Abstract
Purpose:
Our previous study using ex vivo eyes demonstrated that Rose Bengal (RB) plus green light crosslinking (RGX) rapidly increases cornea stiffness in the anterior stroma without causing keratocyte toxicity. This in vivo study of RGX evaluates corneal stiffening and potential effects on other ocular structures.
Methods:
Corneas of young Dutch belted rabbits were de-epithelialized, then treated with RB (0.1% in PBS) and 100 J/cm2 green light (532 nm, 6.6 min) or were controls (no RB, no light.) Surface temperature was measured with a non-contact infrared thermometer during the irradiation. On days 3, 7 and 28, epithelialization was observed and retina fluorescein fluorescence angiography was carried out with a scanning laser ophthalmoscope. On days 1 and 28, eyes were harvested: cornea stiffness was evaluated by uniaxial tensiometry, keratocyte number on H&E-stained sections, viability of cells in the iris by LDH staining and the retinal outer nuclear layer thickness was measured.
Results:
RGX increased the average stromal stiffness by 2.7-fold on day 1 and by 1.6-fold on day 28 compared to controls. During the irradiation, the maximum temperature increase was 7°C. Corneas of both RGX and control groups were >80% re-epithelialized by day 3 and completely re-epithelialized by day 7. RGX did not alter the number of keratocyes in the anterior 100 µm of stroma. Similarly, cell viability in the iris was not affected by RGX and the retinal outer nuclear layer thickness did not differ between control and RGX groups. Fluorescein fluorescence angiography on days 3, 7 and 28 did not show any leakage from retinal vessels or between pigmented epithelial cells indicating that the blood retinal barrier was not affected by RGX. These results are consistent with the calculations indicating that the retinal irradiance and fluence delivered by RGX are below ANSI damage thresholds for thermal or photochemical damage.
Conclusions:
In vivo RGX treatment requiring less than 12 min total time substantially increases cornea stiffness while not decreasing keratocyte numbers or damaging the iris or retina. These results suggest that RGX is a viable approach for treating ectatic cornea conditions.
Keywords: 574 keratoconus •
484 cornea: stroma and keratocytes •
647 photodynamic therapy