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The preservative polyquaternium-1 increases cytoxicity and NF-kappaB linked inflammation in human corneal epithelial cells.

Paimela T, Ryhänen T, Kauppinen A, Marttila L, Salminen A, Kaarniranta K - Mol. Vis. (2012)

Bottom Line: Interestingly, Travatan and Systane Ultra activated NF-κB and elevated the secretion of inflammation markers IL-6 by 3 to eightfold and IL-8 by 1.5 to 3.5 fold, respectively, as analyzed with ELISA.Eye drops containing PQ-1 evoke cytotoxicity and enhance the NF-κB driven inflammation reaction in cultured HCE-2 cells.Our results indicate that these harmful effects of ocular solutions preserved with PQ-1 should be further evaluated in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Institute of Clinical Medicine, University of Eastern Finland, Kuopio, Finland.

ABSTRACT

Purpose: In numerous clinical and experimental studies, preservatives present in eye drops have had detrimental effects on ocular epithelial cells. The aim of this study was to compare the cytotoxic and inflammatory effects of the preservative polyquaternium-1 (PQ-1) containing Travatan (travoprost 0.004%) and Systane Ultra eye drops with benzalkonium chloride (BAK) alone or BAK-preserved Xalatan (0.005% latanoprost) eye drops in HCE-2 human corneal epithelial cell culture.

Methods: HCE-2 cells were exposed to the commercial eye drops Travatan, Systane Ultra, Xalatan, and the preservative BAK. Cell viability was determined using colorimetric MTT (3-(4,5-dimethyldiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and by release of lactate dehydrogenase (LDH). Induction of apoptosis was measured with a using a colorimetric caspase-3 assay kit. DNA binding of the nuclear factor kappa B (NF-κB) transcription factor, and productions of the proinflammatory cytokines, interleukins IL-6 and IL-8, were determined using an enzyme-linked immunosorbent assay (ELISA) method.

Results: Cell viability, as measured by the MTT assay, declined by up to 50% after exposure to Travatan or Systane Ultra solutions which contain 0.001% PQ-1. BAK at 0.02% rather than at 0.001% concentration evoked total cell death signs on HCE-2 cells. In addition, cell membrane permeability, as measured by LDH release, was elevated by sixfold with Travatan and by a maximum threefold with Systane Ultra. Interestingly, Travatan and Systane Ultra activated NF-κB and elevated the secretion of inflammation markers IL-6 by 3 to eightfold and IL-8 by 1.5 to 3.5 fold, respectively, as analyzed with ELISA.

Conclusions: Eye drops containing PQ-1 evoke cytotoxicity and enhance the NF-κB driven inflammation reaction in cultured HCE-2 cells. Our results indicate that these harmful effects of ocular solutions preserved with PQ-1 should be further evaluated in vitro and in vivo.

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Related in: MedlinePlus

Level of cytotoxicity in HCE-2 cells analyzed by MTT assay. Columns represent the viability of cells (mean±SD. The viability of control cells is set as 100%. One-Way ANOVA, followed by Dunnett’s post hoc test, evaluated the statistical differences (n=6, *0.01<p≤0.05, **0.001<p≤0.01, ***p≤0.001, ns=not significant). Experiments were repeated three times.
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f1: Level of cytotoxicity in HCE-2 cells analyzed by MTT assay. Columns represent the viability of cells (mean±SD. The viability of control cells is set as 100%. One-Way ANOVA, followed by Dunnett’s post hoc test, evaluated the statistical differences (n=6, *0.01<p≤0.05, **0.001<p≤0.01, ***p≤0.001, ns=not significant). Experiments were repeated three times.

Mentions: MTT analysis revealed the extensive toxicity associated with BAK 0.02% containing Xalatan and BAK 0.02% (Figure 1). Both solutions evoked total cell death even after 5 min of exposure (followed by 24 h recovery in normal medium). At the same time point, the viability of cells exposed to Travatan, Systane Ultra and BAK 0.001% was near to the control. At later time points (15 and 30 min exposure + 24 h recovery) Travatan, Systane Ultra, and BAK 0.001% exposures reduced cellular viability in a time dependent manner. Based on the MTT analysis results and the extensive protein precipitation caused by the higher concentration of BAK (0.02%) and Xalatan, they were excluded from further experiments.


