J Korean Med Sci.  2015 Dec;30(12):1856-1864. 10.3346/jkms.2015.30.12.1856.

The Toxicity of Nonsteroidal Anti-inflammatory Eye Drops against Human Corneal Epithelial Cells in Vitro

• Lee JS
• Kim YH
• Park YM

• KMID: 2359972
• DOI: http://doi.org/10.3346/jkms.2015.30.12.1856

Abstract

This study investigated the toxicity of commercial non-steroid anti-inflammatory drug (NSAID) eye solutions against corneal epithelial cells in vitro. The biologic effects of 1/100-, 1/50-, and 1/10-diluted bromfenac sodium, pranoprofen, diclofenac sodium, and the fluorometholone on corneal epithelial cells were evaluated after 1-, 4-, 12-, and 24-hr of exposure compared to corneal epithelial cell treated with balanced salt solution as control. Cellular metabolic activity, cellular damage, and morphology were assessed. Corneal epithelial cell migration was quantified by the scratch-wound assay. Compared to bromfenac and pranoprofen, the cellular metabolic activity of diclofenac and fluorometholone significantly decreased after 12-hr exposure, which was maintained for 24-hr compared to control. Especially, at 1/10-diluted eye solution for 24-hr exposure, the LDH titers of fluorometholone and diclofenac sodium markedly increased more than those of bromfenac and pranoprofen. In diclofenac sodium, the Na+ concentration was lower and amount of preservatives was higher than other NSAIDs eye solutions tested. However, the K+ and Cl- concentration, pH, and osmolarity were similar for all NSAIDs eye solutions. Bromfenac and pranoprofen significantly promoted cell migration, and restored wound gap after 48-hr exposure, compared with that of diclofenac or fluorometholone. At 1/50-diluted eye solution for 48-hr exposure, the corneal epithelial cellular morphology of diclofenac and fluorometholone induced more damage than that of bromfenac or pranoprofen. Overall, the corneal epithelial cells in bromfenac and pranoprofen NSAID eye solutions are less damaged compared to those in diclofenac, included fluorometholone as steroid eye solution.

Keyword

Bromfenac Sodium; Corneal Epithelial Cells; Diclofenac Sodium; Fluorometholone; Pranoprofen; Cytotoxicity

MeSH Terms

Anti-Inflammatory Agents, Non-Steroidal/administration & dosage/*toxicity
Benzophenones/administration & dosage/toxicity
Benzopyrans/administration & dosage/toxicity
Bromobenzenes/administration & dosage/toxicity
Cell Movement/drug effects
Cells, Cultured
Diclofenac/administration & dosage/toxicity
Epithelial Cells/drug effects/metabolism/ultrastructure
Epithelium, Corneal/cytology/*drug effects/metabolism
Fluorometholone/administration & dosage/toxicity
Humans
L-Lactate Dehydrogenase/metabolism
Microscopy, Electron, Transmission
Ophthalmic Solutions
Propionates/administration & dosage/toxicity
Anti-Inflammatory Agents, Non-Steroidal
Benzophenones
Benzopyrans
Bromobenzenes
Diclofenac
Fluorometholone
L-Lactate Dehydrogenase
Ophthalmic Solutions
Propionates

Figure

• 

  Fig. 1 The absorption rate of water-insoluble formazan in corneal epithelial cells as measured by scanning spectrometry. The cellular metabolic activity of diclofenac sodium or fluorometholone markedly decreased after 1/10-diluted eye solution for 12-hr of exposure compared to control (P = 0.037, P = 0.021), and was significantly maintained for 24-hr (P = 0.032, P = 0.018), compared with those of bromfenac sodium and pranoprofen (by Kruskal-Wallis test, statistically significant at P < 0.05).

• 

  Fig. 2 Lactate dehydrogenase (LDH) titers of cultured corneal epithelial cells by NSAIDs. Especially, the LDH titers at 1/10-diluted fluorometholone and diclofenac significantly increased after 12-hr exposure (P = 0.029, P = 0.039) and 24-hr exposure compared to control (P = 0.024, P = 0.041) rather than other two NSAIDs, respectively (by Kruskal-Wallis test, statistically significant at P < 0.05).

• 

  Fig. 3 Scratch-wound assay after control (A), fluorometholone (B), pranoprofen (C), bromfenac (D), or diclofenac (E) exposure. Migration was assessed after a 1- (1), 24- (2), and 48- (3) hr exposure to A1-A3, B1-B3, C1-C3, D1-D3, or E1-E3. With time, the diclofenac and fluorometholone-exposed cells did not migrate to the wound region compared to bromfenac and pranoprofen-treated cells. After 1/10 diluted eye solution for 48-hr exposure, the cell migration of diclofenac and fluorometholone significantly inhibited compared with that of control.

• 

  Fig. 4 Corneal epithelial cell transmission electron micrographs after 24-hr exposure to all NSAIDs and control. (A) Diclofenac. (B) Bromfenac. (C) Pranoprofen. (D) Fluorometholone. (E) Control. At 24-hr exposure to 1/50-diluted diclofenac sodium or fluorometholone compared with control, there was more severe cellular morphology of corneal epithelial cell such as microvilli disruption, vacuole formation, plasma membrane, and nuclear changes compared with those of bromofenac sodium or pranoprofen.

• 

  Fig. 5 Corneal epithelial cell transmission electron micrographs after 48-hr exposure to all NSAIDs and control. (A) Diclofenac. (B) Bromfenac. (C) Pranoprofen. (D) Fluorometholone. (E) Control. At 48-hr exposure to 1/50-diluted diclofenac sodium or fluorometholone compared with control, there was more severe cellular morphology of corneal epithelial cell rather than those of bromofenac sodium or pranoprofen.

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