Generic benzalkonium chloride-preserved travoprost eye drops are not identical to the branded polyquarternium-1-preserved travoprost eye drop: Effect on cultured human conjunctival goblet cells and their physicochemical properties

Abstract

Purpose: To investigate the effect of polyquaternium-1 (PQ)-preserved and benzalkonium chloride (BAK)-preserved travoprost eye drops on viability of primary human conjunctival goblet cell (GC) cultures and on secretion of mucin and cytokines. Furthermore, to evaluate the physicochemical properties of the branded travoprost eye drop Travatan® and available generics.

Methods: The effect of travoprost eye drops was evaluated on GC cultures. Cell viability was assessed through lactate dehydrogenase (LDH) and tetrazolium dye (MTT) colorimetric assays. Mucin secretion was evaluated by immunohistochemical staining. Secretion of interleukin (IL)-6 and IL-8 was measured using BD Cytometric Bead Arrays. pH, viscosity, droplet mass, osmolality and surface tension were measured for all included eye drops.

Results: In the LDH assay, BAK travoprost caused significant GC loss after 2 hrs of incubation compared to the control. PQ travoprost caused no GC loss at any time point. Both PQ- and BAK travoprost caused secretion of mucin to the cytoplasma. No difference in IL-6 and IL-8 secretion was identified compared to controls. The pH values for the generics were lower (pH 6.0) than the pH value for Travatan (pH 6.7; p < 0.0001). The viscosity was lowest for Travatan, while the mean droplet mass was higher for Travatan (35 mg) than the generics (28-30 mg; p ≤ 0.0318). The osmolality and surface tension did not differ between the eye drops investigated.

Conclusion: BAK travoprost caused GC loss, indicating that PQ preservation may be preferable in treatment of glaucoma. Furthermore, physicochemical properties of branded and generic travoprost eye drops can not be assumed to be identical.

Keywords: benzalkonium chloride; glaucoma; goblet cells; ocular surface; polyquaternium-1; travoprost.

Fig. 1

Relative cell survival of primary human conjunctival goblet cell (GC) cultures assessed through lactate dehydrogenase (LDH) assays. Results are presented as mean cell survival relative to a fixed 100 % survival for the control ± standard deviation. Cultures were incubated for 30, 60 and 120 min with polyquarternium‐1 (PQ)‐ or benzalkonium chloride (BAK)‐preserved travoprost eye drops. Cultures from three individual donors were included. The large bar shows significant decrease in GC survival after 2 hrs incubation with the generic BAK‐preserved travoprost eye drops compared to the control. There were no differences in survival between the control and Travatan at any time point. Only p‐values <0.05 are shown.

Fig. 2

Relative cell survival of primary human conjunctival goblet cell (GC) cultures assessed through tetrazolium dye (MTT) colorimetric assays. Results are presented as mean cell survival relative to a fixed 100 % GC survival for the control ± standard deviation. Cultures were incubated for 30 min with polyquarternium‐1 (PQ)‐ or benzalkonium chloride (BAK)‐preserved travoprost eye drops. Cultures from three individual donors were included. Significant decrease in GC survival was identified for all travoprost eye drops. Only p‐values <0.05 are shown.

Fig. 3

Secretion of mucin from primary human conjunctival goblet cell (GC) cultures assessed through immunohistochemical stainings. GCs were incubated for 30 minutes with culture medium as a negative control, polyquarternium‐1 (PQ)‐preserved Travatan or benzalkonium chloride (BAK)‐preserved Travoprost Stada. Travoprost Stada represents the BAK‐preserved generics. The nucleus was blue with DAPI (4′,6‐diamidino‐2‐phenylindole), the cytoskeleton green with anti‐cytokeratin‐7 and the mucin red with MUC5AC. The three stainings were merged. Dispersion of mucin to the cytoplasma indicates a secretagogue effect for both BAK‐ and PQ‐preserved travoprost. Stainings were performed on cultures from three individual donors. Scale bar = 100 μm. [Colour figure can be viewed at wileyonlinelibrary.com]

Fig. 4

Secretion of interleukin 6 (IL‐6) and interleukin 8 (IL‐8) from primary human conjunctival goblet cell (GC) cultures. Secretion is presented as mean concentration ± standard deviation after 30 min of incubation with polyquarternium‐1 (PQ)‐preserved Travatan and benzalkonium chloride (BAK)‐preserved Travoprost Stada. Travoprost Stada represents the BAK‐preserved generics. As a negative control GCs were incubated with culture medium. Secretion was measured on GCs from three individual donors using BD Cytometric Bead Arrays. No differences in cytokine secretion were identified compared to the control. Only p‐values <0.05 are shown.

Fig. 5

Physicochemical properties of the branded travoprost eye drop Travatan and the generics Travoprost Stada, Travoprost Teva and Bondulc. pH value, viscosity (mPa.s), drop mass (mg), number of drops per bottle, osmolality (mOsm/kg) and surface tension (mN/m) were measured. Values are shown as mean ± standard deviation. Significant differences in pH value, viscosity, drop mass and number of drop per bottle were identified. Only p‐values ≤0.05 are shown.