Pharmacologic alternatives to riboflavin photochemical corneal cross-linking: a comparison study of cell toxicity thresholds.
OBJECTIVE
The efficacy of therapeutic cross-linking of the cornea using riboflavin photochemistry (commonly abbreviated as CXL) has caused its use to become widespread. Because there are known chemical agents that cross-link collagenous tissues, it may be possible to cross-link tissue pharmacologically. The present study was undertaken to compare the cell toxicity of such agents.


METHODS
Nine topical cross-linking agents (five nitroalcohols, glyceraldehyde [GLYC], genipin [GP], paraformaldehyde [FA], and glutaraldehyde [GLUT]) were tested with four different cell lines (immortalized human corneal epithelial cells, human skin fibroblasts, primary bovine corneal endothelial cells, and immortalized human retinal pigment epithelial cells [ARPE-19]). The cells were grown in planar culture and exposed to each agent in a range of concentrations (0.001 mM to 10 mM) for 24 hours followed by a 48-hour recovery phase. Toxicity thresholds were determined by using the trypan blue exclusion method.


RESULTS
A semiquantitative analysis using five categories of toxicity/fixation was carried out, based on plate attachment, uptake of trypan blue stain, and cellular fixation. The toxicity levels varied by a factor of 10(3) with the least toxic being mononitroalcohols and GLYC, intermediate toxicity for a nitrodiol and nitrotriol, and the most toxic being GLUT, FA, GP, and bronopol, a brominated nitrodiol. When comparing toxicity between different cell lines, the levels were generally in agreement.


CONCLUSIONS
There are significant differences in cell toxicity among potential topical cross-linking compounds. The balance between cross-linking of tissue and cell toxicity should be borne in mind as compounds and strategies to improve mechanical tissue properties through therapeutic tissue cross-linking continue to develop.
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, New York, United States.
MiJung
M
Kim
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, New York, United States.
Anna
A
Takaoka
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, New York, United States.
Quan V
QV
Hoang
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, New York, United States.
Stephen L
SL
Trokel
Department of Ophthalmology, College of Physicians and Surgeons, Columbia University, New York, New York, United States.
David C
DC
Paik
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D003315:Cornea / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D006801:Humans; D007640:Keratoconus / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D017319:Photosensitizing Agents / Q000494:pharmacology; D012256:Riboflavin / Q000494:pharmacology*
D003432:Cross-Linking Reagents; D017319:Photosensitizing Agents; D012256:Riboflavin
cellular toxicity; cornea; keratoconus; protein cross-linking