The Biocompatibility Analysis of Artificial Mucin-Like Glycopolymers.

The ocular surface is covered by a tear film consisting of an aqueous/mucin phase and a superficial lipid layer. Mucins, highly O-glycosylated proteins, are responsible for lubrication and ocular surface protection. Due to contact lens wear or eye disorders, lubrication of the ocular surface can be affected. Artificial glycopolymers which mimic natural mucins could be efficient in ophthalmic therapy. Various neutral, positively, and negatively charged mucin-mimicking glycopolymers were synthesized (n = 11), cultured in different concentrations (1%, 0.1%, and 0.01% w/v) with human corneal epithelial cells (HCE), and analyzed by various cytotoxicity/viability, morphology, and immunohistochemistry (IHC) assays. Six of the eleven glycopolymers were selected for further analysis after cytotoxicity/viability assays. We showed that the six selected glycopolymers had no cytotoxic effect on HCE cells in the 0.01% w/v concentration. They did not negatively affect cell viability and displayed both morphology and characteristic markers as untreated control cells. These polymers could be used in the future as mucin-mimicking semi-synthetic materials for lubrication and protection of the ocular surface.

[New nanofibrous scaffold for corneal tissue engineering]. / Eine neue Nanofaser-Matrix für den kornealen Gewebeersatz.

BACKGROUND: An estimated 10 million people suffer worldwide from vision loss caused by corneal damage. For the worst cases, the only available treatment is transplantation with human donor corneal tissue. However, in numerous countries there is a considerable shortage of corneal tissue of good quality, leading to various efforts to develop tissue substitutes. The present study aims to introduce a nanofibrous scaffold of poly(glycerol sebacate) PGS as a biodegradable implant, for the corneal tissue engineering. MATERIALS AND METHODS: Nanofibrous scaffolds were produced from PGS and poly(ε-caprolactone) (PCL) by a modified electro-spinning process. The biocompatibility of the material was tested in vitro by colorimetric MTT assay on days 3, 5, and 7 to test the cell viability of human corneal endothelium cells (HCEC). To examine a potential immunological reaction of the scaffolds, samples were exposed to mononuclear cells derived from peripheral blood (PBMCs). After an incubation period of 3 days, supernatants were assayed for apoptotic assessment and immunogenic potentials by annexin V FITC//propidium iodide and flow-cytometric analysis. RESULTS: We could successfully demonstrate that cultivation of HCECs on PGS/PCL scaffolds was possible. Compared to day 3, cell density determined by microplate absorbance was significantly higher after 7 days of cultivation (p < 0.0001). According to the MTT data, none of the samples showed toxicity. Apoptotic assessments by FACS analysis showed that no composition stimulated apoptosis or activated PBMCs occurred. All the compositions were inert for native as well as activated T/B/NK cells and monocytes. It can be concluded that leukocytes and their activity was not affected by the scaffolds. CONCLUSION: A tissue-like scaffold mimicking the human stroma could be developed. The results indicate that PGS/PCL scaffolds could be considered as ideal candidates for corneal tissue engineering as they are biocompatible in contact to corneal endothelial cells and blood cells.

Anti-apoptotic gene therapy prolongs survival of corneal endothelial cells during storage.

Corneal transplantation is the most common form of grafting performed worldwide. Corneal endothelial cells (EC) form a monolayer in the posterior portion of the cornea and are essential for corneal transparency. EC loss during storage before transplantation is a principal reason for rendering donor tissue unsuitable for transplantation, and apoptosis has been shown to be the major contributor to EC loss during storage and after transplantation. Therefore, the potential use of anti-apoptotic gene therapy to promote both graft storage and graft survival is of major interest. The goal of this study was to transduce human donor corneas in vitro to enhance EC survival during storage conditions used in eye banking. We utilized a lentiviral vector to perform gene transfer of baculoviral p35 or mammalian Bcl-xL to corneal endothelium in different storage conditions utilizing a lentiviral vector. Our results show significantly enhanced survival and prolonged retention of physiological EC morphology in cells expressing either p35 or Bcl-xL. The clinical application of this technology could lead to a higher availability of donor tissue for transplantation, extend storage periods and reduce graft failure after transplantation.

[Viral vectors for gene delivery to corneal endothelial cells]. / Virale Vektoren für den Gentransfer in korneale Endothelzellen.

