In vitro and in vivo evaluation of a hydrophilized silicone intraocular lens

In vitro and in vivo evaluation of a hydrophilized silicone intraocular lens Hans J. Hettlich, M.D., Robert Kaufmann, Ph.D., Heike Harmeyer, Elke Imkamp, M.D., C. James Kirkpatrick, M.D., Ph.D., Christian Mittermayer, M.D.

The main reason for the development of soft intraocular lens (IOL) materials was to allow implantation through a small incision by folding the lens.1-4 Keeping the corneal/scleral opening as small as possible prevents the risk of high postoperative astigmatism 5 and improves wound healing. It is not clear whether folding the IOL is an advantage,6 but there are reasons for using silicone as an IOL material (chemical stability, autoclavability, interesting geometrical forms, 7 optical properties,8 no biodegeneration, and no adverse reaction if the material is chemically pure 9 ). One disadvantage of silicone as an IOL material is unstable fixation, as described by several authors. 4,lO,1l This may be a consequence of the

hydrophobic surface of the material. From cell culture experiments, we know there is a correlation between cell adhesion and a hydrophilic surface. 12 In this study, we compared a hydrophilized silicone IOL and an untreated but otherwise identical lens. We were particularly interested in lens fixation, foreign-body reaction, and the rate of posterior capsular opacification.

MATERIALS AND METHODS We used disc-shaped silicone IOLs (type 90 D, Adatomed GmbH, Munich, Germany). The lens

From the Institute of Pathology of the RWTH Aachen, Aachen, Germany (Hettlich, Harmeyer, Kirkpatrick, Mittermayer), the German Wool Research Institute, RWTH Aachen, Aachen, Germany (Kaufmann), and the Eye Clinic of the RWTH Aachen, Aachen, Germany (Imkamp). Supported by a grant from the Bundesminister fur Forschung und Technologie, Germany. Reprint requests to H. 140

J.

Hettlich, M.D., Ratzeburger Allee 160, 2400 Lubeck, Germany.

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has a total diameter of 9.6 mm and an optic diameter of6.0 mm. The optic is biconvex and the haptic is plano and disc-shaped. The entire lens is made of poly(dimethylsiloxane) with a refractive index of nD 25 = 1.410 and a specific weight of d 2o • = 1.04 g/cm 3 (product information).

control populations cultured on untreated silicone. By comparing cell morphology (light microscopy) as well as measurements of protein and DNA synthesis, we evaluated the potential cytotoxic effect of the modified material. The DNA syntheSis was examined by scintillation counting after incubation with tritium-labeled thymidine; protein synthesis was measured photometrically according to Lowry and coauthors .14 Scanning Electron Microscopy (SEM). To investigate the morphology of the treated surfaces and to exclude surface damage, we used a scanning electron microscope (Phillips SEM 515). The IOLs were examined after conventional preparation. Contact Angle Measurements. We used contact angle estimations to illustrate the surface modification. Measurements were made with aqua bidest (type Gl, Kruess, Hamburg, Germany).

Surface Modification by Plasma Treatment The surface_was treated with oxygen plasma using a plasma unit (type 30176, Technics Plasma GmbH, Kirchheim/Munich, Germany). In this unit gas is stimulated by microwaves under vacuum conditions. The resulting plasma can be used to bring functional groups onto the surface of silicone IOLs. 13 Using oxygen plasma as described below, one can graft a desired number of oxygen groups onto a polymer surface without inducing surface damage. Surface Analysis X-ray Photoelectron Spectroscopy (XPS) . This method of surface characterization obtains data about the elemental composition and the oxidation states at the outermost layer of a substrate (information depth about 10 nm). We used an X-Probe'" spectrometer (model 206, SSI, Mountain View, CA). Cell Culture. In cell cultures, cell spreading is known to increase on a more hydrophilic surface. 12 We incubated a larynx carcinoma cell line on a plasma-treated silicone IOL for 24 hours at 37°C with BME medium. Spreading was analyzed by planimetric estimation of the cell area. On every surface 120 cells were analyzed and compared to

In Vivo Experiments In a pilot study, 30 New Zealand white rabbits (female) were implanted with a silicone IOL in the left eye. Anesthesia was achieved with an intravenous injection of ketamine/xylazine (5:1). For dilation of the pupil the animals were given tropicamide and phenylephrine hydrochloride drops topically. Fifteen modified lenses (02-plasma, 60s, 600W) and 15 unmodified lenses of the same type were implanted. After corneoscleral opening and radial capsulotomy (12 0'clock/6 o'clock) the crystalline lens was removed by phacoemulsification. Five hundred milliliters of the irrigation solution (Ringer's) contained 5,000 IE heparin and 1 ml

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Oxygen Concentration (atomiC %)

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Fig. 1. 30

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(Hettlich) Oxygen XPS atomic concentration of untreated and 02-plasma-treated poly(dimethylsiloxane) IOLs over treatment period (plasma power: 600W).

