1	>>Lens Refilling for the Restoration of Accommodation and Near-Vision in Human and Other Primate Lenses By Shane Mackey
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5	Table of Contents Slide 6…………….Problem Statement Slide 7-8………...Background Information Slide 9…………….Hypothesis Slide 10-21……..Procedure Slide 22…………..Materials Slide 23…………...Variables Slide 24-46………Graph Data Slide 47……………Results Slide 48……………Application Slide 49……………Conclusion Slide 50……………Acknowledgements Slide 51……………Bibliography
6	Problem Statement Can the symptoms of presbyopia be repaired in Macaca fascicularis (cynomolgous monkey), Macaca mulatta (rhesus monkey), and Homo sapiens (human) lenses without the disadvantages that existing treatments such as spectacles, contact lenses, and intra-ocular lenses present?
7	Accommodation Accommodation is the process by which the eye increases optical power to maintain a focus on an object as it moves closer to the eye. When we view distant objects, our ciliary muscles are relaxed and the zonular fibers are under tension. The tension pulls on the lens and causes the lens' surface to flatten, thus allowing the light rays from the distant object to converge onto the retina. This is known as the unaccommodated state. If a close object is placed in front of the eye while it is unaccommodated, the light rays will converge behind the retina and we will see a blurry image. To bring the image into focus, our eye is able to accommodate the lens. The ciliary muscle contracts and moves closer to the lens, thus releasing the tension on the zonules. When this happens, the lens becomes thicker and its shape becomes more round. This accommodation shortens the focal length, increases the power of the eye, and allows the light rays to converge on the correct spot on the retina.
8	Presbyopia Presbyopia is an age-related disease in which the eye progressively looses its ability to focus on near objects. Although the exact mechanism of presbyopia is unknown, most evidence supports the theory that the lens looses elasticity with age, thus increasing the amount of force required to accommodate. Naturally, the eye begins to lose its ability to focus on near objects almost immediately. While the average child can focus at objects only 50mm (20 diopters) away, by age twenty-five that ability is cut in half to 100mm (10 diopters). By age sixty, the closest the average person can focus is one to two meters away (0.5 to 1 diopter). Presbyopia, which is as natural to old age as grey hair and wrinkles, is first noticed around the ages of forty and fifty as a difficulty reading fine print. Traditionally, presbyopia is treated with eyeglasses, contact lenses, or intra-ocular lenses. The disadvantage to these treatments is that they do not allow for accommodation, they simply allow for a fixed amount of power in the eye that makes reading small print possible. While reading glasses may be fine for reading, they are not good for activities such as driving or walking. The goal of this experiment is to return accommodation to the lenses so that the presbyopic lens has the same optical properties as young lens and can have a full range of focus.
9	Hypothesis It is hypothesized that the vision problems associated with presbyopia can be corrected by restoring accommodation of the lens. It is also hypothesized that accommodation can be restored by the lens refilling procedure of removing the nucleus and cortex of the lens and refilling the capsular bag with a polymeric gel that has optical properties similar to that of a young natural lens.
10	Procedure Step 1: Thirty-seven Macaca fascicularis, twenty Macaca mulatto, and twelve Homo sapiens eyes that were removed posthumously are prepared to undergo the lens refilling procedure. With the pupil of the eye facing downwards, eight PMMA (Polymethyl methacrylate, or Plexiglas) shoes that are designed to fit the curvature of the eye are bonded to form a ring around the iris.
11	"Step 2" Step 2: The posterior hemisphere of the eye is then excised, along with the interior vitreous.
12	"Step 3" Step 3: The remaining segment of the eye is inverted and the cornea and iris are removed. The sclera in between the shoes is sectioned to allow for stretching in eight directions.
13	"Step 4" Step 4: The sample is now ready to mount on the lens stretcher, which is a mechanical device that can apply equal force to the eight PMMA shoes and stretch the eye radially.
14	"Step 5" Step 5: Hooks attached to the lens stretcher are inserted into the shoes.
15	"Step 6" Step 6: The diameter of the lens is measured by a digital camera mounted on the lens stretcher. The stretcher then applies force to the shoes in small increments, enough to stretch the lens 0.25mm in each turn.
16	"The power of the lens..." The power of the lens is determined by a laser underneath where the lens is mounted on the lens stretcher.
