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Optometry & Vision Science:
doi: 10.1097/OPX.0b013e31826c6508
Original Articles

The Impact of Intermittent Air Exposure on Lipid Deposition

Lorentz, Holly*; Heynen, Miriam; Khan, Warda; Trieu, Diana; Jones, Lyndon§

Free Access
Press Release
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Author Information

*PhD

MSc

BSc

§PhD, FCOptom, FAAO

Centre for Contact Lens Research, School of Optometry and Vision Science, University of Waterloo, Waterloo, Ontario, Canada.

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Abstract

Purpose. To analyze the impact of intermittent air exposure on the in vitro deposition of two radioactive lipids on various contact lens (CL) materials, using a custom-designed model blink cell.

Methods. Six different CL materials (balafilcon A, lotrafilcon B, comfilcon A, senofilcon A, etafilcon A, and omafilcon A) were mounted on the model blink cell pistons, which cycled the lenses in and out of a complex artificial tear solution (ATS) that contained a trace amount of 14C-cholesterol or 14C-phosphatidylcholine. For the short-term experiment, air-exposed lenses were continuously cycled in and out of the ATS for 10 h. Longer term incubations for 6 days were tested with lotrafilcon B and balafilcon A materials incubated in 14C-cholesterol ATS. The air-exposed CLs were cycled for 14 h then submerged for 10 h each day. For both experiments, the control lenses were submerged for the entire test period. After incubation, lenses were processed, and deposited masses were quantified.

Results. Exposure to air resulted in increased amounts of cholesterol deposited by 1.6 to 4.3 fold on omafilcon A, balafilcon A, comfilcon A, and senofilcon A (p ≤ 0.03) compared with submerged lenses. No differences in deposition were observed for etafilcon A and lotrafilcon B (p > 0.05). The longer term incubation of lotrafilcon B and balafilcon A showed statistically significant increases in cholesterol deposition for both air-exposed lens materials (p < 0.02) with the increase in deposition 1.8× and 2.8×, respectively. For phosphatidylcholine, all air-exposed lenses had increased masses of deposition. These deposits were statistically greater by 1.1 to 1.6 times for omafilcon A, comfilcon A, lotrafilcon B, and senofilcon A (p < 0.04), but not statistically different for etafilcon A or balafilcon A (p > 0.05).

Conclusions. This study found that lipid deposition profiles are CL material dependent and that intermittent air exposure can influence the mass of lipid deposited.

In vitro models in contact lens research have been used previously to examine lipid deposition, protein deposition, protein denaturation, and interactions with tear film components.1–14 Many of the in vitro models used are simple model systems where an individual lipid or protein was examined in isolation, without the incorporation of the other prevalent tear film components.4,5,9,10,13,14 These in vitro models are too simplistic, and they ignore several key elements of human contact lens wear, including the complexity of the tear film, the exposure to air in-between blinks and the sheering force of the lid. Our laboratory, and others, have begun using a more complex artificial tear film solution containing mixtures of both lipids and proteins, in their in vitro models.2,3,15–21 However, few, if any, researchers have looked at the effect of air exposure or the lid effects on lipid or protein deposition.

The effect of lens drying in the interblink period is especially of interest in lipid deposition on silicone hydrogel contact lens materials, as these lenses are more hydrophobic and less wettable than conventional hydrogel materials. These lens properties are attributed to the silicone components of the contact lenses. Silicone is known for its superior ability to carry oxygen, which is ideal for daily and especially extended wear contact lens materials; however, silicone is known to be very hydrophobic in nature. This hydrophobicity makes the lens material more lipophilic and less wettable, which can lead to increased lipid deposition. In general, deposition of tear film components has been thought to cause lens discomfort and degrade visual clarity for the lens wearer.22–28

When a contact lens material is naturally hydrophobic and unwettable, the anterior tear film that covers the lens surface may not remain stable and intact for a long period. Therefore, the tear film may break, collapse, and recede across the surface of the lens before the next blink. This breaking of the tear film encourages the lipid layer of the tear film to come into direct contact with the lens material itself, which could increase the degree of lipid deposition. This process could be exacerbated by the fact that silicone hydrogels, much like their conventional hydrogel predecessors, have freely rotating polymer structures. This means that when the polymer is exposed directly to air it will rotate to expose its more hydrophobic backbone and “hide” the hydrophilic moieties toward the matrix. This may form an inward attraction for lipid to deposit during the interblink period.

