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The advent of cell culture-based methods for the establishment and expansion of human corneal endothelial cells (CEnC) has provided a source of transplantable corneal endothelium, with a significant potential to challenge the one donor-one recipient paradigm. However, concerns over cell identity remain, and a comprehensive characterization of the cultured CEnC across serial passages has not been performed. To this end, we compared two established CEnC culture methods by assessing the transcriptomic changes that occur during in vitro expansion. In confluent monolayers, low mitogenic culture conditions preserved corneal endothelial cell state identity better than culture in high mitogenic conditions. Expansion by continuous passaging induced replicative cell senescence. Transcriptomic analysis of the senescent phenotype identified a cell senescence signature distinct for CEnC. We identified activation of both classic and new cell signaling pathways that may be targeted to prevent senescence, a significant barrier to realizing the potential clinical utility of in vitro expansion.

Graphical abstract Abstract While tissue form and function is highly dependent upon tissue-specific collagen composition and organization, little is known of the mechanisms controlling the bundling of collagen fibrils into fibers and larger structural designs that lead to the formation of bones, tendons and other tissues. Using the cornea as a model system, our previous 3 dimensional mapping of collagen fiber organization has demonstrated that macrostructural organization of collagen fibers involving interweaving, branching and anastomosing plays a critical role in controlling mechanical stiffness, corneal shape and refractive power. In this work, the cellular and mechanical mechanisms regulating critical events in the assembly of collagen macrostructure are analysed in the developing chicken cornea. We elucidated the temporal events leading to adult corneal structure and determined the effects of intraocular pressure (IOP) on the organization of the collagen macrostructure. Our findings indicate that the complex adult collagen organization begins to appear on embryonic day 10 (E10) after deposition of the primary stroma and full invasion of keratocytes. Importantly, organizational changes in keratocytes appearing at E9 preceded and predicted later changes in collagen organization. Corneal collagen organization remained unaffected when the development of IOP was blocked at E4. These findings support a primary role for keratocytes in controlling stromal organization, mechanical stiffness and corneal shape that are not regulated by the IOP. Our findings also suggest that the avian cornea represents an excellent experimental model for elucidating key regulatory steps and mechanisms controlling the collagen fiber organization that is critical to determining tissue form and function. Statement of significance This work by using an ex ovo model system, begins to investigate the potential mechanisms controlling collagen fibril macrostructure. In particular, this work highlights a convergent role for the corneal keratocytes in organizing the complex collagen macrostructure, necessary to support high visual acuity. Our data supports that the intraocular pressure does not influence collagen fibril macrostructure and suggest that the avian cornea represents an excellent experimental model for elucidating key regulatory steps and mechanisms controlling the collagen fiber organization that is critical to determining tissue form and function. Clearly understanding the cellular and molecular mechanisms that underlie collagen fibril macrostructure will be highly beneficial for future tissue engineering and regenerative medicine applications. [ABSTRACT FROM AUTHOR]

Optisol–GS storage medium allows donor corneas to remain viable until they are transplanted to cornea-blind recipients. In order to remain viable, the corneal endothelial cell density (ECD) must remain above 2 200 cells per mm 2 . The objective of this study is to determine how long donor corneas may remain viable in Optisol-GS based on their ECD. Although our findings show that the ECD of corneas stored in Optisol-GS decreased steadily, it remained above the critical level of 2 200 cells per mm 2 for up to 9 days, suggesting that corneal buttons may remain viable for at least this amount of time. [ABSTRACT FROM AUTHOR]

To describe the natural history, management, and visual outcome in children with congenital primary aphakia (CPA). This is a multicenter retrospective consecutive case series from five academic centers in England and North America. A total of 27 eyes of 14 patients were included (male:female, 1.7:1). Thirteen patients had bilateral CPA, and 1 patient had unilateral CPA. Mean age at diagnosis was 18 months (median, 21; range, 0.5-144). Of 11 patients who underwent genetic testing, 9 had FOXE3 pathogenic variants. In all patients, visual acuity at presentation was not better than fixing and following light. Typical findings included silvery appearance of the cornea with vascularization (96%), glaucoma (81%), iridocorneal adhesions (74%), optic nerve coloboma (55%), abnormal vitreous (33%), retinal detachment (30%), and aniridia with hypoplasia of ciliary body (19%). Surgical interventions in select patients included penetrating keratoplasty (PKP), glaucoma drainage device implantation, and cyclophotocoagulation (CPC). Eyes with corneal ectasia and a silvery appearance of the cornea with vascularization should alert the physician to the possibility of CPA. Glaucoma causes globe enlargement and may increase the risk of corneal perforation, but glaucoma is often refractory to medical treatment, and the threshold for surgical treatment should be low. PKP outcomes are very poor.▪ [ABSTRACT FROM AUTHOR]

