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Purpose: When an eye has become irreversibly blind or painful it is removed by enucleation or evisceration. The resulting anophthalmic socket usually receives a volume replacing implant and is subsequently fitted with a prosthetic shell for adequate cosmesis. Trauma, tumour or immunological pathomechanisms can induce loss of bone, orbital soft tissue volume, and conjunctival contraction or implant exposure, which result in difficult or impossible prosthesis wear. In this situation as well as in numerous diseases limited to the conjunctiva (e.g. Pterygium, or cicatrizing conjunctivitis) strategies to substitute the lost tissue are required. Methods: A review of the literature search using various electronic databases (PubMed and MEDLINE) was performed on indications, surgical techniques and materials used to restore the ocular socket. Results: Amniotic membrane and oral mucosa are still the most commonly used substitutes for the reconstruction of larger conjunctival defects and ocular socket reconstruction. However, due to limitations of clinical available grafts, synthetic scaffolds, biomaterials or tissue-engineered grafts have been described in preclinical studies but most of them have not been investigated adequately in clinical studies yet. In orbital volume replacement, porous and nonporous spheres are used and both show acceptable results. However, more clinical studies are required that directly compare the outcomes in patients with similar conditions. Dermofat graft remains a good option in case of sockets with significant orbital volume and conjunctival surface loss. Conclusion: Beyond established techniques using autologous or allogeneic tissue, various approaches of engineering tissue based on scaffolds and stem cell expansion techniques are currently under investigation and may become alternatives in socket reconstruction in the not too far future. [ABSTRACT FROM AUTHOR]

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Purpose: To examine clinical results up to 10 years after Descemet Membrane Endothelial Keratoplasty (DMEK) and Triple-DMEK., Methods: Prospective study including 201 eyes, of which 54 eyes [38 DMEKs and 16 Triple-DMEKs; Fuchs endothelial dystrophy (n = 45), pseudophakic bullous keratopathy (n = 9)] reached the minimum follow-up of 5 years and were followed up for up to 10 years. We evaluated best-corrected visual acuity (BCVA, logarithm of the minimum angle of resolution [logMAR]), endothelial cell density (ECD, cells/mm2), minimal central corneal thickness (CCT, μm), central retinal thickness (μm), rebubbling, graft survival and re-DMEK rate, chamber angle alterations, and incidence of glaucoma., Results: Fifty-four eyes had a 5-year and 37 eyes an 8- to 10-year follow-up. Mean follow-up was 94.4 ± 12.1 months. Best-corrected visual acuity increased from 0.6 ± 0.3 logMAR to 0.1 ± 0.2 logMAR at 6 months (P ≤ 0.001) and was 539 ± 54 μm at long term. Endothelial cell density decreased from 2488 ± 320.9 (donor) to 980.1 ± 437 cells/mm2 with an average ECD loss/year of 4% and did not correlate with BCVA. Central corneal thickness decreased from 596.9 ± 82.2 μm to 498.6 ± 24.3 μm at 12 months (P ≤ 0.001) and remained stable (P = 1.000). Mean rebubbling rate was 0.3 ± 0.5/eye and did not correlate with BCVA. Eight eyes (14.8%) received at least 1 rebubbling. Primary graft failure/rejection rate was 1.5%/0% within the first postoperative year, and secondary graft failure rate was 12.4% at 7 years. Eight eyes (preoperative n = 3, de novo n = 5) had open-angle glaucoma without chamber angle changes (14.8%, P ≤ 0.001)., Conclusions: Although ECD decreases continuously in the long-term follow-up, excellent visual acuity can be preserved 10 years after DMEK and Triple-DMEK. The final outcome can be estimated 6 months after surgery., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Sustainable treatment of aqueous deficient dry eye (ADDE) represents an unmet medical need and therefore requires new curative and regenerative approaches based on appropriate in vitro models. Tissue specific hydrogels retain the individual biochemical composition of the extracellular matrix and thus promote the inherent cell´s physiological function. Hence, we created a decellularized lacrimal gland (LG) hydrogel (dLG-HG) meeting the requirements for a bioink as the basis of a LG model with potential for in vitro ADDE studies. Varying hydrolysis durations were compared to obtain dLG-HG with best possible physical and ultrastructural properties while preserving the original biochemical composition. A particular focus was placed on dLG-HG´s impact on viability and functionality of LG associated cell types with relevance for a future in vitro model in comparison to the unspecific single component hydrogel collagen type-I (Col) and the common cell culture substrate Matrigel. Proliferation of LG epithelial cells (EpC), LG mesenchymal stem cells, and endothelial cells cultured on dLG-HG was enhanced compared to culture on Matrigel. Most importantly with respect to a functional in vitro model, the secretion capacity of EpC cultured on dLG-HG was higher than that of EpC cultured on Col or Matrigel. In addition to these promising cell related properties, a rapid matrix metalloproteinase-dependent biodegradation was observed, which on the one hand suggests a lively cell-matrix interaction, but on the other hand limits the cultivation period. Concluding, dLG-HG possesses decisive properties for the tissue engineering of a LG in vitro model such as cytocompatibility and promotion of secretion, making it superior to unspecific cell culture substrates. However, deceleration of biodegradation should be addressed in future experiments., (Creative Commons Attribution license.)