17	"Unstretched" Unstretched
18	"Stretched" Stretched
19	"Step 7" Step 7: Once the power, diameter, and force necessary to accommodate the lens is determined for the natural lens, enough information has been gathered to perform the lens refilling procedure. A polymeric gel is chosen that has the same optical properties as those of a young natural lens. The natural lens must first be removed before replacement can take place, so a mini-capsulorhexis is performed. This miniature opening in the lens capsule, which is approximately 1mm in diameter, is where the crystalline lens is removed. The capsular bag remains once the cortex and nucleus of the lens have been removed, and it is this bag that will hold the polymeric gel.
20	"Step 8" Step 8: To prevent leakage of the gel from the lens capsule, a miniature capsulorhexis valve (MCV) is placed against the inner surface of the opening. The gel is then injected into the lens and, depending on the gel type, it may undergo curing, in which a fiber optic light guide is inserted into the lens capsule and 400-460nm blue light is transmitted into the gel. The amount of gel in the lens must be monitored to avoid overfilling or underfilling.
21	"Step 9" Step 9: The same procedure that is used on the natural lenses is used on the refilled lenses when the force, power, and diameters are measured. Step 10: The data is then placed into charts and analyzed to find if refilled lenses have the same or better ability to accommodate than natural lenses.
22	Materials EVAS machine (Ex Vivo Accommodation Simulator) 2 TV screens for surgical and data overlay views of the experiments Surgical microscope Digital camera connected to microscope DMEM (Dulbecco/Vogt modified Eagle's minimal essential medium) 45 Macaca fascicularis eyes 27 Macaca mulatta eyes 12 Homo sapiens eyes Vannas scissors Forceps Scalpel Gloves
23	Variables Experimental Variables Polymer Type: PDMS; 1004TH010-1 H1; SIBS; ISM-1; JJ 68.1.4; JJ 68.1.4; JW1-45a; JW1-22; JJ90.1.3; JJ26,1,4; JJ61,1 AS11A; JJ61,1,4(511B); JJ61,1,4,5; JJ61,1,4(511); JJ100,1,3,4 comb7; UV-SM-1; J61,1,4 (511B); JW1128-102; JW1128-87; 14TL03A-BD7. Species: Cynomolgus Monkey (Macaca fascicularis); Rhesus Monkey (Macaca mulatta); Human (Homo sapiens). Responding Variables Refilled Lens Optical Properties: Unstretched Power (D); Stretched Power (D); Accommodation (D); Accommodation Ratio. Refilled Lens Physical Properties: Unstretched Lens Diameter (mm); Stretched Lens Diameter (mm); Unstretched Lens Thickness (mm); Stretched Lens Thickness (mm); Unstretched Cilliary Body Diameter (mm); Stretched Cilliary Body Diameter (mm); Force at 2 mm stretch (g). Refilled Lens Slopes and Ratios: Load-Diameter Slope (g/mm); Power-Load Slope (D/g); Power-Load Slope Ratio; Load-Diameter Slope Ratio. Variables Held Constant 1) Preparation of eyes for lens refilling procedure. 2) Method of collecting and measuring data. 3) Distance of stretch placed on sclera.
24	Graph Data and Results
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28	Explanation of Previous Graphs The natural versus refilled stretched power scatter plot shows the ratio of the two powers, before and after the Phaco-Ersatz procedure. The dotted line represents a ratio of 1:1, which means that if a point falls on that line, the refilling process induced no changes in the focusing ability of the lens, measured in diopters. Points that fell to the left of the line induced myopia, while points on the right induced hyperopia. The optimum area for a point to land is directly on the line, as this means that the gel that was placed inside the lens has the same optical properties as a natural lens. However, the gels whose points fell into the myopic or hyperopic zones serve a useful purpose as well. These gels can serve to correct vision problems in the cornea. The focusing power of the eye is measured by the sum of the power of the lens and cornea. If a patient has five diopters of hyperopia, then a gel can be placed in the lens that induces five diopters of myopia, thus cancelling out the hyperopia in the cornea and resulting in normal vision.
