Abstracts
The optical and visual effects of tear film break-up
Meredith Jansen
IU School of Optometry
Fri, Jan 25, 2008
PURPOSE: It is well known that the blink rate markedly decreases with tasks requiring concentration, and that contact lens (CL) wear is associated with an increased blink rate. However, blinking patterns of CL wearers engaged in different tasks have not been measured. The purpose of this study was to determine whether CL wear affects the fullness and rate of blinking when subjects are involved in different tasks. METHODS: Four subjects, who had not worn their soft contact lenses for 24 hours were seated behind a slit lamp biomicroscope custom equipped to simultaneously measure blinking (by a customized MATLAB program designed to track a reflective sticker on the upper lid) via a high speed video camera (120 Hz) and retroillumination (RI) of the pupil to monitor tear break-up under infrared light. Three measurements of 55 sec each were taken while subjects listened to music and then played a video game. These procedures were repeated with the subjects' habitual contact lenses. The Current Symptoms Questionnaire (CSQ) was filled out before and after each set of tasks. RESULTS: Without CL, the interblink interval (IBI) significantly increased (AVG music=3.0+/-1.0sec; AVG game=7.5+/-2.1sec), and the blink amplitude (BA) decreased (AVG music=77.1%+/-14.2%; AVG game=53.1%+/-21.5%) when subjects played a video game versus listened to music (paired t-test, p<0.001). With CL, the BA significantly decreased (AVG music=84.0%+/-11.0%; AVG game=65.3%+/-9.6%, p<0.001), but the IBI showed only a minimal increase (AVG music=1.8+/-0.5sec; AVG game=2.5+/-0.9sec). The RI images often showed horizontal lines of tear disruption marking the extent of the blink both with and without CL. Symptoms increased after playing the game with CL. CONCLUSIONS: An increased blink rate with CL wear was especially evident when performing a task requiring concentration, such as playing a video game, and resulted in increased symptoms of ocular discomfort. Increased blinking may be due to vision disruption secondary to drying of the soft contact lens on the eye.
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Corneal curvature at different stages of pterygium
Emad Alsaleh
IU School of Optometry
Fri, Feb 15, 2008
Purpose: To investigate the effect of pterygium on corneal curvature. Methods: Corneal topography was determined using a portable Keratron Scout (EyeQuip Company, Florida, USA). Nineteen points at equal intervals along the horizontal corneal meridian were selected to represent instantaneous corneal curvature (diopters) from limbus to limbus. Photographs of the central, nasal and temporal aspects of the eye were taken using a Nikon D200 digital camera with a Micro-Nikkor 105 mm lens equipped with a close-up flash system. From the photographs the percentage of the corneal diameter occupied by the pterygium was determined. Subjects were volunteers from the clinic at the Indiana University Eye Care Center, Guanajuato, Mexico. This is a region with a high prevalence of pterygium due to altitude, clear skies and an outdoor lifestyle. The age range was 18 to 87 years (27 men and 53 women, mean = 49.3 years). Of the 80 subjects, 73 had a pterygium in at least one eye (6 had one eye affected, 67 had both eyes affected). A total of 154 eyes were enrolled. Fifty-seven (57) eyes had almost the same grade of pterygium in both nasal and temporal sides (bilateral), and 77 eyes had pterygium limited to one side of the cornea (unilateral). Twenty (20) eyes had minimal conjunctival changes, and they were used as controls. Results: All unilateral pterygia were nasal. The measured mean instantaneous curvatures (diopters) for the controls and grades of pterygium were as follows:
|
MED-NASAL |
CENTRAL |
MED-TEMPORAL |
|
CONTROL (n= 20) |
42.2 + 4.2 D |
46.6 + 3.2 D |
43.9 + 3.6 D |
|
GRADE 2 (n= 21) |
42.9 + 7.7 D |
46.3 + 4.4 D |
46.6 + 9.3 D |
|
GRADE 3 (n= 23) |
38.7 + 8.5 D |
46.6 + 3.5 D |
45.2 + 6.1 D |
|
GRADE 4 (n= 21) |
30.4 + 7.8 D |
46.0 + 4.0 D |
46.1 + 5.7 D |
Conclusions: Pterygium causes flattening of the nasal cornea but no change temporally. Pterygium usually does not affect the curvature of the central cornea.
