Bacterial keratitis: Photodynamic inactivation reduced experimental inflammation.

The successful treatment of bacterial keratitis remains an unsolved clinical problem. The current study aimed to establish a murine keratitis model and to investigate the effect of chlorin e6 (Ce6) and photodynamic inactivation (PDI) on corneal inflammation. The cornea of anesthetized mice was scratched and covered with a bacterial suspension of Pseudomonas aeruginosa. A paste containing Ce6 was applied to the cornea with subsequent exposure to specified light. Two days later the animals were sacrificed, and the globes were processed for light microscopy. Evaluation parameters were the maximal corneal thickness and the severity of the hypopyon. The maximal corneal thickness of 290±16 µm in the infected and untreated group was significantly reduced to 220±8 µm in the infected and treated group (P<0.05). In addition, the hypopyon was less severe in the infected and treated group. In conclusion, the present study indicates that PDI using Ce6 may be a potential approach to treat patients suffering with severe bacterial keratitis.

Chlorin e6 mediated photodynamic inactivation for multidrug resistant Pseudomonas aeruginosa keratitis in mice in vivo.

Following corneal epithelium scratches, mouse corneas were infected with the multidrug resistant (MDR) P. aeruginosa strain PA54. 24 hours later, 0% (for control group), 0.01%, 0.05% or 0.1% Chlorin e6 (Ce6), a second generation photosensitizer derived from chlorophyll, was combined with red light, for photodynamic inactivation (PDI). 1 hour or 2 days later, entire mouse eyes were enucleated and homogenized for counting colony forming units (CFU) of P. aeruginosa. For comparison, 0.1% Ce6 mediated PDI was started at 12 hours post infection, and 0.005% methylene blue mediated PDI 24 hours post infection. Clinical scores of corneal manifestation were recorded daily. Compared to the control, CFU 1 hour after PDI started 24 hours post infection in the 0.01% Ce6 and 0.05% Ce6 groups were significantly lower (more than one log10 reduction), the CFU 2 days post PDI higher in the 0.1% Ce6 group, clinical score lower in the 0.1% Ce6 group at 1 day post PDI. These findings suggest that PDI with Ce6 and red light has a transient efficacy in killing MDR-PA in vivo, and repetitive PDI treatments are required to fully resolve the infection. Before its clinical application, the paradoxical bacterial regrowth post PDI has to be further studied.

Gaseous nitric oxide for the local treatment of bacterial keratitis in mice.

The successful treatment of severe bacterial keratitis continues to be a challenge in animals and humans. In the present study the aim was to assess gaseous therapy using gaseous nitric oxide (gNO) in a murine model of Pseudomonas aeruginosa keratitis. The cornea of anesthetized mice was mechanically scratched and covered with a bacterial suspension of P. aeruginosa. One day later, the infected eyes were exposed to 200 ppm NO for 30 min. Three to seven days later the mice were sacrificed and the bulbi were obtained and processed for light microscopy. The read out parameter was the maximal corneal thickness and the severity of the hypopyon. The therapy with NO did not result in either a reduction of inflammation concerning the maximal corneal thickness or the severity of the hypopyon. The bacterial load was not investigated due to technical limitations. Thus, exposure to gNO did not reduce the local inflammation in P. aeruginosa induced murine keratitis at the investigated time-points. This does not exclude effects of NO on the bacterial load, and in experimental and human keratitis.