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Summary The fungus Aspergillus fumigatus is ubiquitous in nature and the most common cause of invasive pulmonary aspergillosis (IPA) in patients with a compromised immune system. The development of IPA in patients under immunosuppressive treatment or in patients with primary immunodeficiency demonstrates the importance of the host immune response in controlling aspergillosis. However, study of the host-microbe interaction has been hampered by the lack of tools for their non-invasive assessment. We developed a methodology to study the response of the host's immune system against IPA longitudinally in vivo by using fluorine-19 magnetic resonance imaging (19F MRI). We showed the advantage of a perfluorocarbon-based contrast agent for the in vivo labeling of macrophages and dendritic cells, permitting quantification of pulmonary inflammation in different murine IPA models. Our findings reveal the potential of 19F MRI for the assessment of rapid kinetics of innate immune response against IPA and the permissive niche generated through immunosuppression., Graphical Abstract, Highlights • Host-pathogen immune response is visualized in vivo and quantified against IPA • Modified PFC-based nanoparticles were used for in vivo labeling of immune cells • Clinical immunosuppression depict dynamic immune response upon fungal challenge • 19F MRI showed follow-up of labeled immune cells in individual animals over time, Infection Control in Health Technology; Medical Imaging; Mycology

Ultrasound is the most commonly used clinical imaging modality. However, in applications requiring cell-labeling, the large size and short active lifetime of ultrasound contrast agents limit their longitudinal use. Here, 100 nm radius, clinically applicable, polymeric nanoparticles containing a liquid perfluorocarbon, which enhance ultrasound contrast during repeated ultrasound imaging over the course of at least 48 h, are described. The perfluorocarbon enables monitoring the nanoparticles with quantitative 19 F magnetic resonance imaging, making these particles effective multimodal imaging agents. Unlike typical core–shell perfluorocarbon-based ultrasound contrast agents, these nanoparticles have an atypical fractal internal structure. The nonvaporizing highly hydrophobic perfluorocarbon forms multiple cores within the polymeric matrix and is, surprisingly, hydrated with water, as determined from small-angle neutron scattering and nuclear magnetic resonance spectroscopy. Finally, the nanoparticles are used to image therapeutic dendritic cells with ultrasound in vivo, as well as with 19 F MRI and fluorescence imaging, demonstrating their potential for long-term in vivo multimodal imaging.

