Final Report Summary - NANOSTEMCELLTRACKING (Nanoparticle probes for photoacoustic tracking of stem cell) Regenerative medicine has a huge potential as a new therapeutic tool for a great variety of diseases, however a large number of stem cell therapies fail before they can be translated to the clinic. A better understanding of the fate of transplanted cells and the ability to appropriately monitor grafting efficiency in preclinical models are factors that would facilitate clinical translation. In this project, we developed a new method to track stem cells in vivo using gold nanorods to label the cells and a novel imaging technique called multispectral optoacoustic tomography (MSOT) to detect them. This technology is based on the photoacoustic effect, which was first reported by A. Graham Bell in 1881. Briefly, after irradiate a material with pulsed light, the absorption of light generates a thermal expansion that creates a pressure wave and ultimately an ultrasound signal that can be transformed into an image. The main advantage of this technique is that uses light as excitation source but the outcome is ultrasounds. Therefore, the technique has the advantages of optical imaging (e.g. sensitivity, real time imaging, and molecular imaging). In addition, excellent resolution at depths up to 5 cm is enabled by measuring ultrasounds instead of light. Note that pure optical-based techniques are limited to no more than 1-2 mm penetration depths due to the scattering of light by tissue. We used gold nanorods as contrast agents for this technique because they have a very strong absorption in the near infrared, which is precisely the region of the spectra where biological tissue absorbs less, one could say it is the region where tissue is more “transparent”. In the first stages of our project, we optimised the synthesis of gold nanorods to be used for stem cell labelling. We observed that after cell uptake nanoparticles were physically packed inside cell vesicles. This altered the optical properties of gold nanorods making them less appealing as contrast agents. To overcome this limitation, we modify our gold nanorods with a silica shell. Silica is transparent and therefore does not modify inherent optical properties of nanorods. We also evaluated the safety of our nanoparticles with the standard toxicity assays, concluding that they did not show any sign of toxicity and they did not affect the differentiation potential of stem cells. In parallel, their photoacoustic properties were studied in phantoms that mimicked biological tissue. It was demonstrated in this stage the efficient labelling of cells with gold nanorods in terms of intensity and preservation of the intrinsic optical properties. The positive results obtained in the first stages of our project allowed us to move to in vivo experiments. We demonstrated an extraordinary sensitivity of cell detection using our system. Hence, ee were able to monitor as few as 10000 cells previously labelled with gold nanorods for 20 days with a resolution of 200 micrometres. Distinctly from other optical-based techniques, MSOT enables to track the evolution of nanorods-labelled cells engraftment with an excellent 3D spatial resolution even if they are deep in tissue. We also exploited the potential of gold nanorods for multilabelling. Different cells were labelled with nanorods with a different aspect ratio, and consequently different optical properties, and monitored in vivo. The preservation of the characteristic absorption band of the nanorods allowed to distinguish the two cell populations and to perform colocalization assays. Development of this particular technology will enable to identify how to cell populations are interacting in vivo in real time which would help to understand the dynamics of biological processes. To assess the potential of this technology in a systemic administration of cells, we labelled macrophages with gold nanorods and followed their accumulation in the liver. We observed a fast initial accumulation during the first two hours followed by a slow progressive accumulation. Our results are in agreement with the reported with other imaging techniques such as magnetic resonance, which validates the use of this technology. In an ongoing work, we are using this technique to monitor stem cells engraftment investigating in parallel the role played by the immune cells in this process.Interestingly, not only regenerative medicine but other fields such as oncology can benefit from the ability to monitor cells. For example, there is a growing interest in using cells of the immune system to treat cancer, as described in the so called immunotherapies. The possibility to track cells in real time might help to understand better this process and therefore being able to optimise the design of drugs. Moreover, the strong absorption of gold nanorods makes them ideal for photothermal therapy. Therefore they could be used as contrast agents and actuators at the same time. All this together, caught the attention of oncology surgeons to start a collaborative project to determine tumour margins to guide surgery in preclinical models. Documents connexes final1-jcomenge-mc-final-report.pdf