The preservative polyquaternium-1 increases cytoxicity and NF-kappaB linked inflammation in human corneal epithelial cells.

Paimela T, Ryhänen T, Kauppinen A, Marttila L, Salminen A, Kaarniranta K - Mol. Vis. (2012)

Level of cytotoxicity in HCE-2 cells analyzed by MTT assay. Columns represent the viability of cells (mean±SD. The viability of control cells is set as 100%. One-Way ANOVA, followed by Dunnett’s post hoc test, evaluated the statistical differences (n=6, *0.01<p≤0.05, **0.001<p≤0.01, ***p≤0.001, ns=not significant). Experiments were repeated three times.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3351434&req=5

f1: Level of cytotoxicity in HCE-2 cells analyzed by MTT assay. Columns represent the viability of cells (mean±SD. The viability of control cells is set as 100%. One-Way ANOVA, followed by Dunnett’s post hoc test, evaluated the statistical differences (n=6, *0.01<p≤0.05, **0.001<p≤0.01, ***p≤0.001, ns=not significant). Experiments were repeated three times.
Mentions: MTT analysis revealed the extensive toxicity associated with BAK 0.02% containing Xalatan and BAK 0.02% (Figure 1). Both solutions evoked total cell death even after 5 min of exposure (followed by 24 h recovery in normal medium). At the same time point, the viability of cells exposed to Travatan, Systane Ultra and BAK 0.001% was near to the control. At later time points (15 and 30 min exposure + 24 h recovery) Travatan, Systane Ultra, and BAK 0.001% exposures reduced cellular viability in a time dependent manner. Based on the MTT analysis results and the extensive protein precipitation caused by the higher concentration of BAK (0.02%) and Xalatan, they were excluded from further experiments.

Bottom Line: Interestingly, Travatan and Systane Ultra activated NF-κB and elevated the secretion of inflammation markers IL-6 by 3 to eightfold and IL-8 by 1.5 to 3.5 fold, respectively, as analyzed with ELISA.Eye drops containing PQ-1 evoke cytotoxicity and enhance the NF-κB driven inflammation reaction in cultured HCE-2 cells.Our results indicate that these harmful effects of ocular solutions preserved with PQ-1 should be further evaluated in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Institute of Clinical Medicine, University of Eastern Finland, Kuopio, Finland.

ABSTRACT

Purpose: In numerous clinical and experimental studies, preservatives present in eye drops have had detrimental effects on ocular epithelial cells. The aim of this study was to compare the cytotoxic and inflammatory effects of the preservative polyquaternium-1 (PQ-1) containing Travatan (travoprost 0.004%) and Systane Ultra eye drops with benzalkonium chloride (BAK) alone or BAK-preserved Xalatan (0.005% latanoprost) eye drops in HCE-2 human corneal epithelial cell culture.

Methods: HCE-2 cells were exposed to the commercial eye drops Travatan, Systane Ultra, Xalatan, and the preservative BAK. Cell viability was determined using colorimetric MTT (3-(4,5-dimethyldiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and by release of lactate dehydrogenase (LDH). Induction of apoptosis was measured with a using a colorimetric caspase-3 assay kit. DNA binding of the nuclear factor kappa B (NF-κB) transcription factor, and productions of the proinflammatory cytokines, interleukins IL-6 and IL-8, were determined using an enzyme-linked immunosorbent assay (ELISA) method.

Results: Cell viability, as measured by the MTT assay, declined by up to 50% after exposure to Travatan or Systane Ultra solutions which contain 0.001% PQ-1. BAK at 0.02% rather than at 0.001% concentration evoked total cell death signs on HCE-2 cells. In addition, cell membrane permeability, as measured by LDH release, was elevated by sixfold with Travatan and by a maximum threefold with Systane Ultra. Interestingly, Travatan and Systane Ultra activated NF-κB and elevated the secretion of inflammation markers IL-6 by 3 to eightfold and IL-8 by 1.5 to 3.5 fold, respectively, as analyzed with ELISA.

Conclusions: Eye drops containing PQ-1 evoke cytotoxicity and enhance the NF-κB driven inflammation reaction in cultured HCE-2 cells. Our results indicate that these harmful effects of ocular solutions preserved with PQ-1 should be further evaluated in vitro and in vivo.

Show MeSH
Related in: MedlinePlus