Posterior capsule opacification (PCO) is a major component of secondary cataract, a complication of current cataract surgery practice. This iatrogenic condition occurs in virtually all pediatric cases and to a lesser extent in adults. PCO correlates with the development in the latter half of the 20th Century of extracapsular cataract extraction (ECCE). In these surgeries, the lens capsule is left intact. During ECCE surgery a circular capsulotomy opening is created in the anterior lens capsule, and the cataractous, proteinaceous lens is removed, often via ultrasonic lens liquefaction i.e. phacoemulsification. The posterior, equatorial and remaining anterior portions of the sac-like capsule are left intact, permitting the insertion of an artificial lens into the emptied capsule. However, cells from the monolayer of epithelium on the inner surface of the capsule often begin to proliferate and migrate onto the normally cell-free inner surface of the posterior capsule, and may obscure the central axis of vision. Subsequently, a second surgery is necessary to create a small capsulotomy in the centre of the posterior capsule, usually employing an Nd:YAG laser. However, up to 5% of patients who have capsulotomies may then develop further serious, vision-threatening complications such as macular edema and retinal detachments. This thesis reports the photodynamic therapy (PDT) conditions required to prevent lens epithelial (LE) cell de novo proliferation and migration onto posterior lens capsules in a euthanized rabbit surgical model in order to predict parameters required to prevent PCO in humans. Experiments with primary in vitro cultures of human LE cells have shown rapid delivery of the photosensitizer benzoporphyrin derivative monoacid ring A (BPD-MA) and efficient killing with low light doses of 690 nm red light. Additional studies have shown the efficacy of various viscous agents in protecting the comeal endothelium. During model phacoemulsification ECCE surgeries, the use of hyaluronate viscoelastic carriers addressed the need for containment necessary for localized delivery of photosensitizer in the emptied capsule. Long-term monitoring of PDT-treated rabbit lens capsules in vitro has demonstrated a phototoxic effect including complete cell kill in this surgical model employing the prophylactic use of PDT.

Posterior capsule opacification (PCO) is a major component of secondary cataract, a complication of current cataract surgery practice. This iatrogenic condition occurs in virtually all pediatric cases and to a lesser extent in adults. PCO correlates with the development in the latter half of the 20th Century of extracapsular cataract extraction (ECCE). In these surgeries, the lens capsule is left intact. During ECCE surgery a circular capsulotomy opening is created in the anterior lens capsule, and the cataractous, proteinaceous lens is removed, often via ultrasonic lens liquefaction i.e. phacoemulsification. The posterior, equatorial and remaining anterior portions of the sac-like capsule are left intact, permitting the insertion of an artificial lens into the emptied capsule. However, cells from the monolayer of epithelium on the inner surface of the capsule often begin to proliferate and migrate onto the normally cell-free inner surface of the posterior capsule, and may obscure the central axis of vision. Subsequently, a second surgery is necessary to create a small capsulotomy in the centre of the posterior capsule, usually employing an Nd:YAG laser. However, up to 5% of patients who have capsulotomies may then develop further serious, vision-threatening complications such as macular edema and retinal detachments. This thesis reports the photodynamic therapy (PDT) conditions required to prevent lens epithelial (LE) cell de novo proliferation and migration onto posterior lens capsules in a euthanized rabbit surgical model in order to predict parameters required to prevent PCO in humans. Experiments with primary in vitro cultures of human LE cells have shown rapid delivery of the photosensitizer benzoporphyrin derivative monoacid ring A (BPD-MA) and efficient killing with low light doses of 690 nm red light. Additional studies have shown the efficacy of various viscous agents in protecting the comeal endothelium. During model phacoemulsification ECCE surgeries, the use of hyaluronate viscoelastic carriers addressed the need for containment necessary for localized delivery of photosensitizer in the emptied capsule. Long-term monitoring of PDT-treated rabbit lens capsules in vitro has demonstrated a phototoxic effect including complete cell kill in this surgical model employing the prophylactic use of PDT.

Posterior capsule opacification (PCO) is a major component of secondary cataract, a complication of current cataract surgery practice. This iatrogenic condition occurs in virtually all pediatric cases and to a lesser extent in adults. PCO correlates with the development in the latter half of the 20th Century of extracapsular cataract extraction (ECCE). In these surgeries, the lens capsule is left intact. During ECCE surgery a circular capsulotomy opening is created in the anterior lens capsule, and the cataractous, proteinaceous lens is removed, often via ultrasonic lens liquefaction i.e. phacoemulsification. The posterior, equatorial and remaining anterior portions of the sac-like capsule are left intact, permitting the insertion of an artificial lens into the emptied capsule. However, cells from the monolayer of epithelium on the inner surface of the capsule often begin to proliferate and migrate onto the normally cell-free inner surface of the posterior capsule, and may obscure the central axis of vision. Subsequently, a second surgery is necessary to create a small capsulotomy in the centre of the posterior capsule, usually employing an Nd:YAG laser. However, up to 5% of patients who have capsulotomies may then develop further serious, vision-threatening complications such as macular edema and retinal detachments. This thesis reports the photodynamic therapy (PDT) conditions required to prevent lens epithelial (LE) cell de novo proliferation and migration onto posterior lens capsules in a euthanized rabbit surgical model in order to predict parameters required to prevent PCO in humans. Experiments with primary in vitro cultures of human LE cells have shown rapid delivery of the photosensitizer benzoporphyrin derivative monoacid ring A (BPD-MA) and efficient killing with low light doses of 690 nm red light. Additional studies have shown the efficacy of various viscous agents in protecting the comeal endothelium. During model phacoemulsification ECCE surgeries, the use of hyaluronate viscoelastic carriers addressed the need for containment necessary for localized delivery of photosensitizer in the emptied capsule. Long-term monitoring of PDT-treated rabbit lens capsules in vitro has demonstrated a phototoxic effect including complete cell kill in this surgical model employing the prophylactic use of PDT. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

