The Impact of Incubation Conditions on In Vitro Phosphatidylcholine Deposition on Contact Lens Materials.

SIGNIFICANCE: Previous in vitro measurements of contact lenses commonly investigate the impact of nonpolar tear film lipids (i.e., sterols). Polar lipids, however, are equally important stabilizing components of the tear film. This research explores and presents further knowledge about various aspects of polar lipid uptake that may impact contact lens performance. PURPOSE: This study evaluated the impact of incubation time, lipid concentration, and replenishment of an artificial tear solution (ATS) on the uptake of phosphatidylcholine (PC) onto conventional hydrogel (CH) and silicone hydrogel (SH) contact lens materials. METHODS: Four SHs and two CH lens materials (n = 4) were soaked in a complex ATS containing radioactive 14C-PC as a probe molecule. Phosphatidylcholine uptake was monitored at various incubation time points (1, 3, 7, 14, and 28 days), with different ATS lipid concentrations (0.5×, 1×, 2×) and with and without regular replenishment of the ATS. Phosphatidylcholine was extracted from the lenses, processed, and counted by a ß counter, and accumulated PC (µg/lens) was extrapolated from standard lipid calibration curves. RESULTS: All materials exhibited increasing PC deposition over time. Conventional hydrogel materials showed significantly lower PC uptake rates (P < .001) than any of the SH materials. Increasing lipid concentration in the ATS resulted in increased PC binding onto the contact lens materials (P < .001). Replenishing the ATS every other day, however, impacted the PC deposition differently, showing increased binding (P < .001) on CHs and reduced PC deposition for SH materials (P < .001). CONCLUSIONS: Length of incubation, lipid concentration in the ATS, and renewal of the incubation solution all influenced the amount of PC that sorbed onto various lens materials and therefore need to be considered when conducting future in vitro deposition studies.

Efficacy of Contact Lens Care Solutions in Removing Cholesterol Deposits From Silicone Hydrogel Contact Lenses.

PURPOSE: To determine the efficacy of multipurpose solutions (MPSs) on the removal of cholesterol deposits from silicone hydrogel (SH) contact lens materials using an in vitro model. MATERIALS AND METHODS: Five SH lens materials: senofilcon A, comfilcon A, balafilcon A, lotrafilcon A, and lotrafilcon B were removed from the blister pack (n=4 for each lens type), incubated for 7 days at 37°C in an artificial tear solution containing C radiolabeled cholesterol. Thereafter, lenses were stored in a preserved saline solution control (Sensitive Eyes Saline Plus) or cleaned with 1 of the 5 MPSs incorporating different preservatives (POLYQUAD/ALDOX, polyquaternium-1/alexidine, polyquaternium-1/PHMB, and 2 based on PHMB alone) using a rub and rinse technique, according to the manufacturer's recommendations, and stored in the MPS for a minimum of 6 hr. Lenses were then extracted with 2:1 chloroform:methanol, analyzed in a beta counter, and µg/lens of cholesterol was determined. RESULTS: Balafilcon A and senofilcon A lens materials showed the highest amounts of accumulated cholesterol (0.93±0.02 µg/lens; 0.95±0.01 µg/lens, respectively), whereas lotrafilcon A and lotrafilcon B deposited the lowest amounts (0.37±0.03 µg/lens; 0.47±0.12 µg/lens, respectively). For all lens materials, the MPS preserved with POLYQUAD/ALDOX removed more deposited cholesterol than any other test solution; however, the amount of removed cholesterol contamination from the individual contact lenses was only statistically significant for balafilcon A and senofilcon A (P=0.006 and P=0.042, respectively). Sensitive eyes and the other evaluated MPSs showed no significant effect on cholesterol removal (P>0.05). CONCLUSION: Cholesterol-removal efficacy varies depending on the combination of lens material and solution. Only 1 MPS showed a statistically significant reduction of cholesterol deposit for only 2 of the 5 tested lens materials.

Depth Profile Assessment of the Early Phase Deposition of Lysozyme on Soft Contact Lens Materials Using a Novel In Vitro Eye Model.

