Tracking immune cells in vivo using magnetic resonance imaging

Abstract

The increasing complexity of in vivo imaging technologies, coupled with the development of cell therapies, has fuelled a revolution in immune cell tracking in vivo. Powerful magnetic resonance imaging (MRI) methods are now being developed that use iron oxide- and ¹⁹F-based probes. These MRI technologies can be used for image-guided immune cell delivery and for the visualization of immune cell homing and engraftment, inflammation, cell physiology and gene expression. MRI-based cell tracking is now also being applied to evaluate therapeutics that modulate endogenous immune cell recruitment and to monitor emerging cellular immunotherapies. These recent uses show that MRI has the potential to be developed in many applications to follow the fate of immune cells in vivo.

Figure 1. Schematic showing ex vivo and in situ labelling of cells with magnetic resonance nanoparticle contrast agents

Cell labelling can be achieved ex vivo in pre-selected and cultured cells by adding the labelling reagent — superparamagnetic iron oxide (SPIO) nanoparticles or perfluorocarbon (PFC) emulsion — directly to the media followed by co-incubation. Often the labelling reagent is complexed with a cationic transfection agent before it is added to the cell culture to label non-phagocytic cells. Self-delivering formulation s of PFC emulsions have also been devised that do not require a transfection agent. Alternatively, electroporation can be used to label cells in culture. After collection and washing, labelled cells are then administered to the subject. In addition, the labelling agent can be intravenously injected; in this in situ labelling approach, the agent is intrinsically taken up by phagocytic cells of the reticuloendothelial (RES) system, particularly by monocytes and macrophages, which then accumulate at sites of inflammation. For both ex vivo and in situ labelling approaches an ¹H magnetic resonance imaging (MRI) scan is then carried out to visualize the anatomy. The resulting images have a decreased signal in regions containing SPIO-labelled cells (known as negative contrast). In the case of PFC probes, a ¹⁹F MRI scan is also acquired in the same imaging session. A composite, pseudo-coloured ¹⁹F and ¹H image is then constructed, in which labelled cells appear in the ¹⁹F colour channel (known as hot spot contrast). By quantifying the ¹⁹F MRI signal in individual organs using using nuclear magnetic resonance (NMR) spectroscopy an inflammation index can be calculated, which is a direct correlation between the ¹⁹F signal and the number of labelled macrophages.

Figure 2. The development of MRI reporter genes

Certain nucleic acid-based reporters encode cell surface receptors that bind to specific ligands that are conjugated to superparamagnetic iron oxide (SPIO) to render them magnetic resonance imaging (MRI)-detectable. They require delivery of the ligand–SPIO complex as an exogenous substrate. An example of this approach is an engineered transferrin receptor that binds transferrin-ligated SPIO. Another exogenous substrate-based approach involves a reporter gene encoding the thymidine kinase enzyme derived from herpes simplex virus (HSV). This enzyme phosphorylates thymidine analogues, which causes them to remain trapped in transduced cells. When thymidine analogues are used that are rich in specific (imino) protons, chemical exchange saturation transfer (CEST) MRI can be used to track the transduced cells. Other approaches use endogenous substrates, whereby the encoded reporter binds iron (Fe²⁺) that is naturally present in the body as the contrasting metal ion. Finally, a CEST reporter (such as lysine-rich protein (LRP)) can be used that does not require a substrate, as the protein itself contains multiple labile amide protons that can be saturated and detected.

Figure 3. Tracking immune cells with MRI using SPIO nanoparticles and PFC emulsions

a | Intracellular labelling of mononuclear cells with magnetoliposomes is shown. The electron micrograph shows superparamagnetic iron oxide (SPIO) particles in secondary lysosomes (small arrows) and in primary lysosomes that are fusing with endosomes (large arrow). The scale bar represents 200 nm. b | Imaging of a non-obese diabetic (NOD) severe combined immunodeficient (SCID) mouse pancreas ex vivo is shown following the adoptive transfer of SPIO-labelled T cells. Image hypointensities represent infiltrating T cells that are observed at 24 hours post transfer. c | Imaging of in vivo antigen capture and trafficking of dendritic cells (DCs) is shown. Sentinel DCs were labelled in situ by intradermal injection of unlabelled (dashed arrow) or SPIO-labelled (solid arrow) irradiated cancer cells, which function as a vaccine. Following phagocytosis of both SPIO particles and tumour antigens in a process known as magnetovaccination, the hypointense DCs migrate into the medulla of the draining popliteal lymph node, as observed on day 8. d | An electron micrograph of a perfluorocarbon (PFC)-labelled DC is shown. Numerous bright spots (PFC droplets) are observed inside the cell. Particles appear as smooth spheroids. Arrowheads indicate vesicles. The scale bar represents 200 nm. e | Inflammatory bowel disease (IBD) in an interleukin-10 (Il10)^−/− mouse model was visualized using in situ PFC labelling and ¹⁹F magnetic resonance imaging (MRI) in vivo. The ¹⁹F image (pseudo-colour) is shown on the far left, the composite ¹H and ¹⁹F image is shown in the middle and a three-dimensional rendering of the in vivo ¹⁹F MRI data from the abdomen is shown on the far right. A reference tube that contains PFC emulsion (R) was placed alongside the torso of the mouse. The images through the abdomen show PFC accumulation (indicating inflammation) in the ascending colon (ac) and descending colon (dc), where (a) is the anus. Part a is reproduced, with permission, from REF. © (1993) John Wiley and Sons. Part b is reproduced, with permission, from REF. © (2002) John Wiley and Sons. Part c is reproduced, with permission, from REF. © (2009) American Association for Cancer Research. Part d is reproduced, with permission, from REF. © (2005) Macmillan Publishers Ltd. All rights reserved. Part e is reproduced, with permission, from REF. © (2012) Macmillan Publishers Ltd. All rights reserved.