Marcin Piejko1,2,3, Piotr Walczak1,2,4, Xiaowei Li5,6, Jeff W M Bulte1,2,7,8,9, Miroslaw Janowski10,11. 1. Russell H. Morgan Department of Radiology and Radiological Science, Division of MR Research, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. 2. Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. 3. 3rd Department of General Surgery, Jagiellonian University Medical College, Krakow, Poland. 4. Department of Neurology and Neurosurgery, University of Warmia and Mazury, Olsztyn, Poland. 5. Translational Tissue Engineering Center, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. 6. Mary and Dick Holland Regenerative Medicine Program, Department of Neurological Sciences, The University of Nebraska Medical Center, Omaha, NE, USA. 7. Department of Biomedical Engineering, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. 8. Department of Chemical and Biomolecular Engineering, Whiting School of Engineering, The Johns Hopkins University, Baltimore, MD, USA. 9. Department of Oncology, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. 10. Russell H. Morgan Department of Radiology and Radiological Science, Division of MR Research, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. mjanows1@jhmi.edu. 11. Cellular Imaging Section and Vascular Biology Program, Institute for Cell Engineering, The Johns Hopkins University School of Medicine, Baltimore, MD, USA. mjanows1@jhmi.edu.
Abstract
PURPOSE: We studied the feasibility of labeling hydrogel scaffolds with a fluorine nanoemulsion for 19F- magnetic resonance imaging (MRI) to enable non-invasive visualization of their precise placement and potential degradation. PROCEDURE: Hyaluronan-based hydrogels (activated hyaluronan, HA) with increasing concentrations of fluorine nanoemulsion (V-sense) were prepared to measure the gelation time and oscillatory stress at 1 h and 7 days after the beginning of gelation. All biomechanical measurements were conducted with an ARES 2 rheometer. Diffusion of fluorine from the hydrogel: Three hydrogels in various Vs to HA volumetric ratios (1:50, 1:10, and 1:5) were prepared in duplicate. Hydrogels were incubated at 37 °C. To induce diffusion, three hydrogels were agitated at 1000 rpm. 1H and 19F MRI scans were acquired at 1, 3, 7 days and 2 months after gel preparation on a Bruker Ascend 750 scanner. To quantify fluorine content, scans were analyzed using Voxel Tracker 2.0. Assessment of cell viability in vitro and in vivo: Luciferase-positive mouse glial-restricted progenitors (GRPs) were embedded in 0:1, 1:50, 1:10, and 1:5 Vs:HA mixtures (final cell concentration =1 × 107/ml). For the in vitro assay, mixtures were placed in 96-wells plate in triplicate and bioluminescence was measured after 1, 3, 7, 14, 21, and 28 days. For in vivo experiments, Vs/HA mixtures containing GRPs were injected subcutaneously in SCID mice and BLI was acquired at 1, 3, 7, and 14 days post-injection. RESULTS: Mixing of V-sense at increasing ratios of 1:50, 1:10, and 1:5 v/v of fluorine/activated hyaluronan (HA) hydrogel gradually elongated the gelation time from 194 s for non-fluorinated controls to 304 s for 1:5 V-sense:HA hydrogels, while their elastic properties slightly decreased. There was no release of V-sense from hydrogels maintained in stationary conditions over 2 months. The addition of V-sense positively affected in vitro survival of scaffolded GRPs in a dose-dependent manner. CONCLUSIONS: These results show that hydrogel fluorination does not impair its beneficial properties for scaffolded cells, which may be used to visualize scaffolded GRP transplants with 19F MRI.
PURPOSE: We studied the feasibility of labeling hydrogel scaffolds with a fluorine nanoemulsion for 19F- magnetic resonance imaging (MRI) to enable non-invasive visualization of their precise placement and potential degradation. PROCEDURE: Hyaluronan-based hydrogels (activated hyaluronan, HA) with increasing concentrations of fluorine nanoemulsion (V-sense) were prepared to measure the gelation time and oscillatory stress at 1 h and 7 days after the beginning of gelation. All biomechanical measurements were conducted with an ARES 2 rheometer. Diffusion of fluorine from the hydrogel: Three hydrogels in various Vs to HA volumetric ratios (1:50, 1:10, and 1:5) were prepared in duplicate. Hydrogels were incubated at 37 °C. To induce diffusion, three hydrogels were agitated at 1000 rpm. 1H and 19F MRI scans were acquired at 1, 3, 7 days and 2 months after gel preparation on a Bruker Ascend 750 scanner. To quantify fluorine content, scans were analyzed using Voxel Tracker 2.0. Assessment of cell viability in vitro and in vivo: Luciferase-positive mouse glial-restricted progenitors (GRPs) were embedded in 0:1, 1:50, 1:10, and 1:5 Vs:HA mixtures (final cell concentration =1 × 107/ml). For the in vitro assay, mixtures were placed in 96-wells plate in triplicate and bioluminescence was measured after 1, 3, 7, 14, 21, and 28 days. For in vivo experiments, Vs/HA mixtures containing GRPs were injected subcutaneously in SCIDmice and BLI was acquired at 1, 3, 7, and 14 days post-injection. RESULTS: Mixing of V-sense at increasing ratios of 1:50, 1:10, and 1:5 v/v of fluorine/activated hyaluronan (HA) hydrogel gradually elongated the gelation time from 194 s for non-fluorinated controls to 304 s for 1:5 V-sense:HA hydrogels, while their elastic properties slightly decreased. There was no release of V-sense from hydrogels maintained in stationary conditions over 2 months. The addition of V-sense positively affected in vitro survival of scaffolded GRPs in a dose-dependent manner. CONCLUSIONS: These results show that hydrogel fluorination does not impair its beneficial properties for scaffolded cells, which may be used to visualize scaffolded GRP transplants with 19F MRI.
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