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Validation of Novel Hartmann-Moiré Wavefront Sensor with large dynamic range
Xin Wei
IU School of Optometry
Fri, Feb 29, 2008
Purpose: Traditional Hartmann-Shack aberrometers have been shown to be accurate and clinically useful. However, this technique occasionally lacks the necessary dynamic range to measure highly aberrated eyes. Recently, a new wavefront sensing technology, the Hartmann-Moiré (HM) wavefront sensor (PixelOptics, Inc.), was developed to meet the need for wide dynamic range. This new wavefront sensor combines two traditional Hartmann screens, with the second screen rotated relative to the first to achieve a Moiré effect that amplifies the movement of the spots of light created by the first Hartmann Screen. The purpose of our study was to validate the accuracy, repeatability, and dynamic range of the instrument with spherical and cylindrical trial lenses.
Methods: Wavefront vergence was measured at a wavelength of 532nm without focus adjustment so that the full range of wavefront vergences was presented to the wavefront sensor. The accuracy and dynamic range of the HM wavefront sensor was evaluated by measuring defocus and astigmatism induced by a series of standard Topcon spherical (77 lenses from -20D to +18D) and cylindrical trial lenses (16 lenses from -8D to 8D). Repeatability was assessed by taking 3 measurements within a 2-minute period. Measured trial lens values with the HM wavefront sensor were compared to lens values verified with a standard lensometer. Analyses were based on a 4-mm pupil diameter specified in the software.
Results: Defocus was accurately measured over a 38D range and astigmatism over a 16D range. The primary limitation to demonstrating an even larger dynamic range was the increasingly critical requrements for optical alignment. Correlation coefficients between mean wavefront measurements (n=3) and expected wavefront vergence for both sphere and cylinder lenses were >0.999. For spherical lenses, the instrument was accurate to within 0.2D over the range from -20D to +18D. For cylindrical lenses, the instrument was accurate to within 0.2D over the range from -8D to +8D. Sensitivity to small changes of vergence was constant over the instrument's full dynamic range. Repeatability of measurements for fixed condition was within 0.04D. Improved accuracy would be expected after an optimized calibration that takes component tolerances into account.
Conclusion: The Hartmann-Moiré wavefront sensor measures defocus and astigmatism accurately and repeatedly over a large dynamic range required for clinically abnormal eyes.
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The role of bicarbonate and carbonic anhydrase in lactic acid transport across the corneal endothelium
Tracy Nguyen
IU School of Optometry
Fri, Mar 28, 2008
Purpose: Previous studies done in various tissues have shown that bicarbonate (HCO3-) along with carbonic anhydrase (CA) facilitates lactate flux. Because of the presence of HCO3- transporters, lactate:H+ cotransport, and several CAs, a similar mechanism could be present in the corneal endothelium (CE). We will test the hypothesis that HCO3- together with CA activity facilitates lactate:H+ flux from the basolateral to apical surface of CE. This hypothesis will be tested using both in vitro and in vivo techniques.
Methods: Bovine CE cells were grown to confluence on tissue culture inserts and placed in a 24 well plate. 500 μl of 5mM (L)-lactate in either bicarbonate free (BF) or bicarbonate rich (BR) ringer solution was added to the basolateral compartment and 350 μl of lactate-free BF or BR ringer to the apical compartment. Samples were taken from the apical compartment at various time points (0, 1, 3, 5, 10, 20 minutes) after addition of lactate. Lactate concentration was analyzed using the BioVision Lactate Assay Kit. Endogenous lactate was collected from the basolateral and apical compartment using lactate free BR (5% CO2, pH7.5) and BF (Air, pH7.5) ringer's on both sides at 37 degrees C. Lactate-dependent H+ flux was determined in CE cultured on coverslips perfused with BF or BR +100uM acetazolamide, a carbonic anhydrase inhibitor by measuring the the rate of change in intracellular pH (pHi) upon addition of 20mM (L)-lactate. Relative changes in cell buffering capacity for each condition was determined by measuring the pHi change to a 5 mM NH4Cl pulse. In vivo, we used lentiviral mediated transfection and shRNA to knockdown carbonic anhydrase II and IV and sodium-bicarbonate cotransporter of the rabbit endothelium and assessed corneal thickness, stromal pH and corneal lactate concentration.
