In vitro study of inflammatory potential and toxicity profile of latanoprost, travoprost, and bimatoprost in conjunctiva-derived epithelial cells.
OBJECTIVE
Conjunctiva-derived epithelial cells were used to investigate, in vitro, the expression of various inflammation-associated markers known to be overexpressed in patients with glaucoma after contact with the three major commercially available eye drops containing prostaglandin analogues. The impact on cellular viability and apoptosis in the same cell line was evaluated, to address the possible proinflammatory and/or toxic origin of the most frequent clinical impairments induced by prostanoids (i.e., conjunctival hyperemia).


METHODS
Conjunctiva-derived cells were treated in vitro with the commercial solutions of latanoprost, travoprost, bimatoprost, prostaglandin (PG)F2alpha, tumor necrosis factor (TNF)-alpha, and different concentrations of benzalkonium chloride (BAC). Expressions of three inflammation- and immune-related markers, intercellular adhesion molecule (ICAM)-1, platelet-endothelial cell adhesion molecule (PECAM)-1 and HLA DR, were evaluated with flow cytometry after 24 to 72 hours of contact at low, subtoxic concentrations. Toxicological tests were also performed with cold-light cytofluorometry, in which cellular viability and apoptosis were evaluated with the neutral red and Hoechst/propidium iodide tests, respectively.


RESULTS
TNFalpha induced or stimulated expression of the three inflammatory markers, whereas the PGF2alpha, latanoprost, travoprost, and bimatoprost solutions did not induce an increase in these markers and even produced a marked reduction of ICAM-1 and PECAM-1 expression in those solutions most concentrated in BAC, thus suggesting a toxic phenomenon in cellular membranes induced by the preservative rather than the medication itself. Cytotoxic assays confirmed this hypothesis and showed significant toxicity with prostaglandin analogues after prolonged contact, proportional to the concentration of BAC in the solution and similar to that of the corresponding concentration of BAC alone, bimatoprost having both the least concentration of BAC and the least cytotoxic in these experimental conditions.


CONCLUSIONS
The comparison of latanoprost, travoprost, and bimatoprost, in their commercial formulations, showed that none of them appeared to induce direct stimulation of the inflammatory pathways involving adhesion molecules or class II antigens, although these markers have been found ex vivo in conjunctival specimens from patients treated with prostaglandins. In fact, their toxicity was mild and seemed to be primarily related to the concentration of BAC, their common preservative, which may be the major factor responsible for long-term ocular surface reactions in patients receiving topical prostaglandins, but most likely is not a factor in early and transient conjunctival hyperemia.
Guenoun
Jean-Marc
JM
Department of Ophthalmology, Quinze-Vingts National Ophthalmology Hospital, Paris, France.
Baudouin
Christophe
C
Rat
Patrice
P
Pauly
Aude
A
Warnet
Jean-Michel
JM
Brignole-Baudouin
Françoise
F
D000577:Amides; D000959:Antihypertensive Agents / Q000633:toxicity; D017209:Apoptosis; D001548:Benzalkonium Compounds / Q000633:toxicity; D000069580:Bimatoprost; D015415:Biomarkers / Q000378:metabolism*; D002470:Cell Survival; D002478:Cells, Cultured; D003008:Cloprostenol / Q000031:analogs & derivatives* / Q000633:toxicity*; D003228:Conjunctiva / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D005434:Flow Cytometry; D006684:HLA-DR Antigens / Q000378:metabolism; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D000077338:Latanoprost; D008055:Lipids / Q000633:toxicity*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D011310:Preservatives, Pharmaceutical / Q000633:toxicity; D011461:Prostaglandins F, Synthetic / Q000633:toxicity*; D000069557:Travoprost; D014409:Tumor Necrosis Factor-alpha / Q000633:toxicity
D000577:Amides; D000959:Antihypertensive Agents; D001548:Benzalkonium Compounds; D015415:Biomarkers; D006684:HLA-DR Antigens; D008055:Lipids; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D011310:Preservatives, Pharmaceutical; D011461:Prostaglandins F, Synthetic; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D003008:Cloprostenol; D000077338:Latanoprost; D000069580:Bimatoprost; D000069557:Travoprost