Abstract: Due to the high demand for donor corneas and their low supply, autologous corneal endothelial cell (CEC) culture and transplantation for treatment of corneal endothelial dysfunction would be highly desirable. Many studies have shown the possibility of culturing CECs in vitro, but lack potential robust substrates for transplantation into the cornea. In this study, we investigate the properties of novel ultrathin chitosan–poly(ethylene glycol) (PEG) hydrogel films (CPHFs) for corneal tissue engineering applications. Cross-linking of chitosan films with diepoxy-PEG and cystamine was employed to prepare ∼50μm (hydrated) hydrogel films. Through variation of the PEG content (1.5–5.9wt.%) it was possible to tailor the CPHFs to have tensile strains and ultimate stresses identical to or greater than those of human corneal tissue while retaining similar tensile moduli. Light transmission measurements in the visible spectrum (400–700nm) revealed that the films were >95% optically transparent, above that of the human cornea (maximum ∼90%), whilst in vitro degradation studies with lysozyme revealed that the CPHFs maintained the biodegradable characteristics of chitosan. Cell culture studies demonstrated the ability of the CPHFs to support the attachment and proliferation of sheep CECs. Ex vivo surgical trials on ovine eyes demonstrated that the CPHFs displayed excellent characteristics for physical manipulation and implantation purposes. The ultrathin CPHFs display desirable mechanical, optical and degradation properties whilst allowing attachment and proliferation of ovine CECs, and as such are attractive candidates for the regeneration and transplantation of CECs, as well as other corneal tissue engineering applications. [Copyright &y& Elsevier]

Copyright of Canadian Journal of Ophthalmology is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Corneal transplantation has been performed successfully for over 100 years. Normally, HLA typing and systemic immunosuppressive drugs are not utilized, yet 90% of corneal allografts survive. In rodents, corneal allografts representing maximal histoincompatibility enjoy >50% survival even without immunosuppressive drugs. By contrast, other categories of transplants are invariably rejected in such donor/host combinations. The acceptance of corneal allografts compared to other categories of allografts is called immune privilege. The cornea expresses factors that contribute to immune privilege by preventing the induction and expression of immune responses to histocompatibility antigens on the corneal allograft. Among these are soluble and cell membrane molecules that block immune effector elements and also apoptosis of T lymphocytes. However, some conditions rob the corneal allograft of its immune privilege and promote rejection, which remains the leading cause of corneal allograft failure. Recent studies have examined new strategies for restoring immune privilege to such high-risk hosts. [ABSTRACT FROM AUTHOR]

Copyright of IRBM is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of IRBM is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The aim of this study was to examine the kinetic profile of bioactive TNF levels in aqueous humour of rabbit eyes undergoing corneal allograft rejection and to investigate the effect of locally blocking TNF activity after corneal transplantation. In a rabbit corneal transplantation, endothelial allograft rejection was identified and correlated with increase in central graft thickness. Samples of aqueous humour obtained on alternate days following transplantation were tested for TNF mRNA and bioactive TNF protein. To investigate the effect of locally blocking TNF activity in allograft recipients, the fusion protein TNFR-Ig was administered by injections into the anterior chamber after transplantation. Pulsatile increases in levels of this cytokine were found in 14 of 15 allograft recipients. Peaks of TNF bioactivity preceded by varying intervals the observed onset of rejection in allograft recipients. TNF levels were not elevated in aqueous humour from corneal autograft recipient controls or in serum of allografted animals. mRNA levels were elevated before onset of and during clinically observed allograft rejection. In three of seven animals receiving TNFR-Ig injections on alternate days from day 8 to day 16 post-transplant, clear prolongation of corneal allograft survival was demonstrated. Bioactive TNF is present in aqueous humour following rabbit corneal allotransplantation. Rather than correlating directly with endothelial rejection onset, pulsatile peak levels of TNF precede and follow the observed onset of endothelial rejection. Blockade of TNF activity prolongs corneal allograft survival in some animals, indicating that this cytokine may be a suitable target in local therapy of corneal allograft rejection. [ABSTRACT FROM AUTHOR]