BACKGROUND: Corneal endothelium is an interesting target for in vitro gene transfer strategies as it is readily accessible thanks to its anatomic structure as a monolayer and its direct contact to culture medium. Whereas the use of adenoviruses as viral vectors (carriers) to endothelial cells (EC) has been described as problematic as to its immunogenicity, lentiviruses and adeno-associated viruses (AAV) are potent vectors for the transfer of genetic DNA into EC. Lentiviral vectors, developed on the basis of HI-viruses, can integrate the transferred gene into the host DNA and thus lead to a permanent protein expression. Evaluating apathogen alternatives to lentiviral vectors for humans, we herein compared non-integrating AAV to lentiviral gene transfer. MATERIALS AND METHODS: A comparison was made of the kinetics of expression of a green fluorescent protein after transduction using a lentiviral vector and AAV 2 / 2 in a murine EC line, human EC line and human primary cells (flow cytometry). A proof of principle experiment was conducted to demonstrate the function after lentiviral gene transfer of the anti-apoptotic gene Bcl-xL. RESULTS: The kinetics of protein expression after transduction of EC using a lentiviral or an AAV vector show fundamental differences. Contrary to gene transfer using AAV, a high expression of the reporter protein was readily detectable only hours after transduction using the lentiral vector. In addition, we could demonstrate distinct differences in protein expression characteristics between human and murine EC as well as human EC line and primary human EC. Function could be demonstrated by showing a significant reduction in apoptosis in both murine and human EC. CONCLUSION: AAV vectors are an alternative to lentiviral vectors for gene transfer to corneal EC. Given a cultivation time of donor corneas of up to 4 weeks before transplantation, translation to eye banking, e. g., to decrease apoptosis in corneal allografts, is conceivable.

[Acute angle-closure glaucoma and effusion syndrome after phacoemulsification]. / Akutes Engwinkelglaukom und Effusionssyndrom nach Phakoemulsifikation.

A 72-year-old female patient developed bilateral secondary iridocorneal angle-closure glaucoma with uveal effusion syndrome after uncomplicated cataract surgery. The postoperative intake of acetazolamide was identified as causative for the development of the effusion syndrome. Taking a sulfonamide-free systemic and local intraocular pressure lowering and anti-inflammatory treatment into account, a rapid improvement of the ocular manifestation was achieved. The case illustrates a rare but clinically severe adverse effect of acetazolamide and outlines efficient treatment options.

[Evaluation of corneal biomechanics in keratoconus using dynamic ultra-high-speed Scheimpflug measurements]. / Evaluation kornealer Biomechanik bei Keratokonus mithilfe dynamischer Ultra-High-Speed-Scheimpflug-Messung.

BACKGROUND: The in vivo analysis of corneal biomechanics in patients with keratoconus is especially of interest with respect to diagnosis, follow-up and monitoring of the disease. OBJECTIVE: For a better understanding it is necessary to describe the potential of dynamic Scheimpflug measurements for the detection and interpretation of biomechanical changes in keratoconus. MATERIAL AND METHODS: The current state of analyzing biomechanical changes in keratoconus with the Corvis ST (Oculus Optikgeräte GmbH, Wetzlar, Germany) is described. This technique represents a new approach for understanding corneal biomechanics. Furthermore, it was investigated whether the device can biomechanically quantify a rigidity increasing effect of therapeutic UV-crosslinking and whether early stages of keratoconus can be detected using dynamic Scheimpflug analysis. RESULTS AND DISCUSSION: In patients with keratoconus, the in vivo analysis of corneal biomechanics using dynamic Scheimpflug measurements as a supplementary procedure can be of advantage with respect to disease management. By optimization of screening of subclinical keratoconus stages, this method widens the analytic spectrum regarding diagnosis and follow-up of the disease; however, further studies are required to evaluate whether visual outcome of affected patients can be improved by earlier diagnosis.

[Critical endothelial procedures during posterior lamellar graft preparation and transplantation]. / Endothelkritische Prozeduren während der Herstellung und Transplantation posteriorer lamellärer Hornhauttransplantate.