Results: In the endogenous lactate test, BR induced a 5x (n=3) greater lactate efflux into the apical compartment compared to BF and 3x greater efflux into the basal compartment with BR than BF. Cell buffering capacity was unaffected by acetazolamide in BF, but reduced 35% in BR. Whereas lactate-dependent pHi changes were faster in BF, lactate-dependent proton flux (j=β*dpHi/dt) was 3 times faster in BR media compared to BF. Proton flux was unaffected by acetazolamide in BF. However, in BR media, proton flux decreased by 55% in the presence of acetazolamide. We are still in the preliminary stages of constructing and testing the lentiviral vectors that express shRNA of NBC with GFP. Preliminary results show that we can successfully transfect the corneal endothelium with NBC shRNA using lentiviral vector.
Conclusion: Transport of lactate from the basolateral (stroma) to the apical (anterior chamber) side of corneal endothelium is facilitated in the presence of HCO3-. Similarly, lactate-dependent proton flux is faster in HCO3- and is stimulated by carbonic anhydrase activity.
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The usable field of multifocal spectacles
Wuthikrai Boonvarute
IU School of Optometry
Fri, Apr 4, 2008
Purpose: If the results of the clinical tests of the new design lens in this research are superior to the optical properties of bifocal, trifocal, and progressive additional lens, then it will give the presbyopic users a new selection of lens to choose from to correct their vision problem.
Methods: This clinical experiment was designed and conducted to deliver the objectives of the new lenses designer. This randomized clinical trial included standard clinical methods and newly developed testing methods including questionnaires. The experiment used small sample size, N =20. Twelve of them were recruited from Optometry School patients list, the rest were recruited from outside. Subjects were bifocal, trifocal, and progressive additional lenses users whose spectacles' add powers were between +1.25D to +2.25D. They must not have ocular diseases be able to do the tests for one and a half hours.
Results: The new design lenses had wider peripheral visual field than progressive additional lenses, but comparable to bifocal and trifocal lenses. The new design lenses also had good depth of field while flat top bifocal does not have it. Overall preferences of the new design lenses were much higher than flat top bifocal.
Conclusion: Results confirm some of our predictions were that width of field for Pixel was larger than for conventional PAL, but comparable to bifocals or trifocals, while still providing intermediate vision.
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Influence of TNF-α on barrier integrity of corneal endothelium
Mahesh Shivanna
IU School of Optometry
Fri, Apr 4, 2008
Purpose: TNF-α is a pro-inflammatory cytokine implicated in corneal endothelial failure during graft rejection. It is known to induce an increase in permeability of corneal endothelial cells but the underlying molecular mechanisms are not well understood. We have investigated the role of microtubules and the influence of cAMP in TNF-α-induced loss of barrier integrity in cultured bovine corneal endothelial cells.
Methods: Changes in the organization of microtubules was examined by staining for α-tubulin. Localization of ZO-1, a marker of TJ assembly, was visualized by immunostaining. Changes in the barrier function were continuously monitored by transendothelial electrical resistance (TER) using ECIS (Applied Biophysics, Inc, NY).
Results: Exposure to TNF-α (20 ng/ml; 6h) induced microtubule disassembly and dispersion of ZO-1. TNF-α also led to a continued decline in TER, which sustained beyond 20 h. These effects of TNF-α were significantly opposed by pretreatment with paclitaxel (a microtubule stabilizing agent; 10 μM; 1 hr) and SB203580 (20 µM; 1 hr), a p38 MAPK (mitogen activated protein kinase) inhibitor. Co-treatment with forskolin (10 μM) and rolipram (PDE4 inhibitor; 50 μM) significantly opposed the TNF-α-induced decrease of TER and prevented the dispersion of ZO-1. Similarly, co-treatment with the nonselective A2B agonist NECA (20 _M) and rolipram also significantly opposed the decrease in TER. Adenosine, however, was found to be less effective when compared to forskolin and NECA in opposing TNF-α response.