When a blink occurs, the tear film will spread over the anterior surface of the contact lens and the hydrophilic polymer moieties flip back to the surface to come in contact with the aqueous phase of the tear film. Deposited lipid on the lens surface may then be encouraged to move further into the lens matrix to avoid the aqueous phase of the tear film and the hydrophilic moieties of the lens polymer. This is a dynamic process that may have a huge impact on the quantity of lipid, types of lipid, state of the lipid, and the mechanism of how the deposition onto and into contact lens materials occurs.

To test this effect of air exposure on lipid deposition using an in vitro model, a novel device was built by our laboratory. This device, called a “model blink cell” (MBC), was designed to mimic the interblink drying time that occurs during contact lens wear. This MBC was placed into an atmospherically controlled chamber, and mounted contact lenses were cycled in and out of an artificial tear solution (ATS). When the lenses are out of the ATS, the artificial tear film will “break” over the surface of the lens and allow for drying, to mimic human contact lens wear. This was completed and tested against an in vitro model where the contact lenses remained submerged in the ATS for the entire incubation time, without cycling, similar to traditional in vitro vial incubations.

In this pilot experiment, two different model lipids, 14C-cholesterol and 14C-phosphatidylcholine (PC), were examined for their deposition onto conventional and silicone hydrogel contact lens materials, using our specially designed MBC in either fully submerged or air exposure mode.

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METHODS

Artificial Tear Solution

The protocol to prepare an ATS solution specifically optimized for closed system or in-vial incubations has previously been published.20,21,29 However, this study involved an experiment that required incubation of contact lenses in an open-to-air system; therefore, an ATS was developed for open-system incubations, which was able to remain stable in pH and osmolality throughout the incubation period, when exposed to air. This ATS is similar in composition and preparation to the previously described system outlined in Table 1, and the changes from the original in-vial ATS components are italicized and remain limited to the complex salt solution. For this study, an ATS with one of two model radioactive lipids, 14C-cholesterol or 14C-phosphatidylcholine, were used as probe molecules to allow for their sensitive and accurate quantification. This technique has been previously optimized in our laboratory.20,21,29 Cholesterol and PC were chosen as model lipids for this study as they represent two structurally and functionally different species of lipids, which are both prevalent in the tear film and within contact lens deposits.30–37

Table 1
Table 1
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Preparing the radioactive open-system, ATS involves the same main steps as is required for in-vial incubations. This protocol is published elsewhere and is therefore not reiterated here.20,21 The 14C-cholesterol was added to the ATS at a concentration of 5.6% of the total cholesterol concentration, and the 14C-phosphatidylcholine was added at a concentration of 27% of the total PC concentration. The radioactive lipids were purchased from Perkin-Elmer (Woodbridge, ON, Canada), and their details can be found in Table 2.

Table 2
Table 2
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Contact Lens Materials

Six contact lens materials were examined: Acuvue 2 (etafilcon A; Vistakon), Acuvue OASYS (senofilcon A; Vistakon), Air Optix (lotrafilcon B; Ciba Vision), Biofinity (comfilcon A; CooperVision), Proclear (omafilcon A; CooperVision), and PureVision (balafilcon A; Bausch & Lomb). The material characteristics of all contact lens materials can be found in Table 3. Each contact lens material, incubation condition, incubation time, and lipid was examined in triplicate.

Table 3
Table 3
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Model Blink Cell

The MBC is a device specially designed and built in our laboratory to mimic the interblink period during contact lens wear (Fig. 1). The blink cell itself is composed of a trough with contact lenses mounted on six form-fitting Teflon pistons (Fig. 1) connected to a motor that cycles the pistons in and out of the ATS contained in the trough. The cycling of the pistons is programmable so that a desired blink frequency can be chosen. The MBC itself is contained within a temperature and humidity controlled chamber so that physiological parameters can be maintained during incubation.

Figure 1
Figure 1
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Experimental Design

All contact lens materials were soaked in saline with agitation for 24 h to remove the blister pack solution components. After presoaking, the lenses were blotted on lens paper before mounting onto the pistons in the MBC.

Two identical MBC units were presoaked with ATS without radioactivity for at least 48 h to reduce any non-specific binding to the MBC of lipid and proteins in subsequent incubations. When the contact lenses were ready for incubation, the ATS was replaced with fresh ATS containing a single radioactive lipid. Three replicates of two contact lens types were incubated in each unit. One unit cycled the lenses in and out of ATS (referred to as “air exposed”), and the lenses in the other unit remained submerged for the duration of the experiment (“submerged”). The blink rate was set such that the lenses were submerged for 2 s and then were out of solution and exposed to the atmosphere for 5 s. The atmosphere in the chamber was set at 36°C ± 1°C and had a relative humidity of 16 ± 1%. Both MBC units were filled with 18 mL of ATS solution, containing a single tracer of radioactive (cholesterol or PC). To compensate for solution evaporation, both units were refilled as needed, maintaining appropriate osmolality and final volume. Uniformity of the ATS between the units was achieved by pooling ATS from both units during the refill.