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32	Explanation of Previous Graphs The power-load slope represents the amount of force required to focus the lens. In the power-load slope scatter plots, the force exerted on the lens in order to accommodate was measured in grams, and the power was measured in diopters. It is important to note that the scale for the vertical axis on the scatter plot decreases in value as it increases in magnitude. Naturally, the power-load slope should increase with age, as it takes more and more force in order to accommodate the lens as we get older. This is most likely a result of decreased elasticity of the lens as we age. The optimum result for the refilled gels' power-load slope should be that there is no change with age (Slope ≤ 0). Therefore, the line of best fit for a scatter plot of all of the power-load slopes of the refilled lenses should be horizontal or negative. If this condition is matched, then it means that the amount of force required to focus the lens stays constant with age. If a refilled lens proves to have a constant power-load slope, then it will negate the effects of the presbyopia-inducing stiffening of the lens.
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36	Explanation of Previous Graphs The load-diameter slope measures how difficult it is to change the shape of the lens. A greater slope means greater hardness of the lens, which results in more force being required to accommodate. This slope should naturally increase with age, but the optimal slope should be constant versus age. Another optimum condition is that the magnitude of the refilled lenses’ slope should be lower than the natural lenses’ slope. A constant slope with age means that the amount of force required to change the shape of the lens into the accommodated state stays the same, whether it is a young or old eye.
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46	Explanation of Previous Graphs When the lens is unstretched, the ciliary muscle is relaxed and the eye is focused at near distances. In this state, the surface of the lens is round the zonular tension is decreased. When the lens is stretched, the ciliary muscle and the zonules tighten, pulling the surface of the lens into a flat surface. At this point the eye can focus on far objects. As we age, our ability to focus greatly diminishes. By the age of twenty, half of our lenses’ power has disappeared. The scatter plots show that naturally, our ability to accommodate from both the stretched and unstretched positions decreases greatly with age. A successful refilled lens is one that shows no change in the power of the lens with respect to age. Subtracting the stretched power from the unstretched power results in the range that the eye is able to accommodate.
47	Results In this experiment, the data collected on the natural lenses was used as the control while the refilled lenses supplied the experimental data. Three polymers were used consistently in all three species of eye: PDMS, 1004TH010-1 H1, and SIBS. These gels had refractive indices of 1.4060, 1.4049, and 1.4066, respectively. The higher the refractive index is, the slower light can travel through the gel. This results in a shorter focal length which means that the light converges further back in the eye than it would if the gel had a lower refractive index. Therefore, 1004TH010-1 H1 resulted in the shortest focal length while SIBS resulted in the longest focal length. As a comparison, the refractive index of glass is 1.5000. The viscosities of the three polymers played a role in the eye’s ability to accommodate. A higher viscosity means that more load must be applied to the lens in order to accommodate, thus causing difficulty and strain on the ciliary muscle if the polymer is too viscous. The viscosities of the three polymers were 10,000cP (PDMS), 10,000cP (1004TH010-1 H1), and 23,400cP (SIBS). As a comparison, the viscosity of water is 1cP. When inserted into a cynomolgus monkey (Macaca fascicularis) lens, PDMS reduced the eye’s ability to accommodate by 37% compared to the natural lens. The refilled lens also required 28% more force per diopter in order to accommodate. However, the PDMS filled lens stretched 21% more millimeters per diopter than it had before it was refilled. When the PDMS was placed into a rhesus monkey (Macaca mulatta) lens, it reduced the range of accommodation by 52%. However, the PDMS increased the amount of diopters per gram the eye accommodated by 2% and the amount of millimeters per gram by 39%. The PDMS gel showed significantly better results when placed into a human (Homo sapiens) lens. While the gel reduced the eye’s ability to accommodate by 13%, it increased the diopters per gram by 32% and the millimeters per gram by 78%. The 1004TH010-1 H1 polymer failed to produce any positive results in any species, decreasing the eye’s accommodation ability by 49% in the cynomolgus monkey, 40% in the rhesus monkey, and 22% in the human. The SIBS polymer was the most effective of the gels used at improving the overall accommodation of the eye. When placed in a cynomolgus monkey lens, SIBS increased the range in which the eye could accommodate by 58%, increased the diopters per gram by 50%, and the millimeters of lens stretching per gram by 12%. In rhesus monkey eyes, the polymer increased accommodation by 21%, increased the number of diopters per gram by 30%, and reduced the force needed to change the diameter of the lens by 18%. In humans, SIBS increased the range of accommodation by 54%, the diopters per gram by 150%, and the diopters per grams by 24%. Overall, the refilled lenses proved to have the same or better ability to accommodate than normal lenses. The refilled human lenses performed 180% of what the normal lenses performed in terms of accommodation, while the refilled cynomolgus monkey lenses reached 111% of what its natural counterpart did, and the rhesus monkey lenses reached 94%. The refilled lenses also required less force to change diameter in order to focus on near objects than the natural lenses did. The human lenses required 65% as much force, the cynomolgus monkey lenses required 92% as much force, and the rhesus monkey lenses required 94% as much force as the natural lenses.
