In Vivo Optical Imaging of Cellular Inflammatory Response in Granuloma Formation Using Fluorescence-Labeled Macrophages

Abstract

Near-infrared imaging such as fluorescence reflectance imaging (FRI) and fluorescence-mediated tomography (FMT) yields high signal-to-noise ratios (SNRs) and should thus be well suited for cell-tracking studies. Extravasation of monocytes or macrophages (Mfs) is one of the earliest events in inflammation. The purpose of this study was to assess whether FRI and FMT allow for the visualization and quantification of early inflammatory processes by tracing the migration of fluorescence-labeled murine Mfs in a cutaneous granuloma model. Methods: Mfs were labeled with a membrane-selective carbocyanine dye (1,3,3,). Cellular viability and function (nitric oxide production, phagocytosis, adherence) were assessed in vitro. Local inflammation was induced in mice by the subcutaneous injection of polyacrylamide gel pellets including or excluding a strong inflammatory stimulus (lipopolysaccharide). Labeled Mfs were injected intravenously, and FRI and FMT were performed up to 7 d. SNRs were calculated for the pellets, and the 3-dimensional distribution of Mfs was assessed using FMT. Cells were harvested from gel pellets and analyzed by flow cytometry. Results: DiR labeling did not affect cell viability or cell function. FRI revealed the migration of labeled Mfs into gel pellets and the homing of Mfs to different body compartments. The lipopolysaccharide-containing pellets exhibited significantly higher SNRs than did pellets without lipopolysaccharide. FMT showed that Mfs distributed mainly in the periphery of the pellets. Opt ical imaging is an appealing concept for studying cell migration in vivo. Specifically, in the near-infrared range (NIR) background fluorescence is minimal, yielding excellent signal-to-noise ratios (SNRs) and thus high sensitivity of the technique for detecting even small amounts of cells. Further, fluorophores emitting in the NIR can sufficiently penetrate the tissue, even from deeper sections, and allow detection over a longer time (1). Moreover, imaging techniques for optical imaging are rapidly developing, ranging from simple surface-weighted reflection (fluorescence reflectance imaging [FRI]) to fluorescence-mediated tomography (FMT) approaches. The latter technology is capable of delivering quantitative information on 3-dimensional fluorochrome distribution in vivo.The recruitment of immune cells and especially monocytes from the blood into tissue is crucial for the development and maintenance of inflammation (2). Interference with the mechanisms of monocyte extravasation represents an emerging new therapeutic strategy (3,4). After extravasation, monocytes can differentiate into macrophages and dendritic cells. They are crucial for nearly every step of an immune reaction including the initiation of inflammation, clearance of infectious agents or tumor cells, initiation of an adaptive immune response, and resolution of inflammation (5,6).Thus, the noninvasive tracing and monitoring of monocytes or macrophages (Mfs) in vivo by optical imaging could allow for the better localization, visualization, and

Extracellular proteolysis in macrophage migration: Losing grip for a breakthrough

“…For the moment, despite the development of new probes to label macrophages for intravital live-animal imaging approaches [62][63][64][65], the migration mode used by macrophages and the contribution of proteases has not been investigated. When neutrophils and/or monocytes/macrophages infiltrate tissues, the presence of MMPs is detected but their role in the migration process has not been studied [66][67][68][69][70]. As cell morphology is indicative of the migration mode (amoeboid or mesenchymal) used by the migrating cells and therefore predictive of the possible involvement of proteases, we have examined this point in published in vivo imaging experiments [71,72].…”

“…When neutrophils and/or monocytes/macrophages infiltrate tissues, the presence of MMPs is detected but their role in the migration process has not been studied [66][67][68][69][70]. As cell morphology is indicative of the migration mode (amoeboid or mesenchymal) used by the migrating cells and therefore predictive of the possible involvement of proteases, we have examined this point in published in vivo imaging experiments [71,72].…”

Reprogramming of Monocytes by GM-CSF Contributes to Regulatory Immune Functions during Intestinal Inflammation

“…For in vivo cell tracking of GMaMs, cells were stained with a commercially available lipophilic tracer 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbocyanine iodide (Life Technologies, Darmstadt, Germany) with an emission maximum of 782 nm as described elsewhere (35). In vivo distribution of labeled cells across the intestine was studied 24 h after i.v.…”

“…In vivo distribution of labeled cells across the intestine was studied 24 h after i.v. injection using a planar smallanimal FMT system (FMT 2500; VisEn Medical) as described previously (35).…”

Trafficking Macrophage Migration Using Reporter Gene Imaging with Human Sodium Iodide Symporter in Animal Models of Inflammation

“…In the era of molecular imaging, optical imaging using enhanced GFP and Fluc genes (17,18) and 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbocyanine iodide cell tracker dyes (19), MRI using superparamagnetic iron oxide (20,21), and CT using a nanoparticulate contrast agent (22) also have been applied for macrophage trafficking.…”