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Short-Term Incubation

The first experimental test mimicked wear of a daily disposable contact lens with one 10 h incubation period. This short-term test was conducted with both radioactive lipids and all lens materials. After the 10 h incubation in either the submerged or air-exposed MBC, the lenses were removed and processed as described in the section titled processing of contact lenses.

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Long-Term Incubation

The long-term incubation experiment was tested on balafilcon A and lotrafilcon B contact lens materials for 6 days to mimic extended wear conditions. Air exposed lenses in this experiment were cycled in and out of the solution for 14 h (to mimic wear time) and were then submerged for 10 h (to mimic the overnight wear). This cycle was repeated for each of the 6 days. Submerged lenses were simply submerged for the entire 6 days. Fresh radioactive ATS replaced the used solution every other day. Following the long-term incubations, the lenses were removed and processed as described in the next section.

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Processing of Contact Lenses

At the end of the incubation, the lenses were extracted with (2:1) chloroform: methanol, processed, and analyzed, as previously published.20 The data were corrected for extraction efficiency38 and radioactive signal in the ATS.

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Statistical Analysis

Statistical analysis was performed using Statistica 9 and independent t-tests. P values of <0.05 were considered to be statistically significant.

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RESULTS

Cholesterol

Exposure to air significantly increased (p ≤ 0.03) the amount of cholesterol deposited on most lenses (omafilcon A, balafilcon A, comfilcon A, senofilcon A), with lotrafilcon B and etafilcon A depositing lower (but statistically identical) amounts (Fig. 2). The order of cholesterol deposits in submerged lenses was omafilcon A = etafilcon A = lotrafilcon B = senofilcon A = comfilcon A < balafilcon A. Air exposure for cholesterol deposits occurred in the following order: etafilcon A < lotrafilcon B < senofilcon A < omafilcon A < comfilcon A < balafilcon A. Generally, the air exposed lenses deposited 1.6 to 4.3× more cholesterol than the submerged lenses, except for etafilcon A and lotrafilcon B materials, which deposited equal or slightly less cholesterol when exposed to air.

Figure 2
Figure 2
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The longer term incubation was tested for both balafilcon A and lotrafilcon B lenses. These specific lenses were chosen as they are both approved for at least 6 days of continuous wear, and they represented the two extremes of silicone hydrogel lens deposition in the short-term 10 h experiment, both in quantity of cholesterol deposited and in the effect of air exposure. After 6 days of incubation in the MBC, both lens materials exhibited a statistically significant increase in cholesterol deposition when exposed to air (p < 0.011) and both materials had deposited significantly more cholesterol in the long-term incubation experiment when compared with the 10 h experiment (Fig. 3). Once again, balafilcon A lenses deposited significantly more lipid than lotrafilcon B lenses, and overall, the air-exposed lenses deposited 2.8× more cholesterol on balafilcon A and 1.8× more cholesterol on lotrafilcon B materials.

Figure 3
Figure 3
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Phosphatidylcholine

For all lenses, air exposure resulted in higher levels of PC being deposited. These levels were statistically significantly higher (p < 0.04) for the neutral charge Food and Drug Administration (FDA) group I (lotrafilcon B, senofilcon A, comfilcon A) and II (omafilcon A) lens materials when exposed to air. However, no differences in deposition were found (p > 0.05) for the ionic FDA group III (balafilcon A) or IV (etafilcon A) lens materials (Fig. 4).

Figure 4
Figure 4
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The masses deposited were all low, reflecting the low concentration of PC in the solution. The relative increase of PC on air-exposed lenses is smaller than the effect seen with cholesterol, in the range of 17% more for senofilcon A and up to 60% more for omafilcon A. The order of deposition for submerged lenses was etafilcon A < omafilcon A < comfilcon A < lotrafilcon B < senofilcon A < balafilcon A. Exposing the lenses to air resulted in the following order of PC deposition: etafilcon A = omafilcon A < comfilcon A < senofilcon A = lotrafilcon B = balafilcon A.