Knowledge of the mechanics of the lens capsule is crucial for improving cataract surgery as well as understanding better the physiological role of the lens capsule in the process of accommodation. Previous research on the mechanical properties of the lens capsule contains many gaps and contradictions due to experimental limitations and inappropriate assumptions. Thus, the goal of this work is to quantify fully the regional, multiaxial mechanical behavior of the lens capsule and to calculate the change in stress and strain fields as a result of cataract surgery. Determining in situ the multiaxial mechanical behavior of the lens capsule required the design and construction of an experimental device capable of altering stresses in the capsule while measuring localized surface deformations. Tests performed on this device reveal that the meridional and circumferential strains align with the principal directions and are equivalent through most of the anterior lens capsule, except close to the equator where the meridional strain is greater. Furthermore, preconditioning effects were also found to be significant. Most importantly, however, these tests provide the data necessary for calculating material properties. This experimental system is advantageous in that it allows reconstruction of 3D geometry of the lens capsule and thereby quantification of curvature changes, as well as measurement of surface deformations that result from various surgical interventions. For instance, a continuous circular capsulorhexis (CCC) is commonly used during cataract surgery to create a hole in the anterior lens capsule (typically with a diameter of 5 mm). After the introduction of a CCC, strain was found to redistribute evenly from the meridional direction (retractional strain) to the circumferential direction (extensional strain), where both directional components of strain reached magnitudes up to 20% near the edge of the CCC. Furthermore, the curvature was found to increase at the edge of the CCC and remain the same near the equator, indicating that the mere introduction of a hole in the lens capsule will alter the focal characteristics of the lens and must therefore be considered in the design of an accommodative intraocular lens.

Abstract This study was undertaken to identify mechanisms of adhesion of intraocular lenses (IOLs) to the capsular bag after cataract surgery and IOL implantation. It was also done to challenge the sandwich theory presented for posterior capsular opacification (PCO): If the IOL is made of a bioactive material it would allow a single lens epithelial cell layer to bond both to the IOL and the posterior capsule at the same time. This would produce a sandwich pattern including the IOL, the cell monolayer and the posterior capsule. The sealed sandwich structure would prevent further epithelial ingrowth. The degree of bioactivity of the IOL could explain the basic difference in the incidence of PCO and capsulotomy rates with different IOL materials. The sandwich theory was put forward on the basis of a search for a keratoprosthesis material, which would allow maximal adhesion of the prosthesis to corneal tissue. Titanium and glass-ceramic coated titanium were found to develop better adhesion than poly (methyl methacrylate) (PMMA). The adhesion of PMMA to the corneal stromal tissue was loose, and down growth of corneal epithelial cells was seen around the prosthesis. The differences between various IOL materials were first tested with rabbit corneal tissue cultures. There was better adhesion of corneal tissue to soft, hydrophobic acrylate than to PMMA, heparin surface modified (HSM)-PMMA, silicone or hydrogel IOLs. To assess differences in protein adhesion to IOL surfaces, different IOLs were incubated for 24 hours with radioactive iodine labeled fibronectin. Soft hydrophobic acrylate (AcrySof®) showed the highest binding of fibronectin, and the differences relative to all the other materials were significant (p < 0.01-0.001), except to PMMA (p = 0.31). The sandwich theory and the results with rabbit corneal tissue cultures and the protein adhesion study in vitro were evaluated against the results found in pseudophakic autopsy eyes. Altogether, 70 autopsy eyes were analyzed. From 38 autopsy eyes containing PMMA, silicone, soft hydrophobic acrylate or hydrogel IOLs histological sections were prepared from the capsular bag and immunohistochemical analyses were performed for fibronectin, vitronectin, laminin and collagen type IV. A total of 152 specimens were analyzed. From 32 autopsy eyes containing IOLs made of PMMA, silicone, acrylate or hydrogel, IOLs were explanted from the capsular bag and immunohistochemical analysis was done on both sides of the IOLs for fibronectin, vitronectin, laminin or collagen type IV. Soft hydrophobic acrylate IOLs had significantly more adhesion of fibronectin to their surfaces than PMMA or silicone IOLs. Also, more vitronectin was attached to acrylate IOLs than to the other IOL materials. Silicone IOLs had more collagen type IV adhesion in comparison to the other IOL materials studied. In histologic sections a sandwich-like structure (anterior or posterior capsule-fibronectin-one cell layer-fibronectin-IOL surface) was seen significantly more often in eyes with acrylate IOLs than in PMMA, silicone or hydrogel IOL eyes. These studies support the sandwich theory for posterior capsule opacification after cataract surgery with IOLs. The results suggest that fibronectin may be the major extracellular protein responsible for the attachment of acrylate IOLs to the capsular bag. This may represent a true bioactive bond between the IOL and the lens epithelial cells, and between the IOL and the capsular bag. This may explain the reason for clinical observations of less posterior capsular opacification and lower capsulotomy rates with the soft hydrophobic acrylate material of AcrySof® IOLs compared to the other IOL materials studied.

