Name: bioluminescence and near-infrared imaging of optic neuritis and brain inflammation in the eae model of multiple sclerosis in mice
Uploaded: 2017-03-01T10:10:00
Description: Watch this Scientific Journal Video about Bioluminescence and Near-infrared Imaging of Optic Neuritis and Brain Inflammation in the EAE Model of Multiple Sclerosis in Mice at JoVE.com

Diese Arbeit wurde von der Deutschen Forschungsgemeinschaft (CRC1039 A3) und der Forschungsförderungsprogramm &quot;Landesoffensive zur Entwicklung wissenschaftlich-ökonomischer Exzellenz&quot; (LOEWE) des Landes Hessen, Forschungszentrum für Translationale Medizin und Pharmakologie TMP und der Else Kröner-Fresenius-Stiftung (EKFS), Graduiertenkolleg Translationale Forschung Innovation - Pharma (TRIP).

1. EAE Induktion in SJL / J-Mäuse <ol><li> Mäuse <ol><li> Verwenden Sie 11-Wochen alte weibliche SJL / J-Mäuse und es ihnen ermöglichen, für ca. 7 Tage zur experimentellen Raum zu gewöhnen. Verwenden Sie n = 10 Mäuse pro Gruppe. </li><li> Für die Beurteilung der Arzneimittelwirkungen, die Verwaltung der Drogen-und Placebo für die Kontrollgruppe kontinuierlich über das Trinkwasser oder über Futterpellets beginnend 3 oder 5 Tage nach der Immunisierung (n = 10 pro Gruppe). Während der Höhepunkt der ...

<ol>
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Die vorliegende Video zeigt Techniken zur Biolumineszenz und Nah-Infrarot - Fluoreszenz in vivo Bildgebung von EAE in SJL / J - Mäusen. Wir zeigen, dass Biolumineszenz-Bildgebung eine entzündungs ​​empfindliche Sonde zeigt hauptsächlich optische Neuritis verwenden, und die Quantifizierung stimmt mit der klinischen Bewertung von EAE Schwere und die Auswirkungen von Medikamenten. Jedoch war die Biolumineszenz - Bildgebungsverfahren nicht in der Lage Entzündung des lumbalen Rückenmarks zu erkennen, die ei...

