Title: Development of Imaging Methods towards Monitoring of Cell Therapy by In Vivo Magnetic Resonance Imaging
Other Titles: De Ontwikkeling van Kern Spin Tomografie Methoden gericht op In Vivo Opvolging van Celtherapie
Authors: Atre, Ashwini
Issue Date: 22-Nov-2013
Abstract: Despite recent advancements in medical research, many disorders still remain beyond the capabilities of current treatments. Cell based therapy is extensively being investigated to solve these unmet clinical needs. Several techniques are under development to assist the monitoring and validation of cell therapy. Of which, non-invasive imaging is one of the prime aides used to follow up on therapy efficacy and safety. By preloading therapeutic cells with MR contrast agents, it is possible to track the location of cells with magnetic resonance imaging (MRI). Current pre-labeling strategies are optimized mainly for non-phagocytic stem cells based on improved nanoparticle (NP) properties. With increasing knowledge about the cellular and molecular biology of tissue/ organ regeneration, extra-embryonic sources (like adult stem cells: MSCs, NSCs, HSCs or immune cells like dendritic cells, T-cells) have also been utilized as therapeutic vehicles. These cells exhibit different regenerative capacities but also different cell cytoskeleton organization and elasticity, cell shape, and adhesion strength and in vitro culture characteristics. Due to these different parameters involved in the uptake mechanism, it is not always feasible to translate currently available labeling protocols from one cell type to other. Therefore, one aim of this PhD thesis was to contribute to the development of cell labeling strategies with iron oxide based nanoparticles (NPs) to follow the fate of different therapeutic cells (adherent, in suspension and in differentiated mixed culture) in in vivo preclinical models. To achieve this goal, we utilize unspecific as well as specific (or targeted) labeling principles.In the context of unspecific labeling, means of NPs uptake is normally achieved by a simple incubation step. Lot of attention has been paid to NP related characteristic where size, surface charge and surface coating of NPs shown to be responsible for differential uptake and toxic responses in different types of cells. Here, we determined that the cell based parameters (cell size, proliferation rate) are responsible for such variation with the help of electron microscopic investigation. This study also emphasizes the need for performing validation experiments by using, for example, transmission electron microscopy to determine the fate of NPs post endocytosis. Next, unspecific labeling approaches were employed for labeling of pancreatic islets (PIs). Here we showed that magnetoliposomes (MLs), due to their positive surface charge and small size, were taken up by PIs in relatively shorter time (2-4 hours) when compared to commercially available agents like Endorem and Resovist (24-72 hours) without showing any apparent negative effects on the viability and functionality of PIs. These preliminary results indicate the potential of MLs as promising alternative for tracking transplanted PIs in diabetes model. Targeted labeling was studied in feasibility experiments to check if galactose functionalized MLs would be specific to hepatocytes in vitro and in vivo. Galactose MLs were tested in mixed cultures generated after differentiation of embryonic stem cells. After particle labeling, cells were separated magnetically and the positive and negative magnetic fractions were tested for the presence of hepatocyte specific marker. Due to the very small percentage of mature hepatocyte population, it was difficult to isolate hepatocytes from mixed cultures. Specificity of galactose MLs in vivo was evaluated by systemic administration of the particles in healthy mice. MR analysis, microscopic evaluation and immunostaining confirmed successful targeting of hepatocytes. The technology can further be implemented to determine the onset of liver diseases. This work contributes to the development of labeling strategies for monitoring cell based therapy in different disease models in vivo. This knowledge is currently being translated to other multiple applications including the development of cell therapy strategies and for determining the onset of diseases. Thus, this work will contribute to a solid basis for future applications and development in in vivo monitoring of cells with MR imaging.
Publication status: published
KU Leuven publication type: TH
Appears in Collections:Biomedical MRI
Stem Cell Biology and Embryology (+)

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