<p>Improved materials for implantation as intraocular lens (IOL) devices are needed to minimize the occurrence of posterior capsule opacification (PCO). In this work, novel polydimethylsiloxane (PDMS) loaded with matrix metalloprotease inhibitors (MMPI) were developed as model IOL materials.</p> <p>PDMS was chosen as silicones are currently used successfully as IOLs. Inhibitor release rates and amount of initial burst of drug-loaded PDMS could be controlled by changing solvent when loading into elastomer base, as well as drug loading method, and release buffer.</p> <p>Two lens epithelial cell lines were characterized for in vitro tests: FHL124 and HLE B3. These cell lines produce different combinations of extracellular matrix proteins when grown on various biomaterial surfaces. Significant differences between the two cell lines were observed both in collagen VIII and α-smooth muscle actin levels, both when cells were unstimulated, and as a result of epithelial to mesenchymal transition induced by treatment with transforming growth factor β2. FHL124 cells were selected in further tests due to their consistent expression of extracellular matrix components when exposed to different materials.</p> <p>Solutions of synthetic MMPI GM6001 and MMP 2/9 Inhibitor II, known to mitigate anterior subcataract formation, were released from PDMS and found to protect in a modest but significant way against protein profile changes and to delay migration. Due to the Zn²⁺ dependence of MMPs, chelators, including EDTA, TPEN and 1-10 phenanthroline were examined as alternative inhibitors. Only the latter was found to have a significant effect on cell migration rates in vitro. Sulfadiazine, due to its chemical resemblance to synthetic MMPI was determined to be the most efficient at reducing migration rates as well as to have the lowest toxicity.</p> <p>Overall, sulfadiazine was determined in this work to be a potentially effective solution to mitigating PCO. These results indicate that releasing MMPI molecules from PDMS as a model IOL is a promising way to mitigate aspects of PCO.</p> / Thesis / Doctor of Philosophy (PhD)

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Hintergrund: RGD-Peptide hemmen kompetitiv die Adhäsionsmoleküle von Linsenepithelzellen (LEC). Ziel unserer Untersuchungen war es herauszufinden, ob diese Peptide in der Lage sind, auch nach Kurzzeitinkubation eine suffiziente Inhibition der Adhäsion bzw. eine Ablösung adhärenter Zellen und damit eine ausreichende Prävention des Nachstars zu bewirken. Außerdem wurde überprüft, ob das von uns verwendete RGD-Peptid eine Toxizität für die Hornhaut aufweist. Material und Methoden: Kulturen boviner und humaner LEC, boviner Hornhautendothelzellen, humane und bovine Linsenkapselexzidate und humane explantierte Hornhäute wurden verwendet. Wir untersuchten die Inhibition der Adhäsion und die Ablösung konfluenter LEC-Layer mittels des zyklischen RGD-Peptids cRGDDFV (Inkubationszeiten von 1 Stunde bzw. 5-7 Tagen und Konzentrationen von 10-4 M, 10-3 M und 2x10-3 M wurden angewandt). Ergebnisse: Wir fanden nach nur einstündiger Inkubation in der Kulturschale eine Adhäsionsinhibition von 48% für bovine LEC und von 100% für humane LEC. Die Differenz zwischen Kontrollpeptid und cRGDDFV war statistisch signifikant (p / Purpose: RGD-peptides competitively inihibit adhesion molecules of the lens epithelial cells (LEC). The purpose of our study was to investigate whether this peptide could be able to inhibit adhesion sufficiently after short term incubation resp. to detach adherent cells and so to prevent posterior capsule opacification (PCO). Also there was proofed if there is any toxicity for the cornea. Methods: Cultures of bovine and human LEC, bovine cornea endothelial cells, humane and bovine fragments of the lens capsule and explanted humane corneas were used. The inhibition of adhesion and the detachment of confluent LEC-layer by the cyclic RGD-peptide cRGDDFV were studied (incubation time was 1 hour resp. 5-7 days and concentrations of 10-4, 10-3 M and 2x10-3 M were used). Results: After one hour incubation time in a culture dish inhibition of adhesion was 48% for bovine LEC resp. 100% for humane LEC. There was a statistically significant difference between the control-peptide-group and cRGDDFV (p