Study of nanocrystals activated by acoustic stimuli for anticancer therapy or tissue engineering

In recent years, several nanotools have been developed as effective therapeutic agents against cancer. These tools can be exploited for their intrinsic toxicity or they can be triggered by external stimuli of a different nature to cause cell death. Ultrasound (US) has emerged as a promising technique for the activation of specific nanomaterials, due to its deep tissue penetration and ability to focus in a specific area. This research work investigates the ultrasound response capacity of a specific nanocrystal. In particular, the system is formed by nanocrystals of zinc oxide, doped with iron atoms to increase the biocompatibility of the nanomaterial, functionalized with amino-propyl groups with the aim of improving the stability of the nanocrystals and increasing the gas pocket present on the surface of nanoparticles, which play an important role in the interaction with US. The final Fe:ZnO nanocrystals were further decorated with a lipid bilayer shell, designed to impart biomimetic properties to Fe:ZnO. Ultrasonic irradiation of these nanocrystals is able to increase the phenomenon of inertial cavitation, defined as the formation and implosion of gas bubbles in a liquid medium. Under the influence of ultrasonic waves, these gas nuclei expand and keep absorbing energy from the waves, and this can lead to the formation of free radicals resulting from the sonolysis of water and oxygen known as ROS (Reacting Oxygen Species). By introducing semiconductor nanocrystals into a liquid medium, the cavitation threshold is reduced, enabling the use of low-intensity ultrasound. This also makes it easy to obtain sonoluminescence, a secondary effect of cavitation which generates photons from the implosion of gas bubbles, which gives specific spectral characteristics to the emitted light. In view of the use of targeted and selective nanoparticles at the site of interest, here you get a customized bio-imaging method against a specific tumor cell or tissue, with high-resolution capabilities. The set up presented for capturing sonoluminescent light is composed of an inverted fluorescence microscope, which creates the path of the light to be captured, and a cooled CCD camera, capable of detecting the signal deriving from sonoluminescence. A comprehensive study was conducted to evaluate the light generated by the interaction of US with pure water, in the presence of both Fe:ZnO NC and lipid-coated ZnO NC. At the biological level, different concentrations of nanomaterials were tested and sonoluminescence measurements were performed not only in water but also in cell culture media. Furthermore, nanocrystal-assisted sonoluminescence was recorded for the first time in living cells, using two different cell lines. The evaluation of cell viability also allowed to monitor the safety of this new imaging method and to compare the results obtained. In addition, various methods have been developed to identify and measure the generation of free radicals, such as the use of sonochemiluminescence measurements with Luminol solutions, allowing for rapid qualitative analysis, making it easier to detect ROS with a camera in a dark environment. Electron paramagnetic resonance spectroscopy has also been employed as a technique to provide a more reliable and quantitative analysis of ROS concentration. In conclusion, the experimental results demonstrate how ZnO NCs enhance the light signal of sonoluminescence, supported by Matlab simulations. This approach therefore allows to define a promising imaging technique.