Conclusion: TNF-α may induce loss of barrier integrity through microtubule disassembly involving activation of p38 MAPK. The other mechanism involved could be through depletion of intracellular cAMP. The influence of elevated cAMP on TNF-α-induced loss of barrier integrity may involve opposition to actomyosin contraction and/or enhancement in actin polymerization at the apical junctional complex.
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Photoreceptor contributions to the photopic flicker electroretinogram of the rat
Christina Gant
IU School of Optometry
Fri, Apr 18, 2008
Purpose: The purpose of this study was to determine the photoreceptor contribution to the photopic flicker electroretinogram (ERG) of the rat.
Methods: Full-field photopic flicker ERGs were recorded using DTL electrodes from anesthetized (ketamine 85mg/kg/hr, i.m. and Xylazine 5mg/kg/hr, i.m.) pigmented rats (FBNF1/Hsd) using an ESPION system. The visual stimuli consisted of sinusoidally modulated green LED light (525nm peak wavelength and 40nm 1/2 band width) with a mean luminance of 500 cd/m.sq and 82% contrast. The stimulus frequency range was 1 - 40 Hz. Pupils were fully dilated to about 4mm diameter. Responses were obtained in the same eyes before and after blocking post-receptoral response with intravitreal injections of a mixture of 4mM L-2-amino-4-phosphonobutyric acid (to block depolarizing bipolar cells) and 4mM cis-piperedine-2, 3-dicaroxylic acid (to block transmission to hyperpolarizing bipolar cells and horizontal cells). Fundamental and second harmonic Fourier components of the flicker ERG responses before and after injection of pharmacological agents were extracted.
Results: The fundamental as well as the second harmonic Fourier components of the flicker ERG responses from control eyes reduced progressively with increase in stimulus temporal frequency to reach a response minimum around a stimulus frequency of 20Hz. Thereafter, amplitudes increased slightly with further increase in stimulus temporal frequency and this increase was relatively larger for the second harmonic compared to the fundamental. The phase of the fundamental and second harmonic components from control eyes showed progressive changes with stimulus frequency up to the frequency that elicited the response minimum and thereafter abruptly shifted to demonstrate a second limb. Following pharmacological blockade of post-receptoral responses, the fundamental and second harmonic amplitudes were reduced relative to the control amplitudes from the same eyes at all stimulus frequencies greater than 1Hz. Also, the amplitudes reduced continuously with increase in stimulus temporal frequency with amplitudes reaching noise levels beyond a stimulus frequency of 11 and 20 Hz for the fundamental and second harmonic respectively. The phase plots of the fundamental and second harmonic components of the isolated photoreceptor responses were monotonic, consistent with the presence of a single cellular mechanism contributing to these responses.
Conclusion: Photoreceptors make significant contributions to both, the fundamental and second harmonic Fourier components of the of the full-field photopic flicker ERG responses. These contributions are restricted to stimulus frequencies below 20Hz. Post-receptoral neurons are largely contributory to the flicker ERG responses at stimulus frequencies greater than 20 Hz. These findings should be useful in the interpretation of the flicker ERGs in different experimental models that employ this species.
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Local and global anisotropies in cone photoreceptor packing
Toco Chui
IU School of Optometry
Fri, Apr 18, 2008
Purpose: To use high resolution retinal imaging to investigate local and global anisotropies in cone densities within the central retina of emmetropic adults.
Methods: A high resolution adaptive optics scanning laser ophthalmoscope (AOSLO) (50nm bandwidth centered at 840nm) was used to image the cone photoreceptors of human eyes. Confocal aperture was fixed at either 12μm or 24μm relative to the retina. Photoreceptor mosaics covering the entire central 10deg x 10deg of retina were obtained by using programmable displacements of scanning mirrors and careful fixation. Cones positions in the entire cone mosaics were determined automatically using a custom Matlab program (MathWorks, Natick, MA). Local anisotropy (meridional variation in sampling within a local region) and global anisotropy (sampling differences across retinal meridians) in spacing of cone photoreceptor were studied by analyzing the density contour maps and the Fourier power spectra at retinal eccentricities along the primary meridians.
