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Descrizione
Second-harmonic generation (SHG) microscopy is based on a coherent non-linear scattering process that allows high-resolution deep-tissue imaging of biological structures having high hyperpolarizability and structural anisotropy. For this reason, SHG represents a powerful tool for imaging collagen and probing its hierarchical organization from molecular scale up to tissue architectural level. In this study, SHG microscopy is used to characterize collagen ultrastructural modifications in murine corneal buttons treated with conventional and FLASH radiotherapy (RT). This evaluation serves as a benchmark of the structural integrity of the cornea, which is essential for the initial passage of light into the visual system. Stacks of optical sections covering the full corneal thickness were characterized using image analysis methods and parameters describing the organization of collagen structures at a supramolecular level. The preliminary results using Electron Flash show an increased off-plane contribution of SHG emitters after RT, with a lower variation in FLASH-RT than in conventional-RT, when compared to control samples indicating potential tissue-sparing effects. Despite additional samples and a larger statistic would be required to confirm the results and to understand the potential of this methodology for monitoring dose-dependent modifications of collagen, this approach could become a useful tool for monitoring the effect of RT on connective tissues and for evaluating the impact of different ionization sources.