Prcis: A main safety concern associated with new glaucoma implants is corneal endothelial cell loss, which could even be observed in already established procedures., Purpose: The purpose of this study was to evaluate the efficacy and corneal safety, especially corneal endothelial cell loss (ECL), after Preserflo MicroShunt implantation., Methods: A monocentric, prospective clinical observational study of patients with a follow-up of at least 12 months who underwent Preserflo MicroShunt implantation at the Department of Ophthalmology, University Hospital of Düsseldorf, Germany, between January 2020 and October 2021. Endothelial cell density (ECD), the distance between the tip of the implant and the corneal endothelium, and the success of the surgery were analyzed., Results: In total, 14 eyes of 12 patients were included. After a mean follow-up of 20±2.7 months intraocular pressure was significantly reduced by 8.2 mm Hg (P=0.0041); in 28.6% of patients, a revision surgery was necessary. Reduction of preoperative and follow-up ECD was not statistically significant (ECL of 45 cells/mm2, ECDpreoperative=2074±703.6 cells/mm2, ECDlast follow-up=2029±742.3 cells/mm2, P=0.42). A longer intracameral shunt tube length correlated significantly with a higher distance between the shunt tip and corneal endothelium (r=0.61, P=0.036)., Conclusions: Preserflo MicroShunt effectively lowered intraocular pressure without substantial ECL after a minimum follow-up period of 17 months., Competing Interests: Disclosure: The authors declare no conflict of interest., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: The aim of this study was to evaluate results after Descemet membrane endothelial keratoplasty (DMEK) and DMEK with cataract surgery (triple DMEK) in eyes with endothelial dysfunction and concomitant macular pathology., Methods: A monocentric, prospective clinical observational study of patients who underwent DMEK or triple DMEK surgery at the Department of Ophthalmology, University Hospital Düsseldorf, Germany, from June 2013 to February 2016 was conducted. Sex, age, best-corrected visual acuity (BCVA), central corneal thickness, endothelial cell density, and central retinal thickness in the 1 millimeter zone were analyzed., Results: A total of 209 eyes were included in the study. Forty-two eyes (20.1%) had vision-limiting concomitant maculopathies. These were age-related macular degeneration (n = 17, 8.1%), epiretinal gliosis (n = 13, 6.2%), chronic macular edema (n = 7, 3.3%), macular holes (n = 3, 1.4%), and macular scarring (n = 2, 1.0%). BCVA significantly increased in patients without maculopathy from 0.6 ± 0.33 logMAR to 0.1 ± 0.15 logMAR ( P < 0.001) and also in patients with maculopathy from 0.9 ± 0.38 logMAR to 0.27 ± 0.23 logMAR 12 months postoperatively ( P < 0.001). There was a significant central retinal thickness increase of 34.1 μm 6 weeks after triple DMEK in the central 1-mm zone ( P = 0.011). This increase was insignificant after DMEK. Postoperative macular edema occurred in 5.9% of cases after DMEK and 8.1% after triple DMEK, which was not significantly different., Conclusions: DMEK and triple DMEK significantly increase BCVA in eyes with concomitant maculopathy. Postoperative macular edema is a common disorder after lamellar keratoplasty; therefore, prophylactic treatment with nonsteroidal antiinflammatory drugs should be considered. Maculopathies did not predispose the development of postoperative macular edema., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Porcine decellularized conjunctiva (PDC) represents a promising alternative source for conjunctival reconstruction. Methods of its re-epithelialization in vitro with primary human conjunctival epithelial cells (HCEC) have already been established. However, a long-term storage method is required for a simplified clinical use of PDC. This study investigates the influence of several storage variants on PDC. PDC were stored in (1) phosphate-buffered saline solution (PBS) at 4 °C, (2) in glycerol-containing epithelial cell medium (EM/gly) at -80 °C and (3) in dimethyl sulfoxide-containing epithelial cell medium (EM/DMSO) at -196 °C in liquid nitrogen for two and six months, respectively. Fresh PDC served as control. Histological structure, biomechanical parameters, the content of collagen and elastin and the potential of re-epithelialization with primary HCEC under cultivation for 14 days were compared (n = 4-10). In all groups, PDC showed a well-preserved extracellular matrix without structural disruptions and with comparable fiber density ( p ≥ 0.74). Collagen and elastin content were not significantly different between the groups ( p ≥ 0.18; p ≥ 0.13, respectively). With the exception of the significantly reduced tensile strength of PDC after storage at -196 °C in EM/DMSO for six months (0.46 ± 0.21 MPa, p = 0.02), no differences were seen regarding the elastic modulus, tensile strength and extensibility compared to control (0.87 ± 0.25 MPa; p ≥ 0.06). The mean values of the epithelialized PDC surface ranged from 51.9 ± 8.8% (-196 °C) to 78.3 ± 4.4% (-80 °C) and did not differ significantly ( p ≥ 0.35). In conclusion, all examined storage methods were suitable for storing PDC for at least six months. All PDC were able to re-epithelialize, which rules out cytotoxic influences of the storage conditions and suggests preserved biocompatibility for in vivo application.