In a rat model of corneal transplantation, Fischer 344 (RT1lv1) rats received orthotopic corneal isografts or Wistar-Furth (RT1u) donor allografts. Rejection was observed in 25 of 26 allograft recipients, at a median time of 18 days, with all isografts surviving > 100 days. Flow cytometric analysis of aqueous humour identified cellular infiltration of the aqueous at the time of allograft rejection, in contrast to the acellular aqueous found in isografts at corresponding limes following transplantation. A higher proportion of CD8+ than CD4+ cells was found at days 1-3 following rejection, whereas there was a higher proportion of CD4+ cells at days 5-8. No changes in peripheral blood T cell subsets were found at the time of rejection. Immunohistochemical analysis of cells infiltrating recipient iris and grafted cornea undertaken at days 1-2, 4 and 7-10 following onset of rejection, demonstrated inflammatory cells in the graft epithelium, stroma and aggregated on the endothelium. Large numbers of macrophages, T cells (CD4+ > CD8+ at all time points), natural killer (NK) cells and neutrophils were detected in graft tissue at days 1-2 and 4, diminishing after that time. Most infiltrating cells expressed MHC class II antigen, and a smaller number expressed IL-2R. Expression of the co-stimulatory marker B7 was identified in a few cells at day 4 in the region of the graft-host wound. The immune response in graft rejection was characterized at day 4 also by expression of intercellular adhesion molecule-1 (ICAM-1) on endothelial cells of iris and cornea, vessels, demonstration of interferon-gamma on mononuctear cells in the peripheral (recipient) cornea, and tumour necrosis factor-alpha on aggregated mononuclear cells on the graft, but not recipient, endothelium. Only sparse cellular infiltrates were found in isograft controls, with inflammation located at the graft-host wound. These findings suggest that inflammatory cells reach a corneal allograft by two routes-from vessels in the peripheral recipient cornea, and from vessels in the recipient iris via the aqueous humour. Different aqueous and intragraft T cell subset proportions were seen early in rejection, although a preponderance of CD4+ cells was found in both aqueous and graft at later times. [ABSTRACT FROM AUTHOR]

The article comments on the 1906 landmark paper of Eduard Zirm, the first doctor who successfully performed a full-thickness corneal transplant. A case report was presented, describing the methods and procedures used in the corneal transplant, as well as the quality of the cornea and the tissue donated by a young boy. Also, Zirm mentioned the other conduct of operation which was important to the successful outcome besides meticulous asepsis and general anesthesia.

Background/aims Diagnosis of rejection in the mouse model of corneal transplantation is based on subjective judgement of loss of graft transparency. The aims of this study were to (1) evaluate a pachymetry technique to measure changes in mouse corneal thickness and (2) correlate increases in transplant thickness with clinical and histological features of rejection. Methods Orthotopic corneal allografts (C57BL/6 strain donor) and syngeneic grafts were performed in A/J mice. Graft transparency was graded and corneal thickness measured by pachymetry on alternate days. Transverse sections of donor cornea excised from eyes representative of clinical opacity grades 1-4 were prepared, photographed, graft section thickness measured and stromal graft-infiltrating cells counted. Intraobserver and interobserver variations in pachymetry were statistically tested. Results Graft thickness, as measured by pachymetry, increased with each clinical opacity grade. Thickness for opacity grades 0, 1 and 2 was less than 300 μm in all recipients. Graft thickness for grades 3 and 4 was greater than 300 μm in all cases. For measurements up to 400 μm, there was a good correlation between thickness as measured by in vivo pachymetry and in histopathological sections. The mean interobserver bias was -11.35 .μm, while the mean intraobserver bias was +3.96 μm. Stromal cellularity increased with increasing corneal thickness up to approximately 300 μm. Conclusion In vivo graft pachymetry provides a new and reliable way to objectively diagnose rejection in the mouse model of corneal transplantation. [ABSTRACT FROM AUTHOR]