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DISCUSSION

Examining lipid deposition on contact lenses in an in vitro model system helps researchers and developers to gain an understanding of the processes and interactions that occur in vivo. Our laboratory has developed an MBC to simulate the interblink tear film thinning and drying that occurs during the interblink period of contact lens wear. This model has demonstrated that a change in the kinetics of lipid deposition occurs when incubating contact lenses using this intermittent air-exposure method versus incubating lenses in a fully submerged aqueous environment.

This experiment specifically found that the effect of intermittent air exposure on lipid deposition is both lipid and lens material dependent. For example, etafilcon A lens materials did not differ from control values when this material was exposed to air during the blink cycle for either lipid examined. However, omafilcon A, senofilcon A, and comfilcon A, all deposited significantly more cholesterol and PC when incubated with intermittent air exposure. Balafilcon A, showed no statistical differences in deposition with PC but did with cholesterol. In contrast, lotrafilcon B had the opposite result with the 10 h incubation, but when lotrafilcon B was incubated to mimic 6 days of continuous wear, significantly higher cholesterol deposition was found.

Cholesterol deposition increased for all lens materials when intermittently exposed to air, with the exception of etafilcon A. Overall, PC deposition increased on all lens materials when exposed to air; however, only four of the six materials had statistically significant differences. Once again, etafilcon A showed no statistically significant differences when exposed to air. Etafilcon A is well known as a material that deposits little amounts of lipid but deposits large amounts of lysozyme, due to its hydrophilic hydroxyl moieties.13 Therefore, it is not a surprise that its lipid deposition profile is not easily manipulated for either cholesterol or PC. The second lens that was found to have no statistical difference in PC deposition when exposed to air was balafilcon A, which may be because of larger standard deviations in the data. On average, exposure to air had a larger impact on cholesterol deposition when compared with PC deposition, and lens materials deposited more cholesterol, on average, than PC.

The deposition of both cholesterol and PC in this experiment is lower than other in vitro and ex vivo experiments conducted.4,5,16,33,39 Recent studies have quantified cholesterol and PC deposition in the microgram range4,5,16,33,39 and not the nanogram range, as seen here. There are several reasons for this: first, the experimental procedure tested for all lenses was the short-term (10 h) incubation, without replenishment of the solution, which is a shorter incubation time than most other in vitro experiments. The short-time frame was used to mimic the wear time on a daily disposable lens and was used to determine the impact that air exposure has on lipid deposition. If statistical differences in deposition could be found after only 10 h of incubation, then it can be deemed a significant effect, and longer incubation times would be expected to find greater masses of deposition and similar if not greater deposition effects with air exposure. Second, the MBC will favor anterior surface deposition of cholesterol and PC, and discourage deposition on the posterior surface. This is because the lens needs to be clipped in place with a specially designed ring manufactured to fit on the MBC pistons. Because of its structure, this ring will reduce the flow of ATS beneath the lens, and it will also reduce the anterior lens surface area exposed and available for deposition. Lastly, PC deposition was specifically lower, which is likely because of its low concentration in the ATS. All of these factors together explain the lower deposition of both lipids examined in this in vitro model. Although the tear film exchange on the posterior side of human-worn soft lenses is reduced when compared with rigid lenses, there is still much more movement and tear film exchange than in the MBC due to lens movement during blinking.

It is hypothesized that the exposure to air increases the lipid deposition on conventional and silicone hydrogel lens materials because of their polymer mobility and chain rotation, as mentioned previously. This chain rotation or hysteresis can be analyzed while measuring wettability through advancing and receding contact angle measurements such as sessile drop, captive bubble, or Wilhelmy plate techniques, just to name a few.40 In human contact lens wear, immediately following a blink, the lens is immersed in the tear film, and lenses hydrophilic moieties face outward with the hydrophobic back bone situated within the matrix.40 As the period between blinks grows longer, the tear film begins to thin and then break over the surface of the contact lens bringing the lipid layer and then air into direct contact with the lens material. When the lens is exposed to lipid and/or air, the previous lens polymer arrangement becomes undesirable and the hydrophilic groups will rotate inward to expose the materials’ hydrophobic backbone to the air.40 This then encourages lipid to deposit and then the air exposure will drive the lipid into the lens matrix. Once another blink occurs, the hydrophilic polymer moieties will again flip to the anterior surface. This occurrence is especially true for lenses that have higher contact angles and for a contact lens wearer with dry eyes, more unstable tear films, and thus shorter tear break up times (TBUT). This wetting/de-wetting cycle can occur after every blink and therefore thousands of times a day, thus allowing lipid to continuously accumulate on and in the lens material.

