Nine human organ donor lenses aged 58 to 75 years of age were exposed to infra-red 800 nm radiation from a femtosecond pulsed Ti:Sapphire laser using pulse energies ranging from 0.10 to 0.55 µJ. The laser beam was scanned across the lens in a line-by-line raster pattern covering a volume of 1mm in height ×1 mm in width ×0.52 mm in depth. Photobleaching was visible to the unaided observer in the treated area with no signs of damage to the untreated parts of the lens (). Using a slit-lamp biomicroscope it was seen that photobleaching was confined to the subvolume inside the lens targeted by the scanning procedure. The effect of the laser treatment remained visible 1 to 2 weeks after the treatment. Continued observation was not possible due to the gradual optical deterioration of the donor lenses post mortem.
Photographic documentation of the laser photobleaching.
After laser treatment the transmission of light through the lens was increased over the entire visible spectrum and most prominently in the blue-green part (), where the age-related loss of transmission is also most pronounced. The effect of the laser treatment showed a clear dose response with the number of scans applied to the lens ().
Changes in the transmission properties after treatment.
The treatment response depends on the laser dosage.
To estimate the clinical relevance of the laser-induced improvement in lens transmission the age-induced changes in lens transmission was assessed in a separate experiment using twenty-five human donor lenses (). We found that the age-related changes in transmission properties of blue-green light from 430 to 530 nm could be approximated by Eq. 1, where Age
is age in years, p<0.0001, R2
Using Eq. 1 to calculate the apparent lens age after laser treatment, the improvement in lens transmission after treatment was found to correspond to a reduction in lens age of 5.2 years (range 3.0 to 7.3 years, ). Notably, the treatment effects presented in was the result of treating a subvolume of the lens of 0.52 mm in thickness, corresponding to 1/10 of the axial thickness of a typical lens ().
Age-dependent decline in the transmission of blue light.
Increase in transmission of human lenses after femtosecond laser photolysis.
Schematic presentation of the laser treatment shown on a cross section of a lens in the sagittal plane.
During 800 nm femtosecond laser irradiation the lenses emitted a strong blue autofluorescence signal indicating that two-photon absorption took place (). The rate of two-photon absorption (RTPA
)depends on the density (ρ
) of the molecules that can be excited by two-photon absorption, the two-photon absorption cross section for the molecule (β
), laser power/area (In
, with n being the number of photons involved in the process, i.e. n
2 for a two-photon process), and the energy of the impinging photons (︀ω
). Assuming that the medium is optically thin, the rate of TPA of a laser beam impinging on a cylindrical volume with a cross sectional area A
and length L
will follow Eq. 2:
If not only the fluorescence but also the photobleaching was the result of two-photon absorption it follows from Eq. 2 that the rate of photobleaching will be faster in more densely coloured lenses and when more laser power is applied to the lens. In other words, older and more densely coloured lenses will, predictably, benefit more from the laser treatment than younger and less coloured lenses.
Fluorescence intensity profiles.
We found that, indeed, the photobleaching was significantly (p
0.78) greater for more densely coloured lenses combined with the square of the laser power, Eq. 3:
denominates the increase (in percent) in transmission after laser treatment, Tpre
denominates the transmission before laser treatment, and I
is the mean laser power (in Watt). All other variables from Eq. 2 were kept constant between experiments, i.e. the energy of the impinging photons, the area of laser spot size, and the two-photon absorption cross section. The transmission measurements before treatment was used as a surrogate measure of the density of the two-photon absorbing molecules in the lens.