Multiple sclerosis is caused by the autoimmune-mediated attack and destruction of the myelin sheath in the brain and the spinal cord1. With an overall incidence of about 3.6 cases per 100,000 people a year in women and about 2.0 in men, MS is the second most common cause of neurological disability in young adults, after traumatic injuries2,3. The disease pathology is contributed to by genetic and environmental factors4 but is still not completely understood. Autoreactive T lymphocytes enter the central nervous system and trigger an inflammatory cascade that causes focal infiltrates in the white matter of the brain, spinal cord, and optic nerve. In most cases, these infiltrates are initially reversible, but persistence increases with the number of relapses. A number of rodent models have been developed to study the pathology of the disease. The relapsing-remitting EAE in SJL/J mice and the primary-progressive EAE in C57BL6 mice are the most popular models.
The clinical EAE scores, which describe the extent of the motor function deficits, and body weight are the gold standards to assess EAE severity. These clinical signs agree with the extent of immune cell infiltration and myelin destruction in the spinal cord and moderately predict drug treatment efficacy in humans5. However, these signs mainly reflect the destruction of the ventral fiber tracts in the spinal cord. Presently, there is no easy, non-invasive, reliable, and reproducible method to assess in vivo brain infiltration and optic neuritis in living mice. 
The in vivo imaging agrees with the 3 "R" principles of Russel and Burch (1959), which claim a Replacement, Reduction, and Refinement of animal experiments6, because imaging increases the readouts of one animal at several time points and allows for a reduction of the overall numbers. Presently, inflammation or myelin status is mainly assessed ex vivo via immunohistochemistry, FACS-analysis, or different molecular biological methods7, all requiring euthanized mice at specific time points. 
A number of in vivo imaging system probes have been developed to assess inflammation in the skin, joints, and vascular system. The techniques rely on the activation of bioluminescent or near-infrared fluorescent substrates by tissue peroxidases, including myeloperoxidase (MPO), matrix metalloproteinases (MMPs)8, and cathepsins9 or cyclooxygenase2. These probes have been mainly validated in models of arthritis or atherosclerosis9,10. A cathepsin-sensitive probe has also been used for fluorescence molecular tomographic imaging of EAE11. MMPs, particularly MMP2 and MMP9, contribute to the protease-mediated BBB disruption in EAE and are upregulated at sites of immune cell infiltration12, suggesting that these probes may be useful for EAE imaging. The same holds true for peroxidase or cathepsin-based probes. Technically, imaging of inflammation in the brain or spinal cord is substantially more challenging because the skull or spine absorb bioluminescent and near-infrared signals. 
In addition to inflammation indicators, fluorescent chemicals have been described, which specifically bind to myelin and may allow for quantification of myelination13. A near-infrared fluorescent probe, 3,3'-diethylthiatricarbocyanine iodide (DBT), was found to specifically bind to myelinated fibers and was validated as a quantitative tool in mouse models of primary myelination defects and in cuprizone-evoked demyelination14. In EAE, the DBT signal was rather increased, reflecting the inflammation of the myelin fibers5. 
An additional hallmark of EAE and MS is the BBB breakdown, resulting in increased vascular permeability and the extravasation of blood cells, extracellular fluid, and macromolecules into the CNS parenchyma. This can lead to edema, inflammation, oligodendrocyte damage, and, eventually, demyelination15,16. Hence, visualization of the BBB leak using fluorescent probes, such as fluorochrome-labeled bovine serum albumin5, which normally distribute very slowly from blood to tissue, may be useful to assess EAE.
In the present study, we have assessed the usefulness of different probes in EAE and show the procedure for the most reliable and robust bioluminescent technique. In addition, we discuss the pros and cons of near-infrared probes for MMP activity and BBB integrity.

<table border="0" cellpadding="0" cellspacing="0" width="1287">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">AngioSpark-680</td>
			<td style="width:267px;">Perkin Elmer, Inc., Waltham, USA</td>
			<td style="width:119px;">NEV10149</td>
			<td style="width:636px;">Imaging probe, pegylated nanoparticles, useful for imaging of blood brain barrier integrity</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MMP-sense 680</td>
			<td>Perkin Elmer, Inc., Waltham, USA</td>
			<td>NEV10126</td>
			<td>Imaging probe, activatable by matrix metalloproteinases, useful for imaging of inflammation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">XenoLight RediJect Inflammation Probe</td>
			<td>Perkin Elmer, Inc., Waltham, USA</td>
			<td>760535</td>
			<td>Imaging probe, activatable by oxidases, useful for imaging of inflammation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">PLP139-151/CFA emulsion&#160;</td>
			<td>Hooke Labs, St Lawrence, MA</td>
			<td>EK-0123</td>
			<td>EAE induction kit</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Pertussis Toxin</td>
			<td>Hooke Labs, St Lawrence, MA</td>
			<td>EK-0123</td>
			<td>EAE induction kit</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">IVIS Lumina Spectrum</td>
			<td>Perkin Elmer, Inc., Waltham, USA</td>
			<td style="width:119px;"></td>
			<td>Bioluminescence and Infrared Imaging System</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">LivingImage 4.5 software&#160;</td>
			<td>Perkin Elmer, Inc., Waltham, USA</td>
			<td>CLS136334</td>
			<td>IVIS analysis software</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Isoflurane</td>
			<td style="width:267px;">Abbott Labs, Illinois, USA</td>
			<td>26675-46-7</td>
			<td>Anaesthetic</td>
		</tr>
	</tbody>
</table>

bioluminescence and near-infrared imaging of optic neuritis and brain inflammation in the eae model of multiple sclerosis in mice

