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Corneal epithelial cell biocompatibility to silicone hydrogel and conventional hydrogel contact lens packaging solutions.

Gorbet MB, Tanti NC, Jones L, Sheardown H - Mol. Vis. (2010)

Bottom Line: Incubation of HCECs with CLs stored in borate-buffered packaging solutions resulted in a significant reduction in cell viability.However, a significant decrease in reactive oxygen species was observed at 24 h.When used in ophthalmic packaging solutions, the antimicrobial properties of borate buffer may be outweighed by its relatively cytotoxic effects on cells.

View Article: PubMed Central - PubMed

Affiliation: Systems Design Engineering, University of Waterloo, Waterloo, Ontario, Canada. mgorbet@uwaterloo.ca

ABSTRACT

Purpose: Although all contact lenses (CLs) are applied initially to the eye directly from a packaging solution, little is known about the effects of these solutions on human corneal epithelial cells (HCECs). Due to the porous nature of CL materials, they have the potential to sorb components of the packaging solution during storage, which could then be subsequently released upon insertion of the CL on the eye. The purpose of this study was to investigate the effect of various packaging solutions on HCECs, using an in vitro model.

Methods: An in vitro assay was developed whereby various silicone hydrogels and conventional, poly-2-hydroxyethylmethacrylate (polyHEMA)-based lens materials were removed directly from their packaging and then incubated for up to 24 h with HCECs. The effect of the retained and released packaging solution components on HCECs was assessed by measuring cell viability, adhesion phenotype, and apoptosis.

Results: Incubation of HCECs with CLs stored in borate-buffered packaging solutions resulted in a significant reduction in cell viability. Adherent cells incubated with these CLs also exhibited reduced levels of beta(1) and alpha(3) integrin. Soaking borate-buffered packaged CLs in PBS before cell incubation resolved viability and integrin expression in all cases, with the exception of galyfilcon A and balafilcon A, from which a 20% reduction in cell viability was still observed. In comparison, CLs stored in phosphate-buffered packaging solutions had cellular viability and expression of integrins similar to control cells (cells incubated in the absence of a lens). When incubated with cells at a 10% concentration in serum-free medium, borate-buffered packaging solutions and borate-containing saline (Unisol 4) significantly reduced cell viability and integrin expression. Neither caspase activation nor annexin V binding was observed on cells following exposure to borate buffer solution. However, a significant decrease in reactive oxygen species was observed at 24 h. These latter results suggest that in vitro exposure to low concentration of borate/boric acid results in cell dysfunction, leading to necrosis rather than apoptosis.

Conclusions: Borate-buffered packaging solutions were shown to adversely affect the viability and integrin expression of HCECs in vitro. When used in ophthalmic packaging solutions, the antimicrobial properties of borate buffer may be outweighed by its relatively cytotoxic effects on cells.

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Related in: MedlinePlus

Caspase activation following incubation with phosphate or borate-buffered solution. No increase in cells staining positive for FITC-VAD-FMK was observed after cells were incubated for 24 h with either phosphate, borate-buffered solutions, or soaked lenses. Dot plots are representative of the three experiments that were performed.
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f4: Caspase activation following incubation with phosphate or borate-buffered solution. No increase in cells staining positive for FITC-VAD-FMK was observed after cells were incubated for 24 h with either phosphate, borate-buffered solutions, or soaked lenses. Dot plots are representative of the three experiments that were performed.

Mentions: As shown in Figure 4, caspase activation, detected using FITC-VAD-FMK, was not observed in cells incubated for 24 h with borate buffer-containing solutions. The percentage of cells staining positive for caspase activity (5±2%) was similar for cells grown in the absence of solution, or in the presence of PBS or Unisol. This was done with concentrations of up to 0.1% borate/boric acid (i.e., 20% Unisol), which were tested at 24 h; caspase activation remained within control levels. After 24 h exposure to diluted borate solutions, there was also no significant increase in binding of annexinV-FITC between cells exposed to diluted borate or phosphate solutions and control cells (cells incubated in the absence of solution and contact lens). For all samples, less than 2% of cells were found to bind Annexin V. In the presence of borate solution, a small but not significant increase in YO-PRO-1 permeability was observed (Table 7). To further characterize the potential mechanisms of borate-induced cytotoxicity, the intracellular levels of reactive oxygen species (ROS) were measured. As shown in Figure 5, a significant underproduction of DCF-A was observed for cells that had been exposed to borate-buffered solution for 24 h, either from products released from a lens or from direct dilution. A 25% reduction in fluorescence intensity of rhodamine 123 (Figure 5) was also observed with these samples. Upon phorbol myristate acetate (PMA) stimulation, all cells were able to increase their production of ROS (Table 8). As expected, the levels of PMA-induced ROS in cells exposed to diluted borate solutions for 24 h were lower than those of controls (since they had lower levels of ROS to start with). Only cells exposed to 0.1% borate/boric acid (20% unisol) for 24 h showed an impaired response in PMA-induced ROS formation. All other cells exhibited 50% increases in ROS production, suggesting that while they had an impaired level of intracellular ROS after 24 h-exposure to borate-buffered solution, they were still able to respond to PMA stimulus.