In the 1990s, Bontempo and Rapp’s in vitro studies found that FDA group II lens materials were the lens materials that were prone to the highest masses of lipid deposition.1 They therefore published a theory for lipid deposition on conventional hydrogel contact lens materials called the “push/pull” theory.1 The theory outlined that the forces involved in lipid deposition onto conventional hydrogel lenses were from the material, which pulled lipid into the lens, and the water component, that pushed the lipid into the matrix of the lens.1 This theory can be modified for silicone hydrogel lens materials, as the incorporation of silicone will provide a strong pull of lipid into the lens material, and then the lipid is forced to “hide” within the matrix because of air exposure during the interblink period. This creates a more powerful push/pull dynamic for lipid deposition on silicone hydrogel lens materials. The mechanism of lipid deposition introduced here is different than experienced with in-vial incubations. During in-vial incubations, the lens remains in an aqueous environment, and the deposition is mainly driven by hydrophobic/hydrophilic interactions with the lens and the ATS.

To date, the effect of intermittent air exposure on in vitro lipid deposition on contact lens materials has not been examined. Peters and Millar analyzed the stabilizing effects of phospholipids on TBUT using a Tearscope and a complex upright model blink eye system.41 Their blinking model eye held a contact lens and was able to spread ATS over the surface of the contact lens using a solenoid and an artificial eyelid. However, this study only examined TBUT, and lipid deposition was not examined. In 2004, Copley et al presented a poster at the Association of Research in Vision Science and Ophthalmology conference, which outlined an experiment that used a model blink to analyze the wettability of lens materials.42 Their model included a blink cell that held a single contact lens and the ATS used was pumped in and out of the cell, thus raising and lowering the liquid levels. In this experiment, a layered ATS was used by spreading a single lipid layer on the anterior surface of the liquid.42 Once again, lipid deposition was not quantified or compared with in-vial incubations.42

In vitro models for lipid deposition onto various contact lens materials have proven to be valuable for the examination of extraction efficiency,38 the effect the tear film composition,21 the effects of ATS concentration,19,21 the effect of ATS replenishment (unpublished data), and the effect of incubation time.18,20,21

The MBC unit introduced in this article has effectively modeled the exposure to air experienced by contact lenses during the interblink period. Further work is needed to determine whether the changes in deposition are a phenomenon specific to early stages of deposition or if the effects are cumulative for a longer period for all lens materials and lipids. It also may be interesting to see if the lenses incubated with MBC contain a higher mass of oxidized lipids than conventional in-vial incubations and to compare that to ex vivo lens data. Additionally, since the MBC elicits a different mode of deposition than in-vial incubations, it may be prudent to examine the depth of lipid penetration into the lens matrix using both incubation models.

The ATS used in this experiment was a homogenous mixture of all incorporated components and was not a layered structure like the human tear film. It is well known that the human tear film is composed of three main phases: the anterior lipid layer, the larger aqueous phase, and the mucin glycocalyx phase that covers the epithelium.35,43,44 The MBC unit could support a layered ATS, with mechanical modifications, to make the device similar to a Langmuir trough. It would be of great interest to see whether the deposition pattern and masses deposited differed with a change in ATS structure. The experiment presented here was a pilot study for this device, and, therefore, further experimentation is needed.

It is true that a laboratory-based model will never be able to fully simulate in vivo conditions, as there are just too many confounding and uncontrollable variables, which can impact deposition onto contact lens materials. However, the incorporation of interblink air exposure has increased the validity of this lipid deposition model.

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CONCLUSIONS

In conclusion, this in vitro blink cell model has demonstrated that lipid deposition kinetics can be impacted by air exposure and that lipid deposition profiles are contact lens material and lipid dependent. These methodologies will provide hitherto unavailable information on the way in which lipid interacts with silicone hydrogel and conventional hydrogel materials and will be of interest to the contact lens industry, clinicians, and other areas of biomaterial research.

Holly Lorentz

Centre for Contact Lens Research

School of Optometry

University of Waterloo

200 University Avenue West

Waterloo, ON N2L 3G1, Canada

e-mail: hmelchin@uwaterloo.ca

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ACKNOWLEDGMENTS

We thank Louie Mansour for the CAD drawings of the model blink cell. This work was sponsored by ALCON Research Ltd.

LJ has received funding over the past 3 years from the following companies who either are directly involved in products used in this manuscript or are involved in the manufacture of competing products—Alcon, AMO, B&L, CIBA Vision, CooperVision, and Johnson & Johnson.

Received February 13, 2012; accepted July 9, 2012.

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Keywords:

lipid; deposition; contact lens; tear film

© 2012 American Academy of Optometry

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