48	Application Presbyopia affects 1.3 billion people worldwide and over a third of the American population. Presbyopia is as common a part of aging as wrinkles and gray hair, and can begin to manifest as early as the age of forty. In fact, our loss of accommodative ability begins the moment we are born, and our range of accommodation shrinks almost exponentially. By age twenty, our amplitude of focus has shrunk to half of what it was at age ten, and by age sixty it is almost non-existent. This loss of accommodation is most likely the result of accumulation of advanced glycation end-products in the crystalline lens that cause the lens to lose its flexibility over time and make accommodation more difficult. By removing the hardened contents of the lens of a presbyopic eye and replacing them with a polymeric gel that has the same optical properties as a young, healthy lens, the presbyopia can be cured and accommodation returned to the lens. Several advantages exist to the lens refilling procedure that cannot be found in any of the existing treatments for presbyopia. Currently, contact lenses or eyeglasses are prescribed in order to restore focusing ability, but these devices are not convenient for everyday use. People who use bifocals must sacrifice field of view and can interfere with those who need distortion-free sight in their professions. Using contact lenses to correct one eye for near vision and one eye for far vision, known as monovision, causes issues with depth perception. Intraocular lenses, which are artificial lenses placed inside the capsular bag of the lens, are mostly fixed monofocal lenses that provide for distance vision. However, this technique does not restore accommodation; it merely acts as an artificial, inflexible lens. The lens refilling procedure uses polymers that are flexible enough that the ciliary muscle can contract and accommodation can occur as it does in a young, healthy lens.
49	Conclusion The results supported the hypothesis. Compared to natural lenses, refilled lenses have the same or better ability to accommodate (Homo sapiens 180%, Macaca fascicularis 111%, Macaca mulatto 94%). The refilled lenses require less force to change diameter than natural lenses (Homo sapiens 65%, Macaca fascicularis 92%, Macaca mulatto 94%). Less significantly different results were expected in the monkey lenses, due to the lack of old lenses tested. PDMS reduced the accommodation ratio of all three species (Homo sapiens 13%, Macaca fascicularis 37%, Macaca mulatto 52%), increased the load-diameter slope ratio (Homo sapiens 78%, Macaca fascicularis 21%, Macaca mulatto 39%), increased the power-load slope ratio in two species (Homo sapiens 32%, Macaca mulatto 2%) and decreased in one species (Macaca fascicularis 28%). 1004TH010-1 H1 reduced the accommodation ratio of all three species (Homo sapiens 22%, Macaca fascicularis 49%, Macaca mulatto 40%) and did not produce successful results in other areas. SIBS greatly increased the accommodation ratio of all three species (Homo sapiens 54%, Macaca fascicularis 58%, Macaca mulatto 21%), increased the load-diameter slope ratio (Homo sapiens 24%, Macaca fascicularis 12%, Macaca mulatto 30%), and increased the power-load slope ratio in all three species (Homo sapiens 150%, Macaca mulatto 50%, Macaca fascicularis 30%). Of the three polymers used throughout the experiment, SIBS was considered a successful substitute for a natural lens, PDMS was a marginal success, and 1004TH010-1 H1 was a failure. The experiment may have been affected by the volume of gel placed in the capsular bag. Care must be taken not to overfill or underfill the capsular bag with polymer when refilling takes place. Any changes in original volume greater than a 20% increase or decrease can have significant effect on the optical properties of the lens. Care must also be taken during the formation of the mini-capsulorhexis not to create a tear in the capsular bag. The small incision must be almost perfectly circular in order to prevent tearing during refilling. The experiment may also have been affected by the health and quality of the eye used in the refilling process. The human eyes were all received from diseased individuals, which may or may not have had an effect on the chemical composition or optical properties of the eye. Several of the cynomolgus monkey eyes used were from diabetic monkeys, and two of the eye specimens from rhesus monkeys were received because the donor had died from neurological disease. Future studies will include the testing of additional Phaco-Ersatz gels with different viscosities and refractive indices.
50	Acknowledgements
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