OBJECTIVE: To characterize the location of fluorescently labeled lysozyme on commercial contact lenses (CLs) using an in vitro eye model that simulates tear volume, tear flow, air exposure, and mechanical wear. METHODS: One commercially available conventional hydrogel CL material (etafilcon A) and three silicone hydrogel CL materials (balafilcon A, lotrafilcon B, and senofilcon A) were evaluated in this study. The CLs were mounted on the in vitro eye model and exposed to artificial tear fluid containing fluorescein isothiocyanate (FITC)-labeled lysozyme for 2 and 10 hrs. After these short incubation periods, circular discs were punched from the CLs at the center and periphery and were prepared for confocal laser scanning microscopy (CLSM). The CLSM captured a series of consecutive images spaced 5 µm apart, and the resulting images were rendered into two dimensional cross-sectional views of the CL. The mean fluorescence at each 5 µm slice was used to generate a histogram depicting the penetration of FITC-lysozyme into CLs. RESULTS: For both incubation periods, the CLSM images and histogram of etafilcon A showed that FITC-lysozyme is more concentrated at the lens surface, with a moderate amount of deposition in the lens matrix. For balafilcon A, FITC-lysozyme was evenly distributed throughout the lens. For lotrafilcon B, there was a greater amount of FITC-lysozyme deposition on the surfaces of the lens versus the matrix. Senofilcon A had differential FITC-lysozyme distribution profiles depending on the location of the lens. At the lens periphery, FITC-lysozyme primarily deposited on the surface, whereas FITC-lysozyme was uniformly distributed at the center of the lens. CONCLUSIONS: With the use of a sophisticated in vitro eye model, the study revealed a complex deposition pattern of FITC-labeled lysozyme on various CL materials after short periods of exposure. An understanding of the early deposition pattern of lysozyme on different CL material may elucidate new insights into the processes behind CL discomfort.

In Vitro Cholesterol Deposition on Daily Disposable Contact Lens Materials.

PURPOSE: The goal of this study was to analyze how various incubation times affect the uptake of cholesterol on silicone hydrogel (SH) and conventional hydrogel (CH) daily disposable (DD) contact lens materials using an in vitro radiochemical detection method. METHODS: Three SH (somofilcon A, delefilcon A, and narafilcon A) and four CH (etafilcon A, nesofilcon A, ocufilcon A, and nelfilcon A) contact lenses were incubated in an artificial tear solution that contained major tear film components and a portion of radioactive C-cholesterol. Lenses (N = 4) were incubated for four incubation times (2, 6, 12, or 16 h) to assess the effects on cholesterol deposition. Subsequent to the incubation, the lenses were extracted using 2:1 chloroform:methanol, and the extracts were analyzed in a beta counter and (in nanograms per lens) extrapolated from standard curves. RESULTS: In general, cholesterol deposited statistically significantly more on SH lenses than CHs (p ≤ 0.033), with the exception of somofilcon A and nesolfilcon A materials (p = 0.067). Within the SH materials, narafilcon A accumulated the largest quantity of cholesterol (p < 0.05) and somofilcon A the lowest (p < 0.05). The uptake of cholesterol ranged from 22.63 ± 2.98 ng/lens to 97.94 ± 4.18 ng/lens for all lens materials. The accumulation of cholesterol was shown to be continuous throughout the 16 h of incubation, without reaching a plateau (p < 0.001). CONCLUSIONS: For the periods that DD lens materials are worn, cholesterol deposits significantly more onto SH contact lenses than CHs. This could have implications for wearers who have higher levels of lipid in their tears that are fitted with SH DD materials.

Factors that influence in vitro cholesterol deposition on contact lenses.