Purpose: Descemet membrane endothelial keratoplasty (DMEK) accounts for >50% of all corneal transplants in Germany. So far, no data from such a large multicenter study have been published., Methods: This retrospective study included 3200 DMEKs at seven departments performed for Fuchs endothelial corneal dystrophy (FECD) or bullous keratopathy (BK). We evaluated best corrected visual acuity (BCVA, logMAR), endothelial cell density (ECD, cells/mm 2 ), minimal corneal thickness (CT, μm), rebubbling-, primary transplant failure- and immune reaction-rate. Changes over time were evaluated by linear mixed models for repeated measures and correlation with case number by center by weighted linear regression., Results: For patients without vision-limiting comorbidities (74% of all analysed eyes, n = 2270), mean BCVA improved from 0.6 ± 0.4 logMAR to 0.2 ± 0.2 logMAR 6 months (p < 0.001, n = 1441) and 0.1 ± 0.2 logMAR 12 months (p = 0.001, n = 1402) postoperatively. BK- had a worse BCVA compared to FECD-patients (0.3 ± 0.5 vs. 0.1 ± 0.2 logMAR [p < 0.001] at 1 year). ECD declined from 2465 ± 259 cells/mm 2 (n = 2876 preoperatively) to 1587 ± 433 cells/mm 2 after 12 months (p < 0.001, n = 1237). Mean rebubbling rate was 0.4 ± 0.7/eye. 784 eyes (25%) received at least one rebubbling. More rebubblings correlated with a lower ECD, a worse BCVA, a higher CT, and higher transplant failure and rejection rates (p < 0.001, p = 0.013 for BCVA at 12 months). A single rebubbling did not influence the BCVA (p = 0.785). Graft failure rate was 3% (n = 67), rejection rate 1.5% (n = 48)., Conclusion: Descemet membrane endothelial keratoplasty increases visual acuity with low transplant failure- and rejection-rates. FECD has a better outcome than BK. Since a quarter of all patients need a rebubbling, this should be included in the informed consent. Remarkably, one rebubbling has no influence on the outcome., (© 2022 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Background: In recent years, an increasing number of surgical microscopes fitted with an OCT module (intraoperative OCT, iOCT) have become available, providing high-resolution images of the surgical site in real time. While a 2018 survey at our hospital showed that iOCT delivered an additional intraoperative benefit in only 2.4% of all operations, considering that the manufacturer had since revised the hardware and software, we conducted a second user evaluation of this technology., Material and Methods: Prospective monocentric analysis of the application and user-friendliness of an EnFocus Ultra-Deep OCT (Leica Microsystems) over a period of 25 (2018) and 20 working days (2021). A standardized questionnaire was used to assess the surgeons' use of iOCT and its influence on the surgical course., Results: 118 operations were performed over a 25-day period in 2018 and 92 operations were performed over a 20-day period in 2021. In 2018, iOCT was used in 24.6% and in 2021 in 48.9% of all surgeries, with iOCT proving to be "critical" to the surgical course in 2.4% and 3.3% of cases, respectively, as assessed by the surgeons in both years. These were operations in which the intraocular view was limited, e.g., with decompensated cornea, vitreous hemorrhage, or after previous surgery, e.g., after penetrating keratoplasty., Conclusion: Further development of the user interface led to an improvement in usability, and the iOCT was used significantly more often. In both years, the iOCT proved to be critical for the course of the surgery in a comparably small number of operations, especially those involving complex situations., Competing Interests: GG: Leica Microsystems, die anderen Autoren und Autorinnen geben keine Interessenkonflikte an./GG: Leica Microsystems, the other authors declare that they have no conflict of interest., (Thieme. All rights reserved.)

In the case of thermal or caustic burns of the ocular surface, loss of limbal epithelial stem cells leads to compromised self-renewal of the corneal epithelium. This results in permanent loss of vision. In these situations, transplantation of cultured limbal epithelial cells on an amniotic membrane or fibrin gel as substrate (Holoclar®) can help to regenerate the corneal surface. The required cells are obtained from the healthy partner eye, if available. Adult stem cells from other parts of the body potentially serve as alternative cell sources: hair follicles, oral mucosa, mesenchymal stromal cells, or induced pluripotent stem cells (originally, e.g., skin fibroblasts). The reprogramming of such cells can be achieved with the help of transcription factors. In addition, work is being done on biosynthetic or synthetic matrices, which not only serve as substrate material for the transplantation but also support the functional properties of these cells (self-renewal, corneal epithelial-typical phenotype)., (© 2022. The Author(s), under exclusive licence to Springer Medizin Verlag GmbH, ein Teil von Springer Nature.)

Background: The demands on conjunctival replacement tissues are high: they need to be elastic, clinically compatible, surgically feasible and support goblet cell growth., Objective: This article provides an overview of currently applied conjunctival replacement tissues and those under investigation., Method: Current publications on clinically applied conjunctival replacement tissues and substrates which are the subject of scientific research and those already tested in animal models are presented and discussed., Results: Replacement tissues in clinical use are autologous and allogenic conjunctiva, nasal and oral mucous membranes, amniotic membrane and decellularized tissues. Autologous conjunctiva shows good results but is not suitable for large defects due to limited availability. In these cases autologous nasal and oral mucous membranes can be used; however, success is limited in cases of autoimmune diseases. Amniotic membranes are frequently applied clinically but goblet cell growth is limited. Different decellularized tissues are used clinically and goblet cell growth was found in vivo. Robust comparative studies are not yet available. Biological matrices such as fibrin, collagen, elastin, gelatin or hyaluronate and synthetic tissues from the group of polyesters are being investigated in the laboratory and in animal models. These studies show good epithelialization and goblet cell growth in vivo., Conclusion: Transplantation of conjunctiva, nasal and oral mucous membranes and amniotic membranes show satisfactory clinical results but exhibit individual weaknesses. Further studies in animal models and clinical settings are required to further evaluate the benefits of other matrices, such as cell-free tissues or other biological and synthetic matrices., (© 2022. The Author(s), under exclusive licence to Springer Medizin Verlag GmbH, ein Teil von Springer Nature.)