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epinephrine hydrochloride. The disc lenses were inserted into the capsule in an unfolded state after enlarging the corneoscleral opening to 8.5 mm and the capsulotomy to 7.0 mm. For disc-lens implantation into an intact rabbit capsule, we preferred a radial capsulotomy as used in all experiments described. Methylcellulose (2%) was used as a viscoelastic substance. The wound was closed with 9-0 polyglactin (Vicryl®) and dexmethasone/neomycin/polymixin B (Isopto-Max®) ointment was instilled into the superior fornix. No treatment was provided postoperatively. During the 12 week follow-up the animals were regularly examined at the slitlamp. As it is very difficult to distinguish clinically between synechias to the capsule and to the IOL surface in rabbit eyes, findings of pupillary mobility and synechia were classified semiquantitatively as follows: o = pupil round, mobile 1 = pupil largely round, not totally mobile 2 = pupil slightly distorted, remnant of mobility 3 = pupil totally distorted, no mobility At the end of the follow-up the animals were sacrificed by an overdose of pentobarbital. The macroscopic findings were photographed from the anterior aspect and, after equatorial sectioning of the globes, posteriorly. The posterior capsular opacification rate was semiquantitatively evaluated as follows: o = no posterior capsular opacification 1 = posterior capsular opacification covering the haptic only 2 = posterior capsular opacification covering part of the optic

3 = posterior capsular opacification covering optic and haptic totally Most of the explanted lenses (26) were fixed in ethanol (50%) with 2 % Carbowachs® and prepared for light microscopy by staining according to Papanicolaou. As coverslips would disturb the exact focusing, we coated the lenses with a thin layer of Kaiser's Glyceringelatine® (Merck). We also fixed two lenses of each group in buffered glutaraldehyde solution (2%) and inspected them by SEM after sputtering. The anterior segments of the eyes were fixed in formalin (5%), embedded in paraffin, and sectioned. After staining with H&E, they were examined by light microscopy. Because of the high regeneration rate of the corneal endothelium in rabbits, we did not investigate endothelial damage. RESULTS

Surface AnalYSis/Cell Culture Surface analysis of the plasma-treated silicone IOL by XPS showed a correlation between the oxygen at the surface and the treatment period (Figure 1). The process was limited to periods of up to 60 seconds (600W) by the occurrence of fissures at the surface as shown by SEM. The longest treatment period without surface damage (60s/600W) involved a decrease of approximately 25 degrees in the contact angle (from 121.86° [SD = 2.28/n = 36] to 96.58° [SD = 2.55/n = 36]). In cell culture experiments we found an increase of spreading correlating with the density of oxygen-containing groups at the surface (Figure 2). A decrease of spreading at treatment periods of 300

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Fig. 2.

(Hettlich) Cell spreading on plasma-treated silicone IOLs. For each treatment period, 120 cell areas were estimated planimetrically.

spreading (X)

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seconds can be explained by the enhanced crack formation. The cell morphology evaluated by light microscopy was normal on every surface. Measurements of DNA and protein synthesis were not significantly influenced by the surface modification (DNA synthesis [n = 5]: contr. 4291 cpm [SD = 21]/60s 02-plasma 4280 cpm [SD = 55]; protein synthesis [n = 5]: contr. 201.4 ~g/culture [SD = 7]/60s 02-plasma 202.6 ~g/culture [SD = 5]).

In Vivo Experiments In clinical examinations we observed total or partial dislocations out of the capsule (Figure 3) of four hydrophilized lenses (28.6%) and eight control group lenses (53.3%). The difference evaluated by the Fisher test and the chi-square test was not significant. Statistical analysis of the semiquantitative

evaluation of synechias and pupil mobility for the entire follow-up showed that the data for the hydrophilized lenses were superior to those of the control lens (Cochran-Mantel-Haenszel test: P = .009) (Figure 4). Intraocular lens cytology 12 weeks postoperatively showed no homogeneous cell distribution on lenses in either group. Counting every foreignbody giant cell on the anterior lens surface with more than 15 nuclei (Figure 5), we found a range from 0 to more than 50 giant cells in both groups (Figure 6). A statistically significant difference could not be found. Scanning electron microscopy images of the cellular reaction on the explanted lenses (Figure 7) as well as the histological examination of the anterior segments showed no signif-

c

A

Fig. 3.

(Hettlich) Silicone disc lens in the rabbit eye, partially dislocated into the anterior chamber. A and B are anterior views, C the view from behind after sectioning the globe. The IOL was pushed out of the capsular bag by regenerative lens material.

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-. Billeono IOL untruted - silicone IOl hydrophlllzed

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average iris score

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0, 5 o~-----+------~------~-----;

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Fig. 4.

Fig. 5.

