB231 cells (MDA-MB231/5HRE-ODD-luc). In vitro HIF-1&alpha; bioluminescence assay is performed by incubating the transfected cells in a hypoxic chamber (0.1% O2) for 24 hr before BLI, while the cells in normoxia (21% O2) serve as a control. Significantly higher photon flux observed for the cells under hypoxia suggests an increased HIF-1&alpha; binding to its promoter (HRE elements), as compared to those in normoxia. Cells are injected directly into the mouse brain to establish a breast cancer brain metastasis model. In vivo bioluminescence imaging of tumor hypoxia dynamics is initiated 2 wks after implantation and repeated once a week. BLI reveals increasing light signals from the brain as the tumor progresses, indicating increased intracranial tumor hypoxia. Histological and immunohistochemical studies are used to confirm the in vivo imaging results. Here, we will introduce approaches of in vitro HIF-1&alpha; bioluminescence assay, surgical establishment of a breast cancer brain metastasis in a nude mouse and application of in vivo bioluminescence imaging to monitor intracranial tumor hypoxia.

In vivo Bioluminescence Imaging of Tumor Hypoxia Dynamics of Breast Cancer Brain Metastasis in a Mouse Model

Bioluminescence imaging of hypoxia inducible factor-1&alpha; activity is applied to monitor intracranial tumor hypoxia development in a breast cancer brain metastasis mouse model.

In-vivo-Biolumineszenz-Bildgebung von Tumorhypoxie Dynamics of Breast Cancer Hirnmetastasen in einem Mausmodell

It is well recognized that tumor hypoxia plays an important role in promoting malignant progression and affecting therapeutic response negatively. There ...

Forschung

Medizin

A Novel High-resolution In vivo Imaging Technique to Study the Dynamic Response of Intracranial Structures to Tumor Growth and Therapeutics

We have successfully integrated previously established Intracranial window (ICW) technology 1-4 with intravital 2-photon confocal microscopy to develop a novel platform that allows for direct long-term visualization of tissue structure changes intracranially. Imaging at a single cell resolution in a real-time fashion provides supplementary dynamic information beyond that provided by standard end-point histological analysis, which looks solely at 'snap-shot' cross sections of tissue.
Establishing this intravital imaging technique in fluorescent chimeric mice, we are able to image four fluorescent channels simultaneously. By incorporating fluorescently labeled cells, such as GFP+ bone marrow, it is possible to track the fate of these cells studying their long-term migration, integration and differentiation within tissue. Further integration of a secondary reporter cell, such as an mCherry glioma tumor line, allows for characterization of cell:cell interactions. Structural changes in the tissue microenvironment can be highlighted through the addition of intra-vital dyes and antibodies, for example CD31 tagged antibodies and Dextran molecules.
Moreover, we describe the combination of our ICW imaging model with a small animal micro-irradiator that provides stereotactic irradiation, creating a platform through which the dynamic tissue changes that occur following the administration of ionizing irradiation can be assessed.
Current limitations of our model include penetrance of the microscope, which is limited to a depth of up to 900 &mu;m from the sub cortical surface, limiting imaging to the dorsal axis of the brain. The presence of the skull bone makes the ICW a more challenging technical procedure, compared to the more established and utilized chamber models currently used to study mammary tissue and fat pads 5-7. In addition, the ICW provides many challenges when optimizing the imaging.

A Novel High-resolution In vivo Imaging Technique to Study the Dynamic Response of Intracranial Structures to Tumor Growth and Therapeutics

We describe a novel in vivo imaging technique that couples fluorescent chimeric mice with intracranial windows and high-resolution 2-photon microscopy. This imaging platform aids studies of dynamic changes in brain tissue and microvasculature, at a single-cell level, following pathological insults and is adaptable to assess intracranial drug delivery and distribution.

A Novel Hochauflösende<em&gt; In vivo</em&gt; Bildtechnik, um die Dynamik der intrakranielle Strukturen zu Tumorwachstum und Therapeutics Studieren

We have successfully integrated previously established Intracranial window (ICW) technology 1-4 with intravital 2-photon confocal microscopy to develop a ...

Osmotic Drug Delivery to Ischemic Hindlimbs and Perfusion of Vasculature with Microfil for Micro-Computed Tomography Imaging

Preclinical research in animal models of peripheral arterial disease plays a vital role in testing the efficacy of therapeutic agents designed to stimulate microcirculation. The choice of delivery method for these agents is important because the route of administration profoundly affects the bioactivity and efficacy of these agents1,2. In this article, we demonstrate how to locally administer a substance in ischemic hindlimbs by using a catheterized osmotic pump. This pump can deliver a fixed volume of aqueous solution continuously for an allotted period of time. We also present our mouse model of unilateral hindlimb ischemia induced by ligation of the common femoral artery proximal to the origin of profunda femoris and epigastrica arteries in the left hindlimb. Lastly, we describe the in vivo cannulation and ligation of the infrarenal abdominal aorta and perfusion of the hindlimb vasculature with Microfil, a silicone radiopaque casting agent. Microfil can perfuse and fill the entire vascular bed (arterial and venous), and because we have ligated the major vascular conduit for exit, the agent can be retained in the vasculature for future ex vivo imaging with the use of small specimen micro-CT3.

