Cross-linking of stromal collagen with Riboflavin and UVA radiation is an alternative treatment of keratoconus. After
the cross-linking a wound healing process starts with the regeneration of the abraded epithelial layer and the stromal
keratocyte-network. To clarify possible side effects by visualization we established an imaging platform for the
multimodal three-dimensional imaging of the cornea and looked for differences between normal and cross-linked rabbit
corneae. The microscopy system utilizes femtosecond laser light for two photon excitation of autofluorescent metabolic
compounds, second harmonic imaging in forward and backward direction for the study of stromal collagen-I structure
and confocal detection of the backscattered femtosecond laser light for cell detection. Preliminary results show
signatures of treatment 5 weeks after the intervention in all imaging modalities.
Cross-linking of the cornea with application of Ribovlavin and UV-A light is an evolving clinical treatment of the eye
disease keratoconus. Despite the positive clinical track record of corneal cross-linking, the complex wound healing
process after the treatment is still under investigation. In this study an animal model was used to clarify the state of
wound healing 5 weeks after treatment. Cross-linked rabbit corneae were imaged with reflective confocal laser scanning
and nonlinear microscopy, namely second harmonic imaging microscopy (SHIM) and two-photon excited
autofluorescence. First results show that the NAD(P)
H-autofluorescence of the corneal keratocytes and their scattering
signal still show a signature of the treatment five weeks after the cross-linking procedure. The SHIM signals show the
structural morphology of the fibrous collagen sheets in the stroma of the cornea. SHIM detected in the forward direction
differs substantially from backward SHIM, but no signature of treatment was found in both detection channels of the
SHIM signal.
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