title: In vitro effects of preserved and unpreserved anti-allergic drugs on human corneal epithelial cells.
abstract: OBJECTIVE
  Treatment with topical eye drops for long-standing ocular diseases like allergy can induce detrimental side effects. The purpose of this study was to investigate in vitro cytotoxicity of commercially preserved and unpreserved anti-allergic eye drops on the viability and barrier function of monolayer and stratified human corneal-limbal epithelial cells.
  
  
  METHODS
  Cells were treated with unpreserved ketotifen solution, benzalkonium chloride (BAC)-containing anti-allergic drugs (ketotifen, olopatadine, levocabastine) as well as BAC alone. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to determine cell viability. Effects of compounds on barrier function were analyzed measuring transepithelial electrical resistance (TEER) to determine paracellular permeability and rose bengal assays to evaluate transcellular barrier formation.
  
  
  RESULTS
  The BAC-preserved anti-allergic formulations and BAC alone significantly reduced cell viability, monolayer cultures being more sensitive to damage by these solutions. Unpreserved ketotifen induced the least diminution in cell viability. The extent of decrease of cell viability was clearly dependent of BAC presence, but it was also affected by the different types of drugs when the concentration of BAC was low and the short time of exposure. Treatment with BAC-containing anti-allergic drugs and BAC alone resulted in increased paracellular permeability and loss of transcellular barrier function as indicated by TEER measurement and rose bengal assays.
  
  
  CONCLUSIONS
  The presence of the preservative BAC in anti-allergic eye drop formulations contributes importantly to the cytotoxic effects induced by these compounds. Stratified cell cultures seem to be a more relevant model for toxicity evaluation induced on the ocular surface epithelia than monolayer cultures.
authors.affiliation: Department of Biochemistry and Molecular Biology IV, Faculty of Optics and Optometry, Universidad Complutense de Madrid , Madrid, Spain .
authors.forename: Ana
authors.initials: A
authors.lastname: Guzman-Aranguez
authors.forename: Patricia
authors.initials: P
authors.lastname: Calvo
authors.forename: Inés
authors.initials: I
authors.lastname: Ropero
authors.forename: Jesús
authors.initials: J
authors.lastname: Pintor
mesh_terms: D018926:Anti-Allergic Agents / Q000008:administration & dosage / Q000633:toxicity*; D001548:Benzalkonium Compounds / Q000737:chemistry / Q000633:toxicity; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003990:Dibenzoxepins / Q000008:administration & dosage / Q000633:toxicity; D017097:Electric Impedance; D004847:Epithelial Cells / Q000187:drug effects* / Q000473:pathology; D019573:Epithelium, Corneal / Q000166:cytology / Q000187:drug effects* / Q000473:pathology; D006801:Humans; D066298:In Vitro Techniques; D007665:Ketotifen / Q000008:administration & dosage / Q000633:toxicity; D000069605:Olopatadine Hydrochloride; D009883:Ophthalmic Solutions; D010880:Piperidines / Q000008:administration & dosage / Q000633:toxicity; D011310:Preservatives, Pharmaceutical / Q000737:chemistry / Q000633:toxicity*
chemical_list: D018926:Anti-Allergic Agents; D001548:Benzalkonium Compounds; D003990:Dibenzoxepins; D009883:Ophthalmic Solutions; D010880:Piperidines; D011310:Preservatives, Pharmaceutical; D000069605:Olopatadine Hydrochloride; C047340:levocabastine; D007665:Ketotifen
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