We show here the in vivo insertion of an osmotic pump for constant local drug delivery and the creation of hindlimb ischemia in a mouse model. Moreover, the hindlimb vasculature is perfused with Microfil, a silicone radiopaque agent, to prepare for micro-computed tomography (micro-CT) imaging.

Osmotic Drug Delivery zu ischämischen Hinterbeine und Perfusion von Blutgefäßen mit Microfil für Mikro-Computertomographie

Preclinical research in animal models of peripheral arterial disease plays a vital role in testing the efficacy of therapeutic agents designed to ...

Bioluminescence Imaging of an Immunocompetent Animal Model for Glioblastoma

In contrast to commonly reported human glioma xenograft animal models, GL261 murine glioma xenografts recapitulate nearly all relevant clinical and histopathologic features of the human disease. When GL261 cells are implanted intracranially in syngeneic C57BL/6 mice, the model has the added advantage of maintaining an intact immune microenvironment. Stable expression of luciferase in GL261 cells allows non-invasive cost effective bioluminescence monitoring of intracranial tumor growth. We have recently demonstrated that luciferase expression in GL261 cells does not affect the tumor growth properties, tumor cell immunomodulatory cytokine expression, infiltration of immune cells into the tumor, or overall survival of animals bearing the intracranial tumor. Therefore, it appears that the GL261 luciferase glioma model can be useful in the study of novel chemotherapeutic and immunotherapeutic modalities. Here we report the technique for generating stable luciferase expression in GL261 cells and how to study the in vitro and in vivo growth of the tumor cells by bioluminescence imaging.

GL261 glioma cells provide a useful immunocompetent animal model of glioblastoma. The goals of this protocol are to demonstrate proper techniques for monitoring intracranial tumor growth using in vivo bioluminescence imaging, and to verify the utility of luciferase-modified GL261 cells for studying tumor immunology and immunotherapeutic approaches for treating glioblastoma.

Biolumineszenz-Bildgebung von einem immunkompetenten Tiermodell für Glioblastoma

In contrast to commonly reported human glioma xenograft animal models, GL261 murine glioma xenografts recapitulate nearly all relevant clinical and ...

Bioluminescence-Based Tumor Quantification Method for Monitoring Tumor Progression and Treatment Effects in Mouse Lymphoma Models

Although bioluminescence imaging (BLI) shows promise for monitoring tumor burden in animal models of cancer, these analyses remain mostly qualitative. Here we describe a method for bioluminescence imaging to obtain a semi-quantitative analysis of tumor burden and treatment response. This method is based on the calculation of a luminoscore, a value that allows comparisons of two animals from the same or different experiments. Current BLI instruments enable the calculation of this luminoscore, which relies mainly on the acquisition conditions (back and front acquisitions) and the drawing of the region of interest (manual markup around the mouse). Using two previously described mouse lymphoma models based on cell engraftment, we show that the luminoscore method can serve as a noninvasive way to verify successful tumor cell inoculation, monitor tumor burden, and evaluate the effects of in situ cancer treatment (CpG-DNA). Finally, we show that this method suits different experimental designs. We suggest that this method be used for early estimates of treatment response in preclinical small-animal studies.

Bioluminescence imaging is a well-known tool for localizing tumors and metastases, but quantification of these images often requires complex calculations and particular instruments. We describe the easy-to-use luminoscore method, based on precise acquisition conditions, requiring no calculations, and enabling tumor burden and treatment response to be monitored in mouse models.

Biolumineszenz-basierte Tumor Quantifizierung Verfahren zur Überwachung Tumorprogression und Therapieeffekte bei den Maus-Lymphom-Modelle

Although bioluminescence imaging (BLI) shows promise for monitoring tumor burden in animal models of cancer, these analyses remain mostly qualitative. ...