Purpose: The aim was to present a case of lumen obstruction after implantation of microshunt with subconjunctival drainage and subsequent management., Case Report: A 56-year-old male with primary open angle glaucoma underwent uneventful PRESERFLO MicroShunt (PMS) implantation in both eyes. Five months after surgery the patient presented with an intraocular pressure (IOP) of 26 mm Hg in the right eye because of iris obstruction., Results: The iris fibers were severed and removed surgically to free the lumen. Follow-up showed a normalized IOP of 10 mm Hg and a free lumen., Conclusion: Uncontrolled IOP because of iris obstruction following PMS was addressed by surgical removal of iris fibers at the lumen entrance without the need for tube repositioning., Competing Interests: Disclosure: The authors declare no conflict of interest., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Background: In recent years, great progress has been made in intraoperative imaging using optical coherence tomography (iOCT). There are now several commercially available iOCT systems that allow high-resolution imaging of all structures of the eye without interrupting surgery. This real-time visualisation can provide additional information to conventional surgical microscopy, but is relatively expensive. The aim of our study was to find out how often OCT integrated into the surgical microscope is used by trained surgeons, or to what extent they consider that iOCT is relevant for intraoperative procedures., Patients and Methods: A prospective monocentric analysis was conducted of the field of application and user-friendliness of the EnFocus Ultra-Deep OCT (Leica Microsystems), a mobile device combination of surgical microscope and OCT. The use and benefit were investigated of iOCT, which was not mandatory. Standardised documentation and evaluation using a questionnaire was performed by the respective surgeon (n = 5) immediately after surgery., Results: Over a period of 25 working days, 118 procedures were performed in the operating theatre equipped with the microscope-OCT combination. The iOCT was used in 24.6% of the 118 procedures performed. iOCT was regarded as crucial to the intraoperative procedure in 3 of the 29 patients. In one patient, it was possible to check graft orientation during a DMEK operation in a very opaque cornea and, in the second patient, to visualise the correct positioning of an iris diaphragm in the capsular bag. In the third patient, the risk of developing a pseudoforamen was assessed, and this led to the decision not to perform a full gliosis peel., Conclusion: Experienced surgeons in a university eye hospital with a full surgical spectrum considered that intraoperative OCT was decisive for the course of surgery in only a few selected surgical situations, e.g. in case of limited corneal transparency. The impact of the use of iOCT on post-operative outcome quality still needs to be evaluated by larger prospective studies. On the basis of this survey, the cost-benefit ratio is still unclear., Competing Interests: The authors declare that they have no conflict of interest./Die Autorinnen/Autoren geben an, dass kein Interessenkonflikt besteht., (Thieme. All rights reserved.)

Purpose: The long-term success of visual rehabilitation in patients with severe conjunctival scarring is reliant on the reconstruction of the conjunctiva with a suitable substitute. The purpose of this study is the development and investigation of a re-epithelialized conjunctival substitute based on porcine decellularized conjunctiva (PDC)., Methods: PDC was re-epithelialized either with pre-expanded human conjunctival epithelial cells (PDC + HCEC) or with a human conjunctival explant placed directly on PDC (PDC + HCEx). Histology and immunohistochemistry were performed to evaluate epithelial thickness, proliferation (Ki67), apoptosis (Caspase 3), goblet cells (MUC5AC), and progenitor cells (CK15, ΔNp63, ABCG2). The superior construct (PDC + HCEx) was transplanted into a conjunctival defect of a rabbit (n = 6). Lissamine green staining verified the epithelialization in vivo. Orbital tissue was exenterated on day 10 and processed for histological and immunohistochemical analysis to examine the engrafted PDC + HCEx. A human-specific antibody was used to detect the transplanted cells., Results: From day-14 in vitro onward, a significantly thicker epithelium and greater number of cells expressing Ki67, CK15, ΔNp63, and ABCG2 were noted for PDC + HCEx versus PDC + HCEC. MUC5AC-positive cells were found only in PDC + HCEx. The PDC + HCEx-grafted rabbit conjunctivas were lissamine-negative during the evaluation period, indicating epithelial integrity. Engrafted PDC + HCEx showed preserved progenitor cell properties and an increased number of goblet cells comparable to those of native conjunctiva., Conclusion: Placing and culturing a human conjunctival explant directly on PDC (PDC + HCEx) enables the generation of a stable, stratified, goblet cell-rich construct that could provide a promising alternative conjunctival substitute for patients with extensive conjunctival stem and goblet cell loss., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Impact Statement: Conjunctival integrity is crucial for a healthy ocular surface and visual acuity. In severe cases of inflammatory surface disorders or after trauma, thermal or chemical burns as well as after ocular surgery, a surgical reconstruction using conjunctival substitutes is required. Due to limitations of currently used substitutes, such as the amniotic membrane, there is a need for the development of new scaffolds of consistent quality for conjunctival reconstruction. This study explored the biocompatibility and surgical usability of plastic-compressed collagen as an alternative conjunctival substitute in a rabbit model.

Severe dry eye syndrome resulting from lacrimal gland (LG) dysfunction can cause blindness, yet treatments remain palliative. In vitro reconstruction of LG tissue could provide a curative treatment. We aimed to combine epithelial cells with endothelial cells and mesenchymal stem cells (MSCs) to form a 3D functional unit. Epithelial cells and MSCs were isolated from porcine LG; endothelial cells were isolated from human foreskin. MSCs were characterised (flow cytometry and differentiation potential assays). All 3 cell types were combined on Matrigel and spheroid formation observed. Spheroids were characterised [immunohistochemistry (IHC) and transmission electron microscopy] and function assessed (β-hexosaminidase assay). Spheroids were transferred to decellularised jejunum (SIS-Muc) in dynamic cultures for 1 week before further characterisation. MSCs did not express CD31 but expressed CD44 and CD105 and differentiated towards osteogenic and adipogenic lineages. Spheroids formed on Matrigel within 18 hr, contracting to ~10% of the well area (p < .005). IHC revealed presence of all 3 cells within spheroids. Transmission electron microscopy revealed cell-cell contacts and polarisation at the apical surface. In static cultures, function was increased in spheroids cf. monolayer controls (p < .05) but over 72 hr, spheroid function (p < .05), viability (p < .05), and proliferation decreased, whilst apoptosis increased. On SIS-Muc under dynamic culture, however, spheroids continued to proliferate to repopulate SIS-Muc. IHC revealed LG epithelial cells coexpressing pan-cytokeratin and lysozyme, as well as endothelial cells and MSCs and cells remained capable of responding to carbachol (p < .05). These spheroids could form the basis of a regenerative medicine treatment approach for dry eye syndrome. In vivo studies are required to evaluate this further., (Copyright © 2017 John Wiley & Sons, Ltd.)