Outcomes of beta cell islet transplantation at subcutaneous sites have been disappointing due to hypoxia-induced oxidative stress by poor vascularization. However, subcutaneous engraftment still has advantages like the simple surgical procedure under local anesthesia and the possibility to biopsy when needed1. Therefore, it is important to have non-invasive imaging techniques available to monitor the exact location and fate of transplanted islets. Subcutaneously transplanted islets offer the unique opportunity to perform in vivo optical imaging, which is often limited by the short penetration depth of light when using intraportal transplantations. For validation purposes, 19F MRI using perfluorocarbon based particles can be used, which draws attention in the field of cellular imaging2 thanks to the lack of background signal and possibility for quantification of transplanted cells. These perfluorocarbon particles could easily be tacked with a fluorescent dye to be also detected by fluorescence imaging. To perform BLI, islets/cells need to be transduced by viral vectors. The present work deals with the use of multimodal in vivo imaging (BLI / Fluorescence /19F MRI) to image and monitor the transplanted pancreatic islets in mice. By combining different imaging techniques, we want to overcome the limitations in sensitivity, resolution and specificity of the individual method. Materials and methods Perfluorocarbon (PFCE) particle and lentiviral vector (LV): Cationic PFCE particles were prepared according to previous report3 with 3.5mol% red DiD fluorescent dye (particle concentration: 4.5*109 particles/ml, particle mean size: ±0.513μm). Lentiviral vector carrying the hEFlα promoter to drive triple flag tagged firefly luciferase (3flagFluc) and a puromycin resistance gene (PuroR) was used for cell/islet transduction4. Cell line and pancreatic islets isolation: Tumor insulin producing cells (INS-1E) were used for uptake validation as well as pancreatic islets isolated from Wistar rats (female, 8-10 weeks) using the collagenase digestion method. In-vitro studies: Phantoms containing PFCE labeled cells/pancreatic islets filled with 2% agarose were made for performing 19F MRI. PFCE labeled or LV transfected cells/islets were placed in 24 well plates for either fluorescent imaging or BLI. Animal model: C57Bl6 mice (male, 8-10 weeks) were subcutaneously injection of either 200 PFCE labeled/LV transfected pancreatic islets or one million PFCE labeled/LV transfected cells at one side with another 200μl PBS injected at the other side as negative control. Microscopy and Confocal imaging: OLYMPUS CKX41 Microscope (Tokyo, Japan) was used for cell/pancreatic islets visualization. Confocal images were also acquired using a Nikon A1R Eclipse Ti confocal microscope (Tokyo, Japan). NMR spectroscopy: NMR experiments were carried out using a 400 MHz Bruker Avance II NMR spectrometer (Bruker Biospin, Rheinstetten, Germany) with the following acquisition parameters: 350 ppm spectral width, 8 s relaxation delay, 128/2048 number of acquisitions and 64k data points. Optical imaging: Both fluorescence images and BLI images were acquired with an IVIS Spectrum system (PerkinElmer, Waltham, USA). For fluorescence imaging, epi-fluorescence illumination mode (FOV = 13.4 cm, Excitation peak = 648 nm, Emission peak = 670 nm) was used. D-luciferin (126 mg/kg) was administered intravenously after the cell/islets transplantation for BLI. MRI experiments: Both in vitro and in vivo MRI were performed using 9.4T Biospec small animal MRI system (Bruker, Ettlingen, Germany) equipped with a home-made surface coil tuneable and matchable to both 1H MRI and 19F MRI with same setups. RARE sequence was used for both in vitro and in vivo studies with some parameters modifications (RARE factor= 8/18; FOV= 6.4 cm*6.4 cm/8 cm*4 cm; TE= 15.9 ms; TR= 6 s (1H)/1 s (19F); slice thickness= 0.5 mm(1H)/2.5 mm(19F); Number of average= 2(1H)/1000(19F)). Results The pancreatic islets/INS-1E cells were incubated with PFCE particles/lentiviral vector and then further used for in vitro and in vivo studies. Viability essays confirmed that PFCE particle and lentiviral vector did not affect cells/pancreatic islets’ function and morphology. Microscopy images show both INS-1E cells and pancreatic islets have excellent uptake of PFCE particles after 24 hours’ incubation. Confocal image confirms the internalization of the PFCE particles by some pancreatic cells. NMR results further confirm and quantify the uptake, which is around 4*1012 19F atoms/cell and 1*1015 19F atoms/pancreatic islet. In vitro BLI shows both INS-1E cells and pancreatic islets could be transduced successfully after a 48 hours’ transduction. Phantoms containing labeled cells or pancreatic islets confirm good visualization by 19F MRI (approx. 200 labeled islets) within an hour acquisition. After subcutaneous transplantation, cells/ islets could be detected and monitored in vivo by all three different imaging modalities, resulting in the following detectability thresholds: approx. 100 islets for 19F MRI and 50 islets for both BLI and fluorescent imaging. Initial follow up results of PFCE labeled islets show the fluorescent signal could be tracked for over two weeks, while the 19F MRI signal can be sustained for over 3 weeks. Conclusion and Discussion By using subcutaneous islet transplantation, we were able to cross-validate their location and quantity using a tri-modal in vivo imaging platform. All three different modalities could provide direct or indirect information regarding the number of engrafted pancreatic islets. Future work will focus on the follow up after the transplantation of islets in diabetic animals using all three imaging modalities. The pancreatic islet’s tri-modal imaging has the potential to overcome the limitations in sensitivity, resolution and specificity of the individual methods. References 1.Sakata N et al., Diabetes Metab Res Rev 30(1), 1-10(2014); 2. Ahrens ET et al., Nature Biotechnology 23, 983-987(2005); 3.Dewitte H et al., Journal of Controlled Release 169, 141-149(2013); 4.Wolfs E et al., Plos One 9, issue 4(2014). ispartof: Molecular Imaging And Biology vol:20 issue:6 pages:940-951 ispartof: location:United States status: published