Results: Cone density was within the bandwidth of the imaging system for all the peri- and parafoveal regions, but cone counting was not possible in the central 1 degree due to the small size of foveal cones. Iso-eccentric samples obtained along the primary retinal meridians revealed a global anisotropy that was stable across eccentricity and revealed highest sampling density along the horizontal meridian.
Conclusion: Throughout the perifoveal and parafoveal retinal regions, our in vivo imaging reveals both local and global anisotropies similar to those reported for a single cadaver eye (Curcio and Sloan, 1992).
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Off-axis monochromatic aberrations and image quality in a wide-angle model eye
Jason Shen
IU School of Optometry
Fri, Apr 25, 2008
Purpose: To validate the measurements of off-axis aberration using clinical wavefront aberrometer (COAS) and verify data analysis using commercial software (CLAS-2D). 2. To study the effects of eccentricity on IQ and the relative contributions of each Zernike components to the wavefront aberration.
Methods: A physical model eye was developed for this project to provide a stable test case that resembled a human eye. Measurements were taken in 5-degree steps up to 45 degrees by COAS (Wavefront Science, Inc.) with auxiliary lens to correct off-axis refractive error. We were using CLAS-2D software to specify a circular analysis pupil circumscribed the elliptical entrance pupil and treat the data between the elliptical pupil and the surrounding circular area as missing data. The wavefront was reconstructed over the circular analysis pupil and IQ was computed by masking off the corresponding missing data area.
Results:Up to 30 degrees of eccentricity, the wavefront sensor recorded slope measurements over the entire entrance pupil. Beyond 35 degrees, the irradiance declined rapidly along the minor axis of the elliptical pupil, possibly due to the range-limiting-aperture in COAS that blocks highly aberrated rays. Image quality metric VSOTF declined slowly as eccentricity increased. VSOTF increased greatly on-axis when C40 was removed, indicating that spherical aberrataion is the main cause of poor image quality centrally in this model eye. VSOTF is much larger centrally and it falls off quickly with eccentricity when setting C40 = 0 at all eccentricities.
Conclusion: Measurements of off-axis wavefront aberrations of a model eye from 0 degrees to 30 degrees of eccentricity are possible with the COAS clinical wavefront aberrometer and auxiliary lenses to correct astigmatism. Data analyzed by COAS and CLAS-2D are mutually consistent. When central IQ is good, the off-axis aberrations will have a powerful effect on peripheral IQ. When central IQ is poor, the additional effect of off-axis aberrations will be minor.
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Microtubules regulate barrier integrity of corneal endothelial monolayer
Supriya Shivakumar
IU School of Optometry
Fri, Apr 25, 2008
Purpose: Increased Myosin Light Chain (MLC) phosphorylation breaks down the barrier integrity of corneal endothelium. This study has examined the effects of microtubule disassembly on MLC phosphorylation and barrier integrity in cultured bovine corneal endothelial cells (BCEC).
Methods: Nocodazole was used to induce microtubule disassembly. Changes in the organization of cytoskeleton were visualized by staining for F-actin and α-tubulin. MLC phosphorylation was assayed by urea-glycerol gel electrophoresis and Western blotting. Barrier integrity was measured in terms of transendothelial electrical resistance (TER).
Results: Exposure to nocodazole led to disassembly of microtubules and also caused disruption of characteristic dense assembly of cortical actin. Nocodazole also led to an increase in MLC phosphorylation as well as a significant decrease in TER. The decrease in TER was opposed by paclitaxel, a microtubule stabilizing agent. Forskolin, which is known to inhibit activation of RhoA through elevated cAMP, and Y-27632 (a Rho kinase inhibitor), attenuated nocodazole-induced MLC phosphorylation as well as the decrease in TER.
Conclusion: Microtubule disassembly breaks down the barrier integrity of BCEC through disruption of the actin cytoskeleton possibly through RhoA activation.
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