Corneal epithelial cell biocompatibility to silicone hydrogel and conventional hydrogel contact lens packaging solutions.

Gorbet MB, Tanti NC, Jones L, Sheardown H - Mol. Vis. (2010)

Caspase activation following incubation with phosphate or borate-buffered solution. No increase in cells staining positive for FITC-VAD-FMK was observed after cells were incubated for 24 h with either phosphate, borate-buffered solutions, or soaked lenses. Dot plots are representative of the three experiments that were performed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2823797&req=5

f4: Caspase activation following incubation with phosphate or borate-buffered solution. No increase in cells staining positive for FITC-VAD-FMK was observed after cells were incubated for 24 h with either phosphate, borate-buffered solutions, or soaked lenses. Dot plots are representative of the three experiments that were performed.
Mentions: As shown in Figure 4, caspase activation, detected using FITC-VAD-FMK, was not observed in cells incubated for 24 h with borate buffer-containing solutions. The percentage of cells staining positive for caspase activity (5±2%) was similar for cells grown in the absence of solution, or in the presence of PBS or Unisol. This was done with concentrations of up to 0.1% borate/boric acid (i.e., 20% Unisol), which were tested at 24 h; caspase activation remained within control levels. After 24 h exposure to diluted borate solutions, there was also no significant increase in binding of annexinV-FITC between cells exposed to diluted borate or phosphate solutions and control cells (cells incubated in the absence of solution and contact lens). For all samples, less than 2% of cells were found to bind Annexin V. In the presence of borate solution, a small but not significant increase in YO-PRO-1 permeability was observed (Table 7). To further characterize the potential mechanisms of borate-induced cytotoxicity, the intracellular levels of reactive oxygen species (ROS) were measured. As shown in Figure 5, a significant underproduction of DCF-A was observed for cells that had been exposed to borate-buffered solution for 24 h, either from products released from a lens or from direct dilution. A 25% reduction in fluorescence intensity of rhodamine 123 (Figure 5) was also observed with these samples. Upon phorbol myristate acetate (PMA) stimulation, all cells were able to increase their production of ROS (Table 8). As expected, the levels of PMA-induced ROS in cells exposed to diluted borate solutions for 24 h were lower than those of controls (since they had lower levels of ROS to start with). Only cells exposed to 0.1% borate/boric acid (20% unisol) for 24 h showed an impaired response in PMA-induced ROS formation. All other cells exhibited 50% increases in ROS production, suggesting that while they had an impaired level of intracellular ROS after 24 h-exposure to borate-buffered solution, they were still able to respond to PMA stimulus.

Bottom Line: Incubation of HCECs with CLs stored in borate-buffered packaging solutions resulted in a significant reduction in cell viability.However, a significant decrease in reactive oxygen species was observed at 24 h.When used in ophthalmic packaging solutions, the antimicrobial properties of borate buffer may be outweighed by its relatively cytotoxic effects on cells.

View Article: PubMed Central - PubMed

Affiliation: Systems Design Engineering, University of Waterloo, Waterloo, Ontario, Canada. mgorbet@uwaterloo.ca

ABSTRACT

Purpose: Although all contact lenses (CLs) are applied initially to the eye directly from a packaging solution, little is known about the effects of these solutions on human corneal epithelial cells (HCECs). Due to the porous nature of CL materials, they have the potential to sorb components of the packaging solution during storage, which could then be subsequently released upon insertion of the CL on the eye. The purpose of this study was to investigate the effect of various packaging solutions on HCECs, using an in vitro model.

Methods: An in vitro assay was developed whereby various silicone hydrogels and conventional, poly-2-hydroxyethylmethacrylate (polyHEMA)-based lens materials were removed directly from their packaging and then incubated for up to 24 h with HCECs. The effect of the retained and released packaging solution components on HCECs was assessed by measuring cell viability, adhesion phenotype, and apoptosis.

Results: Incubation of HCECs with CLs stored in borate-buffered packaging solutions resulted in a significant reduction in cell viability. Adherent cells incubated with these CLs also exhibited reduced levels of beta(1) and alpha(3) integrin. Soaking borate-buffered packaged CLs in PBS before cell incubation resolved viability and integrin expression in all cases, with the exception of galyfilcon A and balafilcon A, from which a 20% reduction in cell viability was still observed. In comparison, CLs stored in phosphate-buffered packaging solutions had cellular viability and expression of integrins similar to control cells (cells incubated in the absence of a lens). When incubated with cells at a 10% concentration in serum-free medium, borate-buffered packaging solutions and borate-containing saline (Unisol 4) significantly reduced cell viability and integrin expression. Neither caspase activation nor annexin V binding was observed on cells following exposure to borate buffer solution. However, a significant decrease in reactive oxygen species was observed at 24 h. These latter results suggest that in vitro exposure to low concentration of borate/boric acid results in cell dysfunction, leading to necrosis rather than apoptosis.

Conclusions: Borate-buffered packaging solutions were shown to adversely affect the viability and integrin expression of HCECs in vitro. When used in ophthalmic packaging solutions, the antimicrobial properties of borate buffer may be outweighed by its relatively cytotoxic effects on cells.

Show MeSH
Related in: MedlinePlus