Abstract
Purpose :
To evaluate photomodulation effects on LC beam microstructure and biomechanics.
Methods :
A multiphoton microscope system was used for structural modification and imaging of sheep LC beams. The system was coupled to a 25× NA 1.05 objective lens and a Ti-Sapphire laser with a 140-fs pulse duration and an 80-MHz repetition rate at a wavelength of 800 nm. For photomodulation, laser pulses with a pulse energy of 1.5 nJ were used and scanned in either a linear or planar pattern. For multiphoton imaging, the laser pulse energy was 0.15 nJ, and detection bandwidths of second harmonic generation (SHG) and two-photon autofluorescence (TPAF) signals were 350-450 nm and 500-550 nm, respectively. Instant polarized light microscopy (IPOL) was used to examine the collagen microstructure and stretch-induced deformation after photomodulation.
Results :
Photomodulation changed SHG and TPAF signals (Fig. 1A). The decreased SHG signal indicates collagen degradation, called "photoscalpel". The increased TPAF signal was related to crosslink enhancement, called "photocrosslinking." The occurrence of photocrosslinking at the border suggests the need for lower doses of laser irradiation. The profiles of SHG and TPAF signals show the lateral resolutions of photoscalpel and photocrosslinking were ~3 µm and ~6 µm, respectively. The IPOL image shows that photoscalpel made a fine cut at the border of the cavity, and the axial resolution was better than 16 µm (section thickness) (Fig. 1B). Stretch testing shows a lower strain in the photocrosslinking region, indicating an increase in tissue stiffness (Fig. 2A). In contrast, stretch testing shows high strain in the photoscalpel region, indicating less ability at carrying loads (Fig. 2B).
Conclusions :
We demonstrated femtosecond laser irradiation can change the microstructure and biomechanics of LC beams without using photosensitizers. The doses of laser irradiation modulate the effects of photocrosslinking and photoscalpel. The precision of photomodulation was at the µm-level in both lateral and axial directions. This approach to modulating tissue biomechanics may be a therapeutic strategy for addressing mechanical damage in glaucoma.
This abstract was presented at the 2024 ARVO Imaging in the Eye Conference, held in Seattle, WA, May 4, 2024.