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

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 (M phi s) 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 M phi s in a cutaneous granuloma model. Methods: M phi s were labeled with a membrane-selective carbocyanine dye (1,1-dioctadecyl-3,3,3,3- tetramethylindotricarbocyanine iodide [DiR]). 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 M phi s 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 M phi s 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 M phi s to different body compartments. The lipopoly-saccharide-containing pellets exhibited significantly higher SNRs than did pellets without lipopolysaccharide. FMT showed that M phi s distributed mainly in the periphery of the pellets. The cellular infiltrates extracted from the harvested pellets revealed the presence of approximately 10%-23% DiR-positive M phi s-expressing typical markers, confirming the transendothelial migration of injected M phi s. Conclusion: The tagging of Mfs with DiR allows the noninvasive tracking of inflammatory cells for several days in vivo. FRI and FMT are versatile techniques to monitor and quantify cellular inflammatory responses in vivo.