PURPOSE: The purpose of this study was to analyze the impact that incubation time, lipid concentration, and solution replenishment have on silicone hydrogel (SiHy) and conventional hydrogel (CH) contact lens cholesterol deposition via in vitro radiochemical experiments. METHODS: Four SiHy (senofilcon A, lotrafilcon B, comfilcon A, balafilcon A) and two CH (etafilcon A and omafilcon A) contact lenses were incubated in an artificial tear solution (ATS) that contained major tear film proteins, lipids, salts, salts, and a trace amount of radioactive C-cholesterol. Lenses were incubated for various incubation times (1, 3, 7, 14, or 28 days), with three concentrations of lipid (0.5×, 1×, 2× tear film concentration) and with or without solution replenishment to assess each variable's impact on cholesterol deposition. After incubation, the lenses were extracted using 2:1 chloroform:methanol, extracts were analyzed in a beta counter and masses (micrograms per lens) were extrapolated from standard curves. RESULTS: Within the SiHy materials, balafilcon A deposited the greatest amount of cholesterol (p < 0.001) and lotrafilcon B the lowest (p < 0.001). The CH lens materials showed significantly lower uptake amounts than any of the SiHy lens materials (p < 0.001). The uptake of cholesterol ranged from 0.01 ± 0.01 µg/lens to 3.22 ± 0.34 µg/lens for all lens materials. Kinetic uptake of cholesterol was shown to be continuous throughout the 28 days of incubation without plateau (p < 0.001), and varying the lipid concentration did impact the resulting cholesterol deposition (p < 0.001). Replenishing the ATS every other day also affected cholesterol deposition throughout the experiment. Overall, the deposition pattern was 2× > replenishing > 1× > 0.5×. CONCLUSIONS: Overall, SiHy lenses deposit significantly more cholesterol than CH lens materials, and the mass of lipid deposited is dependent on the contact lens material, length of incubation, concentration of lipids in the ATS, and the replenishment of ATS.

Development of an In Vitro Blink Model for Ophthalmic Drug Delivery.

PURPOSE: The purpose of this study was to develop an advanced in vitro blink model that can be used to examine the release of a wide variety of components (for example, topical ophthalmic drugs, comfort-inducing agents) from soft contact lenses. METHODS: The model was designed using computer-aided design software and printed using a stereolithography 3D printer. The eyelid and eyeball were synthesized from polyvinyl alcohol and silicone material, respectively. Simulated tear fluid was infused through tubing attached to the eyelid using a syringe pump. With each blink cycle, the eyelid slides and flexes across the eyeball to create an artificial tear film layer. The flow-through fluid was collected using a specialized trough. Two contact lenses, etafilcon A and senofilcon A, were incubated in 2 mL of a water-soluble red dye for 24 h and then placed on the eye model (n = 3). The release of the dye was measured over 24 h using a tear flow rate of 5 µL/min. RESULTS: Approximately 25% of the fluid that flowed over the eye model was lost due to evaporation, nonspecific absorption, and residual dead volume. Senofilcon A absorbed more dye (47.6 ± 2.7 µL) than etafilcon A (22.3 ± 2.0 µL). For etafilcon A, the release of the dye followed a burst-plateau profile in the vial but was sustained in the eye model. For senofilcon A, the release of the dye was sustained in both the vial and the eye model, though more dye was released in the vial (p < 0.05). Overall, the release of the dye from the contact lenses was higher in the vial compared with the eye model (p < 0.05). CONCLUSION: The blink model developed in this study could be used to measure the release of topical ophthalmic drugs or comfort agents from contact lenses. Simulation of a blink mechanism, an artificial tear film, and nonspecific absorption in an eye model may provide better results than a simple, static vial incubation model.

Effects of Temperature and Blinking on Contact Lens Dehydration of Contemporary Soft Lens Materials Using an In Vitro Blink Model.