Purpose: Limbal epithelial stem cells (LESC) reside in a niche in the corneo-scleral transition zone. Deficiency leads to pain, corneal opacity, and eventually blindness. LESC transplantation of ex-vivo expanded human LESC on a carrier such as human amniotic membrane is a current treatment option. We evaluated decellularised human limbus (DHL) as a potential carrier matrix for the transplantation of LESC., Methods: Human corneas were obtained from the local eye bank. The limbal tissue was decellularised by sodium desoxychelate and DNase solution and sterilised by γ-irradiation. Native limbus- and DHL-surface structures were assessed by scanning electron microscopy and collagen ultrastructure using transmission electron microscopy. Presence and preservation of limbal basement membrane proteins in native limbus and DHL were analysed immunohistochemically. Absence of DNA after decellularisation was assessed by Feulgen staining and DNA quantification. Presence of immune cells was explored by CD45 staining, and potential cytotoxicity was tested using a cell viability assay., Results: In the DHL, the DNA content was reduced from 1.5 ± 0.3 μg/mg to 0.15 ± 0.01 μg/mg; the three-dimensional structure and the arrangement of the collagen fibrils were preserved. Main basement membrane proteins such as collagen IV, laminin, and fibronectin were still present after decellularisation and γ-irradiation. CD45-expressing cells were evident neither in the native limbus nor in the DHL. DHL did not convey cytotoxicity., Conclusions: The extracellular matrix (ECM) of the limbus provides a tissue specific morphology and three-dimensionality consisting of particular ECM proteins. It therefore represents a substantial component of the stem cell niche. The DHL provides a specific limbal niche surrounding, and might serve as an easily producible carrier matrix for LESC transplantation.

Conjunctival reconstruction is an integral component of ocular surface restoration. Decellularised tissues are frequently used clinically for tissue engineering. This study identifies porcine decellularised conjunctiva (PDC) and human decellularised conjunctiva (HDC) as promising substitutes for conjunctival reconstruction. PDC and HDC were nearly DNA-free, structurally intact and showed no cytotoxic effects in vitro, which was confirmed by DNA quantification, histology, transmission electron microscopy, collagen quantification and cytotoxicity assay. Comparing the biomechanical properties to amniotic membrane (AM), the most frequently applied matrix for ocular surface reconstruction today, the decellularised conjunctiva was more extensible and elastic but exhibited less tensile strength. The in vivo application in a rabbit model proofed significantly enhanced transplant stability and less suture losses comparing PDC and HDC to AM while none of the matrices induced considerable inflammation. Ten days after implantation, all PDC, 4 of 6 HDC but none of the AM transplants were completely integrated into the recipient conjunctiva with a partially multi-layered epithelium. Altogether, decellularised conjunctivas of porcine and human origin were superior to AM for conjunctival reconstruction after xenogeneic application in vivo., Statement of Significance: Conjunctival integrity is essential for a healthy ocular surface and clear vision. Its reconstruction is required in case of immunological diseases, after trauma, chemical or thermal burns or surgery involving the conjunctiva. Due to limitations of currently used substitute tissues such as amniotic membrane, there is a need for the development of new matrices for conjunctival reconstruction. Decellularised tissues are frequently applied clinically for tissue engineering. The present study identifies porcine and human decellularised conjunctiva as biocompatible and well tolerated scaffolds with superior integration into the recipient conjunctiva compared to amniotic membrane. Decellularised conjunctiva depicts a promising substitute for conjunctival reconstruction in ophthalmology., (Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Purpose: Dry eye syndrome (DES) can cause blindness in severe cases, but mainly palliative treatments exist. A tissue-engineered lacrimal gland (LG) could provide a curative treatment. We aimed to evaluate decellularized porcine jejunum (SIS-Muc) as a scaffold for porcine LG epithelial cells., Methods: To evaluate SIS-Muc as a potential scaffold, basement membrane proteins in SIS-Muc and native LG were compared (immunohistochemistry [IHC]). Porcine LG epithelial cells cultured on plastic were characterized (immunocytochemistry), and their culture supernatant was compared with porcine tears (proteomics). Epithelial cells were then seeded onto SIS-Muc in either a static (cell crown) or dynamic culture (within a perfusion chamber) and metabolic (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) and secretory capacities (β-hexosaminidase assay), protein expression (IHC), and ultrastructure transmission electron microscopy (TEM) compared in each., Results: Collagen IV and laminin were found in both native LG and SIS-Muc. When cultured on plastic, LG epithelial cells expressed pan-cytokeratin, Rab3D, HexA, and produced mucins, but lysozyme and lactoferrin expression was nearly absent. Some porcine tear proteins (lipocalin-2 and lactoferrin) were found in LG epithelial cell culture supernatants. When LG cells were cultured on SIS-Muc, metabolic and β-hexosaminidase activities were greater in dynamic cultures than static cultures (P < 0.05). In both static and dynamic cultures, cells expressed pan-cytokeratin, Rab3D, lysozyme, and lactoferrin and produced mucins, and TEM revealed cell polarization at the apical surface and cell-cell and cell-scaffold contacts., Conclusions: SIS-Muc is a suitable scaffold for LG cell expansion and may be useful toward reconstruction of LG tissue to provide a curative treatment for DES. Dynamic culture enhances cell metabolic and functional activities.

Competing Interests: Interessenkonflikt: Die Autoren bestätigen, dass kein Interessenkonflikt vorliegt.