Objective: to analyze negative laboratory results of cadaver cornea donor screening during preparation of corneas for transplantation according to data from the internal registry of donors of the eye bank (EB) of the Fyodorov Eye Microsurgery Federal State Institution and the European Eye Bank Association (EEBA) from 2011 through 2015. Materials and methods. Data analysis was carried out using the internal registry of EB donors and the EEBA annual directories. The analyzed data included the number of eyeballs obtained, the frequency of incomplete tests (hemolysis for EB) and positive serological results for human immunodeficiency virus (HIV-1 and HIV-2), viral hepatitis B, viral hepatitis C and syphilis. Results. In just 5 years, the EB received 3,479 eyeballs. After hemolysis of donor blood samples, 13.9% (n = 486) of corneas were excluded from the EB. EEBA recorded fewer inconclusive tests during the same period. After hemolysis and positive serological tests, 19.4% (n = 676) of corneas were excluded from the EB. Overall, the number of positive serological tests in EBs was far higher than in the EEBA data. Frequency of positive HIV tests (HIV-1 and HIV-2) and syphilis in EB showed low variability annually, while incidence of hepatitis B increased in 2015. For the analyzed period, positive serology for hepatitis C was found to be prevalent among EB donors. Mixed infections were quite often recorded in blood samples. Conclusion. Based on analysis conducted, positive serology and hemolysis were the main contraindications and led to exclusion of 33.3% (n = 1162) of cadaver donor corneas received in EB. Frequency of positive serological tests for indicated infections in EB was higher than in the EEBA data, with significant predomination of hepatitis C.

Khaled Alzahrani,1,2 Syarifah-Faiza Dardin,1 Fiona Carley,2 Arun Brahma,2 Debbie Morley,2 M Chantal Hillarby1 1Division of Pharmacy and Optometry, School of Health Sciences, University of Manchester, Manchester, UK; 2Manchester Royal Eye Hospital, Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Science Centre, Manchester, UK Purpose: To compare the corneal clarity measurement between penetrating keratoplasty (PK) and deep anterior lamellar keratoplasty (DALK) in patients with keratoconus, using densitometry software for the Oculus Pentacam.Methods: A retrospective comparative study was carried out at Manchester Royal Eye Hospital. Data were collected 12–18 months after corneal transplantation for keratoconus, including postoperative corneal densitometry, best corrected visual acuity (BCVA), central corneal thickness (CCT), and other relevant clinical details.Results: Analysis of 37 keratoconus eyes from 36 patients found there was a significantly higher corneal densitometry measurement after DALK than PK. This was predominantly in the posterior layer of the concentric zone 0–2 mm of the cornea (P=0.0004). A significant correlation was found between postoperative BCVA and corneal densitometry in DALK groups at full thickness (P=0.03). This correlation was seen in the central 0–2 mm (P=0.03) and posterior 0–2 mm (P=0.04) zones. In addition, within the DALK group, a correlation was found between central corneal thickness and densitometry at full thickness 2–6 mm (P=0.007), central 0–2 (P=0.04), central 2–6 mm (P=0.01), and at posterior 2–6 mm (P=0.01) zones.Conclusion: This study showed that corneal densitometry measurement differs depending on the type of corneal transplantation used to treat keratoconus patients. Densitometry may have an important role to play in the final BCVA achieved by patients undergoing corneal transplantation for keratoconus. Analysis of Oculus Pentacam images provides an objective evaluation to monitor the cornea status after the surgery. Keywords: keratocytes, corneal transplantation, corneal wound healing, keratoconus, transparency

Introduction: A significant cause of visual impairment in the developing world is corneal blindness which may be reversed successfully by transplantation. The availability of corneas transplantation however, is way less than the actual requirement in India. Aim: To assess the knowledge of medical students regarding the eye donation. Settings and Design: Questionnaire based cross-sectional study Materials and Methods: One hundred and twenty six newly enrolled medical students in age group 17-23 years were made to answer a pre tested semi-structured questionnaire on eye donation. Questions were based on testing their knowledge as well as their views towards eye donation. Statistical Analysis was done using SPSS 22 Results: It was observed that a majority (70.63%) of the students, though willing to pledge their eyes, were unaware of the technicalities and held a lot of misconceptions. Mass media was revealed to be the main informant. Many, however did not wish to pledge their eyes due to religious and social beliefs. Conclusion: Even though medical students were aware about eye donation and corneal transplantation, there were many lacunae and misconceptions which need to be refined and cleared to enhance their knowledge and hence increase the number of eye donations in our country