Contains fulltext : 153574.pdf (Publisher’s version ) (Open Access) AIM: In vivo imaging using (19)F MRI is advantageous, due to its ability to quantify cell numbers, but is limited for a lack of suitable labels. Here, we formulate two stable and clinically applicable labels for tracking two populations of primary human dendritic cells (DCs) simultaneously. MATERIALS & METHODS: Plasmacytoid and myeloid DCs are able to take up sufficient nanoparticles (200 nm) for imaging (10(12 19)F's per cell), despite being relatively nonphagocytic. RESULTS: Clinically relevant numbers of labeled DCs could be imaged in about 10 min, even on a clinical scanner. CONCLUSION: We demonstrate the use of perfluorocarbon nanoparticles for simultaneous (19)F MRI of distinct cell populations in a clinical setting, without spectroscopic imaging.

A mononuclear Gd-III Schiff base complex of formula [Gd(HL)(2)-(NO3)(3)] (1), where HL = 2-methoxy-6-[(E)-phenyliminomethyl]phenol, has been isolated and its structure was determined by single-crystal X-ray diffraction. H-1 NMR spectroscopic data reveal the zwitterionic nature of the HL ligand upon coordination with the Gd-III ion. Direct current (dc) magnetic susceptibility and X-band EPR measurements were performed to probe the electronic and magnetic properties of complex 1. The susceptibility and magnetization data were fitted simultaneously, resulting in spin-Hamiltonian parameters: S = 7/2, g = 2.018, and D = 0.108 cm(-1). The extracted parameters are in good agreement with the values obtained by fitting the temperature-dependent polycrystalline X-band EPR spectra of 1 (S = 7/2, g = 2.005, and D = 0.0815 cm(-1); E/D = 0.08). EPR dilution experiments performed on 1 indicates strongly that the anisotropy arises solely from the Gd-III ion with undetectably small or no contribution from spin-spin exchange interactions.

Adapting ways to functionalize polymer materials is becoming increasingly important to their implementation in translational biomedical sciences. By tuning the mechanical, chemical, and biological qualities of these materials, their applications can be broadened, opening the door for more advanced integration into modern medical techniques. Here, we report on a method to integrate chemical functionalizations into discrete, microscale polymer structures, which are used for tissue engineering applications, for in vivo localization, and three-dimensional manipulation. Iron oxide nanoparticles were incorporated into the polymer matrix using common photolithographic techniques to create stably functional microstructures with magnetic potential. Using magnetic resonance imaging (MRI), we can promote visualization of microstructures contained in small collections, as well as facilitate the manipulation and alignment of microtopographical cues in a realistic tissue environment. Using similar polymer functionalization techniques, fluorine-containing compounds were also embedded in the polymer matrix of photolithographically fabricated microstructures. The incorporation of fluorine-containing compounds enabled highly sensitive and specific detection of microstructures in physiologic settings using fluorine MRI techniques (19F MRI). These functionalization strategies will facilitate more reliable noninvasive tracking and characterization of microstructured polymer implants as well as have implications for remote microstructural scaffolding alignment for three-dimensional tissue engineering applications. © 2014 American Chemical Society.

Perfluorocarbon nanoemulsions (PFCNEs) have recently been undergoing rigorous study to investigate their ability to improve the therapeutic efficacy of tumor ablation by high intensity focused ultrasound (HIFU). For precise control of PFCNE delivery and thermal ablation, their accumulation and distribution in a tumor should be quantitatively analyzed. Here, we used fluorine-19 (F-19) magnetic resonance imaging (MRI) to quantitatively track PFCNE accumulation in a tumor, and analyzed how intra-tumoral PFCNE quantities affect the therapeutic efficacy of HIFU treatment. Ablation outcomes were assessed by intra-voxel incoherent motion analysis and bioluminescent imaging up to 14 days after the procedure. Assessment of PFCNE delivery and treatment outcomes showed that 2-3 mg/mL of PFCNE in a tumor produces the largest ablation volume under the same HIFU insonation conditions. Histology showed varying degrees of necrosis depending on the amount of PFCNE delivered. F-19 MRI promises to be a valuable platform for precisely guiding PFCNE-enhanced HIFU ablation of tumors.