Aim: To investigate indications, surgical challenges, and outcome of Descemet-membrane endothelial keratoplasty (DMEK) in patients with retinal comorbidities (RC)., Methods: In a prospective cohort study, 8 eyes of 8 DMEK-patients with known RC were compared to 38 eyes of 38 DMEK-patients without RC. The duration of surgery, the degree of difficulty graded by the surgeon, and the complications through DMEK-surgery were analyzed for each patient. The best-corrected visual acuity (BCVA), the endothelial cell count, the intraocular pressure, and the subjective satisfaction was evaluated after a 6-month follow-up. Data were compared applying the non-parametric Wilcoxon-, Chi-square- and Fisheŕs-exact-test with P≤0. 05 as level of significance., Results: RC-patients had dry age-related macular degeneration (n=4) or history of pars-plana vitrectomy (n=4). The main indication for DMEK was pain due to bullous keratopathy for the RC-patients (n=7, 88%) and visual impairment due to Fuchs endothelial keratoplasty for the non-RC-patients (n=33, 87%). The BCVA increased for both groups (P=0.01, P<0.001) and all corneas cleared. For the RC-patients, the subjective satisfaction improved significantly (P=0.02). Oil-filling and missing support of the vitreous body complicated surgery in vitrectomized eyes., Conclusion: DMEK is a favorable technique to treat endothelial disorders even if patients suffer from a retinal comorbidity. By enhancing the corneal clarity, it enables retinal examination or intraocular surgery and increases the patientś satisfaction. However, in vitrectomized or silicone-oil filled eyes, the duration of surgery and degree of complexity are increased. An experienced surgeon should perform DMEK in these patients., Clinical Trial Registration Number: DRKS00007566.

Lacrimal gland (LG) insufficiency is a main cause for severe dry eye leading to pain, visual impairment, and eventually loss of sight. Engineering of transplantable LG tissue with secretory capacity is a desirable goal. In this study, a three-dimensional decellularized LG (DC-LG) scaffold with preserved LG morphology was generated by treatment with 1% sodium deoxycholate and DNase solution using porcine LG tissue. To address clinical applicability, the primary in vitro culture of secretory active LG cells from a small tissue biopsy of 1.5 mm diameter was introduced and compared with an established isolation method by enzymatic digestion. Cells from both isolation methods depicted an epithelial phenotype, maintained their secretory capacity for up to 30 days, and exhibited progenitor cell capacity as measured by aldehyde dehydrogenase-1 activity, side population assay, and colony-forming units. Cells from passage 0 were reseeded into the DC-LG and secretory active cells migrated into the tissue. The cells resembled an LG-like morphology and the constructs showed secretory activity. These results demonstrate the possibility of engineering a secretory competent, three-dimensional LG construct using LG cells expanded from a small tissue biopsy and DC-LG as a matrix that provides the native structure and physiological niche for these cells.

Purpose: To report a case of Urrets-Zavalia syndrome (UZS) after Descemet membrane endothelial keratoplasty (DMEK)., Methods: A 74-year-old woman with Fuchs endothelial dystrophy and inconspicuous ocular history developed UZS after DMEK surgery. The intraoperative and postoperative course is presented., Results: After uneventful DMEK surgery, intraocular pressure was elevated up to 40 mm Hg on the first postoperative day. A small bleed from the peripheral wide-open iridectomy in the 12 o'clock position in the otherwise deep anterior chamber was observed. On the sixth postoperative day, a 4-mm-wide pupil, nonreactive to light, was noted. One year after surgery, the fixed medium mydriasis (4 mm) persisted and best-corrected vision was 0.1 logMAR. No pupillary reaction was noted after application of 0.2% or 2% pilocarpine., Conclusions: Filling the anterior chamber with air to secure fixation of a grafted Descemet membrane carries the risk of early acute postoperative ocular hypertension. This can lead to iris sphincter defects resulting in a fixed dilated pupil after DMEK surgery. Large patent iridectomy in the 12 o'clock position is insufficient to prevent this. Patients undergoing DMEK surgery should be informed about this potential complication.

Purpose: The application of lacrimal gland-derived mesenchymal stem cells (LG-MSC) for the regeneration of lacrimal gland tissue could result in a novel therapy for dry-eye syndrome. To optimize the culture conditions, the purpose of this study was to evaluate the influence of low oxygen on phenotype, differentiation potential, proliferative, and regenerative capacity of murine LG-MSC., Methods: Murine LG-MSC were cultured in 21% and 5% oxygen and characterized by flow cytometry, cell sorter assisted proliferation-, and colony forming unit-assays. Reactive oxygen species (ROS) levels as well as lineage differentiation were evaluated. The effect of conditioned medium of LG-MSC from both oxygen conditions (CM MSC 21%, respectively, CM MSC 5%) on lacrimal gland epithelial cells (LG-EC) was examined in wound healing and proliferation assays., Results: Cells under both culture conditions revealed differentiation potential and presented a MSC-specific flow cytometric phenotype. In 5% oxygen, cells yielded less ROS, showed a stable morphology, higher colony forming potential, and an increased proliferation capacity. Five percent oxygen significantly increased the number of CD44+ LG-MSC. Furthermore, CM MSC 5% significantly enhanced migration and proliferation in LG-EC., Conclusions: In vitro expansion in low oxygen preserves the proliferation capacity and differentiation potential of LG-MSC and increases the effects of conditioned medium on migration and proliferation in LG-EC. Therefore, expansion in low oxygen seems to be an excellent method, to obtain vital MSC. Also, an increased number of LG-MSC expressing CD44 was observed under low oxygen, which might be a valuable marker to identify a potent MSC subpopulation.