Purpose: The purpose of this study was to evaluate the effects of temperature and blinking on contact lens (CL) dehydration using an in vitro blink model. Methods: Three silicone hydrogel (delefilcon A, senofilcon A, and comfilcon A) and two conventional hydrogel (etafilcon A and omafilcon A) CL materials were evaluated at 1 and 16 hours. The water content (WC) of the CLs was measured using a gravimetric method. Lenses were incubated on a blink model, internally heated to achieve a clinically relevant surface temperature of 35°C. An artificial tear solution (ATS) was delivered to the blink model at 4.5 µL/min with a blink rate of 6 blinks/min. A comparison set of lenses were incubated in a vial containing either 2 mL of ATS or phosphate-buffered saline (PBS) at 35°C. Results: Increasing temperature to 35°C resulted in a decrease in WC for all tested CLs over time (P ≤ 0.0052). For most CLs, there was no significant difference in WC over time between ATS or PBS in the vial (P > 0.05). With the vial system, WC decreased and plateaued over time. However, on the blink model, for most CLs, the WC significantly decreased after 1 hour but returned toward initial WC levels after 16 hours (P > 0.05). Conclusions: The reduction in WC of CLs on the eye is likely due to both an increase in temperature and dehydration from air exposure and blinking. Translational Relevance: This study showed that the novel, heated, in vitro blink model could be used to provide clinical insights into CL dehydration on the eye.

Novel in vitro method to determine pre-lens tear break-up time of hydrogel and silicone hydrogel contact lenses.

PURPOSE: To develop an in vitro model to determine pre-lens non-invasive break-up time (NIBUT) and to subsequently use this method to compare the NIBUT over contemporary daily disposable (DD) contact lenses (CL). METHODS: Three silicone hydrogel (SH) and two conventional hydrogel (CH) DD CLs were incubated in an artificial tear solution (ATS). A model blink cell (MBC) was utilised to mimic intermittent air exposure. CLs were repeatedly submerged for 3 seconds (s) and exposed to air for 10 s over periods of 2, 6, 12, and 16 hours (h). NIBUTs (n = 4) were determined out of the blister pack (T0) and at the end of each incubation period. RESULTS: Overall, nesofilcon A showed the longest NIBUTs (p < 0.001). At T0, CHs revealed significantly longer NIBUTs (p ≤ 0.001) than SHs. After 2 h, nesofilcon A showed the longest NIBUT, however, this was only statistically significant compared with delefilcon A (p ≤ 0.001). After 6 h, nesofilcon A NIBUT was significantly longer than all other CLs (p ≤ 0.001). Etafilcon A showed a significantly longer NIBUT (p ≤ 0.001) after 12 h and delefilcon A had the longest NIBUT (p ≤ 0.001) after 16 h. Statistically significant (p ≤ 0.05) changes of NIBUT within the lens materials varied between time points. After 16 h, all CLs showed significant reductions in NIBUTs (p ≤ 0.001) in comparison to T0. CONCLUSION: NIBUT values reduced gradually over time and varying levels of deposition impacted measured pre-lens NIBUTs. While NIBUT of CH materials are longer immediately out of the blister pack, after tear film exposure, the NIBUTs obtained using this methodology became very similar.

Analysis of polyvinyl alcohol release from commercially available daily disposable contact lenses using an in vitro eye model.

The purpose of this work was to determine the release of polyvinyl alcohol (PVA) from etafilcon A, omafilcon A, and nelfilcon A daily disposable hydrogel contact lenses using a novel in vitro model. PVA is an ocular lubricant that can be found in multiple formulations of artificial tears. Nelfilcon A innately contains PVA, so only the release of PVA from this lens was evaluated. Etafilcon A and omafilcon A lenses were incubated in a PBS solution containing PVA. The release of PVA was evaluated using a novel in vitro blink platform with Milli-Q water and PBS under various blink conditions and flow rates. Nelfilcon A lenses significantly released more PVA than other lenses at 0.5 and 1.5 h in both PBS and Milli-Q water (p < 0.001). For nelfilcon A, there was no statistical significance between the release profiles of PVA between the blink and no-blink conditions, or for the various flow rates (p > 0.05). All tested groups and lenses showed a burst release within the first 4.5 h and rapidly plateaued thereafter. The current study demonstrates that releasable PVA (whether through uptake or through being inherently available from the material) is loosely bound on hydrogel lenses, and the majority is released within 4.5 h. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1662-1668, 2019.

Differential Deposition of Fluorescently Tagged Cholesterol on Commercial Contact Lenses Using a Novel In Vitro Eye Model.

