Cette étude a été financée par la Deutsche Forschungsgemeinschaft à SW (DFG WA 2804) et une bourse universitaire pour SW à partir du Centre de recherche expérimentale et clinique, une coopération du Centre Max Delbrück de médecine moléculaire et de la faculté de médecine Charité à Berlin. Les bailleurs de fonds n&#39;ont joué aucun rôle dans la conception de l&#39;étude, la collecte de données et d&#39;analyse, la décision de publier ou de la préparation du manuscrit. Nous remercions M. Robert Westphal pour le support technique lors de son stage dans notre laboratoire.

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Les auteurs déclarent n&#39;avoir aucun conflit d&#39;intérêts financiers.

Cette méthode d&#39;employer 19 F / 1 H IRM pour suivre le mouvement de DC dans le ganglion lymphatique donne l&#39;occasion d&#39;étudier les schémas de migration des cellules immunitaires in vivo. Les cellules dendritiques sont d&#39;excellents exemples de la migration rapide des cellules immunitaires qui sont capables de manœuvrer à travers des structures tridimensionnelles sans bien adhérentes à des substrats spécifiques 17. Bien que la faible résolution spatiale (?...

The tracking of cells in vivo is a crucial aspect in several fields of biomedicine. For this, noninvasive imaging techniques that can selectively localize cells over a period of time are extremely valuable. Prior to the development of three-dimensional magnetic resonance imaging (MRI), the tracking of immune cell migration was limited to microscopic analyses or tissue biopsies. Cell tracking with the help of MRI has gained immense attention in the past few years, not only for immunologists studying immune cell behavior in vivo, but also for clinical and stem cell researchers. During the mid-90s, the first studies on iron oxide nanoparticles 1 initiated a cascade of developments for tracking cells with MRI. Iron oxide particles shorten the MR relaxation time (T2*) of the labeled cells and thus cause signal depletion in MR images. Iron oxide particles have been employed to label macrophages 2, oligodendrocyte progenitors 3 and many other cell types. Some of these particles have also been clinically approved by the FDA for labeling cellular vaccines in melanoma patients 4. Since in vivo or ex vivo labeling of cells with iron oxide particles relies on a shortening of the T2* signal and the latter could be also brought about by in vivo susceptibility-related T2* effects such as micro bleeds, iron deposits or air bubbles, it might be difficult to identify labeled cells in vivo from other background T2* signal extinctions 5.
In this article, we describe a technique for tracking dendritic cells (DC) in vivo by employing 19F/1H magnetic resonance imaging (MRI). This cell tracking technology was only introduced in 2005 6, several years after the first recognized applications for 19F in MRI had been reported 7. One important advantage of 19F over iron oxide particle cell labeling is the low biological occurrence of 19F in tissue; this makes it possible to track cells very selectively with basically background-free images. Furthermore, it is possible to overlay the 19F MR signal from the transplanted labeled cells with anatomical images obtained from conventional 1H MRI. 19F/1H MRI is therefore considerably relevant for studies investigating cell migration in vivo. Cells studied with this method are labeled with 19F-rich particles. Synthetically-derived perfluorocarbons (PFCs) consisting primarily of carbon and fluorine atoms are commonly used to prepare the particles. These compounds are insoluble in water and need to be emulsified prior to application in vitro or in vivo. The usual size of the PFC particles that have been employed by other groups for in vivo 19F-MRI tracking experiments ranges between 100 nm and 245 nm 6,8-10. We have however shown that the efficiency in labeling dendritic cells with perfluoro-15-crown-5-ether (PFCE) particles increases with increasing particle size (&gt;560 nm).11

<table border="1"><tbody><tr><td></td><td></td><td></td><td> REACTIFS </td></tr><tr><td> Souris C57BL / 6 </td><td> Charles River, Berlin </td><td></td><td></td></tr><tr><td> RPMI </td><td> Gibco </td><td> 21875-091 </td><td></td></tr><tr><td> FBS supérieure </td><td> Biochrom AG </td><td> S 0615 </td><td></td></tr><tr><td> HEPES </td><td> Gibco Invitrogen- </td><td> 15630-056 </td><td></td></tr><tr><td> Pénicilline-streptomycine </td><td> Gibco </td><td> 15140-122 </td><td></td></tr><tr><td> L-glutamine </td><td> Gibco </td><td> 25030-024 </td><td></td></tr><tr><td> PBS Dulbecco </td><td> Sigma Aldrich </td><td> D8662 </td><td></td></tr><tr><td> PFA </td><td> Santa Cruz </td><td> sc-281692 </td><td></td></tr><tr><td> Perfluoro-15-couronne-5-éther </td><td> ChemPur </td><td> 391 -1996 </td><td></td></tr><tr><td> Pluronic F-68 </td><td> Sigma Aldrich </td><td> P5556 </td><td></td></tr><tr><td> Des boîtes de Pétri (35 x 10 mm) </td><td> VWR, Allemagne </td><td> 391-1996 </td><td></td></tr><tr><td> 27 ½ seringues G </td><td> VWR, Allemagne </td><td> 612-0151 </td><td></td></tr><tr><td> Tamis en nylon de cellules um (100 mesh) </td><td> VWR, Allemagne </td><td> 734-0004 </td><td></td></tr><tr><td> RMN tubes </td><td> VWR, Allemagne </td><td> 634-0461 </td><td></td></tr><tr><td></td><td></td><td></td><td> ÉQUIPEMENT </td></tr><tr><td> Outils de dissection </td><td> FST </td><td></td><td></td></tr><tr><td> Incubateur à CO 2 </td><td> Liant </td><td></td><td></td></tr><tr><td> Petit animal MR système </td><td> Bruker BioSpin </td><td></td><td> 9.4T BioSpec 94/20 USR,Acquisition ParaVision et de logiciels de traitement </td></tr><tr><td> 1 H / 19 F à double bobine RF accordable de volume </td><td> Rapid Biomed, Würzburg, Allemagne </td><td></td><td> 35 mm de diamètre intérieur, 50 mm de longueur </td></tr><tr><td> 19 bobine spectroscopie F </td><td> interne </td><td></td><td> air / match bobine en boucle, 4 tours, diamètre intérieur 5 mm, 10 mm de long, deux condensateurs pour le réglage et l&#39;appariement </td></tr><tr><td> Système d&#39;inhalation d&#39;isoflurane </td><td> Föhr Medical Instruments GmbH </td><td></td><td></td></tr><tr><td> Modèle du système de surveillance des animaux 1025 </td><td> SA Instruments Inc, New York, USA </td><td></td><td></td></tr></tbody></table>