Experimentelle Autoimmun-Enzephalomyelitis (EAE) in SJL / J-Mäusen ist ein Modell für rezidivierend-remittierender Multipler Sklerose (RRMS). Klinische EAE Scores Motorik Defizite beschreiben, sind Grund Auslesungen der immunvermittelte Entzündung des Rückenmarks. Allerdings Gewicht Scores und Körper nicht erlauben , für eine in vivo Beurteilung der Entzündung des Gehirns und Neuritis. Letzteres ist eine frühe und häufige Erscheinung in etwa 2/3 MS-Patienten. Hier zeigen wir Methoden für die Biolumineszenz und Nah-Infrarot - Live - Bildgebung zur Beurteilung EAE Neuritis hervorgerufen, Gehirnentzündung und Blut-Hirn - Schranke (BHS) Störung in lebenden Mäusen unter Verwendung eines in vivo - Bildgebungssystem. Ein Biolumineszenz-Substrat aktiviert durch Oxidasen zeigte in erster Linie optische Neuritis. Das Signal war spezifisch und erlaubt die Visualisierung von Arzneimittelwirkungen und Krankheitszeitkurse, die die klinischen Bewertungen einher. Pegyliertem fluoreszierende Nanopartikel, die innerhalb der vasculatur bliebe für längere Zeiträume verwendet wurden, die BBB Integrität zu beurteilen. Nah-Infrarot-Bildgebung ergab einen BBB-Leck auf dem Höhepunkt der Krankheit. Das Signal war das stärkste um die Augen. Ein Nahinfrarot-Substrat für Matrixmetalloproteinasen wurde verwendet EAE-evozierte Entzündung zu beurteilen. Auto-Fluoreszenz mischte sich mit dem Signal, erfordern Entmischung zur Quantifizierung. Insgesamt war Biolumineszenz-Bildgebung eine zuverlässige Methode EAE-assoziierte optische Neuritis und Medikamente Effekte und war besser als die Nah-Infrarot-Techniken in Bezug auf Signal Spezifität, Robustheit, einfache Quantifizierung und Kosten zu bewerten.

Zeitverlauf der Biolumineszenz von Neuritis Das Biolumineszenz-Signal der Entzündung Sonde war das stärkste um die Augen und trat ausschließlich in EAE Mäuse mit Optikusneuritis. Ein Signal, trat bei keiner der nicht-EAE Mäuse noch die Mäuse nicht mit der Entzündung Sonde injiziert. Das Signal verschwunden, wenn die Mäuse gewonnen. Daher ist das Signal für Optikusneuritis spezifisch und die Spitze des...

Watch this Scientific Journal Video about Bioluminescence and Near-infrared Imaging of Optic Neuritis and Brain Inflammation in the EAE Model of Multiple Sclerosis in Mice at JoVE.com

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

Wir zeigen eine Technik für die in vivo lebenden Biolumineszenz und Nahinfrarot-Bildgebung von Optikusneuritis und Enzephalitis in der experimentellen autoimmunen Enzephalomyelitis (EAE) -Modell für Multiple Sklerose in SJL / J - Mäusen.

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 ...

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

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRI&#8212;Application in Premanifest Huntington's Disease

Whole-brain Segmentation and Change-point Analysis of Anatomical Brain MRIÃƒÆ’Ã‚Â¢ÃƒÂ¢Ã¢â‚¬Å¡Ã‚Â¬ÃƒÂ¢Ã¢â€šÂ¬Ã‚ÂApplication in Premanifest Huntington's Disease

Gesamte Gehirn Segmentierung und Change-Point-Analyse der anatomischen Gehirn MRI-Anwendung in Prämanifesten Huntington-Krankheit

Recent advances in MRI offer a variety of useful markers to identify neurodegenerative diseases. In Huntington&#39;s disease (HD), regional brain atrophy ...