PURPOSE: We evaluate the differences in lipid uptake and penetration in daily disposable (DD) contact lenses (CL) using a conventional "in-vial" method compared to a novel in vitro eye model. METHODS: The penetration of fluorescently labelled 22-(N-(7-Nitrobenz-2-Oxa-1,3-Diazol-4-yl)Amino)-23,24-Bisnor-5-Cholen-3beta-Ol (NBD)-cholesterol on three silicone hydrogel (SH) and four conventional hydrogel (CH) DD CLs were investigated. CLs were incubated for 4 and 12 hours in a vial, containing 3.5 mL artificial tear solution (ATS), or were mounted on an in vitro eye-blink platform designed to simulate physiologic tear flow (2 mL/24 hours), tear volume and "simulated" blinking. Subsequently, CLs were analyzed using laser scanning confocal microscopy and ImageJ. RESULTS: Penetration depth and fluorescence intensities of NBD-cholesterol varied between the incubation methods as well as lens materials. Using the traditional vial incubation method, NBD-cholesterol uptake occurred equally on both sides of all lens materials. However, using our eye-blink model, cholesterol penetration was observed primarily on the anterior surface of the CLs. In general, SH lenses showed higher intensities of NBD-cholesterol than CH materials. CONCLUSIONS: The traditional "in-vial" incubation method exposes the CLs to an excessively high amount of ATS, which results in an overestimation for cholesterol deposition. Our model, which incorporates important ocular factors, such as intermittent air exposure, small tear volume, and physiological tear flow between blinks, provides a more natural environment for in vitro lens incubation. TRANSLATIONAL RELEVANCE: In vitro measurements of CLs are a common approach to predict their interactions and performance on the eye. Traditional methods, however, are rudimentary. Therefore, this study presents a novel in vitro model to evaluate CLs, which consequently will enhance elucidations of the interactions between CLs and the eye.

Determination of the release of PEG and HPMC from nelfilcon A daily disposable contact lenses using a novel in vitro eye model.

The traditional method to measure release of components from CLs is a vial containing a static volume of PBS (phosphate buffered saline). However, this model does not simulate physiologically relevant tear volume and natural tear flow, air exposure, and mechanical rubbing. These factors can significantly impact release kinetics. We have developed an in vitro eye model (OcuFlow) that simulates these parameters. The aim of the study was to measure the release of PEG (polyethylene glycol), and HPMC (hydroxypropyl methylcellulose) from a daily disposable hydrogel contact lens material (nelfilcon A; Dailies AquaComfort PLUS; DACP;) over 24 hrs using the OcuFlow platform. The elution of PEG and HPMC from DACP lenses was analyzed using LCMS (liquid chromatography mass spectrometry). The release of all wetting agents from the lenses followed a burst release pattern, which occurred within the first 1.5 hrs (P < 0.05). The release of PEG was greater than that of HPMC (P < 0.05). The amount of PEG and HPMC released at any given time was less than 1% of the amount in the blister pack solution. Our results suggest that HPMC and PEG are rapidly released from the CL.

Development of an In Vitro Ocular Platform to Test Contact Lenses.

Currently, in vitro evaluations of contact lenses (CLs) for drug delivery are typically performed in large volume vials, which fail to mimic physiological tear volumes. The traditional model also lacks the natural tear flow component and the blinking reflex, both of which are defining factors of the ocular environment. The development of a novel model is described in this study, which consists of a unique 2-piece design, eyeball and eyelid piece, capable of mimicking physiological tear volume. The models are created from 3-D printed molds (Polytetrafluoroethylene or Teflon molds), which can be used to generate eye models from various polymers, such as polydimethylsiloxane (PDMS) and agar. Further modifications to the eye pieces, such as the integration of an explanted human or animal cornea or human corneal construct, will permit for more complex in vitro ocular studies. A commercial microfluidic syringe pump is integrated with the platform to emulate physiological tear secretion. Air exposure and mechanical wear are achieved using two mechanical actuators, of which one moves the eyelid piece laterally, and the other moves the eyeballeyepiece circularly. The model has been used to evaluate CLs for drug delivery and deposition of tear components on CLs.