Background: An elevated intraocular pressure remains the main risk factor for the development and progression of glaucoma but other factors such as fluctuations of the intraocular pressure, the blood pressure, and the ocular perfusion pressure are gaining in importance. The influence of the anti-glaucomatous therapies on these parameters is barely investigated. This study investigates 1) a correlation between the fluctuations of the intraocular pressure, the blood pressure and the ocular perfusion pressure with the severity of the glaucoma damage and 2) if these parameters can be influenced by different anti-glaucomatous therapies., Material and Methods: Data from 121 glaucoma patients who had been hospitalised for 24-hour measurements of the intraocular pressure and blood pressure between 2003 and 2012 were analysed retrospectively. The patients were divided into groups by the severity of the glaucoma damage and by their anti-glaucomatous therapy. Mean values for the intraocular pressure, the blood pressure and the ocular perfusion pressure were calculated and correlated with the individual groups. The statistical analysis was performed with SPSS21.0 using Kruskal-Wallis and Mann-Whitney U tests for the comparison of the groups. RESULTS with p < 0.05 were considered statistically significant., Results: The severity of the glaucoma damage correlated significantly with higher mean values of the intraocular pressure and its fluctuations, as well as with a nocturnal decrease of the blood pressure and low mean values of the ocular perfusion pressure (p < 0.0001). The anti-glaucomatous therapy decreased the intraocular pressure to normal values. A monotherapy with carboanhydrase inhibitors correlated with the lowest fluctuations of the ocular perfusion pressure while a quadri-therapy was associated with higher values of the intraocular pressure and its fluctuations. The anti-glaucomatous therapy did not show an association with the blood pressure or the ocular perfusion pressure., Conclusion: Glaucoma is negatively influenced by high intraocular pressure, nocturnal blood pressure decreases and ocular perfusion pressure fluctuations. A monotherapy with carboanhydrase inhibitors seems to be sufficient to decrease fluctuations of the intraocular pressure. Patients with a quadri-therapy should be monitored regularly as they present high values of intraocular pressure and its fluctuations. A hospitalisation of the patients with 24-hour measurements of the intraocular pressure and blood pressure provides information needed to improve the therapy or decide about surgical interventions., (Georg Thieme Verlag KG Stuttgart · New York.)

Objective: Congenital nasolacrimal duct obstruction (CNDO) is the most common cause of neonatal epiphora. Persistence can lead to chronic dacryocystitis and amblyopia. This study analyzed the association between the incidence of CNDO and delivery by cesarean section., Study Design: This was a retrospective cohort study of 386 children with CNDO (born between 2000 and 2008). The incidence of the delivery mode in patients with CNDO was compared with data from a corresponding population derived from annual birth statistics., Results: There was no statistically significant association between the overall cesarean section rate and the incidence of CNDO, but primary cesarean section was significantly more frequent among patients with CNDO (73.15%, p < 0.05). The difference was significant for both genders for the period from 2000 to 2008 (p < 0.05%). The relative risk for CNDO was 1.7-fold increased in children delivered by primary cesarean section., Conclusion: Primary cesarean section may be a risk factor for CNDO., (Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.)

Background: The potential negative influence of angiogenic gene therapy on the development or progression of retinal pathologies such as diabetic retinopathy (DR) or age-related macular degeneration (AMD) has led to the systematic exclusion of affected patients from trials. We investigated the role of nonviral fibroblast factor 1 (NV1FGF) in two phase II, multinational, double-blind, randomized, placebo-controlled, gene therapy trials (TALISMAN 201 and 211)., Methods: One hundred and fifty-two subjects with critical limb ischemia or claudication were randomized to receive eight intramuscular injections of 2.5 ml of NV1FGF at 0.2 mg/ml or 0.4 mg/dl or placebo. One hundred and fifty-two patients received a plasmid dose of NV1FGF of up to 32 mg or placebo. All patients underwent a systematic ophthalmologic examination at baseline and at 3, 6 or 12 months following gene therapy. Twenty-six of these patients (Münster subgroup) received a retinal fluorescence angiography at baseline and at final examination., Results: Among those 26 patients, four of nine patients with diabetes suffered from nonproliferative DR. Three patients showed non-exsudative AMD. No change of retinal morphology or function was observed in Münster subgroup of both TALISMAN trials independent of the intramuscular NV1FGF dosage applied., Conclusions: Angiogenic gene therapy using NV1FGF is safe even in diabetics., (Copyright © 2014 John Wiley & Sons, Ltd.)

Purpose: Autologous serum eye drops are used for therapy of severe ocular surface disorders by patients with visual and manual impairments. Until recently, they were prepared under sterile conditions from open blood sampling systems. Closed blood donation systems simplify production. This study compares handling and costs of a new day dosage vial ("Meise-vial") and a single-dose tube system ("Maco-tube") based on closed production systems with conventional dropper bottles., Methods: Nonimpaired volunteers and patients with visual or manual impairment (n = 10 each group) single-handedly tested the applicators filled with 1.5 ml sterile isotonic saline solution. Participants rated convenience of opening the containers and applying eye drops on a scale from 1.0 (very good) to 6.0 (very bad). Number of retrievable drops was counted. Participants were asked which system they prefer, both with and without knowledge of the price for the systems., Results: The median for convenience of opening (eye drop application) was 2.0 (1.0) for Meise-vials, 5.0 (4.0) for Maco-tubes, and 2.0 (2.0) for dropper bottles (p < 0.001). Median number of drops retrieved from the systems was 30.5 (vials), 2 (tubes), and 30 (bottles). Ranking did not differ between nonimpaired and impaired participants. Assuming equal prices, 16 participants chose Meise-vials, 14 dropper bottles, and no tubes. With knowledge of pricing, preference changed (p = 0.001), 20 participants (67%) opted for dropper bottles and 5 (17%) preferred the other containers., Conclusion: Convenience of opening, applying eye drops, and number of drops retrieved was substantially better for dropper bottles and Meise-vials compared with Maco-tubes. Bottles and vials were equally well received. With regard to price, nonimpaired as well as impaired participants preferred dropper bottles. While closed systems simplify production, patients preferred dropper bottles for daily application of autologous serum eye drops for a number of reasons.