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(Hettlich) Mean values of the semiquantitative iris score. As we only assessed cases in which the IOL was well placed in the capsular bag, the number decreased from 15 control IOLs and 15 surface-treated IOLs at the beginning to 7 and 11 eyes at the end of the follow-up.

(Hettlich) Cell population on an explanted silicone IOL (plasma-treated) 12 weeks postoperatively. Some giant cells and macrophages at different stages of spreading can be seen . Staining: Papanicolaou; original magnification: 250x.

icant difference. Cells analyzed on the explanted lens surfaces by SEM showed all the morphological criteria of macrophages. Various stages of adhesion and of confluence leading to giant cell formation could be seen. Neither lymphocytes nor other cellular components were detected. Light microscopy examination of the corneas revealed an infiltration of the stroma with polymorphonuclear granulocytes showing eosinophilic granules in some cases. These findings did not correspond to one of the investigated lens surfaces or to any other clinical or pathological parameter. We found this infiltration also in an eye that had had the same procedure but that had not received an IOL. 144

Fig. 6.

(Hettlich) Survey of a cell population on an explanted silicone IOL (untreated) 12 weeks postoperatively. On most lenses, as in this case, we found a nonhomogeneous cell population. Staining: Papanicolaou; original magnification: 20X .

In the capsular bag the disc-shaped haptic prevented the formation of a Soemmering's ring. In cross sections of many capsular bags, we found a mold of the disc-shaped haptic in the equator region where it was placed before explantation (Figure 8). The amount of posterior capsular opacification ranged from an almost cell-free capsular bag (Figure 9) to capsules in which the IOL was completely embedded in regenerative crystalline lens material (Figure 10). Twelve weeks postoperatively the posterior capsular opacification rate was 2.0 in the control group and 1.7 for the hydrophilized IOL group. This difference was not statistically significant; however, we found special patterns on the posterior capsule in some cases with hydrophilized IOLs. The fibrosis was localized to a discrete area in which contact with the posterior capsule was minimal because of the lens geometry (Figure II). Adherence of the IOL surface to the lens capsule in the clear areas could not be demonstrated. DISCUSSION This study was designed to investigate the potential benefits of hydrophilized of silicone IOLs and the effect of the wettability of an IOL surface on the intraocular reaction in general. Like other authors who have not found a difference in foreign-body reaction between poly(methyl methacrylate) and silicone IOLs,l5,16 we found no difference in the reaction between hydrophilized and untreated silicone IOLs. Because our lenses as well as our surgical procedure were otherwise identical, we conclude that the wettability of an IOL is not one of the most important

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Fig. 7.

(Hettlich) Scanning electron microscopic images of cells on an explanted silicone IOL 12 weeks postoperatively. Left: untreated control lens. Right: plasma-treated silicone IOL. Morphologically these cells appear to be macrophages in different stages of spreading with initial giant cell formation.

Fig. 8.

(Hettlich) Equatorial region of a lens capsule 12 weeks postoperatively. The disc-shaped haptic formed a mold and prevented the formation of a Soemmering's ring. Staining: H&E, original magnification: 200x .

Fig. 9. (Hettlich) Survey of a lens capsule 12 weeks postoperatively. The capsular bag of this eye is nearly free of cellular material. Staining: H&E, original magnification: BOx .

influences on foreign-body reaction after IOL implantation. The cellular reaction on the explanted IOL investigated by light microscopy and SEM support the hypothesis of a macrophagic origin of the so-called fibroblast-like cells described in the literature. 17 We found single cells in different stages of adhesion as well as confluent cells forming giant cells. Corneal infiltration with granulocytes appeared to be caused more by the procedure (corneal/scleral opening, lens extraction, wound suturing) than by one of the tested lens surfaces. The finding of eosinophilic granules in the granulocytes does not mean that these cells are identical to eosinophilic granulocytes from adverse reactions in humans. Silicone is known to be nonadhesive to

tissue. 5 We assume from our results that the tendency to develop posterior synechias to the lens can be reduced even more by a hydrophilized IOL surface. A correlation between IOL dislocation and posterior capsular opacification has been described. 1B , 19 Our results for dislocation rate and posterior capsular opacification rate also indicate that these phenomena should be evaluated together. They suggest that both are influenced by the ability of an IOL surface to establish firm contact to the posterior lens capsule. This contact can influence the posterior capsular opacification rate. 20 As our results are not conclusive, this correlation should be investigated in more detail. If

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Fig. 10.

(Rettlich) Same technique as Figure 9. A large mass of regenerative lens material can be seen on the posterior capsule of this eye (12 weeks postoperatively).

Fig. 11.

(Rettlich) View of a posterior capsule after removing a hydrophilized silicone disc IOL (12 weeks postoperatively). The fibrosis was found in a sharply limited and peripheral area; the rest of the capsule remained clear.

more evidence for this hypothesis can be obtained,

it might be possible to improve contact between

the IOL surface and the capsule by specific surface modifications.

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