monitoring dendritic cell migration using 19f / 1h magnetic resonance imaging

Progrès continus en matière modalités d&#39;imagerie non invasives comme l&#39;imagerie par résonance magnétique (IRM) ont grandement amélioré notre capacité à étudier les processus physiologiques ou pathologiques dans les organismes vivants. IRM s&#39;avère également être un outil précieux pour la capture des cellules transplantées in vivo. Les premières stratégies de marquage cellulaire pour utilisation d&#39;agents de contraste IRM qui influencent les temps de relaxation MR (T1, T2, T2 *) et conduire à une amélioration (T1) ou l&#39;épuisement (T2 *) du signal où les cellules marquées sont présentes. T2 agents d&#39;amélioration * tels que les agents ultrapetites d&#39;oxyde de fer (USPIO) ont été utilisés pour étudier la migration cellulaire et certains ont également été approuvé par la FDA pour une application clinique. Un inconvénient de T2 * agents est la difficulté de distinguer l&#39;extinction du signal créé par les cellules marquées d&#39;autres artefacts tels que des caillots de sang, des saignements ou des micro-bulles d&#39;air. Dans cet article, nous décrivons une nouvelle technique pour le suivi in vivo des cellules quisont basées sur le marquage des cellules avec du fluor (19 F)-particules riches. Ces particules sont préparées par émulsification d&#39;hydrocarbures perfluorés (PFC) et de composés ensuite utilisé pour marquer les cellules, qui peuvent ensuite être visualisées par 19 F IRM. Les avantages importants de PFC pour le suivi des cellules in vivo comprennent (i) l&#39;absence de carbone lié à 19 F in vivo, ce qui donne alors des images sans fond et complète de cellules selectivityand (ii) la possibilité de quantifier le signal de la cellule de 19 F spectroscopie MR .

Dix-huit à vingt et un heures suivant l&#39;application intradermique, 19 F-étiquetés cellules dendritiques (DC) migrent dans le ganglion lymphatique poplité drainant. Le mouvement des DC via les canaux lymphatiques dans le ganglion drainant politeal peut être apprécié par superposition des images 1 H anatomiques avec les 19 images DC F (figure 2A). Nous avons déjà fait état ​​de la migration de ces cellules in vivo, ainsi que l&#39;impact de la <s...

Watch this Scientific Journal Video about Monitoring Dendritic Cell Migration using 19F / 1H Magnetic Resonance Imaging at JoVE.com

Monitoring Dendritic Cell Migration using 19F / 1H Magnetic Resonance Imaging

Suivi des cellules en utilisant l&#39;IRM a attiré l&#39;attention remarquable dans les dernières années. Ce protocole décrit le marquage des cellules dendritiques avec le fluor (<sup&gt; 19</sup&gt; Particules F)-riches, l&#39;application in vivo de ces cellules, et la surveillance de l&#39;étendue de leur migration vers le ganglion drainant avec<sup&gt; 19</sup&gt; F /<sup&gt; 1</sup&gt; H IRM et<sup&gt; 19</sup&gt; F MRS.

La migration des cellules dendritiques de surveillance utilisant l&#39;imagerie par résonance 19F / 1H magnétique

Continuous  advancements in noninvasive imaging modalities such as magnetic resonance  imaging (MRI) have greatly improved our ability to study ...

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Immunology and Infection

Monitoring Dendritic Cell Migration using 19F / 1H Magnetic Resonance Imaging

11.7K vues.  A joint cooperation between the Charité Medical Faculty and the Max Delbrück Center for Molecular Medicine. Suivi des cellules en utilisant l'IRM a attiré l'attention remarquable dans les dernières années. Ce protocole décrit le marquage des cellules dendritiques avec le fluor ( 19 Particules F)-riches, l'application in vivo de ces cellules, et la surveillance de l'étendue de leur migration vers le ganglion drainant avec 19 F / 1 H IRM et 19 F MRS.