BACKGROUND: In view of the very low proliferation rate and functional importance of the corneal endothelium in maintaining corneal transparency, safeguarding the integrity of this monolayer plays a central role in posterior lamellar corneal transplantation. Several critical endothelial procedural stages are necessary to carry out such a transplantation. OBJECTIVE: This article presents various preparatory and operative approaches for carrying out the necessary and critical stages within the framework of posterior lamellar corneal transplantation and concentrates on the question of optimization. METHODS: A review of our own studies and studies of other groups is presented. RESULTS: For the performance of critical endothelial procedural steps, a variety of approaches are available. These range from preparation and insertion of the transplant, through the manipulation during centralization up to the effects of postoperative air or gas bubble tamponade. CONCLUSION: Because endothelial damage can permanently impair the integrity of lamellar transplants, a minimal handling and no touch policy should be strived for in all critical procedures. Long-term data on the follow-up course will show which of the procedures favored by various authors lead to the best postoperative results.

[Adjuvant topical interferon-alpha-2b treatment in epithelial tumors of the ocular surface]. / Adjuvante topische Interferon-alpha-2b-Therapie bei epithelialen Tumoren der Augenoberfläche.

PURPOSE: The aim of this study was to evaluate the role of topical interferon-alpha-2b in the adjuvant treatment of corneal and conjunctival tumors. METHOD: In this noncomparative, prospective, interventional case series, five patients with histologically proven conjunctival intraepithelial neoplasia (CIN) after primary excision and amniotic membrane transplantation were treated with interferon (IFN)-alpha-2b eye drops five times daily over a period of 6 weeks (1 million IU/ml Intron A, Schering). An in situ hybridization technique was used to detect human papillomavirus (HPV) in all specimens. Frequent follow-up was undertaken clinically and photographically for evidence of tumor recurrence. RESULTS: In the follow-up period (13.2+/-4,97 months) no clinical evidence of recurrence with only limited treatment side effects such as mild conjunctival hyperemia were recorded after 6 weeks of interferon. In one CIN specimen HPV 16/18 could be detected. CONCLUSIONS: The combination of excisional biopsy and topical interferon-alpha-2b application seems to be an effective and safe treatment for conjunctival intraepithelial neoplasias. Therefore, we prefer this combined treatment to topical interferon-alpha-2b treatment alone, more destructive approaches such as radiation and cryotherapy, or treatment with antimetabolites such as 5-fluorouracil or mitomycin C.

[Protection of corneal endothelium from apoptosis by gene and cell therapy]. / Protektion von Hornhautendothel vor Zelluntergang durch Gen- und Zelltherapie.

Protection of corneal endothelium from apoptosis using gene and cell therapy is in a translational phase. This approach offers advantages for eye banking and after transplantation. Safe vehicles for gene or cell therapeutic transduction of corneal endothelium with nucleic acids are available. This strategy will be further developed in consultation with the Paul Ehrlich Institute and European regulatory authorities.

[Neurotrophic keratopathy--a post-LASIK case report]. / Neurotrophe Keratopathie--Ein Fallbeispiel nach LASIK.

BACKGROUND: Neurotrophic keratopathy is a degenerative affection of the cornea caused by impairment of corneal sensitivity and represents a therapeutic challenge for ophthalmologists. The present article concentrates on the etiology, pathogenesis, diagnosis, therapeutic aspects and future therapeutic models for treating neurotrophic keratopathy and will present a case report: Also we evaluate the effects of autologous serum eye drops in bilateral LASIK-induced neurotrophic keratopathy with epithelial breakdown revealed by positive fluorescence and rose bengal staining and reduced tear film break-up time. METHODS: We treated a 42-year-old patient with post-LASIK neurotrophic keratopathy and tear film instability with autologous serum eye drops (5 x daily) and emulsion eye drops (Refresh Endura, Allergan, Irvine, CA, USA) after insertion of punctal plugs. RESULTS: Stabilization of vision, healing of the epithelium and reduction of the previously experienced symptoms like redness, itching and burning were achieved within 6 weeks. 10 months after changing therapy, the patient only complained about slight pain during lid movement. CONCLUSIONS: Severe denervation after bilateral LASIK disrupts ocular tear film dynamics and causes irritation symptoms of the ocular surface. Autologous serum eye drops may be an effective treatment of severe epithelial breakdown and be helpful to reestablish the disturbed ocular surface integrity, as shown by negative vital staining. Therefore, autologous serum eye drops represent a significant approach in the therapy of LASIK-induced severe dry eye and associated pain.