Combined Near-infrared Fluorescent Imaging and Micro-computed Tomography for Directly Visualizing Cerebral Thromboemboli

Direct thrombus imaging visualizes the root cause of thromboembolic infarction. Being able to image thrombus directly allows far better investigation of stroke than relying on indirect measurements, and will be a potent and robust vascular research tool. We use an optical imaging approach that labels thrombi with a molecular imaging thrombus marker&#160;&#8212; a Cy5.5 near-infrared fluorescent (NIRF) probe that is covalently linked to the fibrin strands of the thrombus by the fibrin-crosslinking enzymatic action of activated coagulation factor XIIIa during the process of clot maturation. A micro-computed tomography (microCT)-based approach uses thrombus-seeking gold nanoparticles (AuNPs) functionalized to target the major component of the clot: fibrin. This paper describes a detailed protocol for the combined in vivo microCT and ex vivo NIRF imaging of thromboemboli in a mouse model of embolic stroke. We show that in vivo microCT and fibrin-targeted glycol-chitosan AuNPs (fib-GC-AuNPs) can be used for visualizing both in situ thrombi and cerebral embolic thrombi. We also describe the use of in vivo microCT-based direct thrombus imaging to serially monitor the therapeutic effects of tissue plasminogen activator-mediated thrombolysis. After the last imaging session, we demonstrate by ex vivo NIRF imaging the extent and the distribution of residual thromboemboli in the brain. Finally, we describe quantitative image analyses of microCT and NIRF imaging data. The combined technique of direct thrombus imaging allows two independent methods of thrombus visualization to be compared: the area of thrombus-related fluorescent signal on ex vivo NIRF imaging vs. the volume of hyperdense microCT thrombi in vivo.

This protocol describes the application of combined near-infrared fluorescent (NIRF) imaging and micro-computed tomography (microCT) for visualizing cerebral thromboemboli. This technique allows the quantification of thrombus burden and evolution. The NIRF imaging technique visualizes fluorescently labeled thrombus in excised brain, while the microCT technique visualizes thrombus inside living animals using gold-nanoparticles.

Kombinierte Near-Infrarot-Fluoreszenz-Bildgebung und Mikrocomputertomographie für Direkt Visualizing Cerebral Thromboembolie

Direct thrombus imaging visualizes the root cause of thromboembolic infarction. Being able to image thrombus directly allows far better investigation of ...

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). Impaired uptake and metabolism of glucose in multiple organ systems due to insulin resistance is a hallmark of type 2 diabetes. The ability of this technique to extract an image-derived input function from the vena cava using an iterative deconvolution method eliminates the requirement of the collection of arterial blood samples. Fitting of tissue and vena cava time activity curves to a two-tissue, three compartment model permits the estimation of kinetic micro-parameters related to the 18F-FDG uptake from the plasma to the intracellular space, the rate of transport from intracellular space to plasma and the rate of 18F-FDG phosphorylation. This methodology allows for multiple measures of glucose uptake and metabolism kinetics in the context of longitudinal studies and also provides insights into the efficacy of therapeutic interventions.

Determining Glucose Metabolism Kinetics Using 18F-FDG Micro-PET/CT

This study describes a protocol that uses 18F-FDG and positron emission tomography/computed tomography (PET/CT) imaging, together with kinetic modelling, to quantify the in vivo, real-time uptake of 18F-FDG into tissues.

Die Bestimmung Glucose Metabolism Kinetics Verwendung<sup&gt; 18</sup&gt; F-FDG Micro-PET / CT

This paper describes the use of 18F-FDG and micro-PET/CT imaging to determine in vivo glucose metabolism kinetics in mice (and is transferable to rats). ...

Bioluminescence and Near-infrared Imaging of Optic Neuritis and Brain Inflammation in the EAE Model of Multiple Sclerosis in Mice

Experimental autoimmune encephalomyelitis (EAE) in SJL/J mice is a model for relapsing-remitting multiple sclerosis (RRMS). Clinical EAE scores describing motor function deficits are basic readouts of the immune-mediated inflammation of the spinal cord. However, scores and body weight do not allow for an in vivo assessment of brain inflammation and optic neuritis. The latter is an early and frequent manifestation in about 2/3 of MS patients. Here, we show methods for bioluminescence and near-infrared live imaging to assess EAE evoked optic neuritis, brain inflammation, and blood-brain barrier (BBB) disruption in living mice using an in vivo imaging system. A bioluminescent substrate activated by oxidases primarily showed optic neuritis. The signal was specific and allowed the visualization of medication effects and disease time courses, which paralleled the clinical scores. Pegylated fluorescent nanoparticles that remained within the vasculature for extended periods of time were used to assess the BBB integrity. Near-infrared imaging revealed a BBB leak at the peak of the disease. The signal was the strongest around the eyes. A near-infrared substrate for matrix metalloproteinases was used to assess EAE-evoked inflammation. Auto-fluorescence interfered with the signal, requiring spectral unmixing for quantification. Overall, bioluminescence imaging was a reliable method to assess EAE-associated optic neuritis and medication effects and was superior to the near-infrared techniques in terms of signal specificity, robustness, ease of quantification, and cost.

