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WIREs Nanomed Nanobiotechnol
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Tracking and evaluation of dendritic cell migration by cellular magnetic resonance imaging

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Cellular magnetic resonance imaging (MRI) is a means by which cells labeled ex vivo with a contrast agent can be detected and tracked over time in vivo. This technology provides a noninvasive method with which to assess cell‐based therapies in vivo. Dendritic cell (DC)‐based vaccines are a promising cancer immunotherapy, but its success is highly dependent on the injected DC migrating to a secondary lymphoid organ such as a nearby lymph node. There the DC can interact with T cells to elicit a tumor‐specific immune response. It is important to verify DC migration in vivo using a noninvasive imaging modality, such as cellular MRI, so that important information regarding the anatomical location and persistence of the injected DC in a targeted lymph node can be provided. An understanding of DC biology is critical in ascertaining how to label DC with sufficient contrast agent to render them detectable by MRI. While iron oxide nanoparticles provide the best sensitivity for detection of DC in vivo, a clinical grade iron oxide agent is not currently available. A clinical grade 19Fluorine‐based perfluorcarbon nanoemulsion is available but is less sensitive, and its utility to detect DC migration in humans remains to be demonstrated using clinical scanners presently available. The ability to quantitatively track DC migration in vivo can provide important information as to whether different DC maturation and activation protocols result in improved DC migration efficiency which will determine the vaccine's immunogenicity and ultimately the tumor immunotherapy's outcome in humans. WIREs Nanomed Nanobiotechnol 2013. doi: 10.1002/wnan.1227 This article is categorized under: Diagnostic Tools > In Vivo Nanodiagnostics and Imaging Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials

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Monocyte‐derived dendritic cell (DC) can be labeled and then matured in vitro for use in both preclinical and clinical cellular magnetic resonance imaging (MRI) studies. Hematopoietic precursors isolated from the bone marrow of mice or monocytes from the peripheral blood of humans when treated with granulocyte‐macrophage colony stimulating factor (GM‐CSF) and interleukin 4 (IL‐4) differentiate into immature DC. These cells can then be incubated with MRI contrast agents (e.g., iron oxide nanoparticles, Fe; or perfluorocarbon emulsions, 19 F) before further differentiation to mature DC by incubation with a cocktail of maturation cytokines. Mature labeled DCs can be injected into either (I) animal models (area of DC‐associated superparamagnetic iron oxide (SPIO)‐induced signal loss has been pseudo‐color red at the flank injection site and in the draining inguinal lymph node (inset) or (II) human subjects to enable the visualization of DC migration by cellular MRI (Ia, IIa–c). (a) Gradient echo image before vaccination showing a right inguinal lymph node with a hyperintense signal (arrow). (b) Spin echo (an MR image sequence much less sensitive for SPIO) image obtained from the same lymph node (arrow) after vaccination showing an area of signal void. (c) Gradient echo image after vaccination in same position as (b) showing a more pronounced decrease in signal intensity in the lymph node (arrow). (Panels IIa–c: Reprinted with permission from Ref . Copyright 2005 Nature Publishing Group)
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Impact of 19F‐perfluorocarbon Cell Sense on mouse dendritic cell (DC) maturation and phenotype. Bone marrow‐derived DC cultures were prepared and enriched as per usual and then labeled overnight with 4.5 mg/mL of Cell Sense. Labeling CD11c+ DC with 19F Cell Sense does not affect CD11c+DC maturation (a) or the phenotype of mature CD11c+CD86+ (b) or immature CD11c+CD86 DC (c) with respect to cell surface proteins involved in antigen uptake (CD36), antigen presentation and co‐stimulation [major histocompatibility complex (MHC) I, MHC II, CD86, CD80, and CD54]. However, DC matured in the presence of Cell Sense did exhibit a small but significant reduction in the percentage of CD11c+CD86+ DC expressing CD40, a DC activation marker, and CCR7, a chemokine receptor that chemotactically directs DC to secondary lymphoid tissues. Expression of CD38, which is necessary for the activation of CCR7, was not affected by Cell Sense.
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Labeling immature dendritic cells (DC) with superparamagnetic iron oxide (SPIO) (FeREX) does not affect immature bone marrow‐derived DC transduction by a lentiviral vector or, following maturation, alter their migration to a lymph node. (a) The gating strategy is illustrated that defines the CD11c+CD86+ mature DC population (i) and the CD11c+CD86 immature DC (ii). (b) SPIO does not affect the frequency of mature and immature DC following transduction before or after lentiviral vector transduction. (c, d) The percentage of EGFP+ mature and immature DCs is independent of whether DCs are transduced before or after SPIO loading. (E) Either 3 × 105 or 1 × 106 100%‐SPIO‐labeled mature DCs were injected into the footpad of two groups of four C57BL/6 mice and MRI conducted 2 days later. Similarly, equal numbers of SPIO‐loaded and then lentivirus‐transduced DCs were injected into the contralateral hind footpad to serve as a control. Two days later the mice were MR imaged. There were no differences observed for (f) lymph node volume, (g) signal void volume and (h) fractional signal loss as compared to SPIO‐labeled DCs that were lentiviral transduced as compared with control. Scale bar = 1 mm.
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Mouse bone marrow‐derived dendritic cells (DCs) are not labeled efficiently with USPIO as compared to those labeled with superparamagnetic iron oxide (SPIO) nanoparticles. (a) Mouse bone marrow‐derived DCs were prepared as previously described and following enrichment on day 4 were labeled overnight (for 20 h) with either 200 µg/mL of SPIO (FeREX®) or 400 µg/mL of ultrasmall superparamagnetic iron oxide (USPIO) (Feraheme). (a) A portion of the DC culture was collected on day 5 to determine SPIO labeling efficiency compared to USPIO in immature DC. Some USPIO‐treated DCs were left in culture and matured along with protamine sulfate and more USPIO (400 µg/mL). These DCs were left to culture for an additional 20 hours. On day 6, efficiency of USPIO labeling was determined in mature DC by magnetic separation. The efficiency of USPIO uptake by immature and mature DCs was significantly less when compared to the labeling efficiency of SPIO. After 24 h of maturation DC tended to have a further reduction in USPIO content and this was not improved by relabeling. (b,c) For MRI studies, bone marrow‐derived DC cultures were prepared and DCs were enriched on day 4 as usual. Cells were labeled overnight with either 400 µg/mL of USPIO or 200 µg/mL of SPIO and concurrently matured as above in vitro. Some cells were matured but were left untreated (UT; no USPIO or SPIO) to serve as a control. On day 5, cells were collected and magnetically separated to select for populations of either 100% USPIO‐labeled or 100% SPIO‐labeled mature DC. Either 4 × 105 (b) or 1 × 106 (c) 100%‐USPIO labeled mature DCs were injected into the right hind footpad of C57 BL/6 mice (N = 2, n = 3). (d) Some mice were administered 1 × 106 100%‐SPIO‐labeled mature DC. Equal numbers of untreated mature DCs were injected into the contralateral left hind footpads of each mouse to serve as a control. No signal loss was observed in any popliteal lymph nodes of mice that received either dose of USPIO‐labeled mature DC (b.c). However, signal loss was readily detectable (blue arrow) in the popliteal lymph nodes of mice that received SPIO‐labeled DC (d). Lymph nodes are outlined in red. Spatial resolution of MRI is 100 × 100 × 200 µm.
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