In recent years, a special type of cancer cell--the cancer stem cell (CSC)--has been identified and characterized for different tumors. CSCs may be responsible for the recurrence of a tumor following a primarily successful therapy and are thought to bear a high metastatic potential. For the development of efficient treatment strategies, the establishment of reliable methods for the identification and effective isolation of CSCs is imperative. Similar to their stem cell counterparts in bone marrow or small intestine, different cluster of differentiation surface antigens have been characterized, thus enabling researchers to identify them within the tumor bulk and to determine their degree of differentiation. In addition, functional properties characteristic of stem cells can be measured. Side population analysis is based on the stem cell-specific activity of certain ATP-binding cassette transporter proteins, which are able to transport fluorescent dyes out of the cells. Furthermore, the stem cell-specific presence of aldehyde dehydrogenase isoform 1 can be used for CSC labeling. However, the flow cytometric analysis of these CSC functional features requires specific technical adjustments. This review focuses on the principles and strategies of the flow cytometric analysis of CSCs and provides an overview of current protocols as well as technical requirements and pitfalls. A special focus is set on side population analysis and analysis of ALDH activity. Flow cytometry-based sorting principles and future flow cytometric applications for CSC analysis are also discussed., (Copyright © 2012 International Society for Advancement of Cytometry.)

Polo-like kinase 1 (PLK1) is a regulator of mitosis and its upregulation in tumours is often associated with poor prognosis. Although PLK1 inhibitors have already entered phase 1 clinical trials, little is known about their impact on the treatment of paediatric malignancies. Thus, we evaluated the concept of PKL1 inhibition by testing the effects of the PLK1 inhibitor GW843682X alone and in combination with the topoisomerase 1 inhibitor, camptothecin, against a panel of 18 paediatric tumour cell lines. Cytotoxicity was evaluated by MTT test and by caspase 3/7 activation. Expression of target was confirmed by western blot analysis. Expression of ATP binding cassette transporters was analysed by quantitative real-time reverse transcription PCR. GW843682X significantly inhibited cell growth in all 18 cell lines. Concentrations, which inhibited cell growth by 50% compared with untreated controls after 72 h, ranged from 0.02 to 11.7 μmol/l. Apart from the N-Myc-amplified neuroblastoma cell lines, the osteosarcoma cell lines MNNG-HOS and OST, which are highly resistant to standard anticancer drugs, were sensitive to GW843682X. The toxicity of GW843682X was dependent neither on the ATP binding cassette drug transporter expression nor on the p53 mutation status. Neither synergistic nor antagonistic effects were observed for the combination of GW843682X and camptothecin in 14 cell lines. GW843682X showed considerable toxicity against a panel of paediatric tumour cell lines suggesting that PLK1 inhibitors under clinical development should be evaluated against paediatric malignancies too.

Background: Basal cell carcinoma (BCC) is the most common malignancy of the skin and of the eyelid in the Caucasian population. Our study evaluated the long-term results of 5% imiquimod cream therapy for nodular BCC of the eyelid as an alternative to surgical approaches., Methods: Five patients suffering from histologically proven nodular BCC of the eyelid who had refused surgical treatment were included in this interventional off-label use study. The patients applied 5% imiquimod cream topically five times a week for 6 weeks at the site of the tumorous lesion. Patients were followed up regularly for up to 7 years to check for tumor disappearance or recurrence, and for local and systemic side-effects., Results: Complete long-term clinical clearance was obtained in four of the five patients, with no tumor recurrence after 7 years of follow-up. Cosmetic results were excellent. One patient refused to continue the treatment 2 weeks after therapy onset, due to significant subjective discomfort. No serious local side-effects, and no systemic side-effects at all were observed following treatment., Conclusions: Imiquimod cream (5%) provides an effective alternative therapy for the treatment of nodular BCC of the eyelid, although surgical treatment remains the gold standard at the present time. This study is the first to assess the long-term efficacy and safety of this approach. Trial studies are necessary.

Background: Irrigation of the lacrimal duct with silicone tube intubation, endoscopic microdrill dacryoplasty and external dacryocystorhinostomy (DCR) are commonly used in lacrimal duct surgery. Dacryocystography (DCG) is one of the standard procedures in preoperative imaging. This study evaluates the influence of individual parameters and the prognostic meaning of preoperative DCG on the success rates after lacrimal duct surgery, in order to predict more precisely the long-term outcome after different operative paradigms., Materials and Methods: 145 patients (mean age 60 years; 32 % male) with lacrimal stenosis, who underwent lacrimal duct surgery in the University Eye Hospital Münster during 2002 were included in this retrospective study. Individual parameters (age, gender, preoperative duration of symptoms, lacrimal sac area and operative procedures) were recorded. The lacrimal sac area of each patient was calculated from preoperative DCG imaging. The individual parameters and the prognostic relevance of preoperative DCG were statistically analysed with regards to the long-term success rate after different surgical procedures. Criteria for success were an asymptomatic patient and a successful probing and irrigation of the lacrimal duct system at follow-up at 24 to 36 months after lacrimal duct surgery., Results: With a mean success rate of 79 %, DCR was most effective regarding long-term outcome. Particularly in elderly patients DCG was superior to minimal-invasive procedures. The mean area of the lacrimal sac, calculated from DCG imaging, was 18 mm (2). There was a significant correlation (p = 0.001) between the area of the lacrimal sac and long-term success of DCR., Conclusion: Endoscopic microdrill dacryoplasty and DCR have the most successful options in the treatment of lacrimal stenosis. DCG was the most effective procedure in elderly patients. An enlargred lacrimal sac proved to be a positive prognostic factor for the long-term success of DCR., ((c) Georg Thieme Verlag KG Stuttgart New York.)