We show a technique for in vivo live bioluminescence and near-infrared imaging of optic neuritis and encephalitis in the experimental autoimmune encephalomyelitis (EAE) model for multiple sclerosis in SJL/J mice.

Biolumineszenz und Nah-Infrarot-Imaging von Neuritis und Gehirnentzündung im EAE-Modell der Multiplen Sklerose in Mäusen

Experimental autoimmune encephalomyelitis (EAE) in SJL/J mice is a model for relapsing-remitting multiple sclerosis (RRMS). Clinical EAE scores describing ...

Induction and Micro-CT Imaging of Cerebral Cavernous Malformations in Mouse Model

Mutations in the CCM1 (aka KRIT1), CCM2, or CCM3 (aka PDCD10) gene cause cerebral cavernous malformation (CCM) in humans. Mouse models of CCM disease have been established by tamoxifen induced deletion of Ccm genes in postnatal animals. These mouse models provide invaluable tools to investigate molecular mechanism and therapeutic approaches for CCM disease. An accurate and quantitative method to assess lesion burden and progression is essential to harness the full value of these animal models. Here, we demonstrate the induction of CCM disease in a mouse model and the use of the contrast enhanced X-ray micro computed tomography (micro-CT) method to measure CCM lesion burden in mouse brains. At postnatal day 1 (P1), we used 4-hydroxytamoxifen (4HT) to activate Cre recombinase activity from the Cdh5-CreErt2 transgene to cleave the floxed allele of Ccm2. CCM lesions in mouse brains were analyzed at P8. For micro-CT, iodine based Lugol&#39;s solution was used to enhance contrast in brain tissue. We have optimized the scan parameters and utilized a voxel dimension of 9.5 &#181;m, which lead to a minimum feature size of approximately 25 &#181;m. This resolution is sufficient to measure CCM lesion volume and number globally and accurately, and provide high-quality 3-D mapping of CCM lesions in mouse brains. This method enhances the value of the established mouse models to study the molecular basis and potential therapies for CCM and other cerebrovascular diseases.

This protocol demonstrates the induction of cerebral cavernous malformation disease in a mouse model and uses contrast enhanced micro computed tomography to measure lesion burden. This method enhances the value of established mouse models to study the molecular basis and potential therapies for cerebral cavernous malformation and other cerebrovascular diseases.

Induktion und Mikro-CT-Bildgebung des zerebralen höhlenartigen Fehlbildungen im Mausmodell

Mutations in the CCM1 (aka KRIT1), CCM2, or CCM3 (aka PDCD10) gene cause cerebral cavernous malformation (CCM) in humans. Mouse models of CCM disease have ...

Gene Regulation and Targeted Therapy in Gastric Cancer Peritoneal Metastasis: Radiological Findings from Dual Energy CT and PET/CT

Gastric cancer remains fourth in cancer incidence worldwide with a five-year survival of only 20%-30%. Peritoneal metastasis is the most frequent type of metastasis that accompanies unresectable gastric cancer and is a definitive determinant of prognosis. Preventing and controlling the development of peritoneal metastasis could play a role in helping to prolong the survival of gastric cancer patients. A non-invasive and efficient imaging technique will help us to identify the invasion and metastasis process of peritoneal metastasis and to monitor the changes in tumor nodules in response to treatments. This will enable us to obtain an accurate description of the development process and molecular mechanisms of gastric cancer. We have recently described experiment using dual energy CT (DECT) and positron emission tomography/computed tomography (PET/CT) platforms for the detection and monitoring of gastric tumor metastasis in nude mice models. We have shown that weekly continuous monitoring with DECT and PET/CT can identify dynamic changes in peritoneal metastasis. The sFRP1-overexpression in gastric cancer mice models showed positive radiological performance, a higher FDG uptake and increasing enhancement, and the SUVmax (standardized uptake value) of nodules demonstrated an obvious alteration trend in response to targeted therapy of TGF-&#946;1 inhibitor. In this article, we described the detailed non-invasive imaging procedures to conduct more complex research on gastric cancer peritoneal metastasis using animal models and provided representative imaging results. The use of non-invasive imaging techniques should enable us to better understand the mechanisms of tumorigenesis, monitor tumor growth, and evaluate the effect of therapeutic interventions for gastric cancer.

This protocol describes the value of dual energy CT and PET/CT imaging methods in tumor imaging and efficacy evaluation. This article demonstrates the research methods and results acquired by dual energy CT and PET/CT to evaluate the gene regulation and targeted treatment of gastric cancer peritoneal metastasis.

Genregulation und gezielten Therapie bei Magenkrebs peritonealen Metastasen: radiologische Befunde von Dual Energy CT und PET/CT

Gastric cancer remains fourth in cancer incidence worldwide with a five-year survival of only 20%-30%. Peritoneal metastasis is the most frequent type of ...